阅读说明：本技术 一种聚乙二醇偶联药物、其制备方法及用途 (Polyethylene glycol coupled drug, preparation method and application thereof ) 是由 李高全 李大军 张倩 卫玉松 彭永陈 阳享伟 曾下凡 梅刚 李蝶蝶 高承志 丁小 于 2020-11-18 设计创作，主要内容包括：本发明涉及一种聚乙二醇偶联药物、其制备方法及用途,具体涉及如式I所示的聚乙二醇偶联药物或其药学上可接受的盐。本发明还涉及所述聚乙二醇偶联药物或其药学上可接受的盐的制备方法和中间体,包含所述聚乙二醇偶联药物或其药学上可接受的盐的药物组合物和在制备药物中的用途。(The invention relates to a polyethylene glycol coupled drug, a preparation method and application thereof, in particular to a polyethylene glycol coupled drug shown as a formula I or pharmaceutically acceptable salt thereof. The invention also relates to a preparation method and an intermediate of the polyethylene glycol conjugate drug or the pharmaceutically acceptable salt thereof, a pharmaceutical composition containing the polyethylene glycol conjugate drug or the pharmaceutically acceptable salt thereof and application of the polyethylene glycol conjugate drug or the pharmaceutically acceptable salt thereof in preparing drugs.)

1. A polyethylene glycol coupling drug shown as a formula (I) or pharmaceutically acceptable salt thereof,

wherein:

m is-C (═ O) C_1-6Alkylene group C (O) -, -NHC_1-6Alkylene NH-, -C (═ O) C_1-6Alkylene NH-, or PEG_m(ii) a Wherein, PEG_mIs a single-arm or multi-arm (e.g. four-arm, eight-arm, preferably four-arm) polyethylene glycol segment, PEG_mThe number average molecular weight of (a) is 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 5k or 10 k;

a1, A1' are independently

A2 is independently a direct bond or

L1 is independently a direct bond or-C (═ O) C_1-6Alkylene C (═ O) -;

w1, W1' and W2 are independently Q1, -Z0—(Q1)₂、

Z4, Z3, Z2, Z1 and Z0 are independently

Q is-N-AC;

q1 is-N1-AC 1;

q2 is-N2-AC 2;

n, N1 and N2 are independently GFLG, G,(preferably, it is)；

AC. AC1, AC2 are independently drug molecules (e.g. drug molecules with anti-tumor activity), preferably PCB, DOX, LPT, SB7, PKA, SN38, PTX or NPB;

n1, n2 are independently 0, 1, 2, 3, 4, 5 or 6;

y, Y' are independently a direct bond, GLFG, -C (═ O) C₁-₆Alkylene group C (═ O) -),

X, X' is independently

PEG is a single-arm polyethylene glycol chain segment, and the number average molecular weight of the PEG is independently 5k-40 k;

j1 is 1, 2, 3, 4, 5 or 6;

j2, j3 are independently 0 or 1.

2. The polyethylene glycol conjugate drug of claim 1, having a structure represented by formula (II), or a pharmaceutically acceptable salt thereof,

wherein:

m is-C (═ O) C_1-6Alkylene group C (═ O) -),-NHC_1-6Alkylene NH-, or PEG_mPreferably, it is Or PEG_mWherein, PEG_mIs a single-arm or multi-arm (e.g. four-arm, eight-arm, preferably four-arm) polyethylene glycol segment, PEG_mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 5k or 10k,

a1 is independently Preferably, it is

Y is independently a direct bond,GLFG、-C(＝O)C_1-6Alkylene group C (═ O) -), Preferably a direct bond,GLFG、

X is independently

W1 is independently

Z4, Z3, Z2, Z1 and Z0 are independently Preferably, it is

Q is-N-AC, and Q is-N-AC,

q1 is-N1-AC 1,

q2 is-N2-AC 2,

n, N1 and N2 are independently G, GFLG,(preferably, it is)，

AC. AC1, AC2 are independently SN38, PKA, PCB, LPT, SB7, PTX or NPB,

the number average molecular weight of the PEG is independently 5k to 40 k.

3. The polyethylene glycol conjugate drug of claim 2, or a pharmaceutically acceptable salt thereof, wherein:

m is-C (═ O) C_1-6Alkylene radicals C (═ O) -, A1 areY isX isW1 isZ4, Z2 and Z1 areZ3 and Z0 are

Preferably, M isA1 isY isX isW1 isZ4, Z2 and Z1 areZ3 and Z0 areQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is PCB, and AC2 is PKA;

Alternatively, M is-NHC_1-6Alkylene NH-, A1 beingY is GLFG and X isW1 isZ2 isZ1 isZ0 is

Preferably, M isA1 isY is GLFG and X isW1 isZ2 isZ1 isZ0 isQ is-N-AC, N is G, AC is SN 38;

or, M isA1 isY is a direct bond or-C (═ O) C_1-6Alkylene C (═ O) -, X isW1 isZ1 isZ0 is

Preferably, M isA1 isY is a direct bond and X isW1 isZ1 isZ0 isQ is-N-AC, N is GFLG, AC is LPT; or

Preferably, M isA1 isY isX isW1 isZ1 isZ0 isQ is-N-AC, N is GFLG, AC is PCB;

or, M isA1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 is

Preferably, M isA1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is NPB, and AC2 is PCB;

alternatively, M is PEGm, PEG_mIs a single-arm polyethylene glycol chain segment, PEG_mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 5k, and A1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 is

Preferably, M is PEGm, PEG_mIs a single-arm polyethylene glycol chain segment, PEG_mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 5k, and A1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 is Q is-N-AC, N is G, AC is SN 38;

alternatively, M is PEGm, PEG_mIs a multi-arm (e.g., four-arm, eight-arm, preferably four-arm) polyethylene glycol segment, PEG_mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 10k, and A1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 is

Preferably, M is PEGm, PEG_mIs a multi-arm (e.g., four-arm, eight-arm, preferably four-arm) polyethylene glycol segment, PEG_mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 10k, and A1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is LPT, and AC2 is PCB;

or, M isA1 isY isX isW1 isZ3 isZ2 isZ1 isZ0 is

Preferably, M isA1 isY isX isW1 isZ3 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is LPT, and AC2 is PCB;

or, M isA1 isY isX isW1 isZ2 isZ1 isZ0 is

Preferably, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is LPT, and AC2 is PCB;

or, M isA1 isY is a direct bond and X isW1 isZ1 isZ0 is

Preferably, M isA1 isY is a direct bond and X is W1 isZ1 isZ0 isQ is-N-AC, N isAC is PTX;

or, M isA1 isY isX isW1 isZ2 isZ1 isZ0 is

Preferably, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ is-N-AC, N is G, and AC is SN 38.

4. The polyethylene glycol conjugate drug of claim 2, or a pharmaceutically acceptable salt thereof, wherein:

m is-C(＝O)C_1-6Alkylene group C (═ O) -),A1 isY is a direct bond, GLFG, -C (═ O) C_1-6Alkylene C (═ O) -orX is W1 isZ2 isZ1 isZ0 is

Preferably, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB;

or, preferably, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB;

or, preferably, M isA1 isY is GLFG and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB;

or, preferably, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB;

or, preferably, M isA1 isY is GLFG and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB;

Or, preferably, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is LPT;

or, preferably, M isA1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1, AC2 are independently PCB, SB7 or LPT;

or, preferably, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ is-N-AC, N is GFLG, AC is PCB;

or, preferably, M isA1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 isQ is-N-AC, N is GFLG, and AC is NPB.

5. The polyethylene glycol conjugate drug of claim 1, having a structure represented by formula (III), or a pharmaceutically acceptable salt thereof,

wherein:

m is-C(＝O)C_1-6Alkylene NH-or-C (═ O) C_1-6Alkylene C (═ O) -, preferably

A1, A1' are independentlyPreferably, it is

Y, Y' are independently a direct bond or

X and X' are

W1, W1' are independentlyQ1、 —Z0-(01)₂、

Z4, Z3, Z2, Z1 and Z0 are independently Preferably, it is

Q is-N-AC, and Q is-N-AC,

q1 is-N1-AC 1,

q2 is-N2-AC 2,

n, N1 and N2 are independently GFLG, G,

AC. AC1, AC2 are independently SN38, PKA, PCB, PTX, LPT, SB7 or DOX,

The number average molecular weight of the PEG is independently 5k to 40 k.

6. The polyethylene glycol conjugate drug of claim 5, or a pharmaceutically acceptable salt thereof, wherein:

m isA1 isA1' isY is a direct bond and Y' isW1 isW1' isZ4, Z2 and Z0 areZ3 and Z1 are

Preferably, M isA1 isA1' isY is a direct bond and Y' isW1 isW1' isZ4, Z2 and Z0 areZ3 and Z1 areQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 is G, N2 is GFLG, AC1 is SN38, and AC2 is PKA;

or, M is-C (═ O) C_1-6Alkylene NH-, A1 beingA1' isY is a direct bond and Y' isW1 isW1' is Q1, Z2 and Z0 areZ1 is

Preferably, M isA1 isA1' isY is a direct bond and Y' isW1 isW1' is Q1, Z2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, and N1 is G orN2 is GFLG, AC1 is PTX or DOX, AC2 is PCB or LPT;

or, M is-C (═ O) C_1-6Alkylene radicals C (═ O) -, A1 and A1' areY and Y' being a direct bond, W₁Is composed ofW1' is-ZO- (Q1)₂Z3 and Z1 areZ2 and Z0 are

Preferably, M isA1 and A1' areY and Y' are direct bonds, W1 isW1' is-ZO- (Q1)₂Z3 and Z1 areZ2 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB;

or, M is-C (═ O) C_1-6Alkylene radicals C (═ O) -, A1 and A1' are Y and Y' are direct bonds, W1 isW1' is Q1, Z1 and Z0 are

Preferably, M isA1 and A1' areY and Y' are direct bonds, W1 isW1' is Q1, Z1 and Z0 areQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is PCB, and AC2 is LPT;

or, M is-C (═ O) C_1-6Alkylene NH-, A1 beingA1' isY and Y 'are direct bonds, W1 and W1' areZ2 isZ1 isZ0 is

Preferably, M isA1 isA1' isY and Y 'are direct bonds, W1 and W1' areZ2 isZ1 isZ0 isQ is-N-AC, N is GFLG, and AC is SB 7.

7. The polyethylene glycol conjugate drug of claim 1, having a structure represented by formula (IV),

wherein:

m is

A1 isPreferably, it is

X is

W1 is independently

Z4, Z3, Z2, Z1 and Z0 are independentlyPreferably, it is

Q is-N-AC, and Q is-N-AC,

q1 is-N1-AC 1,

q2 is-N2-AC 2,

n1 and N2 are GFLGs,

AC1, AC2 are independently PCB, SB7, LPT, PKA,

the number average molecular weight of the PEG is independently 5k to 40 k.

8. The polyethylene glycol conjugate drug of claim 7, or a pharmaceutically acceptable salt thereof, wherein:

m isW1 isZ2 isZ1 isZ0 is

Preferably, M isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is LPT, and AC2 is SB 7;

Or, M isW1 isZ4 and Z1 areZ3, Z2 and Z0 are

Preferably, M isW1 isZ4 and Z1 areZ3, Z2 and Z0 areQ1 is-N1-AC 1, Q2 is-N2-AC2, N1 and N2 are GFLG, AC1 is PCB, AC2 is PKA;

or, M isW1 isZ1 isZ0 is

Preferably, M isW1 isZ1 isZ0 isQ is-N-AC, N is GFLG, and AC is LPT or PCB.

9. The polyethylene glycol conjugate drug of claim 1, having a structure represented by formula (V), or a pharmaceutically acceptable salt thereof,

wherein:

l1 is-C (═ O) C_1-6Alkylene C (═ O) -, preferably

A2 isPreferably, it is

M is

A1 isPreferably, it is

Y is

X is

W1, W2 are independently

Z2, Z1, Z0 are independentlyPreferably, it is

Q1 is-N1-AC 1,

q2 is-N2-AC 2,

n1 and N2 are GFLGs,

AC1, AC2 are independently PCB or SB7,

the number average molecular weight of PEG is 5k-40 k.

10. The polyethylene glycol conjugate drug of claim 9, or a pharmaceutically acceptable salt thereof, wherein:

w1 and W2 areZ2 isZ1 isZ0 is

Preferably, W1 and W2 areZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is PCB, and AC2 is SB 7.

11. A polyethylene glycol conjugate drug selected from the group consisting of:

12. An intermediate for the preparation of a polyethylene glycol conjugate drug or a pharmaceutically acceptable salt thereof as claimed in any one of claims 2 to 4, said intermediate being selected from the following:

13. a process for preparing a polyethylene glycol conjugate drug or a pharmaceutically acceptable salt thereof according to any one of claims 2 to 4, comprising the steps of:

(1) preparation of intermediatesM, A1, W1, Y and j1 are as defined in any one of claims 2 to 4, wherein:

preparation of intermediatesWhen Y is not a direct bond,a carboxyl group at the end of-Y-OH, when Y is a direct bond,the tail end of the compound has a carboxyl group,

preparation of intermediatesWhen Y is not a direct bond, -Y-H has an amino group at the terminal, and when Y is a direct bond,the tail end of the compound has an amino group;

(2) reacting PEG with amino or activated amino with said intermediateAmidation reaction, or reacting PEG with carboxyl or activated carboxyl with the intermediateCarrying out amidation reaction to obtain the polyethylene glycol coupled drug as claimed in any one of claims 2 to 4.

14. An intermediate for the preparation of a polyethylene glycol conjugate drug or a pharmaceutically acceptable salt thereof according to any one of claims 5 to 6, said intermediate being selected from the group consisting of:

15. a process for preparing a polyethylene glycol conjugate drug or a pharmaceutically acceptable salt thereof according to any one of claims 5 to 6, comprising the steps of:

(1) Preparation of intermediatesM, A1, A1 ', W1, W1 ', Y, Y ' and j1 are as defined in any one of claims 5 to 6, wherein:

when Y or Y 'is not a direct bond, -Y-H or-Y' -H is terminally provided with an amino group,

when Y or Y' is a direct bond,the tail end of the compound has an amino group;

(2) reacting PEG having carboxyl group or activated carboxyl group with the intermediateCarrying out amidation reaction to obtain the polyethylene glycol coupled drug as claimed in any one of claims 5 to 6.

16. An intermediate for the preparation of a polyethylene glycol conjugate drug or a pharmaceutically acceptable salt thereof according to any one of claims 7 to 8, said intermediate being selected from the group consisting of:

17. a process for preparing a polyethylene glycol conjugate drug or a pharmaceutically acceptable salt thereof according to any one of claims 7 to 8, comprising the steps of:

(1) preparation of intermediatesM, A1, W1 and j1 are as defined in any one of claims 7 to 8,the tail end of the compound has an amino group;

(2) reacting PEG having carboxyl group or activated carboxyl group with the intermediateCarrying out amidation reaction to obtain the polyethylene glycol coupled drug as claimed in any one of claims 7 to 8.

18. An intermediate for preparing the polyethylene glycol conjugate drug or the pharmaceutically acceptable salt thereof according to any one of claims 9 to 10, wherein the intermediate is

19. A process for preparing a polyethylene glycol conjugate drug or a pharmaceutically acceptable salt thereof according to any one of claims 9 to 10, comprising the steps of:

(1) preparation of intermediatesM, A1, A2, W1, W2, L1, Y and j1 are as defined in any one of claims 9 to 10, wherein the-Y-OH terminus bears a carboxyl group;

(2) reacting PEG with amino or activated amino with said intermediateAmidation to obtain the polyethylene glycol conjugate drug of any one of claims 9 to 10.

20. A pharmaceutical composition comprising a therapeutically and/or prophylactically effective amount of the polyethylene glycol conjugate drug of any one of claims 1-11, or a pharmaceutically acceptable salt thereof; the composition also contains one or more pharmaceutically acceptable auxiliary materials.

21. Use of a polyethylene glycol conjugate drug according to any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment and/or prevention of a disease for which the active ingredient of the polyethylene glycol conjugate drug is indicated;

preferably, the disease is a cancer selected from the group consisting of: colon cancer, leukemia, lymphoma, bladder cancer, bone cancer, brain tumor, medulloblastoma, glioma, breast cancer, adenoma/carcinoid cancer, adrenocortical cancer, islet cell cancer, cervical cancer, endometrial cancer, ovarian cancer, colorectal cancer, skin cancer, esophageal cancer, eye cancer, gall bladder cancer, gastric cancer, head and neck cancer, liver cancer, melanoma, kaposi's sarcoma, kidney cancer, oral cancer, lung cancer, nasopharyngeal cancer, neuroblastoma, ovarian cancer, pancreatic cancer, thyroid cancer, parathyroid penis cancer, prostate cancer, urinary tract cancer, vaginal cancer, vulvar cancer, anal cancer, sarcoma, and metastases of the cancers.

Technical Field

The invention belongs to the technical field of medicines, and relates to a polyethylene glycol coupled drug, a preparation method and application thereof.

Background

The pegylation drug has great advantages relative to the original drug, and can increase the water solubility of drug molecules (important for molecules with extremely low solubility such as paclitaxel, camptothecin or platinum); can prevent or reduce drug agglomeration, immunogenicity and antigenicity. Most of small molecule anticancer drugs can only be kept for a few minutes in blood circulation, while the macromolecule coupling anticancer drugs can be kept for dozens, hundreds of hours or even longer, which is beneficial to the effect of 'enhancing penetration and retention', namely EPR effect, generated by the leakage of tumor capillaries; the elimination of the kidney of the medicine is weakened due to the increase of the hydrodynamic volume, the medicine is protected from being degraded by enzyme, the half-life period of the medicine in blood plasma is prolonged, and the bioavailability of the medicine is increased; the anticancer drugs are highly enriched in cancerated organs, tissues or cells through EPR passive targeting or active targeting, so that the toxic and side effects caused by the fact that small-molecule anticancer drugs fill the whole body are greatly reduced; the cell absorption of the drug is limited to an endocytosis path, which is beneficial to the drug transmission to the lysosome direction, thereby avoiding drug resistance caused by pumping out p-glycoprotein; stimulating or restoring immunity, and killing cancer cells.

The polyethylene glycol conjugated drug technology of the united states NEKTAR and ENZON has had great success, and up to now 15 drugs have been approved by the united states FDA for marketing, and in addition 36 new clinical drugs are in clinical phase one, two, three or enter NDA. However, all of the pegylated drugs above are pegylated single drugs.

Chinese patent ZL201510996205.4 discloses that by simultaneously grafting gemcitabine, a chemotherapeutic agent, and AZD7762, an inhibitor of Chk1, onto a four-armed polyethylene glycol carrier, the Chk1 inhibitor has no anticancer effect by itself, but can enhance the effect of the chemotherapeutic agent when used in combination with gemcitabine. Chinese patents ZL201710761441.7 and ZL201710761572.5 disclose that two anticancer drugs are grafted on a grafting site of polyethylene glycol simultaneously, so that different cancer cell biological signal channels and target inhibition are realized, and different treatment modes are freely combined.

Disclosure of Invention

The present invention is directed to solving, at least to some extent, one of the technical problems in the related art. To this end, the present invention proposes a polyethylene glycol conjugate drug having excellent tumor suppressive activity. The preparation method can efficiently and conveniently prepare the polyethylene glycol coupled drug.

Polyethylene glycol medicine

In a first aspect of the invention, the invention provides a polyethylene glycol conjugate drug shown as formula (I) or a pharmaceutically acceptable salt thereof.

The polyethylene glycol conjugate drug of the invention takes amino acid or polypeptide as a connecting chain, takes dicarboxylic acid with amino group (such as natural amino acid with two carboxyl groups) as a connecting bridge, and couples a plurality of same or different drug molecules together. The type, proportion and drug loading of the drug can be adjusted. In formula (I), PEG is a single-arm polyethylene glycol segment having a number average molecular weight of 2k to 40k, such as 5k to 10k or 10k to 40k, such as about 2k, about 5k, about 10k, about 20k, about 30k, or about 40 k. In certain embodiments, PEG reacts with carboxyl groups on the backbone through terminal amino groups to form amide bonds. Alternatively, PEG reacts with an amino group on the main chain via a terminal carboxyl group to form an amide bond. In certain embodiments, the molecular weight of the PEG comprises the amino group at the terminus thereof (i.e., the PEG derivative with the reactive group), and in some embodiments, the molecular weight of the PEG may be 2k to 40k, such as 2k to 3k, 3k to 5k, 5k to 10k, or 10k to 40k, such as about 2k, about 3k, about 5k, about 10k, about 40k, of the polyethylene glycol and X or X' as a whole.

Wherein:

m is-C (═ O) C_1-6Alkylene group C (O) -, -NHC_1-6Alkylene NH-, -C (═ O) C_1-6Alkylene NH-, or PEG_m(ii) a Wherein, PEG_mIs a single-arm or multi-arm (e.g. four-arm, eight-arm, preferably four-arm) polyethylene glycol segment, PEG_mThe number average molecular weight of (a) is 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 5k or 10 k;

a1, A1' are independently

A2 is independently a direct bond or

L1 is independently a direct bond or-C (═ O) C_1-6Alkylene C (═ O) -;

w1, W1' and W2 are independently Q1,

Z4, Z3, Z2, Z1 and Z0 are independently

Q is-N-AC;

q1 is-N1-AC 1;

q2 is-N2-AC 2;

n, N1 and N2 are independently GFLG, G,(preferably, it is

AC. AC1, AC2 are independently drug molecules (e.g. drug molecules with anti-tumor activity), preferably PCB, DOX, LPT, SB7, PKA, SN38, PTX or NPB;

n1, n2 are independently 0, 1, 2, 3, 4, 5 or 6;

y, Y' are independently a direct bond, GLFG, -C (═ O) C_1-6Alkylene group C (═ O) -),

X, X' is independently

PEG is a single-arm polyethylene glycol chain segment, and the number average molecular weight of the PEG is independently 5k-40 k;

j1 is 1, 2, 3, 4, 5 or 6;

j2, j3 are independently 0 or 1.

It is noted that, in some embodiments, PEG_mThe remaining structure of the compound of formula (I) is linked via the carbonyl group.

Alternatively, the polyethylene glycol conjugate drug of the present invention comprises a plurality of the same or different drug molecules coupled together by using an amino acid or polypeptide as a connecting chain and a dicarboxylic acid having an amino group (e.g., a natural amino acid having two carboxyl groups) as a connecting bridge. The type, proportion and drug loading of the drug can be adjusted. In formula (I), PEG is a single-arm polyethylene glycol segment having a number average molecular weight of 2k to 40k, such as 5k to 10k or 10k to 40k, such as about 2k, about 5k, about 10k, about 20k, about 30k, or about 40 k. In certain embodiments, PEG reacts with carboxyl groups on the backbone through terminal amino groups to form amide bonds. Alternatively, PEG reacts with an amino group on the main chain via a terminal carboxyl group to form an amide bond. In certain embodiments, the molecular weight of the PEG comprises the amino group at the terminus thereof (i.e., the PEG derivative with the reactive group), and in some embodiments, the molecular weight of the PEG may be 2k to 40k, such as 2k to 3k, 3k to 5k, 5k to 10k, or 10k to 40k, such as about 2k, about 3k, about 5k, about 10k, about 40k, of the polyethylene glycol and X or X' as a whole.

Wherein:

m is-C (═ O) C_1-6Alkylene group C (O) -, -NHC_1-6Alkylene NH-, -C (═ O) C _1-6Alkylene NH-,

a1, A1' are independently

A2 is independently a direct bond or

L1 is independently a direct bond or-C (═ O) C_1-6Alkylene C (═ O) -;

w1, W1' and W2 are independently Q1,

Z4, Z3, Z2, Z1 and Z0 are independently

Q is-N-AC;

q1 is-N1-AC 1;

q2 is-N2-AC 2;

n, N1 and N2 are independently GFLG, G,

AC. AC1, AC2 are independently drug molecules (e.g. drug molecules with anti-tumor activity), preferably PCB, DOX, LPT, SB7, PKA, SN38 or PTX;

n1, n2 are independently 0, 1, 2, 3, 4, 5 or 6;

y, Y' are independently a direct bond, GLFG, -C (═ O) C_1-6Alkylene group C (═ O) -),

X, X' is independently

PEG is a single-arm polyethylene glycol chain segment, and the number average molecular weight of the PEG is independently 5k-40 k;

j1 is 1, 2, 3, 4, 5 or 6;

j2, j3 are independently 0 or 1.

In some embodiments, the polyethylene glycol conjugate drug has a structure as shown in formula (II), formula (III), formula (IV), or formula (V):

in some embodiments, the polyethylene glycol-conjugated drug has a structure represented by formula (II), wherein:

m is-C (═ O) C_1-6Alkylene group C (═ O) -),-NHC_1-6Alkylene NH-, or PEG_mPreferably, it is Or PEG_mWherein, PEG_mIs a single-arm or multi-arm (e.g. four-arm, eight-arm, preferably four-arm) polyethylene glycol segment, PEG _mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 5k or 10k,

a1 is independently Preferably, it is

Y is independently a direct bond,GLFG、-C(＝O)C_1-6Alkylene group C (═ O) -), Preferably a direct bond,GLFG、

X is independently

W1 is independently

Z4, Z3, Z2, Z1 and Z0 are independently Preferably, it is

Q is-N-AC, and Q is-N-AC,

q1 is-N1-AC 1,

q2 is-N2-AC 2,

n, N1 and N2 are independently G, GFLG,(preferably, it is

AC. AC1, AC2 are independently SN38, PKA, PCB, LPT, SB7, PTX or NPB,

the number average molecular weight of the PEG is independently 5k to 40 k.

Alternatively, in some embodiments, the polyethylene glycol-conjugated drug has a structure represented by formula (II), wherein: m is-C (═ O) C_1-6Alkylene group C (═ O) -),-NHC_1-6Alkylene NH-, preferably, it is

A1 is independentlyPreferably, it is

Y is independently a direct bond, GLFG, -C (═ O) C_1-6Alkylene group C (═ O) -), Preferably a direct bond, GLFG,X is independentlyW1 is independently

Z4, Z3, Z2, Z1 and Z0 are independentlyPreferably, it is

Q is-N-AC, and Q is-N-AC,

q1 is-N1-AC 1,

q2 is-N2-AC 2,

n, N1 and N2 are independently G, GFLG,

AC. AC1, AC2 are independently SN38, PKA, PCB, LPT or SB7,

the number average molecular weight of the PEG is independently 5k to 40 k.

In some embodiments, wherein:

M is-C (═ O) C_1-6Alkylene radical C (O) -, a1 isY isX isW1 isZ4, Z2 and Z1 areZ3 and Z0 are

In some embodiments, M isA1 isY isX isW1 isZ4, Z2 and Z1 areZ3 and Z0 areQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is PCB, and AC2 is PKA.

In some embodiments, M is-NHC_1-6Alkylene NH-, A1 beingY is GLFG and X isW1 isZ2 isZ1 isZ0 is

In some embodiments, M isA1 isY is GLFG and X isW1 isZ2 isZ1 isZ0 isQ is-N-AC, N is G, and AC is SN 38.

In some embodiments, M isA1 isY is a direct bond or-C (═ O) C_1-6Alkylene C (═ O) -, X isW1 isZ1 isZ0 is

In some embodiments, M isA1 isY is a direct bond and X isW1 isZ1 isZ0 isQ is-N-AC, N is GFLG, and AC is LPT.

In some embodiments, M isA1 isY isX isW1 isZ1 isZ0 isQ is-N-AC, N is GFLG, and AC is PCB.

In some embodiments, M isA1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 is

In some embodiments, M isA1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is NPB, and AC2 is PCB.

In some embodiments, M is PEGm, PEG_mIs a single-arm polyethylene glycol chain segment, PEG_mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 5k, and A1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 is

In some embodiments, M is PEGm, PEG_mIs a single-arm polyethylene glycol chain segment, PEG_mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 5k, and A1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 isQ is-N-AC, N is G, and AC is SN 38.

In some embodiments, M is PEGm, PEG_mIs a multi-arm (e.g., four-arm, eight-arm, preferably four-arm) polyethylene glycol segment, PEG_mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 10k, and A1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 is

In some embodiments, M is PEGm, PEG_mIs a multi-arm (e.g., four-arm, eight-arm, preferably four-arm) polyethylene glycol segment, PEG_mHas a number average molecular weight of 5k to 40k, preferably 5k to 10k or 10k to 40k, more preferably 10k, and A1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is LPT, and AC2 is PCB.

In some embodiments, M is A1 isY isX isW1 isZ3 isZ2 isZ1 isZ0 is

In some embodiments, M isA1 isY isX isW1 isZ3 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is LPT, and AC2 is PCB.

In some embodiments, M isA1 isY isX isW1 isZ2 isZ1 isZ0 is

In some embodiments, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is LPT, and AC2 is PCB.

In some embodiments, M isA1 isY is a direct bond and X isW1 isZ1 isZ0 is

In some embodiments, M isA1 isY is a direct bond and X isW1 isZ1 isZ0 isQ is-N-AC, N isAC is PTX.

In some embodiments, M isA1 isY isX isW1 isZ2 isZ1 isZ0 is

In some embodiments, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ is-N-AC, N is G, and AC is SN 38.

In some embodiments, wherein:

m is-C(＝O)C_1-6Alkylene group C (═ O) -),A1 is Y is a direct bond, GLFG, -C (═ O) C_1-6Alkylene C (═ O) -orX isW1 is Z2 isZ1 isZ0 is

Or, in some embodiments, wherein:

m is-C(＝O)C_1-6Alkylene group C (═ O) -),A1 is Y is a direct bond, GLFG, -C (═ O) C_1-6Alkylene C (═ O) -orX isW1 isZ2 isZ1 isZ0 is

In some embodiments, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB.

In some embodiments, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB.

In some embodiments, M isA1 isY is GLFG and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB.

In some embodiments, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB.

In some embodiments, M isA1 isY is GLFG and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB.

In some embodiments, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is LPT.

In some embodiments, M is A1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, and AC1 and AC2 are independently PCB, SB7 or LPT.

In some embodiments, M isA1 isY isX isW1 isZ2 isZ1 isZ0 isQ is-N-AC, N is GFLG, and AC is PCB.

In some embodiments, M isA1 isY is a direct bond and X isW1 isZ2 isZ1 isZ0 isQ is-N-AC, N is GFLG, and AC is NPB.

In some embodiments, the polyethylene glycol-conjugated drug has a structure represented by formula (III), wherein:

m is-C(＝O)C_1-6Alkylene NH-or-C (═ O) C_1-6Alkylene C (═ O) -, preferably

A1, A1' are independentlyPreferably, it is

Y, Y' are independently a direct bond or

X and X' are

W1, W1' are independentlyQ1、

Z4, Z3, Z2, Z1 and Z0 are independently Preferably, it is

Q is-N-AC, and Q is-N-AC,

q1 is-N1-AC 1,

q2 is-N2-AC 2,

n, N1 and N2 are independently GFLG, G,

AC. AC1, AC2 are independently SN38, PKA, PCB, PTX, LPT, SB7 or DOX,

the number average molecular weight of the PEG is independently 5k to 40 k.

Alternatively, in some embodiments, the polyethylene glycol-conjugated drug has a structure represented by formula (III), wherein: m is-C(＝O)C_1-6Alkylene NH-or-C (═ O) C_1-6Alkylene C (═ O) -, preferably

A1, A1' are independentlyPreferably, it is

Y, Y' are independently a direct bond or

X and X' are

W1 and W1' are independently Q1,

Z4, Z3, Z2, Z1 and Z0 are independently Preferably, it is

Q1 is-N1-AC 1,

q2 is-N2-AC 2,

n1 and N2 are independently GFLG, G,

AC1, AC2 are independently SN38, PKA, PCB, PTX, LPT, SB7 or DOX,

the number average molecular weight of the PEG is independently 5k to 40 k.

In some embodiments, wherein:

m isA1 isA1' isY is a direct bond and Y' isW1 isW1' isZ4, Z2 and Z0 areZ3 and Z1 are

In some embodiments, M isA1 isA1' isY is a direct bond and Y' isW1 isW1' isZ4, Z2 and Z0 areZ3 and Z1 areQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 is G, N2 is GFLG, AC1 is SN38, and AC2 is PKA.

In some embodiments, M is-C(＝O)C_1-6Alkylene NH-, A1 beingA1' isY is a direct bond and Y' isW1 isW1' is Q1, Z2 and Z0 areZ1 is

In some embodiments, M isA1 isA1' isY is a direct bond and Y' isW1 isW1' is Q1, Z2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, and N1 is G orN2 is GFLG, AC1 is PTX or DOX, and AC2 is PCB or LPT.

In some embodiments, M is-C (═ O) C_1-6Alkylene radicals C (═ O) -, A1 and A1' are Y and Y' are direct bonds, W1 isW1' isZ3 and Z1 areZ2 and Z0 are

In some embodiments, M isA1 and A1' areY and Y' are direct bonds, W1 isW1' isZ3 and Z1 areZ2 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is SB7, and AC2 is PCB.

In some embodiments, M is-C (═ O) C_1-6Alkylene radicals C (═ O) -, A1 and A1' areY and Y' are direct bonds, W1 isW1' is Q1, Z1 and Z0 are

In some embodiments, M isA1 and A1' areY and Y' are direct bonds, W1 isW1' is Q1, Z1 and Z0 areQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is PCB, and AC2 is LPT.

In some embodiments, M is-C (═ O) C_1-6Alkylene NH-, A1 beingA1' isY and Y 'are direct bonds, W1 and W1' areZ2 isZ1 isZ0 is

In some embodiments, M isA1 isA1' isY and Y 'are direct bonds, W1 and W1' areZ2 isZ1 isZ0 isQ is-N-AC, N is GFLG, and AC is SB 7.

In some embodiments, the polyethylene glycol-conjugated drug has a structure represented by formula (IV), wherein:

m is

A1 isPreferably, it is

X is

W1 is independently

Z4, Z3, Z2, Z1 and Z0 are independentlyPreferably, it is

Q is-N-AC, and Q is-N-AC,

q1 is-N1-AC 1,

Q2 is-N2-AC 2,

n1 and N2 are GFLGs,

AC1, AC2 are independently PCB, SB7, LPT, PKA,

the number average molecular weight of the PEG is independently 5k to 40 k.

In some embodiments, wherein:

m isW1 isZ2 isZ1 isZ0 is

In some embodiments, M isW1 isZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is LPT, and AC2 is SB 7.

In some embodiments, M isW1 isZ4 and Z1 areZ3, Z2 and Z0 are

In some embodiments, M isW1 isZ4 and Z1 areZ3, Z2 and Z0 areQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is PCB, and AC2 is PKA.

In some embodiments, M isW1 isZ1 isZ0 isIn some embodiments, M isW1 isZ1 isZ0 isQ is-N-AC, N is GFLG, and AC is LPT or PCB. In some embodiments, the polyethylene glycol-conjugated drug has a structure represented by formula (V), wherein:

l1 is-C (═ O) C_1-6Alkylene C (═ O) -, preferably

A2 isPreferably, it is

M is

A1 isPreferably, it is

Y is

X is

W1, W2 are independently

Z2, Z1, Z0 are independentlyPreferably, it is

Q1 is-N1-AC 1,

q2 is-N2-AC 2,

n1 and N2 are GFLGs,

AC1, AC2 are independently PCB or SB7,

the number average molecular weight of PEG is 5k-40 k.

In some embodiments, wherein:

w1 and W2 arez2 isZ1 isZ0 is

In some embodiments, W1 and W2 areZ2 isZ1 isZ0 isQ1 is-N1-AC 1, Q2 is-N2-AC 2, N1 and N2 are GFLG, AC1 is PCB, and AC2 is SB 7.

In a second aspect of the invention, the invention provides a polyethylene glycol conjugate drug, or a pharmaceutically acceptable salt thereof, selected from the group consisting of:

intermediates and methods of preparation

In a third aspect of the invention, the invention provides an intermediate for the preparation of a polyethylene glycol conjugate drug of the aforementioned formula (II) or a pharmaceutically acceptable salt thereof,

the intermediate is selected from the following:

in a fourth aspect of the present invention, the present invention provides a process for preparing a polyethylene glycol conjugate drug of the aforementioned formula (II) or a pharmaceutically acceptable salt thereof, comprising the steps of:

(1) preparation of intermediatesM, A1, W1, Y and j1 are as previously defined, wherein:

preparation of intermediatesWhen Y is not a direct bond, the-Y-OH terminus has a carboxyl group, and when Y is a direct bond,the tail end of the compound has a carboxyl group,

preparation of intermediatesWhen Y is not a direct bond, -Y-H has an amino group at the terminal, and when Y is a direct bond,the terminal has carboxyl;

(2) reacting PEG with amino or activated amino with said intermediate Amidation reaction, or reacting PEG with carboxyl or activated carboxyl with the intermediateAmidation reaction to obtain the said polyglycol conjugated medicine.

In a fifth aspect of the invention, the invention provides intermediates useful in the preparation of the foregoing polyethylene glycol conjugate drug of formula (III), or a pharmaceutically acceptable salt thereof,

the intermediate is selected from the following:

in a sixth aspect of the present invention, the present invention provides a process for preparing a polyethylene glycol conjugate drug of formula (III) as described above, or a pharmaceutically acceptable salt thereof, comprising the steps of:

(1) preparation of intermediatesM, A1, A1 ', W1, W1 ', Y, Y ' and j1 are as previously defined, wherein:

when Y or Y 'is not a direct bond, -Y-H or-Y' -H is terminally provided with an amino group,

when Y or Y' is a direct bond,the tail end of the compound has an amino group;

(2) reacting PEG having carboxyl group or activated carboxyl group with the intermediateAmidation reaction to obtain the said polyglycol conjugated medicine.

In a seventh aspect of the invention, the invention provides an intermediate for the preparation of a polyethylene glycol conjugate drug of formula (IV) as hereinbefore described or a pharmaceutically acceptable salt thereof,

the intermediate is selected from the following:

In an eighth aspect of the present invention, there is provided a process for preparing a polyethylene glycol conjugate drug of formula (IV) as described above, or a pharmaceutically acceptable salt thereof, comprising the steps of:

(1) preparation of intermediatesM, A1, W1, and j1 are as previously defined, wherein,the tail end of the compound has an amino group;

(2) reacting PEG having carboxyl group or activated carboxyl group with the intermediateAmidation reaction to obtain the said polyglycol conjugated medicine.

In a ninth aspect of the invention, the invention provides an intermediate for the preparation of a polyethylene glycol conjugate drug of formula (V) as hereinbefore described or a pharmaceutically acceptable salt thereof,

the intermediate is

In a tenth aspect of the present invention, the present invention provides a process for preparing a polyethylene glycol conjugate drug of formula (V) as described above, or a pharmaceutically acceptable salt thereof, comprising the steps of:

(1) preparation of intermediatesM, A1, A2, W1, W2, L1, Y and j1 are as previously defined, wherein the-Y-OH terminus bears a carboxyl group;

(2) reacting PEG with amino or activated amino with said intermediateAmidation reaction to obtain the said polyglycol conjugated medicine.

Pharmaceutical composition and pharmaceutical use

In one aspect of the present invention, the present application provides a pharmaceutical composition comprising a therapeutically and/or prophylactically effective amount of a polyethylene glycol conjugate drug of the present invention, or a pharmaceutically acceptable salt thereof; the composition further comprises one or more pharmaceutically acceptable adjuvants, such as carriers and/or excipients. Such carriers and/or excipients include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins such as human serum albumin, buffer substances such as phosphates, glycerol, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulosic substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, beeswax, polyethylene-polyoxypropylene block polymers and lanolin.

The pharmaceutical composition can be prepared into any pharmaceutically acceptable dosage form. The pharmaceutical composition may also be administered to an individual in need of such treatment by any suitable administration, e.g., oral, parenteral, rectal or pulmonary administration. For oral administration, the pharmaceutical composition can be made into conventional solid preparations such as tablet, capsule, pill, granule, etc.; it can also be made into oral liquid, such as oral solution, oral suspension, syrup, etc. When the composition is formulated into oral preparations, appropriate filler, binder, disintegrating agent, lubricant, etc. can be added. For parenteral administration, the pharmaceutical composition can be prepared into injections, including injection solutions, sterile powders for injection, and concentrated solutions for injection. The injection can be prepared by conventional method in the existing pharmaceutical field, and can be prepared without adding additives or adding appropriate additives according to the properties of the medicine. For rectal administration, the pharmaceutical composition may be formulated as a suppository or the like. For pulmonary administration, the pharmaceutical composition may be formulated as an inhalant or a spray. Preferably, the pharmaceutical composition of the present invention can be prepared into injections, such as injection solutions. Optionally, physiological saline is used as a carrier of the injection.

In another aspect, the present application provides the use of a polyethylene glycol conjugate drug of the present invention, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment and/or prevention of a disease (e.g., cancer) to be treated by the active ingredient in the polyethylene glycol conjugate drug.

In another aspect, the present application provides a polyethylene glycol conjugate drug of the present invention, or a pharmaceutically acceptable salt thereof, for use in the treatment and/or prevention of a disease (e.g., cancer) for which the active ingredient in the polyethylene glycol conjugate drug is indicated.

In the present invention, cancer refers to a cell proliferative disease state, including but not limited to: colon cancer, leukemia, lymphoma, bladder cancer, bone cancer, brain tumor, medulloblastoma, glioma, breast cancer, adenoma/carcinoid cancer, adrenocortical cancer, islet cell cancer, cervical cancer, endometrial cancer, ovarian cancer, colorectal cancer, skin cancer, esophageal cancer, eye cancer, gall bladder cancer, gastric cancer, head and neck cancer, liver cancer, melanoma, kaposi's sarcoma, kidney cancer, oral cancer, lung cancer, nasopharyngeal cancer, neuroblastoma, ovarian cancer, pancreatic cancer, thyroid cancer, parathyroid penis cancer, prostate cancer, urinary tract cancer, vaginal cancer, vulvar cancer, anal cancer, sarcoma, and the like, including metastases of the foregoing cancers.

In another aspect, the present application provides a method for treating and/or preventing a disease (e.g., cancer) comprising administering to a subject in need thereof an effective amount of a polyethylene glycol conjugate drug of the present invention or a pharmaceutically acceptable salt thereof. The dosing regimen may be adjusted to provide the best desired response. For example, a single bolus may be administered, several divided doses may be administered over time, or the dose may be proportionally reduced or increased as indicated by the exigencies of the therapeutic situation. It is noted that dosage values may vary with the type and severity of the condition being alleviated, and may include single or multiple doses. It is further understood that for any particular individual, the specific dosage regimen will be adjusted over time according to the individual need and the professional judgment of the person administering the composition or supervising the administration of the composition.

In the present invention, "individual" includes a human or a non-human animal. Exemplary human individuals include human individuals (referred to as patients) having a disease (e.g., a disease described herein) or normal individuals. In the present invention, "non-human animal" includes all vertebrates, such as non-mammals (e.g., birds, amphibians, reptiles) and mammals, such as non-human primates, livestock, and/or domesticated animals (e.g., sheep, dogs, cats, cows, pigs, etc.).

Term interpretation or definition

In the present invention, the active ingredient suitable for coupling with polyethylene glycol may be a drug molecule having at least one amino, hydroxyl, carboxyl or acyl group, for example, a drug molecule having anti-tumor activity having at least one amino, hydroxyl, carboxyl or acyl group, such as MK2, LPT, PCB, SB7, PKI, NPB, etc., which represent the following meanings:

unless defined otherwise below, all technical and scientific terms used herein are intended to have the same meaning as commonly understood by one of ordinary skill in the art. Reference to the techniques used herein is intended to refer to those techniques commonly understood in the art, including those variations of or alternatives to those techniques that would be apparent to those skilled in the art. While the following terms are believed to be well understood by those skilled in the art, the following definitions are set forth to better explain the present invention.

As used herein, "PEG" is an abbreviation for polyethylene glycol, meaning that the repeating unit is-CH₂CH₂Homopolymers of O-include single-arm polyethylene glycol, multi-arm polyethylene glycol and derivatives thereof, such as derivatives with reactive functional groups such as amino or carboxyl at the end group. In the present invention, each arm of the multi-arm polyethylene glycol preferably has the same degree of polymerization. When referring to the molecular weight of a multi-armed polyethylene glycol, the molecular weight means the total molecular weight of each arm. In the structural formula of the invention, the letter m or n marked below the repeating unit of the polyethylene glycol represents the polymerization degree of the polyethylene glycol, and when the polyethylene glycol is multi-arm polyethylene glycol, the letter m or n represents the polymerization degree of each arm.

In the various parts of this specification, substituents of the disclosed compounds are disclosed in terms of group type or range. It is specifically intended that the invention includes each and every independent subcombination of the various members of these groups and ranges. For example, the term "C₁-C₆Alkyl "means in particular independently disclosed methyl, ethyl, C₃Alkyl radical, C₄Alkyl radical, C₅Alkyl and C₆An alkyl group.

Unless otherwise indicated, the term "alkyl" denotes 1 to 6 carbon atoms (C)₁-C₆Alkyl), or 1 to 4 carbon atoms (C)₁-C₄Alkyl), or 1 to 3 carbon atoms (C)₁-C₃Alkyl) wherein the alkyl may be independently and optionally substituted with one or more substituents described herein, including, but not limited to, deuterium, amino, hydroxy, cyano, F, Cl, Br, I, mercapto, nitro, oxo (═ O), and the like. Examples of alkyl groups include, but are not limited to, methyl (Me, -CH)₃) Ethyl (Et-CH)₂CH₃) N-propyl (n-Pr, -CH)₂CH₂CH₃) Isopropyl (i-Pr, -CH (CH)₃)₂) N-butyl (n-Bu, -CH)₂CH₂CH₂CH₃) Isobutyl (i-Bu, -CH)₂CH(CH₃)₂) Sec-butyl (s-Bu, -CH (CH)₃)CH₂CH₃) T-butyl (t-Bu, -C (CH)₃)₃) N-pentyl (-CH)₂CH₂CH₂CH₂CH₃) 2-pentyl (-CH (CH)₃)CH₂CH₂CH₃) 3-pentyl (-CH (CH)₂CH₃)₂) 2-methyl-2-butyl (-C (CH) ₃)₂CH₂CH₃) 3-methyl-2-butyl (-CH (CH)₃)CH(CH₃)₂) 3-methyl-1-butyl (-CH)₂CH₂CH(CH₃)₂) 2-methyl-1-butyl (-CH)₂CH(CH₃)CH₂CH₃) N-hexyl (-CH)₂CH₂CH₂CH₂CH₂CH₃) 2-hexyl (-CH (CH)₃)CH₂CH₂CH₂CH₃) 3-hexyl (-CH (CH)₂CH₃)(CH₂CH₂CH₃) 2-methyl-2-pentyl (-C (CH))₃)₂CH₂CH₂CH₃) 3-methyl-2-pentyl (-CH (CH)₃)CH(CH₃)CH₂CH₃) 4-methyl-2-pentyl (-CH (CH)₃)CH₂CH(CH₃)₂) 3-methyl-3-pentyl (-C (CH)₃)(CH₂CH₃)₂) 2-methyl-3-pentyl (-CH (CH)₂CH₃)CH(CH₃)₂) 2, 3-dimethyl-2-butyl (-C (CH)₃)₂CH(CH₃)₂) 3, 3-dimethyl-2-butyl (-CH (CH)₃)C(CH₃)₃) N-heptyl, n-octyl, and the like. The term "alkyl" and its prefix "alkane" as used herein, both include straight and branched saturated carbon chains.

The term "alkylene" denotes a saturated divalent hydrocarbon radical derived from a straight or branched chain saturated alkyl radical by the removal of two hydrogen atoms; for example, "C₁-C₆Alkylene "is derived from C₁-C₆Alkyl radical, "C₁-C₄Alkylene "is derived from C₁-C₄Alkyl radical, "C₁-C₃Alkylene group "Derived from C₁-C₃An alkyl group. And the alkylene group may be substituted or unsubstituted, wherein the substituent may be, but is not limited to, deuterium, hydroxyl, amino, halogen, cyano, aryl, heteroaryl, alkoxy, alkyl, alkenyl, alkynyl, heterocyclic, mercapto, nitro, or aryloxy. Examples include, but are not limited to, methylene (-CH)₂-) ethylene (-CH ₂-CH₂-) isopropylidene (-CH₂-CH(CH₃) -), ethane-1, 1-diyl, 2-methoxypropane-1, 1-diyl, 2-hydroxypropane-1, 1-diyl, 2-methyl-2-hydroxypropane-1, 1-diyl, and the like. Wherein the meaning of alkyl is as described above.

In addition, unless otherwise expressly indicated, the descriptions "… independently for each … and" … and … independently for each … and … "used throughout this document are interchangeable and should be construed broadly to mean that particular items expressed between the same symbols in different groups do not affect each other, or that particular items expressed between the same symbols in the same groups do not affect each other.

As used herein, the variable being a "direct bond" means that the linking group is absent, with a corresponding absence of substituents for the linking group. For example, in-X-Y-Z, if Y is a direct bond, it is denoted as-X-Z, and accordingly, the substituents on Y are simultaneously absent.

As used herein, "pharmaceutically acceptable salts" of the compounds of the present invention include acid addition salts and base addition salts of the compounds. Such as hydrochloride, hexafluorophosphate, meglumine salts and the like.

As used herein, wavy lines in a structural formulaMeaning the position at which other groups are bonded to the structure represented by the structural formula.

As used herein, the term "effective amount" refers to an amount of a compound that, when administered, will alleviate one or more symptoms of the condition being treated to some extent.

As used herein, the term "treating" means reversing, alleviating, inhibiting the progression of, or preventing such a disorder or condition, or one or more symptoms of such a disorder or condition, to which such term applies.

Advantageous effects

The polyethylene glycol coupled drug has excellent tumor inhibition activity. The preparation method can efficiently and conveniently prepare the polyethylene glycol coupled drug.

Drawings

FIG. 1 shows the drug configuration of drugs 44-2 and 27-134;

FIG. 2 shows the absorbance measurements for drugs 44-2 and 27-134;

FIG. 3 shows the inhibitory effect of drug 44-2 on MDA-MB-231 cancer cells, wherein inhibition is indicated by inhibition and concentration is indicated by concentration;

FIG. 4 shows the effect of drugs 27-134 on inhibiting Colo205 cancer cells, wherein inhibition is indicated by inhibition rate and concentration is indicated by concentration;

FIG. 5 shows the IC's of drugs 44-2 and 27-134₅₀Calculating a result;

FIG. 6, FIG. 7 and FIG. 8 show the growth tendency of tumors in each group in example 3;

FIG. 9, FIG. 10 and FIG. 11 are graphs showing the tumor weight inhibition ratios of the respective groups in example 3;

FIG. 12 is a photograph of groups of euthanized animals from example 3;

FIG. 13 is a photograph of the tumors of each group in example 3;

FIG. 14 shows the growth trend of tumors in each group in example 4;

FIG. 15 is a graph showing the inhibition ratios of tumor weights in each group in example 4.

Detailed Description

Embodiments of the present invention will be described in detail below with reference to examples, but those skilled in the art will appreciate that the following examples are only illustrative of the present invention and should not be construed as limiting the scope of the present invention. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products commercially available.

The meanings of the abbreviations in the examples are as follows:

the sources and structures of part of the raw materials are as follows:

M-NH₂-2K.HCl

bond Kai, mPEG-CH₂CH₂-NH₂HCl

M-NH₂-3K.HCl

Bond Kai, mPEG-CH₂CH₂-NH₂CHl

M-NH₂-5K.HCl

Bond Kai, mPEG-CH₂CH₂-NH₂CHl

M-SCM-10K

The key of the key is a key of the key,

synthesis of the Compound of example 1

1. Synthesis of 37-14 (Compound No. 10)

The route is as follows:

The method comprises the following specific steps:

25-71

Boc-Glu-OH (ex Ark Pharm, 15.0g, 60.6673mmol), HBTU (ex Aladdin, 69.0225g, 182.0022mmol), HOBT (ex Innochem, 24.5921g, 182.0022mmol) and H-Glu (OBn)₂TosOH (available from Ark Pharm, 63.6473g, 127.4014mmol) was added to a 1000mL round-bottomed flask, then DMF (300mL) was added to dissolve it, the reaction flask was left to stir at-5 ℃ for about 30 minutes, then DIEA (90.2mL, 546.0066mmol) was added slowly dropwise, after which the reaction flask was allowed to stir at-5 ℃ for 1 hour and finally at room temperature overnight. After completion of the reaction, the reaction solution was transferred to a 2L separatory funnel, and a saturated sodium bicarbonate solution (400mL) and ethyl acetate (300mL) were added thereto, shaken, allowed to stand, extracted, and the aqueous phase and the organic phase were separated. Then, a saturated sodium chloride solution (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous phase and the organic phase were separated. Then, saturated sodium chloride (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous and organic phases were separated. Finally, the organic phase is concentrated and evaporated to dryness and is placed in an oven for drying. Obtaining products 25-71: 67.9 g.

25-73

25-71(52.5355g, 60.6673mmol) was charged into a 500mL round bottom flask, dichloromethane (10mL) was added to dissolve, trifluoroacetic acid (TFA, 67.6mL, 910.0101mmol) was added with stirring, and the reaction was left to stir at room temperature overnight. After the reaction, the reaction mixture was concentrated under reduced pressure and evaporated to dryness to remove methylene chloride. The reaction solution was transferred to a 2L separatory funnel, to which a saturated sodium bicarbonate solution (400mL) and ethyl acetate (300mL) were added, shaken, allowed to stand, extracted, and the aqueous phase and the organic phase were separated. Subsequently, a saturated sodium bicarbonate solution (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous and organic phases were separated. Then, deionized water (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous and organic phases were separated. Finally, the organic phase is concentrated and evaporated to dryness. Dissolving the organic phase with 20% methanol/dichloromethane mixed solvent (100mL), adding 150mL silica gel powder, evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (60% -100% ethyl acetate: 40% -0% petroleum ether and 1% -4% methanol: 99% -96% ethyl acetate), collecting liquid, concentrating, evaporating, and drying. Obtaining products 25-73: 44.4g, yield: 95.56 percent.

16-34

Weighing 2- (-2 aminoethoxy) ethanol (18.8680g,190.2226mmol) and pouring into a 500mL round bottom flask, diluting with dichloromethane (100mL), adding triethylamine (38.4972mL,380.4452mmol), and slowly adding (Boc) while stirring₂O (49.8261g,228.2671mmol), and the reaction was stirred at ordinary temperature. After the reaction is finished, evaporating the reaction solution to dryness, adding sodium bicarbonate powder, diluting with dichloromethane, adding silica gel powder, evaporating to dryness, loading by a dry method, and carrying out column chromatography. Elution with 50% ethyl acetate/petroleum ether gave 27.3g of product in 70% yield.

16-36

Weighing 16-24(27.3g,132.8144mmol), adding into a 500mL reaction flask, introducing nitrogen for protection, adding a THF solution of potassium tert-butoxide, placing the reaction at 0 ℃, adding ethyl bromoacetate (17.6265mL,159.3773mmol), stirring for three hours, and moving to room temperature for reaction. And (3) after the reaction is finished, evaporating the reaction solution to dryness, adding deionized water and ethyl acetate, separating an organic phase, extracting a water phase by using ethyl acetate until no product exists, combining the organic phase, drying by using anhydrous sodium sulfate powder, carrying out suction filtration, carrying out dry-method sample loading on the filtrate, and carrying out column chromatography. Gradient elution is carried out by using 30 percent to 100 percent ethyl acetate/petroleum ether to obtain 20g of product with the yield of 52 percent.

24-36

16-36(17.9g,61.4402mmol) was weighed into a 250mL reaction flask, 1, 4-dioxane was added, lithium hydroxide (3.2386g,135.1685mmol) was added with stirring, and after 30 minutes deionized water was added to clarify. At the end of the reaction, with methyl tert-butyl ether: the reaction solution was extracted 3 times (100mL × 3) with n-hexane 1: 1. The aqueous phase was adjusted to PH 1 with concentrated hydrochloric acid, extracted three times with ethyl acetate (300mL × 3), the ethyl acetate phases combined, washed 3 times with saturated sodium chloride (100mL × 3), concentrated, dry loaded, column chromatographed, eluted with 40% -100% ethyl acetate/petroleum ether to give 10.1g, 62% yield.

