8-azacyclo-substituted chromone derivative and preparation method and pharmaceutical application thereof
阅读说明：本技术 8-氮杂环取代色酮类衍生物及其制备方法与制药用途 (8-azacyclo-substituted chromone derivative and preparation method and pharmaceutical application thereof ) 是由 汪豪 龙桓 胡晓龙 熊非 叶文才 于 2021-07-23 设计创作，主要内容包括：本发明公开了一种8-氮杂环取代色酮类衍生物及其制备方法与制药用途,属于药物化学领域。本发明提供了一种具有选择性PARP1/2抑制活性的8-氮杂环取代色酮类化合物,可用于制备预防或治疗PARP1/2缺陷型肿瘤的药物,具有广泛的市场应用和开发前景。(The invention discloses an 8-azacyclo-substituted chromone derivative, a preparation method and pharmaceutical application thereof, belonging to the field of pharmaceutical chemistry. The invention provides an 8-azacyclo-substituted chromone compound with selective PARP1/2 inhibitory activity, which can be used for preparing a medicament for preventing or treating PARP1/2 defective tumors and has wide market application and development prospect.)
1. An 8-azacyclo-substituted chromone derivative shown in formula 1, or a pharmaceutically acceptable salt, ester or solvate thereof:
wherein R is1Selected from: H. OH, N3、F、C1-C8Alkyl, carboxyl, C1-C8Alkoxy radical, C2-C8Alkenyl radical, C2-C8Alkynyl, C1-C8Alkylcarbonyl group, C1-C8Alkoxycarbonyl group, C1-C8Alkylsulfonyl radical, C1-C8Alkylsulfinyl radical, C1-C8Alkylthio radical, C1-C5Alkylcarbonylamino, C1-C8Alkylaminocarbonyl, di (C)1-C3) Alkylaminocarbonyl radical, C3-C8Cycloalkyl radical, C3-C8Cycloalkyl oxy, C3-C8Heterocycloalkyl radical, C3-C8Heterocycloalkyloxy, phenyl, phenyloxy, phenyl (C)1-C2) Alkyl, phenyl (C)1-C2) Alkoxy, phenylsulfonyl, phenylsulfinyl, C5-C6Heteroaryl group, C5-C6Heteroaryloxy radical, C5-C6Heteroaryl (C)1-C3) Alkyl radical, C5-C6Heteroaryl (C)1-C3) Alkoxy radical, C3-C8Cycloalkyl (C)1-C2) Alkyl radical, C3-C8Cycloalkyl (C)1-C2) Alkoxy radical, C3-C8Heterocycloalkyl (C)1-C2) Alkyl, heterocycloalkyl (C)1-C2) Alkyloxy, condensed ring aryl or substituted condensed ring aryl, wherein R1May be unsubstituted or substituted with one or more substituents selected from halogen, hydroxy, cyano, unsubstituted or halogenated C1-C8Alkyl and unsubstituted or halogenated C1-C8An alkoxy group;
R2selected from: H. f, Cl, Br, I, CN, NO2、NH2、OH、N3Open chain C2-C8Saturated or unsaturated alkyl, substituted C3-C8Cycloalkyl, substituted C3-C8Heterocycloalkyl radical, C1-C9Alkenyl radical, C1-C9Alkyl radical, wherein R2May be unsubstituted or substituted with one or more substituents selected from halogen, hydroxy, cyano, unsubstituted or halogenated C1-C8Alkyl and unsubstituted or halogenated C1-C8An alkoxy group;
R3、R4、R5independently selected from: H. f, Cl, Br, I, CN, NO2、NH2、OH、N3、Me、C1-C8Alkoxy radical, C1-C8Alkylthio radical, C1-C5Alkylcarbonylamino, C1-C8An alkoxycarbonyl group;
R6selected from: open chain C2-C8Saturated or unsaturated alkyl, substituted C3-C8Cycloalkyl, substituted C3-C8Heterocycloalkyl, substituted phenyl, heteroaryl, substituted heteroaryl, fused ring aryl, or substituted fused ring aryl, wherein said substituted phenyl, substituted heteroaryl, or substituted fused ring aryl may independently be substituted with 1 to 3 of the following substituents: H. f, Cl, Br, I, CN, NO2、NH2、N3、OH、-S(O)2NH2、-S(O)2NHCOCH3、C1-C6Alkyl radical, C1-C6Alkoxy radical, C2-C6Alkenyl radical, C2-C6Alkynyl, 1H-tetrazol-5-yl, 3H- [1,3,4 ]]Oxadiazol-2-one-5-yl, 3H- [1,3,4 ]]Oxa-diAzole-2-thione-5-yl, 4H- [1,2,4 [ ]]Oxadiazol-5-one-3-yl, 4H- [1,2,4 ]]Oxadiazole-5-thione-3-yl, 3H- [1,2,3,5]Oxathiadiazol-2-oxide-4-yl, wherein each alkyl or alkoxy group may be unsubstituted or substituted by one or more substituents selected from halogen and unsubstituted or fluorinated C1-C3Alkyl or C1-C3An alkoxy group;
a is selected from: furan, thiophene, piperidine, pyridine, pyrrole, imidazole, thiazole, oxazole, piperazine, tetrahydrofuran, tetrahydrothiophene, tetrahydropyridine, imidazoline, thiazoline, oxazoline, C1-C6Triazole, C1-C6Triazole, pyrazine, pyridazine, the said heterocycles being independently substituted by 2-4 carbon atoms1-C3Alkyl, halogen, haloalkyl, cycloalkyl, halocycloalkyl, hydroxyalkyl or alkoxyalkyl;
x is selected from: -S (O)2-, -C (O) -or- (CH)2) n-; n is 0, 1 or 2;
y is selected from: 0 to 1N or C;
wherein m is 0 or 1.