25-75

Boc-LC-OH (synthesized by 24-36, 15.0236g, 57.0608mmol), HBTU (32.4596g, 85.5912mmol), HOBT (11.5651g, 85.5912mmol) and 25-73(43.7g, 57.0608mmol) were added to a 500mL round bottom flask, dissolved in DMF (150mL), stirred for about 30 minutes with the reaction flask at-5 ℃ and then DIEA (61.5mL, 342.3648mmol) was slowly added dropwise, after which the reaction flask was allowed to continue to stir at-5 ℃ for 2 hours and then transferred to room temperature overnight. After the reaction was completed, the reaction solution was transferred to a 2L separatory funnel, and a saturated sodium bicarbonate solution (400mL) and ethyl acetate (300mL) were added thereto, shaken, allowed to stand, extracted, and the aqueous phase and the organic phase were separated. Then, a saturated sodium chloride solution (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous phase and the organic phase were separated. Then, deionized water (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous and organic phases were separated. Finally, the organic phase is concentrated and evaporated to dryness. Dissolving the organic phase with 20% methanol/dichloromethane mixed solvent (100mL), adding 150mL silica gel powder, evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (1% ammonia water: 1% -2% methanol: 98% -97% dichloromethane), collecting liquid, concentrating and evaporating to dryness. Obtaining products 25-75: 42.1g, yield: 72.97 percent.

22-181

25-75(26.7g, 26.4064mmol) was placed in a 500mL round-bottom flask, dichloromethane (10mL) was added and dissolved, trifluoroacetic acid (TFA, 19.6mL, 264.064mmol) was added with stirring, and the reaction flask was left to stir at room temperature overnight. After the reaction, the reaction mixture was concentrated under reduced pressure and evaporated to dryness to remove methylene chloride. Then, the reaction solution was transferred to a 2L separatory funnel, and a saturated sodium bicarbonate solution (400mL) and ethyl acetate (300mL) were added thereto, shaken, allowed to stand, extracted, and the aqueous phase and the organic phase were separated. Then, a saturated sodium chloride solution (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous phase and the organic phase were separated. Then, deionized water (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous and organic phases were separated. Finally, the organic phase is concentrated and evaporated to dryness and is placed in an oven for drying. Obtaining products 22-181: 20.3g, yield: 84.2 percent.

35-3

Fmoc-L-Lys (Boc) -OH (purchased from Accela, 1.39g, 2.967mmol), 22-181(2.70g, 2.967mmol), HBTU (1.69g, 4.4505mmol), HOBT (0.6g, 4.4505mmol) were added to a 100mL flask, the reaction was stirred at-5 ℃ for about 30 minutes, DIEA (2.21mL, 13.3515mmol) was slowly added dropwise, the reaction was allowed to react at-5 ℃ for 1 hour after the addition was complete, and then the reaction was allowed to stir at room temperature overnight. After the reaction, the reaction solution was transferred to a 1L separatory funnel, and saturated saline (200mL) and ethyl acetate (250mL) were added to extract the mixture to obtain an organic phase, the aqueous phase was washed with ethyl acetate (200 mL. times.1), the organic phases were combined, the organic phase was washed with saturated saline (200 mL. times.1), concentrated and evaporated to dryness to obtain 35-3:4.0g of a product.

35-4

35-3(4.04g, 2.967mmol) was charged into a 250mL round-bottom flask, dichloromethane (10mL) was added to dissolve it, then trifluoroacetic acid (3.31mL, 44.505mmol) was added with stirring, and finally the reaction was left to stir at room temperature overnight. After the reaction, the reaction mixture was concentrated to a small amount. Adding n-hexane (120mL), stirring and washing to remove trifluoroacetic acid, pouring out supernatant, adding n-hexane (120mL) into lower oily product, repeating the steps for 3 times to obtain oily product, and drying to obtain 35-4: 3.74 g.

25-102

Boc-GFLG-OBn (synthesized according to the literature, 6.8282g, 11.7184mmol) and 10% Pd/C (50mg) were charged into a hydrogenation reactor, DMF (30mL) was added to dissolve the Boc-GFLG-OBn, the hydrogenation reactor was closed, hydrogen was then introduced to make the pressure in the reactor 1.6MPa, and finally the hydrogenation reactor was left to stir at room temperature overnight. After the reaction is finished, the reaction kettle is taken out, the reaction solution is uniformly dripped into a sand core funnel filled with compacted diatomite, the filtration is carried out, and the diatomite is cleaned by DMF (60mL) until the diatomite does not contain the product, thus obtaining the reaction product solution.

35-6

35-4(3.74g, 32.967mmol), 25-102(1.75g, 3.560mmol), HBTU (1.69g, 4.451mmol) and HOBT (0.6g, 4.451mmol) were added to a 250mL flask, 30mL DMF was added to dissolve the mixture, the mixture was stirred at-5 ℃ for 30min, DIEA (4.4mL, 26.703mmol) was slowly added dropwise, the reaction was continued for 1 hour after the dropwise addition was completed, and then the mixture was stirred at room temperature overnight. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding deionized water (200mL) and ethyl acetate (200mL) for extraction to obtain an organic phase, washing the water phase with ethyl acetate (200mL x 2), combining the organic phases, washing the organic phase with saturated saline (200mL x 2), evaporating to dryness, and drying in an oven to obtain a product 35-6: 5.1 g.

35-7

Putting 35-6(5.1g, 2.967mmol) into a 250mL flask, adding DMF (20mL) to dissolve the DMF, adding morpholine (3.9mL, 44.505mmol), stirring at room temperature for 1 hour, finishing the reaction, transferring the reaction solution into a 1L separating funnel, adding saturated saline (200mL) and ethyl acetate (200mL) to extract to obtain an organic phase, washing an aqueous phase with ethyl acetate (200mL x 2), combining the organic phases, washing the organic phase with saturated saline (200mL x 1), concentrating and evaporating to dryness, adding silica gel powder (30g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a mixed solution of 1% ammonia water and 3% -5% methanol in dichloromethane, collecting and concentrating, evaporating to dryness, and drying in an oven to obtain 35-7:3.4g of a product, wherein the yield is 77%.

35-11

35-7(3.4g, 2.27mmol), succinic acid (0.12g, 1.03mmol, from ImmunoKat), HBTU (1.17g, 3.09mmol), HOBT (0.42g, 3.09mmol) were added to a 500mL flask, the reaction was left to stir at-5 ℃ for about 30 minutes, DIEA (1.53mL, 9.27mmol) was slowly added dropwise, the reaction was allowed to react at-5 ℃ for 1 hour after completion of the addition, and the reaction was allowed to stir at room temperature overnight. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding saturated sodium bicarbonate (200mL) and ethyl acetate (250mL) for extraction to obtain an organic phase, washing a water phase with ethyl acetate (200mL x 3), combining the organic phases, washing the organic phase with saturated saline (200mL x 2), concentrating and evaporating to dryness, adding silica gel powder (30g), evaporating to dryness to obtain a powdery solid, loading by a dry method, carrying out column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 5% methanol, collecting, concentrating, evaporating to dryness, and drying by an oven to obtain a product 35-11:3.0g, wherein the yield is 96%.

¹H-NMR(400MHz,DMSO-d₆)δ8.59–8.43(m,1H),8.36–8.26(m,2H),8.20–8.14(m,1H),8.01–7.87(m,6H),7.78–7.61(m,3H),7.38–7.11(m,55H),6.93-6.95(m,2H),5.21–4.97(m,15H),4.53-4.52(m,1H),4.42–4.22(m,7H),4.15-4.13(m,1H),3.89-3.87(m,3H),3.76–3.43(m,19H),3.18-3.16(m,5H),2.31-2.98(m,7H),2.84–2.64(m,6H),2.44–2.38(m,8H),2.37–2.28(m,8H),2.21–2.10(m,5H),2.08–1.81(m,11H),1.81–1.68(m,3H),1.51–1.40(m,4H),1.50–1.41(m,7H),1.29–1.16(m,18H),1.28–1.16(m,12H),0.93–0.75(m,12H).

30-30

Boc-GFLG-OH (synthesized by 25-102, 13.97mmol), PCB (5g, 11.17mmol), HBTU (6.35g, 16.76mmol) and HOBT (2.26g, 16.76mmol) were placed in a 500mL round-bottomed flask and then DMF (100mL) was added to dissolve it and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (8.31mL, 50.28mmol) was then added slowly dropwise, and after the addition was complete, the reaction was allowed to proceed at low temperature for 2 hours, after which the reaction apparatus was allowed to stand at room temperature and stirred overnight. After the reaction was completed, deionized water (1000mL) was added to wash the filter cake, and a yellow solid was precipitated and dried to give 10.3g of the product.

30-33

30-30(10.3g, 11.17mmol) was placed in a 500mL round-bottom flask, dissolved in dichloromethane (50mL), followed by addition of TFA (12.45mL, 167.55mmol), and the reaction was stirred at room temperature overnight with the reaction stopped, the reaction solution was concentrated under reduced pressure, n-hexane (50mL) and methyl t-butyl ether (400mL) were added for precipitation, which was repeated 3 times, and the solid product was filtered to obtain a solid, which was dissolved in dichloromethane and methanol, dry-loaded, column-chromatographed, eluted with a mixed solution of 5% methanol and 1% aqueous ammonia in dichloromethane, collected, concentrated, and dried to obtain 8.6g of a product with a yield of 94%.

28-258

Fmoc-Glu-OtBu (3.223g, 7.5915mmol, available from Ark pharm), 30-33 (synthesized by 30-33) (5.2g, 6.3263mmol), HBTU (3.5988g, 9.4895mmol), HOBT (1.282g, 9.4895mmol) were placed in a 500mL flask, stirred for 10 min, DIEA (4.7mL, 28.4684mmol) was slowly added dropwise, and the reaction was stirred at 0 ℃ overnight. After the reaction is finished, adding n-hexane (300mL) and methyl tert-butyl ether (50mL), separating layers, pouring the upper layer, adding n-hexane (300mL) and methyl tert-butyl ether (50mL) into the lower oily liquid phase, continuing to settle, repeating the process for 5 times to obtain a viscous oily substance, and drying to obtain a product 28-258:7.78 g.

28-260

28-258(7.78g, 6.3263mmol) was placed in a 500mL flask, dichloromethane (30mL) was added and dissolved, and TFA (9/396mL, 126.52mmol) was added with stirring, and the reaction was stirred at room temperature overnight. After the reaction, the reaction solution was concentrated, methyl tert-butyl ether (300mL) was added to precipitate a solid, which was filtered, the filter cake was washed with methyl tert-butyl ether (80 mL. times.3), and the filter cake was dried to give 28-260:8.1 g.

25-130

Boc-GFLG-OH (synthesized by 25-102, 4.9738g, 10.0977mmol), HBTU (4.4186g, 11.6513mmol), HOBT (1.5743g, 11.6513mmol) and SB-743291(4.0162g, 7.7675mmol) were added to a 500mL round bottom flask, dissolved in DMF (60mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (5.8mL, 34.9538mmol) was slowly added dropwise, and after the addition was complete, the reaction flask was allowed to continue to stir at-5 ℃ for 1 hour and then transferred to room temperature overnight with stirring. After the reaction was completed, the reaction solution was transferred to a 2L separatory funnel, and a saturated sodium bicarbonate solution (400mL) and ethyl acetate (300mL) were added thereto, shaken, allowed to stand, extracted, and the aqueous phase and the organic phase were separated. Then, a saturated sodium chloride solution (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous phase and the organic phase were separated. Then, deionized water (300mL) was added to the organic phase, shaken, allowed to stand, extracted, and the aqueous and organic phases were separated. Finally, the organic phase is concentrated and evaporated to dryness and is placed in an oven for drying. Obtaining products 25-130: 7.7023 g.

25-132

25-130(7.7023g, 7.7675mmol) was charged into a 500mL round-bottom flask, dichloromethane (5mL) was added to dissolve, and TFA (8.7mL, 116.5125mmol) was added with stirring, and the reaction was stirred at room temperature overnight. After the reaction, the reaction mixture was concentrated and evaporated to dryness to remove methylene chloride. First, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and the mixture was settled, the supernatant was poured, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and the mixture was settled, and the operation was repeated three times to obtain an oily solid. The oily solid was dissolved with dichloromethane (10mL) and methyl tert-butyl ether (150mL) was added thereto to settle to a powdery solid, which was filtered to give the solid product. Dissolving the solid product with 20% methanol/dichloromethane mixed solvent (50mL), adding 80mL silica gel powder, evaporating to dryness, loading by dry method, and performing column chromatography. Eluting with eluent (1% ammonia water: 3% -4% methanol: 96% -95% dichloromethane), collecting liquid, concentrating, and evaporating to dryness. Obtaining products 25-132: 5.4g, yield: 77.98 percent.

28-264

28-260(7.4g, 6.3263mmol), 25-132(5.9218g, 6.6426mmol), HBTU (3.5988g, 9.4895mmol) and HOBT (1.2823g, 9.4895mmol) were charged into a 500mL flask, DMF (110mL) was added to dissolve the mixture, the reaction was left at 0 ℃ and stirred for 10 minutes, DIEA (4.7mL, 28.4684mmol) was slowly added dropwise, and the reaction was stirred at 0 ℃ overnight. After the reaction is finished, adding n-hexane (300mL) and methyl tert-butyl ether (50mL), separating layers, pouring an upper layer, adding the n-hexane (300mL) and the methyl tert-butyl ether (50mL) into a lower oily liquid phase, continuing to settle, repeating the operation for 6 times, finally obtaining oily viscous oily matter, adding methanol (10mL) and dichloromethane (40mL) to dissolve, adding the methyl tert-butyl ether (450mL), separating out a solid, filtering, washing a filter cake by using the methyl tert-butyl ether (100mL x 3), and drying to obtain a product 28-264:12.9 g.

28-265

28-264(12.9g, 6.3263mmol) was charged into a 500mL flask, DMF (50mL) was added and dissolved, morpholine (16.5mL, 189.789mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction is finished, adding n-hexane (300mL) and methyl tert-butyl ether (50mL) for settling, carrying out layering, pouring the upper layer, adding the n-hexane (300mL) and the methyl tert-butyl ether (50mL) into the lower oily liquid phase, continuing to settle, repeating the process for 5 times, adding dichloromethane (20mL) and methanol (80mL) for dissolving, adding the methyl tert-butyl ether (350mL), precipitating a solid, filtering, washing a filter cake with the methyl tert-butyl ether (120mL x 2), and drying the filter cake to obtain a product 28-265:11.5 g.

37-1

28-265(11.54g, 6.3263mmol), Boc-Gly-OH (1.3299g, 7.5916mmol, available from Ark pharm), HBTU (3.5988g, 9.4895mmol), HOBT (1.2823g, 9.4895mmol) were added to a 250mL flask, DMF (80mL) was added to dissolve, the reaction was left to stir at-5 ℃ for 10 min, DIEA (4.7053mL, 28.4684mmol) was slowly added dropwise, the reaction was continued at-5 ℃ for 30 min after completion of the addition, and then the reaction was left to stir at room temperature overnight. After the reaction is finished, adding n-hexane (300mL) and methyl tert-butyl ether (50mL), demixing, pouring the upper layer, adding n-hexane (300mL) and methyl tert-butyl ether (50mL) into the lower oily liquid phase, continuing to settle, repeating the process for 4 times to obtain a viscous oily substance, adding dichloromethane (70mL) and methyl tert-butyl ether (350mL), separating out a solid, filtering, washing a filter cake with methyl tert-butyl ether (50mL x 3), and drying the filter cake in a vacuum oven to obtain a product 37-1:12.5 g.

37-2

37-1(12.537g, 6.3263mmol) was charged to a 500mL flask, dichloromethane (30mL), TFA (14.09mL, 189.789mmol) were added, and the reaction was stirred at room temperature overnight. After the reaction is finished, adding methyl tert-butyl ether (250mL), precipitating a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (100mL x 3), dissolving the filter cake with a methanol (60mL) dichloromethane (240mL), adding silica gel powder (25g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a dichloromethane mixed solution of 15 ammonia water and 7% -8% methanol, collecting and concentrating, evaporating to dryness to obtain a solid, and drying in a vacuum oven to obtain a product 37-2: 9.7g, yield 82%.

35-13

35-11(0.38g, 0.124mmol) was charged into a hydrogenation reactor, and Pd/C (0.0210g) was added, DMF (30mL) was added and dissolved, and hydrogen gas was introduced under a pressure Pa of 1.8MPa, and the reaction was stirred at room temperature overnight. After the reaction was complete, the reaction solution was filtered through celite, washing with DMF (20 mL. times.3) to give a DMF solution containing 35-13.

35-14

37-2(2.0g, 1.063mmol), HBTU (0.56g, 1.4832mmol), HOBT (0.2g, 1.4832mmol) were added to a solution of 35-13(0.364g, 0.1236mmol) in DMF (90mL) and the reaction was allowed to stand at-5 ℃ with stirring for about 10 minutes, followed by the slow dropwise addition of DIEA (0.74mL, 4.4496mmol) which was completed and allowed to react at-5 ℃ for 60 minutes, after which the reaction was allowed to warm to room temperature and stirred overnight. After the reaction is finished, adding n-hexane (100mL) and methyl tert-butyl ether (30mL), shaking for layering, pouring out the supernatant, adding n-hexane (100mL) and methyl tert-butyl ether (30mL) into the lower layer liquid, repeating the steps for 6 times, adding dichloromethane (30mL) and methyl tert-butyl ether (250mL) to separate out a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (100mL x 3), adding a methanol (30mL) dichloromethane (120mL) solution to dissolve, adding silica gel powder (15g), evaporating to dryness to form a powdery solid, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 5% -7% methanol, collecting and concentrating, evaporating to dryness to obtain a product 35-14:1.2g, wherein the yield is 56%.

35-16

35-14(1.2g, 0.0695mmol) was charged to a 250mL flask, dichloromethane (15mL) was added, TFA (1.0322mL, 13.6mmol) was added, and the reaction was stirred at room temperature overnight. After the reaction is finished, concentrating the reaction solution to a small amount, adding methyl tert-butyl ether (100mL) to separate out a powdery solid, performing suction filtration, cleaning a filter cake with methyl tert-butyl ether (50mL x 3), dissolving with a mixed solution of methanol (30mL) and dichloromethane (120mL), adding silica gel powder (10g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, and performing column chromatography with 1% ammonia water: eluting with 6-7% methanol dichloromethane mixed solution, collecting, concentrating and evaporating to dryness to obtain 35-16: 0.74g, yield 67%.

37-14

35-16(0.3011g, 0.01754mmol) was placed in 250mL of DMF (25mL) and dissolved, M-SCM-40K (1.5227g, 0.0369mmol, from KeKao) was added and dissolved by sonication, and the reaction was stirred at room temperature in the dark for 7 days at a low speed. After the reaction, adding n-hexane (120mL) and methyl tert-butyl ether (40mL), pouring the supernatant, adding n-hexane (120mL) and methyl tert-butyl ether (40mL) into the lower layer of liquid, repeating the reaction for 3 times to obtain a viscous oily substance, adding methyl tert-butyl ether (100mL) to separate out a solid, filtering, washing the filter cake with methyl tert-butyl ether (50mL x 3), dissolving the solid with a solution of methanol (30mL) and dichloromethane (120mL), adding silica gel powder (25g), evaporating to dryness to obtain a powdery solid, performing dry sampling, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 6% -7% methanol, collecting and concentrating, evaporating to obtain a solid, drying in a vacuum oven for 1 hour, adding absolute ethyl alcohol (5mL) and dichloromethane (20mL) to dissolve, adding methyl tert-butyl ether (80mL), separating out the solid, filtering, washing the filter cake with methyl tert-butyl ether (50mL, drying the filter cake in a vacuum oven to obtain a product 37-14: 0.98g, yield: 58 percent.

¹H-NMR(600MHz,DMSO-d₆)δ10.18-10.11(m,4H),9.21–8.97(m,4H),8.24–7.89(m,61H),7.58–7.43(m,12H),7.30–7.08(m,99H),5.91–5.77(m,4H),4.64–4.68(m,23H),4.38–4.14(m,57H),4.07–3.89(m,81H),3.60–3.45(m,7482H),3.24–2.96(m,7H),2.82–2.65(m,43H),2.42–2.40(m,12H),2.35–2.26(m,32H),2.25–2.03(m,11H),1.95–1.67(m,26H),1.63–1.41(m,40H),1.37–1.15(m,19H),1.17–1.05(m,17H),0.90–0.77(m,72H),0.52–0.46(m,8H).

2. Synthesis of 10-109 (Compound No. 16)

The route is as follows:

the details are as follows

23-210

Boc-Glu- (OH) -OBn (purchased from Shaoshao Yuan, 10g, 29.64mmol), Glu- (OBn)₂TosOH (16.3g, 32.61mmol), HBTU (16.9g, 44.46mmol) and HOBT (6.1g, 44.46mmol) were placed in a 500mL round-bottomed flask and then DMF (200mL) was added and allowed to dissolve, and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (22mL, 133.4mmol) was then added slowly dropwise, and after the addition was complete, the reaction was allowed to proceed at low temperature for 2 hours, after which the reaction apparatus was allowed to stand at room temperature and stirred overnight. After completion of the reaction, the reaction mixture was taken out, and a saturated sodium bicarbonate solution (200mL) was added thereto, followed by extraction with ethyl acetate three times (200 mL. times.3), and the organic phases were combined. The organic phase was added with saturated sodium chloride solution (200 mL. times.2), and extracted twice with ethyl acetate (100 mL. times.2). And finally, anhydrous sodium sulfate is used for removing water, and finally, evaporation is carried out to dryness and concentration are carried out to obtain 31.4g (weight) of the product.

29-1

23-210(31.4g, 29.64mmol) was dissolved in dichloromethane (100mL) in a 500mL round-bottom flask, followed by addition of TFA (33mL, 444.6mmol), reaction was stirred at room temperature overnight, the reaction was stopped, the reaction mixture was concentrated under reduced pressure and evaporated to dryness, a saturated sodium bicarbonate solution (200mL) was added to the reaction mixture, extraction was carried out three times with ethyl acetate (200 mL. times.3), and the organic phases were combined. The organic phase was added with saturated sodium chloride solution (200mL) and extracted twice with ethyl acetate (100 mL. times.2). And finally, removing water by using anhydrous sodium sulfate, and finally evaporating to prepare for the next reaction.

29-3

29-1(16.2g, 29.64mmol), 10-102 (synthesized by the method of 24-36, 38.5g, 38.5mmol), HBTU (16.8g, 44.4mmol) and HOBT (6g, 44.4mmol) were placed in a 500mL round-bottomed flask and then dissolved by adding DMF (200mL), and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (22mL, 133.2mmol) was then added slowly dropwise, and after the addition was complete, the reaction was allowed to proceed at low temperature for 2 hours, after which the reaction apparatus was allowed to stand at room temperature and stirred overnight. After completion of the reaction, the reaction mixture was taken out, and a saturated sodium bicarbonate solution (200mL) was added thereto, followed by extraction with ethyl acetate three times (200 mL. times.3), and the organic phases were combined. The organic phase was added with saturated sodium chloride solution (200 mL. times.2), and extracted twice with ethyl acetate (100 mL. times.2). And finally, removing water by using anhydrous sodium sulfate, evaporating to dryness and concentrating to obtain 34.1g (weighed) of a product, performing dry-method sample loading, performing column chromatography, eluting by using 45% ethyl acetate, collecting the product, concentrating to dryness, collecting a pure spot 11.1g, putting the pure spot into a refrigerator for later use, mixing the pure spot 10.7g, and putting the mixed spot into the next step for deprotection.

30-28

The starting materials Boc-GFLG-OBn (synthesized according to the literature, 19.0g,32.6mmol) and 10% Pd/C palladium on carbon catalyst (300mg) were charged into a hydrogenation reactor, DMF (50mL) was added to dissolve the catalyst, the hydrogenation reactor was closed with the solvent over a stirrer, three-pump three-fill (air in the reaction system was pumped by a vacuum pump for about 3 minutes-hydrogen-fill-hydrogen-pump-fill-hydrogen-fill) was performed, and then the pressure reading on the hydrogenation reactor was 18Psi, followed by overnight reaction at room temperature. After completion of the reaction was observed on the next day by TLC plate spotting, post-treatment was carried out, the reaction solution was taken out and uniformly added dropwise to a suction filtration funnel containing compacted diatomaceous earth, and the reaction apparatus was washed with DMF (90mL) until the reactor was washed clean and free of the product, to obtain a reaction product.

30-29

30-28(17.9mmol), LPT (8g,13.77mmol), HBTU (7.83g,20.65mmol) and HOBT (2.79g,20.65mmol) were placed in a 500mL round bottom flask and then dissolved by adding DMF (100mL), and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (10.24mL,61.96mmol) was then added slowly dropwise, and after the addition was complete, the reaction was allowed to proceed at low temperature for 2 hours, after which the reaction apparatus was allowed to stand at room temperature and stirred overnight. After the reaction was completed, deionized water (1000mL) was added to wash DMF, and a pale yellow solid was precipitated and dried to obtain 14.53g of a product.

14-128

30-29(14.53g,13.77mmol) was placed in a 500mL round-bottom flask, dissolved in dichloromethane (150mL), followed by addition of TFA (15.34mL,206.55mmol) and reaction with stirring at room temperature overnight, the reaction was stopped, the reaction solution was concentrated under reduced pressure, saturated sodium bicarbonate (200mL) was added to neutralize TFA, the product in the aqueous phase was extracted with ethyl acetate 3 times (150 mL. times.3), the combined organic phases were dried over anhydrous sodium sulfate, filtered with suction, concentrated, dried, dry loaded, column chromatographed, eluted with 5% methanol/0.5% ammonia/dichloromethane, and concentrated to give 13.15g of pure product.

10-85

29-3(1.95g, 2.46mmol) and 10% Pd/c (0.1g) were put into a hydrogenation reactor, DMF (30mL) was added and dissolved, the hydrogenation reactor was closed, three-pump three-fill was performed so that the pressure on the hydrogenation reactor was read at 0.18MPa, and then the reaction was carried out overnight at room temperature. After the reaction was completed, the reaction solution was filtered through celite, and the filter cake was washed with DMF (20mL x 3) to give 10-85% as a product with a yield of 100%.

10-86

10-85(1.28g, 2.46mmol), GFLG-LPT (synthesized by 14-128 methods, 8.0g, 8.37mmol), HBTU (4.20g, 11.07mmol) and HOBT (1.49g, 11.07mmol) were put in a 500mL flask, dissolved by adding an appropriate amount of DMF (40mL), and the reaction was stirred at 0 ℃ for 30 minutes. DIEA (5.69mL, 34.44mmol) was then added slowly dropwise over 3 minutes. The reaction was continued at-5 ℃ with stirring overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, and drying in a vacuum oven to obtain a product 10-86:8.2g with the yield of 100%.

10-87

10-86(8.2g, 2.46mmol) was charged to a 250mL flask, dichloromethane (8mL) and TFA (2.7mL, 36.90mmol) were added and dissolved, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating the reaction solution to be oily by using a rotary evaporator, adding methyl tert-butyl ether (60mL), precipitating powdery solid in the reaction solution, performing suction filtration, cleaning a filter cake by using methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake by using a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (40g), evaporating to be powdery, performing dry loading, performing column chromatography, performing gradient elution by using a dichloromethane mixed solution of 1% of ammonia water and 4.5% -6% of methanol, collecting and concentrating, and drying in a vacuum oven to obtain 18-87:5.56g of a product with the yield of 70%.

10-83

After partial dissolution of 29-3(9.9g, 12.5019mmol) in dichloromethane (10mL), TFA (18.5716mL, 250.0379mmol) was added and the solution was dissolved completely by sonication, and the mixture was stirred at room temperature with a ground glass stopper. Detecting by TLC, developing phosphomolybdic acid, stopping reaction, evaporating reaction liquid to dryness, removing dichloromethane, dissolving with ethyl acetate (200mL), adding saturated sodium bicarbonate solution until aqueous phase becomes alkaline, separating organic phase, extracting aqueous phase with ethyl acetate until the aqueous phase has no product, combining organic phase, washing with saturated saline solution for 3 times (100mL multiplied by 3), concentrating and evaporating to dryness to obtain 8.7g of product, and 0.1g of over-yield.

10-92

10-83(1.6g, 2.31mmol), Fmoc-Lys (Boc) -OH (1.14g, 2.43mmol), HBTU (1.32g, 3.47mmol) and HOBT (0.47g, 3.47mmol) were put in a 500mL flask, dissolved by adding an appropriate amount of DMF (40mL), and the reaction was stirred at 0 ℃ for 30 minutes. DIEA (1.72mL, 10.39mmol) was then added slowly dropwise over 3 minutes. The reaction was continued at-5 ℃ with stirring overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, collecting a filter cake, dissolving the filter cake with a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (30g), evaporating to be powdery, performing dry sampling, performing column chromatography, performing gradient elution with a 1-5% methanol dichloromethane mixed solution, collecting and concentrating, and drying in a vacuum oven to obtain a product 10-92:2.4g with a yield of 92.3%.

10-88

10-92(0.57g, 0.506mmol) and 10% Pd/c (0.1g) were put into a hydrogenation reactor, DMF (30mL) was added and dissolved, the hydrogenation reactor was closed, three-pump three-fill was performed so that the pressure on the hydrogenation reactor was read at 0.18MPa, and then the reaction was carried out overnight at room temperature. After the reaction was completed, the reaction solution was filtered through celite, and the filter cake was washed with DMF (20mL x 3) to give a product 10-88: 0.434, yield 100%.

10-89

10-88(0.434g, 0.506mmol), 10-87(5.56g, 1.722mmol), HBTU (0.864g, 2.277mmol) and HOBT (0.307g, 2.277mmol) were put in a 500mL flask, dissolved by adding an appropriate amount of DMF (40mL), and the reaction was stirred at 0 ℃ for 30 minutes. DIEA (1.171mL, 7.084mmol) was added slowly dropwise over 30 min. The reaction was continued at-5 ℃ with stirring overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring out the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring out the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, and drying in a vacuum oven to obtain a product 10-89: 5.34g, yield 100%.

10-90

10-89(5.34g, 0.506mmol) was put into a 250mL flask, DMF (10mL) was added thereto and dissolved, and morpholine (0.88mL, 10.12mmol) was added thereto and the reaction was stirred at ordinary temperature for 1 hour. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (30g), evaporating to be powdery, performing dry sampling, performing column chromatography, performing gradient elution with a dichloromethane mixed solution of 1% ammonia water and 3% -7% methanol, collecting and concentrating, and drying in a vacuum oven to obtain a product 10-90:2.65g with the yield of 51%.

10-71

30-33(3.5g, 4.26mmol), Fmoc-Lys (Boc) -OH (2.2g, 4.69mmol), HBTU (2.42g, 6.39mmol), and HOBT (0.86g, 6.39mmol) were put in a 250mL flask, dissolved by adding an appropriate amount of DMF (40mL), and the reaction was stirred at 0 ℃ for 30 minutes. DIEA (3.17mL, 19.17mmol) was added slowly dropwise over 30 min. The reaction was continued at-5 ℃ with stirring overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring out the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring out the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, and drying in a vacuum oven to obtain a product 10-71: 5.42g, yield 100%.

10-76

10-71(5.42g, 4.26mmol) was put into a 250mL flask, DMF (10mL) was added and dissolved, and morpholine (7.42mL, 85.2mmol) was added and the reaction was stirred at room temperature for 1 hour. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, then dissolving with methanol/dichloromethane (1:4) solution (100mL), concentrating, and drying in a vacuum oven to obtain 10-76:4.473g of a product with the yield of 100%.

10-77

10-76(4.473g, 4.26mmol) and succinic anhydride (1.29g, 12.89mmol) were put into a 250mL flask, and an appropriate amount of DMF (40mL) was added thereto to dissolve the mixture, and the reaction was stirred at 0 ℃ for 30 minutes. DIEA (2.82mL, 17.04mmol) was then added slowly dropwise over 30 minutes. The reaction was continued at-5 ℃ with stirring overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (30g), evaporating to be powdery, performing dry sampling, performing column chromatography, performing gradient elution with a dichloromethane mixed solution of 1% ammonia water and 5% -8% methanol, collecting and concentrating, and drying in a vacuum oven to obtain a product 10-77:2.98g with the yield of 61%.

10-97

10-77(0.28g, 0.245mmol), 10-90(2.65g, 0.62mmol) HBTU (0.139g, 0.368mmol) and HOBT (0.049g, 0.368mmol) were put in a 250mL flask, dissolved by adding DMF (40mL) and reacted at 0 ℃ for 30 minutes with stirring. DIEA (0.18mL, 1.1mmol) was then added slowly dropwise over 30 minutes. The reaction was continued at-5 ℃ with stirring overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring out the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring out the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, and drying in a vacuum oven to obtain a product 10-97: 2.8g, yield 100%.

10-98

10-97(2.8g, 0.245mmol) was charged to a 250mL flask, dichloromethane (8mL) and TFA (0.546mL, 7.35mmol) were added and dissolved, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating the reaction solution to be oily by using a rotary evaporator, adding methyl tert-butyl ether (60mL), precipitating powdery solid in the reaction solution, performing suction filtration, cleaning a filter cake by using methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake by using a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (30g), evaporating to be powdery, performing dry loading, performing column chromatography, performing gradient elution by using a dichloromethane mixed solution of 1% of ammonia water and 5% -10% of methanol, collecting and concentrating, and drying in a vacuum oven to obtain a product 10-98:1.38g, wherein the yield is as follows: 60 percent of

10-109

10-98(1.29g, 0.1134mmol) was placed in a 250mL flask, dissolved in DMF (10mL), and M-SCM-20K (0.54g, 0.054mmol, from KEY) was added to the flask, and the reaction was stirred at low speed at room temperature in the dark for one week. After the reaction is finished, adding methyl tert-butyl ether (40mL), precipitating a solid, performing suction filtration, cleaning a filter cake with methyl tert-butyl ether (40mL x 3), dissolving the filter cake with a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (10g), evaporating to dryness to obtain a powdery solid, performing dry sampling, performing column chromatography, performing gradient elution with a dichloromethane mixed solution of 1% ammonia water and 6% -12% methanol, collecting and concentrating, evaporating to obtain a solid, and drying in a vacuum oven to obtain a product 10-109:0.7g, wherein the yield is 51%.

¹H-NMR(600MHz,DMSO-d₆)δ7.46–6.89(m,225H),4.21–4.08(m,111H),3.55-3.49(m,3882H),3.08–2.64(m,209H),2.45-2.40(m,10H),2.34-2.28(m,28H),0.88-0.83(m,60H).

3. 39-17 Synthesis (Compound No. 14)

The route is as follows

The method comprises the following specific steps:

41-1

Boc-Glu-OH (1.2g, 4.8mmol), GFLF-PCB (synthesized by 30-33, 8.2g, 9.7mmol), HBTU (5.5g, 14.4mmol) and HOBT (1.9g, 14.4mmol) were placed in a 500mL round-bottomed flask and then dissolved by adding DMF (50mL), and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (7.1mL, 43.2mmol) was slowly added dropwise and the reaction was continued for 2h and stirred at room temperature overnight. After the reaction is finished, deionized water (200mL) is added into the reaction liquid, a light yellow solid is separated out, and the product 22g (weighed) is obtained after suction filtration and drying.

41-2

41-1(22g, 11.8mmol) was placed in a 500mL round-bottom flask, and dichloromethane (10mL) was added to dissolve it, followed by dropwise addition of TFA (26.3mL, 354mmol), and stirring at room temperature overnight. After the reaction, the reaction solution was concentrated, n-hexane (100mL) and methyl tert-butyl ether (300mL) were added to the reaction solution for precipitation, the solution was filtered, dichloromethane (80mL) and methanol (20mL) were added to the filter cake to dissolve the filter cake, and silica gel powder was added to the solution and evaporated to dryness. And (3) performing column chromatography by using a dry method, performing gradient elution by using 1% ammonia water/3% methanol/dichloromethane-1% ammonia water/5% methanol/dichloromethane, collecting and concentrating to obtain 5.9g of a product, wherein the yield is 70.2%.

41-4

41-2(5.9g, 3.36mmol), Boc-LC-OH (0.88g, 3.36mmol), HBTU (1.9g, 5.04mmol) and HOBT (0.68g, 5.04mmol) were placed in a 250mL round bottom flask and then DMF (20mL) was added to dissolve it and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (2.5mL, 15.12mmol) was slowly added dropwise and the reaction was continued for 2h and stirred at room temperature overnight. After the reaction, n-hexane (100mL) and methyl tert-butyl ether (300mL) were added to the reaction mixture to precipitate a solid, which was then filtered off with suction and dried in vacuo.

41-6

41-4(7.9g, 3.9mmol) was placed in a 250mL round-bottom flask, and dichloromethane (10mL) was added to dissolve it, followed by dropwise addition of TFA (8.7mL, 117mmol) and stirring at room temperature overnight. After the reaction is finished, the reaction solution is concentrated, n-hexane (100mL) and methyl tert-butyl ether (300mL) are added for sedimentation, and the mixture is filtered, solid is taken out and dried for later use. And (3) performing dry column chromatography, and performing column chromatography by using 1% ammonia water: 3% methanol/dichloromethane-1% ammonia: gradient elution is carried out on 5 percent methanol/dichloromethane, and the product is obtained after collection and concentration.

24-205

Weighing reactant 15-91 (synthesized according to 35-3 route, 0.33g, 0.1503mmol), putting into a micro reaction kettle, adding 10% Pd/C (30mg), adding DMF (30mL) for dissolving, and introducing H₂(20psi), stir the reaction. After the reaction is finished, diatomite is used as a filter cake, the reaction liquid is filtered by suction to remove Pd/C, the diatomite is washed 4 times by DMF to obtain a DMF solution of 24-205, and the next step of reaction is carried out.

41-7

41-6(0.84g, 0.44mmol), 24-247 (synthesized by 24-205, 0.88g, 0.091mmol), HBTU (0.07g, 0.546mmol) and HOBT (0.2g, 0.546mmol) were placed in a 250mL round-bottomed flask and then DMF (20mL) was added to dissolve it, and the reaction was stirred at-5 ℃ for 30 minutes. DIEA (0.27mL, 1.638mmol) was added slowly dropwise and the reaction was continued for 2h and stirred at room temperature overnight. After the reaction, n-hexane (100mL) and methyl tert-butyl ether (300mL) were added to the reaction mixture to precipitate a solid, which was then filtered off with suction. And (5) drying in vacuum.

41-9

Reaction product 41-7(0.7g, 0.082mmol) was dissolved in DMF (30mL), morpholine (0.214mL, 2.46mmol) was added, the reaction was stirred, methyl tert-butyl ether (100mL) was added after the reaction was completed, n-hexane (200mL) was added to precipitate as a powder, which was filtered under suction. Column chromatography, dry loading, loading with 1% ammonia water: 5% methanol/dichloromethane-1% ammonia: elution with a 12% methanol/dichloromethane gradient. The product weighed 0.5 grams.

39-1

Boc-Asp-OH (purchased from damas-beta, 0.18g, 0.8mmol), GFLG-SB7 (synthesized by 25-132, 1.5g, 1.68mmol), HBTU (0.9g, 2.4mmol) and HOBT (0.32g, 2.4mmol) were placed in a 250mL round bottom flask and then dissolved by addition of DMF (20mL), and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (1.19mL, 7.2mmol) was slowly added dropwise to continue the reaction for 2h, after which it was allowed to stir at room temperature. After the reaction was completed, the reaction mixture was taken out, deionized water (200mL) was added thereto, extraction was carried out three times with ethyl acetate (100 mL. times.3), the organic phases were combined, and the organic phase was washed twice with saturated sodium chloride solution (200mL), concentrated and evaporated to dryness. Loading the sample by a dry method, carrying out column chromatography, eluting the eluent by using 1% ammonia water, 2% methanol/dichloromethane-1% ammonia water and 3% methanol/dichloromethane for gradient elution, collecting the product, concentrating, evaporating to dryness to obtain the product, and putting the product into the next step for reaction.

39-5

39-1(1.5g, 0.8mmol) was dissolved in dichloromethane (20mL) in a 250mL round-bottom flask and then TFA (1.78mL, 24mmol) was added and the reaction stirred at room temperature overnight. After completion of the reaction, the reaction mixture was concentrated, and a saturated sodium bicarbonate solution (200mL) and ethyl acetate (100mL) were added to extract the mixture, the organic phase was separated, the aqueous phase was extracted three times with ethyl acetate (100 mL. times.3), and the organic phases were combined. The organic phase was washed twice with saturated sodium chloride solution (200 mL). The organic phase was evaporated to dryness to give 1.1g of product in 73.3% yield.

39-6

39-5(1.1g, 0.58mmol), Fmoc-Lys (Boc) -OH (0.28g, 0.609mmol), HBTU (0.32g, 0.87mmol) and HOBT (0.11g, 0.87mmol) were placed in a 250mL round bottom flask and then dissolved by adding DMF (20mL), and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (0.43mL, 2.61mmol) was slowly added dropwise to continue the reaction for 2h, after which it was allowed to stir at room temperature. After completion of the reaction, the reaction mixture was taken out and extracted with deionized water (200mL) and ethyl acetate (100mL), the organic phase was separated, the aqueous phase was extracted three times with ethyl acetate (100 mL. times.3), and the organic phases were combined. The organic phase was washed twice with saturated sodium chloride solution (200 mL). Concentrating and evaporating to dryness. 1.3 g of the product is obtained.

39-8

39-6(1.5g, 0.58mmol) was placed in a 250mL round bottom flask, dissolved in DMF (20mL), and morpholine (1.51mL, 17.4mmol) was added and the reaction stirred at room temperature for 1 hour. After completion of the reaction, the reaction mixture was taken out and extracted with deionized water (200mL) and ethyl acetate (100mL), the organic phase was separated, the aqueous phase was extracted three times with ethyl acetate (100 mL. times.3), and the organic phases were combined. The organic phase was washed twice with saturated sodium chloride solution (200 mL). Concentrating and evaporating to dryness. And (5) putting the next step of reaction.

39-10

39-8(0.58mmol) was placed in a 250mL round-bottom flask, dissolved in DMF (20mL), DIEA (0.38mL, 2.32mmol) was added, and after stirring at room temperature for 30 minutes, succinic anhydride (0.17g, 2.32mmol) was added and the reaction was continued. After completion of the reaction, the reaction mixture was taken out and extracted with deionized water (200mL) and ethyl acetate (100mL), the organic phase was separated, the aqueous phase was extracted three times with ethyl acetate (100 mL. times.3), and the organic phases were combined. The organic phase was washed twice with saturated sodium chloride solution (200 mL). Concentrating and evaporating to dryness. And (5) putting the next step of reaction.

39-14

Reactants 41-9(0.5g, 0.06mmoL), 39-10(0.134g, 0.060mmoL), HBUT (0.034g, 0.09mmoL), HOBT (0.012g, 0.09mmoL) were weighed into a 250mL reaction flask, DMF (30mL) was added to dissolve, the mixture was stirred at-5 ℃ for 0.5 hour, DIEA (0.044mL, 0.27mmoL) was slowly added dropwise, and the mixture was taken out after 1 hour and stirred at room temperature for reaction. After the reaction, methyl tert-butyl ether (100mL) was added, and n-hexane (200mL) was added to the mixture to precipitate the mixture as a powder. Column chromatography, dry loading, loading with 1% ammonia water: 4% methanol/dichloromethane-1% ammonia: 10% methanol in dichloromethane. The product was evaporated to dryness to give 0.5g, 80% yield.

39-15

39-14(0.5g) was weighed out, dichloromethane (5mL) was added, TFA (0.106mL, 1.42mmol) was added, the mixture was dissolved by sonication, and the reaction was stirred. After the reaction, the reaction mixture was concentrated, and methyl t-butyl ether (100mL) was added to the concentrated solution, followed by addition of n-hexane (200mL) to the concentrated solution and precipitation to obtain a powder. Column chromatography, dry loading, loading with 1% ammonia water: 6% methanol/dichloromethane-1% ammonia: 10% methanol/dichloromethane gradient. The product weighed 0.1 gram.

¹H-NMR(600MHz,DMSO-d₆)δ10.19–10.15(m,7H),9.11–8.89(m,7H),8.36–8.00(m,37H),7.94–7.85(m,13H),7.54–7.47(m,11H),7.30–7.11(m,89H),6.74–6.63(m,2H),5.87–5.78(m,6H),4.66–3.37(m,118H),3.29–2.53(m,123H),2.45–2.39(m,24H),2.34–2.18(m,59H),2.14–1.42(m,107H),0.94–0.76(m,72H).

39-17

The reaction mass 39-15(1g) was weighed, dissolved in DMF (20mL), and M-SCM-20K (0.4g) was added and stirred at low speed with the exclusion of light. After the reaction, methyl tert-butyl ether (50mL) and n-hexane (100mL) were added to precipitate, and a solid was precipitated and filtered off. The filter cake was dissolved in dichloromethane (50mL), silica gel powder was added and evaporated to dryness. Column chromatography, dry loading, eluting with dichloromethane-1% ammonia: elution with a gradient of 6% methanol in dichloromethane. Collecting the product, evaporating to dryness, adding anhydrous ethanol (10mL), performing ultrasonic treatment to obtain a homogeneous phase, and adding n-hexane (50mL) for settling. The sedimentation was repeated three times. Vacuum drying to obtain 0.15 g of product.

¹H-NMR(600MHz,DMSO-d₆)δ10.20–10.12(m,2H),9.00–8.90(m,2H),8.16–7.83(m,40H),7.42–7.04(m,123H),4.64–4.13(m,129H),4.03–3.97(m,35H),3.81–3.80(m,26H),3.70–3.66(m,149H),3.51–3.50(m,876H),3.27–2.64(m,20H),2.43–2.24(m,54H),2.17–1.41(m,24H),0.97–0.73(m,72H).

4. Synthesis of 43-27 (Compound No. 6)

The route is as follows

34-15

Boc-L-Lys (Fmoc) -OH (purchased from Aladdin, 2.78g, 5.927mmol), 22-181(5.4g, 5.927mmol), HBTU (3.38g, 8.891mmol), HOBT (1.2g, 8.891mmol) was added to a 100mL flask, dissolved in DMF (50mL), and after the reaction was allowed to stand at-5 ℃ for about 30 minutes, DIEA (5.878mL, 35.565mmol) was slowly added dropwise, after which the reaction was allowed to continue to stir at-5 ℃ for 3 hours. After completion of the reaction, the reaction mixture was transferred to a 1L separatory funnel, and deionized water (200mL) and ethyl acetate (200mL) were added thereto, followed by shaking and extraction. The aqueous phase was washed with ethyl acetate (150mL x 2), the organic phases were combined, washed with saturated brine (200mL x 1), concentrated to dryness and dried in a vacuum oven to give product 34-15: 8.07 g.

34-17

34-15(8.0g, 5.927mmol) was added to a 500mL flask, dichloromethane (15mL) was added, trifluoroacetic acid (4.4mL, 59.271mmol) was added, and the reaction was stirred at room temperature overnight. After the reaction is finished, concentrating the reaction solution to a small amount, adding n-hexane (150mL), shaking, layering, pouring the supernatant, adding n-hexane (150mL) into the lower oily substance, repeating the operation for 6 times to finally obtain an oily substance, and drying to obtain a product 34-17: 7.5 g.

34-19

34-17(7.5g, 5.92mmol), 34-18 (synthesized by 25-102, 3.5g, 7.11mmol), HBTU (3.36g, 8.88mmol), HOBT (1.2g, 8.88mmol) was charged into a 500mL flask, dissolved in DMF (50mL), and the reaction was stirred at-5 ℃ for about 30 minutes, then DIEA (8.8mL, 53.28mmol) was slowly added dropwise, and after the addition was complete, the reaction was stirred at-5 ℃ for 3 hours. After completion of the reaction, the reaction mixture was transferred to a 1L separatory funnel, and deionized water (200mL) and ethyl acetate (200mL) were added thereto, followed by shaking and extraction. The aqueous phase was washed with ethyl acetate (150mL x 2), the organic phases were combined, washed with saturated brine (200mL x1), concentrated to dryness and dried in a vacuum oven to give product 34-19: 10.3 g.

34-21

Putting 34-19(10.3g, 5.92mmol) into a 500mL flask, adding DMF (20mL) to dissolve, adding morpholine (7.8mL, 88.8mmol), reacting at room temperature for 1 hour, finishing the reaction, transferring the reaction solution into a 1L separating funnel, adding saturated saline (150mL) and ethyl acetate (250mL) to extract to obtain an organic phase, washing the aqueous phase with ethyl acetate (200mL x1), combining the organic phases, washing the organic phase with saturated saline (200mL x 2), concentrating and evaporating to dryness, adding methanol (30mL) and dichloromethane (120mL) to dissolve, adding silica gel powder (15g), evaporating to dryness to obtain a powdery solid, performing dry sampling, performing column chromatography, and eluting with a mixed solution of 3% methanol and dichloromethane to obtain a product 34-21:5.37g, wherein the yield is 60%.

43-8

34-21(1.6g, 1.06mmol) was placed in a 250mL flask, dichloromethane (30mL) was added to dissolve the mixture, triethylamine (0.29mL, 2.12mmol) was added, the reaction was left to stir at 0 ℃ for 15 minutes, phenyl chloroformate (0.13mL, 1.06mmol) was added dropwise, and the reaction was continued to stir at 0 ℃ overnight. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding dichloromethane (250mL) and saturated sodium chloride solution (200mL) for extraction to obtain an organic phase, washing an aqueous phase with dichloromethane (200mL x 1), combining the organic phases, adding silica gel powder (20g), evaporating to obtain a powdery solid, loading the powdery solid by a dry method, carrying out column chromatography, and eluting with a mixed solution of 4% methanol and dichloromethane to obtain a product 43-8:0.9g, wherein the yield is 56%.

43-11

Putting 43-8(0.9g, 0.55mmol) and 34-21(0.83g, 0.55mmol) into a 100mL flask, adding DMF (20mL) for dissolving, adding triethylamine (0.0771mL, 0.55mmol) for reaction, stirring at 80 ℃ overnight, finishing the reaction, transferring the reaction solution into a 1L separating funnel, adding ethyl acetate (200mL) and saturated saline (200mL) for extraction to obtain an organic phase, washing the organic phase with saturated saline (150mL), adding silica gel powder (10g), evaporating to dryness to obtain a powdery solid, loading by a dry method, carrying out column chromatography, eluting with a mixed solution of 1% ammonia water and 5% -36% methanol in dichloromethane, and obtaining a product 43-11:0.95g with a yield of 57%.

¹H-NMR(400MHz,DMSO-d₆)δ8.57–8.48(m,2H),8.37–8.24(m,2H),8.21–8.12(m,2H),8.08–7.95(m,4H),7.94–7.80(m,4H),7.76–7.65(m,2H),7.51(s,1H),7.44–7.25(m,40H),7.24–7.10(m,11H),7.01–6.81(m,2H),5.17–5.01(m,15H),4.54(d,J＝4.4Hz,2H),4.41–4.16(m,10H),3.94–3.86(m,4H),3.78–3.68(d,J＝4.4Hz,4H),3.66–3.49(m,13H),3.45(d,J＝6.0Hz,2H),3.42–3.36(m,6H),3.24–3.15(d,J＝5.5Hz,4H),3.08–2.97(m,2H),2.94–2.86(m,4H),2.83–2.71(m,3H),2.46–2.32(m,9H),2.23–2.11(m,4H),1.81–1.68(m,4H),1.68–1.53(m,5H),1.53–1.42(m,7H),1.34(d,J＝5.9Hz,18H),1.31–1.10(m,19H),0.93–0.75(m,13H).

43-15

34-23 (synthesized by 43-11, 0.377g, 0.1236mmol) was charged into a hydrogenation reactor, 10% Pd/C (0.0400g) was added thereto, DMF (30mL) was added thereto and dissolved therein, hydrogen was introduced thereinto under a pressure of 1.8MPa, and the reaction was stirred at room temperature overnight. After the reaction was completed, the reaction vessel was taken out, and the reaction solution was filtered through celite, and washed with DMF (20mL × 3) to be used as a raw material for the next reaction.

43-21

43-15(0.288g, 0.1236mmol), 37-2(2.0g, 1.063mmol), HBTU (0.56g, 1.4832mmol) and HOBT (0.2g, 1.4832mmol) were charged into a 250mL flask, DMF (40mL) was added to dissolve the mixture, the reaction was left at-5 ℃ and stirred for 20 minutes, DIEA (0.74mL, 4.4496mmol) was slowly added dropwise, the reaction was stirred at-5 ℃ for 30 minutes after completion of the addition, and then stirred at room temperature overnight. After the reaction is finished, adding n-hexane (120mL) and methyl tert-butyl ether (30mL), settling for 3 times to obtain a viscous oily substance, adding methyl tert-butyl ether (250mL), separating out a solid, filtering, washing a filter cake with methyl tert-butyl ether (50mL x 3), dissolving the filter cake with dichloromethane (150mL) and methanol (30mL), adding silica gel powder (10g), evaporating to obtain a powdery solid, loading by a dry method, performing column chromatography, eluting with a dichloromethane mixed solution of 1% of ammonia water and 5% -6% of methanol, collecting, concentrating and drying to obtain a product 43-21:1.6g, wherein the yield is 75%.

43-24

43-21(1.6g, 0.09mmol) was charged into a 250mL flask, dichloromethane (10mL) and trifluoroacetic acid (1mL, 13.47mmol) were added, and the reaction was stirred at room temperature overnight. After the reaction is finished, concentrating the reaction solution to a small amount, adding methyl tert-butyl ether (150mL) to precipitate a solid, performing suction filtration, cleaning a filter cake with methyl tert-butyl ether (50mL x 3), dissolving with a methanol (30mL) dichloromethane (120mL), adding silica gel powder (20g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a mixed solution of 1% ammonia water and 6% -8% methanol dichloromethane, collecting and concentrating, evaporating to dryness to obtain a solid, and drying in a vacuum oven to obtain a product 43-24: 1.11g, 73% yield.

43-27

43-24(0.4g, 0.023mmol) was put into 250mL, DMF (25mL) was added and dissolved, M-SCM-40K (1.983g, 0.048mmol) was added and dissolved by ultrasound, and the reaction was stirred at low speed at room temperature for 7 days in the dark. After the reaction, adding n-hexane (150mL) and methyl tert-butyl ether (50mL), pouring the supernatant, adding n-hexane (150mL) and methyl tert-butyl ether (50mL) into the lower layer liquid, repeating the reaction for 3 times to obtain a viscous oily substance, adding methyl tert-butyl ether (120mL), separating out a solid, filtering, washing the filter cake with methyl tert-butyl ether (50mL x 3), dissolving the filter cake with a solution of methanol (30mL) and dichloromethane (120mL), adding silica gel powder (15g), evaporating to dryness to obtain a powdery solid, performing dry sampling, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 5% -6% methanol, collecting and concentrating, evaporating to obtain a solid, performing vacuum oven drying for 1 hour, adding absolute ethyl alcohol (5mL) and dichloromethane (20mL) to dissolve, adding methyl tert-butyl ether (80mL), separating out a solid, filtering, washing the filter cake with methyl tert-butyl ether (50mL x 2, drying the filter cake in a vacuum oven to obtain a product 43-27: 0.9g, yield: 38 percent.

¹H-NMR(600MHz,DMSO-d₆)δ10.16–10.13(m,5H),8.99–8.93(m,6H),8.29–7.95(m,84H),7.92–7.85(m,14H),7.59–7.44(d,J＝3.3Hz,70H),7.33–7.03(m,154H),4.53–4.41(m,55H),3.72–3.41(m,7336H),3.20–3.03(m,123H),2.42–2.38(m,25H),2.36–2.15(48,71H),1.94–1.69(m,48H),1.65–1.39(m,71H),1.36–1.08(m,235H),0.99–0.68(m,124H),0.56–0.39(m,16H).

5. Synthesis of 27-134 (Compound No. 18)

The route is as follows

27-120

DMF (10mL) was added to a flask containing 27-119 (synthesized by 35-3, 1g, 0.7345mmol), and morpholine (0.64mL, 7.345mmol) was added after complete dissolution by sonication, and the reaction was stirred at room temperature for 2 h. After the reaction is finished, adding saturated sodium bicarbonate (100mL), adding ethyl acetate (200mL), extracting, standing for layering, and collecting an organic phase; washing the water phase with ethyl acetate (200mL x 3), collecting the combined organic phases, evaporating to dryness and concentrating to about 50mL, taking out, adding saturated sodium chloride (100mL x 3) for washing, collecting the combined organic phases, concentrating, evaporating to dryness, and vacuum oven drying to obtain 27-120(1.4317g, 100%)

27-121

Succinic acid (0.039g, 0.3339mmol), HBTU (0.3798g, 1.0017mmol) and HOBT (0.1354g, 1.0017mmol) were weighed into a flask containing 27-120(0.8368g, 0.7345mmol), an appropriate amount of DMF was added to dissolve the mixture, DIEA (0.50mL, 3.0051mmol) was slowly added dropwise at-5 ℃, after completion of the dropwise addition, the mixture was taken out after half an hour of the reaction, and the reaction was stirred at room temperature overnight. After the reaction is finished, adding excessive saturated sodium bicarbonate until the solution is alkaline, adding ethyl acetate (200mL), extracting, standing for layering, and collecting an organic phase; washing the water phase with ethyl acetate (200mL x 3), combining the organic phases, evaporating to dryness and concentrating to about 100mL, taking out, adding saturated sodium chloride (100mL x 3) for washing, collecting the combined organic phases, evaporating to dryness, dissolving with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (50mL), evaporating to dryness, performing dry loading, performing column chromatography, eluting with a mixed solution of 1% ammonia water and 6% methanol in dichloromethane, collecting and concentrating, evaporating to dryness, and drying in a vacuum oven to obtain 27-121(0.5811g, 73.72%) of the product

27-123

27-121(0.5811g, 0.2462mmol) was added to the hydrogenation reactor, Pd/C (0.1g) was added, DMF (20mL) was added, hydrogen was introduced under a pressure of 1.4MPa, and the reaction was stirred at room temperature overnight. After the reaction was complete, the reaction solution was filtered through celite, the filter cake was washed with DMF (15 mL. times.3), and the filtrate was charged into a 250mL round bottom flask to give 27-123 as the next starting material.

27-128

35-12 (synthesized by 37-2, 1.8g, 0.9566mmol), HBTU (0.54g, 1.4348mmol), HOBT (0.19g, 1.4348mmol) were weighed into a flask containing 27-123(0.1782g, 0.1087mmol), an appropriate amount of DMF was added to dissolve the mixture, DIEA (0.23mL, 4.3045mmol) was slowly added dropwise at-5 ℃, the reaction was completed after completion of the dropwise addition, the reaction was taken out after half an hour, and the mixture was stirred at room temperature overnight. After the reaction is finished, methyl tert-butyl ether is added: shaking 250mL of mixed solution of n-hexane and 1:5, standing, pouring out supernatant, repeating for three times, adding 200mL of methyl tert-butyl ether, precipitating solid, performing suction filtration, dissolving filter cake with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (40mL), evaporating to dryness to obtain powdery solid, loading by a dry method, performing column chromatography, eluting with mixed solution of 1% ammonia water and 3-5% methanol in dichloromethane, collecting and concentrating, and drying by a vacuum oven to obtain 27-128(0.8g, 44.69%)

27-132

Methylene chloride was added to a flask containing 27-128(0.8g, 0.0483mmol), and after complete dissolution by sonication, TFA (0.107mL, 1.4503mmol) was added and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating dichloromethane in the reaction liquid to dryness, adding methyl tert-butyl ether (100mL) for oscillation, precipitating a solid, performing suction filtration, dissolving a filter cake by using a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (40mL), evaporating to dryness, performing dry loading, performing column chromatography, eluting by using a mixed solution of 1% ammonia water and 7-10% methanol in dichloromethane, collecting, concentrating, and drying in a vacuum oven to obtain a product 27-132(0.5633g, 71.34%).

¹H-NMR(600MHz，DMSO-d₆)δ10.21-10.17(m,4H)，9.15–8.87(m，8H)，8.47–7.69(m，91H)，7.60–6.88(m，202H)，6.78–6.62(m，4H)，5.93–5.60(m，13H)，5.35-5.29(m，1H)，4.57-4.53(m，9H)，4.27(m，23H)，4.07–3.22(m，166H)，3.21–2.85(m，40H)，2.85–2.53(m，28H)，2.44–2.04(m，87H)，2.04–1.95(m，7H)，1.83-1.79(m，41H)，1.55-1.49(m，55H),1.38–0.99(m，92H)，0.86-0.82(m，103H)，0.52-0.47(m，39H).

27-134

27-132(0.2633g, 0.01610mmol) was placed in a 250mL flask, and dissolved in DMF (20mL), M-SCM-20K (0.7955g, 0.03703mmol, from KeKao) was added thereto, and dissolved by ultrasonic oscillation, and the reaction was stirred at a low speed at room temperature for one week while keeping out of the sun. After the reaction, methyl tert-butyl ether (200mL) and n-hexane (70mL) were added, the solid was precipitated, filtered, the filter cake was washed with methyl tert-butyl ether (40mL x 3), the filter cake was dissolved in methanol/dichloromethane (1:4) solution (100mL), silica gel powder (50mL) was added, the mixture was evaporated to dryness to give a powdery solid, the solid was loaded by dry method, column chromatography was performed, elution was performed with a mixed solution of 1% ammonia and 7% methanol in dichloromethane, the mixture was collected and concentrated, and the product was dried in a vacuum oven to obtain 27 to 134(0.5876g, 61.85%).

¹H-NMR(600MHz,DMSO-d₆)δ9.06-9.02(m,19H),8.98-8.92(m,11H),8.28–7.93(m,69H),7.87-7.82(m,25H),7.55-7.49(m,32H),7.44–6.95(m,116H),6.92-6.88(m,10H),6.76–6.63(m,26H),5.40–5.25(m,29H),4.62–4.43(m,38H),4.39–4.07(m,64H),3.72-3.68(m,3897H),3.19–3.10(m,20H),2.91-2.86(m,19H),2.83–2.57(m,62H),2.36–2.06(m,66H),2.05–1.89(m,38H),1.51-1.45(m,50H),1.39–0.97(m,242H),0.95–0.64(m,87H).

6. Synthesis of 44-2 (Compound No. 3)

The route is as follows

25-189

7-Ethyl-10-hydroxycamptothecin (abbreviated SN38, 20g, 50.9684mmol) and tert-butyldiphenylchlorosilane (available from Innochem, TBDPSCl, 79.5mL, 305.8104mmol) were charged into a 1L round bottom flask, and dichloromethane (250mL) and triethylamine (Et)₃N, 42.5mL, 305.8104mmol) was dissolved and the reaction flask was placed in a 37 ℃ oil bath and mechanically stirred overnight. After the reaction is finished, the reaction solution is concentrated under reduced pressure and evaporated to dryness to remove dichloromethane, dichloromethane (20mL) is added to dissolve the dichloromethane, then n-hexane (200mL) is added to the reaction solution to precipitate, a solid product is separated out, the filtration is carried out, the filter cake is washed by the n-hexane (100mL), the filtrate is placed in a refrigerator to be refrigerated for 30 minutes at the temperature of 2-8 ℃, the filter cake is taken out and filtered, the n-hexane (100mL) is used for washing the filter cake, and the operation is repeated for 5 times to obtain the solid product. It was dried in an oven. Obtaining products 25-189: 23.9g, yield: 74.34 percent.

25-199

Boc-glycine (Boc-Gly-OH, 7.9647g, 45.4661mmol), 25-189(23.9g, 37.8884mmol) and DMAP (0.9258g, 7.5777mmol) were added to a 500mL round bottom flask, after dissolution with dichloromethane (100mL), the reaction flask was stirred at 0 ℃ for about 30 minutes, followed by dicyclohexylcarbodiimide (DCC, 15.6350g, 75.7767mmol) and the reaction flask was stirred at 0 ℃ for 3 hours. After the reaction was completed, the reaction solution was first filtered to remove DCC, and the filter cake was washed with dichloromethane (60mL), and the filtrate was collected, evaporated to dryness, and dried. Obtaining products 25-199: 29.8549 g.

25-200

25-199(29.8549g, 37.8884mmol) was charged into a 100mL round-bottom flask, methylene chloride (20mL) was added and dissolved, TFA (42.2mL, 568.326mmol) was added with stirring, and the reaction flask was stirred at room temperature overnight. After the reaction is finished, the reaction solution is firstly decompressed, concentrated and evaporated to dryness to remove dichloromethane, then n-hexane (150mL) is added into the reaction solution for sedimentation, the supernatant is poured, n-hexane (150mL) is added into the lower oily substance for sedimentation, and the operation is repeated three times to obtain oily solid. The viscous oily solid was dissolved in dichloromethane (20mL), silica gel powder (100mL) was added, evaporated to dryness, and subjected to dry loading and column chromatography. Eluting with eluent (1% -4% methanol: 99% -96% dichloromethane), collecting, concentrating, evaporating, and drying. Obtaining products 25-200: 23.4557g, yield: 90.04 percent.

25-211

Boc-Gly-OH (purchased from alatin, 3.0g, 17.1252mmol), HBTU (9.7418g, 25.6878mmol), HOBT (3.4709g, 25.6878mmol) and H-Glu (OBzl) -OBzl.TosOH (8.5554g, 17.1252mmol) were added to a 500mL round bottom flask, dissolved in DMF (50mL), the reaction flask was then left to stir at-5 ℃ for about 30 minutes, DIEA (12.7mL, 77.0634mmol) was then added slowly dropwise, after which the reaction flask was allowed to continue to stir at-5 ℃ for 1 hour and finally transferred to room temperature for 2 hours. After completion of the reaction, the reaction solution was transferred to a 2L separatory funnel, and a saturated sodium bicarbonate solution (400mL) and ethyl acetate (300mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase and the organic phase. Saturated sodium chloride solution (300mL) was then added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Saturated sodium chloride (300mL) was then added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. The organic phase is concentrated and evaporated to dryness. Dissolving the extract with 20% methanol/dichloromethane (50mL), adding silica gel powder (50mL), evaporating, dry-loading, and performing column chromatography. Eluting with eluent (1% -2% methanol: 99% -98% ethyl acetate), collecting liquid, concentrating, evaporating, and drying. Obtaining products 25-211: 6.9213g, yield: 83.41 percent.

25-220

Adding 25-211(2.6890g, 5.5382mmol) and 10% Pd/C (50mg) into a hydrogenation reaction kettle, adding DMF (30mL) to dissolve, sealing the hydrogenation reaction kettle, introducing hydrogen to make the pressure in the kettle reach 1.6MPa, and placing the hydrogenation reaction kettle at room temperature to stir overnight. After the reaction is finished, the reaction kettle is taken out, the reaction solution is uniformly dripped into a sand core funnel filled with compacted diatomite, the filtration is carried out, and the diatomite is cleaned by DMF (60mL) until the diatomite does not contain the product, thus obtaining the reaction product solution.

25-222

25-220(1.6853g, 5.5382mmol), HBTU (6.3009g, 16.6146mmol), HOBT (2.2450g, 16.6146mmol) and 25-200(8.3810g, 12.1841mmol) were added to a 500mL round bottom flask, dissolved in DMF (50mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (8.2mL, 49.8438mmol) was slowly added dropwise, after which the reaction flask was stirred at-5 ℃ for 1 hour and finally transferred to room temperature for 2 hours. After the reaction, the reaction mixture was transferred to a 1L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (200mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase and the organic phase. Saturated sodium chloride solution (300mL) was then added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Deionized water (300mL) was then added to the organic phase, shaken, and extracted. Finally, the organic phase is concentrated and evaporated to dryness and then is dried in a vacuum oven. Obtaining products 25-222: 9.0g, yield: 98.85 percent.

MALDI-TOF MS:[M-H⁺]1641.76。

25-224

25-222(9.0g, 5.4745mmol) was placed in a 100mL round-bottom flask, dichloromethane (10mL) was added and dissolved, TFA (6.1mL, 82.1178mmol) was added with stirring, and the reaction flask was left to stir at room temperature overnight. After the reaction, the reaction mixture was concentrated under reduced pressure to dryness to remove methylene chloride, and then transferred to a 2L separatory funnel, and a saturated sodium chloride solution (400mL) and ethyl acetate (200mL) were added thereto, shaken, extracted, and the aqueous phase and the organic phase were separated. Deionized water (300mL) was then added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. The organic phase is concentrated and evaporated to dryness. Dissolving the extract with 20% methanol/dichloromethane (70mL), adding 50mL silica gel powder, evaporating, dry-loading, and performing column chromatography. Eluting with eluent (2% -7% methanol: 98% -93% ethyl acetate), collecting liquid, concentrating, evaporating, and drying. Obtaining products 25-224: 6.0g, yield: 70.99 percent.

MALDI-TOF MS:[M+Na⁺]1565.64。

25-203

Fmoc-Glu (OtBu) -OH (available from Ark Pharm, 4.0646g, 9.5134mmol), HBTU (5.4118g, 14.2701mmol), HOBT (1.9282g, 14.2701mmol) and 25-201 (synthesized by 22-181, 9.1g, 9.9890mmol) were added to a 500mL round-bottomed flask, dissolved in DMF (60mL), the reaction flask was then placed at-5 ℃ and stirred for about 30 minutes, DIEA (7.1mL, 42.8103mmol) was slowly added dropwise, after which the reaction flask was allowed to continue to stir at-5 ℃ for 2 hours, and finally transferred to room temperature and stirred overnight. After the reaction, the reaction mixture was transferred to a 2L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (200mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase and the organic phase. Deionized water (300mL) was then added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated and evaporated to dryness and then is dried in a vacuum oven. Obtaining products 25-203: 12.5430 g.

25-210

25-203(12.5430g, 9.5134mmol) was placed in a 100mL round-bottom flask, dichloromethane (10mL) was added to dissolve, trifluoroacetic acid (TFA, 10.6mL, 142.701mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature overnight. After the reaction, the reaction mixture was concentrated under reduced pressure and evaporated to dryness to remove methylene chloride. Then, the reaction solution was transferred to a 1L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (300mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Saturated sodium chloride solution (300mL) was added to the organic phase, shaken, extracted and the aqueous phase separated. Next, deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous phase separated. Finally, the organic phase is concentrated to dryness. Dissolving the extract with 20% methanol/dichloromethane mixed solvent (50mL), adding silica gel powder (50mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (1% -5% methanol: 99% -95% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 25-210: 9.0g, yield: 74.94 percent.

MALDI-TOF MS:[M+Na⁺]1284.24。

25-221

25-210(9.0g, 7.1295mmol), HBTU (3.9376g, 10.3828mmol), HOBT (1.4029g, 10.3828mmol) and ethylenediamine monohydrate (from TCl, 0.2mL, 3.4609mmol) were added to a 500mL round bottom flask, dissolved in DMF (60mL), stirred at-5 ℃ for about 30 minutes, then DIEA (5.1mL, 31.1483mmol) was added slowly dropwise, after which the reaction was allowed to continue stirring at-5 ℃ for 2 hours and then allowed to stir at room temperature overnight. After completion of the reaction, the reaction mixture was transferred to a 1L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (300mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Saturated sodium chloride solution (300mL) was added to the organic phase, shaken, extracted and the aqueous phase separated. Next, deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous phase separated. Finally, the organic phase is concentrated and evaporated to dryness and then is dried in a vacuum oven. Obtaining products 25-221: 8.8211 g.

MALDI-TOF MS:[M+Na⁺]2570.22。

25-226

25-221(8.8211g, 3.4609mmol) was charged into a 500mL round-bottomed flask, DMF (10mL) was added and dissolved, morpholine (4.5mL, 51.9135mmol) was added with stirring, and the reaction was stirred at room temperature for 2 hours. After completion of the reaction, the reaction mixture was transferred to a 1L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (300mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Saturated sodium chloride solution (300mL) was added to the organic phase, shaken, extracted and the aqueous phase separated. Next, deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous phase separated. Finally, the organic phase is concentrated and evaporated to dryness and then is dried in a vacuum oven. Obtaining products 25-226: 7.2828 g.

MALDI-TOF MS:[M+Na⁺]2125.39。

25-227

25-226(3.2368g, 1.5382mmol), HBTU (1.7500g, 4.6146mmol), HOBT (0.6235g, 4.6146mmol) and 25-207 (synthesized by 25-102) (1.7426g, 3.5379mmol) were added to a 500mL round bottom flask, after dissolution in DMF (60mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, then DIEA (2.3mL, 13.8438mmol) was slowly added dropwise, after which the reaction flask was stirred at-5 ℃ for 1 hour and finally transferred to room temperature for 2 hours. After completion of the reaction, the reaction mixture was transferred to a 2L separatory funnel, and a saturated sodium bicarbonate solution (300mL) and ethyl acetate (200mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Saturated sodium chloride solution (300mL) was added to the organic phase, shaken, extracted and the aqueous phase separated. Then, deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous phase was separated. Finally, the organic phase is concentrated and evaporated to dryness. Dissolving the extract with 20% methanol/dichloromethane mixed solvent (50mL), adding silica gel powder (50mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (1% ammonia water: 2% -6% methanol: 97% -93% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 25-227: 2.0618g, yield: 43.9 percent.

MALDI-TOF MS:[M+Na⁺]3076.43。

25-236

25-227(0.6519g, 0.2135mmol) and 10% Pd/C (50mg) were put into a hydrogenation reactor, DMF (30mL) was added to dissolve the mixture, the hydrogenation reactor was closed, hydrogen was introduced to the reactor so that the pressure in the reactor became 1.6MPa, and the hydrogenation reactor was left at room temperature and stirred overnight. After the reaction is finished, the reaction kettle is taken out, the reaction solution is uniformly dripped into a sand core funnel filled with compacted diatomite, the filtration is carried out, and the diatomite is cleaned by DMF (50mL) until the diatomite does not contain the product, thus obtaining the reaction product solution.

25-238

A solution of 25 to 236 (0.4980g, 0.2135mmol), HBTU (0.9716g, 2.5620mmol), HOBT (0.3462g, 2.5620mmol) and 25 to 224(2.9g, 1.8784mmol) were added to a 500mL round bottom flask, which was then stirred for about 30 minutes at-5 ℃ after dissolution with DMF (30mL), DIEA (1.3mL, 7.6860mmol) was slowly added dropwise, and after the addition, the reaction flask was stirred at-5 ℃ for 3 hours. After completion of the reaction, the reaction mixture was transferred to a 1L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (200mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Then, deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous phase was separated. Finally, the organic phase is concentrated to dryness. Dissolving the extract with 20% methanol/dichloromethane mixed solvent (50mL), adding silica gel powder (30mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (3% -10% methanol: 97% -90% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 25-238: 1.9233g, yield: 61.96 percent.

25-243

25-238(1.9233g, 0.1323mmol) was placed in a 100mL round-bottom flask, dichloromethane (5.0mL) was added and dissolved, trifluoroacetic acid (TFA, 0.2mL, 1.9843mmol) was added with stirring, and the reaction flask was left to stir at room temperature overnight. After the reaction, the reaction mixture was concentrated and evaporated to dryness to remove methylene chloride. Then, the reaction solution was transferred to a 1L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (300mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Saturated sodium chloride solution (300mL) was added to the organic phase, shaken, extracted and the aqueous phase separated. Finally, the organic phase is concentrated to dryness. Dissolving the extract with 20% methanol/dichloromethane mixed solvent (50mL), adding 30mL silica gel powder, evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (1% -5% methanol: 99% -95% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 25-243: 0.7924g, yield: 71.01 percent.

25-260

25-243(0.4738g, 0.033mmol) and M-SCM-40K (available from Jenkem Technology, 3.0g, 0.0727mmol) were added to a 500mL round-bottomed flask, dissolved in DMF (20mL), and reacted at room temperature with exclusion of light for one week. After the reaction, n-hexane (100mL) and methyl tert-butyl ether (10mL) were added to the reaction solution, and the mixture was settled, the supernatant was decanted, and n-hexane (300mL) and methyl tert-butyl ether (50mL) were added to the lower oily liquid phase, and the settling was continued, and this was repeated 3 times to give a viscous oily substance. Dichloromethane (5mL) was then added to the viscous mass, followed by methyl tert-butyl ether (60mL) and allowed to settle as a powdery solid which was filtered. The filter cake was washed with methyl tert-butyl ether (60mL) to give the solid product. The solid product was dissolved in a 20% methanol/dichloromethane mixed solvent (50mL), silica gel powder (40mL) was added, evaporated to dryness, dry loaded, and subjected to column chromatography. Eluting with eluent (4% -12% methanol: 96% -88% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 25-260: 1.6g, yield: 50.17 percent.

44-2

25-260(0.8233g, 0.0085mmol) was dissolved in THF (20 mL); taking TBAF.3H₂O (0.0860g, 0.2726mmol) was dissolved in THF (10 mL); the THF solution containing TBAF was added to the THF solution containing 25-260 and a certain amount of diluted hydrochloric acid solution (30mL, 0.05mol/L) was added, and the reaction was stirred at room temperature overnight protected from light. After the reaction is finished, the reaction solution is added,firstly, evaporating reaction liquid to dryness, adding absolute ethyl alcohol (20mL), decompressing, concentrating and evaporating to dryness, and circulating for 3 times; dissolving the solid obtained by the last step of evaporation by distillation in DMF (1.0mL), adding isopropanol (50mL) for sedimentation to form powdery solid, filtering, dissolving the filter cake again in DMF (3.0mL), adding isopropanol (50mL) for sedimentation to form powdery solid, filtering, and washing the filter cake with isopropanol (30 mL); the solid in the second step, dichloromethane (2.0mL) was dissolved and methyl tert-butyl ether (50mL) was added to settle to form a powdered solid, filtered, the filter cake was dissolved again with dichloromethane (2.0mL), methyl tert-butyl ether (50mL) was added to settle to form a powdered solid, filtered, the filter cake was washed with methyl tert-butyl ether (30mL) and dried. To obtain a product 44-2: 0.5519g, yield: 69.79 percent.

¹H-NMR(600MHz,DMSO-d₆)δ9.45-9.29(m,80H),9.09-7.93(m,103H),7.65-7.56(m,20H),7.30-7.19(m,81H),6.97-6.95(s,40H),5.46-5.42(m,59H),5.15-5.12(m,87H),4.05-3.54(m,6944H),2.93-2.89(m,97H),2.74–2.66(m,39H),2.34-2.31(m,20H),2.09-2.06(m,106H),1.90-1.72(m,104H),1.48-1.45(m,31H),1.31-1.21(m,119H),0.95-0.84(m,96H).

7. Synthesis of 42-52 (Compound No. 1)

The route is as follows

42-20

Fmoc-Glu-OtBu (available from Accela, 3.2226g, 7.5741mmol), HBTU (4.3086g, 11.3612mmol), HOBT (1.5351g, 11.3612mmol) and 44-19 (synthesized by 22-181, 6.9g, 7.5741mmol) were added to a 500mL round-bottomed flask, dissolved in DMF (60mL), the reaction was stirred at-5 ℃ for about 30 minutes, DIEA (7.5mL, 45.4460mmol) was slowly added dropwise, and after addition, the reaction flask was stirred at-5 ℃ for 2 hours. After the reaction, the reaction mixture was transferred to a 2L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (200mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Then, a saturated sodium chloride solution (200mL) was added to the organic phase, and the aqueous phase was separated by shaking and extraction. Finally, the organic phase is concentrated to dryness. Dissolving the extract with 20% methanol/dichloromethane mixed solvent (50mL), adding silica gel powder (80mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (4% -12% methanol: 96% -88% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-20: 6.9g, yield: 69.10 percent.

42-25

42-20(5.0g, 3.7923mmol) was placed in a 500mL round-bottom flask, dichloromethane (10mL) was added and dissolved, TFA (4.2mL, 56.8845mmol) was added with stirring, and the reaction was stirred at room temperature overnight. After the reaction, the reaction mixture was concentrated under reduced pressure and evaporated to dryness to remove methylene chloride. The reaction solution was transferred to a 2L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (200mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Then, a saturated sodium chloride solution (200mL) was added to the organic phase, and the aqueous phase was separated by shaking and extraction. Deionized water (300mL) was then added to the organic phase, shaken, extracted, and the aqueous phase separated. The organic phase is concentrated to dryness and dried in a vacuum oven. Obtaining products 42-25: 4.7872 g.

33-143

1, 2-bis (2-aminoethoxy) ethane (purchased from TCl, 50mL, 340.7327mmol) was charged into a 500mL flask, methylene chloride (150mL) was added and dissolved, triethylamine (94.8928mL, 681.4654mmol) was added, di-tert-butyl dicarbonate (purchased from Innochem, 74.3751g, 340.7321mmol) was slowly added dropwise with stirring at room temperature, and after the addition, the mixture was stirred at room temperature overnight. And after the reaction is finished, adding silica gel powder, evaporating to obtain a powdery solid, loading the powdery solid by a dry method, carrying out column chromatography, eluting by a mixed solution of 1% of ammonia water and 2% -3% of methanol in dichloromethane, collecting, concentrating and drying to obtain the product 33-143 with the yield of 10%.

42-27

42-25(4.7872g, 3.7923mmol), HBTU (2.1573g, 5.6885mmol), HOBT (0.7686g, 5.6885mmol) and 33-143(0.9888g, 3.9819mmol) were added to a 500mL round bottom flask, dissolved in DMF (60mL), the reaction was stirred at-5 ℃ for about 30 minutes, DIEA (2.8mL, 17.0654mmol) was slowly added dropwise, and after the addition, the reaction flask was stirred at-5 ℃ for 2 hours. After the reaction, the reaction mixture was transferred to a 2L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (200mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Then, a saturated sodium chloride solution (200mL) was added to the organic phase, and the aqueous phase was separated by shaking and extraction. Finally, the organic phase is concentrated and evaporated to dryness and placed in a vacuum oven for drying. Obtaining products 42-27: 5.4g, yield: 95.40 percent.

42-30

42-27(5.4g, 3.6177mmol) was placed in a 500mL round-bottom flask, DMF (10mL) was added and dissolved, morpholine (4.7mL, 54.2635mmol) was added with stirring, and the reaction flask was left to react with stirring at room temperature for 2 hours. After the reaction was completed, the reaction solution was transferred to a 2L separatory funnel, and a saturated sodium chloride solution (300mL) and ethyl acetate (200mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Then, a saturated sodium chloride solution (200mL) was added to the organic phase, and the aqueous phase was separated by shaking and extraction. Finally, the organic phase is concentrated to dryness. Dissolving the extract with 20% methanol/dichloromethane mixed solvent (50mL), adding silica gel powder (80mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (1% ammonia water: 1% -6% methanol: 98% -93% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-30: 3.0140g, yield: 65.58 percent.

42-34

42-30(3.0140g, 2.3724mmol), HBTU (1.2269g, 3.2352mmol), HOBT (0.4371g, 3.2352mmol) and glutaric acid (0.1425g, 1.0784mmol) were added to a 500mL round bottom flask, dissolved in DMF (50mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (1.6mL, 9.7056mmol) was slowly added dropwise, and after the addition, the reaction flask was stirred at-5 ℃ for 2 hours and finally transferred to room temperature and stirred overnight. After completion of the reaction, the reaction mixture was transferred to a 2L separatory funnel, and a saturated sodium bicarbonate solution (300mL) and ethyl acetate (200mL) were added thereto, followed by shaking, extraction and separation of the aqueous phase. Then, a saturated sodium chloride solution (200mL) was added to the organic phase, and the aqueous phase was separated by shaking and extraction. Finally, the organic phase is concentrated to dryness. Dissolving the extract with 20% methanol/dichloromethane mixed solvent (50mL), adding silica gel powder (50mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (1% ammonia water: 2% -5% methanol: 97% -94% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-34: 2.8437g, yield: 100 percent.

¹H-NMR(400MHz,DMSO-d₆)δ8.60–8.22(m,5H),8.19–7.63(m,11H),7.55–7.11(m,40H),5.10-5.07(m,16H),3.56-3.47(m,20H),3.19-3.17(m,8H),3.09–3.01(m,4H),2.99–2.65(m,16H),2.43-2.42(m,8H),2.17-2.09(m,32H),1.36-1.34(m,18H).

42-15

42-14 solution (synthesized by 25-102, 5.5320g, 11.2308mmol), HBTU (5.8080g, 15.3147mmol), HOBT (2.0693g, 15.3147mmol) and PKA (PKA is a truncated bis-methyl derivative of PKI-587, a new clinical drug, 6.0g, 10.2098mmol) were added to a 500mL round-bottomed flask, dissolved in DMF (80mL), the reaction flask was then left to stir at-5 ℃ for about 30 minutes, DIEA (7.6mL, 45.9441mmol) was slowly added dropwise, after which the reaction was allowed to continue stirring at-5 ℃ for 1 hour and finally allowed to shift to room temperature and stir overnight. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction solution to settle, the supernatant was poured, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower liquid to settle, and the supernatant was poured, and this was repeated 3 times to obtain a viscous substance. The viscous mass was then dissolved in dichloromethane (5mL), and methyl tert-butyl ether (100mL) was added to settle to a solid as a powder, which was filtered, the filter cake was washed with methyl tert-butyl ether (60mL), and the filter cake was dried in a vacuum oven. Obtaining products 42-15: 10.8450 g.

42-18

42-15(10.8450g, 10.2098mmol) was placed in a 500mL round-bottom flask, dichloromethane (5mL) was added and dissolved, TFA (11.4mL, 153.1470mmol) was added with stirring, and the reaction flask was stirred at room temperature overnight. After the reaction, the reaction mixture was concentrated under reduced pressure and evaporated to dryness to remove methylene chloride. Methyl tert-butyl ether (150mL) was added to the reaction solution and allowed to settle to form a powdery solid, which was filtered, the filter cake was washed with methyl tert-butyl ether (60mL), and the filter cake was dried in an oven. Obtaining products 42-18: 9.8230g, yield: 100 percent.

42-24

42-20(1.8683g, 1.4170mmol) and 10% Pd/C (50mg) are put into a hydrogenation reaction kettle, DMF (30mL) is added to dissolve the mixture, the hydrogenation reaction kettle is sealed, then hydrogen is introduced to ensure that the pressure in the hydrogenation reaction kettle reaches 1.6MPa, and finally the hydrogenation reaction kettle is placed at room temperature and stirred overnight. After the reaction is finished, the reaction kettle is taken out, the reaction solution is uniformly dripped into a sand core funnel filled with compacted diatomite, the filtration is carried out, and the diatomite is cleaned by DMF (60mL) until the diatomite does not contain the product, thus obtaining the reaction product solution.

42-26

42-24 solution (1.3574g, 1.4170mmol), HBTU (3.2243g, 8.5020mmol), HOBT (1.1488g, 8.5020mmol) and 42-18(8.0g, 6.2350mmol) were added to a 500mL round bottom flask, dissolved in DMF (80mL), and the reaction flask was stirred at-5 ℃ for about 30 minutes, followed by the slow dropwise addition of DIEA (4.2mL, 25.5060mmol) and, after the dropwise addition, the reaction flask was stirred at-5 ℃ for 3 hours. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction solution to settle, the supernatant was poured, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower liquid to settle, and the supernatant was poured, and this was repeated 3 times to obtain a viscous substance. The viscous mass was then dissolved in dichloromethane (5mL), and methyl tert-butyl ether (100mL) was added to settle to a solid as a powder, which was filtered, the filter cake was washed with methyl tert-butyl ether (60mL), and the filter cake was dried in a vacuum oven. Obtaining products 42-26: 6.7086 g.

42-28

42-26(6.7086g, 1.4170mmol) was placed in a 100mL round-bottom flask, dichloromethane (10mL) was added and dissolved, and TFA (1.6mL, 21.2550mmol) was added with stirring, and the reaction was stirred at room temperature overnight. After the reaction is finished, the reaction solution is decompressed, concentrated and evaporated to dryness to remove dichloromethane; methyl tert-butyl ether (150mL) was added and the mixture was allowed to settle to a powdery solid, filtered and the filter cake washed with methyl tert-butyl ether (60 mL). The filter cake was dissolved in 20% methanol/dichloromethane (60mL), silica gel powder (70mL) was added, evaporated to dryness, dry loaded and chromatographed. Eluting with eluent (2% -11% methanol: 98% -89% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-28: 5.4g, yield: 81.46 percent.

42-32

42-28(4.4g, 0.9405mmol), HBTU (0.5350g, 1.4108mmol), HOBT (0.1906g, 1.4108mmol) and 25-254 (synthesized by 30-33) (0.8504g, 1.0346mmol) were added to a 500mL round bottom flask, dissolved in DMF (60mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (0.7mL, 4.2324mmol) was slowly added dropwise, and after the addition, the reaction flask was stirred at-5 ℃ for 3 hours. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction solution to settle, the supernatant was poured, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower liquid to settle, and the supernatant was poured, and this was repeated 3 times to obtain a viscous substance. The viscous mass was then dissolved in dichloromethane (5mL), and methyl tert-butyl ether (100mL) was added to settle to a solid as a powder, which was filtered and the filter cake was washed with methyl tert-butyl ether (60 mL). The filter cake was dissolved in 20% methanol/dichloromethane (60mL), silica gel powder (50mL) was added, evaporated to dryness, dry loaded and chromatographed. Eluting with eluent (3% -15% methanol: 97% -85% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-32: 3.6g, yield: 69.82 percent.

42-36

42-32(3.6g, 0.6567mmol) was placed in a 500mL round-bottom flask, DMF (30mL) was added and dissolved, morpholine (0.9mL, 9.8505mmol) was added with stirring, and the reaction flask was left to react with stirring at room temperature for 2 hours. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction solution to settle, the supernatant was poured, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower liquid to settle, and the supernatant was poured, and this was repeated 3 times to obtain a viscous substance. The viscous mass was then dissolved in dichloromethane (5mL), and methyl tert-butyl ether (100mL) was added to settle to a solid as a powder, which was filtered and the filter cake was washed with methyl tert-butyl ether (60 mL). The filter cake was dissolved in 20% methanol/dichloromethane (60mL), silica gel powder (30mL) was added, evaporated to dryness, dry loaded and chromatographed. Eluting with eluent (1% ammonia water: 2% -7% methanol: 97% -92% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-36: 3.2g, yield: 92.64 percent.

42-40

42-36(3.2g, 0.6084mmol), HBTU (0.3461g, 0.9126mmol), HOBT (0.1233g, 0.9126mmol) and Boc-LC-OH (synthesized by 24-36, 0.1762g, 0.6692mmol) were added to a 500mL round bottom flask, dissolved in DMF (60mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (0.5mL, 2.7378mmol) was added slowly and dropwise, after addition, the reaction flask was stirred at-5 ℃ for 1 hour and finally transferred to room temperature and stirred overnight. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction solution to settle, the supernatant was poured, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower liquid to settle, and the supernatant was poured, and this was repeated 3 times to obtain a viscous substance. The viscous mass was then dissolved in dichloromethane (5mL), and methyl tert-butyl ether (100mL) was added to settle to a solid as a powder, which was filtered, the filter cake was washed with methyl tert-butyl ether (60mL), and the filter cake was dried in a vacuum oven. Obtaining products 42-40: 3.3473 g.

42-41

42-40(3.3473g, 0.6084mmol) was placed in a 100mL round-bottom flask, dichloromethane (10mL) was added and dissolved, TFA (0.7mL, 9.1260mmol) was added with stirring, and the reaction flask was stirred at room temperature overnight. After the reaction, the reaction mixture was concentrated under reduced pressure and evaporated to dryness to remove methylene chloride. To this was added methyl tert-butyl ether (100mL) and the mixture was allowed to settle to a solid in the form of a powder, filtered, and the filter cake was washed with methyl tert-butyl ether (60 mL). The filter cake was dissolved in 20% methanol/dichloromethane (60mL), silica gel powder (40mL) was added, evaporated to dryness, dry loaded and chromatographed. Eluting with eluent (1% ammonia water: 3% -10% methanol: 96% -89% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-41: 2.1g, yield: 63.86 percent.

42-37

42-34(1.0g, 0.3792mmol) and 10% Pd/C (40mg) were placed in a hydrogenation reactor, DMF (30mL) was added to dissolve, the hydrogenation reactor was closed, hydrogen was then introduced to bring the pressure in the reactor to 1.6MPa, and finally the hydrogenation reactor was left at room temperature and stirred overnight. After the reaction is finished, the reaction kettle is taken out, the reaction solution is uniformly dripped into a sand core funnel filled with compacted diatomite, the filtration is carried out, and the diatomite is cleaned by DMF (60mL) until the diatomite does not contain the product, thus obtaining the reaction product solution.

42-43

42-41(2.1g, 0.3885mmol), HBTU (0.2007g, 0.5292mmol), HOBT (0.0715g, 0.5292mmol) and 42-37 solution (0.0845g, 0.0441mmol) were added to a 500mL round bottom flask, after dissolution in DMF (50mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (0.3mL, 1.5876mmol) was slowly added dropwise, after which the reaction flask was stirred at-5 ℃ for 1 hour and finally transferred to room temperature overnight. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction solution to settle, the supernatant was poured, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower liquid to settle, and the supernatant was poured, and this was circulated 5 times to obtain a viscous substance. The viscous mass was then dissolved in dichloromethane (5mL), and methyl tert-butyl ether (100mL) was added to settle to a solid as a powder, which was filtered and the filter cake was washed with methyl tert-butyl ether (60 mL). Dissolving the filter cake with 20% methanol/dichloromethane mixed solvent (80mL), adding silica gel powder (30mL), evaporating to dryness, loading by dry method, and performing column chromatography. Eluting with eluent (1% ammonia water: 4% -8% methanol: 95% -91% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-43: 1.0g, yield: 50.38 percent.

¹H-NMR(600MHz,DMSO-d₆)δ10.31-10.27(m,52H),8.53–7.91(m,88H),7.80-7.71(m,28H),7.62–7.26(m,55H),7.04-6.98(m,32H),5.76(s,48H),5.44-5.22(m,80H),4.78–4.27(m,76H),4.07-4.02(m,61H),3.92–3.20(m,2292H),3.17-3.03(m,53H),2.67-2.59(m,118H),2.33-2.09(m,98H),1.78-1.61(m,54H),1.41–0.70(m,127H).

42-50

42-43(1.0g, 0.0222mmol) was placed in a 250mL round-bottom flask, dichloromethane (5mL) was added and dissolved, TFA (0.1mL, 0.6615mmol) was added under stirring, and the reaction flask was left to stir at room temperature overnight. After the reaction, the reaction mixture was concentrated and evaporated to dryness to remove methylene chloride. Additional methyl tert-butyl ether (150mL) was added and the mixture was allowed to settle to form a powdery solid, which was filtered and the filter cake washed with methyl tert-butyl ether (60 mL). The filter cake was dissolved in 20% methanol/dichloromethane (50mL), silica gel powder (30mL) was added, evaporated to dryness, dry loaded and chromatographed. Eluting with eluent (1% ammonia water: 4% -10% methanol: 95% -89% dichloromethane), collecting liquid, concentrating, and evaporating to dryness. Obtaining products 42-50: 0.4772g, yield: 47.97 percent.

42-52

42-50(0.4472g, 0.0106mmol) and M-SCM-20K (0.5004g, 0.0233mmol) were placed in a 500mL round-bottomed flask, dissolved in DMF (30mL), and then left to react at room temperature with exclusion of light for 7 days. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction solution to settle, the supernatant was poured, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower liquid to settle, and the supernatant was poured, and this was repeated 3 times to obtain a viscous substance. The viscous mass was then dissolved in dichloromethane (5mL), and methyl tert-butyl ether (100mL) was added to settle to a solid as a powder, which was filtered and the filter cake was washed with methyl tert-butyl ether (60 mL). Dissolving the filter cake with 20% methanol/dichloromethane mixed solvent (80mL), adding silica gel powder (40mL), evaporating to dryness, loading by dry method, and performing column chromatography. Eluting with eluent (1% ammonia water: 4% -10% methanol: 95% -89% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-52: 0.547g, yield: 58.96 percent.

¹H-NMR(600MHz,DMSO-d₆)δ9.36–8.90(m,144H),8.53–7.94(m,349H),7.94–7.48(m,250H),7.32-7.22(m,231H),7.15-7.09(m,27H),7.03-6.96(m,16H),4.54-4.23(m,380H),4.10–3.36(m,3418H),3.19-3.03(m,106H),2.90-2.73(m,50H),2.63-2.59(m,380H),2.41-2.30(m,61H),2.22-2.12(m,95H),1.98-1.49(m,288H),1.40-1.35(m,59H),1.15-1.03(m,75H),0.91-0.85(m,240H).

8. Synthesis of 37-200 (Compound No. 2)

The route is as follows

37-192

Pentamine dithionate (4.33g, 13.20mmol, available from Conrong chemical) is put into a 500mL flask, dichloromethane (30mL) is added to dissolve the dithionate, triethylamine (14.8mL, 105.6mmol) is added, the reaction is stirred for 30 minutes at 0 ℃, phenyl chloroformate (9.95mL, 79.2mmol) is slowly and dropwise added, the reaction is stirred overnight at room temperature, the reaction is finished, the reaction solution is transferred into a 1L separating funnel, dichloromethane (200mL) and deionized water (200mL) are added to extract to obtain an organic phase, a water phase is washed by dichloromethane (200mL x 1), the organic phases are combined, silica gel powder (15g) is added, powdery solid is evaporated to dryness, a sample is loaded, column chromatography is carried out, and elution is carried out by using a mixed solution of 5% methanol and dichloromethane, so that 37-192:3.3g of a product is obtained, and the yield is 42%.

37-45

35-31(2.73g, 2.196mmol, synthesized by the method 34-17), tert-butyl succinate (0.459g, 2.635mmol, ex Ilykok), HBTU (1.249g, 3.294mmol), HOBT (0.445g, 3.294mmol) were added to a 500mL flask, dissolved in DMF (70mL), the reaction was stirred at-5 ℃ for about 30 minutes, DIEA (1.633mL, 9.882mmol) was slowly added dropwise, and after the addition was complete, the reaction was stirred at-5 ℃ for 3 hours. After completion of the reaction, the reaction mixture was transferred to a 1L separatory funnel, and deionized water (200mL) and ethyl acetate (200mL) were added thereto, followed by shaking and extraction. The aqueous phase was washed with ethyl acetate (150mL x 2), the organic phases were combined, washed with saturated brine (200mL x 1), concentrated to dryness and dried in a vacuum oven to give product 37-45: 3.1g.

¹H-NMR(400MHz,DMSO-d₆)δ8.52(d,J＝7.3Hz,1H),8.30(d,J＝7.4Hz,1H),7.99–7.93(m,4H),7.88(d,J＝7.3Hz,2H),7.68(t,J＝7.2Hz,3H),7.51(t,J＝7.6Hz,1H),7.43–7.18(m,22H),5.16–5.01(m,7H),4.45–4.07(m,7H),3.70–3.46(m,4H),3.43–3.29(m,10H),3.24–3.12(m,4H),2.46–2.28(m,7H),2.17(t,J＝7.9Hz,2H),1.39–1.24(m,9H).

47-19

37 to 45(0.5137g, 0.3624mmol) was charged into a hydrogenation reactor, and Pd/C (0.010g) was added thereto, and DMF (30mL) was added thereto and dissolved therein, and hydrogen gas was introduced under a pressure Pa of 1.8MPa, and the reaction was stirred at room temperature overnight. After the reaction was completed, the reaction vessel was taken out, the reaction solution was filtered through celite, washed with DMF (20mL × 3), and the DMF solutions were combined as the raw material for the next reaction.

47-21

47-19(0.38g, 0.3624mmol), 37-2(3g, 1.5944mmol), HBTU (0.8246g, 2.1744mmol) and HOBT (0.2938g, 2.1744mmol) were charged into a 250mL flask, DMF (95mL) was added to dissolve the mixture, the reaction was left at-5 ℃ and stirred for 20 minutes, DIEA (1.078mL, 6.523mmol) was slowly added dropwise, the reaction was stirred at-5 ℃ for 30 minutes after completion of the addition, and then stirred at room temperature overnight. After the reaction is finished, adding n-hexane (120mL) and methyl tert-butyl ether (30mL), settling for 3 times to obtain a viscous oily substance, adding methyl tert-butyl ether (250mL), separating out a solid, filtering, washing a filter cake with methyl tert-butyl ether (50mL x 3), dissolving the filter cake with dichloromethane (150mL) and methanol (30mL), adding silica gel powder (10g), evaporating to obtain a powdery solid, loading by a dry method, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 4% -8% methanol, collecting, concentrating and drying to obtain a product 47-21:2.5g, wherein the yield is 83%.

43-123

47-21(2.5g, 0.2938mmol) was charged into a 250mL flask, DMF (30mL) was added and dissolved, morpholine (7.67mL, 88.14mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction is finished, adding n-hexane (100mL) and methyl tert-butyl ether (20mL) for settling, carrying out layering, pouring an upper layer, adding n-hexane (100mL) and methyl tert-butyl ether (20mL) into a lower oily liquid phase, continuing to settle, repeating the process for 3 times, finally adding methyl tert-butyl ether (150mL) into a viscous oily substance, separating out a solid, filtering, washing a filter cake with methyl tert-butyl ether (40mL x 2), dissolving the filter cake with dichloromethane (20mL) and methanol (80mL), adding silica gel powder (10g), evaporating to dryness to obtain a powdery solid, carrying out dry loading, carrying out column chromatography, and eluting with a dichloromethane mixed solution of 1% ammonia water and 6% -10% methanol to obtain a product 43-123:1.85g, wherein the yield is 77%.

37-195

43-123(1.85g, 0.2628mmol), 37-192(0.0326g, 0.0531mmol) were charged to a 250mL flask, DMF (30mL) was added to dissolve, triethylamine (0.0336mL, 0.239mmol) was added, and the reaction was mechanically stirred at 80 ℃ overnight. After the reaction is finished, adding n-hexane (100mL) and methyl tert-butyl ether (20mL) for settling, carrying out layering, pouring an upper layer, adding n-hexane (100mL) and methyl tert-butyl ether (20mL) into a lower oily liquid phase, continuing to settle, repeating the process for 3 times, finally adding methyl tert-butyl ether (150mL) into a viscous oily substance, separating out a solid, filtering, washing a filter cake with methyl tert-butyl ether (40mL x 2), dissolving the filter cake with dichloromethane (20mL) and methanol (80mL), adding silica gel powder (10g), evaporating to dryness to obtain a powdery solid, carrying out dry loading, carrying out column chromatography, and eluting with a dichloromethane mixed solution of 1% ammonia water and 6% -7% methanol to obtain a product 37-195:0.7g, wherein the yield is 40%.

37-199

37-195(0.7g, 0.021mmol) was charged to a 100mL flask, dichloromethane (10mL), TFA (4.67mL, 6.29mmol) were added, and the reaction was stirred at room temperature overnight. And (3) after the reaction is finished, adding methyl tert-butyl ether (100mL), separating out a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (100mL x 3), dissolving with a methanol (20mL) and dichloromethane (80mL) solution, adding silica gel powder (10g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 6% -8% methanol, collecting and concentrating, evaporating to dryness to obtain a solid, and drying in a vacuum oven to obtain a product 37-199: 0.697g, 43% yield.

37-200

37-199(0.3g, 0.009mmol) was put into 100mL, DMF (40mL) was added thereto and dissolved, and M-NH was added thereto₂-5k.hcl (0.2772g, 0.0543mmol, ex key k), HBTU (0.21g, 0.543mmol), HOBT (0.073g, 0.543mmol), stirring at-5 ℃ for about 10 min at low speed, then slowly adding DIEA (0.7mL, 4.32mmol) dropwise, continuing the reaction at-5 ℃ for 20 min, then moving the reaction to room temperature and stirring at low speed for 7 days in the absence of light. After the reaction is finished, adding n-hexane (100mL) and methyl tert-butyl ether (30mL), pouring supernatant, adding n-hexane (100mL) and methyl tert-butyl ether (30mL) into lower layer liquid, repeating the steps for 3 times to obtain viscous oily substance, adding methyl tert-butyl ether (100mL), separating out solid, filtering, washing filter cake with methyl tert-butyl ether (30mL x 3), dissolving with methanol (40mL) and dichloromethane (160mL), adding silica gel powder (10g), evaporating to obtain powdery solid, dry loading, performing column chromatography, eluting with dichloromethane mixed solution of 1% ammonia water and 4% -7% methanol, collecting, concentrating, evaporating to obtain solid, drying in a vacuum oven for 1 hour, adding absolute ethyl alcohol (2mL) and dichloromethane (30mL) to dissolve, adding methyl tert-butyl ether (200mL), separating out solid, filtering, washing filter cake with methyl tert-butyl ether (50mL x 2), drying the filter cake in a vacuum oven to obtain a product 37-200: 0.27g, yield: 56 percent.

¹H-NMR(600MHz,DMSO-d₆)δ10.19–10.05(m,16H),9.06–8.84(m,15H),8.32–7.77(m,251H),7.59–7.46(m,45H),7.31–7.05(m,313H),5.92–5.66(m,25H),4.61–4.49(m,25H),4.43–4.29(m,14H),4.26–4.16(m,40H),4.05–3.84(m,31H),3.70–3.61(m,113H),3.58–3.41(m,1856H),3.26–3.24(m,28H),3.20–3.17(m,44H),3.14–3.11(m,32H),3.07–2.87(m,99H),2.81–2.69(m,75H),2.67–2.60(m,35H),2.45–2.37(m,87H),2.36–2.28(m,118H),2.27–2.08(m,116H),1.94–1.69(m,145H),1.62–1.42(m,151H),1.30–1.16(m,44H),0.94–0.75(m,247H),0.53–0.46(m,35H).

9. Synthesis of 35-78 (Compound No. 4)

The route is as follows

45-57

Pentaerythritol (5g, 36.7242mmol, available from Aladdin) was charged into a 500mL flask, added to a solution of potassium tert-butoxide (146.8968mL, 146.8968mmol) in tetrahydrofuran, the reaction was left to stir at 0 ℃ for 20 minutes, benzyl bromoacetate (24.5129mL, 161.5865mmol) was added slowly dropwise, stirring was continued at 0 ℃ for 2 hours after the addition was complete, and the mixture was allowed to move to room temperature for overnight reaction. After completion of the reaction, the reaction mixture was transferred to a 2L separatory funnel, and deionized water (300mL) and ethyl acetate (300mL) were added, followed by shaking and extraction. Washing the water phase with ethyl acetate (250mL x 2), combining the organic phases, washing with saturated saline (200mL x 1), concentrating and evaporating to dryness, adding dichloromethane (300mL) for dissolving, adding silica gel powder (55g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a 20% ethyl acetate petroleum ether mixed solution, concentrating and evaporating to dryness, and drying in a vacuum oven to obtain a product 45-57: 12.3g, yield 46%.

35-33

35-32 (synthesized by 42-25, 9.364g, 7.418mmol), Gly-OtBu.HCl (1.243g, 7.418mmol, available from Ilykok), HBTU (4.219g, 11.126mmol), HOBT (1.503g, 11.126mmol) were added to a 250mL flask, 30mL of DMF was added to dissolve the DMF, the mixture was stirred at-5 ℃ for 30min, DIEA (11.034mL, 66.758mmol) was slowly added dropwise, the reaction was continued for 1 h after the dropwise addition was completed, and then the mixture was stirred at room temperature overnight. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding deionized water (200mL) and ethyl acetate (200mL) into the separating funnel, extracting to obtain an organic phase, washing a water phase by using ethyl acetate (200mL x 2), combining the organic phases, washing the organic phase by using saturated saline (200mL x 2), evaporating to dryness, and drying in an oven to obtain a product 35-33: 10.2 g.

35-34

Putting 35-33(10.203g, 7.418mmol) into a 250mL flask, adding DMF (30mL) to dissolve, adding morpholine (9.69mL, 111.27mmol), stirring at room temperature for 1 hour, finishing the reaction, transferring the reaction solution into a 1L separating funnel, adding saturated saline (200mL) and ethyl acetate (200mL) to extract to obtain an organic phase, washing an aqueous phase with ethyl acetate (200mL x 2), combining the organic phases, washing the organic phase with saturated saline (200mL x 1), concentrating and evaporating to dryness, adding silica gel powder (35g), evaporating to dryness to obtain a powdery solid, loading by a dry method, performing column chromatography, eluting with a mixed solution of 0.5% ammonia water and 3% methanol in dichloromethane, collecting and concentrating, evaporating to dryness, and drying in an oven to obtain 35-34:5.2g of a product, wherein the yield is 61%.

35-35

35-34(5.2g, 4.509mmol), succinic anhydride (1.35g, 13.527mmol, available from ImmunoKat) were added to a 250mL flask, 30mL of DMF was added to dissolve, the mixture was stirred at-5 ℃ for reaction for 30min, DIEA (4.47mL, 27.053mmol) was slowly added dropwise, the reaction was continued for 1 hour after the addition was complete, and the mixture was stirred at room temperature for reaction overnight. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding deionized water (200mL) and ethyl acetate (200mL) into the separating funnel, extracting to obtain an organic phase, washing a water phase by using ethyl acetate (200mL x 2), combining the organic phases, washing the organic phase by using saturated saline (200mL x 2), evaporating to dryness, and drying in an oven to obtain a product 35-35: 4.7g, yield 83%.

35-36

35-35(4.7g, 3.75mmol), 35-31 (synthesized by 34-17, 5.203g, 4.125mmol), HBTU (2.133g, 5.625mmol) and HOBT (0.76g, 5.625mmol) were added to a 250mL flask, 30mL of DMF was added to dissolve, the mixture was stirred at-5 ℃ for reaction for 30min, DIEA (2.789mL, 16.875mmol) was slowly added dropwise, the reaction was continued for 1 h after the dropwise addition was complete, and then the mixture was stirred at room temperature for overnight reaction. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding a saturated sodium chloride solution (270mL) and ethyl acetate (240mL) into the separating funnel, extracting to obtain an organic phase, washing a water phase by using ethyl acetate (200mL x 2), combining the organic phases, washing the organic phase by using a saturated saline solution (200mL x 2), evaporating to dryness, adding silica gel powder (15g), evaporating to dryness to obtain a powdery solid, loading by a dry method, carrying out column chromatography, eluting by using a dichloromethane mixed solution of 4% -6% methanol, collecting, concentrating, evaporating to dryness, and drying by an oven to obtain a product 35-36: 6.3g, yield 67%.

¹H-NMR(400MHz,DMSO-d₆)δ8.52(d,J＝6.8Hz,2H),8.31(d,J＝6.8Hz,3H),8.20(s,1H),8.08(s,1H),7.96(m,2H),7.86(m,3H),7.78–7.54(m,4H),7.54–7.46(m,2H),7.39–7.26(m,42H),4.26–4.16(m,11H),4.12–3.84(m,3H),3.72–3.47(m,15H),3.25–3.10(m,10H),2.99–2.84(m,3H),2.81–2.67(m,2H),2.45–2.34(m,13H),2.23–2.11(m,6H),1.42–1.34(m,9H),1.29–1.24(d,J＝6.6Hz,28H).

35-48

35-36(0.463g, 0.185mmol) was charged into a hydrogenation reactor, Pd/C (0.030g) was added thereto, DMF (30mL) was added thereto and dissolved therein, hydrogen gas was introduced under a pressure Pa of 1.8MPa, and the reaction was stirred at room temperature overnight. After the reaction was complete, the reaction vessel was removed, and the reaction solution was filtered through celite and washed with DMF (20mL × 3) to give a DMF solution containing C7.