2. The 8-azacyclo-substituted chromone derivative or the pharmaceutically acceptable salt, ester or solvate thereof according to claim 1, wherein:
R1selected from: H. c1-C8Alkyl radical, C3-C8Cycloalkyl radical, C3-C8Heterocycloalkyl, phenyl, substituted phenyl, heteroaryl, substituted heteroaryl, wherein said substituted phenyl or substituted heteroaryl may independently be substituted with 1 to 3 substituents as follows: F. cl, Br, I, CN, NO2、NH2、OH、C1-C3An alkyl group;
R2selected from: H. f, Cl, Br, I, CN, NO2、NH2、OH、N3Substituted C3-C8A heterocycloalkyl group;
R3、R4、R5independently selected from: H. c1-C8Alkoxy radical, C1-C8Alkylthio radical, C1-C5Alkylcarbonylamino, C1-C8An alkoxycarbonyl group;
R6selected from: open chain C2-C8Saturated or unsaturated alkyl, substituted C3-C8Cycloalkyl, substituted C3-C8Heterocycloalkyl, substituted phenyl, heteroaryl, substituted heteroaryl, fused ring aryl, or substituted fused ring aryl, wherein said substituted phenyl, substituted heteroaryl, or substituted fused ring aryl may independently be substituted with 1 to 3 of the following substituents: H. f, Cl, Br, I, CN, NO2、NH2、N3；
A is selected from: piperidine, pyridine, pyrrole, imidazole, thiazole, oxazole, piperazine, tetrahydropyridine, imidazoline, thiazoline, oxazoline, optionally substituted with 2 to 4 carbon atoms1-C3Alkyl, halogen, haloalkyl, cycloalkyl, halocycloalkyl, hydroxyalkyl or alkoxyalkyl;
x is-C (O) -or- (CH)2)n-；n＝1；
Y is selected from: 0 to 1N or C;
wherein m is 0 or 1.
3. The 8-azacyclo-substituted chromone derivative or the pharmaceutically acceptable salt, ester or solvate thereof according to claim 1 or 2, wherein the derivative is selected from the following compounds 1-90:
。
4. a process for the preparation of a compound of formula (I) according to any one of claims 1 to 3, characterized in that the reaction scheme is selected from any one of the following:
the method comprises the following steps:
the second method comprises the following steps:
5. a pharmaceutical composition comprising a compound according to any one of claims 1 to 3, or a pharmaceutically acceptable salt, ester or solvate thereof, as an active ingredient, and a pharmaceutically acceptable excipient.
6. The pharmaceutical composition of claim 5, wherein the pharmaceutical composition is in the form of a capsule, powder, tablet, granule, pill, injection, syrup, oral liquid, inhalant, cream, ointment, suppository, or patch.
7. Use of the derivative of any one of claims 1-3 or a pharmaceutically acceptable salt, ester or solvate thereof, and the pharmaceutical composition of claim 5 for preparing a medicament for treating tumor and other PARP related diseases.
8. The use of claim 7 wherein said other PARP related disease is stroke, myocardial infarction, inflammation, hypertension, atherosclerosis or diabetes.
9. The use of claim 7, wherein the tumor is a PARP-1/2 mediated tumor deficient in the BRCA1 gene.
10. The use of claim 7, wherein the tumor is ovarian cancer, breast cancer and fallopian tube cancer, malignant peritoneal tumor, metastatic breast cancer, ovarian epithelial cancer, primary peritoneal cancer, advanced malignant germline or somatic BRCA-mutated advanced epithelial ovarian cancer, advanced malignant germline or somatic BRCA-mutated fallopian tube cancer, advanced malignant germline or somatic BRCA-mutated peritoneal cancer refractory advanced ovarian cancer.
Technical Field
The invention relates to a pharmaceutical chemistry technology, in particular to 8-azacyclo-substituted chromone derivatives, a preparation method and pharmaceutical application thereof.
Background
Poly (adenosine diphosphate ribose) polymerase-1 (PARP-1) is a cellular ribozyme that catalyzes poly (nuclear glycation) in eukaryotic cells and plays an important role in mediating base excision repair pathways, repairing intracellular abnormal DNA damage, and maintaining genome integrity. The repair mechanism of PARP-1 to DNA single-strand damage is well studied at present: when DNA is damaged by breakage, PARP-1 can rapidly recognize and combine with a damage gap to activate a poly adenosine diphosphate ribose (PAR) synthesis pathway so as to repair the damaged DNA. (Science 2016,353,45-50) several studies have shown that inhibition of PARP-1 reduces DNA repair and enhances the effectiveness of chemotherapeutic treatments on a variety of tumors. Meanwhile, the system for repairing abnormal DNA in cells is not unique, the activity of PARP-1 in cells is removed or inhibited under normal conditions, and although single-stranded DNA in cells can be accumulated in a broken way to a certain extent, another repair system in cells, namely a Homologous Recombination (HR) path mediated by breast cancer susceptibility genes BRCA-1 and BRCA-2, can repair double-stranded DNA damage and maintain the stability of chromosomes. (Nature reviews, molecular cell biology 2017,18,610-621) breast cancer is the most common cancer among female patients, the mortality rate of the patients is extremely high, and the BRCA-1 gene-related breast cancer patients account for about 60-80% of the causes of breast malignant tumors. In the cells for repairing abnormal DNA, breast cancer and ovarian cancer cells with defects or mutations of the cancer suppressor gene BRCA-1/2 are sensitive to PARP-1 inhibitors by 1000 times of normal cells, the PARP-1 inhibition activity can target and kill tumor cells, (Nature 2005,434,917-21; European journal of clinical chemistry 2018,145, 389-one 403) to cause synthetic lethal, and the synthetic lethal effect between PARP-1 and BRCA-1/2 is verified by clinical tests. In summary, PARP-1 inhibitors are key points in the treatment of tumor-associated diseases, especially for BRCA-1 deficient tumors.
The PARP-1 inhibitors Olaparib (Olaparib) and Rucapenberg camphorsulfonate (Rucaparib) which are on the market at present have good effects whether being used in combination with chemotherapy or being used for BRCA-deficient breast cancer and ovarian cancer independently. Although PARP-1 inhibitors are of great interest in tumor therapy and chemosensitization, there are some deficiencies (Precision clinical medicine 2020,3,187-201) such as: 1) the selectivity is poor, most PARP-1 inhibitors also have certain inhibitory effect on PARP-2, and the risk of cerebral apoplexy, embryo malformation and anemia is increased after the PARP-2 is deficient or inhibited; 2) the half-life period is short, frequent administration is needed, and the compliance of patients is poor; 3) side effects are high, such as blood and lymphatic system disorder and gastrointestinal reaction, and pneumonia, embryotoxicity and the like are rarely caused after stopping taking the medicine; 4) the administration dosage is large, and whether the occult toxicity exists or not needs to be further examined. Therefore, novel PARP-1 inhibitors with novel structure, better action effect, less toxicity and better pharmacokinetic profile are still expected.
Disclosure of Invention
The purpose of the invention is as follows: aiming at the prior art, the invention provides an 8-azacyclo-substituted chromone derivative with selective PARP-1 inhibitory activity, a preparation method and pharmaceutical application thereof.
The technical scheme is as follows: the structure of the 8-azacyclo-substituted chromone derivative or the pharmaceutically acceptable salt, ester or solvate thereof is shown as the formula I:
wherein R is1Selected from: H. OH, N3、F、C1-C8Alkyl, carboxyl, C1-C8Alkoxy radical, C2-C8Alkenyl radical, C2-C8Alkynyl, C1-C8Alkylcarbonyl group, C1-C8Alkoxycarbonyl group, C1-C8Alkylsulfonyl radical, C1-C8Alkylsulfinyl radical, C1-C8Alkylthio radical, C1-C5Alkylcarbonylamino, C1-C8Alkylaminocarbonyl, di (C)1-C3) Alkylaminocarbonyl radical, C3-C8Cycloalkyl radical, C3-C8Cycloalkyl oxy, C3-C8Heterocycloalkyl radical, C3-C8Heterocycloalkyloxy, phenyl, phenyloxy, phenyl (C)1-C2) Alkyl, phenyl (C)1-C2) Alkoxy, phenylsulfonyl, phenylsulfinyl, C5-C6Heteroaryl group, C5-C6Heteroaryloxy radical, C5-C6Heteroaryl (C)1-C3) Alkyl radical, C5-C6Heteroaryl (C)1-C3) Alkoxy radical, C3-C8Cycloalkyl (C)1-C2) Alkyl radical, C3-C8Cycloalkyl (C)1-C2) Alkoxy radical, C3-C8Heterocycloalkyl (C)1-C2) Alkyl, heterocycloalkyl (C)1-C2) Alkyloxy, condensed ring aryl or substituted condensed ring aryl, wherein R1Each group in (1) may be unsubstitutedSubstituted or substituted by one or more substituents selected from halogen, hydroxy, cyano, unsubstituted or halogenated C1-C8Alkyl and unsubstituted or halogenated C1-C8An alkoxy group;
R2selected from: H. f, Cl, Br, I, CN, NO2、NH2、OH、N3Open chain C2-C8Saturated or unsaturated alkyl, substituted C3-C8Cycloalkyl, substituted C3-C8Heterocycloalkyl radical, C1-C9Alkenyl radical, C1-C9Alkyl radical, wherein R2May be unsubstituted or substituted with one or more substituents selected from halogen, hydroxy, cyano, unsubstituted or halogenated C1-C8Alkyl and unsubstituted or halogenated C1-C8An alkoxy group;
R3、R4、R5independently selected from: H. f, Cl, Br, I, CN, NO2、NH2、OH、N3、Me、C1-C8Alkoxy radical, C1-C8Alkylthio radical, C1-C5Alkylcarbonylamino, C1-C8An alkoxycarbonyl group;
R6selected from: open chain C2-C8Saturated or unsaturated alkyl, substituted C3-C8Cycloalkyl, substituted C3-C8Heterocycloalkyl, substituted phenyl, heteroaryl, substituted heteroaryl, fused ring aryl, or substituted fused ring aryl, wherein said substituted phenyl, substituted heteroaryl, or substituted fused ring aryl may independently be substituted with 1 to 3 of the following substituents: H. f, Cl, Br, I, CN, NO2、NH2、N3、OH、-S(O)2NH2、-S(O)2NHCOCH3、C1-C6Alkyl radical, C1-C6Alkoxy radical, C2-C6Alkenyl radical, C2-C6Alkynyl, 1H-tetrazol-5-yl, 3H- [1,3,4 ]]Oxadiazol-2-one-5-yl, 3H- [1,3,4 ]]Oxadiazole-2-thione-5-yl, 4H- [1,2,4 [ ]]Oxadiazoles as fungicides-5-keto-3-yl, 4H- [1,2,4 [ ]]Oxadiazole-5-thione-3-yl, 3H- [1,2,3,5]Oxathiadiazol-2-oxide-4-yl, wherein each alkyl or alkoxy group may be unsubstituted or substituted by one or more substituents selected from halogen and unsubstituted or fluorinated C1-C3Alkyl or C1-C3An alkoxy group;
a is selected from: furan, thiophene, piperidine, pyridine, pyrrole, imidazole, thiazole, oxazole, piperazine, tetrahydrofuran, tetrahydrothiophene, tetrahydropyridine, imidazoline, thiazoline, oxazoline, C1-C6Triazole, C1-C6Triazole, pyrazine, pyridazine, the said heterocycles being independently substituted by 2-4 carbon atoms1-C3Alkyl, halogen, haloalkyl, cycloalkyl, halocycloalkyl, hydroxyalkyl or alkoxyalkyl;
x is selected from: -S (O)2-, -C (O) -or- (CH)2) n-; n is 0, 1 or 2;
y is selected from: 0 to 1N or C;
wherein m is 0 or 1.