35-49

35-48(0.329g, 0.185mmol), 37-2 (synthesized by 37-2, 3.0g, 1.594mmol), HBTU (0.842g, 2.22mmol), HOBT (0.3g, 2.22mmol) were charged into a 250mL flask, DMF (90mL) was added to dissolve the mixture, the reaction was left at-5 ℃ and stirred for 20 minutes, DIEA (1.1mL, 6.66mmol) was slowly added dropwise, the dropwise addition was complete, the reaction was stirred at-5 ℃ for 30 minutes, and then stirred at room temperature overnight. After the reaction is finished, adding n-hexane (150mL) and methyl tert-butyl ether (40mL), settling for 3 times to obtain a viscous oily substance, adding methyl tert-butyl ether (250mL), separating out a solid, filtering, washing a filter cake with methyl tert-butyl ether (50mL x 3), dissolving the filter cake with dichloromethane (130mL) and methanol (25mL), adding silica gel powder (15g), evaporating to obtain a powdery solid, loading by a dry method, performing column chromatography, eluting with a dichloromethane mixed solution of 0.5% of ammonia water and 5% -12% of methanol, collecting, concentrating and drying to obtain a product 35-49:2.7g, wherein the yield is 87%.

35-73

35-49(2.7g, 0.1615mmol) was placed in a 250mL flask, DMF (30mL) was added and dissolved, piperidine (0.479mL, 4.8443mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction is finished, adding n-hexane (100mL) and methyl tert-butyl ether (20mL) for settling, carrying out layering, pouring an upper layer, adding n-hexane (100mL) and methyl tert-butyl ether (20mL) into a lower oily liquid phase, continuing to settle, repeating the process for 3 times, finally adding methyl tert-butyl ether (150mL) into a viscous oily substance, separating out a solid, filtering, washing a filter cake with methyl tert-butyl ether (40mL x 2), dissolving the filter cake with dichloromethane (20mL) and methanol (80mL), adding silica gel powder (10g), evaporating to dryness to form a powdery solid, carrying out dry loading, carrying out column chromatography, and eluting with a dichloromethane mixed solution of 1% ammonia water and 5% -7% methanol to obtain a product 35-73:1.2g, wherein the yield is 45%.

35-74

45-57(0.118g, 0.1655mmol) was charged into a hydrogenation reactor, and Pd/C (0.01g) was added thereto, and DMF (30mL) was added thereto and dissolved therein, and hydrogen gas was introduced under a pressure of 1.8MPa, and the reaction was stirred at room temperature overnight. After the reaction was complete, the reaction vessel was removed, and the reaction solution was filtered through celite and washed with DMF (20 mL. times.3) to provide a DMF solution containing 35-74.

35-75

35-73(1.2g, 0.0727mmol), 35-74(0.375g, 0.0165mmol), HBTU (0.375g, 0.99mmol), HOBT (0.134g, 0.99mmol) were charged into a 250mL flask, DMF (30mL) was added to dissolve the mixture, the reaction was left at-5 ℃ and stirred for 20 minutes, DIEA (0.492mL, 2.975mmol) was slowly added dropwise, the reaction was stirred at-5 ℃ for 30 minutes after completion of the addition, and then stirred at room temperature overnight. After the reaction is finished, adding n-hexane (150mL) and methyl tert-butyl ether (40mL), settling for 3 times to obtain a viscous oily substance, adding methyl tert-butyl ether (250mL), separating out a solid, filtering, washing a filter cake with methyl tert-butyl ether (50mL x 3), dissolving the filter cake with dichloromethane (130mL) and methanol (25mL), adding silica gel powder (10g), evaporating to obtain a powdery solid, loading by a dry method, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 6% -9% methanol, collecting, concentrating and drying to obtain a product 35-75:0.8g, wherein the yield is 73%.

35-77

35-75(0.8g, 0.012mmol) was charged into a 100mL flask, dichloromethane (10mL), TFA (10mL, 134.65mmol) were added, and the reaction was stirred at room temperature overnight. After the reaction is finished, adding methyl tert-butyl ether (70mL), precipitating a solid, performing suction filtration, cleaning a filter cake with methyl tert-butyl ether (40mL x 3), dissolving with a methanol (20mL) and dichloromethane (80mL) solution, adding silica gel powder (10g), evaporating to dryness to obtain a powdery solid, loading by a dry method, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 4% -8% methanol, collecting and concentrating, evaporating to dryness to obtain a solid, and drying by a vacuum oven to obtain a product 35-77: 0.4g, yield 51%.

35-78

35-77(0.4g, 0.0061mmol) was placed in 250mL, DMF (25mL) was added and dissolved, and M-NH was added₂-20k. hcl (0.379g, 0.036mmol, from key kelly), HBTU (0.137g, 0.36mmol), HOBT (0.049g, 0.36mmol), stirring at-5 ℃ for about 10 min at low speed, then slowly adding DIEA (0.1313mL, 0.7945mmol) dropwise, continuing the reaction at-5 ℃ for 20 min, then moving the reaction to room temperature and stirring at low speed away from light for 7 days. After the reaction, n-hexane (100mL) and methyl tert-butyl ether (30mL) were added, the supernatant was poured off, the lower liquid was added with n-hexane (100mL) and methyl tert-butyl ether (30mL), and this was repeated 3 times to give a viscous oil, and methyl tert-butyl ether was added (100mL), precipitating a solid, filtering, washing a filter cake with methyl tert-butyl ether (30mL x3), dissolving with a methanol (40mL) and dichloromethane (160mL), adding silica gel powder (10g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 4% -7% methanol, collecting and concentrating, evaporating to dryness to obtain a solid, drying in a vacuum oven for 1 hour, adding absolute ethanol (2mL) and dichloromethane (30mL) to dissolve, adding methyl tert-butyl ether (140mL), precipitating a solid, filtering, washing the filter cake with methyl tert-butyl ether (50mL x 2), drying the filter cake in the vacuum oven to obtain a product 35-78: 0.46g, yield: 70 percent.

¹H-NMR(600MHz,DMSO-d₆)δ10.23–9.98(m,34H),9.12–8.76(m,32H),8.25–8.13(m,116H),8.12–7.95(m,240H),7.84–7.64(m,95H),7.57–7.31(m,99H),7.21–6.91(m,630H),5.88–5.63(m,56H),4.76–4.48(m,64H),4.33–4.26(m,154H),4.05–3.83(m,116H),3.71–3.62(m,241H),3.59–3.41(m,3840H),3.42–3.37(m,91H),3.27–3.21(m,393H),3.19–3.11(m,149H),3.09–3.93(m,72H),2.85–2.73(m,102H),2.52–2.42(m,116H),2.34–2.07(m,369H),1.91–1.84(m,120H),1.78–1.63(m,123H),1.61–1.52(m,265H),1.29–0.92(m,89H),0.85–0.63(m,461H),0.60–0.47(m,81H).

10. Synthesis of 33-200 (Compound No. 5)

The route is as follows

45-1

24-143 (5 g, 3.6723mmol, synthesized by 34-17) was placed in a 250mL round bottom flask, dissolved in DMF (20mL), placed in a low temperature thermostatic bath at-5 deg.C, DIEA (2.5390mL, 18.3615mmol) was added, the reaction was stirred at room temperature for 30 minutes, succinic anhydride (1.1025g, 11.0170mmol) was added, and after 1 hour, the reaction was allowed to proceed at room temperature. After the reaction, the reaction solution was transferred to a 1L separatory funnel, ethyl acetate (200mL) and deionized water (150mL) were added, the organic phase was separated, the aqueous phase was extracted 2 times with ethyl acetate (100mL × 2), the product was not found in the aqueous phase, the organic phases were combined, washed 1 time with saturated saline (100mL), concentrated, and evaporated to dryness to obtain 5g of product, 0.1g of over-yield, and 100% yield.

45-6

Taking reactant 45-1(4.9g, 3.5990mmol), NH₂-Gly-OtBu & HCl (available from Accela, 0.4721g, 3.5990mmol), HBTU (2.0473g, 5.3985mmol), HOBT (0.7294g, 5.3985mmol) were placed in a 250mL reaction flask, dissolved in DMF (30mL), stirred at-5 ℃ for 30 min, DIEA (4mL, 24.2933mmol) was slowly added dropwise, and the reaction was terminated at low temperature. After the reaction, the reaction solution was transferred to a 1L separatory funnel, ethyl acetate (200mL) and deionized water (150mL) were added, the organic phase was separated, the aqueous phase was extracted 2 times with ethyl acetate (100mL × 2), no product was found in the aqueous phase, the organic phases were combined, washed 1 time with saturated saline (100mL), concentrated, added with silica gel powder, evaporated to dryness, and subjected to column chromatography. The product 3.5g is obtained by 50% -100% ethyl acetate/petroleum ether, and the yield is 66%.

¹H-NMR(400MHz,DMSO-d₆)δ8.34–8.18(m,2H),8.01–7.85(m,4H),7.72–7.64(m,3H),7.43–7.22(m,26H),5.14–5.03(m,8H),4.41–4.06(m,8H),3.94–3.86(m,2H),3.71–3.66(m,2H),3.58–3.49(m,4H),3.42–3.36(m,2H),3.22–3.13(m,3H),2.98–2.88(m,2H),2.47–1.81(m,17H),1.39–1.37(m,9H),1.28–1.20(m,3H).

MALDI-TOF MS:[M+H⁺]1474.70,[M+Na⁺]1496.70.

33-190

Weighing reactant 45-6(0.4702g, 0.3189mmol) and putting into a micro reaction kettle, adding 10% Pd/C (50mg), adding DMF (30mL) for dissolving, and introducing H₂(20psi), stir the reaction. And after the reaction is finished, using diatomite as a filter cake, carrying out suction filtration on the reaction liquid to remove Pd/C, washing the diatomite for 3-4 times by using DMF to obtain a DMF solution of a product, and carrying out the next step of reaction.

33-191

Weighing 33-162 (3 g synthesized by 37-2 method, 1.5944mmol), HBTU (0.7256g, 1.9134mmol) and HOBT (0.2585g, 1.9134mmol), putting into a 250mL reaction flask, adding 33-190 DMF solution for dissolving, performing ultrasonic treatment to completely dissolve the reactants, stirring for 30 minutes at-5 ℃, slowly dropwise adding DIEA (0.9487mL, 5.7402mmol), and reacting at low temperature until the reaction is finished. After the reaction, methyl tert-butyl ether (100mL) and n-hexane (150mL) were added to the reaction mixture, the mixture was sonicated for 5 minutes, the mixture was placed in a refrigerator and allowed to stand for 20 minutes, the supernatant was poured off, ethyl acetate (20mL) was added to the lower layer, sonicated for 2 minutes, n-hexane (100mL) was added thereto, the mixture was suction-filtered, dissolved in 20% methanol/dichloromethane (70mL) and then silica gel powder was added thereto and evaporated to dryness. And (5) performing column chromatography. With 1% ammonia: 5% methanol/dichloromethane-1% ammonia: elution with a 10% methanol/dichloromethane gradient gave 1.9g of product in 70% yield.

33-192

33-191(1.9g, 0.2218mmol) was placed in a 250mL round bottom flask, dissolved in DMF (20mL) and morpholine (0.5796mL, 6.6540mmol) was added and the reaction stirred at room temperature. After the reaction, methyl tert-butyl ether (150mL) and n-hexane (100mL) were added to the reaction mixture, sonicated for 5 minutes, filtered, dissolved in 20% methanol/dichloromethane (50mL), and then silica gel powder was added and evaporated to dryness. And (5) performing column chromatography. The product was eluted with 1% aqueous ammonia, 7%/dichloromethane-1% aqueous ammonia, 15% methanol/dichloromethane to give 1.3g, 72% yield.

45-2

Dipentaerythritol (purchased from ACROS, 5g, 19.6634mmol) was weighed into a 500mL reaction flask, nitrogen was introduced, a THF solution of potassium tert-butoxide (141.57mL, 141.5762mmol) was added, the reaction was stirred at 0 ℃ for 1 hour, phenyl bromoacetate (30.4445g, 141.5762mmol) was added, and after stirring for three hours, the mixture was allowed to cool to room temperature. And (3) after the reaction is finished, evaporating the reaction solution to dryness, adding deionized water and ethyl acetate, separating an organic phase, extracting a water phase by using ethyl acetate until no product exists, combining the organic phase, drying by using anhydrous sodium sulfate powder, carrying out suction filtration, carrying out dry-method sample loading on the filtrate, and carrying out column chromatography. The product 13.9g is obtained by gradient elution with 3 percent to 5 percent of ethyl acetate/petroleum ether, and the yield is 62 percent.

45-12

Weighing reactant 45-2(0.2541g, 0.0223mmol) and putting into a micro reaction kettle, adding 10% Pd/C (50mg), adding DMF (30mL) for dissolving, and introducing H₂(20psi), stir the reaction. And after the reaction is finished, using diatomite as a filter cake, carrying out suction filtration on the reaction liquid to remove Pd/C, washing the diatomite for 3-4 times by using DMF to obtain a DMF solution of a product, and carrying out the next step of reaction.

33-195

33-192(1.3g, 0.1558mmol), HBTU (0.0761g, 0.2007mmol) and HOBT (0.0271g, 0.2007mmol) were weighed out and put into a 250mL reaction flask, and dissolved in 45-12 DMF, and the reaction mixture was stirred for 30 minutes at-5 ℃ with ultrasound, DIEA (0.0995mL, 0.6021mmol) was slowly added dropwise, and the reaction was completed at low temperature. After the reaction, methyl tert-butyl ether (100mL) and n-hexane (150mL) were added to the reaction mixture, the mixture was sonicated for 5 minutes, the mixture was placed in a refrigerator and allowed to stand for 20 minutes, the supernatant was poured off, ethyl acetate (20mL) was added to the lower layer, sonicated for 2 minutes, n-hexane (100mL) was added thereto, the mixture was suction-filtered, dissolved in 20% methanol/dichloromethane (70mL) and then silica gel powder was added thereto and evaporated to dryness. And (5) performing column chromatography. With 1% ammonia: 5% methanol/dichloromethane-1% ammonia: elution with a 15% methanol/dichloromethane gradient gave 0.8g of product in 53% yield.

33-197

After adding dichloromethane (10mL) to 33-195(0.8g, 0.0158mmol), TFA (0.03572mL, 0.4749mmol) was added and dissolved completely by sonication, and the reaction was stirred at room temperature with a ground glass stopper. After the reaction is finished, methyl tert-butyl ether (150mL) and n-hexane (100mL) are directly added into the reaction solution, and the product is obtained by suction filtration and vacuum drying, wherein the yield is 75 percent and 0.6 g.

33-200

Reactants 33-197(0.6g, 0.0119mmoL), M-NH were weighed₂HCl-10K (0.1g, 0.0099mmoL), HBTU (0.0056g, 0.0149mmoL), and HOBT (0.0020g, 0.0149mmoL) were put into a 250mL reaction flask, dissolved with DMF (15mL) by sonication, stirred at-5 ℃ for 30 minutes, then DIEA (0.0073mL, 0.0446mmoL) was slowly added dropwise, stirred for 1 hour, and then reacted at room temperature in a low-speed dark place. After the reaction is finished, adding methyl tert-butyl ether (200mL) into the reaction solution for sedimentation, performing suction filtration to obtain a powder product, dissolving the powder product by using a mixed solvent of 20% methanol/dichloromethane, adding silica gel (3g), evaporating to dryness, performing dry-method sample loading, and performing column chromatography. With 6% methanol/dichloromethane-1% ammonia: gradient eluting with 10% methanol/dichloromethane, collecting product, evaporating to dryness, adding dichloromethane (5mL), dissolving with ultrasound, adding methyl tert-butyl ether (150mL), adding n-hexane (50mL), vacuum filtering, dissolving with dichloromethane, precipitating with methyl tert-butyl ether and n-hexane, dissolving repeatedly for three times,0.5g of product is obtained, yield 71%.

¹H-NMR(600MHz,DMSO-d₆)δ10.17–10.13(m,18H),8.97–8.93(m,32H),8.25–7.88(m,412H),7.59–7.48(m,93H),7.32–7.16(m,459H),5.92–5.67(m,58H),4.40–3.99(m,672H),3.51–3.50(m,3560H),3.29–2.99(m,344H),2.94–2.64(m,264H),2.45–2.05(m,133H),1.93–1.06(m,301H),0.99–0.74(m,432H).

11. Synthesis of 40-123 (Compound No. 7)

The route is as follows

45-8

The reaction product 42-27 was placed in a 250mL flask, DMF solution (20mL) was added, morpholine (7.2mL, 83.4mmoL) was added, and the reaction was stirred at room temperature for 3 hours to complete the reaction. Methyl tert-butyl ether (100mL) was added, and n-hexane (200mL) was added to the mixture to precipitate the mixture as a powder. Column chromatography, dry loading, loading with 1% ammonia water: 2% methanol/dichloromethane-1% aqueous ammonia 6% methanol/dichloromethane gradient. The product weighed 4 grams.

45-9

The reaction products E4(4g, 3.14mmoL), SA-OtBu (purchased from Shaoyuan, 0.54, 3.14mmoL), HBUT (1.78g, 4.71mmoL), HOBT (0.63g, 4.71mmoL) were put into a 250mL reaction flask, DMF (40mL) was added to dissolve, the mixture was placed at 0 ℃ and stirred for 0.5 hour, DIEA (2.3mL, 14.13mmoL) was slowly added dropwise, the reaction was stirred until the reaction was completed, deionized water (100mL) and ethyl acetate (100mL) were added, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (50 mL. times.3), the aqueous phase was free of product, the organic phase was combined, washed with saturated saline (50 mL. times.2), the organic phase was evaporated to dryness to obtain 3.5 g of product, and the yield was 79%.

¹H-NMR(400MHz,DMSO-d₆)δ8.58–8.43(m,1H),8.35–8.23(m,1H),8.06–7.76(m,4H),7.74–7.65(m,1H),7.37–7.27(m,20H),5.13–5.02(m,8H),4.41–4.13(m,4H),3.95–3.85(m,2H),3.53–3.46(m,9H),3.25–2.99(m,8H),2.89-2.87(m,1H),2.73-2.71(m,1H),2.46–2.30(m,9H),2.21–1.79(m,12H),1.37–1.34(m,18H).

MALDI-TOF MS:[M+H⁺]1426.75,[M+Na⁺]1448.75.

33-145

Reactants Fmoc-E (OtBu) (OH) (1.39g, 3.29mmoL), GFLG-LPT (synthesized by 14-128, 3g, 3.13mmoL), HBUT (1.78g, 4.7mmoL), and HOBT (0.63g, 4.7mmoL) were weighed into a 250mL reaction flask, dissolved in DMF (30mL), placed at-5 deg.C, stirred for 0.5 hour, DIEA (2.34mL, 14.2mmoL) was slowly added dropwise, and after 1 hour, the reaction was stirred at room temperature. After the reaction was complete, deionized water (100mL) and ethyl acetate (100mL) were added, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (50 mL. times.3), the aqueous phase was freed of product, the organic phases were combined, washed 2 times with saturated brine (50 mL. times.2), and the organic phase was evaporated to dryness. The yield was 100%.

33-147

33-145(3.13mmol) was weighed out, dichloromethane (5mL) was added, TFA (6.63mL, 89.4mmol) was added, the mixture was dissolved by sonication, and the reaction was stirred. After the reaction, the reaction mixture was concentrated, and methyl t-butyl ether (100mL) was added to the concentrated solution, followed by addition of n-hexane (200mL) to the concentrated solution and precipitation to obtain a powder. Column chromatography, dry loading, eluting with 2% methanol/dichloromethane to 6% methanol/dichloromethane. The weight of the product was 2.8g, and the yield was 70%.

39-11

Reactants 33-147(2.8g, 2.14mmoL), GFLG-SB7 (synthesized by 25-132 method, 2g, 2.247mmoL), HBUT (1.21g, 3.21mmoL), HOBT (0.43g, 3.21mmoL) were weighed into a 250mL reaction flask, dissolved in DMF (30mL), stirred at-5 ℃ for 0.5 hour, DIEA (1.59mL, 9.63mmoL) was slowly added dropwise, and the reaction was stirred at room temperature after 1 hour. After the reaction, deionized water (100mL) and ethyl acetate (100mL) are added for extraction, the organic phase is separated, the aqueous phase is extracted by ethyl acetate for 3 times (50mL multiplied by 3), the product does not exist in the aqueous phase, the organic phases are combined, the organic phase is washed by saturated saline solution for 2 times (50mL multiplied by 2), and the organic phase is evaporated to dryness to obtain 4.6 g of the product. The yield was 100%.

39-12

Reaction products 39-11(2.14mmoL) are placed in a 250mL flask, DMF solution (20mL) is added, morpholine (5.59mL, 64.2mmoL L) is added, stirring is carried out at normal temperature for reaction, after 3 hours, the reaction is finished, deionized water (300mL) and ethyl acetate (150mL) are added, an organic phase is separated, an aqueous phase is extracted for 3 times by ethyl acetate (50mL multiplied by 3), products are not contained in the aqueous phase, and the organic phases are combined. The yield was 100%.

39-13

Reactants 39-12(2.14mmoL), Boc-Gly-OH (0.39, 2.247mmoL), HBUT (1.21g, 3.21mmoL), and HOBT (0.43g, 3.21mmoL) were weighed into a 250mL reaction flask, dissolved in DMF (50mL), stirred at-5 ℃ for 0.5 hour, DIEA (1.59mL, 9.63mmoL) was slowly added dropwise, and the reaction was stirred at room temperature after 1 hour. After the reaction was completed, deionized water (300mL) and ethyl acetate (150mL) were added, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (50 mL. times.3), the aqueous phase was free of product, the organic phases were combined, washed 2 times with saturated brine (50 mL. times.2), and the organic phase was evaporated to dryness with a yield of 100%.

39-20

39-13(2.14mmol) was weighed out, dichloromethane (5mL) was added, TFA (4.7mL, 64.2mmol) was added, the mixture was dissolved by sonication, and the reaction was stirred. After the reaction, the reaction mixture was concentrated, and methyl t-butyl ether (100mL) was added to the concentrated solution, followed by addition of n-hexane (200mL) to the concentrated solution and precipitation to obtain a powder. Column chromatography, dry loading, loading with 1% ammonia water: 3% methanol/dichloromethane-1% ammonia: 6% methanol in dichloromethane. The product weighed 3g, yield 69%.

45-29

Reactant 45-9(0.22g, 0.1551mmoL) was weighed into a hydrogenation reactor, DMF was added dropwise along the inner wall to dissolve the reactant, 10% Pd/C (0.05g) was added, hydrogen was added, and the mixture was stirred overnight. After the reaction is finished, adding diatomite into a sand core funnel, compacting, filtering the reaction solution by suction, removing Pd/C, washing the product solution of the diatomite for three or four times by DMF (40ml), and putting the product solution into the next step for reaction.

45-30

Reactants 39-20(1.5g, 0.7445mmoL), 45-29(0.10551mmoL), HBUT (0.3529g, 0.9306mmoL) and HOBT (0.1257g, 0.9306mmoL) were weighed into a 250mL reaction flask, dissolved in DMF (50mL), stirred at-5 ℃ for 0.5 hour, DIEA (0.4614mL, 2.7918mmoL) was slowly added dropwise, and the mixture was taken out after 1 hour and stirred at room temperature. After the reaction, methyl tert-butyl ether (100mL) was added, and n-hexane (200mL) was added to the mixture to precipitate the mixture as a powder. Column chromatography, dry loading, loading with 1% ammonia water: 4% methanol/dichloromethane-1% ammonia: 10% methanol in dichloromethane. The product was evaporated to dryness to give 1.4g, 40% yield.

45-35

45-35(1.4g) was weighed out, dichloromethane (5mL) was added, TFA (0.5741mL, 7.73mmol) was added, the mixture was dissolved by sonication, and the reaction was stirred. After the reaction, the reaction mixture was concentrated, and methyl t-butyl ether (100mL) was added to the concentrated solution, followed by addition of n-hexane (200mL) to the concentrated solution and precipitation to obtain a powder. Column chromatography, dry loading, loading with 1% ammonia water: 5% methanol/dichloromethane-1% ammonia: 10% methanol in dichloromethane. The product weighed 1.2 g, yield 80%.

45-65

Reactants 35-74(2g, 5.4mmol), succinic alcohol (5.5082g, 47.86mmol), DMAP (0.2922g, 2.392) were weighed into a 250mL round bottom flask, dichloromethane (50mL) was added, stirring was carried out at-5 ℃ and DCC (9.8750g, 47.86mmol) was added portionwise and rapidly. Stirring and reacting at room temperature, filtering the reaction solution after the reaction is finished, and evaporating the filtrate to dryness. Performing column chromatography and dry-method sample loading. The product was eluted with a gradient of 10% ethyl acetate/petroleum ether-ethyl acetate to give 3 g, 81% yield.

40-121

Reactants 45 to 35(1.2g, 0.101mmol) were dissolved in DMF (20mL) in a 250mL flask, and then placed in a low-temperature constant-temperature bath (-5 ℃) and stirred for 30 minutes, followed by dropwise addition of DIEA (0.1 mL). Adding 45-65(0.015g, 0.021mmoL), dissolving, reaction at normal temp. in dark, and low-speed stirring. After the reaction, methyl tert-butyl ether was added to the flask(150mL), the reaction mixture was poured into the flask, and n-hexane (200mL) was further added. And (4) separating out a product. And (5) suction filtration. And (5) performing column chromatography. Column height 5cm, eluent 7% methanol/dichloromethane-1% ammonia: elution with a 10% methanol/dichloromethane gradient gave 0.7 g of product.¹H-NMR(600MHz,DMSO-d₆)δ10.14-10.12(m,12H),9.02-9.00(m,4H),8.95-8.94(m,14H),8.18-8.16(m,41H),8.13-8.05(m,90H),7.86-7.83(m,30H),7.55–7.47(m,41H),7.26–7.17(m,274H),7.11-7.08(m,65H),7.02-7.00(m,44H),6.73-6.64(m,6H),4.39–4.13(m,71H),4.03–3.45(m,268H),3.22–2.66(m,207H),2.41-2.38(m 59H),2.31-2.28(m,99H),2.26–1.97(m,127H),1.88-1.47(m,281H),0.97–0.73(m,288H).

40-123

Weighing M-NH₂HCL-10K (0.0528g, 0.05mmoL), 40-121(0.37g, 0.012mmoL), HOBT (0.009g, 0.072mmoL), HBTU (0.027g, 0.072mmoL), in a 250mL flask, DMF solution (20mL) was added, the flask was placed in a low temperature constant temperature bath, DIEA (0.035mL, 0.216mmoL) was added dropwise after 30 minutes, and the reaction was stirred at room temperature and low speed after 1 hour. And (4) avoiding light. After the reaction was completed, methyl t-butyl ether (50mL) was added, and n-hexane (150mL) was further added. Precipitating and filtering. The solid was dissolved in 50mL (20% methanol: 80% dichloromethane), dry-loaded, and subjected to column chromatography. Column height 5cm, 1% ammonia: gradient eluting with 5-10% methanol/dichloromethane, evaporating to dryness, adding anhydrous ethanol (5mL), adding dichloromethane (2mL), dissolving, ultrasonic treating to obtain homogeneous phase, adding n-hexane (100mL), and filtering. Repeatedly dissolving and settling for three times. And (5) drying in vacuum. 0.45 g of product is obtained.

¹H-NMR(600MHz,DMSO-d₆)δ10.12-10.10(m,16H),8.93-8.91(m,16H),8.19-8.06(m,172H),7.84-7.67(m,65H),7.21-7.11(m,354H),4.54-3.89(m,134H),3.48-3.46(m,3578H),3.21-3.11(m,175H),2.89-2.87(m,28H),2.79-2.66(m,80H),2.43-2.07(m,353H),1.93-1.43(m,340H),0.96–0.74(m,288H).

12. Synthesis of 41-40 (Compound No. 13)

The route is as follows

Reactants Fmoc-Glu-OH (purchased from Aladdin, 0.7719g, 2.0899mmol), 31-155 (synthesized by 27-120 method, 5g, 4.3887mmol), HBUT (2.3777g, 6.2697mmol) and HOBT (0.8472g, 6.2697mmol) are put into a 250mL reaction flask, DMF (40mL) is added for dissolving, the mixture is placed at-5 ℃ and stirred for 30 minutes, DIEA (3.1208mL, 18.8091mmol) is slowly added dropwise, and the mixture is transferred to the room temperature for reaction after 2 hours of low temperature reaction. After the reaction, the reaction mixture was transferred to a 1L separatory funnel, deionized water (200mL) and ethyl acetate (300mL) were added for extraction, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (100 mL. times.3), the organic phases were combined, washed 2 times with saturated brine (150 mL. times.2), concentrated and evaporated to dryness to obtain 5.5g of a product with a yield of 100%.

39-29

Substance 39-27(5.4g, 2.0899mmol) was placed in a 250mL round-bottomed flask and then DMF (20mL) was added to dissolve it, and the mixture was stirred at room temperature. Morpholine (4.5544mL, 52.2475mmol) was added dropwise thereto, and the reaction was carried out at room temperature for 2 hours after the addition. After the reaction, the reaction solution was transferred to a 1L separatory funnel, deionized water (200mL) and ethyl acetate (300mL) were added for extraction, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (100mL × 3), the organic phases were combined, washed 2 times with saturated saline (150mL × 2), concentrated to 100mL, silica gel powder (15g) was added to evaporate to dryness, column chromatography was performed with 1% ammonia: 0.5% methanol/dichloromethane-1% ammonia: the product was 3.9848g, 73% yield, eluting with a 2% methanol/dichloromethane gradient.

41-26

Reactants 31-155 (synthesized by 27-120, 1g, 0.87mmol) were dissolved in DMF (20mL), stirred at 0 deg.C, and DIEA (0.58mL, 3.51mmol) was added dropwise. Succinic anhydride (0.26g, 2.16mmol) was added after 30 minutes. After the reaction, the reaction solution was transferred to a 1L separatory funnel, deionized water (100mL) and ethyl acetate (100mL) were added, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (50 mL. times.3), the aqueous phase was free of product, and the organic phases were combined. Evaporated to dryness, yield 100%.

41-27

The reaction products 39 to 29(2.0g, 0.87mmol), 41 to 26(0.4721g, 0.87mmol), HBTU (0.49g, 1.30mmol) and HOBT (0.17g, 1.30mmol) were taken and placed in a 250mL reaction flask, DMF (30mL) was added and dissolved, the mixture was stirred at-5 ℃ for 30 minutes, DIEA (0.64mL, 3.91mmol) was slowly added dropwise and the reaction was terminated at low temperature. After the reaction, the reaction solution was transferred to a 1L separatory funnel, ethyl acetate (200mL) and deionized water (150mL) were added for extraction, the organic phase was separated, the aqueous phase was extracted 2 times (100mL × 2) with ethyl acetate, the organic phases were combined, washed 1 time (100mL) with saturated saline, concentrated, added with silica gel powder, evaporated to dryness, and subjected to column chromatography. 1.8g of product is obtained by using 50-100% ethyl acetate/petroleum ether, and the yield is 60%.

¹H-NMR(400MHz,DMSO-d₆)δ8.57–8.47(m,3H),8.35–7.88(m,10H),7.75–7.65(m,3H),7.39–7.27(m,60H),6.76–6.61(m,3H),5.15–5.02(m,24H),4.42–4.29(m,9H),3.93–3.87(m,5H),3.61–3.47(m,13H),3.42–3.36(m,7H),2.90–2.80(m,6H),2.47–2.39(m,12H),2.23–1.68(m,31H),1.38–1.14(m,55H).

41-33

Reactants 41-27(0.41g, 0.113mmol), 10% Pd/C (0.166g) were weighed, dissolved in DMF (30mL), placed in a hydrogenation reactor, equipped with a device, charged with hydrogen, maintained at a pressure of 18Ps, and stirred for reaction. After the reaction, the reaction mixture was filtered through celite to remove Pd/C, and the celite was washed 4 times (25 mL. times.4) with DMF to obtain a product solution.

41-34

The reaction products 39-20(3.0g, 1.48mmol), 41-33(0.2858g, 0.113mmol), HBTU (0.77g, 2.034mmol) and HOBT (0.27g, 2.034mmol) were taken and placed in a 250mL reaction flask, DMF (30mL) was added to dissolve the mixture, the mixture was stirred at-5 ℃ for 30 minutes, DIEA (1.0mL, 6.102mmol) was slowly added dropwise, and the reaction was terminated at low temperature. After the reaction, methyl tert-butyl ether (100mL) was added, and n-hexane (200mL) was added to the mixture to precipitate the mixture as a powder. Column chromatography, dry loading, loading with 1% ammonia water: 2% methanol/dichloromethane-1% ammonia: 10% methanol in dichloromethane. The product weighed 1.64g, yield 55%.

41-35

41-34(1.64g, 0.06215mmol) was weighed out, dichloromethane (5mL) was added, TFA (0.41mL, 5.5935mmol) was added, the mixture was dissolved by sonication, and the reaction was stirred. After the reaction, the reaction mixture was concentrated, and methyl t-butyl ether (100mL) was added to the concentrated solution, followed by addition of n-hexane (200mL) to the concentrated solution and precipitation to obtain a powder. Column chromatography, dry loading, loading with 1% ammonia water: 5% methanol/dichloromethane-1% ammonia: 10% methanol in dichloromethane. The product weighed 1.13g, and the yield was 70%.

41-40

The reaction mixture 41-35(1.1g, 0.04350mmol) was dissolved in M-SCM-10K (1.43g, 0.14355mmol) and DMF (20mL) and stirred at low speed with the exclusion of light. At the end of the reaction, methyl tert-butyl ether (50mL) was added and n-hexane (100mL) was allowed to settle. And (5) suction filtration. Column chromatography, dry loading, eluting with dichloromethane-1% ammonia: elution with a gradient of 8% methanol in dichloromethane. The product is evaporated to dryness, absolute ethyl alcohol (10mL) is added, ultrasonic treatment is carried out to form a homogeneous phase, and n-hexane (50mL) is added for sedimentation. The sedimentation was repeated three times. Vacuum drying to obtain 1g of product. The yield was 41%.

¹H-NMR(600MHz,DMSO-d₆)δ8.58–8.53(m,5H),8.13–7.92(m,111H),7.72–7.65(m,34H),7.58–7.46(m,51H),7.41–7.33(m,88H),7.30–7.16(m,241H),5.29–5.23(m,12H),5.16–5.08(s,19H),4.15–4.12(m,48H),4.05–3.97(m,75H),3.51–3.50(m,3285H),3.25–3.23(m,173H),3.16–2.98(m,242H),2.93–2.85(m,54H),2.76–2.72(m,50H),2.68–2.65(m,55H),2.36–2.27(m,74H),0.94–0.75(m,216H).

13. 39-55 Synthesis (Compound No. 12)

The route is as follows

39-31

33-22 (synthesized by 22-181, 0.5g, 0.54mmol) was placed in a 100mL round-bottomed flask, and then, dichloromethane (10mL) was added thereto and dissolved by sonication, phenyl chloroformate (0.3mL, 2.19mmol) was added thereto, and the reaction mixture was stirred at 0 ℃ for 30 minutes. Phenyl chloroformate (0.13mL, 1.09mmol) was slowly added dropwise and reacted at low temperature. After 2 hours, the reaction was terminated, deionized water (200mL) and ethyl acetate (300mL) were added and extracted, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (100 mL. times.3), the organic phases were combined, washed 2 times with saturated brine (150 mL. times.2), and evaporated to dryness to give 0.5g of product.

39-30

39-29(1.4g, 0.59mmol) and 39-31(0.5g, 0.54mmol) were weighed into a 250mL reaction flask, a DMF (30mL) solution and DIEA (0.1mL) were added, and the reaction was stirred at 80 ℃ until completion. After the reaction, the reaction solution was transferred to a 1L separatory funnel, deionized water (100mL) and ethyl acetate (150mL) were added for extraction, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (50mL × 3), the organic phases were combined, washed 2 times with saturated saline (100mL × 2), concentrated to 50mL, silica gel powder (6g) was added for evaporation, and the product was subjected to column chromatography and gradient elution with 2% methanol/dichloromethane-3% methanol/dichloromethane to obtain 1.3g, the yield was 65.38%.

¹H-NMR(400MHz,DMSO-d₆)δ8.57–8.47(m,3H),8.40–8.06(m,6H),8.01–7.65(m,4H),7.38–7.28(s,60H),7.22–7.11(m,1H),5.14–5.03(m,24H),4.42–4.31(m,8H),3.94–3.86(m,5H),3.60–3.48(m,13H),2.88–2.81(m,4H),2.47–2.37(m,11H),2.22–2.13(m,6H),2.10–1.72(m,23H),1.37–1.31(m,29H),1.26–1.21(m,18H),0.88–0.82(m,2H).

39-51

Weighing 39-30(0.16g, 0.048mmol) and 10% Pd/C (70mg), dissolving with DMF (30mL), placing in a hydrogenation reaction kettle, installing the device, introducing hydrogen, keeping the pressure at 18Ps, and stirring for reaction. After the reaction, the reaction mixture was filtered through celite to remove Pd/C, and the celite was washed 4 times (25 mL. times.4) with DMF to obtain a product solution.

39-52

39-20(1.25g, 0.625mmol), 39-51(0.048mmol), HBTU (0.32g, 0.864mmol) and HOBT (0.11g, 0.864mmol) were weighed and placed in a 250mL reaction flask, DMF (30mL) was added and sonicated to dissolve the reaction product completely, stirring was carried out at-5 ℃ for 30 minutes, DIEA (0.42mL, 2.59mmol) was slowly added dropwise and stirred at low temperature for 2 hours, and then the reaction was carried out at room temperature until completion. After the reaction, methyl tert-butyl ether (250mL) was added to the reaction mixture, the mixture was sonicated for 5 minutes, the upper layer was poured, ethyl acetate (20mL) was added to the lower layer, the mixture was sonicated for 3 minutes, methyl tert-butyl ether (150mL) and n-hexane (100mL) were added to precipitate a solid, the mixture was filtered, dissolved in 20% methanol/dichloromethane (20mL), silica gel powder (10g) was added, evaporated to dryness, and subjected to column chromatography. With 1% ammonia: 6% methanol/dichloromethane-1% ammonia: elution with a 15% methanol/dichloromethane gradient gave 0.6g of product.

39-53

After methylene chloride (15mL) and TFA (0.2267mL, 3.0521mmol) were added to 39-52(0.6g), the mixture was dissolved completely by sonication, and the reaction mixture was stirred at room temperature with a ground glass stopper. At the end, the reaction solution is evaporated to dryness, dichloromethane is removed, ethyl acetate (20mL) is added, ultrasonic treatment is carried out for 2 minutes, methyl tert-butyl ether (150mL) is added, n-hexane (70mL) is added for suction filtration, 20% methanol/dichloromethane is added to the solid, ultrasonic dissolution is carried out, silica gel powder (6g) is added, evaporation is carried out by using a rotary evaporator, and column chromatography is carried out. With 1% ammonia: 6% methanol/dichloromethane-1% ammonia: elution with a 15% methanol/dichloromethane gradient gave 0.3g of product.

39-55

39-53(0.6g) was weighed out and dissolved in DMF (20mL), followed by addition of M-SCM-20K (0.9378g) and ultrasonic dissolution. The reaction was carried out at low speed in the absence of light. After the reaction, methyl tert-butyl ether (150mL) and n-hexane (70mL) were added to the reaction mixture to precipitate a solid, which was then filtered off, dissolved in 20% methanol/dichloromethane, added with silica gel powder (10g), evaporated to dryness, and subjected to column chromatography. Gradient with/dichloromethane-1% ammonia: gradient elution is carried out on 15% methanol/dichloromethane, the collected product is dried by distillation, absolute ethyl alcohol (3mL) is added, ultrasonic treatment is carried out uniformly, methyl tert-butyl ether (150mL) and n-hexane (50mL) are added, suction filtration is carried out, dissolution of absolute ethyl alcohol (3mL) is continued to be added, the methyl tert-butyl ether and the n-hexane are settled, repeated dissolution and settlement are carried out for three times, suction filtration and drying are carried out, and 0.8g of the product is obtained.

¹H-NMR(600MHz,DMSO-d₆)δ9.93–9.81(m,7H),9.09–9.01(m,14H),8.79–7.94(m,163H),7.87–6.50(m,404H),5.79–5.70(m,5H),5.29–5.23(m,18H),4.81–4.00(m,138H),3.51–3.50(m,2227H),3.25–2.64(m,177H),2.36–1.91(m,101H),1.55–1.32(m,293H),0.88–0.72(m,216H).

14. Synthesis of 29-235 (Compound No. 15)

26-250

Boc-Lys (Boc) -OH (available from Arm Pharm, 0.3769g, 1.0879mmol), 26-232 (synthesized by 22-181, 0.989mmol), HBTU (0.5627g, 1.4835mmol), HOBT (0.2005g, 1.4835mmol) was added to a 250mL flask containing it, dissolved by adding the appropriate amount of DMF (40mL), and stirred at-5 ℃ for 30 minutes. DIEA (0.7356mL, 4.4505mmol) was added slowly dropwise over 3 min. After the reaction was continued at-5 ℃ with stirring for 1 hour, it was left to stir at room temperature overnight. After the reaction was completed, the reaction solution was transferred to a 1L separatory funnel, pure water (120mL) and ethyl acetate (80mL) were added for extraction to give an organic phase, the aqueous phase was extracted with ethyl acetate (80mL x 3), the organic phases were combined, washed with a saturated sodium chloride solution (80mL x 3), concentrated to dryness, and dried in a vacuum oven to give a crude product 26-250: yield 100%.

26-255

26-250(0.989mmol) was charged to a 250mL flask, dichloromethane (10mL) and TFA (2.2182mL, 29.87mmol) were added and dissolved, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating the reaction liquid to dryness to obtain oil, dissolving the residue with ethyl acetate (50mL), adding excessive solid sodium bicarbonate to neutralize until no bubbles exist, filtering, adding silica gel powder (15g), evaporating to dryness to obtain powdery solid, loading the powdery solid by a dry method, carrying out column chromatography, carrying out gradient elution by using a dichloromethane mixed solution containing 1% ammonia water and 3% -6% methanol, collecting and concentrating, and drying in a vacuum oven to obtain a product 26-255:0.8g with the yield of 81%.

26-258

26-255(0.8g, 0.7698mmol), 41-26(2.0990g, 1.6936mmol), HBTU (0.8761g, 2.3095mmol) and HOBT (0.3121g, 2.3095mmol) were put in a 250mL flask equipped with DMF (30mL) and dissolved therein, and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (1.1458mL, 6.9285mmol) was added slowly dropwise over 3 min. After the reaction was continued at-5 ℃ with stirring for 1 hour, it was left to stir at room temperature overnight. After the reaction, the reaction solution was transferred to a 1L separatory funnel, pure water (120mL) and ethyl acetate (80mL) were added for extraction to obtain an organic phase, the aqueous phase was extracted with ethyl acetate (80mL x 3), the organic phases were combined, washed with a saturated sodium chloride solution (80mL x 3), concentrated to dryness, then dissolved with a methanol/dichloromethane (1:4) solution (200mL), silica gel powder (15g) was added, evaporated to dryness to obtain a powdery solid, dry-loading, column chromatography, gradient elution with a 3% to 5% methanol-dichloromethane mixed solution, collected, concentrated to dryness to a solid, and vacuum oven-dried to obtain 26 to 258:1.6884g of a product with a yield of 63%.

29-152

29-147 (synthesized by the method of 42-20, 1.7952g, 1.3616mmol) and 10% Pd/C (0.03g) were put into a hydrogenation reactor, then DMF (30mL) was added to dissolve it, the hydrogenation reactor was closed, after three pumping and three charging, the pressure reading on the hydrogenation reactor was 0.18MPa, and then the reaction was carried out overnight at room temperature. After the reaction was completed, the reaction solution was filtered through celite, and the filter cake was washed with DMF (20mL x 3) to give 29-152% as a product with a yield of 100%.

29-153

29-152(1.3616mmol), 42-18(6.0g, 6.2632mmol), HBTU (3.0990g, 8.1696mmol) and HOBT (1.1040g, 8.1696mmol) were put in a 500mL flask, dissolved in DMF (60mL) and reacted at 0 ℃ for 30 minutes with stirring. DIEA (4.0532mL, 24.5088mmol) was added slowly dropwise over 7 min. The reaction was continued to stir at 0 ℃ overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (30g), evaporating to be powdery, performing dry sampling, performing column chromatography, performing gradient elution with 1% ammonia water and 5% -8% methanol dichloromethane mixed solution, collecting and concentrating, and drying in a vacuum oven to obtain the product 29-153:3.6g with the yield of 47%.

29-157

29-153(3.6g, 1.3493mmol) was charged to a 250mL flask, dichloromethane (10mL) and TFA (1.5031mL, 20.2401mmol) were added to dissolve, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating the reaction liquid to dryness to obtain oil, adding powdery solid in the reaction liquid of methyl tert-butyl ether (40mL), separating out, performing suction filtration, cleaning a filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, and drying in a vacuum oven to obtain a product 29-157 with the yield of 100%.

29-162

29-157(1.3493mmol), GFLG-PCB (synthesized by 30-33, 1.220g, 1.4842mmol), HBTU (0.7676g, 2.0240mmol), HOBT (0.2735g, 2.0240mmol) were placed in a 250mL flask, dissolved in DMF (60mL), and the reaction was stirred at 0 ℃ for 30 minutes. DIEA (1.0mL, 6.0719mmol) was added slowly dropwise over 3 minutes. The reaction was continued to stir at 0 ℃ overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, and drying in a vacuum oven to obtain the product 29-162 with the yield of 100%.

29-164

29-162(1.3493mmol) was placed in a 250mL flask, DMF (15mL) was added and dissolved, morpholine (1.2mL, 13.493mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (15g), evaporating to be powdery, performing dry sampling, performing column chromatography, performing gradient elution with 1% ammonia water and 5% -9% methanol dichloromethane mixed solution, collecting and concentrating, and drying in a vacuum oven to obtain the product 29-164:4.1g with the yield of 59%.

29-167

29-164(4.1g, 0.7795mmol), Boc-Gly-OH (0.1502g, 8574mmol), HBTU (0.4434g, 1.1692mmol) and HOBT (0.1578g, 0.1.1692mmol) were put in a 250mL flask, dissolved by adding an appropriate amount of DMF (40mL), and the reaction was stirred at-5 ℃ for 30 minutes. DIEA (0.5797mL, 3.5076mmol) was added slowly dropwise and the reaction was stirred at-5 ℃ for 1 hour and then allowed to stir at room temperature overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, and drying in a vacuum oven to obtain the product with the yield of 100%.

29-194

29-167(0.7795mmol) was charged to a 250mL flask, dichloromethane (5mL), TFA (0.8683mL, 11.6921mmol) were added, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating the reaction solution to dryness until the reaction solution is oily, adding methyl tert-butyl ether (250mL) to precipitate a solid, performing suction filtration, cleaning a filter cake with methyl tert-butyl ether (100mL x 3), dissolving the filter cake with a methanol (60mL) dichloromethane (240mL), adding silica gel powder (15g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a dichloromethane mixed solution of 1% of ammonia water and 3% -8% of methanol, collecting and concentrating, evaporating to dryness to obtain a solid, and drying in a vacuum oven to obtain a product: 3.2g, yield 76%.

29-196

Reactants 26 to 258(0.2545g, 0.073mmol) and 10% Pd/C (40mg) were weighed into a hydrogenation reactor, DMF (30mL) was added, hydrogen was introduced under a pressure Pa of 1.8MPa, and the reaction was stirred at room temperature overnight. After the reaction was complete, the reaction solution was filtered through celite, washed with DMF (20mL × 3), and the DMF solutions were combined as the starting material for the next reaction.

29-197

The reaction products 29-194(3.2g, 0.9211mmol), 29-196(0.073mmol), HBTU (0.5g, 1.3159mmol) and HOBT (0.1778g, 1.3159mmol) were taken and placed in a 250mL reaction flask, DMF (30mL) was added to dissolve the mixture, the mixture was stirred at-5 ℃ for 30 minutes, DIEA (0.6525mL, 3.9477mmol) was slowly added dropwise, the reaction mixture was stirred at-5 ℃ for 1 hour, and then the mixture was stirred at room temperature overnight. After the reaction, methyl tert-butyl ether (100mL) was added, the mixture was placed in a refrigerator for 30 minutes, taken out, a solid precipitated, filtered, collected by suction, and a filter cake was dissolved in a methanol/dichloromethane (1:4) solution (100mL), and then silica gel powder (15g) was added and evaporated to dryness to a powder. Dry loading, column chromatography, eluting with 1% ammonia water: 3% methanol/dichloromethane-1% ammonia: 10% methanol in dichloromethane. The product weighed 2.2g, 67% yield.

¹H-NMR(600MHz,DMSO-d₆)δ10.17-10.13(m,10H),9.10-8.84(m,88H),8.35–7.43(m,676H),7.37-6.69(m,420H),5.87-5.73(m,12H),4.60–4.09(m,217H),3.96–3.43(m,1173H),3.21–2.65(m,524H),2.30–1.66(m,440H),1.59–1.08(m,541H),0.92-0.75(m,360H).

29-201

29-197(2.2g) was weighed out, dichloromethane (10mL) was added, TFA (4mL) was added, the mixture was dissolved by sonication, and the reaction was stirred. After the reaction, the reaction solution was evaporated to dryness to an oil, and after methyl t-butyl ether (100mL) was added, a solid precipitated, and the solid was filtered off, and the filter cake was collected by suction filtration, then dissolved in a methanol/dichloromethane (1:4) solution (100mL), and silica gel powder (15g) was added and evaporated to a powder. Column chromatography, dry loading, loading with 1% ammonia water: 4% methanol/dichloromethane-1% ammonia: 10% methanol in dichloromethane. The product weighed 1.4g, and the yield was 52%.

¹H-NMR(600MHz,DMSO-d₆)δ10.13-10.07(m,10H),9.37-9.29(m,63H),9.09–8.84(m,51H),8.55-8.49(m,20H),8.03-7.38m,560H),7.12-7.06(m,376H),6.74–6.64(m,13H),5.36-5.28(m,12H),4.57–4.19(m,153H),3.85-3.60(m,803H),3.16–3.01(m,317H),2.30–2.10(m,150H),1.91-1.72(m,143H),1.54-1.44(m,127H),1.34-1.15(m,940H),0.93–0.76(m,360H).

29-235

M-SCM-20K (1.2163g, 0.0589mmol) was added to the reaction mixture 29-201(1.0g), and DMF solution (20mL) was added to dissolve the mixture, and the mixture was stirred at low speed and protected from light for one week. After the reaction, methyl tert-butyl ether (100mL) was added, the mixture was placed in a refrigerator for 30 minutes, taken out, a solid precipitated, filtered, collected by suction, and a filter cake was dissolved in a methanol/dichloromethane (1:4) solution (100mL), and then silica gel powder (15g) was added and evaporated to dryness to a powder. Column chromatography, dry loading, loading with 1% ammonia water: 3% methanol/dichloromethane-1% ammonia: elution with a gradient of 8% methanol in dichloromethane. The product is evaporated to dryness, absolute ethyl alcohol (10mL) is added, ultrasonic treatment is carried out to form a homogeneous phase, and n-hexane (50mL) is added for sedimentation. The sedimentation was repeated three times. Vacuum drying to obtain 1.3g of product. The yield was 61%.

¹H-NMR(600MHz,DMSO-d₆)δ10.05-9.88(m,10H),9.41-9.30(m,62H),9.11–8.89(m,50H),8.58-8.51(m,20H),8.11-7.40m,562H),7.15-7.08(m,378H),6.78–6.66(m,13H),5.38-5.31(m,15H),4.58–4.21(m,151H),3.95-3.65(m,796H),3.52–3.43(m,3706H),3.19–3.07(m,321H),2.32–2.13(m,148H),1.93-1.76(m,146H),1.58-1.46(m,125H),1.36-1.16(m,935H),0.98–0.74(m,366H).

15. Synthesis of 37-108 (Compound No. 17)

The route is as follows

37-39

Placing erythritol (purchased from Aladdin, 2.5g, 20.4717mmol) into a 500mL flask, adding tetrahydrofuran (130mL) to dissolve, placing the reaction at 0 ℃, slowly adding a tetrahydrofuran solution (98.2642mL, 98.2642mmol) of potassium tert-butoxide (1mol/L) under nitrogen protection, stirring for 5 hours, slowly adding benzyl bromoacetate (14.5937mL, 92.1225mmol), stirring for 2 hours, and moving the reaction to room temperature to stir overnight. After completion of the reaction, the reaction mixture was transferred to a 1L separatory funnel, and saturated sodium hydrogencarbonate (200mL) and ethyl acetate (250mL) were added thereto, followed by shaking and extraction. Washing the water phase with ethyl acetate (200mL x 2), combining the organic phases, washing with saturated saline (200mL x 2), concentrating and evaporating to dryness, adding dichloromethane for dissolution, adding silica gel powder (50g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a 1.5-5% petroleum ether mixed solution of ethyl acetate, collecting and concentrating, evaporating to dryness, and drying in a vacuum oven to obtain a product 37-39: 4.8g, yield 33%.

37-49

37-39(1.03g, 1.441mmol) was charged into a hydrogenation reaction vessel, 10% Pd/C (0.040g) was added thereto, DMF (30mL) was added thereto and dissolved therein, and hydrogen gas was introduced under a pressure Pa of 1.8MPa, and the reaction was stirred at room temperature overnight. After the reaction was complete, the reaction solution was filtered through celite, washed with DMF (20mL × 3), and the DMF solutions were combined as the starting material for the next reaction.

37-67

37-49(0.14g, 0.3013mmol), 37-66 (synthesized by 35-4, 1.9g, 1.506mmol), HBTU (1.537g, 4.0528mmol), HOBT (0.5473g, 4.0528mmol) were added to a 250mL flask, 30mL of DMMF was added to dissolve, the mixture was stirred at-5 ℃ for 30 minutes, DIEA (2.01mL, 12.1585mmol) was slowly added dropwise, the reaction was continued for 1 hour after the dropwise addition was completed, and the mixture was left to stir at room temperature overnight. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding deionized water (200mL) and ethyl acetate (200mL) into the separating funnel, extracting to obtain an organic phase, washing the water phase with ethyl acetate (200mL x 2), combining the organic phases, washing the organic phase with saturated saline (200mL x 2), evaporating to dryness, and drying in an oven to obtain a product 37-67: 1.61 g.

37-68

Putting 37-67(1.6053g, 0.3013mmol) into a 250mL flask, adding DMF (30mL) to dissolve, adding morpholine (2.6245mL, 30.125mmol), stirring at room temperature for 1 hour, finishing the reaction, transferring the reaction solution into a 1L separating funnel, adding saturated saline (200mL) and ethyl acetate (200mL) to extract to obtain an organic phase, washing an aqueous phase with ethyl acetate (200mL x 2), combining the organic phases, washing the organic phase with saturated saline (200mL x 1), concentrating and evaporating to dryness, adding silica gel powder (20g), evaporating to dryness to obtain a powdery solid, loading by a dry method, carrying out column chromatography, eluting with a mixed solution of 0.5% ammonia water and 5% -8% methanol in dichloromethane, collecting and concentrating, evaporating to dryness, and drying in an oven to obtain 37-68:0.59g of a product, wherein the yield is 45%.

37-79

Putting 33-143(3.7248g, 15.0mmol) into a 250mL flask, adding dichloromethane (30mL) to dissolve, adding triethylamine (2.598mL, 18.4912mmol), stirring at 0 ℃ for 30 minutes, slowly adding phenyl chloroformate (1.39mL, 11.0947mmol) dropwise, stirring at room temperature overnight, ending the reaction, transferring the reaction liquid into a 1L separating funnel, adding dichloromethane (200mL) and deionized water (200mL) to extract to obtain an organic phase, washing an aqueous phase with dichloromethane (200mL x 1), combining the organic phases, adding silica gel powder (10g), evaporating to obtain a powdery solid, loading by a dry method, performing column chromatography, and eluting with a mixed solution of 3% methanol and dichloromethane to obtain a product 37-79:2.34g, wherein the yield is 69%.