In certain preferred embodiments, the compounds of formula I, pharmaceutically acceptable salts or esters, or solvates thereof, of the present invention:
R1preferably selected from: H. c1-C8Alkyl radical, C3-C8Cycloalkyl radical, C3-C8Heterocycloalkyl, phenyl, substituted phenyl, heteroaryl, substituted heteroaryl, wherein said substituted phenyl or substituted heteroaryl may independently be substituted with 1 to 3 substituents as follows: F. cl, Br, I, CN, NO2、NH2、OH、C1-C3An alkyl group.
R2Preferably selected from: H. f, Cl, Br, I, CN, NO2、NH2、OH、N3Substituted C3-C8A heterocycloalkyl group.
R3、R4、R5Independently preferably from: H. c1-C8Alkoxy radical, C1-C8Alkylthio radical, C1-C5Alkylcarbonylamino, C1-C8An alkoxycarbonyl group.
R6Preferably selected from: open chain C2-C8Saturated or unsaturated alkyl, substituted C3-C8Cycloalkyl, substituted C3-C8Heterocycloalkyl, substituted phenyl, heteroaryl, substituted heteroaryl, fused ring aryl, or substituted fused ring aryl, wherein said substituted phenyl, substituted heteroaryl, or substituted fused ring aryl may independently be substituted with 1 to 3 of the following substituents: H. f, Cl, Br, I, CN, NO2、NH2、N3。
A is preferably selected from: piperidine, pyridine, pyrrole, imidazole, thiazole, oxazole, piperazine, tetrahydropyridine, imidazoline, thiazoline, oxazoline, optionally substituted with 2 to 4 carbon atoms1-C3Alkyl, halogen, haloalkyl, cycloalkyl, halocycloalkyl, hydroxyalkyl or alkoxyalkyl.
X is-C (O) -or- (CH)2) n-; n is 1; y is selected from: 0 to 1N or C; wherein m is 0 or 1.
Further preferably, the derivatives of the invention are selected from the following compounds 1 to 90:
the compounds of the present invention may also be used as pharmaceutically acceptable salts. The salt may be an acid salt of at least one of the following acids: galactaric acid, D-glucuronic acid, glycerophosphoric acid, hippuric acid, isethionic acid, lactobionic acid, maleic acid, 1, 5-naphthalenedisulfonic acid, naphthalene-2-sulfonic acid, pivalic acid, terephthalic acid, thiocyanic acid, cholic acid, n-dodecylsulfuric acid, benzenesulfonic acid, citric acid, D-glucose, glycolic acid, lactic acid, malic acid, malonic acid, mandelic acid, phosphoric acid, propionic acid, hydrochloric acid, sulfuric acid, tartaric acid, succinic acid, formic acid, hydroiodic acid, hydrobromic acid, methanesulfonic acid, nicotinic acid, nitric acid, orotic acid, oxalic acid, picric acid, L-pyroglutamic acid, saccharinic acid, salicylic acid, gentisic acid, p-toluenesulfonic acid, valeric acid, palmitic acid, sebacic acid, stearic acid, lauric acid, acetic acid, adipic acid, carbonic acid, 4-benzenesulfonic acid, ethanedisulfonic acid, ethylsuccinic acid, fumaric acid, 3-hydroxynaphthalene-2-carboxylic acid, 1-hydroxynaphthalene-2-carboxylic acid, oleic acid, undecylenic acid, ascorbic acid, camphoric acid, camphorsulfonic acid, dichloroacetic acid, ethanesulfonic acid. Alternatively, the salts may be formed with metal (including but not limited to sodium, potassium, calcium, etc.) ions or pharmaceutically acceptable amines (including but not limited to ethylenediamine, tromethamine, etc.), ammonium ions or choline of the compounds of the present invention.
The compounds of the present invention may also be formulated as esters, pro-drugs, N-oxides or solvates thereof as active ingredients in pharmaceutical compositions. The dosage form of the pharmaceutical composition is capsule, powder, tablet, granule, pill, injection, syrup, oral liquid, inhalant, cream, ointment, suppository or patch.
The application of the compound of the formula I or the pharmaceutically acceptable salt, ester or solvate thereof in preparing the medicament for preventing or treating the PARP1/2 mediated BRCA1 deficient tumor is also within the protection scope of the invention.
The compound of the formula I or the pharmaceutically acceptable salt, ester or solvate thereof is a novel PARP1/2 inhibitor, and therefore can be used for preparing medicines for preventing or treating PARP1/2 mediated BRCA1 deficient tumors. The PARP-associated disease is cancer, stroke, myocardial infarction, inflammation, hypertension, atherosclerosis or diabetes. The PARP1/2 mediated BRCA-1 deficient tumors are ovarian, breast and fallopian tube cancers, malignant peritoneal tumors, metastatic breast cancers, ovarian epithelial cancers, primary peritoneal cancers, advanced malignant germline or somatic BRCA-mutated advanced epithelial ovarian cancers, advanced malignant germline or somatic BRCA-mutated fallopian tube cancers, advanced malignant germline or somatic BRCA-mutated peritoneal cancers refractory advanced ovarian cancers.
Preferred compounds are in combination with at least one of the following compounds: platinum-based antineoplastic agents (including cisplatin, carboplatin, dicycloplatin, nedaplatin, oxaliplatin, picoplatin, satraplatin), nucleoside analogs and antimetabolites (including arabinoside, fludarabine, gemcitabine, 5FU), DNA intercalators (including daunorubicin, doxorubicin, epirubicin and idarubicin, camptothecin), alkylated neoplastic agents (including cyclophosphamide, melphalan, bendamustine, carmustine, lomustine, ifosfamide), topoisomerase inhibitors (including etoposide, topotecan), PARP inhibitors (including olapanil, nilapanib, lucapanib), agents that interfere with microtubule dynamics (including combretastatin, eribulin, docetaxel, taxanes, vinblastine, vincristine), agents that block the interaction between p53 and MDM2 or MDM4 (including norlin, sardallin, sardalbergine, docetaxel, taxotene, vinorelbine, and vinorelbine), HDM-201, DS3032b, AMG-232, ALRN-6924), kinase inhibitors (including BRAF inhibitors vemomfenib, dabrafenib), PI3K and/or mTOR inhibitors (including LY294002, dacrolimus, rapamycin and the rapamycin analogue sirolimus, everolimus, diphospholimus), MRP1 inhibitors (including indomethacin, meloxicam, sulindac sulfide, GSK1904529A, MK571, verapamil), hypomethylating agents (including azacitidine, decitabine), histone deacetylase inhibitors (including chromaffins, hydroxamates (including vorinostat, belinostat, darsita, panobinostat), valproic acid, benzamides (including entinostat, motricitabine), proteasome inhibitors (including bortezomib, ritonavir, carfilzomib), antiangiogenic or antiangiogenic agents (including 2aG 4), Bevacizumab), tyrosine kinase inhibitors (including lapatinib), EGFR inhibitors (including gefitinib), CDK inhibitors, PLK inhibitors, MEK inhibitors (including pimatinib), immune checkpoint inhibitors (including antibodies to PD-1 (including nivolumab, pembrolizumab), PD-L1 (avizumab, atelizumab), PDL2, CTLA-4 (including ipilimumab, tremelimumab), GITR, IL-40, CD-40, LAG3/CD-223 (including BMS-986016, REGN3767), OX-40 (including palivizumab, PF-04518600)), antibody-binding protein tyrosine kinase receptors, NFE2L2 inhibitors (including 385, brucellol, trigonelline, luteolin, ascorbic acid, ATRA), genetically engineered to express recognition extracellular cancer targets (including CD19, ML) PSMA or mesothelin), a glucocorticoid receptor agonist (including dexamethasone), thiamine, folic acid, metformin, sorafenib, sulfasalazine, bleomycin, erlotinib, tunicamycin, wortmannin, pidilizumab, Devolumab, GSK3174998, tavoriximab, deazaadenine a or piperlongumine.
Preferably the compound is administered alone or in combination with other active pharmaceutical ingredients, and wherein the administration is optionally also in combination with: external beam radiation by gamma radiation or neutron radiation; or targeted therapy with antibodies labeled with beta or alpha emitting radionuclides including I-131, Y-90, Lu-177, Bi-213, Ac-225, Th-227; or radiation therapy with Ra-223.
The adjuvant which can be arbitrarily mixed in the pharmaceutical composition of the present invention may vary depending on the dosage form, administration form, etc. The adjuvants include excipient, binder, disintegrating agent, lubricant, correctant, flavoring agent, colorant, sweetener, etc. The administration route of the pharmaceutical composition can be oral, sublingual, transdermal, intramuscular or subcutaneous, cutaneous mucosa or vein, etc. The pharmaceutical composition can be in the form of capsules, powders, tablets, granules, pills, injections, syrups, oral liquids, inhalants, creams, ointments, suppositories, patches and other pharmaceutically conventional preparations.
The preparation of the compounds of the invention can be carried out with reference to the following synthetic routes or modified methods.
Scheme 1.
Wherein R is1，R2，R3，R4,R5And R6The definition is the same as before.