37-81

Adding 37-68(0.59g, 0.1329mmol) and 37-79(0.2938g, 0.7975mmol) into a 250mL flask, adding DMF (40mL) to dissolve, stirring overnight at 80 ℃, after the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding ethyl acetate (200mL) and saturated saline (200mL) to extract to obtain an organic phase, washing the organic phase with saturated saline (150mL), adding silica gel powder (15g), evaporating to obtain a powdery solid, performing dry loading, performing column chromatography, and eluting with a mixed solution of 6% -8% methanol and dichloromethane to obtain a product 37-81:0.27g, wherein the yield is 39%.

¹H-NMR(400MHz,DMSO-d₆)δ8.32–8.14(m,20H),7.41–7.38(m,80H),7.26–7.10(m,4H),6.21–5.93(m,8H),5.34–5.20(m,32H),4.51–4.45(m,16H),3.96–3.81(m,24H),3.63–3.04(m,32H),2.21–2.05(m,64H),1.84–1.55(m,16H),1.48–1.33(m,36H),1.25(m,9H).

37-89

37-81(0.27g, 0.0488mmol) was charged into a hydrogenation reactor, 10% Pd/C (0.070g) was added thereto, DMF (30mL) was added thereto and dissolved therein, hydrogen gas was introduced thereinto under a pressure of 1.8MPa, and the reaction was stirred at room temperature overnight. After the reaction was complete, the reaction solution was filtered through celite, washed with DMF (20mL × 3), and the DMF solutions were combined as the starting material for the next reaction.

37-91

37-89(0.1998g, 0.0488mmol), 37-38 (1.769 g, 0.87796mmol synthesized by 39-20 method), HBTU (0.4442g, 1.1712mmol) and HOBT (0.1583g, 1.1712mmol) are added into a 250mL flask, 30mL of DMF is added to dissolve the materials, the mixture is stirred at-5 ℃ for 30 minutes, DIEA (0.58mL, 3.5136mmol) is slowly added dropwise, the reaction is continued for 1 hour after the dropwise addition is finished, and then the mixture is stirred at room temperature for overnight reaction. After the reaction is finished, adding n-hexane (250mL) and methyl tert-butyl ether (40mL), layering, pouring the upper layer, adding n-hexane (250mL) and methyl tert-butyl ether (40mL) into the lower oily liquid phase, continuing to settle, repeating the process for 4 times, finally obtaining a viscous oily substance, adding methyl tert-butyl ether (150mL), separating out a solid, filtering, adding silica gel powder (15g), evaporating to obtain a powdery solid, loading by a dry method, performing column chromatography, eluting by a mixed solution of 1% of ammonia water and 8% -10% of methanol in dichloromethane, collecting, concentrating, evaporating to dryness, and drying in an oven to obtain a product 37-91: 1.2g, yield: 68%.

37-101

37-91(1.2g, 0.0488mmol) was charged into a 250mL flask, dichloromethane (20mL), trifluoroacetic acid (0.7248mL, 9.76mmol) was added, and the reaction was stirred at room temperature overnight. After the reaction is finished, adding methyl tert-butyl ether (100mL), precipitating a solid, performing suction filtration, cleaning a filter cake with methyl tert-butyl ether (40mL x 3), dissolving the filter cake with a methanol (20mL) dichloromethane (80mL), adding silica gel powder (15g), evaporating the silica gel powder to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 8% -12% methanol, collecting and concentrating the eluent, evaporating the eluent to dryness to obtain a solid, and drying the solid in a vacuum oven to obtain a product 37-101: 0.69g, yield 40%.

37-108

37-101(0.3g, 0.0084mmol) was placed in 250mL, dissolved in DMF (35mL), dissolved with M-SCM-10K (0.5348g, 0.0505mmol from Kenky) by sonication, and the reaction was stirred at low speed for 7 days at room temperature in the dark. After the reaction, adding n-hexane (150mL) and methyl tert-butyl ether (30mL), pouring the supernatant, adding n-hexane (130mL) and methyl tert-butyl ether (30mL) into the lower layer liquid, repeating the reaction for 3 times to obtain a viscous oily substance, adding methyl tert-butyl ether (100mL), separating out a solid, filtering, washing a filter cake with methyl tert-butyl ether (50mL x 3), dissolving the filter cake with a methanol (30mL) and dichloromethane (120mL), adding silica gel powder (15g), evaporating to dryness to obtain a powdery solid, performing dry sampling, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 6% -9% methanol, collecting, concentrating, evaporating to a solid, drying in a vacuum oven for 1 hour, adding absolute ethanol (7mL) and dichloromethane (20mL) to dissolve, adding methyl tert-butyl ether (100mL), separating out a solid, filtering, washing the filter cake with methyl tert-butyl ether (50mL x 2), drying the filter cake in a vacuum oven to obtain a product 37-108: 0.3g, yield: 52 percent.

¹H-NMR(600MHz,DMSO-d₆)δ8.49–8.22(m,49H),8.12–8.03(m,212H),7.91–7.66(m,48H),7.36–7.01(m,388H),6.97–6.85(m,16H),6.70–6.01(m,40H),5.34–5.16(m,64H),4.92–4.26(m,144H),4.09–4.03(m,152H),3.76–3.54(m,86H),3.51–3.41(m,3722H),3.44–3.19(m,160H),2.93–2.72(m,48H),2.46–2.05(m,192H),1.84–1.55(m,144H),1.25–0.91(m,297H).

16. Synthesis of 29-226 (Compound No. 8)

The route is as follows

29-168

Fmoc-Glu-OtBu (4g, 9.401mmol, available from Ilykok), H-Gly-OtBu HCl (1.7g, 10.3415mmol, available from Ilykok), HBTU (5.3g, 14.1021mmol) and HOBT (1.9g, 14.1021mmol) were placed in a 500mL round bottom flask, which was then dissolved by adding DMF (70mL) and the mixture was stirred at 0 ℃ for 30 minutes. DIEA (7.0mL, 42.3061mmol) was then added slowly dropwise and after completion the mixture was stirred at 0 ℃ overnight. After completion of the reaction, the reaction solution was transferred to a 1L separatory funnel, pure water (100mL) and ethyl acetate (80mL) were added for extraction to give an organic phase, the aqueous phase was extracted with ethyl acetate (80mL x 2), the organic phases were combined, extracted with a saturated sodium chloride solution (150mL x 3), concentrated to dryness, and dried to give a crude product.

29-171

29-168(9.401mmol) was placed in a 500mL flask, dichloromethane (10mL) and TFA (7.0mL, 94.01mmol) were added and dissolved, and the reaction was stirred at room temperature overnight. After the reaction was completed, the reaction solution was evaporated to dryness to obtain an oil, and then transferred to a 1L separatory funnel, a saturated sodium bicarbonate solution (100mL) was added to adjust the pH to alkaline, and then ethyl acetate (80mL) was added to extract to obtain an organic phase, the aqueous phase was extracted with ethyl acetate (80mL x 2), the organic phases were combined, extracted with a saturated sodium chloride solution (100mL x 3), concentrated to dryness, and dried in a vacuum oven to obtain a crude product with a yield of 100%.

29-173

Mixing 29-171(9.401mmol), H-Glu- (OtBu)₂(2.6789g, 10.3415mmol), HBTU (5.3g, 14.1021mmol) and HOBT (1.9g, 14.1021mmol) were placed in a 500mL round bottom flask, then DMF (70mL) was added and dissolved, and the mixture was stirred at 0 ℃ for 30 minutes. DIEA (7.0mL, 42.3061mmol) was then added slowly dropwise, and after the addition was complete, the reaction was stirred at 0 deg.C overnight. After the reaction, the reaction mixture was transferred to 1LTo the liquid funnel, purified water (100mL) and ethyl acetate (80mL) were added for extraction to give an organic phase, the aqueous phase was extracted with ethyl acetate (80mL x 2), the organic phases were combined, extracted with saturated sodium chloride solution (150mL x 3), concentrated to dryness and dried to give the crude product.

29-184

29-173(9.401mmol) was placed in a 250mL flask, DMF (10mL) was added and dissolved, morpholine (8.2mL, 94.01mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction was completed, the reaction solution was transferred to a 1L separatory funnel, pure water (100mL) and ethyl acetate (80mL) were added for extraction to obtain an organic phase, an aqueous phase was extracted with ethyl acetate (80mL x 2), the organic phases were combined, extracted with a saturated sodium chloride solution (150mL x 3), and then concentrated, and silica gel powder (20g) was added to dry-evaporate to a powder, dry-loading, column chromatography, gradient elution with a mixed solution of 1% ammonia and 4% to 8% methanol in dichloromethane, collection, concentration, and vacuum oven drying to obtain 3.7g of a product, with a yield of 74%.

29-185

Fmoc-Gly-OH (2.2g, 7.5985mmol, available from Ilykok), 29-184(3.7g, 6.9078 mmol), HBTU (3.9g, 10.3616mmol), HOBT (1.400g, 10.3616mmol) were placed in a 500mL round bottom flask, which was then dissolved by adding DMF (70mL), and the mixture was stirred at 0 ℃ for 30 minutes. DIEA (5.1mL, 31.0849mmol) was then added slowly dropwise, and after the addition was complete, the reaction was stirred at 0 deg.C overnight. After the reaction was completed, the reaction solution was transferred to a 1L separatory funnel, pure water (100mL) and ethyl acetate (80mL) were added for extraction to obtain an organic phase, an aqueous phase was extracted with ethyl acetate (80mL x 2), the organic phases were combined, extracted with a saturated sodium chloride solution (150mL x 3), and then concentrated, and silica gel powder (25g) was added to dry-evaporate to a powder, dry-loading, column chromatography, gradient elution with a mixed solution of 3% to 7% methanol in dichloromethane, collection, concentration, and vacuum oven drying to obtain 4.5g of a product, with a yield of 77%.

29-209

29-185(0.9281g, 1.0354mmol), 10% Pd/C catalyst (30mg) were added to the hydrogenation apparatus, then DMF (30mL) was added to dissolve, the hydrogenation apparatus was closed, hydrogen was charged to allow the pressure on the hydrogenation apparatus to read 18psi, and then stirring was carried out overnight at ambient temperature. After the reaction is finished, taking out the reaction liquid, uniformly dropwise adding the reaction liquid onto a suction filter funnel filled with compacted diatomite, cleaning the reaction device and the diatomite with DMF (30mL x 3), and collecting filtrate to obtain a reaction product.

K-7

Reacting Boc-Glu- (OH)₂(0.39g, 1.5738mmol, ex ImmunoKat), 34-17(4.5g, 3.3051mmol), HBTU (1.8g, 4.7216mmol) and HOBT (0.6380g, 4.7216mmol) were placed in a 250mL round bottom flask, then DMF (50mL) was added and dissolved, and the mixture was stirred at 0 ℃ for 30 minutes. DIEA (2.3mL, 14.1647mmol) was then added slowly dropwise, and after the addition was complete, the reaction was stirred at 0 deg.C overnight. After completion of the reaction, the reaction solution was transferred to a 1L separatory funnel, and pure water (100mL) and ethyl acetate (80mL) were added to conduct extraction to obtain an organic phase, and the aqueous phase was extracted with ethyl acetate (80mL x 2), and the organic phases were combined, extracted with a saturated sodium chloride solution (150mL x 3), concentrated to dryness, and dried to obtain a crude product.

K-8

K-7(1.5738mmol) was charged to a 500mL flask, dichloromethane (10mL) and TFA (1.7531mL, 15.738mmol) were added to dissolve, and the reaction was stirred at room temperature overnight. After the reaction, the reaction solution was evaporated to dryness to obtain an oil, the oil was transferred to a 1L separating funnel, a saturated sodium bicarbonate solution (100mL) was added to adjust the pH to alkaline, then ethyl acetate (80mL) was added to extract the oil to obtain an organic phase, the aqueous phase was extracted with ethyl acetate (80mL x 2), the organic phases were combined, extracted with a saturated sodium chloride solution (100mL x 3), concentrated again, silica gel powder (20g) was added to evaporate the oil to a powder, dried to obtain a sample, subjected to column chromatography, gradient-eluted with a mixed solution of 3% to 8% methanol and dichloromethane, collected, concentrated, dried in a vacuum oven to obtain 3.2g of a product, and the yield was 77%.

29-210

K-8(3.0g, 1.1389mmol), 29-209(1.0354mmol), HBTU (0.5891g, 1.5531mmol) and HOBT (0.2099g, 1.5531mmol) were placed in a 250mL round bottom flask, then DMF (50mL) was added and dissolved, and the mixture was stirred at 0 ℃ for 30 minutes. DIEA (0.7701mL, 4.6595mmol) was then added slowly dropwise, and after the addition was complete, the reaction was stirred at 0 deg.C overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out solids in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x3), collecting the filter cake, dissolving the filter cake with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (10g), evaporating to be powdery, performing dry sampling, performing column chromatography, performing gradient elution with 3% -8% methanol dichloromethane mixed solution, collecting, concentrating, and drying in a vacuum oven to obtain 2.7g of a product with the yield of 79%.

K-10

Mixing 24-36(4g, 15.1924mmol), H-Glu- (OBn)₂(6.9g, 13.8112mmol, ex ImmunoKat), HBTU (7.8567g, 20.7169mmol), HOBT (2.8g, 20.7169mmol) were placed in a 500mL round bottom flask, then DMF (70mL) was added and dissolved, and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (10.2723mL, 62.1506mmol) was then added slowly dropwise, and after stirring at-5 ℃ for 1 hour, the reaction flask was transferred to room temperature and stirred overnight. After the reaction, the reaction solution was transferred to a 1L separatory funnel, and saturated sodium bicarbonate solution (100mL) and ethyl acetate (80mL) were added to extract to obtain an organic phase, an aqueous phase was extracted with ethyl acetate (80mL x 2), the organic phases were combined, extracted with saturated sodium chloride solution (150mL x3), and then concentrated and added with silica gel powder (20g), evaporated to powder, dried to sample, subjected to column chromatography, gradient eluted with a mixed solution of 1% to 3% methanol in dichloromethane, collected and concentrated, and dried in a vacuum oven to obtain 8.7g of a product with a yield of 62%.

K-11

K-10(2.7g, 4.7469mmol), 10% Pd/C catalyst (30mg) were added to the hydrogenation apparatus, then DMF (30mL) was added to dissolve, the hydrogenation apparatus was closed, hydrogen was charged to allow the pressure on the hydrogenation apparatus to read 18psi, and then stirred at ambient temperature overnight. After the reaction is finished, taking out the reaction liquid, uniformly dropwise adding the reaction liquid onto a suction filter funnel filled with compacted diatomite, cleaning the reaction device and the diatomite with DMF (30mL x 3), and collecting filtrate to obtain a reaction product.

K-12

K-11(2.1484mmol), G-SN38-TBDPS (synthesized by 25-200 methods, 3.0G, 4.3614mmol), HBTU (2.4443G, 6.4453mmol) and HOBT (0.8710G, 6.4453mmol) were placed in a 500mL round-bottomed flask, then DMF (50mL) was added and dissolved, and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (3.1959mL, 19.3360mmol) was then added slowly dropwise, and after completion of the addition, the reaction was stirred at-5 ℃ for 1 hour, and the reaction flask was transferred to room temperature and stirred overnight. And after the reaction is finished, adding methyl tert-butyl ether (80mL), putting the reaction bottle into a refrigerator for 30 minutes, taking out, separating out a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (30mL x 3), collecting the filter cake, and drying to obtain a crude product.

K-13

K-12(2.1484mmol) was charged to a 500mL flask, dichloromethane (5mL) and TFA (2.3932mL, 32.226mmol) were added and dissolved, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating the reaction liquid to be oily, adding methyl tert-butyl ether (60mL), precipitating a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (30mL x 3), collecting the filter cake, dissolving the filter cake with a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (20g), evaporating to be powdery, performing dry loading, performing column chromatography, performing gradient elution with a dichloromethane mixed solution of 3% -8% methanol, collecting, concentrating, and drying in a vacuum oven to obtain 2.7g of a product with the yield of 72%.

K-14

K-11(2.5985mmol), 42-18(6.0g, 6.2363mmol), HBTU (2.9563g, 7.7953mmol) and HOBT (1.0534g, 7.7953mmol) were placed in a 500mL round bottom flask, then DMF (50mL) was added and dissolved, and the mixture was stirred at-5 ℃ for 30 minutes. DIEA (3.8653mL, 23.360mmol) was then added slowly dropwise, and after completion of the addition, the reaction was stirred at-5 ℃ for 1 hour, and the reaction flask was transferred to room temperature and stirred overnight. And after the reaction is finished, adding methyl tert-butyl ether (80mL), putting the reaction bottle into a refrigerator for 30 minutes, taking out, separating out a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (30mL x 3), collecting the filter cake, and drying to obtain a crude product.

K-15

K-14(2.5985mmol) was charged to a 500mL flask, dichloromethane (5mL) and TFA (2.8946mL, 38.9775mmol) were added and dissolved, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating the reaction liquid to be oily, adding methyl tert-butyl ether (60mL), precipitating a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (30mL x 3), collecting the filter cake, dissolving the filter cake with a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (20g), evaporating to be powdery, performing dry loading, performing column chromatography, performing gradient elution with a dichloromethane mixed solution of 1% of ammonia water and 3% -8% of methanol, collecting, concentrating, and drying in a vacuum oven to obtain 3.3g of a product with the yield of 61%.

29-212

29-210(0.6579g, 0.1972mmol), 10% Pd/C catalyst (30mg) were added to the hydrogenation apparatus, then DMF (30mL) was added to dissolve, the hydrogenation apparatus was closed, hydrogen was charged to allow the pressure on the hydrogenation apparatus to read 18psi, and then stirring was carried out overnight at ambient temperature. After the reaction is finished, taking out the reaction liquid, uniformly dropwise adding the reaction liquid onto a suction filter funnel filled with compacted diatomite, cleaning the reaction device and the diatomite with DMF (30mL x 3), and collecting filtrate to obtain a reaction product.

29-213

K-15(3.3g, 1.6566mmol), 29-212(0.5391g, 0.1972mmol), HBTU (0.8975g, 2.3667mmol) and HOBT (0.3198g, 2.3667mmol) were placed in a 500mL round-bottomed flask, then DMF (50mL) was added and dissolved, and the mixture was stirred at 0 ℃ for 30 minutes. DIEA (1.1735mL, 7.1mmol) was then added slowly dropwise, and after the addition was complete, the reaction was stirred at 0 deg.C overnight. And after the reaction is finished, adding methyl tert-butyl ether (80mL), putting the reaction bottle into a refrigerator for 30 minutes, taking out, separating out a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (30mL x 3), collecting the filter cake, and drying to obtain a crude product.

29-214

29-213(0.1972mmol) was placed in a 500mL flask, dichloromethane (5mL) and TFA (0.22mL, 2.958mmol) were added and dissolved, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating the reaction liquid to be oily, adding methyl tert-butyl ether (60mL), precipitating a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (30mL x 3), collecting the filter cake, dissolving the filter cake with a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (20g), evaporating to be powdery, performing dry loading, performing column chromatography, performing gradient elution with a dichloromethane mixed solution of 3% -9% methanol, collecting, concentrating, and drying in a vacuum oven to obtain 2.7g of a product with the yield of 69%.

29-216

29-214(2.7g, 0.1405mmol), K-13(0.5117g, 0.2951mmol), HBTU (0.1599g, 0.4215mmol) and HOBT (0.057g, 0.4215mmol) were placed in a 500mL round-bottomed flask, then DMF was added to dissolve it, and the mixture was stirred at 0 ℃ for 30 minutes. DIEA (0.2090g, 1.2645mmol) was then added and stirring continued at 0 deg.C overnight. And after the reaction is finished, adding methyl tert-butyl ether (80mL), putting the reaction bottle into a refrigerator for 30 minutes, taking out, separating out a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (30mL x 3), collecting the filter cake, and drying to obtain a crude product.

29-220

29-216(0.1405mmol) was placed in a 250mL flask, DMF (8mL) was added and dissolved, morpholine (0.3378mL, 3.8778mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving with methanol/dichloromethane (1:4) solution (10mL), settling with methyl tert-butyl ether, and repeating the operation for 3 times to obtain a crude product.

29-221

M-SCM-5K (0.8773g, 0.1667mmol, available from Kerkin) was charged into a 250mL round-bottomed flask, DMF (20mL) was added to dissolve, DIEA (0.3443mL, 2.0833mmol) was added, the reaction flask was stirred at 0 ℃ for 10 minutes, and then a solution of 29-220(1.0g, 0.0416mmol) in DMF (20mL) was slowly added dropwise over 2 hours from the dropping funnel. After the dropwise addition, the mixture was stirred at 0 ℃ for 15 minutes, and then the reaction flask was transferred to room temperature and stirred at a low speed in the dark for one week. After the reaction is finished, adding methyl tert-butyl ether (80mL), putting the reaction bottle into a refrigerator for 30 minutes, taking out, separating out a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (30mL x 3), collecting the filter cake, dissolving the filter cake with a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (10g), drying to obtain a powder, performing dry sampling, performing column chromatography, performing gradient elution with a 4-15% methanol dichloromethane mixed solution, collecting and concentrating, and drying in a vacuum oven to obtain 1.6g of a product with the yield of 58%.

¹H-NMR(600MHz,DMSO-d₆)δ9.27–9.02(m,62H),8.36-7.95(m,171H),7.77-7.51(m,115H),7.34–7.12(m,157H),6.69-6.65(m,13H),4.57-4.50(m,34H),4.31-4.17(m,71H),3.91-3.80(m,207H),3.55-3.45(m,1430H),3.26-3.20(m,12H),3.12-3.05(m,71H),2.93-2.88(m,12H),2.81-2.65(m,50H),2.25-2.19(m,61H),1.89–1.19(m,298H),0.85-0.79(m,96H).

29-226

To a 250mL flask containing 29-221, tetrahydrofuran (10mL) was added, diluted hydrochloric acid (10mL, 0.05mol/L) was added and dissolved, TBAF (67mg, 0.256mmol) was added, and the reaction was stirred at room temperature for 3 hours. After the reaction is finished, evaporating the reaction solution to be oily, adding absolute ethyl alcohol to remove water, evaporating to dryness, repeating for 3 times to obtain a solid substance, and dissolving the solid substance with DMF (20 mL); it was then precipitated with isopropanol (30mL) and repeated 3 times. Dissolving the solid in dichloromethane and small amount of anhydrous ethanol, settling with methyl tert-butyl ether, repeating for 3 times, and drying to obtain final product 1.1g with yield of 72%

¹H-NMR(600MHz,DMSO-d₆)δ9.06-9.00(m,40H),8.29-8.22(m,50H),8.15–7.95(m,96H),7.73–7.47(m,121H),7.29-7.08(m,170H),4.59-4.11(m,99H),3.95–3.73(m,240H),3.53-3.48(m,1434.3H),3.24–3.00(m,110H),2.92-2.87(m,9H),2.80-5.65(m,45H),2.14-2.08(m,45H),1.91–1.28(m,244H),0.85-0.81(m,96H)

17. Synthesis of 27-253 (Compound No. 9)

The route is as follows

27-181

29-168(3.33g, 5.8079mmol) was placed in a 500mL round bottom flask, DMF (20mL) was added and sonicated to dissolve completely, then morpholine (5.06mL, 58.079mmol) was added and the reaction was allowed to stir at room temperature for 2 hours. After the reaction is finished, adding saturated sodium chloride solution (200mL) and ethyl acetate (200mL) for extraction, collecting an organic phase, extracting an aqueous phase with ethyl acetate (200mL multiplied by 3), collecting and combining the organic phases, concentrating under reduced pressure, and drying to obtain 4.2g of an overweight product.

27-184

27-181(2.03g, 5.8079mmol), Fmoc-Gly-OH (1.72g, 5.8079mmol, available from Ikano), HBTU (3.3g, 8.7119mmol) and HOBT (1.13g, 8.7119mmol) were placed in a 500mL round-bottomed flask, then DMF (100mL) was added to dissolve it, the reaction was stirred at-5 ℃ for 30 minutes, then DIEA (4.32mL, 26.1356mmol) was slowly added dropwise, after which time DIEA (4.32mL, 26.1356mmol) was reacted at-5 ℃ for 2 hours, and then the reaction was left to stir at room temperature overnight. After the reaction is finished, adding saturated sodium chloride solution (200mL) and ethyl acetate (200mL) for extraction, collecting an organic phase, extracting an aqueous phase with ethyl acetate (200mL multiplied by 3), collecting and combining the organic phases, concentrating under reduced pressure, adding 60mL of silica gel powder, evaporating to dryness, loading by a dry method, and carrying out column chromatography. Eluting with eluent (2% -3% methanol: 97% -98% dichloromethane), collecting liquid, concentrating under reduced pressure, and drying. Product 2.68g was obtained, yield: 73.42 percent.

27-201

27-184(2.68g, 4.9669mmol) and 10% Pd/C catalyst (100mg) were added to a hydrogenation reaction apparatus, then DMF (50mL) was added to dissolve it, the hydrogenation reaction apparatus was closed, hydrogen gas was introduced under a pressure of 18psi, then stirring was carried out overnight at room temperature, after completion of the reaction was found by TLC, the reaction solution was taken out and uniformly dropped into a sand core funnel containing compacted diatomaceous earth, suction filtration was carried out, and the diatomaceous earth was washed with DMF (30 mL. times.3) to obtain a reaction product solution.

27-202

27-201(1.57g, 2.9123mmol), 34-17(3.34g, 2.6475 mmol), HBTU (1.50g, 3.9713mmol) and HOBT (0.54g, 3.9713mmol) were placed in a 500mL round-bottomed flask, then DMF (100mL) was added and dissolved, the reaction was stirred at-5 ℃ for 30 minutes, then DIEA (1.97mL, 11.9138mmol) was slowly added dropwise, after which it was reacted at-5 ℃ for 2 hours, and then the reaction was left to stir at room temperature overnight. After the reaction is finished, adding saturated sodium chloride solution (200mL) and ethyl acetate (200mL) for extraction, collecting an organic phase, extracting an aqueous phase with ethyl acetate (200mL multiplied by 3), collecting and combining the organic phases, concentrating under reduced pressure, adding 60mL of silica gel powder, evaporating to dryness, loading by a dry method, and carrying out column chromatography. Eluting with eluent (3% methanol: 97% dichloromethane), collecting liquid, concentrating under reduced pressure, and drying. Product 3.58g was obtained, yield: 75.84 percent.

27-204

27-202(3.58g, 1.9456mmol) and 10% Pd/C catalyst (100mg) were added to a hydrogenation reaction apparatus, then DMF (50mL) was added to dissolve it, the hydrogenation reaction apparatus was closed, hydrogen gas was charged under a pressure of 18psi, then stirring was carried out overnight at room temperature, after completion of the reaction was found by TLC, the reaction solution was taken out and uniformly dropped into a sand core funnel containing compacted diatomaceous earth, suction filtration was carried out, and the diatomaceous earth was washed with DMF (30 mL. times.3) to obtain a reaction product solution.

M-6

K-11(0.65g, 1.6590mmol), 30-33(3g, 3.6498mmol), HBTU (0.94g, 2.4885mmol) and HOBT (0.34g, 2.4885mmol) were placed in a 500mL round-bottomed flask, then DMF (100mL) was added and dissolved, the reaction was stirred at-5 ℃ for 30 minutes, then DIEA (1.23mL, 7.4655mmol) was slowly added dropwise, after which time DIEA (1.23mL, 7.4655mmol) was reacted at-5 ℃ for 2 hours, and then the reaction was left to stir at room temperature overnight. After the reaction is finished, adding n-hexane (50mL) and methyl tert-butyl ether (400mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, performing suction filtration to obtain a solid product, and drying to obtain a product 5.9g which is overweight.

M-7

M-6(3.32g, 1.6590mmol) is placed in a 500mL round bottom flask, and is dissolved by dichloromethane (50mL), then TFA (2.35mL, 16.590mmol) is added to be stirred at room temperature for overnight reaction, after the reaction is completed, the reaction liquid is concentrated under reduced pressure, n-hexane (50mL) and methyl tert-butyl ether (400mL) are added for sedimentation, the supernatant is poured, n-hexane and methyl tert-butyl ether are added for sedimentation, the reaction is repeated for 3 times, a solid product is obtained by suction filtration, the obtained solid product is dissolved by dichloromethane and methanol, dry sampling and column chromatography are carried out, elution is carried out by using 7% methanol/1% ammonia water/dichloromethane, collection, concentration and drying are carried out, and 2.5g of the product is obtained, and the yield is 79.36.

27-212

M-7(2.5g, 1.3244mmol), 27-204(0.43g, 0.3010mmol), HBTU (0.69g, 1.806mmol) and HOBT (0.24g, 1.806mmol) were placed in a 500mL round-bottomed flask, then DMF (100mL) was added and dissolved, the reaction was stirred at-5 ℃ for 30 minutes, then DIEA (0.90mL, 5.418mmol) was slowly added dropwise, after which time it was reacted at-5 ℃ for 2 hours, and then the reaction was left to stir at room temperature overnight. After the reaction is finished, adding n-hexane (50mL) and methyl tert-butyl ether (400mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, performing suction filtration to obtain a solid product, and drying to obtain a product 4.5g which is overweight.

27-234

27-212(2.69g, 0.3010mmol) was placed in a 500mL round bottom flask, dissolved in dichloromethane (50mL), followed by addition of TFA (0.22mL, 3.010mmol) and reaction with stirring at room temperature overnight, after completion of the reaction, the reaction solution was concentrated under reduced pressure, n-hexane (50mL) and methyl t-butyl ether (400mL) were added and settled, the supernatant was poured, n-hexane and methyl t-butyl ether were added and settled, repeating 3 times, suction filtration to obtain a solid product, the solid product was dissolved in dichloromethane and methanol, dry-loading, column chromatography, elution with 8% methanol/dichloromethane, collection, concentration and drying to obtain 1.9g of the product, yield 71.76%.

¹H-NMR(600MHz,DMSO-d₆)δ10.17-10.13(m,8H),8.99-8.91(m,8H),8.34–8.14(m,16H),8.14–7.94(m,24H),7.88-7.67(m,30H),7.45–7.05(m,49H),5.83-5.77(m,10H),5.35-5.28(m,5H),4.56–3.98(m,51H),3.74–3.49(m,60H),3.15–2.99(m,48H),2.81–2.66(m,19H),2.41–2.03(m,68H),1.89-1.85(m,50H),1.57-1.51(m,43H),1.34-1.23(m,18H),0.98–0.77(m,72H).

27-214

Paclitaxel (abbreviated as PTX, 0.5g, 0.5855mmol), TBDMS-Cl (from Innochem, 0.53g, 3.5310mmol), imidazole (0.2g, 2.9425mmol) were placed in a 500mL round bottom flask, then anhydrous DMF (20mL) was added and dissolved, and then the reaction was left to stir at room temperature overnight. After completion of the reaction, methylene chloride (200mL) was added to dilute the reaction solution, and saturated NH was added₄Cl (50mL), then deionized water (50mL), then saturated sodium chloride solution (100mL), and the organic phase with anhydrous sulfuric acidSodium drying, dry loading, column chromatography with petroleum ether: ethyl acetate ═ 1: 1, collected, concentrated and dried to obtain 0.35g of product with a yield of 62.5%, wherein the 2' hydroxyl position of paclitaxel is protected by TBDMS.

27-235

Fmoc-Gly-OH (0.44g, 1.4476mmol), EDCI (0.28g, 1.4476mmol), DMAP (0.02g, 0.1810mmol) were placed in a 500mL round-bottom flask, which was then dissolved by addition of anhydrous dichloromethane (50mL) and anhydrous DMF (25mL), and the reaction was stirred at room temperature for 15 min. Further 27-214(0.35g, 0.3619mmol) was added and the reaction mixture was reacted at room temperature overnight. After completion of the reaction, methylene chloride (200mL) was added to dilute the reaction solution, and saturated NH was added₄Washing with Cl solution (100mL), then washing with deionized water (50mL), finally washing with saturated sodium chloride solution (100mL), drying the organic phase with anhydrous sodium sulfate, dry loading, column chromatography, eluting with 3% methanol/dichloromethane, collecting, concentrating, and drying to obtain 0.31g of product with 68.89% yield, wherein the esterification position is hydroxyl at position 7.

¹H-NMR(400MHz,DMSO-d₆)δ8.83(m,1H),7.99(m,2H),7.92–7.81(m,4H),7.76–7.59(m,6H),7.57–7.27(m,12H),7.21(m,1H),5.99(s,1H),5.86(m,1H),5.56–5.40(m,3H),5.01(m,1H),4.80(s,2H),4.26(s,3H),3.66(m,3H),2.45(s,3H),1.99(s,6H),1.65(s,5H),1.55(s,3H),1.00(m,6H),0.80(s,9H),0.05(m,6H).

27-239

27-235(0.31g, 0.2487mmol) was placed in a 500mL round bottom flask, piperidine (0.25mL, 2.487mmol) and DMF (5mL) were added, and the reaction was allowed to stir at room temperature for 2 h. After the reaction is finished, the reaction solution is decompressed and concentrated under a vacuum pump, and piperidine is removed to obtain a reaction product DMF solution.

27-240

27-234(1.9g, 0.2135mmol), 27-239(0.2487mmol), HBTU (0.12g, 0.3203mmol) and HOBT (0.04g, 0.3203mmol) were placed in a 500mL round-bottomed flask, then DMF (30mL) was added and dissolved, the reaction was stirred at-5 ℃ for 30 minutes, then DIEA (0.17mL, 0.9608mmol) was slowly added dropwise, after which time DIEA (0.17mL, 0.9608mmol) was reacted at-5 ℃ for 2 hours, and then the reaction was left to stir at room temperature overnight. After the reaction is finished, adding n-hexane (50mL) and methyl tert-butyl ether (400mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, performing suction filtration to obtain a solid product, adding a small amount of mixed solvent of methanol and dichloromethane for dissolving, adding n-hexane (50mL) and methyl tert-butyl ether (400mL) for settling, performing suction filtration, dissolving in the way, and settling for 3 times to obtain 1.6g of a product with the yield of 75.82%.

27-243

27-240(1.6g, 0.1615mmol) was placed in a 500mL round bottom flask, DMF (10mL) was added and sonicated to dissolve completely, then morpholine (0.14mL, 1.615mmol) was added and the reaction was left to stir at room temperature for 2 hours. After the reaction is finished, adding n-hexane (25mL) and methyl tert-butyl ether (200mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, performing suction filtration to obtain a solid product, dissolving the obtained solid product with dichloromethane and methanol, performing dry sampling, performing column chromatography, eluting with 7% methanol/dichloromethane, collecting, concentrating and drying to obtain 1.2g of a product, wherein the yield is 78.43%.

27-247

27-243(1.2g, 0.1268mmol) was placed in a 250mL flask, dissolved in DMF (20mL), M-SCM-5K (1.45g, 0.2790mmol, from Keka) was added and dissolved by ultrasonic oscillation, and the reaction was stirred at low speed for one week at room temperature in the absence of light. After the reaction is finished, adding n-hexane (25mL) and methyl tert-butyl ether (200mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, performing suction filtration to obtain a solid product, dissolving the obtained solid product with dichloromethane and methanol, performing dry sampling, performing column chromatography, eluting with 9% methanol/dichloromethane, collecting, concentrating and drying to obtain 1.9g of a product, wherein the yield is 75.70%.

¹H-NMR(600MHz,DMSO-d₆)δ10.16-10.12(m,8H),9.02-8.98(m,11H),8.08-7.98(m,55H),7.72-7.64(m,14H),7.56–7.09(m,49H),7.05-6.98(m,2H),6.69-6.65(m,3H),5.80-5.37(m,21H),5.14–4.75(m,20H),4.56-4.30(m,35H),4.03-4.38(m,45H),3.51(s,1120H),3.44–3.04(m,65H),2.77-2.68(m,13H),2.41–2.04(m,55H),2.02–1.69(m,39H),1.68–1.26(m,66H),1.18–1.12(m,5H),1.06–0.69(m,45H),0.09-0.06(m,3H).

27-253

27-247(1.9g, 0.0958mmol), TBAF (0.06g, 0.1916 mmol) were placed in a 500mL round bottom flask, then THF (20mL) was added to dissolve it, and the reaction was left to stir at room temperature for 1.5 h. After the reaction is finished, carrying out reduced pressure concentration on the reaction liquid to obtain a solid product, adding DMF (5mL) for dissolving, adding isopropanol for settling, repeating for three times, and filtering to obtain a solid product; and adding dichloromethane (10mL) for dissolution, adding methyl tert-butyl ether for sedimentation, and repeating for 3 times to obtain a solid product. Drying to obtain 0.75g of product with the yield of 62.5%.

¹H-NMR(400MHz,DMSO-d₆)δ10.16-10.10(m,8H),9.02-8.97(m,11H),8.55–7.81(m,58H),7.70-7.62(m,9H),7.21-6.99(m,53H),6.71-6.66(m,5H),5.88-5.84(m,10H),5.37-4.96(m,10H),4.59-4.28(m,27H),4.14–3.81(m,32H),3.57-3.48(s,1101H),3.19-3.15(m,61H),2.81-2.76(m,7H),2.44-2.27(s,68H),1.82–1.43(m,89H),1.36-1.32(m,36H),0.90–0.79(m,40H).

18. Synthesis of 49-5 (Compound No. 11)

The route is as follows

27-216

27-202(3g, 1.6826mmol) was placed in a 500mL round bottom flask, dissolved in dichloromethane (50mL), followed by the addition of TFA (1.25mL, 16.826mmol) and stirred overnight at room temperature, after completion of the reaction, the reaction was concentrated under reduced pressure, extracted with saturated sodium chloride solution (200mL) and ethyl acetate (200mL), the organic phase was collected, the aqueous phase was extracted with ethyl acetate (200 mL. times.3), the combined organic phases were collected, concentrated under reduced pressure and dried to give 4.3g, extra heavy product.

27-218

27-216(2.90g, 1.6826mmol), Gly-OtBu (0.24g, 1.8509mmol, ex Ilykok), HBTU (0.96g, 2.5239mmol) and HOBT (0.34g, 2.5239mmol) were placed in a 500mL round bottom flask, then DMF (100mL) was added to dissolve it, the reaction was stirred at-5 ℃ for 30 minutes, then DIEA (1.25mL, 7.5717mmol) was slowly added dropwise, after which time DIEA (1.25mL, 7.5717mmol) was reacted at-5 ℃ for 2 hours, and then the reaction was left to stir at room temperature overnight. After the reaction, adding saturated sodium chloride solution (200mL) and ethyl acetate (200mL) for extraction, collecting an organic phase, extracting an aqueous phase with ethyl acetate (200mL multiplied by 3), collecting and combining the organic phases, concentrating under reduced pressure, and drying in vacuum to obtain 6.6g of the product which is overweight.

27-221

27-218(2g, 1.0867mmol) was placed in a 500mL round bottom flask, dissolved in dichloromethane (50mL), followed by the addition of TFA (0.81mL, 10.867mmol) and stirred overnight at room temperature, after completion of the reaction, the reaction was concentrated under reduced pressure, extracted with saturated sodium chloride solution (200mL) and ethyl acetate (200mL), the organic phase was collected, the aqueous phase was extracted with ethyl acetate (200 mL. times.3), the combined organic phases were collected, concentrated under reduced pressure and dried to give 4.3g, extra weight, of the product.

27-224

27-221(1.94g, 1.0867mmol) was placed in a 500mL round bottom flask, dissolved by sonication with dichloromethane (20mL), and Boc-NHNH was weighed₂(0.16g, 1.1954mmol), DCC (0.67g, 3.2601mmol) and DMAP (0.026g, 0.2173mmol) were added to the flask and the reaction was allowed to stir at room temperature overnight. After the reaction was completed, saturated brine (200mL) and ethyl acetate (200mL) were added for extraction, the organic phase was collected, the aqueous phase was extracted with ethyl acetate (200mL × 3), the organic phases were combined, concentrated under reduced pressure, dry-loaded, column-chromatographed, eluted with 3% methanol/dichloromethane, collected, concentrated, and dried to obtain 1.62g of a product with a yield of 78.64%.

27-259

27-224(1.62g, 0.8535mmol) and 10% Pd/C catalyst (100mg) were added to a hydrogenation reaction apparatus, then DMF (30mL) was added to dissolve it, the hydrogenation reaction apparatus was closed, hydrogen gas was introduced under a pressure of 18psi, the mixture was stirred at room temperature overnight, after completion of the reaction was observed by TLC, the reaction solution was taken out and uniformly added dropwise to a sand core funnel containing compacted diatomaceous earth, suction filtration was carried out, and the diatomaceous earth was washed with DMF (30 mL. times.3) to obtain a reaction product solution.

27-205

K-11(0.69g, 1.4272mmol), 14-128(3g, 3.1398mmol), HBTU (1.62g, 4.2816mmol) and HOBT (0.58g, 4.2816mmol) were placed in a 500mL round-bottomed flask, then DMF (50mL) was added and dissolved, the reaction was stirred at-5 ℃ for 30 minutes, then DIEA (2.12mL, 12.8448mmol) was slowly added dropwise, after which time DIEA (2.12mL, 12.8448mmol) was reacted at-5 ℃ for 2 hours, and then the reaction was left to stir at room temperature overnight. After the reaction is finished, adding n-hexane (50mL) and methyl tert-butyl ether (400mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, performing suction filtration to obtain a solid product, drying to obtain 5.1g of the product, and weighing.

27-231

Placing 27-205(3.03g, 1.4272mmol) in a 500mL round bottom flask, dissolving with dichloromethane (50mL), adding TFA (1.06mL, 14.272mmol) and stirring at room temperature for reaction overnight, after the reaction is completed, concentrating the reaction solution under reduced pressure, adding n-hexane (50mL) and methyl tert-butyl ether (400mL) for sedimentation, pouring the supernatant, adding n-hexane and methyl tert-butyl ether for sedimentation, repeating for 3 times, performing suction filtration to obtain a solid product, dissolving the obtained solid product with dichloromethane and methanol, performing dry loading, performing column chromatography, eluting with 8% methanol/1% ammonia water/dichloromethane, collecting, concentrating and drying to obtain 2g of a product with the yield of 69.20%.

27-260

27-259(0.35g, 0.2248mmol), 27-231(2g, 0.9891 mmol), HBTU (0.51g, 1.3488mmol) and HOBT (0.18g, 1.3488mmol) were placed in a 500mL round-bottomed flask, then DMF (100mL) was added and dissolved, the reaction was stirred at-5 ℃ for 30 minutes, then DIEA (0.67mL, 4.0464mmol) was slowly added dropwise, after which time DIEA (0.67mL, 4.0464mmol) was reacted at-5 ℃ for 2 hours, and then the reaction was left to stir at room temperature overnight. After the reaction is finished, adding n-hexane (50mL) and methyl tert-butyl ether (400mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, dissolving the obtained solid product with dichloromethane and methanol, carrying out dry loading, carrying out column chromatography, eluting with 6% methanol/dichloromethane, collecting, concentrating and drying to obtain 1.3g of a product, wherein the yield is 60.46%.

27-265

27-260(1.3g, 0.1361mmol) was placed in a 500mL round bottom flask, DMF (10mL) was added and sonicated to dissolve completely, then morpholine (0.12mL, 1.361mmol) was added and the reaction was left to stir at room temperature for 2 hours. After the reaction is finished, adding n-hexane (25mL) and methyl tert-butyl ether (200mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, performing suction filtration to obtain a solid product, dissolving the obtained solid product with dichloromethane and methanol, performing dry sampling, performing column chromatography, eluting with 6% methanol/dichloromethane, collecting, concentrating and drying to obtain 0.91g of a product with the yield of 73.39%.

49-1

49-265(0.91g, 0.0999mmol) was placed in a 250mL flask, dissolved in DMF (20mL), M-SCM-5K (1.14g, 0.2198mmol, from KEY) was added and dissolved by ultrasonic oscillation, and the reaction was stirred at low speed for one week at room temperature in the dark. After the reaction is finished, adding n-hexane (25mL) and methyl tert-butyl ether (200mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, performing suction filtration to obtain a solid product, dissolving the obtained solid product with dichloromethane and methanol, performing dry loading, performing column chromatography, eluting with 9% methanol/dichloromethane, collecting, concentrating and drying to obtain 1.23g of a product, wherein the yield is 63.07%.

¹H-NMR(600MHz,DMSO-d₆)δ9.85-9.79(m,6H),8.77-8.71(m,7H),8.55–7.91(m,63H),7.89–7.63(m,23H),7.47–6.98(m,93H),6.69-6.54(s,7H),5.27-5.21(m,14H),4.72-4.57(m,20H),4.41–4.12(m,32H),3.85-3.79(m,43H),3.55-3.48(m,1115H),3.27–3.12(m,19H),3.09–2.95(m,32H),2.90–2.60(m,32H),2.14–1.70(m,31H),1.54–1.14(m,95H),0.86-0.79(m,51H).

49-3

49-1(1.23g, 0.0632mmol) was placed in a 500mL round bottom flask, dissolved in dichloromethane (20mL), followed by addition of TFA (0.094mL, 1.264mmol) and reaction with stirring at room temperature overnight, after completion of the reaction, n-hexane (25mL) and methyl tert-butyl ether (200mL) were added and the mixture was allowed to settle, the supernatant was poured, n-hexane and methyl tert-butyl ether were added and allowed to settle 3 times, suction filtration was performed to obtain a solid product, the solid product was dissolved in dichloromethane and methanol, dry-loaded, column chromatography was performed, elution was performed with 9% methanol/dichloromethane, collected, concentrated and dried to obtain 0.98g of product in 80.33% yield.

49-5

Placing 49-3(0.98g, 0.0506mmol) in a dry 500mL round bottom flask, adding anhydrous methanol (10mL) for dissolution, then adding TFA (0.018mL, 0.253mmol), adriamycin (DOX for short, 0.028g, 0.0506mmol) for stirring at room temperature for reaction overnight, after the reaction is completed, adding n-hexane (25mL) and methyl tert-butyl ether (200mL) for sedimentation, pouring the supernatant, adding n-hexane and methyl tert-butyl ether for sedimentation, repeating for 3 times, performing suction filtration to obtain a solid product, dissolving the obtained solid product with dichloromethane and methanol, performing dry loading, performing column chromatography, eluting with 9% methanol/dichloromethane, collecting, concentrating, drying, adding anhydrous alcohol (5mL) and dichloromethane (1mL) for dissolution, adding methyl tert-butyl ether (40mL) for sedimentation, and repeating for three times. Filtration and drying gave 0.65g of the final product in 65% yield, where the hydrazone bond formation was the 13-carbonyl group of doxorubicin.

¹H-NMR(600MHz,DMSO-d₆)δ10.28(s,5H),8.98-8.94(m,8H),8.58-8.51(m,8H),8.34-7.92(m,79H),7.49-7.45(m,10H),7.46-7.10(m,82H),6.68(s,5H),6.54(s,3H),5.26(s,20H),4.80-4.71(m,16H),4.56(s,10H),4.41-4.06(m,42H),3.95-3.74(m,45H),3.52-3.50m,1289H),3.11-2.97(m,57H),2.78-2.74(m,26H),2.11(s,26H),1.60-1.52(m,40H),1.00-0.61(m,72H).

19. 39-42 Synthesis (Compound No. 19)

The route is as follows

39-40

GFLG-LPT (1.0g, 1.04mmol, synthesized by 14-128 methods), 39-51(0.08mmol), HBTU (0.54g, 1.44mmol) and HOBT (0.19g, 1.44mmol) were weighed into a 250mL reaction flask, DMF (30mL) was added and the reaction was stirred for 30 minutes at-5 ℃ with ultrasound, DIEA (0.71mL, 4.32mmol) was slowly added dropwise and stirred at low temperature for 2 hours, and then the reaction was allowed to proceed to completion at room temperature. After the reaction, methyl tert-butyl ether (250mL) was added to the reaction mixture, the mixture was sonicated for 5 minutes, the upper layer was poured, ethyl acetate (20mL) was added to the lower layer, the mixture was sonicated for 3 minutes, methyl tert-butyl ether (150mL) and n-hexane (100mL) were added to precipitate a solid, the mixture was filtered, dissolved in 20% methanol/dichloromethane (20mL), silica gel powder (10g) was added, evaporated to dryness, and subjected to column chromatography. Elution with a gradient of 3% methanol/dichloromethane to 10% methanol/dichloromethane afforded 0.2g of product.

39-41

After methylene chloride (15mL) and TFA (0.033mL, 0.444mmol) were added to 39-40(0.2g) and dissolved completely by sonication, the reaction mixture was stirred at room temperature with a ground glass stopper. After the reaction is finished, the reaction solution is evaporated to dryness, dichloromethane is removed, ethyl acetate (20mL) is added, ultrasonic treatment is carried out for 2 minutes, methyl tert-butyl ether (150mL) and n-hexane (70mL) are added for sedimentation, solid is separated out, suction filtration is carried out, and vacuum drying is carried out to obtain 0.2g of a product.

39-42

39-41(0.6g) was weighed out and dissolved in DMF (20mL), followed by addition of M-SCM-5K (0.9378g) and ultrasonic dissolution. The reaction was carried out at low speed in the absence of light. After the reaction, methyl tert-butyl ether (150mL) and n-hexane (70mL) were added to the reaction mixture to precipitate a solid, which was then filtered off, dissolved in 20% methanol/dichloromethane, added with silica gel powder (10g), evaporated to dryness, and subjected to column chromatography. With 1% ammonia: 4% methanol dichloromethane gradient-1% ammonia: gradient elution is carried out on 8% methanol/dichloromethane, the collected product is dried by distillation, absolute ethyl alcohol (3mL) is added, ultrasonic treatment is carried out uniformly, methyl tert-butyl ether (150mL) and n-hexane (50mL) are added, suction filtration is carried out, dissolution of absolute ethyl alcohol (3mL) is continued to be added, the methyl tert-butyl ether and the n-hexane are settled, repeated dissolution and settlement are carried out for three times, suction filtration and drying are carried out, and 0.15g of the product is obtained.¹H-NMR(400MHz,DMSO-d₆)δ10.06–9.84(m,3H),8.84–8.36(m,4H),8.29–7.65(m,85H),7.57–6.79(m,187H),6.73–6.51(m,15H),4.83–4.07(m,86H),3.50–3.49(m,512H),3.08–2.99(m,81H),1.70–1.17(m,125H),0.90–0.80(m,72H).

20. Synthesis of 41-32 (Compound No. 20)

The route is as follows

41-29

The reaction products GFLG-LPT (1.0g, 1.48mmol, synthesized by the method of 14-128), 41-33(0.077mmol), HBTU (0.35g, 0.924mmol) and HOBT (0.1g, 0.924mmol) were taken into a 250mL reaction flask, DMF (30mL) was added to dissolve the reaction product, the mixture was stirred for 30 minutes at-5 ℃, DIEA (0.45mL, 2.772mmol) was slowly added dropwise, and the reaction was terminated at low temperature. After the reaction, methyl tert-butyl ether (100mL) was added, and n-hexane (200mL) was added to the mixture to precipitate the mixture as a powder. Column chromatography, dry loading, loading with 1% ammonia water: 2% methanol/dichloromethane-1% ammonia: 10% methanol in dichloromethane. The weight of the product is 0.6 g.

41-31

41-29(0.6g) was weighed out, dichloromethane (5mL) was added, TFA (0.17mL, 2.31mmol) was added, the mixture was dissolved by sonication, and the reaction was stirred. After the reaction, the reaction mixture was concentrated, and methyl t-butyl ether (100mL) was added to the concentrated solution, followed by addition of n-hexane (200mL) to the concentrated solution and precipitation to obtain a powder. Vacuum drying to obtain 0.6g of product.

41-32

M-SCM-5K (1.6g, 0.30mmol) was added to reactants 41-31(0.6g, 0.044mmol), and DMF solution (20mL) was added to dissolve, and the mixture was stirred at low speed and protected from light. At the end of the reaction, methyl tert-butyl ether (50mL) was added and n-hexane (100mL) was allowed to settle. And (5) suction filtration. Column chromatography, dry loading, eluting with dichloromethane-1% ammonia: elution with a gradient of 8% methanol in dichloromethane. The product is evaporated to dryness, absolute ethyl alcohol (10mL) is added, ultrasonic treatment is carried out to form a homogeneous phase, and n-hexane (50mL) is added for sedimentation. The sedimentation was repeated three times. Vacuum drying to obtain 0.4 g of product.

¹H-NMR(600MHz,DMSO-d₆)δ9.91–9.83(m,8H),8.80–8.72(m,10H),8.58–8.50(m,10H),8.27–7.96(m,82H),7.63–7.49(m,128H),7.24–7.14(m,51H),6.71–6.50(m,7H),5.29–5.21(m,22H),4.52–4.40(m,82H),4.04–3.98(m,54H),3.51–3.50(m,600H),3.17–3.14(m,83H),3.03–2.97(m,122H),2.88–2.86(m,15H),0.91–0.70(m,72H).

21. Synthesis of 33-207 (Compound No. 21)

The route is as follows

33-194

Weighing GFLG-PCB (synthesized by a method of 30-33, 2g, 2.43mmol), HBTU (1.15g, 3.03mmol) and HOBT (0.41g, 3.03mmol), putting into a 250mL reaction bottle, adding 33-190 DMF solution for dissolving, performing ultrasonic treatment to completely dissolve reactants, stirring for 30 minutes at-5 ℃, slowly dropwise adding DIEA (1.5mL, 9.108mmol), and reacting at low temperature until the reaction is finished. After the reaction is finished, adding methyl tert-butyl ether (100mL) and n-hexane (150mL) into the reaction solution, carrying out ultrasonic treatment for 5 minutes, placing the mixture into a refrigerator, standing the mixture for 20 minutes, pouring out the supernatant, adding ethyl acetate (20mL) into the lower layer, carrying out ultrasonic treatment for 2 minutes, adding the n-hexane (100mL), carrying out suction filtration, and carrying out vacuum drying to obtain 1.4g of a product.

33-196

33-194(1.4g, 0.32mmol) was placed in a 250mL round bottom flask, dissolved in DMF (70mL), and morpholine (0.84mL, 9.6mmol) was added and the reaction stirred at room temperature. After the reaction is finished, methyl tert-butyl ether (150mL) and n-hexane (100mL) are added into the reaction solution, ultrasonic treatment is carried out for 5 minutes, solid is separated out, suction filtration is carried out, and vacuum drying is carried out to obtain 1.3g of a product.

33-199

33-196(1.3g, 0.316mmol), HBTU (0.153g, 0.406mmol), and HOBT (0.0541g, 0.406mmol) were weighed out and placed in a 250mL reaction flask, and 45-12 DMF solution (19mL, 0.0452mmol) was added, DMSO (80mL) was added thereto and the reaction was dissolved completely by sonication, and stirred at-5 ℃ for 30 minutes, DIEA (0.2mL, 1.22mmol) was slowly added dropwise, and after 1 hour, the reaction was stirred at room temperature. After the reaction, deionized water (100mL) was added to the reaction mixture to precipitate a solid, which was then filtered with suction. Vacuum drying to obtain 1.3g of product.

33-204

After adding dichloromethane (10mL) and TFA (0.7mL, 9.48mmol) to 33-199(1.3g, 0.316mmol), the mixture was dissolved completely by sonication, and the reaction mixture was stirred at room temperature with a ground glass stopper. After the reaction, methyl tert-butyl ether (150mL) and n-hexane (100mL) were added directly to the reaction mixture, which was filtered and dried under vacuum to give 1.2 g.