Firstly, substituted alpha chloroacetophenone (II) is taken as a raw material, and is subjected to addition cyclization reaction with aldehyde under an alkaline condition to obtain corresponding chromone intermediates IV of various substitution types, and then the chromone intermediates IV are subjected to Mannich reaction with formaldehyde and a substituted secondary alcohol V to obtain corresponding Mannich base VI. Wherein, IV is prepared by the reaction of the compound II and III, and the used alkali is selected from sodium hydroxide, potassium hydroxide, lithium hydroxide, potassium carbonate and sodium hydride, preferably sodium hydroxide. The reaction solvent is selected from methanol, ethanol, isopropanol, tert-butanol and perfluoroisopropanol, preferably methanol; the reaction temperature is 10-60 deg.C, preferably 30-50 deg.C.
Preparing compound VI from compound IV, wherein the formaldehyde is selected from 10-50% aqueous solution, preferably 37-40% aqueous solution, and the solvent is selected from methanol, ethanol, isopropanol, diethyl ether, methyl tert-butyl ether, tetrahydrofuran and acetonitrile, preferably tetrahydrofuran.
Scheme 2.
Wherein R is1，R3，R4，R5And R6The definition is the same as before.
Firstly, substituted acetophenone (VI) is taken as a raw material and subjected to a Claisen condensation reaction with aldehyde under an alkaline condition, a crude product is subjected to ring closure under the condition that iodine simple substance is taken as a catalyst to generate VII, and the rest steps are similar to the scheme I.
Compound VII is prepared from compound VI using a base selected from the group consisting of sodium hydroxide, potassium hydroxide, lithium hydroxide, potassium carbonate, sodium hydride, preferably potassium hydroxide. The reaction solvent is selected from methanol, ethanol, isopropanol and tert-butanol, preferably ethanol; the reaction temperature is 10-50 ℃, preferably 10-30 ℃; the solvent used in the ring closing step is selected from: dimethyl sulfoxide, N-dimethylformamide, toluene, preferably dimethyl sulfoxide. The reaction temperature is 60 to 120 ℃ and preferably 80 to 110 ℃.
In the agent for preventing or treating PARP1/2 mediated diseases according to the present invention, the amount of the compound of formula I or a pharmaceutically acceptable salt or ester or solvate thereof may be appropriately changed depending on the age, body weight, symptoms and administration route of the patient, etc. When administered orally to an adult (about 60kg), the compound of formula I or a pharmaceutically acceptable salt or ester or solvate thereof is preferably administered in an amount of 1mg to 500mg per time, more preferably 5mg to 60mg per time, 1 to 3 times per day. This dosage range may also vary depending on the degree of disease and dosage form.
Has the advantages that: the invention provides an 8-azacyclo-substituted chromone compound with selective PARP1/2 inhibitory activity, which can be used for preparing a medicament for preventing or treating PARP1/2 defective tumors and has wide market application and development prospect.
Drawings
FIG. 1 shows the results of the anti-SK-OV-3 cell proliferation assay;
FIG. 2 is the results of the in vivo antitumor effect of the compounds;
FIG. 3 shows the results of in vivo chemosensitization of compounds;
FIG. 4 shows the results of the in vivo anti-tumor and chemosensitization effects of the compounds.
Detailed Description
The present invention will be described in detail with reference to examples.
Example 18- ((4- (Cyclopropanecarbonyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 1)
Powder II-1 (1.01g,5mmol) was dissolved in methanol (20mL) to prepare a solution, and potassium hydroxide (2.52g,15mmol) and p-hydroxybenzaldehyde (0.62g,5.05mmol) were added to the solution. The mixture was left at room temperature and stirred for 18 h. After monitoring the complete disappearance of II-1 by thin layer chromatography, the mixture was diluted with ice-cooled water (50mL) and acidified to pH 5 with 2M hydrochloric acid. Excess methanol was removed on a rotary evaporator. The residue was extracted three times with dichloromethane and water. The combined organic layers were taken over Na2SO4Drying and removing the solvent by a rotary evaporator. The residue was purified by silica gel column chromatography (dichloromethane and methanol from 99: 1 to 93: 7) to give pure compound IV (0.82g, 61%).
A solution of 1-cyclopropylformylpiperazine (142. mu.L, 1mmol) in acetonitrile (10ml) was added dropwise at room temperature to a solution of paraformaldehyde (94.6mg, 1.05mmol) and compound IV-1(0.27g, 1.0mmol) in acetonitrile (5 ml). The reaction mixture was stirred until compound IV-1 completely disappeared. The reaction solvent was then removed using a rotary evaporator. Purifying the residue by flash column chromatography with DCM and MeOH as eluent and silica gel as column material to obtain compound 1;1H NMR(300MHz,DMSO–d6)δ13.04(bs,1H),7.95(d,J＝8.8Hz,2H),6.95(d,J＝8.8Hz,2H),6.80(s,1H),6.25(s,1H),3.85(bs,2H),3.68(bs,2H),3.48(s,2H),2.63–2.50(m,4H),2.04–1.85(m,1H),0.94–0.52(m,4H)；13C NMR(75MHz,DMSO–d6)δ182.3,171.3,164.2,163.9,161.6,160.7,155.6,128.8(C×2),121.7,116.4(C×2),103.9,103.0,101.5,99.0,54.5(C×2),52.6(C×2),50.7,10.6,7.3(C×2).
example 28- ((4- (Cyclobutanecarbonyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 2)
Preparing a compound 2 by the method of the reference compound 1;1H NMR(300MHz,DMSO–d6)δ13.03(s,1H),10.33(bs,1H),7.94(d,J＝8.7Hz,2H),6.94(d,J＝8.7Hz,2H),6.80(s,1H),6.24(s,1H),3.83(s,2H),3.50(bs,2H),3.39–3.25(m,3H),2.51(bs,4H),2.24–1.98(m,4H),1.97–1.81(m,1H),1.81–1.65(m,1H)；13C NMR(75MHz,DMSO–d6)δ182.4,172.3,164.1,164.0,161.6,160.8,155.6,128.8(C×2),121.8,116.4(C×2),104.0,103.1,101.4,99.0,52.9,52.5,50.9,44.7,40.8,36.6,24.9(C×2),17.8.
example 38- ((4- (Cyclohexanecarbonyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 3)
Compound 3 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.00(bs,1H),7.92(d,J＝8.7Hz,2H),6.94(d,J＝8.8Hz,2H),6.77(s,1H),6.23(s,1H),3.83(bs,2H),3.47(bs,4H),2.65–2.38(m,5H),1.75–1.52(m,5H),1.40–1.12(m,5H)；13C NMR(75MHz,DMSO–d6)δ182.0,173.3,163.7,163.5,161.2,160.4,155.2,128.4(C×2),121.4,116.0(C×2),103.6,102.7,101.0,98.6,52.8,52.2,50.4(C×2),44.7,40.9,29.1(C×2),25.6,25.1(C×2).
example 48- ((4-Acetylpiperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 4)
Compound 4 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.04(s,1H),10.34(bs,1H),7.95(d,J＝8.8Hz,2H),6.95(d,J＝8.8Hz,2H),6.81(s,1H),6.25(s,1H),3.84(s,2H),3.44(bs,4H),2.53(bs,4H),1.98(s,3H)；13C NMR(75MHz,DMSO–d6)δ181.9,168.0,163.6,163.5,161.1,160.3,155.1,128.4(C×2),121.4,115.9(C×2),103.6,102.6,101.0,98.5,52.3,51.9,50.3,45.5,40.7,21.0.
example 58- ((4-Benzoylpiperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 5)
Compound 5 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.03(bs,1H),7.94(d,J＝7.9Hz,2H),7.66–7.22(m,5H),6.95(d,J＝8.0Hz,2H),6.79(s,1H),6.24(s,1H),3.84(s,2H),3.72–3.26(m,4H),2.57(bs,4H)；13C NMR(75MHz,DMSO–d6)δ182.0,168.9,163.6,163.5,161.2,160.4,155.2,135.8,129.5,128.4(C×2),128.4(C×2),126.9(C×2),121.4,116.0(C×2),103.6,102.6,101.1,98.5,52.2(C×2),50.3,47.3,41.5.
example 65, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (4-methoxybenzoyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 6)
Compound 6 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.01(s,1H),7.90(d,J＝8.4Hz,2H),7.33(d,J＝8.2Hz,2H),7.02–6.89(m,4H),6.69(s,1H),6.20(s,1H),3.81(s,2H),3.75(s,3H),3.49(bs,4H),2.55(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.8,168.9,165.3,163.4,161.7,160.4,160.2,155.5,129.0(C×2),128.3(C×2),127.8,121.2,116.2(C×2),113.6(C×2),103.1,102.4,101.4,98.9,55.2,52.4,50.3(C×2),47.0,41.6.
example 75, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (3- (trifluoromethyl) benzoyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 7)
Compound 7 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.04(s,1H),10.34(s,1H),7.93(d,J＝8.6Hz,2H),7.81(d,J＝6.7Hz,1H),7.77–7.63(m,3H),6.95(d,J＝8.6Hz,2H),6.78(s,1H),6.24(s,1H),3.86(s,2H),3.72–3.55(m,2H),3.46–3.27(m,2H),2.61(s,4H)；13C NMR(75MHz,DMSO–d6)δ182.0,167.4,163.6(C×2),161.2,160.4,155.3,136.9,130.9,130.0(q,JCF＝31.7Hz),129.6,128.4(C×2),129.2(q,JCF＝271.0Hz),126.2(q,JCF＝3.6Hz),123.7(q,JCF＝3.5Hz),121.4,116.0(C×2),103.7,102.7,101.0,98.5,52.0,50.2(C×2),47.0,41.4；19F NMR(376MHz,DMSO–d6)δ-62.7.
example 88- ((4- (4-chlorobenzoyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl-yl) -4H-benzopyran-4-one (Compound 8)
Compound 8 was prepared according to the procedure for compound 1;1H NMR(300MHz,DMSO–d6)δ7.91(d,J＝8.6Hz,2H),7.61–7.34(m,4H),6.95(d,J＝8.6Hz,2H),6.73(s,1H),6.23(s,1H),3.83(s,2H),3.74–3.19(m,4H),2.58(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,167.9,163.6,163.6,161.20 160.5,155.3,134.6,134.3,128.9(C×2),128.5(C×2),128.4(C×2),121.5,116.1(C×2),103.7,102.7,101.0,98.6,52.2(C×2),50.3,46.6,41.4.
example 95, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4-nicotinoylpiperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 9)
Compound 9 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.02(s,1H),8.70–8.55(m,2H),7.92(d,J＝8.6Hz,2H),7.81(d,J＝7.8Hz,1H),7.45(dd,J＝7.6,4.8Hz,1H),6.93(t,J＝8.2Hz,2H),6.76(s,1H),6.23(s,1H),3.83(s,2H),3.68(bs,2H),3.36(bs,2H),2.57(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,166.7,164.1,163.6,161.1,160.4,155.2,150.4,147.6,134.7,131.6,128.4(C×2),123.4,121.4,116.0(C×2),103.6,102.6,101.1,98.5,52.1(C×2),50.2,47.1,41.6.