33-207

Weighing reactants 33-199(1.20g, 0.048mmoL) and M-NH₂HCl-5K (2.0g, 0.38mmoL), HBTU (0.21g, 0.576mmoL), and HOBT (0.077g, 0.576mmoL) were put into a 250mL reaction flask, and DMF (75mL) was added thereto for ultrasonic dissolution, followed by stirring at-5 ℃ for 30 minutes, followed by slow dropwise addition of DIEA (0).0073mL, 0.0446mmoL), stirring for 1 hr, and reacting at room temperature under low speed and in dark. After the reaction is finished, adding methyl tert-butyl ether (200mL) into the reaction solution for sedimentation, performing suction filtration to obtain a powder product, dissolving the powder product by using a mixed solvent of 20% methanol/dichloromethane, adding silica gel (6g), drying by distillation, performing dry-method sampling, and performing column chromatography. With 6% methanol/dichloromethane-1% ammonia: gradient elution is carried out on 10% methanol/dichloromethane, the product is collected and evaporated to dryness, dichloromethane (5mL) is added, ultrasonic dissolution is carried out, methyl tert-butyl ether (150mL) is added, n-hexane (50mL) is added, suction filtration is carried out, dichloromethane is continuously added for dissolution, methyl tert-butyl ether and n-hexane are settled, repeated dissolution and settlement are carried out for three times, and 0.5g of the product is obtained.

¹H-NMR(400MHz,DMSO-d₆)δ10.58–10.39(m,14H),9.05–8.89(m,13H),8.23–7.66(m,169H),7.26–6.91(m,165H),3.51–3.50(m,1957H),3.26–3.22(m,148H),2.97–2.83(m,232H),2.34–2.28(m,86H),1.94–1.45(m,282H),0.94–0.75(m,144H).

22. Synthesis of 29-160 (Compound No. 22)

The route is as follows

29-150

The reaction products 30-33(3.5g, 4.2581mmol), 29-149 (synthesized by W3, 0.3275mmol), HBTU (2.2359g, 5.8958mmol) and HOBT (0.7967g, 5.8958mmol) were taken and placed in a 250mL reaction flask, DMF (50mL) was added to dissolve the reaction products, the reaction flask was placed at-5 ℃ and stirred for 30 minutes, DIEA (2.9mL, 17.6873mmol) was slowly added dropwise, the reaction mixture was stirred at-5 ℃ for 1 hour, and then the reaction mixture was stirred at room temperature overnight. After the reaction is finished, adding methyl tert-butyl ether (100mL), putting the mixture into a refrigerator for 30 minutes, taking out the mixture, separating out a solid, performing suction filtration, collecting a filter cake, and drying to obtain a crude product.

29-154

Weighing 29-150(0.3275mmol), adding dichloromethane (10mL), adding TFA (0.3648mL), dissolving with ultrasound, and stirring the reaction. After the reaction, the reaction solution was evaporated to dryness to an oil, and after methyl t-butyl ether (100mL) was added, a solid precipitated, and the solid was filtered off, and the filter cake was collected by suction filtration, then dissolved in a methanol/dichloromethane (1:4) solution (100mL), and silica gel powder (15g) was added and evaporated to a powder. Column chromatography, dry loading, loading with 1% ammonia water: 4% methanol/dichloromethane-1% ammonia: 10% methanol in dichloromethane. The weight of the product was 1.0g, and the yield was 30%.

29-160

M-SCM-10K (0.75g, 0.07mmol) was added to the reaction mixture 29-154(0.4g), and DMF solution (20mL) was added to dissolve the mixture, followed by stirring at low speed and protecting from light for one week. After the reaction, methyl tert-butyl ether (100mL) was added, the mixture was placed in a refrigerator for 30 minutes, taken out, a solid precipitated, filtered, collected by suction, and a filter cake was dissolved in a methanol/dichloromethane (1:4) solution (100mL), and then silica gel powder (15g) was added and evaporated to dryness to a powder. Column chromatography, dry loading, loading with 1% ammonia water: 3% methanol/dichloromethane-1% ammonia: elution with a gradient of 8% methanol in dichloromethane. The product was evaporated to dryness, taken up in absolute ethanol (8mL) and a small amount of dichloromethane and then precipitated by addition of methyl tert-butyl ether (50 mL). The sedimentation was repeated three times. Vacuum drying to obtain 0.6g product. The yield was 55%.

¹H-NMR(600MHz,DMSO-d₆)δ10.18-10.09(m,8H),9.12-8.90(m,17H),8.25–7.82(m,72H),7.503-7.44(m,13H),7.19-7.15(m,41H),7.05–6.97(m,11H),6.90-6.81(m,7H),6.69-6.64(m,5H),5.36-5.26(m,9H),4.59–4.32(m,27H),4.05-3.84(m,31H),3.52-3.48(m,1925H),2.32-2.85(m,28H),2.76-2.69(m,39H),2.43-2.38(m,23H),2.33-2.22(m,50H),1.99–1.74(m,85H),1.60-1.45(m,59H),1.37–1.32(m,48H),1.25-1.18(m,40H),0.89-0.78(m,72H).

23. Synthesis of 35-98 (Compound No. 23)

37-88

Diglycolamine (25.84mL,260.59mmol, from TCI) was placed in a 1L flask, dichloromethane (50mL) and triethylamine (72.64mL,521.18mmol) were added, the mixture was stirred at 0 ℃ for 1 hour, tert-butyl dicarbonate was dissolved in dichloromethane, slowly added dropwise to the flask, and after the dropwise addition, the stirring was continued at 0 ℃ overnight. After the reaction is finished, concentrating the reaction liquid to a small amount, adding silica gel powder (50g), evaporating to dryness to obtain a powdery solid, carrying out dry-method sample loading, carrying out column chromatography, and eluting with a petroleum ether mixed solution of 50% -60% ethyl acetate to obtain a product 37-88:53.4861 g.

37-148

37-88(53.4861g,260.59mmol) is placed in a 1L flask, stirred for 30 minutes at 0 ℃, a tetrahydrofuran solution of potassium tert-butoxide (286.64mL,1mol/L,286.64mmol) is slowly added dropwise, stirring is continued for 1 hour after the addition is completed, ethyl bromoacetate (28.82mL,260.59mmol) is slowly added dropwise, stirring is continued for 30 minutes at 0 ℃, and then the flask is moved to room temperature and stirred overnight. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding deionized water (200mL) and ethyl acetate (200mL) for extraction to obtain an organic phase, washing a water phase with ethyl acetate (200mL multiplied by 1), combining the organic phases, concentrating and evaporating to dryness, adding methanol (30mL) and dichloromethane (120mL) for dissolution, adding silica gel powder (100g), evaporating to dryness to obtain a powdery solid, performing dry-method sample loading, performing column chromatography, and eluting with a mixed solution of 20% ethyl acetate and petroleum ether to obtain a product 37-148:52.1g, wherein the yield is 68.6%.

37-151

37-148(52.1g,178.8mmol) was placed in a 1L flask, 1, 4-dioxane (100mL) was added, lithium hydroxide (9.4g,393.4mmol) was further added, the reaction was stirred at room temperature for 30 minutes, pure water (200mL) was further added, and stirring was continued for 2 hours. After the reaction is finished, transferring the reaction solution to a 1L separating funnel, adding n-hexane (125mL) and methyl tert-butyl ether (125mL) for extraction, adjusting the pH of a water phase to 2 by using concentrated hydrochloric acid, then adding ethyl acetate (200mL multiplied by 2), combining organic phases, adding silica gel powder, evaporating to dryness, performing dry-method sampling, performing column chromatography, and eluting by using a 50-60% ethyl acetate petroleum ether mixed solution to obtain 37-151: 45.1g, yield: 96 percent.

35-82

Boc-Glu-OH (20.0g, 80.89mmol, ex Ark Pharm), HBTU (92.02g, 242.66mmol), HOBT (32.8g, 242.66mmol) and H-Glu (OBn)₂TsOH (84.861g, 161.8mmol, ex Ark Pharm) was charged to a 1000mL flask, DMF (200mL) was added to dissolve, the reaction was allowed to stir at-5 ℃ for about 30 minutes, DIEA (120.32mL, 728mmol) was added slowly dropwise, after which the reaction was allowed to stir at-5 ℃ for 1 hour and finally transferred to room temperature to stir overnight. After the reaction, the reaction mixture was transferred to a 2L separatory funnel, and a saturated sodium bicarbonate solution (600mL) and ethyl acetate (300mL) were added to extract the mixture, the aqueous phase was washed with ethyl acetate (200 mL. times.1), the organic phases were combined, the organic phase was concentrated with a saturated saline solution (200 mL. times.2) and evaporated to dryness to obtain 35-82:70g of a product.

35-84

35-82(70g, 80.89mmol) was placed in a 1000mL round bottom flask, dichloromethane (50mL) was added to dissolve it, trifluoroacetic acid (300mL, 4044.5mmol) was added with stirring, and the reaction was left to stir at room temperature overnight. After the reaction, the reaction solution was concentrated, ethyl acetate (300mL) and saturated sodium bicarbonate solution (300mL) were added to generate a large amount of bubbles, sodium bicarbonate solid was added continuously until pH was greater than 7, followed by extraction, the aqueous phase was washed with ethyl acetate (200mL × 1), the organic phases were combined and evaporated to dryness to obtain 35-84: 62 g.

35-85

37-151(19.36g, 73.5364mmol), HBTU (41.83g, 110.3045mmol), HOBT (14.91g, 110.3045mmol) and 35-84(61.95g, 80.89mmol) were added to a 1000mL round bottom flask, after dissolution in DMF (200mL), the reaction was stirred at-5 ℃ for about 30 minutes, DIEA (54.69mL, 330.9136mmol) was slowly added dropwise, after which the reaction was allowed to continue stirring at-5 ℃ for 1 hour and then allowed to stir at room temperature overnight. After the reaction is finished, transferring the reaction solution to a 2L separating funnel, adding a saturated sodium bicarbonate solution (500mL) and ethyl acetate (300mL), shaking, extracting to obtain an organic phase, washing a water phase with ethyl acetate (200mL multiplied by 1), combining the organic phases, concentrating to a small amount, adding silica gel powder, evaporating to dryness, loading the sample by a dry method, carrying out column chromatography, eluting with a petroleum ether mixed solution of 50% -70% ethyl acetate to obtain 35-85: 51g, yield: and 69 percent.

35-86

35-85(23.4g, 23.14mmol) was charged into a 500mL round-bottom flask, dichloromethane (30mL) was added to dissolve it, trifluoroacetic acid (85.93mL, 1157.13mmol) was added with stirring, and finally the reaction was left to stir at room temperature overnight. After the reaction, the reaction solution was concentrated, ethyl acetate (300mL) and saturated sodium bicarbonate solution (300mL) were added to generate a large amount of bubbles, sodium bicarbonate solid was added continuously until PH was greater than 7, followed by extraction, the aqueous phase was washed with ethyl acetate (200mL × 1), the organic phases were combined and evaporated to dryness to obtain 35-86: 18.2g, yield: 86 percent.

35-88

35-86(18.2g, 19.98mmol), HBTU (11.36g, 29.97mmol), HOBT (4.05g, 29.97mmol) and Boc-Lys (Fmoc) -OH (8.5g, 18.16mmol, from Accela) were charged to a 500mL round bottom flask, dissolved in DMF (100mL), the reaction was stirred at 0 deg.C for about 30 minutes, DIEA (14.86mL, 89.90mmol) was slowly added dropwise, and after addition, the reaction was allowed to continue stirring at 0 deg.C overnight. After the reaction is finished, transferring the reaction solution to a 1L separating funnel, adding a saturated sodium bicarbonate solution (300mL) and ethyl acetate (200mL), shaking, extracting to obtain an organic phase, washing a water phase with ethyl acetate (200mL multiplied by 1), combining the organic phases, concentrating to a small amount, adding silica gel powder, evaporating to dryness, loading the sample by a dry method, carrying out column chromatography, eluting with a petroleum ether mixed solution of 80-100% ethyl acetate to obtain 35-88: 19.6g, yield: 79 percent.

35-89

35-88(7.0g,5.1413mmol) was placed in a 250mL flask, dichloromethane (20mL) was added and dissolved, trifluoroacetic acid (5.7270mL,77.1191mmol) was added with stirring, and the reaction was stirred at room temperature overnight. After the reaction, the reaction solution was concentrated, transferred to a 1L separatory funnel, added with saturated sodium bicarbonate solution (300mL) and ethyl acetate (200mL), shaken and extracted to obtain an organic phase, the aqueous phase was washed with ethyl acetate (150 mL. times.2), the organic phases were combined, washed with saturated saline (200 mL. times.2), concentrated and evaporated to dryness to obtain 35-89:6.4853g of a product.

35-90

35-89(4.2g, 3.33mmol), HBTU (1.72g, 4.54mmol), HOBT (0.61g, 4.54mmol) and mono-tert-butyl succinate (0.53g, 3.03mmol) were added to a 250mL flask, DMF (40mL) was added to dissolve, the reaction was stirred at 0 deg.C for about 30 minutes, DIEA (2.25mL, 13.62mmol) was slowly added dropwise, and after the addition was complete, the reaction was allowed to continue stirring at 0 deg.C overnight. After the reaction is finished, transferring the reaction solution to a 1L separating funnel, adding deionized water (300mL) and ethyl acetate (200mL), shaking and extracting to obtain an organic phase, washing a water phase with ethyl acetate (200mL multiplied by 1), combining the organic phases, concentrating to a small amount, adding silica gel powder, evaporating to dryness, loading by a dry method, carrying out column chromatography, eluting with a dichloromethane mixed solution of 3% methanol to obtain 35-90: 4.3 g.

¹H-NMR(600MHz,DMSO-d₆)δ8.62–8.53(m,1H),8.40–8.24(d,J＝7.4Hz,1H),7.95–7.81(m,1H),7.88–7.82(d,J＝7.5Hz,3H),7.77–7.64(m,3H),7.51–7.43(m,2H),7.38–7.29(m,22H),7.24–7.12(t,J＝5.6Hz,1H),5.15–5.01(m,9H),4.46–4.15(m,8H),3.92–3.82(m,2H),3.64–3.50(m,4H),3.38–3.27(m,2H),3.20-3.13(d,J＝5.0Hz,2H),3.01–2.89(m,3H),2.48–2.32(m,10H),2.18–2.10(t,J＝8.0Hz,2H),2.05–1.71(m,7H),1.56–1.46(d,J＝5.9Hz,1H),1.40–1.37(m,2H),1.36–1.21(m,10H).

35-93

35-90(0.622g, 0.4390mmol) was charged into a hydrogenation reactor, Pd/C (0.0300g) was added, DMF (30mL) was added and dissolved, hydrogen was introduced under a pressure of 1.8MPa, and the reaction was stirred at room temperature overnight. After completion of the reaction, the reaction mixture was filtered through celite, the celite was washed with DMF (20mL × 3), and the DMF solutions were combined to be used as a raw material for the next reaction.

37-53

Boc-Leu-OH.H₂O (40g,160.44mmol, ex ImmunoKay), Gly-OBn.TsOH (56.837g,168.462mmol, ex Ark pharm), HBTU (66.93g, 176.48mmol), HOBT (23.85g, 176.48mmol) were added to a 1000mL flask, DMF (250mL) was added to dissolve, the reaction was left to stir at-5 ℃ for about 20 minutes, DIEA (145.85mL,882.4356mmol) was then added slowly dropwise, stirring was continued at-5 ℃ for 1 hour after the addition was complete, and then left to stir at room temperature overnight. After the reaction is finished, transferring the reaction solution into a 2L separating funnel, adding a saturated sodium bicarbonate solution (250mL) and ethyl acetate (300mL) into the separating funnel, extracting to obtain an organic phase, washing a water phase with ethyl acetate (200mLx3), combining the organic phases, washing the organic phase with a saturated saline solution (200mL x 2), concentrating and evaporating to dryness, loading the organic phase by a dry method, performing column chromatography, eluting with a petroleum ether mixed solution of 30-40% EA to obtain a product 37-53: 60.7 g.

37-54

37-53(60.7g,160.44mmol) is put into a 1000mL flask, dichloromethane (40mL) and TFA (95mL, 1283.52.9mmol) are added, the reaction is stirred at room temperature overnight, the reaction is finished, the reaction solution is transferred into a 2L separating funnel, saturated sodium bicarbonate solution (350mL) and ethyl acetate (300mL) are added for extraction, an organic phase is obtained, an aqueous phase is washed by ethyl acetate (1500mL x 2), the organic phases are combined, and the concentrated and evaporated to dryness, so that 37-54:45g of the product is obtained.

37-56

37-54(45g,160.44mmol), Boc-Phe-OH (40.438g,152.42mmol, available from aladdin), HBTU (66.93g, 1276.48mmol), HOBT (23.85g, 176.48mmol) were added to a 1000mL flask, DMF (250mL) was added to dissolve, the reaction was stirred at-5 ℃ for about 20 minutes, DIEA (119.85mL,722mmol) was slowly added dropwise, stirring was continued at-5 ℃ for 1 hour after the addition was complete, and then the flask was allowed to stir at room temperature overnight. After the reaction, transferring the reaction solution to a 2L separating funnel, adding saturated sodium bicarbonate solution (350mL) and ethyl acetate (300mL), extracting to obtain an organic phase, washing the water phase with ethyl acetate (200mL x 3), combining the organic phases, washing the organic phase with saturated saline (250mL x 2), concentrating and evaporating to dryness to obtain a product 37-56: 84 g.

37-59

37-56(84g,160.44mmol) was placed in a 1000mL flask, dichloromethane (40mL) and TFA (95mL, 1283.52.9mmol) were added, the reaction was stirred overnight at room temperature, the reaction was terminated, the reaction mixture was transferred to a 2L Erlenmeyer flask, saturated sodium bicarbonate solution (350mL) was added to generate a large amount of bubbles, sodium bicarbonate solid was added slowly until pH >7, solid precipitated, filtered, and the filter cake was dried with pure water (100Mlx2) to obtain 37-59:68g of product.

37-62

37-59(68.27g,152.42mmol), Boc-Gly-OH (25.37g,144.799mmol, available from aladdin), HBTU (63.58g, 167.66mmol), HOBT (63.58g, 167.66mmol) were added to a 1000mL flask, DMF (250mL) was added to dissolve, the reaction was stirred at-5 ℃ for about 20 minutes, DIEA (113.3mL,685.89mmol) was slowly added dropwise, stirring was continued at-5 ℃ for 1 hour after the addition was complete, and then allowed to stir at room temperature overnight. After the reaction, the reaction mixture was transferred to a 2L separatory funnel, and saturated sodium bicarbonate solution (300mL) and ethyl acetate (350mL) were added to extract the mixture to obtain an organic phase, the aqueous phase was washed with (200 mL. times.3), the organic phases were combined, the organic phase was washed with saturated brine (250 mL. times.2), the organic phase was left at room temperature for 1.5 hours to precipitate a solid, which was then filtered, and the cake was washed with ethyl acetate: petroleum ether (3: 7) (150mLx5) to obtain 37-62: 72.8 g.

37-149

37-62(30g,51.4871mmol) was placed in a 250mL flask, dichloromethane (20mL) was added and dissolved, trifluoroacetic acid (30.6mL,411.9mmol) was added with stirring, and the reaction was stirred at room temperature overnight. After the reaction is finished, the reaction solution is concentrated, saturated sodium bicarbonate (200mL) is added to generate a large amount of bubbles, sodium bicarbonate solid is slowly added, the pH value is adjusted to be more than 7, solid is separated out, the filtration is carried out, a filter cake is washed by deionized water (150mL multiplied by 4), and the filter cake is dried to obtain a product 37-149:24.85 g.

37-152

Boc-Glu-OH (5.7864g,23.4032mmol, available from Ark pharm), 37-149(24.85g,51.4871mmol), HBTU (26.626g, 70.2096mmol), HOBT (9.4874g, 70.2096mmol) was added to a 500mL flask, dissolved in DMF (150mL), stirred at-5 ℃ for about 10 minutes, DIEA (34.8mL, 210.628mmol) was slowly added dropwise, and after addition, the reaction was stirred at-5 ℃ for 1 hour and then allowed to stir at room temperature overnight. After completion of the reaction, the reaction mixture was transferred to a 1L separatory funnel, and saturated sodium bicarbonate (200mL) and ethyl acetate (300mL) were added thereto, followed by shaking and extraction. Washing the water phase with ethyl acetate (150mL multiplied by 1), combining the organic phases, concentrating and evaporating to dryness, adding methanol (20mL) and dichloromethane (100mL) for dissolution, adding silica gel powder (50g), evaporating to dryness to form a powdery solid, loading by a dry method, carrying out column chromatography, eluting with a mixed solution of 4% -8% methanol and dichloromethane, and drying in a vacuum oven to obtain a product 37-152: 19.2g, yield 69.8%.

37-154

37-152(19.2g,16.3215mmol) was charged into a 1000mL flask, dichloromethane (30mL), trifluoroacetic acid (9.6966mL,130.5722mmol) was added, and the reaction was stirred at room temperature overnight. After the reaction, concentrating the reaction solution to a small amount, adding methyl tert-butyl ether (200mL), precipitating a solid, filtering, washing a filter cake with methyl tert-butyl ether (100mL x 3), and drying the filter cake to obtain a product 37-154: 17.5 g.

37-156

Boc-Gly-OH (3.4310g,19.5858mmol, available from Ark Pharm), 37-154(17.5g,16.3215mmol), HBTU (9.2847g, 24.4823mmol), HOBT (3.3083g, 24.4823mmol) was added to a 1000mL flask, dissolved in DMF (150mL), stirred at-5 ℃ for about 10 minutes, DIEA (12.14mL, 73.4468mmol) was slowly added dropwise, and after addition, the reaction was stirred at-5 ℃ for 1 hour and then allowed to stir at room temperature overnight. After the reaction, adding ethyl acetate (200mL), methyl tert-butyl ether (200mL) and n-hexane (300mL), precipitating a solid, performing suction filtration, washing a filter cake with methyl tert-butyl ether (100mL x 3), and drying the filter cake to obtain a product 37-156: 19.2 g.

37-158

37-156(10.499g, 8.5125mmol) was charged into a hydrogenation reactor, 10% Pd/C (0.10g) was added thereto, DMF (50mL) was added thereto and dissolved therein, hydrogen gas was introduced thereinto under a pressure of 1.8MPa, and the reaction was stirred at room temperature overnight. After completion of the reaction, the reaction mixture was filtered through celite, the celite was washed with DMF (20mL × 3), and the DMF solutions were combined to be used as a raw material for the next reaction.

37-159

37-158(8.965g,0.4390mmol), Palbociclib (8g,17.8763mmol, the Chinese name of Palbociclib, abbreviated as PCB), HBTU (9.6848g,25.5375mmol), HOBT (3.4509g,25.5375mmol) were added to a 500mL flask, DMF (130mL) was added to dissolve the mixture, the reaction was left at-5 ℃ and stirred for 20 minutes, DIEA (12.6626mL,76.6125mmol) was slowly added dropwise, after completion of the dropwise addition, the reaction was stirred at-5 ℃ for 1 hour, and then stirred at room temperature overnight. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (100mL) were added to precipitate a solid, which was filtered, and the filter cake was washed with methyl tert-butyl ether (100 mL. times.3) and dried to obtain 37-159:16.3g of a product.

37-161

Putting 37-159(16.3g,8.5125mmol) into a 500mL flask, adding dichloromethane (30mL) to dissolve, adding trifluoroacetic acid (18.964mL,255.375mmol), stirring overnight at room temperature after the reaction is finished, adding methyl tert-butyl ether (250mL), precipitating a solid, filtering, washing a filter cake with methyl tert-butyl ether (50mL multiplied by 3), dissolving the filter cake with dichloromethane (240mL) and methanol (60mL), adding silica gel powder (50g), evaporating to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 5% -7% methanol, collecting, concentrating and drying to obtain a product 37-161:11.2g, wherein the yield is 73%.

35-94

35-93(0.464g,0.4390mmol), 37-161(3.5g,1.9315mmol), HBTU (0.9988g,2.6338mmol) and HOBT (0.3559g,2.6338mmol) were charged into a 250mL flask, DMF (50mL) was added to dissolve the mixture, the reaction was left at-5 ℃ and stirred for 20 minutes, DIEA (1.306mL,7.9015mmol) was slowly added dropwise, the reaction was stirred at-5 ℃ for 40 minutes after completion of the addition, and then stirred at room temperature overnight. After the reaction, n-hexane (100mL) and methyl tert-butyl ether (60mL) were added, and the mixture was settled for 2 times to give a viscous oil, and methyl tert-butyl ether (100mL) was added to separate out a solid, followed by filtration, washing of the filter cake with methyl tert-butyl ether (50 mL. times.2), and drying of the filter cake to obtain 35-94:3.614g of a product.

35-95

Putting 35-94(3.614g,0.4390mmol) into a 250mL flask, adding DMF (25mL) to dissolve, adding morpholine (1.7mL,19.7507mmol), stirring at room temperature for 1 hour, finishing the reaction, adding methyl tert-butyl ether (150mL), precipitating a solid, filtering, washing a filter cake with methyl tert-butyl ether (50mL multiplied by 3), dissolving the filter cake with dichloromethane (200mL) and methanol (50mL), adding silica gel powder (20g), evaporating to obtain a powdery solid, performing dry sampling, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 5% -9% methanol, collecting, concentrating and drying to obtain a product 35-95:0.7g with the yield of 20%.

37-172

45-57(0.0412g, 0.0565mmol) was charged into a hydrogenation reactor, 10% Pd/C (0.10g) was added thereto, DMF (50mL) was added thereto and dissolved therein, hydrogen was introduced thereinto under a pressure of 1.8MPa, and the reaction was stirred at room temperature overnight. After completion of the reaction, the reaction mixture was filtered through celite, the celite was washed with DMF (20mL × 3), and the DMF solutions were combined to be used as a raw material for the next reaction.

35-96

37-172(0.007g,0.0199mmol), 35-95(0.7g,0.0874mmol), HBTU (0.45g,1.192mmol) and HOBT (0.16g,1.192mmol) were added to a 250mL flask, DMF (35mL) was added to dissolve the mixture, the reaction was allowed to stand at-5 ℃ and stirred for 20 minutes, DIEA (0.59mL,3.575mmol) was slowly added dropwise, after the dropwise addition, the reaction was stirred at-5 ℃ for 20 minutes and then stirred at room temperature overnight. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (60mL) were added, and the mixture was settled for 1 time to give a viscous oil, and methyl tert-butyl ether (100mL) was added to precipitate a solid, followed by filtration, washing of the filter cake with methyl tert-butyl ether (50 mL. times.2) and drying of the filter cake to obtain 35-96:0.6434g of a product.

35-97

35-96(0.6436g,0.0199mmol) was charged in a 250mL flask, dichloromethane (25mL) and trifluoroacetic acid (2.073mL,27.92mmol) were added, and the reaction was stirred at room temperature overnight. After the reaction is finished, concentrating the reaction solution to a small amount, adding methyl tert-butyl ether (150mL) to precipitate a solid, performing suction filtration, cleaning a filter cake with methyl tert-butyl ether (50mL x 3), dissolving with a methanol (30mL) dichloromethane (120mL), adding silica gel powder (10g), evaporating to dryness to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with a dichloromethane mixed solution of 1% ammonia water and 5% -8% methanol, collecting and concentrating, and evaporating to dryness to obtain a solid, thus obtaining a product 35-97: 0.23g, yield 36%.

35-98

35-97(0.23g,0.0072mmol) was put into 250mL of the flask, DMF (10mL) was added thereto and dissolved therein, and M-NH was added thereto₂HCl-10K (0.452g,0.043mmol from KeyKa), HBTU (0.163g,0.43mmol), HOBT (0.058g,0.43mmol), the reaction was stirred at-5 ℃ for about 15 min at low speed, DIEA (0.1226mL,0.7415mmol) was then added slowly dropwise, and after 20 min at-5 ℃ the reaction was allowed to proceed and then transferred to room temperature with low stirring for 7 days in the dark. After the reaction was complete, n-hexane (150mL) and methyl tert-butyl were addedButyl ether (40mL), pouring the supernatant, adding n-hexane (150mL) and methyl tert-butyl ether (40mL) into the lower layer of liquid, repeating the steps for 2 times to obtain a viscous oily substance, adding methyl tert-butyl ether (100mL) to obtain a viscous substance, dissolving the viscous oily substance by using a methanol (30mL) and dichloromethane (120mL), adding silica gel powder (15g), evaporating to obtain a powdery solid, loading the powdery solid by a dry method, performing column chromatography, eluting by using a dichloromethane mixed solution of 1% ammonia water and 8% -11% methanol, collecting and concentrating, evaporating to obtain a solid, drying in a vacuum oven, adding anhydrous ethanol (10mL) and dichloromethane (15mL) to dissolve, adding methyl tert-butyl ether (200mL), separating out the solid, filtering, washing a filter cake by using methyl tert-butyl ether (50mL x 3), and drying the filter cake by using the vacuum oven to obtain a product 35-: 0.3g, yield 57%.

¹H-NMR(600MHz,DMSO-d₆)δ9.24–9.10(m,5H),8.53–8.13(m,134H),7.96-7.82(m,85H),7.86–7.64(m,68H),7.59-7.42(m,68H),7.41–7.34(m,71H),7.30–7.21(m,69H),7.17–7.04(m,28H),6.99–6.73(m,5H),6.68–6.53(m,5H),4.45–4.23(m,8H),4.14–4.01(m,21H),3.79–3.54(m,304H),3.51–3.42(m,3763H),3.14–3.03(m,205H),2.98–2.73(m,298H),2.79–2.70(m,270H),2.62–2.51(m,17H),2.40–2.35(m,26H),2.31–2.24(m,22H),2.12–1.90(m,21H),1.84–1.62(m,17H),1.44–1.29(m,221H),1.25–1.16(m,173H).

24. Synthesis of 44-172 (Compound No. 24)

25-241

Boc-Gly-OH (3.0g, 17.1252mmol, available from Ark Pharm), HBTU (9.7418g, 25.6878mmol), HOBT (3.4709g, 25.6878mmol) and H-Glu (OBzl) -OBzl. TosOH (8.5554g, 17.1252mmol, available from Ark Pharm) were added to a 500mL round bottom flask, which was dissolved in DMF (50mL), the reaction flask was stirred at-5 deg.C for about 30 minutes, then DIEA (12.7mL, 77.0634mmol) was slowly added dropwise, after which the reaction flask was stirred at-5 deg.C for 1 hour and finally transferred to room temperature for 2 hours. After completion of the reaction, the reaction solution was transferred to a 2L separatory funnel, and a saturated sodium bicarbonate solution (400mL) and ethyl acetate (300mL) were added thereto, shaken, extracted, and the aqueous phase and the organic phase were separated. Saturated sodium chloride (300mL) was added to the organic phase, shaken, extracted and the aqueous and organic phases separated. Saturated sodium chloride (300mL) was then added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated and evaporated to dryness. Dissolving the organic phase with 20% methanol/dichloromethane (50mL), adding silica gel powder (50mL), evaporating, dry-loading, and performing column chromatography. Eluting with eluent (1% -2% methanol: 99% -98% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 25-241: 6.9213g, yield: 83.41 percent.

42-105

25-241 g (2.0g, 4.1276mmol) and 10% Pd/C (100mg) were put into a hydrogenation reactor, DMF (30mL) was added to dissolve the mixture, the hydrogenation reactor was closed, hydrogen was introduced to make the pressure in the reactor 2.0MPa, and the hydrogenation reactor was left at room temperature and stirred overnight. And after the reaction is finished, taking out the reaction kettle, uniformly dropwise adding the reaction solution into a sand core funnel filled with compacted diatomite, performing suction filtration, and cleaning the diatomite with DMF until the diatomite does not contain the product to obtain a reaction product solution.

43-116

Boc-GFLG-OBn (9.9g,17mmol, synthesized by the method 37-62) was added to the hydrogenation reactor, 10% Pd/C (0.025g) was added, DMF (40mL) was added to dissolve it, hydrogen was introduced under a hydrogen pressure of 2.1MPa, and the reaction was stirred at room temperature overnight. After completion of the reaction, the reaction mixture was filtered through Celite, which was washed 3 times with DMF (20mL x 3), and the filtrate was charged into a 500mL round-bottomed flask as the starting material for the next reaction.

42-89

A solution of 43-116 DMF (8.4534g, 17.1618mmol), Niraparib (4.5821g, 14.3015mmol, NPB for short), HBTU (8.1356g, 21.4523mmol) and HOBT (2.8986g, 21.4523mmol) were added to a 250mL round bottom flask, dissolved in DMF (60mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (10.6mL, 64.3568mmol) was slowly added dropwise, and after the addition, the reaction flask was allowed to continue to stir at-5 ℃ for 1 hour and then transferred to room temperature for overnight stirring. After the reaction, the reaction solution was transferred to a 1L separatory funnel, and a saturated aqueous sodium chloride solution (300mL) and ethyl acetate (200mL) were added thereto, shaken, extracted, and the aqueous phase was separated; ethyl acetate (200mL) was added to the aqueous phase, which was shaken, extracted and the aqueous phase separated. The organic phases were combined, and then saturated aqueous sodium chloride (300mL) was added to the organic phase, followed by shaking, extraction and separation of the aqueous phase. Then, deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous phase was separated. Finally, the organic phase is concentrated and evaporated to dryness and then put into an oven for drying. To products 42-89: 11.3695 g.

42-90

42-89(11.3695g, 14.3015mmol) was placed in a 250mL round-bottom flask, dichloromethane (30mL) was added and dissolved, TFA (15.9mL, 214.5225mmol) was added with stirring, and the reaction flask was left to stir at room temperature overnight. After the reaction, the reaction mixture was concentrated and evaporated to dryness to remove methylene chloride. To the reaction solution was added methyl tert-butyl ether (150mL) and the mixture was settled to give a powdery solid, which was then filtered and the filter cake was washed with methyl tert-butyl ether (60 mL). The filter cake was dissolved in a 20% methanol/dichloromethane mixed solvent (60mL), silica gel powder (65mL) was added, evaporated to dryness, and subjected to dry loading and column chromatography. Eluting with eluent (0-2% methanol: 100-98% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-90: 8.6g, yield: 86.6 percent.

42-107

42-105 solution (1.2560g, 4.1276mmol), 42-90(6.3095g, 9.0807mmol), HBTU (4.6961g, 12.3828mmol) and HOBT (1.6732g, 12.3828mmol) were added to a 500mL round bottom flask, which, after dissolution in DMF (60mL), was stirred at-5 deg.C for about 30 minutes, DIEA (6.1mL, 37.1484mmol) was slowly added dropwise, after which the reaction flask was stirred at-5 deg.C for 1 hour and finally the flask was transferred to room temperature and stirred overnight. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and settled, the supernatant was poured off, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and settled, and this was repeated three times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid, which was filtered to give the solid product, which was washed with methyl tert-butyl ether (60mL) and finally dried in an oven. Obtaining products 42-107: 6.8432 g.

42-110

42-107(6.4832g, 4.1276mmol) was placed in a 250mL round bottom flask, dichloromethane (15mL) was added and dissolved, TFA (4.6mL, 61.9140mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. To the reaction solution was added methyl tert-butyl ether (150mL) and the mixture was settled to give a solid as a powder, which was filtered and the solid product was washed with methyl tert-butyl ether (100 mL). Dissolving the solid product with 20% methanol/dichloromethane mixed solvent (100mL), adding silica gel powder (70mL), evaporating to dryness, dry loading, and performing column chromatography. Eluting with eluent (0.5% ammonia water: 2% -10% methanol: 97.5% -89.5% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-110: 2.9609g, yield: 46.05 percent.

44-156

Fmoc-Lys (Boc) -OH (3.6002g, 7.6839mmol, available from Accela), 35-86(9.1g, 9.9890mmol), HBTU (4.3710g, 11.5258mmol) and HOBT (1.5574g, 11.5258mmol) were added to a 500mL round bottom flask, which, after dissolution with DMF (80mL), was stirred for about 30 minutes at-5 deg.C, DIEA (7.6mL, 46.4032mmol) was slowly added dropwise, after which the reaction flask was stirred for 1 hour at-5 deg.C and finally transferred to room temperature for 3 hours. After the reaction, the reaction solution was transferred to a 1L separatory funnel, and a saturated aqueous sodium chloride solution (300mL) and ethyl acetate (200mL) were added, shaken, extracted, and the aqueous phase and the organic phase were separated; ethyl acetate (100mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. Then, the organic phases were combined, and a saturated aqueous solution of sodium chloride (300mL) was further added to the organic phase, followed by shaking, extraction, and separation of the aqueous and organic phases. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated, evaporated to dryness and dried. Obtaining products 44-148: 10.4619 g.

44-157

44-156(10.4619g, 7.6839mmol) was charged into a 500mL round-bottom flask, DMF (10mL) was added and dissolved, morpholine (10.0mL, 115.2583mmol) was added with stirring, and the reaction flask was left to stir at room temperature for 2 hours. After the reaction was completed, n-hexane (150mL) and methyl t-butyl ether (30mL) were added to the reaction solution to precipitate, the supernatant was poured, and n-hexane (150mL) and methyl t-butyl ether (30mL) were added to the lower oily substance to precipitate, and the reaction was repeated five times to obtain a viscous solid. Dissolving the viscous solid with 20% methanol/dichloromethane mixed solvent (60mL), adding silica gel powder (50mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (1% -5% methanol: 99% -95% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 44-157: 8.7542g, yield: 100 percent.

45-91

Glycerol (5g,54.295mmol from Innochem) was weighed into a 500mL reaction flask, nitrogen was introduced, a THF solution of potassium tert-butoxide (211mL, 211.7505mmol) was added, the reaction was stirred at 0 ℃ for 3 hours, benzyl bromoacetate (30.964g,195.46mmol) was added, and after stirring for 2 hours, the reaction was allowed to cool to room temperature. And (3) after the reaction is finished, evaporating the reaction solution to dryness, adding deionized water and ethyl acetate, separating an organic phase, extracting a water phase by using ethyl acetate until no product exists, combining the organic phase, drying by using anhydrous sodium sulfate powder, carrying out suction filtration, carrying out dry-method sample loading on the filtrate, and carrying out column chromatography. The product 9g is obtained by gradient elution with 1 percent to 2 percent of ethyl acetate/petroleum ether, and the yield is 31 percent.

36-186

Adding 45-91(0.5g,0.9318mmol) into a reaction kettle, adding 10% Pd/C (0.0200g), adding DMF (30mL) to dissolve, pumping off the air in the reaction kettle by using a water pump to form vacuum, introducing hydrogen at the hydrogen pressure of 0.16MPa, then discharging the hydrogen, pumping to vacuum by using the water pump, introducing the hydrogen again, repeating the steps for 3 times, finally introducing the hydrogen again, and stirring the reaction at room temperature overnight. After the reaction, the reaction mixture was filtered through celite, the filter cake was washed with DMF (20 mL. times.3), and the DMF solution was combined and used as the starting material for the next step.

44-161

The 36-186 solution (0.5021g, 1.8862mmol), 44-157(8.5957g, 7.5448mmol), HBTU (3.2190g, 8.4879mmol) and HOBT (1.1469g, 8.4879mmol) were added to a 250mL round bottom flask, which was dissolved with DMF (80mL), the reaction flask was stirred at-5 deg.C for about 30 min, DIEA (4.2mL, 25.4637mmol) was slowly added dropwise, and after the addition, the reaction flask was stirred at-5 deg.C for 1 h and then transferred to room temperature for 3 h. After the reaction, transferring the reaction solution to a 1L separating funnel, adding a saturated sodium bicarbonate aqueous solution (400mL) and ethyl acetate (300mL), shaking, extracting, and separating an aqueous phase and an organic phase; ethyl acetate (200mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases were separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated and evaporated to dryness. Dissolving the organic phase with 20% methanol/dichloromethane mixed solvent (60mL), adding silica gel powder (50mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (2% -8% methanol: 98% -92% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 44-161: 3.2551g, yield: 47.54 percent.

¹H–NMR(600MHz,DMSO-d₆)δ8.33–8.02(m,18H),7.50–7.17(m,60H),5.36–4.94(m,24H),4.55–4.50(m,12H),3.95–3.87(m,6H),3.67–3.47(m,25H),3.39–3.16(m,16H),2.47–2.38(m,24H),2.25–1.94(m,12H),1.94–1.79(m,6H),1.57–1.46(m,6H),1.38(s,27H),1.26–1.23(m,6H).

44-166

Putting 44-161(1.0g, 0.2755mmol, self made) and 10% Pd/C (50mg) into a hydrogenation reaction kettle, adding DMF (30mL) to dissolve, closing the hydrogenation reaction kettle, introducing hydrogen to make the pressure in the kettle reach 1.6MPa, and placing the hydrogenation reaction kettle at room temperature to stir overnight. And after the reaction is finished, taking out the reaction kettle, uniformly dropwise adding the reaction solution into a sand core funnel filled with compacted diatomite, performing suction filtration, and cleaning the diatomite with DMF until the diatomite does not contain the product to obtain a reaction product solution.

44-167

44-166 solution (0.3181g, 0.1248mmol), 42-110(2.8g, 1.7974mmol), HBTU (0.8519g, 2.2464mmol) and HOBT (0.3035g, 2.2464mmol) were added to a 250mL round bottom flask, which was dissolved with DMF (30mL), the reaction flask was stirred at-5 deg.C for about 30 minutes, DIEA (1.1mL, 6.7392mmol) was slowly added dropwise, after which the reaction flask was stirred at-5 deg.C for 1 hour, and finally the reaction flask was transferred to room temperature and stirred. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and settled, the supernatant was poured off, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and settled, and this was repeated three times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid which was filtered to give the solid product. The solid product was washed with methyl tert-butyl ether (60mL) and finally dried in an oven. Obtaining products 44-167: 2.4242 g.

44-168

44-167(1.7663g, 0.0840mmol) was placed in a 250mL round-bottom flask, dichloromethane (20mL) was added and dissolved, TFA (1.0mL, 12.6010mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. To the reaction solution was added methyl tert-butyl ether (150mL) and the mixture was settled to give a powdery solid, which was then filtered and the filter cake was washed with methyl tert-butyl ether (60 mL). The filter cake was dissolved in a 20% methanol/dichloromethane mixed solvent (100mL), silica gel powder (50mL) was added, evaporated to dryness, and subjected to dry loading and column chromatography. Eluting with eluent (1% ammonia water: 3% -10% methanol: 96% -89% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 44-168: 0.9538g, yield: 54.79 percent.

¹H–NMR(600MHz,DMSO-d₆)δ8.61–8.57(m,20H),8.46–7.80(m,196H),7.77–7.44(m,53H),7.44–7.04(m,166H),5.33–5.31(m,3H),4.67–4.18(m,77H),4.11–3.92(m,46H),3.86–3.84(m,40H),3.79–3.57(m,44H),3.19–3.04(m,45H),2.94–2.89(m,52H),2.88–2.71(m,102H),2.17–1.90(m,60H),1.61–1.51(m,96H),1.28–1.21(m,178H),0.96–0.78(m,144H).

44-172

44-168(0.8105g, 0.0420mmol) and M-SCM-10K (2.0019g, 0.1890mmol, from Kerkin) were added to a 500mL round bottom flask, which was dissolved in DMF (10mL), stirred at-5 deg.C for about 30 minutes, DIEA (0.7mL, 4.2mmol) was slowly added dropwise, and after completion of the addition, the reaction was stirred at-5 deg.C for 10 minutes and then allowed to shift to room temperature for one week. After completion of the reaction, n-hexane (150mL) and methyl t-butyl ether (20mL) were added to the reaction mixture and the mixture was settled, the supernatant was poured off, and n-hexane (150mL) and methyl t-butyl ether (30mL) were added to the lower oily substance and the settling was repeated five times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (60mL) as a powdery solid which was filtered to give the solid product. Dissolving the filter cake with 20% methanol/dichloromethane mixed solvent (100mL), adding 50mL silica gel powder, evaporating to dryness, loading by dry method, and performing column chromatography. Eluting with eluent (1% ammonia water: 2% -10% methanol: 97% -89% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 44-172: 0.6468g, yield: 29.53 percent.

¹H–NMR(600MHz,DMSO-d₆)δ8.61–8.57(m,20H),8.46–7.80(m,196H),7.77–7.44(m,53H),7.44–7.04(m,166H),5.33–5.31(m,3H),4.67–4.18(m,77H),4.11–3.92(m,46H),3.86–3.84(m,40H),3.79–3.57(m,44H),3.58–3.44(m,2853H),3.24–3.04(m,45H),2.94–2.89(m,52H),2.88–2.71(m,102H),2.17–1.90(m,60H),1.61–1.51(m,96H),1.28–1.21(m,178H),0.94–0.77(m,144H).

25. Synthesis of 41-126 (Compound No. 25)

39-81

N-Boc-L-glutamic acid-5-benzyl ester (10g, 29.6mmoL from alatin), H-Glu (Obzl) -Obzl. TsOH (16.2g, 32.6mmoL from Ark Pharm), HOBT (6g,44.4mmoL), HBTU (16.8g,44.4mmoL) were weighed out and put into a 250mL reaction flask, dissolved in DMF solution (80mL), and the reaction mixture was stirred for 30 minutes at-5 ℃ by sonication, followed by slow addition of DIEA (22mL, 133.4mmoL) thereto until the reaction was completed. After the reaction was completed, the reaction mixture was taken out, deionized water (100mL) was added thereto, and extracted with ethyl acetate several times (100 mL. times.3), and the organic phases were combined, washed twice with a saturated sodium chloride solution (100 mL. times.2), and finally concentrated to dryness. Loading the sample by dry method, performing column chromatography, eluting with 30% ethyl acetate/petroleum ether, collecting the product, concentrating, and evaporating to dryness to obtain the final product.

39-83

After methylene chloride (5mL) and TFA (22mL, 296mmol) were added to the reaction mixture (39-81 (29.6 mmol)), the mixture was dissolved completely by sonication, and the reaction mixture was stirred at room temperature with a ground glass stopper. After the reaction was completed, the reaction mixture was taken out, added with a saturated sodium bicarbonate solution (300mL), extracted with ethyl acetate several times (100 mL. times.3), the organic phases were combined, added with a saturated sodium chloride solution (100mL), washed twice, and finally concentrated to dryness.

41-115

Fmoc-L-Lys (Boc) -OH (5.1430g,10.9767mmol, available from Accela), 39-83(6g,10.9767mmol), HBTU (6.2442g,16.4651mmol), HOBT (1.8257g,16.4651mmol) was weighed into a 250mL reaction flask, dissolved in DMF (50mL), and stirred at-5 ℃ for 30 minutes. DIEA (8.2mL, 19.3954mmol) was slowly added dropwise and the reaction was allowed to proceed overnight at-5 ℃. After the reaction, the reaction was extracted with purified water and ethyl acetate, and the organic phase was concentrated to give 4.6g of a product.

41-116

The reaction product 41-115(4.6g,4.6132mmoL) was placed in a 250mL flask, DMF solution (30mL) was added, morpholine (8mL,92.2639mmoL) was added, and the reaction was stirred at room temperature for 3 hours to complete the reaction. Saturated brine (150mL) and ethyl acetate (200mL) were added, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (50 mL. times.3), the aqueous phase was freed of product, the organic phases were combined, washed 2 times with saturated brine (50 mL. times.2), and the organic phase was evaporated to dryness.

4g of product is obtained.

41-117

41-116(4g,5.1620mmol), 39-114(1.8g,5.1620mmol), HBTU (2.9364g,7.7429mmol) and HOBT (1.0462g,7.7429mmol) were weighed into a 250mL reaction flask, dissolved in DMF (40mL) and stirred at-5 ℃ for 30 minutes. DIEA (8.45mL, 50.9319mmol) was slowly added dropwise and the reaction was allowed to proceed overnight at-5 ℃. After the reaction is finished, adding petroleum ether for settling, collecting a lower layer, adding pure water, performing suction filtration, repeating for three times, collecting and transferring solids, and drying to obtain a product 5g with the yield of 87%.

41-118

The reaction mixture 41-117(5g,5.6303mmoL) was placed in a 250mL flask, DMF solution (30mL) was added, morpholine (9mL,103.24mmoL) was added, and the reaction was stirred at room temperature. After the reaction is finished, adding petroleum ether for settling, collecting a lower layer, adding pure water, performing suction filtration, collecting and transferring solids, and drying to obtain 2.6g of a product with the yield of 58%.

41-119

41-118(2.6g,2.9353mmol), HBTU (1.4312g,3.7740mmol) and HOBT (0.5099g,3.7740mmol) were weighed into a 250mL reaction flask, dissolved in 45-121 (i.e., 36-186, 0.8387mmol) DMF and stirred at-5 ℃ for 30 minutes. DIEA (2.1mL, 12.852mmol) was slowly added dropwise and the reaction was allowed to proceed overnight at-5 ℃. After the reaction, saturated brine (150mL) and ethyl acetate (200mL) were added, the organic phase was separated, the aqueous phase was extracted 3 times with ethyl acetate (50 mL. times.3), the aqueous phase was freed from the product, the organic phases were combined, washed 2 times with saturated brine (50 mL. times.2), and the organic phase was evaporated to dryness. And (4) carrying out dry loading and column chromatography. Eluting with 1% -7% methanol/dichloromethane to obtain 0.5g product with 55.6% yield.

41-120

41-119(0.1g,0.0348mmol) and 10% Pd/C (150mg) were weighed into a hydrogenation reactor, DMF (35mL) was slowly added and dissolved with stirring, hydrogen (300psi) was introduced, and the mixture was stirred at room temperature overnight. The next day, the reaction solution was filtered with a sand core funnel filled with diatomaceous earth, Pd/C was removed to obtain DMF solution of the product, which was directly used for the next reaction.

41-93

Fmoc-Glu-OH (1.6433g, 4.4485mmol, available from Aladdin), GFLG-NPB (synthesized by 42-90, 6.8g, 9.7867mmol), HBTU (5.0615g, 13.3455mmol), HOBT (1.8032g, 13.3455mmol) were added to a 250mL reaction flask, dissolved in DMF (50mL), stirred at-5 deg.C for 30 min, DIEA (6.6mL, 40.0365mmol) was added slowly and dropwise, and the reaction was continued for 3 h under this condition. After the reaction is finished, adding methyl tert-butyl ether (30mL) and n-hexane for settling (200mL), filtering by suction, transferring solid powder to a 500mL round-bottom flask, and evaporating to dryness to obtain 10.2g of a product with the yield of 100%.

41-95

41-93(7.6g,4.4485mmol) was dissolved in DMF (40mL), morpholine (7.8mL, 88.97mmol) was added, and the reaction was stirred at ambient temperature for 1 hour. After the reaction, ethyl acetate (100mL) and n-hexane were added for precipitation (200mL), the mixture was filtered with suction and the solid powder was transferred to a 500mL round-bottom flask, and silica gel powder was added, evaporated to dryness and subjected to column chromatography. Elution with 3% methanol/dichloromethane afforded 4.4g of product in 65.7% yield.

36-81

Adding 37-62(14.95g,25.654 mmol) into a reaction kettle, adding 10% Pd/C (0.300g), adding DMF (40mL) to dissolve, pumping off the air in the reaction kettle by a water pump to form vacuum, introducing hydrogen with the pressure of 0.16MPa, then discharging the hydrogen, pumping to vacuum by a water pump, introducing the hydrogen again, repeating the steps for 3 times, finally introducing the hydrogen again, stirring at room temperature overnight after the reaction is finished, filtering the reaction liquid by using kieselguhr, washing a filter cake by using DMF (20mL multiplied by 3), and combining DMF solution to be used as a raw material of the next step.

36-84

36-81(5.3g,10.7255mmol), Palbociclib (4g,8.9380mmol, abbreviated as PCB), HBTU (5g,13.4069mmol) and HOBT (1.8g,13.4069mmol) were put into 500mL, the reaction was stirred at-5 ℃ for about 20 minutes, DIEA (6.6mL,40.2208mmol) was slowly added dropwise, the reaction was continued at-5 ℃ for 1 hour, and then the reaction was transferred to room temperature and stirred overnight. And (3) after the reaction is finished, adding n-hexane (150mL) and methyl tert-butyl ether (30mL), settling, pouring the upper layer of liquid, continuously adding n-hexane (150mL) and methyl tert-butyl ether (50mL) into the lower layer of oily substance, repeating the operation for 3 times to obtain oily substance, adding methyl tert-butyl ether (200mL) to precipitate solid, performing suction filtration, and drying to obtain 36-84: 15.9 g.

36-98

36-84(15.9g,8.938mmol) was charged to a 500mL flask, dichloromethane (10mL) was added, TFA (6.6mL,89.38mmol) was added, and the reaction was stirred at room temperature overnight. After the reaction, the reaction mixture was concentrated to 10mL, methyl t-butyl ether (200mL) was added to precipitate a powder, which was filtered, the filter cake was washed with methyl t-butyl ether (50mL × 3), and then dissolved in a (20% methanol: 80% dichloromethane) solution (200mL), silica gel powder (60mL) was added to the solution, which was evaporated to dryness to give a solid powder, which was loaded by dry method, column chromatography was performed, and the mixture was purified by using 0.5% ammonia: eluting with 2% -5% methanol dichloromethane mixed solution, collecting, concentrating and evaporating to dryness to obtain a product 36-98: yield 8.5 g: 91.4 percent.

41-92

Fmoc-Glu (OtBu) -OH (3.3522g, 7.7879mmol, available from Ark Pharm), 36-98(6.8g, 8.2728mmol), HBTU (4.7061g, 12.4092mmol) and HOBT (1.6767g, 12.4092mmol) were taken and added to a 250mL reaction flask, dissolved in DMF (20mL), stirred at-5 ℃ for half an hour at a constant temperature, and DIEA (6.2mL, 37.2276mmol) was added dropwise slowly and stirred for 3 hours. After the reaction, methyl tert-butyl ether (30mL) and n-hexane are added for sedimentation (200mL), suction filtration is carried out, solid powder is transferred to a 500mL round-bottom flask, 20% methanol/dichloromethane is added for dissolution, silica gel powder is added, evaporation is carried out, and column chromatography is carried out. Elution with 2% methanol in dichloromethane afforded 8.9g of product in 100% yield.

41-97

41-92(8.9g,7.2447mmol) was taken, methylene chloride (20mL) was taken in and added to the flask followed by TFA (8mL, 108.6709 mmol). The reaction was stirred at room temperature for 3 hours, the reaction was complete, the reaction was evaporated to dryness, the dichloromethane was removed, methyl tert-butyl ether (30mL) and n-hexane were added and settled (200mL), suction filtered and the solid powder was transferred to a 500mL round bottom flask and evaporated to dryness to give 8.5g of product with 100% yield.

41-98

41-95(4.4g,2.9319mmol), 41-97(3.6121g,3.0785mmol), HBTU (1.6679g, 4.3979mmol) and HOBT (0.5492g, 4.3979mmol) are taken and added into a 250mL reaction flask, DMF (20mL) is added for dissolving, the reaction is stirred at the low temperature of 0 ℃ for 30 minutes at the constant temperature, DIEA (2.2mL, 13.1936mmol) is slowly added dropwise, and the reaction is stirred for 3 hours under the condition. After the reaction, methyl tert-butyl ether (30mL) and n-hexane were added and the mixture was settled (200mL), filtered with suction and the solid powder was transferred to a 2L round bottom flask and evaporated to dryness to obtain 7.7873g of product.

41-103

41-98(6.2g,2.3343) was dissolved in DMF (20mL), and morpholine (6mL, 70.0298mmol) was added thereto, and the reaction was stirred at room temperature for 1 hour. After the reaction, methyl tert-butyl ether (100mL) and n-hexane were added and the mixture was settled (200mL), filtered with suction and the solid powder was transferred to a 500mL round bottom flask and evaporated to dryness to obtain 5.68g of product.

41-104

41-103(5.5g,2.2598mmol), Boc-Gly-OH (0.4157g, 2.3728mmol, available from Aladdin), HBTU (1.2855g, 3.3897mmol) and HOBT (0.4580g, 3.3897mmol) were taken and added to a 250mL reaction flask, dissolved in DMF (20mL), stirred at-5 ℃ for 30 min, DIEA (1.7mL, 10.1692mmol) was slowly added dropwise, and the reaction was stirred for 3 h. After the reaction, ethyl acetate (30mL) and n-hexane were added to precipitate (200mL), suction filtration was repeated several times and the solid powder was transferred to a 2L round-bottom flask, evaporated to dryness and subjected to the next reaction.

41-105

41-104(5.8g,2.2598mmol) was taken and dichloromethane (20mL) was metered into the flask and TFA (5mL, 67.2110mL) was slowly added dropwise. Stirring and reacting for 3 hours under the condition of normal temperature, after the reaction is finished, removing dichloromethane by a rotary evaporator, adding 30mL of ethyl acetate) and n-hexane for sedimentation (200mL), repeatedly carrying out suction filtration for many times, transferring solid powder to a 500mL round-bottom flask, and evaporating to dryness to obtain 5.5g of a product with the yield of 98%.

41-122

41-105(0.8660g,0.3476mmol), HBTU (0.1975g,0.5215mmol) and HOBT (0.0705g,0.5215mmol) were weighed out and put into a 250mL reaction flask, and dissolved in a DMF solution of 41-120(0.0348mmol), and stirred at-5 ℃ for 30 minutes. DIEA (2mL, 12.1005mmol) was slowly added dropwise and the reaction was allowed to proceed overnight at 5 ℃. After the reaction is finished, pure water is added to precipitate solid, and the solid is filtered, filtered and dried to obtain 0.84g of product with the yield of 100 percent.

41-125

41-122(0.84g,0.0348mmol) was placed in a 250mL reaction flask, dichloromethane (30mL) was added and dissolved by sonication, and then TFA (0.3mL,0.2326mmol) was added and stirred at room temperature overnight. The next day, the reaction solution was evaporated to dryness, dichloromethane was removed, methyl tert-butyl ether was added for precipitation, and suction filtration was performed. Oven drying to obtain 0.8g product with yield of 100%.

41-126

41-125(0.8g,0.0348mmol) was weighed out and dissolved in DMF (20mL), and M-SCM-10K (1.1058g, 0.1044mmol, from KeKao) was added and dissolved by sonication. The reaction was stirred at low speed in the dark. After the reaction, methyl tert-butyl ether (150mL) and n-hexane (70mL) were added to the reaction mixture to precipitate a solid, which was then filtered off, dissolved in 20% methanol/dichloromethane, added with silica gel powder (3g), evaporated to dryness and subjected to column chromatography. The product 0.35g is obtained by gradient elution with 1% ammonia water and 5% -10% methanol/dichloromethane, and the yield is 19.4%.

¹H-NMR(600MHz,DMSO-d₆)δ10.13-10.12(m,7H),9.29-9.27(m,16H),8.95-8.92(m,7H),8.57-8.56(m,17H),8.18-8.16(m,37H),8.14–7.95(m,137H),7.89-7.87(m,50H),7.59-7.54(m,43H),7.35-7.25(m,176H),5.83-5.81(m,7H),5.16-5.08(m,6H),4.57-4.35(m,58H),4.13-4.10(m,25H),4.08-4.06(m,49H),3.84-3.82(m,26H),3.72-3.65(m,128H),3.51-3.49(m,2888H),3.24-3.19(m,30H),3.13-3.10(m,44H),3.06-3.02(m,40H),2.91-2.90(m,14H),2.78-2.74(m,35H),2.65-2.60(m,26H),2.41-2.30(m,47H),2.21-1.96(m,84H),1.83-1.75(m,91H),1.64-1.39(m,156H),1.20-1.16(m,24H),0.90-0.85(m,162H).

26. Synthesis of 49-136 (Compound No. 26)

49-96

Fmoc-Lys (Boc) -OH (3g, 6.4029mmol, available from Accela), HBTU (3.64g, 9.6044mmol), HOBT (1.30g, 9.6044mmol) were weighed into H-Glu (OBn)₂A flask (3.36g, 6.7230mmol from Ark Pharm) was charged with DMF and dissolved, DIEA (4.76mL, 28.8131mmol) was added dropwise slowly at-5 deg.C, and after dropping, the reaction mixture was taken out after half an hour and stirred at room temperature overnight. After the reaction is finished, adding a saturated NaCl solution (200mL) and EA (200mL), extracting, standing for layering, and collecting an organic phase; the aqueous phase was washed with EA (200mL × 3), the organic phases were combined, evaporated to dryness to a solid and dried in a vacuum oven to give the product (6.96g, high yield)

49-118

49-96(2g, 2.5710mmol) was added to the hydrogenation kettle, 10% Pd/C (0.05g) was added, DMF (20mL) was added, hydrogen was bubbled through at 300Psi, and the reaction stirred at room temperature overnight. After completion of the reaction, the reaction mixture was filtered through celite, the cake was washed 3 times with DMF (15 mL. times.3), and the filtrate was charged into a 250mL round-bottomed flask to obtain the product as the next starting material.

49-101

39-84(8g, 10.4459mmol), HBTU (5.94g, 15.6689mmol) and HOBT (2.12g, 15.6689mmol) were weighed into a flask containing 49-17(3.69g, 10.4459mmol), then an appropriate amount of DMF was added to dissolve the mixture, DIEA (7.77mL, 47.0066mmol) was slowly added dropwise at-5 ℃ and after completion of the dropwise addition, the mixture was taken out after half an hour of reaction and stirred at room temperature overnight. After the reaction is finished, adding a saturated NaCl solution (200mL) and EA (200mL), extracting, standing for layering, and collecting an organic phase; washing the water phase with EA (200mL × 3), combining the organic phases, evaporating to dryness to obtain a solid, dissolving with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (100mL), evaporating to obtain a powdery solid, performing dry loading, performing column chromatography, eluting with 100% dichloromethane, collecting and concentrating, and drying in a vacuum oven to obtain the product (9.6g, 83.48%)

49-104

DMF was added to a flask containing 49-101(9.6g, 8.7193mmol), and after complete dissolution by ultrasonic oscillation, morpholine (7.59mL, 87.193mmol) was added and the reaction was stirred at room temperature for 2 h. After the reaction is finished, adding a saturated NaCl solution (200mL) and EA (200mL), extracting, standing for layering, and collecting an organic phase; washing the water phase with EA (200mL × 3), combining the organic phases, evaporating to dryness to obtain a solid, dissolving with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (100mL), evaporating to obtain a powdery solid, performing dry-method sampling, performing column chromatography, eluting with 3% -5% methanol-dichloromethane mixed solution, collecting and concentrating, and drying in a vacuum oven to obtain a product (5.3g, 69.19%)

49-119

49-104(4.74g, 5.3991mmol), HBTU (2.92g, 7.7130mmol) and HOBT (1.04g, 7.7130mmol) were weighed into a flask containing 49-118(1.53g, 2.5710mmol), an appropriate amount of DMF was added to dissolve the mixture, DIEA (4.76mL, 28.8131mmol) was slowly added dropwise at-5 ℃ and after completion of the dropwise addition, the reaction was taken out after half an hour and stirred at room temperature overnight. After the reaction is finished, adding a saturated NaCl solution (200mL) and EA (200mL), extracting, standing for layering, and collecting an organic phase; the aqueous phase was washed with EA (200mL × 3), the organic phases were combined, evaporated to dryness to a solid and dried in a vacuum oven to give the product (8.65g, high yield)

49-123

DMF was added to a flask containing 49-119(5.96g, 2.5710mmol), and after complete dissolution by ultrasonic oscillation, morpholine (5.19mL, 25.710mmol) was added and the reaction stirred at room temperature for 2 h. After the reaction is finished, adding a saturated NaCl solution (200mL) and EA (200mL), extracting, standing for layering, and collecting an organic phase; the aqueous phase was washed with EA (200mL × 3), the organic phases were combined, evaporated to dryness to a solid and dried in a vacuum oven to give the product (7.6g, high yield)

49-124

Boc-Gly-OH (0.49g, 2.8281mmol, ex alatin), HBTU (1.46g, 3.8565mmol), HOBT (0.52g, 3.8565mmol) were weighed into a flask containing 49-123(5.39g, 2.5710mmol), an appropriate amount of DMF was added to dissolve, DIEA (4.76mL, 28.8131mmol) was slowly added dropwise at-5 deg.C, after completion of the addition, the reaction was taken out after half an hour and stirred at room temperature overnight. After the reaction is finished, adding a saturated NaCl solution (200mL) and EA (200mL), extracting, standing for layering, and collecting an organic phase; washing the water phase with EA (200mL × 3), combining the organic phases, evaporating to dryness to obtain a solid, dissolving with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (100mL), evaporating to obtain a powdery solid, performing dry-method sampling, performing column chromatography, eluting with 2% -5% methanol-dichloromethane mixed solution, collecting and concentrating, and drying in a vacuum oven to obtain a product (3.2g, 55.27%)

49-125

49-124(1g, 0.4437mmol) was added to the hydrogenation kettle, 10% Pd/C (0.05g) was added, DMF (20mL) was added, hydrogen was bubbled through at 300Psi, and the reaction stirred at room temperature overnight. After completion of the reaction, the reaction mixture was filtered through celite, the cake was washed 3 times with DMF (15 mL. times.3), and the filtrate was charged into a 250mL round-bottomed flask to obtain the product as the next starting material.

49-128

47-98(2.56g, 3.7271mmol), HBTU (2.02g, 5.3244mmol) and HOBT (0.72g, 5.3244mmol) were weighed into a flask containing 49-125(0.68g, 0.4437mmol), an appropriate amount of DMF was added to dissolve the mixture, DIEA (2.64mL, 15.9732mmol) was slowly added dropwise at-5 ℃, the reaction was stopped after half an hour, the mixture was taken out and stirred at room temperature overnight. After the reaction is finished, adding a saturated NaCl solution (200mL) and EA (200mL), extracting, standing for layering, and collecting an organic phase; the aqueous phase was washed with EA (200mL × 3), the organic phases were combined, evaporated to dryness to a solid and dried in a vacuum oven to give the product (4.3g, high yield)

49-134

Methylene chloride was added to a flask containing 49-128(3.06g, 0.4437mmol), and after complete dissolution by ultrasonic oscillation, TFA (2.27mL, 4.437mmol) was added, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating dichloromethane and most TFA, adding methyl tert-butyl ether (200mL) for sedimentation, pouring supernate, repeating for 3 times, performing suction filtration to obtain a solid product, dissolving the obtained solid product with a mixed solvent of dichloromethane and methanol, performing dry sampling, performing column chromatography, eluting with a mixed solution of 3-6% methanol and dichloromethane, collecting and concentrating, and drying in a vacuum oven to obtain a product (1.3g, 43.92%). ¹H NMR(400MHz,DMSO-d₆)δ10.39(s,4H),8.05-7.98(m,24H),7.81-7.36(m,99H),7.10-7.04(m,6H),5.39-5.24(m,14H),3.16-2.65(m,122H),1.37-1.16(m,62H),1.05-0.80(m,72H).

49-135

49-134(1.3g, 0.1943mmol) was put into a 250mL flask, dissolved with DMF (20mL), M-SCM-5K (2.27g, 0.4274mmol, from KeKai) was added, dissolved by ultrasonic oscillation, placed at-5 deg.C, DIEA (0.90mL, 5.4401mmol) was slowly added dropwise, after dropping, taken out after half an hour of reaction, and the reaction was left at room temperature under dark conditions and stirred at low speed for one week. After the reaction is finished, adding n-hexane (25mL) and methyl tert-butyl ether (200mL) for settling, pouring out supernate, adding n-hexane and methyl tert-butyl ether for settling, repeating for 3 times, performing suction filtration to obtain a solid product, dissolving the obtained solid product with a mixed solvent of dichloromethane and methanol, performing dry sampling, performing column chromatography, eluting with a mixed solution of 6-10% methanol and dichloromethane, collecting and concentrating, and drying in a vacuum oven to obtain a product (2.1g, 63.25%).

49-136

49-135(2.1g, 0.1229mmol), TBAF (0.51g,1.9671 mmol), THF (20mL) and dilute HCl (20mL) were placed in a 500mL round bottom flask, and the reaction was allowed to stir at room temperature overnight. After the reaction is finished, carrying out reduced pressure concentration on the reaction liquid to obtain a solid product, adding DMF (5mL) for dissolving, adding isopropanol for settling, repeating for three times, and filtering to obtain a solid product; and adding dichloromethane (10mL) for dissolution, adding methyl tert-butyl ether for sedimentation, and repeating for 3 times to obtain a solid product. Drying to obtain the product (1.2g, 64.17%).

¹H NMR(600MHz,DMSO-d₆)δ8.10-7.91(m,21H),7.81-7.24(m,25H),7.06-6.95(m,8H),5.36-5.10(m,22H),3.55-3.46(m,1056H),3.18-2.36(m,133H),1.37-1.16(m,62H).

27. Synthesis of 40-176 (Compound No. 27)

36-143

36-81(8.45g,17.1618mmol), Lapatinib (8.3g,14.3015mmol, LPT for short), HBTU (8.1g,21.4522mmol) and HOBT (2.9g,21.4522mmol) were put into 500mL, the reaction was stirred at-5 ℃ for about 20 minutes, DIEA (10.6mL,64.3567mmol) was slowly added dropwise, the reaction was continued at-5 ℃ for 1 hour, and then the reaction was transferred to room temperature and stirred overnight. After the reaction, the reaction solution was transferred to a 1L separatory funnel and saturated NaHCO was added₃Extracting the solution (200mL) with ethyl acetate (200mL) to obtain an organic phase, and cleaning the aqueous phase with ethyl acetate (200 mL. times.2)Washing, combining organic phases, washing the organic phases with saturated saline (200mL multiplied by 2), combining the organic phases, washing the organic phases with saturated sodium chloride solution (100mL multiplied by 2), evaporating to dryness to obtain 36-143: 14g, yield 92.8%.

36-145

36-143(14g,14.3015mmol) was charged to a 500mL flask, dichloromethane (20mL) was added, TFA (10.6mL,143.015mmol) was added, and the reaction was stirred at room temperature overnight. After the reaction, the reaction mixture was concentrated to 10mL, methyl t-butyl ether (200mL) was added to precipitate a powder, which was filtered, the filter cake was washed with methyl t-butyl ether (50mL × 3), and then dissolved in a (20% methanol: 80% dichloromethane) solution (200mL), silica gel powder (60mL) was added to the solution, which was evaporated to dryness to give a solid powder, which was loaded by dry method, column chromatography, purification with 1% ammonia: eluting with 5% -10% methanol dichloromethane mixed solution, collecting, concentrating and evaporating to dryness to obtain a product 36-145: 6.2g, yield 92.1%.

40-164

Fmoc-L-Lys (Boc) -OH (0.18g,0.3855mmol, available from Accela), 44-208(2g,0.3855mmol, from compound No. 29), HBTU (0.2193g,0.5783mmol), HOBT (0.0781g,0.5783mmol) were weighed into a 250mL reaction flask, dissolved in DMF (40mL), and stirred at-5 ℃ for 30 minutes. DIEA (0.2878mL, 1.7348mmol) was slowly added dropwise and the reaction was allowed to proceed overnight at-5 ℃. After the reaction is finished, adding methyl tert-butyl ether and n-hexane into the reaction solution for settling, performing suction filtration, adding 10% methanol/dichloromethane into the solid for dissolving, adding silica gel powder, evaporating to dryness, performing dry-method sample loading, and performing column chromatography. The product 1.8g is obtained by gradient elution with 1% ammonia water and 4% -8% methanol/dichloromethane, and the yield is 82.9%.

40-165

40-164(1.8g,0.3193mmol) was added to a 250mL round bottom flask, DMF (30mL) was added, morpholine (0.5563mL,6.3852mmol) was added, the mixture was sonicated to dissolve completely, and the reaction was stirred at ambient temperature. After the reaction is finished, methyl tert-butyl ether (150mL) and n-hexane (100mL) are added into the reaction solution, a solid product is separated out, and the product 1.7293g is obtained after suction filtration and drying, wherein the yield is 100%.

40-167

Fmoc-Gly-OH (0.0949g, 0.3193mmol, available from Accela), 40-165(1.7g, 0.3193mmol), HBTU (0.1817g, 0.4790mmol), HOBT (0.0647g, 0.4790mmol) were weighed into a 250mL round bottom flask, dissolved in DMF (50mL), the reaction was stirred at 0 ℃ for about 30 min, DIEA (0.2384mL, 1.4369mmol) was slowly added dropwise, and after addition, the reaction was allowed to continue stirring at 0 ℃ overnight. After the reaction is finished, adding methyl tert-butyl ether (150mL) and n-hexane (100mL) into the reaction solution, separating out a solid product, performing suction filtration, and drying to obtain 1.8g of a product with the yield of 100%.

40-169

40-167(1.8g,0.3193mmol) was charged into a 250mL round-bottomed flask, DMF (30mL) was added, morpholine (0.5563mL,6.3852mmol) was added, and the mixture was dissolved completely by sonication and the reaction was stirred at room temperature. After the reaction is finished, adding methyl tert-butyl ether (150mL) and n-hexane (100mL) into the reaction solution, separating out a solid product, carrying out suction filtration, adding 10% methanol/dichloromethane into the solid for dissolving, adding silica gel powder, evaporating to dryness, carrying out dry-method sample loading, and carrying out column chromatography. The product 1.5g is obtained by gradient elution with 1% ammonia water and 4% -8% methanol/dichloromethane, and the yield is 85.85%.

40-171

40-169(1.5g, 0.2741mmol) was weighed into a 250mL flask, dissolved in DMF (20mL), added DIEA (0.1895mL, 1.1420mmol), added 4ARM-SCM-10K (0.6222g, 0.0571mmol) and dissolved by sonication. The reaction was carried out at low speed in the absence of light. After the reaction, methyl tert-butyl ether (200mL) was added to the reaction mixture to precipitate a solid, which was then filtered, dissolved in 10% methanol/dichloromethane, added silica gel powder (5g), evaporated to dryness, and subjected to column chromatography. The product 1.2g is obtained by gradient elution with 5 percent to 15 percent methanol/dichloromethane, and the yield is 66.67 percent.

40-175

40-171(1.2g,0.0371mmol) was placed in a 250mL flask, dichloromethane (15mL) was added, TFA (0.1379mL, 1.8562mmol) was added, and the mixture was dissolved completely by sonication, and the reaction was stirred at room temperature with a ground glass stopper. And (3) after the reaction is finished, evaporating the reaction solution to dryness, removing dichloromethane, adding methyl tert-butyl ether (150mL), performing ultrasonic treatment for 2 minutes to separate out a solid product, performing suction filtration, dissolving the solid product by using a 20% methanol/dichloromethane solvent, adding silica gel powder, evaporating to dryness by using a rotary evaporator, performing dry sampling, performing column chromatography, and performing gradient elution by using 1% ammonia water and 5% -15% methanol/dichloromethane to obtain 0.8g of a product with the yield of 80.21%. ¹H-NMR(600MHz,DMSO-d₆)δ10.15-9.90(m,14H),9.05-9.03(m,1H),8.94-8.93(m,2H),8.76-8.71(m,12H),8.61-8.55(m,11H),8.25-7.58(m,155H),7.52-7.41(m,16H),7.36-6.99(m,198H),6.71-6.63(m,10H),6.57-6.50(m,4H),5.29-5.22(m,26H),4.80–4.52(m,39H),4.42-4.10(m,58H),4.06–3.89(m,14H),3.86–3.74(m,31H),3.70-3.56(m,65H),3.54–3.42(m,907H),3.22-2.98(m,81H),2.91-2.60(m,37H),2.43–2.09(m,68H),1.93-1.44(m,124H),1.39-1.14(m,144H),0.90-0.75(m,120H).

40-176

40-175(0.8g, 0.0282mmol) was weighed into a 250mL flask, dissolved in DMF (20mL), added DIEA (0.0932mL, 0.5640mmol), added M-SCM-5K (0.6206g, 0.1186mmol) and dissolved by sonication. The reaction was carried out at low speed in the absence of light. After the reaction is finished, adding methyl tert-butyl ether (150mL) into the reaction solution, separating out a solid, performing suction filtration, adding 10% methanol/dichloromethane for dissolution, adding silica gel powder, evaporating to dryness, and performing column chromatography. The product is 0.9g and the yield is 69.23 percent by using 5 to 15 percent methanol/dichloromethane gradient elution.

¹H-NMR(600MHz,DMSO-d₆)δ9.88-9.81(m,9H),8.94-8.55(m,17H),8.21-7.98(m,61H),7.87-7.71(m,21H),7.47-7.45(m,12H),7.23-7.14(m,292H),6.67-6.53(m,11H),5.26-5.23(m,19H),4.75-4.57(m,17H),4.35-4.19(m,35H),3.51-3.33(m,2730H),3.06-3.01(m,111H),2.62-2.60(m,62H),2.40-2.37(m,68H),2.18–1.76(m,221H),1.60-1.48(m,91H),1.24-1.16(m,57H),0.88-0.78(m,120H).

28. Synthesis of 46-51 (Compound No. 28)

39-227

H-Glu (oBzl) -oBzl (15g, 30.02mmol), Boc-Gly-OH (5.25g, 30.02mmol), HBTU (6.08g, 45.03mmol) and HOBT (17.07g, 45.03mmol) were added to a 250mL flask, 30mL of DMF was added to dissolve, the mixture was stirred at-5 ℃ for 30min, DIEA (22.33mL, 135.11mmol) was slowly added dropwise, the reaction was continued for 1H after the addition was completed, and then the mixture was stirred at room temperature overnight. After the reaction, transferring the reaction solution into a 1L separating funnel, adding pure water (200mL) and ethyl acetate (200mL) to extract to obtain an organic phase, washing the water phase with ethyl acetate (200mL x 2), combining the organic phases, washing the organic phases with saturated saline (200mL x 2), combining the organic phases, washing the organic phases with saturated sodium chloride solution (100mL x 2), concentrating the organic phases, performing dry sample loading column chromatography, and performing gradient elution with 20% -50% ethyl acetate/petroleum ether to obtain 10g of a product with the yield of 69%

33-232

39-227(2g,4.28mmol) was added to the reaction vessel, 10% Pd/C (0.075g) was added, DMF (40mL) was added and dissolved, and hydrogen was introduced under a pressure of 0.16MPa, and the mixture was stirred at room temperature overnight. After the reaction, the reaction mixture was filtered through celite, the filter cake was washed with DMF (20mL x 3), and the DMF solutions were combined and used as the starting material for the next step

33-233

14-128(6g, 6.27mmol), HBTU (3.2g, 8.55mmol) and HOBT (1.15g, 8.55mmol) were added to a 250mL flask, 30mL of DMF was added to dissolve the mixture, the mixture was stirred at-5 ℃ for reaction for 30min, DIEA (4.2mL, 25.65mmol) was slowly added dropwise, the reaction was continued for 1 h after the dropwise addition was complete, and the mixture was left at room temperature and stirred for reaction overnight. After the reaction is finished, n-hexane (100mL multiplied by 3) is added for sedimentation, the lower oily solid is dissolved by a small amount of dichloromethane, methyl tert-butyl ether is added, the solid is separated out, and the product 8g with over yield is obtained after drying.

33-235

33-233(8g,9.4mmol) was charged to a 500mL flask, dichloromethane (20mL) was added, TFA (13mL,188mmol) was added, and the reaction was stirred at room temperature overnight. After the reaction, the reaction solution was concentrated to 10mL, methyl tert-butyl ether (200mL) was added to precipitate a powder, the powder was filtered, the filter cake was washed with methyl tert-butyl ether (50mL × 3), and then dissolved in a (20% methanol: 80% dichloromethane) solution (200mL), silica gel powder (100mL) was added to the solution, the solution was evaporated to dryness to give a solid powder, and the solid powder was loaded by dry method, column chromatography, purified with 0.5% to 1% ammonia: eluting with 4% -7% methanol dichloromethane mixed solution, collecting, concentrating and evaporating to dryness to obtain 6g of product.

44-180

35-85(5.4828g, 5.4225mmol) was charged into a 250mL round bottom flask, dichloromethane (10mL) was added to dissolve, TFA (6.0mL, 81.3375mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. Transferring the reaction solution into a 1L separating funnel, adding saturated sodium bicarbonate aqueous solution (400mL) and ethyl acetate (200mL), shaking, extracting, and separating an aqueous phase and an organic phase; ethyl acetate (100mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, deionized water (400mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases were separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated, evaporated to dryness and dried. Obtaining products 44-180: 4.7556g, yield: 96.27 percent.

44-181

44-180(4.7556g, 5.2202mmol), Boc-L-Lys (Fmoc) -OH (2.2235g, 4.7456mmol), HBTU (2.6996g, 7.1185mmol) and HOBT (0.9618g, 7.1185mmol) were added to a 500mL round bottom flask, dissolved with DMF (80mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (4.7mL, 46.1032mmol) was slowly added dropwise, and after completion of the addition, the reaction flask was stirred at-5 ℃ for 4 hours. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding a saturated sodium chloride solution (400mL) and ethyl acetate (200mL), shaking, extracting, and separating an aqueous phase and an organic phase; ethyl acetate (100mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, and to the organic phase was added further saturated sodium chloride solution (300mL), shaken, extracted, and the aqueous and organic phases separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated, evaporated to dryness and dried. Obtaining products 44-181: 6.4613 g.

44-182

44-181(6.4613g, 4.7456mmol) was charged into a 250mL round-bottom flask, dichloromethane (10mL) was added to dissolve, TFA (5.3mL, 71.184mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. Transferring the reaction solution into a 1L separating funnel, adding a saturated sodium chloride solution (300mL) and ethyl acetate (200mL), shaking, extracting, and separating an aqueous phase and an organic phase; ethyl acetate (100mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, and to the organic phase was added further saturated sodium chloride solution (300mL), shaken, extracted, and the aqueous and organic phases separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated, evaporated to dryness and dried. Obtaining products 44-182: 5.9861 g.

44-183

44-182(5.9861g, 4.7456mmol), mono-tert-butyl succinate (0.9920g, 4.7456mmol), HBTU (2.6996g, 7.1185mmol) and HOBT (0.9618g, 7.1185mmol) were added to a 500mL round-bottomed flask, which was dissolved in DMF (80mL), and the reaction flask was stirred at-5 ℃ for about 30 minutes, followed by the slow dropwise addition of DIEA (4.7mL, 46.1032mmol), and after the dropwise addition, the reaction flask was stirred at-5 ℃ for 4 hours. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding a saturated sodium chloride solution (400mL) and ethyl acetate (200mL), shaking, extracting, and separating an aqueous phase and an organic phase; ethyl acetate (100mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, and to the organic phase was added further saturated sodium chloride solution (300mL), shaken, extracted, and the aqueous and organic phases separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated and evaporated to dryness. Dissolving the organic phase with 20% methanol/dichloromethane mixed solvent (100mL), adding silica gel powder (60mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (1% -2% methanol: 99% -98% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 44-183: 4.6123g, yield: 69.45 percent.

46-36

44-183(1.9313g, 1.3624mmol) and 10% Pd/C (80mg) were placed in a hydrogenation reactor, DMF (30mL) was added and dissolved, the hydrogenation reactor was sealed, hydrogen was introduced so that the pressure in the reactor became 2.0MPa, and the hydrogenation reactor was left at room temperature and stirred overnight. And after the reaction is finished, taking out the reaction kettle, uniformly dropwise adding the reaction solution into a sand core funnel filled with compacted diatomite, performing suction filtration, and cleaning the diatomite with DMF until the diatomite does not contain the product to obtain a reaction product solution.

46-39

46-36 solution (0.4349g, 0.4114mmol), 46-35(4.1058g, 1.9748mmol, synthesized by 33-235 methods), HBTU (0.9361g, 2.4684mmol) and HOBT (0.3335g, 2.4684mmol) were added to a 500mL round bottom flask, which was dissolved with DMF (80mL), and the reaction flask was stirred at-5 ℃ for about 30 minutes, then DIEA (1.2mL, 7.4054mmol) was slowly added dropwise, and after completion of the addition, the reaction flask was stirred at-5 ℃ for 1 hour, and finally transferred to room temperature and stirred overnight. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and settled, the supernatant was poured off, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and settled, and this was repeated three times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid, which was filtered to give the solid product, which was washed with methyl tert-butyl ether (60mL) and finally dried in an oven. Obtaining products 46-39: 3.8267 g.

46-41

46-39(3.8267g, 0.4114mmol) was charged into a 500mL round-bottom flask, dichloromethane (10mL) was added and dissolved, TFA (4.6mL, 61.71mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. To the reaction solution was added methyl tert-butyl ether (150mL) and the mixture was settled to give a solid as a powder, which was filtered and the solid product was washed with methyl tert-butyl ether (100 mL). Obtaining products 46-41: 2.794g, yield: 73.46 percent.

46-42

46-41(2.794g, 0.3022mmol), 36-98(0.3726g, 0.0.4533mmol), HBTU (0.1719g, 0.4533mmol) and HOBT (0.0613g, 0.4533mmol) were added to a 500mL round bottom flask, which was dissolved in DMF (80mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (0.3mL, 1.8132mmol) was slowly added dropwise, and after completion of the addition, the reaction flask was stirred at-5 ℃ for 1 hour, and finally transferred to room temperature and stirred overnight. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and settled, the supernatant was poured off, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and settled, and this was repeated three times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid, which was filtered to give the solid product, which was washed with methyl tert-butyl ether (60mL) and finally dried in an oven. Obtaining products 46-42: 3.0369 g.

46-44

46-42(3.0369g, 0.3022mmol) was charged into a 500mL round-bottom flask, DMF (10mL) was added and dissolved, morpholine (0.5mL, 5.4395mmol) was added with stirring, and the reaction flask was left to stir at room temperature for 2 hours. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and settled, the supernatant was poured off, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and settled, and this was repeated three times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid, which was filtered to give the solid product which was washed with methyl tert-butyl ether (60 mL). Products 46-44 are obtained: 0.4886g, yield: 16.5 percent.

42-92

Boc-L-Lys (Fmoc) -OH (9.0g, 19.2086mmol), glycine benzyl ester hydrochloride (4.2608g, 21.1295mmol), HBTU (10.9270g, 28.8129mmol) and HOBT (3.8932g, 28.8129mmol) were added to a 500mL round bottom flask, which, after dissolution with DMF (60mL), was stirred at-5 ℃ for about 30 minutes, DIEA (14.3mL, 86.4387mmol) was slowly added dropwise, and after completion of the addition, the reaction flask was allowed to continue to react at-5 ℃ for 3 hours with stirring. After the reaction, the reaction solution was transferred to a 1L separatory funnel, to which a saturated aqueous sodium chloride solution (300mL) and ethyl acetate (200mL) were added, shaken, extracted, and the aqueous phase was separated; ethyl acetate (100mL) was added to the aqueous phase, shaken, extracted and the aqueous phase separated. The organic phases were combined, and then saturated aqueous sodium chloride (300mL) was added to the organic phase, followed by shaking, extraction and separation of the aqueous phase. Then, deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous phase was separated. Finally, the organic phase is concentrated and evaporated to dryness and placed in an oven for drying. To products 42-92: 11.8271 g.

42-93

42-92(11.8271g, 19.2086mmol) was placed in a 500mL round bottom flask, dichloromethane (40mL) was added and dissolved, TFA (21.4mL, 288.129mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature overnight. After the reaction, the reaction solution is concentrated and evaporated to dryness to remove dichloromethane, then methyl tert-butyl ether (150mL) is added into the reaction solution to be settled to form powdery solid, the powdery solid is filtered, the filter cake is washed by methyl tert-butyl ether (60mL), and finally the filter cake is placed in an oven to be dried. To products 42-93: 9.9040 g.

42-128

42-93(9.9040g, 19.2086mmol), mono-tert-butyl succinate (3.6805g, 21.1295mmol), HBTU (10.9270g, 28.8129mmol) and HOBT (3.8932g, 28.8129mmol) were added to a 500mL round-bottomed flask, which was dissolved in DMF (100mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (14.2mL, 86.4387mmol) was slowly added dropwise, and after completion of the addition, the reaction flask was stirred at-5 ℃ for 1 hour, and finally transferred to room temperature and stirred. After the reaction, the reaction solution was transferred to a 1L separatory funnel, and a saturated aqueous sodium chloride solution (400mL) and ethyl acetate (300mL) were added, shaken, extracted, and the aqueous phase and the organic phase were separated; ethyl acetate (200mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, aqueous sodium chloride (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated, evaporated to dryness and dried. Obtaining products 42-128: 12.9040 g.

42-133

42-128(12.9040g, 19.2086mmol) was charged into a 500mL round-bottom flask, DMF (20mL) was added and dissolved, morpholine (25.1mL, 288.1290mmol) was added with stirring, and the reaction flask was left to stir at room temperature for 2 hours. After the reaction, transferring the reaction solution to a 1L separating funnel, adding a saturated sodium bicarbonate aqueous solution (400mL) and ethyl acetate (300mL), shaking, extracting, and separating an aqueous phase and an organic phase; ethyl acetate (200mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, deionized water (400mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases were separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated, evaporated to dryness and dried. Obtaining products 42-133: 8.6350 g.

42-134

42-133(7.5191g, 16.7263mmol), 44-149 solution (i.e. 36-186, 2.4930g, 4.6462mmol), HBTU (7.9291g, 20.9079mmol) and HOBT (2.8251g, 20.9079mmol) were added to a 500mL round bottom flask, after dissolution with DMF (80mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (10.4mL, 62.7237mmol) was slowly added dropwise, after completion of the addition, the reaction flask was stirred at-5 ℃ for 1 hour, and finally transferred to room temperature and stirred. After the reaction, transferring the reaction solution to a 1L separating funnel, adding a saturated sodium bicarbonate aqueous solution (400mL) and ethyl acetate (300mL), shaking, extracting, and separating an aqueous phase and an organic phase; ethyl acetate (200mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases were separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated. Dissolving the organic phase with 20% methanol/dichloromethane mixed solvent (60mL), adding silica gel powder (40mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (2% -8% methanol: 98% -92% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 42-134: 4.8g, yield: 66.19 percent.

¹H NMR(400MHz,DMSO-d₆)δ8.38–8.35(m,3H),8.11–8.01(m,3H),7.56(t,J＝5.2Hz,3H),7.44–7.27(m,15H),5.12(s,6H),4.36–4.17(m,3H),3.97–3.80(m,6H),3.33(s,11H),3.06(m,6H),2.45–2.31(m,12H),1.69–1.46(m,12H),1.36(m,27H),1.32–1.26(m,6H).

46-46

42-134(0.5g, 0.3204mmol) and 10% Pd/C (50mg) were placed in a hydrogenation reactor, DMF (30mL) was added to dissolve, the hydrogenation reactor was closed, hydrogen was introduced to make the pressure in the reactor 2.0MPa, and the hydrogenation reactor was left at room temperature and stirred overnight. And after the reaction is finished, taking out the reaction kettle, uniformly dropwise adding the reaction solution into a sand core funnel filled with compacted diatomite, performing suction filtration, and cleaning the diatomite with DMF until the diatomite does not contain the product to obtain a reaction product solution.

46-47

46-46 solution (0.0194g, 0.0151mmol), 46-44(0.4886g, 0.0497mmol), HBTU (0.0257g, 0.0678mmol) and HOBT (0.0092g, 0.0678mmol) were added to a 500mL round bottom flask, which, after dissolution with DMF (100mL), was stirred for about 30 minutes at-5 deg.C, then DIEA (0.1mL, 1.057mmol) was slowly added dropwise, after which the reaction flask was stirred for 30 minutes at-5 deg.C and finally transferred to room temperature and stirred overnight. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and settled, the supernatant was poured off, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and settled, and this was repeated three times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid, which was filtered to give the solid product, which was washed with methyl tert-butyl ether (60mL) and finally dried in an oven. Obtaining products 46-47: 0.4628 g.

46-49

46-47(0.4628g, 0.0151mmol) was placed in a 250mL round-bottom flask, dichloromethane (10mL) was added to dissolve, TFA (5mL, 2.265mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. To the reaction solution was added methyl tert-butyl ether (150mL) and the mixture was settled to give a solid as a powder, which was filtered and the solid product was washed with methyl tert-butyl ether (100 mL). The solid product was dissolved in a 20% methanol/dichloromethane mixed solvent (80mL), silica gel powder (50mL) was added, evaporated to dryness, dry loaded, and subjected to column chromatography. Eluting with eluent (1% ammonia water: 6% -10% methanol: 93% -89% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 46-49: 0.2613g, yield: 56.64 percent.

46-51

Mixing 46-49(0.2613g, 0.0085mmol), M-NH₂HCl-10K (0.3253g, 0.0308mmol), HBTU (0.0146g, 0.0385mmol) and HOBT (0.0052g, 0.0385mmol) were added to a 250mL round-bottomed flask, which was dissolved in DMF (20mL), the reaction flask was stirred at-5 ℃ for about 30 min, DIEA (0.1mL, 1.9422mmol) was added dropwise slowly, and after completion of the addition, the reaction flask was stirred at-5 ℃ for 30 min and finally transferred to room temperature and stirred at low speed for one week away from light. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction solution to precipitate, the supernatant was poured off, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance to precipitate, and the precipitation was repeated five times to obtain a powdery solid, and the solid product was filtered and washed with methyl tert-butyl ether (100 mL). Dissolving the solid product with 20% methanol/dichloromethane mixed solvent (60mL), adding silica gel powder (50mL), evaporating to dryness, dry loading, and performing column chromatography. Eluting with eluent (1% ammonia water: 5% -15% methanol: 94% -84% dichloromethane), collecting liquid, concentrating, evaporating, and drying. Obtaining products 46-51: 0.2251g, yield: 42.66 percent. ¹H NMR(600MHz,DMSO-d₆)δ8.97–8.53(m,41H),8.32–7.90(m,183H),7.90–7.60(m,57H),7.54–7.45(m,27H),7.38–6.99(m,254H),6.67–6.53(m,21H),5.26–5.24(m,42H),4.87–4.03(m,168H),3.87–3.40(m,2876H),3.24–3.11(m,110H),3.10–2.93(m,142H),2.92–2.70(m,108H),2.69–2.61(m,20H),2.43–2.00(m,104H),1.86–1.47(m,162H),1.23-1.11(m,61H),0.99–0.66(m,162H).

29. Synthesis of 44-234 (Compound No. 29)

36-200

35-84(8.99g, 11.7386mmol), Boc-L-Lys (Fmoc) -OH (5.0g, 10.6714mmol), HBTU (6.0706g, 16.0072mmol) and HOBT (2.1629g, 16.0072mmol) were added to a 500mL round bottom flask, which, after dissolution with DMF (80mL), was stirred at-5 ℃ for about 30 minutes, DIEA (10.6mL, 64.0287mmol) was slowly added dropwise and, after completion of the addition, the reaction flask was stirred at-5 ℃ for 3 hours. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding a saturated sodium chloride solution (300mL) and ethyl acetate (200mL), shaking, extracting, and separating an aqueous phase and an organic phase; ethyl acetate (100mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, and to the organic phase was added further saturated sodium chloride solution (300mL), shaken, extracted, and the aqueous and organic phases separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated, evaporated to dryness and dried. Obtaining products 36-200: 12.9804 g.

44-207

36-200(11.9g, 9.7832mmol) was charged into a 500mL round bottom flask, dichloromethane (20mL) was added to dissolve, TFA (10.9mL, 146.748mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. To the reaction solution was added methyl tert-butyl ether (150mL) and the mixture was settled to give a solid as a powder, which was filtered and the solid product was washed with methyl tert-butyl ether (100 mL). Obtaining products 44-207: 8.351g, yield: 76.47 percent.

44-208

Mono-tert-butylsuccinate (1.5638g, 8.9775mmol), 44-207(8.351g, 7.4812mmol), HBTU (4.2558g, 11.2218mmol) and HOBT (1.5163g, 11.2218mmol) were weighed into a 500mL round-bottomed flask, which was dissolved with DMF (80mL), and the reaction flask was stirred at-5 ℃ for about 30 minutes, followed by the slow dropwise addition of DIEA (9.9mL, 59.8499mmol), and after the dropwise addition was complete, the reaction flask was stirred at-5 ℃ for 3 hours. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding a saturated sodium chloride solution (300mL) and ethyl acetate (200mL), shaking, extracting, and separating an aqueous phase and an organic phase; ethyl acetate (100mL) was added to the aqueous phase, shaken, extracted and the aqueous and organic phases separated. The organic phases were then combined, and to the organic phase was added further saturated sodium chloride solution (300mL), shaken, extracted, and the aqueous and organic phases separated. Deionized water (300mL) was added to the organic phase, shaken, extracted, and the aqueous and organic phases separated. Finally, the organic phase is concentrated and evaporated to dryness. Dissolving the organic phase with 20% methanol/dichloromethane mixed solvent (100mL), adding silica gel powder (50mL), evaporating to dryness, dry-loading, and performing column chromatography. Eluting with eluent (2% -8% methanol: 98% -92% dichloromethane). Collecting liquid, concentrating, evaporating, and drying. Obtaining products 44-208: 2.5268g, yield: 35.85 percent.

44-212

44-208(2.5268g, 1.9858mmol) and 10% Pd/C (60mg) are put into a hydrogenation reaction kettle, DMF (30mL) is added to dissolve the mixture, the hydrogenation reaction kettle is sealed, hydrogen is introduced to ensure that the pressure in the hydrogenation reaction kettle reaches 2.2MPa, and the hydrogenation reaction kettle is placed at room temperature and stirred overnight. And after the reaction is finished, taking out the reaction kettle, uniformly dropwise adding the reaction solution into a sand core funnel filled with compacted diatomite, performing suction filtration, and cleaning the diatomite with DMF until the diatomite does not contain the product to obtain a reaction product solution.

44-213

44-212 solution (1.0846g, 1.1893mmol), 36-145(5.0g, 5.2329mmol), HBTU (2.7062g, 7.1358mmol) and HOBT (0.9642g, 7.1358mmol) were added to a 500mL round bottom flask, which was dissolved in DMF (80mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (3.5mL, 21.4074mmol) was slowly added dropwise, and after completion of the addition, the reaction flask was stirred at-5 ℃ for 1 hour, and finally was transferred to room temperature and stirred for 3 hours. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and settled, the supernatant was poured off, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and settled, and this was repeated three times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid, which was filtered to give the solid product, which was washed with methyl tert-butyl ether (60mL) and finally dried in an oven. Obtaining products 44-213: 5.5443 g.

44-215

44-213(5.5443g, 1.1893mmol) was placed in a 500mL round bottom flask, dichloromethane (20mL) was added and dissolved, TFA (1.3mL, 17.8395mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. To the reaction solution was added methyl tert-butyl ether (150mL) and the mixture was settled to give a solid as a powder, which was filtered and the solid product was washed with methyl tert-butyl ether (100 mL). Obtaining products 44-215: 5.0g, yield: 91.28 percent.

44-218

44-215(5.0g, 1.0856mmol), 36-98(1.16g, 1.4113mmol), HBTU (0.6176g, 1.6284mmol) and HOBT (0.22g, 1.6284mmol) were added to a 500mL round bottom flask, which was dissolved with DMF (80mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (1.1mL, 6.5136mmol) was slowly added dropwise, and after this addition, the reaction flask was stirred at-5 ℃ for 1 hour and finally transferred to room temperature for 3 hours. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and settled, the supernatant was poured off, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and settled, and this was repeated three times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid, which was filtered to give the solid product, which was washed with methyl tert-butyl ether (60mL) and finally dried in an oven. Obtaining products 44-218: 5.8728 g.

44-219

44-218(5.8728g, 1.0856mmol) was charged into a 500mL round-bottom flask, DMF (20mL) was added and dissolved, morpholine (1.9mL, 21.7120mmol) was added with stirring, and the reaction flask was left to stir at room temperature for 2 hours. After the reaction was completed, n-hexane (150mL) and methyl t-butyl ether (30mL) were added to the reaction mixture, followed by settling, the supernatant was poured off, and n-hexane (150mL) and methyl t-butyl ether (30mL) were added to the lower oily substance, followed by settling, and the operation was repeated five times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid, which was filtered to give the solid product which was washed with methyl tert-butyl ether (60 mL). Dissolving the solid product with 20% methanol/dichloromethane mixed solvent (100mL), adding silica gel powder (60mL), evaporating to dryness, dry loading, and performing column chromatography. Eluting with eluent (1% ammonia water: 3% -15% methanol: 96% -84% dichloromethane). Collecting liquid, concentrating, evaporating, and drying. Obtaining products 44-219: 0.8841g, yield: 15.7 percent.

44-221

Adding 46-46 solution (0.0611g, 0.0473mmol), 44-219((0.8841g, 0.1704mmol), HBTU (0.0808g, 0.2130mmol) and HOBT (0.0288g, 0.2130mmol) into a 500mL round-bottom flask, dissolving the solutions with DMF (30mL), stirring the reaction flask at-5 ℃ for about 30 minutes, slowly dropwise adding DIEA (0.1mL, 0.6628mmol), stirring the reaction flask at-5 ℃ for 30 minutes after the dropwise addition is finished, transferring the reaction flask to room temperature, keeping out of the light, stirring the reaction solution overnight, adding n-hexane (150mL) and methyl tert-butyl ether (30mL) into the reaction solution, settling the reaction solution, pouring the supernatant, adding n-hexane (150mL) and methyl tert-butyl ether (30mL) into the lower oily substance, settling the oily solid three times, dissolving the oily solid with dichloromethane (10mL), adding methyl tert-butyl ether (150mL) into the reaction solution, and settling the oily solid, as a powdery solid, filtered to give a solid product, and the solid product was washed with methyl t-butyl ether (60mL) and dried in an oven. Obtaining products 44-221: 0.7946 g.

44-232

44-221(0.7946g, 0.0473mmol) was placed in a 250mL round bottom flask, dichloromethane (20mL) was added and dissolved, TFA (0.5mL, 7.095mmol) was added with stirring, and finally the reaction flask was left to stir at room temperature. After the reaction, the reaction mixture was concentrated under reduced pressure to remove methylene chloride. To the reaction solution was added methyl tert-butyl ether (150mL) and the mixture was settled to give a solid as a powder, which was filtered and the solid product was washed with methyl tert-butyl ether (100 mL). Dissolving the solid product with 20% methanol/dichloromethane mixed solvent (60mL), adding silica gel powder (30mL), evaporating to dryness, loading by dry method, and performing column chromatography. Eluting with eluent (1% ammonia water: 3% -10% methanol: 96% -89% dichloromethane). Collecting liquid, concentrating, evaporating, and drying. Obtaining products 44-232: 0.2239g, yield: 28.46 percent.

44-234

Mixing 44-232(0.2239g, 0.0135mmol), M-NH₂HCl-10K (0.5547g, 0.0525mmol), HBTU (0.0230g, 0.0606mmol) and HOBT (0.0082g, 0.0606mmol) were added to a 250mL round-bottomed flask, dissolved in DMF (80mL), the reaction flask was stirred at-5 ℃ for about 30 minutes, DIEA (0.1mL, 0.5385mmol) was slowly added dropwise, and after completion of the addition, the reaction flask was stirred at-5 ℃ for 1 hour, and finally transferred to room temperature and stirred for one week away from light. After the reaction, n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the reaction mixture and settled, the supernatant was poured off, and n-hexane (150mL) and methyl tert-butyl ether (30mL) were added to the lower oily substance and settled, and this was repeated three times to obtain an oily solid. The oily solid was dissolved in dichloromethane (10mL) and then precipitated by addition of methyl tert-butyl ether (150mL) as a powdery solid, which was filtered to give the solid product which was washed with methyl tert-butyl ether (60 mL). The solid product was dissolved in a 20% methanol/dichloromethane mixed solvent (100mL), silica gel powder (50mL) was added, evaporated to dryness, dry loaded, and subjected to column chromatography. Eluting with eluent (1% ammonia water: 3% -10% methanol: 96% -89% dichloromethane). Collecting liquid, concentrating, evaporating, and drying. Obtaining products 44-234: 0.2549g, yield: 39.2 percent.

¹H-NMR(600MHz,DMSO-d₆)δ8.95–8.51(m,17H),8.26–7.92(m,115H),7.53–7.46(m,16H),7.35–7.25(m,38H),7.19–7.17(m,84H),4.84–4.07(m,110H),4.05–3.98(m,13H),3.53–3.51(m,2874H),3.25–2.94(m,117H),2.89–2.85(m,73H),2.77–2.73(m,47H),2.69–2.61(m,16H),2.41–2.29(m,52H),2.23–1.75(m,71H),1.59–1.48(m,77H),1.34–1.23(m,30H),0.85–0.83(m,60H)

30. Synthesis of 35-167 (Compound No. 32)

35-124

Fmoc-Gl-OH (3.57g, 9.66mmol), HBTU (10.99g, 28.9mmol), HOBT (3.91g, 28.9mmol) and H-Glu- (OtBu) -OtBu.HCl (6.57g, 20.28mmol) were added to a 500mL flask, then DMF (150mL) was added to dissolve it, the reaction was left to stir at 0 ℃ for about 30 minutes, DIEA (8.78mL, 53.11mmol) was slowly added dropwise, and after the addition, the reaction was continued to stir at 0 ℃ overnight. After the reaction, the reaction solution was transferred to a 1L separatory funnel, deionized water (200mL) and ethyl acetate (250mL) were added for extraction, the aqueous phase was washed with ethyl acetate (200mL), the organic phases were combined, the organic phase was concentrated with saturated saline (200mL) and evaporated to dryness to obtain 35-124:8.2 g.

35-125

35-124(8.2g,9.62mmol) was placed in a 500mL flask, dichloromethane (30mL) was added to dissolve it, trifluoroacetic acid (21.4mL,288.6mmol) was added, the reaction was stirred overnight at room temperature, after the reaction was complete, methyl t-butyl ether (200mL) was added to precipitate a solid, the solid was filtered, the filter cake was washed with methyl t-butyl ether (50 mL. times.2), and the filter cake was dried to obtain 35-125:6g of a product.

35-121

Fmoc-Gly-OH (9g, 30.27mmol), HBTU (16g, 42.19mmol), HOBT (6g, 44.4mmol) and H-Glu- (OtBu) -OtBu.HCl (8g, 27.05mmol) were added to a 500mL flask, then DMF (80mL) was added to dissolve it, the reaction was left to stir at 0 ℃ for about 30 minutes, DIEA (25mL, 148.5mmol) was slowly added dropwise, and after addition, the reaction was allowed to stir at 0 ℃ overnight. After the reaction, the reaction solution was transferred to a 1L separatory funnel, deionized water (200mL) and ethyl acetate (300mL) were added for extraction, the aqueous phase was washed with ethyl acetate (200mL × 2), the organic phases were combined, the organic phase was concentrated with saturated saline (200mL × 2) and evaporated to dryness to obtain 35-121:14g of product, yield: 87.5 percent.

35-122

Putting 35-121(14g,26mmol) into a 250mL flask, adding DMF (200mL) to dissolve, adding morpholine (14mL,260mmol), reacting at room temperature for 1 hour, finishing the reaction, transferring the reaction solution into a 1L separating funnel, adding deionized water (200mL) and ethyl acetate (200mL) to extract to obtain an organic phase, washing the aqueous phase with ethyl acetate (200mL × 2), combining the organic phases, washing the organic phase with saturated saline (200mL × 1), concentrating and evaporating to dryness to obtain a product 35-122:68.2 g.

35-126

35-122(8.1g, 25.6mmol), HBTU (12.1g, 31.98mmol), HOBT (4.3g, 31.98mmol) and 35-125(3.35g, 5.33mmol) were added to a 500mL flask, DMF (50mL) was added to dissolve, the reaction was stirred at 0 ℃ for about 20 minutes, DIEA (15.9mL, 95.94mmol) was slowly added dropwise, and after the addition, the reaction was stirred at 0 ℃ overnight. After the reaction is finished, transferring the reaction solution into a 2L separating funnel, adding deionized water (250mL) and ethyl acetate (300mL) for extraction, washing an aqueous phase with ethyl acetate (150mL multiplied by 1), combining organic phases, using saturated saline (200mL multiplied by 1) for the organic phases, concentrating and evaporating to dryness, adding methanol (20mL) and dichloromethane (80mL) for dissolution, adding silica gel powder (30g), evaporating to dryness to form powdery solid, loading by a dry method, carrying out column chromatography, and eluting with a mixed solution of 5% -8% methanol and dichloromethane to obtain a product 35-126:9.7 g.

35-128

35-126(9.7g,5.33mmol) was placed in a 250mL flask, DMF (150mL) was added and dissolved, morpholine (4.6mL,53.3mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction is finished, adding n-hexane (200mL) and methyl tert-butyl ether (50mL) into the reaction solution, shaking to separate layers, pouring out the supernatant, adding n-hexane (200mL) and methyl tert-butyl ether (50mL) into the lower oily substance continuously, repeating the steps for 4 times, and finally obtaining oily substance which is dried to obtain 35-128: 6.36g, yield: 75 percent.

35-111

Fmoc-Gl-OtBu (5g, 11.8mmol), HBTU (6.7g, 17.7mmol), HOBT (2.4g, 17.7mmol) and H-Gly-OBn.HCl (2.5g, 12.37mmol) were added to a 500mL flask, DMF (80mL) was added to dissolve, the reaction was allowed to stand at 0 ℃ and stirred for about 30 minutes, DIEA (11mL, 64.9mmol) was slowly added dropwise, and after dropwise addition, the reaction was allowed to continue stirring at 0 ℃ overnight. After the reaction, the reaction mixture was transferred to a 1L separatory funnel, deionized water (200mL) and ethyl acetate (300mL) were added to extract the mixture, the aqueous phase was washed with ethyl acetate (200 mL. times.2), the organic phases were combined, the organic phase was concentrated with saturated brine (200 mL. times.2) and evaporated to dryness to obtain 35-111:5.7g of a product.

35-112

35-111(6.7g,11.8mmol) was placed in a 250mL flask, DMF (45mL) was added and dissolved, morpholine (8.22mL,94.4mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction, the reaction solution was transferred to a 1L separatory funnel, deionized water (150mL) and ethyl acetate (250mL) were added to extract the solution to obtain an organic phase, the aqueous phase was washed with ethyl acetate (200 mL. times.3), the organic phases were combined, the organic phase was washed with saturated saline (200 mL. times.1), concentrated and evaporated to dryness to obtain 35-112:3.2g of a product with a yield of 78%.

35-113

35-112(3.2g, 9.1mmol), HBTU (5.2g, 13.65mmol), HOBT (1.8g, 13.65mmol) and Fmoc-Gly-OH (3.2g, 10.92mmol) were added to a 250mL flask, DMF (30mL) was added to dissolve, the reaction was allowed to stand at 0 ℃ and stirred for about 20 minutes, DIEA (7mL, 42.39mmol) was slowly added dropwise, and after the addition was complete, the reaction was allowed to continue stirring at 0 ℃ overnight. After the reaction is finished, transferring the reaction solution into a 2L separating funnel, adding deionized water (250mL) and ethyl acetate (300mL) into the separating funnel, extracting, washing an aqueous phase by using ethyl acetate (150mL multiplied by 1), combining organic phases, concentrating and evaporating the organic phases by using saturated saline (200mL multiplied by 1), adding methanol (20mL) and dichloromethane (80mL) into the organic phases for dissolving, adding silica gel powder (30g), evaporating the silica gel powder to be powdery solid, loading the mixture by a dry method, performing column chromatography, and eluting the mixture by using a mixed solution of 30% -70% of petroleum ether and ethyl acetate to obtain a product 35-113:5.7 g.

35-114

Putting 35-113(5.7g,9.1mmol) into a 250mL flask, adding dichloromethane (30mL) to dissolve, adding trifluoroacetic acid (20.4mL,273mmol), stirring at room temperature overnight to complete the reaction, concentrating the reaction solution to a small amount, adding n-hexane (150mL) and methyl tert-butyl ether (40mL) to separate layers, pouring the upper layer liquid, adding n-hexane (150mL) and methyl tert-butyl ether (40mL) to the lower layer liquid to form a viscous oil, and drying to obtain 35-114:3.59g of a product with a yield of 68%.

35-142

35-128(10.2g, 6.4mmol), HBTU (1.36g, 3.59mmol), HOBT (0.49g, 3.59mmol) and 35-114(3g, 5.3mmol) were added to a 250mL flask, DMF (40mL) was added to dissolve, the reaction was allowed to stir at 0 ℃ for about 20 minutes, DIEA (5mL, 29.15mmol) was slowly added dropwise, and after the addition, the reaction was allowed to continue stirring at 0 ℃ overnight. After the reaction is finished, transferring the reaction solution into a 1L separating funnel, adding deionized water (250mL) and ethyl acetate (300mL) for extraction, washing an aqueous phase with ethyl acetate (150mL multiplied by 1), combining organic phases, using saturated saline (200mL multiplied by 1) for the organic phases, concentrating and evaporating to dryness, adding methanol (20mL) and dichloromethane (80mL) for dissolution, adding silica gel powder (15g), evaporating to dryness to form powdery solid, loading by a dry method, performing column chromatography, and eluting with a mixed solution of 4% -8% methanol and dichloromethane to obtain 35-142:4g of a product, wherein the yield is 81%.

35-144

35-142(9.2g,1.86mmol) was charged into a hydrogenation reactor, 10% Pd/C (0.050 g) was added thereto, DMF (40mL) was added thereto and dissolved therein, and hydrogen gas was introduced thereinto under a pressure Pa of 1.8MPa, and the reaction was stirred at room temperature overnight. After completion of the reaction, the reaction mixture was filtered through celite, the celite was washed with DMF (20mL × 3), and the DMF solutions were combined to be used as a raw material for the next reaction.

35-115

Boc-Lys (Fmoc) -OH (4.7g, 10mmol), HBTU (5.7g, 15mmol), HOBT (2.0g, 15mmol) and H-Gly-OBn.HCl (2.2g, 11mmol) were added to a 500mL flask, then DMF (50mL) was added to dissolve it, the reaction was left to stir at-5 ℃ for about 30 minutes, DIEA (9.1mL, 55mmol) was slowly added dropwise, after which the reaction was allowed to continue stirring at-5 ℃ for 1 hour and finally transferred to room temperature for stirring overnight. After the reaction, the reaction mixture was transferred to a 1L separatory funnel, and a saturated sodium chloride solution (200mL) and ethyl acetate (250mL) were added to extract the mixture, the aqueous phase was washed with ethyl acetate (200 mL. times.1), the organic phases were combined, the organic phase was concentrated with a saturated saline solution (200 mL. times.2) and evaporated to dryness to obtain 35-115:6.5g of a product.

35-116

35-115(7.9g,12.81mmol) was placed in a 250mL flask, methylene chloride (20mL) was added and dissolved, trifluoroacetic acid (7.4mL,100mmol) was added with stirring, and the reaction was stirred at room temperature overnight. After the reaction, the reaction solution was concentrated to a small amount, and n-hexane (150mL) and methyl t-butyl ether (40mL) were added, and the mixture was settled for 2 times to give a viscous oily substance, and dried to obtain 35-116:5.2g of a product.

35-146

35-144(8.9g, 4.3mmol), HBTU (2.45g, 6.45mmol), HOBT (0.9g, 6.45mmol) and 35-116(2.7g, 4.3mmol) were added to a 250mL flask, DMF (200mL) was added to dissolve, the reaction was stirred at 0 ℃ for about 20 minutes, DIEA (3.9mL, 23.65mmol) was slowly added dropwise, and after the addition, the reaction was stirred at 0 ℃ overnight. After the reaction is finished, n-hexane (200mL) and methyl tert-butyl ether (40mL) are added into the reaction liquid, layers are separated, the upper layer liquid is poured, n-hexane (200mL) and methyl tert-butyl ether (40mL) are added into the lower layer oily matter continuously, the steps are repeated for 3 times, and the oily viscous matter is dried to obtain 35-146:11g of the product.

35-148

35-146(11g,4.3mmol) was charged into a hydrogenation reactor, 10% Pd/C (0.050g) was added thereto, DMF (40mL) was added thereto and dissolved therein, and hydrogen gas was introduced under a pressure Pa of 1.8MPa, and the reaction was stirred at room temperature overnight. After completion of the reaction, the reaction mixture was filtered through celite, the celite was washed with DMF (20mL × 3), and the DMF solutions were combined to be used as a raw material for the next reaction.

35-155

35-148(6.8g, 2.75mmol), HBTU (1.4g, 3.75mmol), HOBT (0.51g, 3.75mmol) and 35-128(4g, 2.5mmol) were added to a 250mL flask, DMF (150mL) was added to dissolve, the reaction was allowed to stir at 0 ℃ for about 20 minutes, DIEA (1.86mL, 11.25mmol) was slowly added dropwise, and after the addition, the reaction was allowed to continue stirring at 0 ℃ overnight. After the reaction is finished, adding n-hexane (200mL) and methyl tert-butyl ether (40mL) into the reaction solution, carrying out layering, pouring the upper layer liquid, continuously adding n-hexane (200mL) and methyl tert-butyl ether (40mL) into the lower layer oily substance, repeating the steps for 3 times to obtain an oily viscous substance, adding dichloromethane and methanol to dissolve the oily viscous substance, adding silica gel powder (20g), evaporating the oily viscous substance to obtain a powdery solid, carrying out dry loading, carrying out column chromatography, eluting with a mixed solution of 0.5% ammonia water and 4% -10% methanol in dichloromethane, collecting and drying to obtain a product 35-155:1.2g of yield: 12 percent. ¹H-NMR(600MHz,DMSO-d₆)δ8.13–8.05(m,22H),7.85–7.55(m,8H),7.38–7.28(m,8H),4.71–4.33(m,7H),4.31–4.09(m,14H),3.85–3.42(m,12H),3.18–2.35(m,4H),2.31–2.29(m,32H),2.07–1.91(m,26H),1.89–1.55(m,6H),1.45–1.38(m,144H),1.25–1.11(m,2H).

35-157

Putting 35-155(1.2g,0.3mmol) into a 250mL flask, adding DMF (25mL) to dissolve, adding morpholine (4mL,43.68mmol), stirring at room temperature for 1 hour, finishing the reaction, adding n-hexane (100mL) and methyl tert-butyl ether (30mL) into the reaction solution, layering, pouring the upper layer liquid, adding n-hexane (100mL) and methyl tert-butyl ether (30mL) into the lower layer oily substance, repeating the steps for 3 times to obtain oily viscous substance, and drying to obtain 35-157:1.02g of the product.

35-159

Putting 35-157(1.02g,0.3mmol) into a 250mL flask, adding dichloromethane (30mL) to dissolve, adding trifluoroacetic acid (10mL,134mmol), stirring at room temperature overnight, finishing the reaction, concentrating the reaction solution to a small amount, adding n-hexane (100mL) and methyl tert-butyl ether (30mL) to separate layers, pouring the upper layer liquid, adding n-hexane (100mL) and methyl tert-butyl ether (30mL) to the lower layer liquid to form a viscous oily substance, and drying to obtain 35-159:0.8g of a product.

35-161

35-159(0.1g,0.0367mmol) was placed in a 250mL flask, DMF (20mL) was added to dissolve, DIEA (0.6mL,3.63mmol) was added and stirred for 30 minutes, M-SCM-10K (0.85g,0.081mmol, from Keka) was added and the reaction was transferred to room temperature and stirred at low speed in the dark for 7 days. After the reaction is finished, adding n-hexane (100mL) and methyl tert-butyl ether (30mL) into the reaction solution, layering, pouring the upper layer of liquid, continuously adding n-hexane (100mL) and methyl tert-butyl ether (30mL) into the lower layer of oily substance, repeating the steps for 3 times to obtain oily substance, dissolving the oily substance by using methanol (30mL) and dichloromethane (120mL), adding silica gel powder (20g), evaporating to dryness to obtain powdery solid, performing dry sampling, performing column chromatography, eluting by using a dichloromethane mixed solution of 1% ammonia water and 6% -10% methanol, collecting and concentrating, evaporating to dryness to obtain solid, and drying in a vacuum oven to obtain a product 35-161: 0.87g, yield: 58 percent.

35-167

35-161(0.5g,0.02mmol), 35-99(0.45g,0.5mmol, synthesized by 25-132 method), HBTU (0.l5g,0.05mmol), HOBT (0.06g,0.05mmol) were added to a 250mL flask, DMF (35mL) was added to dissolve it, the reaction was left at-5 ℃ and stirred for 20 minutes, DIEA (0.22mL,1.4mmol) was slowly added dropwise, after completion of dropwise addition, the reaction was stirred at-5 ℃ for 30 minutes, and then stirred at room temperature for 6 days under low speed protected from light. After the reaction is finished, adding n-hexane (100mL) and methyl tert-butyl ether (30mL) to generate layering, pouring the upper layer of liquid, continuously adding n-hexane (100mL) and methyl tert-butyl ether (30mL) into the lower layer of oily substance, repeating the steps for 3 times to obtain a viscous oily substance, dissolving the viscous oily substance by using methanol (30mL) and dichloromethane (120mL), adding silica gel powder (15g), evaporating to dryness to obtain a powdery solid, loading the powdery solid by a dry method, performing column chromatography, eluting by using a dichloromethane mixed solution of 1% of ammonia water and 7% -10% of methanol, collecting and concentrating, evaporating to obtain a solid, and drying by a vacuum oven to obtain a product

35-167: 0.24g, yield: 50 percent.¹H-NMR(600MHz,DMSO-d₆)δ9.04-9.03(m,2H),8.37-8.34(m,29H),8.18–8.02(m,42H),8.01–7.80(m,48H),7.61-7.59(m,38H),7.39-7.36(m,25H),7.27–6.95(m,190H),6.69-6.66(m,4H),4.67–4.49(m,13H),4.38–4.23(m,16H),4.19–4.12(m,10H),3.90-3.87(m,11H),3.79-3.75(m,9H),3.70-3.68(m,16H),3.61-3.58(m,20H),3.51-3.49(m,1941H),3.14–3.03(m,68H),2.68–2.59(m,73H),2.35-2.34(m,39H),1.61–1.54(m,16H),1.48(m,34H),1.34-1.32(m,31H),1.23-1.21(m,186H),0.95–0.81(m,139H),0.52-0.05(m,28H).

31. Synthesis of 41-137 (Compound No. 30)

6-aminocaproic acid (4.61g, 35.1407mmol) was weighed into a 1L flask, 150mL THF H was added₂The solution was mixed 1:1 to dissolve completely, stirred at 0 ℃, and then solid sodium carbonate (7.45g, 70.2814mmol) was added, dissolved by sonication, and stirred at 0 ℃. After 30 minutes of reaction, Fmoc-Cl (10g, 38.6548mmol) was dissolved in 30mL THF and slowly added dropwise to the reaction mixture, which was taken out and left to stir at room temperature. After the reaction is finished, weighing 50g of citric acid, dissolving in 450mL of deionized water, adding the reaction solution, adjusting the pH to 3, transferring to a 1L separating funnel, adding EA (300mL multiplied by 3) for extraction, collecting the organic phase, concentrating and evaporating to dryness, dissolving with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (50mL), evaporating to dryness to form a powdery solid, loading by a dry method, carrying out column chromatography, eluting with 2% methanol/dichloromethane solution, collecting and concentrating, and drying in a vacuum oven to obtain a product (7.7g, 86.51%)

45-158

Paclitaxel (10g,11.71mmol, abbreviated as PTX, available from Chongqing orange), imidazole (3.986g,58.5mmol, available from Yinaokai) were weighed into a 500mL reaction flask, dissolved in DMF (100mL), added with tert-butyldimethylsilyl chloride (10.59g,70.26mmol, available from Yinaokai) under nitrogen protection, and stirred at room temperature for reaction. After the reaction is finished, extracting with saturated ammonium chloride solution and dichloromethane, concentrating the organic phase, and evaporating to dryness to obtain 11.3g of product with 100% yield.

45-161

49-17(4.2g,11.9297mmol), 45-158(11g,11.3616mmol) and DMAP (0.2776g,2.2723mmol) were weighed out and put into a 500mL reaction flask, and methylene chloride (50mL) was added to dissolve them, DCC (11.7212mL, 56.808mmol) was added in portions under stirring at-5 ℃ and reacted overnight at-5 ℃. After the reaction is finished, adding a saturated sodium chloride solution and ethyl acetate, separating an organic phase, extracting a water phase for 1 time by using ethyl acetate, combining the organic phases, washing the organic phase for 2 times by using the saturated sodium chloride solution, adding silica gel powder to the organic phase, drying the organic phase by distillation, loading the sample, carrying out column chromatography, and carrying out gradient elution by using 20-25% ethyl acetate/petroleum ether to obtain 8 g of a product, wherein the yield is 54.05%.

45-163

45-161(7.5g,5.7535mmol) was weighed and put into a 500mL reaction flask, DMF was added to dissolve, morpholine (10.024mL, 115.0695mmol) was added, and the reaction was stirred at room temperature for 2 hours. After the reaction is finished, adding a saturated sodium chloride solution and ethyl acetate, separating an organic phase, extracting a water phase for 1 time by using ethyl acetate, combining the organic phases, washing the organic phase for 2 times by using the saturated sodium chloride solution, adding silica gel powder to the organic phase, evaporating to dryness, loading, carrying out column chromatography, and carrying out gradient elution by using 20-25% ethyl acetate/petroleum ether to obtain 4.5g of a product, wherein the yield is 72.58%.

41-123

Taking Fmoc-E (OH)₂(2g, 5.34142mmol from alatin), E (OtBu)2(3.3632g, 11.3698mmol from innochem), HBTU (6.1598g,16.2426mmol) and HOBT (2.1947g, 16.2426mmol) were put into a 250mL reaction flask, dissolved in DMF (20mL), and stirred at 0 ℃ for 30 minutes at a constant temperature, then DIEA (8mL, 48.7277mmol) was slowly added dropwise, and the reaction was continued for 3 hours under this condition. At the end of the reaction, the reaction solution was transferred to a 2L separatory funnel, ethyl acetate (150mL) and deionized water (300mL) were added, the organic phase was separated, and ethyl acetate (300mL) was added to the aqueous phase for extraction several times until the organic phase was free of product. Mixing organic phases, concentrating, adding silica gel powder, evaporating to dryness, and performing column chromatography. Elution with 4% methanol/dichloromethane gave 5g of product, 0.4g of over yield.

41-124

41-123(5g,5.8684mmol) was taken, dissolved in DMF, morpholine (10.2mL, 117.638mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction is finished, adding petroleum ether for settling, discarding supernatant, taking the lower layer, adding pure water, performing suction filtration, collecting solids, mixing, adding DMF for dissolving, adding pure water, performing suction filtration, and drying to obtain a product 3.3g, wherein the yield is 89%.

41-127

Fmoc-Lys (Boc) -OH (2.455g, 5.2399mmol, purchased from alatin), 41-124(3.3g,5.2399mmol), HBTU (2.9528g, 7.8760mmol), HOBT (1.0520g, 7.8760mmol) was taken and added to a 250mL reaction flask, dissolved in DMF (20mL), stirred at 0 ℃ for 30 min at constant temperature, DIEA (3.9mL, 23.5797mmol) was slowly added dropwise, and the reaction was continued for 3 h under this condition. After the reaction, the reaction solution was transferred to a 2L separatory funnel, ethyl acetate (150mL) and deionized water (300mL) were added, the organic phase was separated, and the aqueous phase was extracted several more times with additional ethyl acetate (300mL) until the organic phase was free of product. Mixing organic phases, concentrating, adding silica gel powder, evaporating to dryness, and performing column chromatography. Elution with 50% ethyl acetate/petroleum ether gave 3.4g of product in 77% yield.

41-128

Weighing 45-91(0.6183g,1.1523mmol) and adding into a hydrogenation reaction kettle; DMF (30mL) was added to dissolve, 10% Pd/C (100mg) was added, and DMF (30mL) was added to dissolve the reaction. Introduction of H₂(300psi) and stirred overnight. Obtaining the product. The next reaction was carried out.

41-130

41-127(3.4g,4.0764mmol) was taken, dissolved in DMF, morpholine (7.1mL, 81.5280mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction, the reaction solution was transferred to a 2L separatory funnel, ethyl acetate (150mL) and deionized water (300mL) were added, the organic phase was separated, and the aqueous phase was extracted several more times with additional ethyl acetate (300mL) until the organic phase was free of product. And combining organic phases, and drying to obtain 3.49g of a product.

41-131

41-130(3.5g,4.0764mmol), 41-128(1.523mol), HBTU (1.9664g, 5.1852mmol) and HOBT (0.7006g, 5.1852mmol) are added into a 250mL reaction flask, DMF (20mL) is added for dissolving, the reaction is stirred at the low temperature and the constant temperature of 0 ℃ for 30 minutes, DIEA (2.6mL, 15.5556mmol) is slowly added dropwise, and the reaction is continued to be stirred for 3 hours under the condition. The reaction was complete. The reaction was transferred to a 2L separatory funnel, extracted with ethyl acetate (150mL) and deionized water (300mL), the organic phase was separated, and the aqueous phase was extracted multiple times with additional ethyl acetate (300mL) until the organic phase was free of product. The organic phases are combined, evaporated to dryness and dried, and the next reaction is carried out for 3.5g, and the over yield is 0.3 g.

41-132

41-131(3.2g,1.1523mmol) was taken, dichloromethane (20mL) was measured and added to the reaction flask, and TFA (20mL,276.552mmol) was added. Taking out, stirring and reacting at normal temperature overnight, after the reaction is finished, distilling part of TFA, adding EA (200mL) and n-hexane for settling (200mL), filtering, transferring solid powder to a 2L round-bottom flask, and evaporating to dryness to obtain 2g of a product.

41-133

Adding 41-132(0.28g,0.1545) and M-SCM-5K (3.279g,0.6178mmol, purchased from Keka) into a 250mL reaction bottle, adding DMF (30mL) for dissolving, stirring and reacting for 30 minutes at the low temperature and the constant temperature of 0 ℃, then slowly dropwise adding DIEA (1mL, 6.0172mmol), and stirring and reacting for one week at normal temperature and in the absence of light. After the reaction, methyl tert-butyl ether (200mL) and n-hexane were added for precipitation (50mL), and the product was filtered under suction to obtain 1g of a powder with a yield of 38.46%.

41-134

41-133(0.1064g,0.0062mmol), 45-163(0.1g,0.0925mmol), HBTU (0.0421g, 0.1109mmol) and HOBT (0.0150g, 0.1109mmol) were taken and put into a 100mL reaction flask, DMF (20mL) was added to dissolve the components, the reaction was stirred at 0 ℃ for 30 minutes at a constant temperature, DIEA (0.5mL, 0.3329mmol) was slowly added dropwise, and the reaction was stirred overnight under this condition. And after the reaction is finished, adding methyl tert-butyl ether (200mL) and n-hexane into the reaction solution for settling (50mL), performing suction filtration to obtain a powder product, concentrating, adding silica gel powder, evaporating to dryness, performing column chromatography, and eluting with 4% methanol/dichloromethane to obtain a product with the yield of 0.1g and 55.56%.

41-137

Adding 41-134(0.7g, 0.0234mmol) and TBAF (1.4684g, 5.6160mmol) into a 100mL reaction bottle, adding THF (20mL) for dissolving, stirring at normal temperature for reacting for 3 hours, adding methyl tert-butyl ether (200mL) and n-hexane for settling (50mL) after the reaction is finished, filtering to obtain a powder product, loading the powder product by a dry method, and carrying out column chromatography. Elution with 4% methanol/dichloromethane afforded 0.35g of product, 50% yield.

¹H-NMR(600MHz,DMSO-d₆)δ8.55-8.54(m,8H),7.96-7.94(m,3H),7.82-7.80(m,7H),7.57-7.39(m,21H),7.29-7.24(m,18H),7.22-7.21(m,28H),6.97-6.90(m,48H),6.78-6.61(m,86H),5.77-5.76(m,9H),5.37-5.32(m,3H),5.09–5.04(m,5H),4.52–4.42(m,3H),4.35-4.23(m,9H),3.51-3.49(m,1414H),3.32-3.31(m,41H),3.06-3.01(m,77H),2.74-2.71(m,26H),2.61-2.59(m,14H),2.05–1.96(m,55H),1.74-1.67(m,44H),1.46–1.39(m,76H),1.24-1.21(m,91H),1.16-1.14(m,22H),1.05-1.01(m,112H),0.87-0.79(m,28H).

32. Synthesis of Compounds 49-166 (Compound No. 31)

39-80

Boc-Glu-OH (5.0g, 20.22mmol from alatin), H-Glu (OBzl) -OBzl. TsOH (21.2g, 42.46mmol from Ark Pharm), HOBT (8g,60.66mmol), HBTU (23g,60.66mmol) were weighed out and put into a 250mL reaction flask, dissolved in DMF (80mL), and the reaction was dissolved completely by sonication, stirred at-5 ℃ for 30 minutes, DIEA (30mL, 181mmol) was slowly added dropwise, and the reaction was terminated at low temperature. After the reaction, the reaction mixture was taken out, deionized water (100mL) was added, extraction was carried out with ethyl acetate several times (100 mL. times.3), the organic phases were combined, the organic phase was washed twice with saturated sodium chloride solution (100mL), and finally concentrated to dryness. Loading by dry method, performing column chromatography, gradient eluting with 40% ethyl acetate/petroleum ether-50% ethyl acetate/petroleum ether, collecting product, concentrating, and evaporating to dryness.

39-84

After methylene chloride (5mL) and TFA (14mL, 192mmol) were added to the reaction mixture (39-80) (19.2mmol), the mixture was dissolved completely by sonication, and the reaction mixture was stirred at room temperature with a ground glass stopper. After the reaction was completed, the reaction mixture was taken out, added with a saturated sodium bicarbonate solution (300mL), extracted with ethyl acetate several times (100 mL. times.3), the organic phases were combined, added with a saturated sodium chloride solution (100mL), washed twice, and finally concentrated to dryness.

47-96

7-Ethyl-10-hydroxycamptothecin (15.00g, 38.23mmol, abbreviated as SN38) was placed in a 1000ml round bottom flask, dichloromethane (150ml) was added to dissolve it, tert-butyldiphenylchlorosilane (59.64ml, 229.36mmol, ex Shaoshao) and triethylamine (31.88ml, 229.36mmol) were added and the reaction was stirred overnight in an oil bath at 37 ℃. After the reaction is finished, the reaction solution is steamed to be viscous, normal hexane (150ml) is added to precipitate the product to be solid, and the product is obtained after suction filtration and drying (23.15g, 96%).

47-97

47-96(23.15g,36.70mmol), Boc-Gly-OH (8.71g, 49.70mmol, from alatin), DMAP (0.94g,7.65mmol) were placed in a 1000mL round bottom flask, then dichloromethane (150mL) was added and dissolved, and the mixture was stirred at 0 ℃ for 30 minutes. DCC (39.41g,191.15mmol) was added in three portions, with 30min intervals between each DCC addition, and after completion of the reaction at 0 ℃ for 2 hours, the reaction apparatus was allowed to stand at room temperature and stirred overnight. After the reaction was completed, n-hexane (200mL) and petroleum ether (50mL) were added to the reaction solution to precipitate, and the precipitation was repeated 3 times, and the solid product was obtained by filtration and dried in a vacuum oven to obtain a product (27.53g, 94%).

47-98

47-97(27.53g, 34.50mmol) was placed in a 1000mL round bottom flask, and after adding methylene chloride (50mL) to dissolve it, trifluoroacetic acid (28.40mL, 382.30mmol) was added and the reaction was allowed to proceed at room temperature. After the reaction is finished, taking out the reaction liquid, adding deionized water (200mL), extracting with ethyl acetate (100mL multiplied by 3), combining organic phases, adding saturated sodium chloride solution (200mL multiplied by 2) into the organic phase, cleaning, concentrating, adding silica gel powder, evaporating to dryness to be powdery, carrying out column chromatography, and performing column chromatography by using (1% -3% CH)₃OH, remaining CH₂Cl₂) Gradient elution, collection and concentration gave the product (16.98g, 72%).

29-242

Fmoc-Lys (Boc) -OH (5.0g, 10.6714mmol, available from alatin), H-Gly-OBn (3.7802g, 11.2050mmol, available from innochem), HBTU (6.0705g,16.0072mmol), HOBT (2.1630g, 16.0072mmol) was added to a 500mL flask, dissolved by adding DMF (50mL), and the reaction was stirred at 0 deg.C for 30 min. DIEA (7.9371mL, 48.0215mmol) was added slowly dropwise and the reaction was continued to stir at 0 deg.C overnight. After the reaction is finished, taking out the reaction liquid, adding deionized water (200mL), extracting with ethyl acetate (100mL multiplied by 3), combining organic phases, adding saturated sodium chloride solution (200mL) into the organic phases, washing twice, concentrating, evaporating to dryness, and drying in a vacuum box to obtain a crude product.

29-243

29-242(6.57g, 10.6714mmol) was charged to a 500mL flask, dissolved by addition of appropriate amounts of dichloromethane and TFA (7.9248mL, 106.714mmol), and the reaction was stirred at room temperature overnight. After the reaction is finished, taking out the reaction liquid, evaporating to be oily, adding a saturated sodium bicarbonate solution to adjust the pH to be alkaline, extracting with ethyl acetate (100mL multiplied by 3), combining organic phases, adding a saturated sodium chloride solution (200mL) into the organic phase, washing twice, concentrating, evaporating to dryness, and drying in a vacuum box to obtain a crude product.

29-245

Boc-Lys (Boc) -OH (4.2805g, 11.7385mmol, available from Ark Pharm), 29-243(5.50g, 10.6714mmol), HBTU (6.0705g,16.0072mmol), HOBT (2.1630g, 16.0072mmol) was charged into a 500mL flask, dissolved by adding DMF (50mL), and the reaction was stirred at 0 ℃ for 30 minutes. DIEA (7.9371mL, 48.0215mmol) was added slowly dropwise and the reaction was continued overnight at 0 deg.C. After the reaction is finished, taking out the reaction liquid, adding deionized water (200mL), extracting with ethyl acetate (100mL multiplied by 3), combining organic phases, adding saturated sodium chloride solution (200mL) into the organic phases, washing twice, concentrating, evaporating to dryness, and drying in a vacuum box to obtain a crude product.

29-246

Morpholine (9.24mL, 106.714mmol) and DMF (10mL) were added to a 500mL flask containing 29-245(9.0g, 10.6714mmol) and allowed to dissolve, and the reaction was stirred at room temperature for 1 hour. After the reaction is finished, taking out the reaction liquid, adding deionized water (200mL), extracting with ethyl acetate (100mL multiplied by 3), combining organic phases, adding saturated sodium chloride solution (200mL) into the organic phases, washing twice, concentrating, adding silica gel powder, evaporating to dryness to form powder, carrying out column chromatography, carrying out gradient elution with (1% ammonia water, 4% -5% methanol and the rest dichloromethane), collecting and concentrating to obtain the product 29-246:3.7g, wherein the yield is 56%.

21-221

Weighing erythritol (4.5g,36.849mmol), putting the erythritol into a 500mL two-neck reaction bottle, adding tetrahydrofuran (60mL) for uniform ultrasonic treatment, stirring at 0 ℃, introducing nitrogen for protection, adding potassium tert-butoxide (200mL,184.245mmol), stirring at 0 ℃ for 2 hours, adding benzyl bromoacetate (29.187mL,184.245mmol), stirring for 3 hours, and moving to the normal temperature for reaction. After the reaction is finished, extracting the reaction solution by pure water and ethyl acetate, concentrating the organic phase, loading the sample by a dry method, and carrying out column chromatography. Gradient elution is carried out by using 1% -2% ethyl acetate/petroleum ether to obtain 5g of product with 20% of yield.

38-120

21-221(0.64g,0.9793mmol) and 10% Pd/C (100mg) were charged into a hydrogenation reactor, DMF (30mL) was slowly added and dissolved with stirring, hydrogen (300psi) was introduced, and the mixture was stirred at room temperature overnight. The next day, the reaction solution was filtered with a sand core funnel filled with diatomaceous earth, Pd/C was removed to obtain DMF solution of the product, which was directly used for the next reaction.

29-248

38-120(0.39g, 1.0966mmol), 29-246(3.0g, 4.8249mmol), HBTU (2.4951g,6.5795mmol) and HOBT (0.8891g,6.5795mmol) were charged into a 500mL flask, and DMF (50mL) was added thereto for dissolution, followed by stirring at-5 ℃ for 30 minutes. DIEA (3.2624mL, 19.7384mmol) was added slowly dropwise, stirred at-5 deg.C for 1 hour, and then allowed to stir at room temperature overnight. After the reaction is finished, taking out the reaction liquid, adding deionized water (200mL), extracting with ethyl acetate (100mL multiplied by 3), combining organic phases, adding saturated sodium chloride solution (200mL) into the organic phase, washing twice, concentrating, adding silica gel powder, evaporating to dryness to form powder, performing column chromatography, performing gradient elution with (50% -80% ethyl acetate and the rest petroleum ether), collecting and concentrating to obtain a product (1.6g, 53%).

49-79

29-248(0.52g, 0.1878mmol) and 10% Pd/C (0.0,5g) were put into a hydrogenation reactor, DMF (30mL) was added and dissolved, the hydrogenation reactor was closed, three-pump three-fill was performed, the pressure on the hydrogenation reactor was read at 0.18MPa, and then the reaction was carried out overnight at room temperature. After the reaction, the reaction mixture was filtered through celite, and the filter cake was washed with DMF (20mL x 3) to give a DMF solution of the product as the next starting material.

49-80

49-79(0.45g, 0.1878mmol), M-NH₂HCl-5K (4.91g, 0.9389mmol, Kjeldahl), HBTU (0.43g,1.1237mmol), HOBT (0.15g,1.1267mmol) were charged into a 500mL flask, dissolved by adding DMF (60mL), and the reaction was stirred at-5 ℃ for 30 minutes. DIEA (0.56mL, 3.3780mmol) was slowly added dropwise, stirred at-5 ℃ for 1 hour, and then transferred toStirring at room temperature in low speed and dark for three days. After the reaction is finished, shaking the mixture by using normal hexane (100mL), pouring off supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of normal hexane (10mL) for shaking, pouring off supernatant, repeating the operation for three times, separating out powdery solid in reaction liquid, performing suction filtration, cleaning a filter cake by using methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake by using methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (20g), evaporating the mixture to be powdery, performing dry sampling, performing column chromatography, performing gradient elution by using a dichloromethane mixed solution of 1% ammonia water and 3% -7% methanol, collecting and concentrating, and drying the product in a vacuum oven to obtain the product (3.2g, 73.73%).

49-153

49-80(3.2g, 0.1383mmol) was placed in a 250mL flask, dichloromethane (5mL) and TFA (0.82mL,11.0640mmol) were added to dissolve, and the reaction was stirred overnight at room temperature in the dark at low speed. After the reaction is finished, evaporating the reaction solution to be oily by using a rotary evaporator, adding methyl tert-butyl ether (60mL), precipitating powdery solid in the reaction solution, performing suction filtration, cleaning a filter cake by using methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake by using a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (20g), evaporating to be powdery, performing dry loading, performing column chromatography, performing gradient elution by using a dichloromethane mixed solution of 3% -12% methanol, collecting and concentrating, and drying in a vacuum oven to obtain a product (1.74g, 56.35%).

49-157

39-84(5.16g, 6.7433mmol), mono-tert-butylsuccinic acid (1.40g, 8.0920mmol, available from Accela), HBTU (3.84g,10.1149mmol), HOBT (1.36g, 10.1149mmol) were charged into a 500mL flask, dissolved by adding DMF (50mL), and the reaction was stirred at-5 ℃ for 30 minutes. DIEA (10.03mL, 60.6897mmol) was added slowly dropwise, the reaction stirred at-5 ℃ for 1 hour, then moved to room temperature and stirred overnight. After the reaction is finished, taking out the reaction liquid, adding deionized water (200mL), extracting with ethyl acetate (100mL multiplied by 3), combining organic phases, adding saturated sodium chloride solution (200mL) into the organic phase, washing twice, concentrating, adding silica gel powder, evaporating to dryness to form powder, performing column chromatography, performing gradient elution with (50% -90% ethyl acetate and the rest petroleum ether), collecting and concentrating to obtain a product (5.66g and 90.99%).

49-158

49-157(2.8g, 3.0367mmol) and 10% Pd/c (0.08g) were charged into a hydrogenation reactor, DMF (30mL) was added and dissolved, the hydrogenation reactor was closed, three-pump three-fill was performed, the pressure on the hydrogenation reactor was read at 0.18MPa, and the reaction was allowed to proceed overnight at ambient temperature. After the reaction, the reaction mixture was filtered through celite, and the filter cake was washed with DMF (20mL x 3) to give a DMF solution of the product as the next starting material.

49-159

49-158(1.71g, 3.0367mmol), 47-98(8.77g, 12.7541mmol), HBTU (6.90g,18.2202mmol) and HOBT (2.46g,18.2202mmol) were charged into a 500mL flask, dissolved by adding DMF (60mL), and the reaction was stirred at-5 ℃ for 30 minutes. DIEA (13mL, 78.9542mmol) was added slowly dropwise, the reaction stirred at-5 ℃ for 1 hour, then transferred to room temperature and stirred overnight. After the reaction is finished, shaking with n-hexane (100mL), pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL), shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (60g), evaporating to be powdery, performing dry sampling, performing column chromatography, performing gradient elution with 3% -7% methanol dichloromethane mixed solution, collecting, concentrating, and drying in a vacuum oven to obtain the product (11.3g, over-yield).

49-161

49-159(9.84g, 3.0367mmol) was charged to a 250mL flask, dichloromethane (8mL) and TFA (8mL) were added to dissolve, and the reaction was stirred at room temperature overnight. After the reaction is finished, evaporating the reaction solution to be oily by using a rotary evaporator, adding methyl tert-butyl ether (60mL), precipitating powdery solid in the reaction solution, performing suction filtration, cleaning a filter cake by using methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake by using a methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (60g), evaporating the mixture to be powdery, performing dry loading, performing column chromatography, performing gradient elution by using a 1-4% methanol dichloromethane mixed solution, collecting and concentrating, and drying the product in a vacuum oven to obtain a product (2.7g, 27.92%).

49-162

49-153(1.19g, 0.0535mmol), 49-161(1.5g, 0.4710mmol), HBTU (0.24g,0.6422mmol), HOBT (0.08g,0.6422mmol) were charged into a 500mL flask, dissolved by adding DMF (60mL), and the reaction was stirred at-5 ℃ for 30 minutes. DIEA (0.60mL, 3.6395mmol) was slowly added dropwise, stirred at-5 ℃ for 1 hour, and then transferred to room temperature and stirred overnight in the dark at low speed. After the reaction is finished, adding n-hexane (100mL) for shaking, pouring off the supernatant, repeating the operation for three times, adding methyl tert-butyl ether (80mL) and a small amount of n-hexane (10mL) for shaking, pouring off the supernatant, repeating the operation for three times, separating out powdery solid in the reaction solution, performing suction filtration, cleaning the filter cake with methyl tert-butyl ether (40mL x 3), collecting the filter cake, dissolving the filter cake with methanol/dichloromethane (1:4) solution (100mL), adding silica gel powder (20g), evaporating the silica gel powder to be powdery, performing dry sampling, performing column chromatography, performing gradient elution with 3% -15% methanol dichloromethane mixed solution, collecting, concentrating, and drying in a vacuum oven to obtain the product (0.95g, 37.25%).

49-166

THF (10ml) and dilute hydrochloric acid (10ml, 0.05mmol/L) were added to a 500ml flask containing 49-162(0.95g, 0.0199mmol), dissolved by sonication, TBAF (0.5g,1.9133mmol) was added thereto, and the reaction was stirred at room temperature for 3 hours with exclusion of light. After the reaction is finished, evaporating the reaction solution to dryness, dissolving the reaction solution by using DMF (5ml), then using isopropanol for sedimentation, and repeating the operation for three times; and then dissolved in absolute ethanol and a little dichloromethane, and then the solution is added with methyl tert-butyl ether for sedimentation, and the process is repeated three times, and the solid is collected and dried in a vacuum box to obtain the product (0.75g, 93.75%).

¹H NMR(600MHz,DMSO-d6)δ8.36-8.18(m,32H),8.12-7.89(m,56H),7.81-7.76(m,16H),7.35(s,100H),7.05-6.96(m,27H),5.95-5.18(m,124H),4.25-3.81(m,156H),3.53-3.49(m,3706H),3.12-2.78(m,123H),2.08(s,109H),1.47-1.20(m,211H),0.99-0.78(m,136H).

Example 2 Activity test

Hereinafter, the test methods for other compounds are the same as those for 27-134 (compound No. 18) and 44-2 (compound No. 3), and the inhibitory effect on cancer cells is also significant.

Firstly, experimental materials:

1. customer samples: colo205, MDA-MB-231 cancer cell, 2 kinds of medicine (27-134, 44-2)

2. Experimental equipment and reagents: PerkinElmer multifunctional microplate reader, CCK8 cell proliferation and cytotoxicity detection kit

Second, Experimental methods

The preparation of the medicine comprises the following steps: the drug was prepared in 0.1g/ml physiological saline corresponding to 7.4537mM of the molar concentration of each drug.

1. Culturing cells, and digesting with pancreatin. Cells were blown into single cell suspensions with the corresponding medium. 100ul of the cells are inoculated in a 96-well plate per well, and the cell density is about 10-15%. 37 ℃ and 5% CO ₂Culture boxThe culture is continued for 6 hours until the cells are treated after the adherence.

2. According to the requirement of the drug concentration, the multiple dilution method is used for preparing drug-containing culture media with different concentrations. Two different sets of gradient combinations of the same drug were screened on one plate as shown in figure 1.

3. Drug gradient: 100ul of the fresh culture medium containing the corresponding drug prepared in step 2 (total volume 200ul, drug concentration as above table) was added, and the culture was continued for 48 hours.

4. After 48 hours of drug treatment, the original medium was discarded.

5. Add 100. mu.l (10% CCK8) of medium to each well and incubate for an additional 2 hours in the cell culture chamber.

6. The absorbance at 450nm was measured, and the results are shown in FIG. 2.

Thirdly, analyzing an experimental result:

1. the cell inhibition rate at each drug concentration was calculated from the readings as follows:

2. drawing:

drug 44-2 had inhibitory effects on MDA-MB-231 cancer cells, as shown in FIG. 3.

The inhibiting effect of the medicines 27-134 on Colo205 cancer cells is shown in figure 4.

3. Calculating individual drug IC₅₀

The results are shown in FIG. 5.

EXAMPLE 3 in vivo efficacy prediction of anti-tumor Agents against human Breast cancer BT474 cell NPG mouse subcutaneous transplantation tumor model with multiple test samples

1. Experimental materials:

the preparation method comprises the following steps:

test sample diluent: taking a proper amount of PEG300 and absolute alcohol, and adding sodium chloride injection to ensure that the content of the PEG300 and the absolute alcohol is 20% (v/v).

And (3) testing the sample: respectively measuring a proper volume of 41-32, 41-40 and 39-55, adding a proper amount of test sample diluent, and respectively preparing solutions with the concentrations of 8.3mg/mL, 4.6mg/mL and 5.5 mg/mL.

SB7. HCL: weighing an appropriate amount of SB7.HCL (reduced coefficient of 93.4%), adding a certain volume of ethanol (5%, V/V) for dissolving, adding an appropriate amount of sodium chloride injection after complete dissolution, and preparing into a solution with the concentration of 0.5 mg/mL.

LPT: weighing a proper amount of LPT, adding a certain volume of 0.5% CMC-Na solution, and uniformly stirring on a magnetic stirrer to prepare a solution with the concentration of 0.56 mg/mL.

Negative control: the test sample diluent was used directly.

Preparing a solution, preserving and treating: the prepared test sample and the prepared reference sample are stored at the temperature of 2-8 ℃ or in an ice box before administration, and the rest of the test sample and the reference sample after administration are treated according to medical waste.

Human breast cancer cells BT 474: is derived from cell resource center of Shanghai Life sciences institute of Chinese academy of sciences, and is cultured under RPMI1640+ 10% FBS, 37 deg.C and 5% CO₂And (5) culturing.

Animal species & strain: NPG mice

Animal grade: SPF stage

The experimental animal source is as follows: beijing Wintoda Biotechnology Ltd

Week age at tumor inoculation: about 4-5 weeks.

Body weight at tumor inoculation: about 15-18 g. The body weight of animals of the same sex is between 80% and 120% of the average body weight.

Sex and quantity: female, 60 mice, model animals 36 were screened for final experiments, and the remaining animals were either handed over to veterinarian or euthanasia.

Animals are raised in an independent ventilation system (IVC), 6 animals in the same group and an SPF animal house are raised in each cage at most, the environmental conditions are controlled to be 20-26 ℃ at room temperature, the relative humidity is 40-70%, and the light and the shade alternate within 12 hours of illumination.

Feeding: qualified rat feed (production unit: Australian cooperative feed Co., Ltd., Beijing, family) was provided daily and animals were allowed free access to food. Animals had free access to water.

2. The experimental method comprises the following steps:

resuscitating BT474 cellsAnd (4) carrying out cell passage amplification, collecting cells in a logarithmic growth phase when the cells are amplified to a sufficient cell number, and carrying out cell inoculation. Estrogen-sustained release tablets (17 β -estrogen, cat # SE-121, Innovative Research of America) were implanted subcutaneously in the neck backs of NPG mice prior to cell inoculation. The concentration is 7.5 multiplied by 10⁷BT474 cell suspension/mL was mixed with Matrigel (Matrix base Membrane Matrix, BD Co.) at a volume ratio of 1:1 to give a concentration of 3.75X 10⁷one/mL cell suspension, 0.2mL was inoculated into the right breast pad of the mouse. Observing the growth condition of the tumor after inoculation, and finally screening to obtain the tumor with the volume of 66.83-324.35 mm ³36 of the above were used for the test.

The tumor volumes and body weights were randomly divided into 6 groups, which were: group 1 (negative control group, test sample diluent), group 2 (LPT, 20mg/kg), group 3 (SB7+ LPT, 5+5.6mg/kg), group 4 (41-32 i.e. compound No. 20, 83mg/kg), group 5 (41-40 i.e. compound No. 13, 46mg/kg), group 6 (39-55 i.e. compound No. 12, 55mg/kg), and 6 subjects per group at a dose capacity of 10 mL/kg. Negative control, SB7, 41-32, 41-40, 39-55 intravenous injection; and (4) performing LPT intragastric administration. Group 2 and 4 were administered 1 time per 3 days (D19 was stopped once due to insufficient drug), and the remaining groups were each administered 1 time/week for 4 consecutive weeks, and D29 animals were euthanized. General clinical symptoms of animals were observed 2 times a day during the trial, 2 weight and tumor size measurements were taken weekly, tumors were removed after euthanasia, and tumor weights were weighed. Calculating tumor volume, relative tumor volume RTV, relative tumor proliferation rate T/C%, and tumor weight inhibition rate IR_TW% of the total weight of the composition. The relative tumor proliferation rate T/C% of the administration group is less than or equal to 40%, and the P of the RTV of the administration group is less than or equal to 0.05 compared with the RTV of the negative control group, and IR is used for treating_TWMore than or equal to 60 percent is an auxiliary reference index of effectiveness.

A. Measurement of tumor diameter:

and (3) detecting animals: all animals

Detection time: grouping (i.e. D1 on the day of first administration), 2 times per week after first administration, before euthanasia, slide caliper measurements and recording tumor length and length, and tumor volume calculation.

Tumor volume was calculated according to the following formula:

v is 1/2X long diameter X short diameter²

B. Evaluation of efficacy based on tumor volume

Relative Tumor Volume (RTV) and relative tumor proliferation rate T/C% were calculated according to the following formula:

RTV＝V_t/V₀

V_t: tumor volume obtained by daily measurement of tumors

V₀: initial tumor volume (before administration)

T/C% -mean RTV of the administered group/mean RTV of the control group X100%

If the T/C% is less than or equal to 40%, and the statistical treatment P of the experimental group RTV and the model group RTV is less than 0.05, the tumor growth inhibition effect is achieved; on the other hand, if the T/C% is more than 40%, the tumor growth is not inhibited.

C. Evaluation of therapeutic effects based on tumor weight

After the experiment, tumor nodules were detached and weighed, and the difference in tumor weight between groups was compared to further calculate the tumor inhibition rate IR_TWBy IR_TW>60% is an effectiveness reference index, and the calculation formula is as follows:

IR_TW(％)＝(W_{model set}-W_{Administration set})/W_{Model set}×100％

3. The experimental results are as follows:

throughout the experiment, 1 animal in group 2D 19 died; in group 3, 2 animals died at D15 and D25, respectively, of which 1 had been dissected and observed to have a bladder full of white sand-like particles, the cause of death being considered to be related to the inoculation of estrogen tablets; the cause of death in the other 2 animals was presumed to be related to control toxicity. The weight of the animal bodies in 1-6 groups gradually increases, and no significant difference (P >0.05) is found among the groups.

The negative control group (group 1) showed a gradual tumor growth throughout the experiment, and by the end of the experiment (D29), the mean tumor volume of group 1 was 1938.55. + -. 511.20mm³The average RTV is 14.98 +/-8.63; the mean tumor volumes of the 2-6 groups were 1313.80 + -241.46 mm, respectively³、1463.75±1088.95mm³、1232.77±652.36mm³、1616.52±985.05mm³、346.96±203.35mm³The average RTV is respectively 8.13 +/-2.92, 8.80 +/-3.57, 8.27 +/-3.92, 11.61 +/-8.28 and 2.26 +/-1.32, and the tumor volume is obviously lower than that of the group 1 (P is less than or equal to 0.05) when the groups 6D 15-D29 are adopted; the RTV of group 6D 12 is significantly lower than that of group 1 (P is less than or equal to 0.05), and no significant difference is found between tumor volume and RTV among other groups (P is less than or equal to 0.05)>0.05)。

The tumor growth trend of each group is shown in fig. 6, fig. 7 and fig. 8.

By the end of the experiment (D29), the T/C% values of groups 2-6 were 54.27%, 58.74%, 55.21%, 77.54%, 15.07%, respectively, and their IR values were_TV% 45.73%, 41.26%, 44.79%, 22.46%, 84.93%, respectively. The T/C% of the group 6 is reduced to 40% or less at D12-D29.

At the end of the test, the tumor weight of the animals is weighed after euthanasia, and the average tumor weight of 1-6 groups is 1.807 + -0.350 g, 1.226 + -0.370 g, 1.185 + -0.934 g, 0.899 + -0.428 g, 1.287 + -0.746 g and 0.266 + -0.167 g. Group 6 was significantly lower than group 1 (P.ltoreq.0.05). 2 to 6 groups of IR_TW% 32.15%, 34.42%, 50.25%, 28.78%, 85.28%, respectively.

The tumor weight inhibition ratios of the groups are shown in fig. 9, fig. 10 and fig. 11. Fig. 12 is a photograph of groups of euthanized animals. Fig. 13 is a photograph of each group of tumors.

And (4) conclusion: under the experimental condition, each administration group has tumor growth inhibition effects of different degrees on NPG mouse subcutaneous transplantation tumor models of human breast cancer BT474 cells. The test article 39-55 has obvious inhibiting effect on the tumor growth of NPG mouse subcutaneous transplantation tumor model of human breast cancer BT474 cells by tail vein injection administration at 55mg/kg dose, and the effect is superior to that of SB7 by tail vein injection at 5mg/kg dose and LPT by intragastric administration at 5.6mg/kg dose. The LPT has certain inhibition effect on the tumor growth of NPG mouse subcutaneous transplantation tumor models of human breast cancer BT474 cells by intragastric administration at the dose of 20mg/kg, intravenous administration at the dose of 83mg/kg by 41-32 and intravenous administration at the dose of 46mg/kg by 41-40.

Example 4 in vivo Pre-test of the Effect of the test article on antitumor Activity of human Colon cancer COLO-205 cells in vivo in nude mouse model of subcutaneous transplantation tumor

1. Experimental Material

Test article

Name: 39-17 (i.e., compound No. 14); concentration: 10 mg/mL.

Vehicle/negative control

Name: sodium chloride injection; batch number: 4B 19091206; specification: 0.9g in 100 mL; the characteristics are as follows: a colorless clear liquid; storage conditions are as follows: sealing and storing; production unit: shandong Qi is a pharmaceutical company, Inc.

Preparing a test sample and a reference sample:

the preparation method comprises the following steps:

and (3) testing the sample: can be used directly without dilution.

SB7. HCL: weighing an appropriate amount of SB7.HCL (reduced coefficient of 93.4%), adding a certain volume of ethanol (5%, V/V) for dissolving, adding an appropriate amount of sodium chloride injection after complete dissolution, and preparing into a solution with the concentration of 0.3 mg/mL.

PCB: weighing a proper amount of PCB, adding a certain volume of 0.5% CMC-Na solution, and uniformly stirring on a magnetic stirrer to prepare a solution with the concentration of 1.04 mg/mL.

Negative control: sodium chloride injection is directly used.

Preparing a solution, preserving and treating: the prepared test sample and the prepared reference sample are stored at the temperature of 2-8 ℃ or in an ice box before administration, and the rest of the test sample and the reference sample after administration are treated according to medical waste.

Human colon cancer cell COLO-205: derived from cell resource center of foundation institute of Chinese medical science and sciences, and cultured under RPMI1640+ 10% FBS, 37 deg.C, and 5% CO₂。

Animal species & strain: BALB/c nude mice

Animal grade: SPF stage

The experimental animal source is as follows: beijing Wittiulihua laboratory animal technology Co Ltd

Week age at tumor inoculation: about 4-5 weeks.

Body weight at tumor inoculation: about 15-18 g. The body weight of animals of the same sex is between 80% and 120% of the average body weight.

Sex and quantity: male, 25 mice, 18 model animals were screened for the final experiment, and the remaining animals were kept on stock or euthanized.

Animals are raised in an independent ventilation system (IVC), 6 animals in the same group and an SPF animal house are raised in each cage at most, the environmental conditions are controlled to be 20-26 ℃ at room temperature, the relative humidity is 40-70%, and the light and the shade alternate within 12 hours of illumination.

Feeding: qualified rat feed (production unit: Australian cooperative feed Co., Ltd., Beijing, family) was provided daily and animals were allowed free access to food. Animals had free access to water.

2. The experimental method comprises the following steps:

recovering COLO-205 cells, performing cell passage expansion, collecting cells in logarithmic growth phase when the cells are expanded to a sufficient cell number, and preparing for cell inoculation. Adjusting the cell concentration to 5 × 10 according to the actual cell number⁷one/mL, at 0.2 mL/mouse inoculated in 25 mice right side axillary subcutaneous. Observing the growth condition of the tumor after inoculation, and waiting for the tumor volume to be about 100-300 mm³In the process, screening is carried out according to the size of tumor volume, and the patient with the excessive tumor volume and without tumor formation is not selected, and finally 18 tumor-forming animals are obtained for testing through screening.

18 qualified tumor-bearing mice are screened, and are randomly divided into 3 groups, namely: 1 group (negative control group, sodium chloride injection), 2 groups (SB7+ PCB, 3+10.4mg/kg) and 3 groups (39-17, 153mg/kg), wherein 6 of each group had a negative control, SB7 and PCB administration volume of 10mL/kg and a 39-17 administration volume of 15.3 mL/kg. Negative control, SB7, 39-17 intravenous injection, PCB gavage, 1 time per week for 3 weeks, D22 animals were euthanized. General clinical symptoms of animals were observed 2 times a day during the trial, 2 weight and tumor size measurements were taken weekly, tumors were removed after euthanasia, and tumor weights were weighed. Calculating tumor volume, relative tumor volume RTV, relative tumor proliferation rate T/C%, and tumor weight inhibition rate IR _TW% of the total weight of the composition. The relative tumor proliferation rate T/C% of the administration group is less than or equal to 40%, and the P of the RTV of the administration group is less than or equal to 0.05 compared with the RTV of the negative control group, and IR is used for treating_TWMore than or equal to 60 percent is an effectiveness reference index.

A. Measurement of tumor diameter

And (3) detecting animals: all animals

Detection time: grouping (i.e. D1 on the day of first administration), 2 times per week after first administration, before euthanasia, slide caliper measurements and recording tumor length and length, and tumor volume calculation.

Tumor volume was calculated according to the following formula:

v is 1/2X long diameter X short diameter²

B. Evaluation of efficacy based on tumor volume

Relative Tumor Volume (RTV) and relative tumor proliferation rate T/C% were calculated according to the following formula:

RTV＝V_t/V₀

V_t: tumor volume obtained by daily measurement of tumors

V₀: initial tumor volume (before administration)

T/C% -mean RTV of the administered group/mean RTV of the control group X100%

If the T/C% is less than or equal to 40%, and the statistical treatment P of the experimental group RTV and the model group RTV is less than 0.05, the tumor growth inhibition effect is achieved; on the other hand, if the T/C% is more than 40%, the tumor growth is not inhibited.

C. Evaluation of therapeutic effects based on tumor weight

After the experiment, tumor nodules were detached and weighed, and the difference in tumor weight between groups was compared to further calculate the tumor inhibition rate IR _TWBy IR_TW>60% is an effectiveness reference index, and the calculation formula is as follows:

IR_TW(％)＝(W_{model set}-W_{Administration set})/W_{Model set}×100％

3. The experimental results are as follows:

during the whole experiment, 1 animal in the 3D 8 group died, weighed 14.5g after death, and lost much before death, which is presumed to be related to 39-17 toxicity. In group 1, 2 animals died at D19 and D20, respectively, with tumor ulceration observed before 1 death, presumably associated with greater tumor burden. The weight of the animals in the 1 group gradually decreases, the weight of the animals in the 2 groups slightly increases and then decreases, the weight of the animals in the 3 groups gradually increases, and the weight of the animals in the 3 groups is obviously higher than that in the 1 group (P is less than or equal to 0.05) from D8 to D22; the ratio of D12-D22 is significantly higher than that of group 2 (P is less than or equal to 0.05).

The tumors grew gradually throughout the experiment in the negative control group (group 1), and by the end of the experiment (D22), the mean tumor volume in group 1 was 1900.58. + -. 489.19mm³The average RTV is 16.03 +/-6.43; the mean tumor volumes of the 2-3 groups were 774.06. + -. 228.43mm, respectively³、33.83±37.01mm³The average RTV is respectively 6.38 +/-1.81 and 0.24 +/-0.21, the tumor volume is obviously lower than that of the group 1 (P is less than or equal to 0.05) when the groups 2D 5-D22 are arranged, and the tumor volume is obviously lower than that of the groups 1 and 2 (P is less than or equal to 0.05) when the groups 3D 5-D22 are arranged; the RTV is significantly lower than that in the 1 st group (P is less than or equal to 0.05) in the 2 nd groups D5-D19, and the tumor volume is significantly lower than that in the 1 st and 2 nd groups (P is less than or equal to 0.05) in the 3 rd groups D5-D22.

The tumor growth trend for each group is shown in fig. 14.

By the end of the experiment (D22), the T/C% values of 2-3 groups were 59.68% and 1.88%, respectively, and their IR values were_TV% is 40.32%, 98.12%, respectively. The T/C% of the group 3 is reduced to below 40% at D5-D22 and is significantly lower than that of the group 1 (P ≦ 0.05).

At the end of the test, the tumor weights of the animals were weighed after euthanasia, and the average tumor weights of 1-3 groups were 1.551 + -0.290 g, 0.607 + -0.173 g and 0.036 + -0.036 g, respectively. Groups 2 and 3 are significantly lower than group 1 (P.ltoreq.0.05). 2 to 3 groups of IR_TW% is 60.86%, 97.68%, respectively.

The tumor weight inhibition rate of each group is shown in FIG. 15.

And (4) conclusion: under the experimental condition, the tail vein injection administration of the test sample 39-17 at the dose of 153mg/kg has obvious tumor growth inhibition effect on human colon cancer cell COLO-205 subcutaneous transplanted tumor models, the tail vein injection administration of SB7 at the dose of 3mg/kg and the intragastric lavage combined administration of PCB at the dose of 10.4mg/kg have obvious growth inhibition effect on tumor models, and the 39-17 effect is obviously better than the combination of PCB and SB 7.

While specific embodiments of the invention have been described in detail, those skilled in the art will understand that: various modifications and changes in detail can be made in light of the overall teachings of the disclosure, and such changes are intended to be within the scope of the present invention. The full scope of the invention is given by the appended claims and any equivalents thereof.