example 105, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4-isonicotinoylpiperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 10)
Compound 10 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ8.65(s,2H),7.91(s,2H),7.37(s,2H),6.94(s,2H),6.76(s,1H),6.24(s,1H),3.83(s,2H),3.65(bs,2H),3.28(bs,2H),2.62(bs,2H),2.51(bs,2H)；13C NMR(300MHz,DMSO–d6)δ182.3,167.0,163.9,161.5,161.1,160.7,155.6,150.4(C×2),143.7,128.8(C×2),121.5,121.4(C×2),116.4(C×2),104.0,103.0,101.5,98.9,52.2,50.5,47.3,47.2,41.6.
example 115, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4-pyridinylpiperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 11)
Compound 11 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.03(bs,1H),10.28(bs,1H),8.56(d,J＝3.9Hz,1H),8.03–7.82(m,3H),7.55(d,J＝7.6Hz,1H),7.51–7.39(m,1H),6.94(d,J＝8.3Hz,2H),6.77(s,1H),6.23(s,1H),3.84(s,2H),3.66(bs,2H),3.41(bs,2H),2.63(bs,2H),2.54(bs,2H)；13C NMR(75MHz,DMSO–d6)δ182.3,167.0,165.0,164.1,161.6,160.8,155.6,154.4,148.7,137.6,128.8(C×2),124.9,123.5,121.9,116.4(C×2),104.1,103.1,101.5,99.0,52.9,52.4,50.7,46.8,41.9.
example 125, 7-dihydroxy-2- (4-hydroxyphenyl) -8- (((4- (thiophene-2-carbonyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 12)
Compound 12 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ7.92(d,J＝8.7Hz,2H),7.71(dd,J＝4.9,0.6Hz,1H),7.42–7.32(m,1H),7.09(dd,J＝4.8,3.8Hz,1H),6.94(d,J＝8.7Hz,2H),6.75(s,1H),6.23(s,1H),3.84(s,2H),3.65(bs,4H),2.59(bs,4H)；13C NMR(75MHz,DMSO–d6)δ182.0,163.7,163.6,162.3,161.2,160.4,155.2,137.1,129.4,129.1,128.4(C×2),127.1,121.4,116.1(C×2),103.7,102.7,101.1,98.6,52.6(C×2),50.6,46.8,41.9.
EXAMPLE 138- (((4- (furan-2-carbonyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 13)
Compound 13 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ7.96(d,J＝8.0Hz,2H),7.82(s,1H),7.06–6.90(m,3H),6.80(s,1H),6.62(s,1H),6.27(s,1H),3.88(s,2H),3.70(bs,4H),2.62(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.8,163.5,163.4,162.1,161.0,160.3,155.1,136.9,129.0,128.8,128.2(C×2),126.8,121.3,115.9(C×2),103.6,102.6,100.9,98.4,52.1(C×2),50.2,44.6,40.3.
EXAMPLE 144 benzyl- (((5, 7-dihydroxy-2- (4-hydroxyphenyl) -4-oxo-4H-benzopyran-8-yl) methyl) piperazine-1-carboxylate (Compound 14)
Compound 14 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.04(bs,1H),10.04(bs,1H),7.93(d,J＝8.4Hz,2H),7.34(bs,5H),6.95(d,J＝8.5Hz,2H),6.78(s,1H),6.24(s,1H),5.07(s,2H),3.82(s,2H),3.42(s,4H),2.53(s,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,163.6,163.5,161.1,160.3,155.1,154.3,136.8,128.3(C×2),128.3(C×2),127.7,127.4(C×2),121.3,115.9(C×2),103.6,102.6,100.9,98.5,66.1,51.9(C×2),50.4,43.4(C×2).
example 153, 5-dimethylbenzyl 4- ((5, 7-dihydroxy-2- (4-hydroxyphenyl) -4-oxo-4H-benzopyran-8-yl) methyl) piperazine-1-carboxylic acid (Compound 15)
Compound 15 was prepared according to compound 1;1H NMR(500MHz,DMSO–d6)δ13.03(bs,1H),9.80(bs,1H),7.99–7.85(m,2H),6.94(d,J＝8.7Hz,2H),6.77(s,1H),6.48(s,2H),6.42(s,1H),6.23(s,1H),4.99(s,2H),3.82(s,2H),3.71(s,6H),3.42(bs,4H),2.53(bs,4H)；13C NMR(125MHz,DMSO–d6)δ181.9,163.6,163.5,161.1,160.5(C×2),160.4,155.2,154.2,139.2,128.4(C×2),121.4,116.0(C×2),105.1(C×2),103.6,102.6,100.9,99.4,98.5,65.9,55.1(C×2),51.9(C×2),50.4,43.4(C×2).
EXAMPLE 168- (((4- (cyclopropylmethyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 16)
Compound 16 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ12.99(bs,1H),7.91(bs,2H),6.94(d,J＝8.3Hz,2H),6.75(s,1H),6.16(s,1H),3.90(s,2H),2.61(bs,4H),2.51(bs,4H),2.17(d,J＝6.1Hz,2H),0.79(s,1H),0.51–0.34(m,2H),0.12–0.01(m,2H)；13C NMR(300MHz,DMSO–d6)δ181.9,164.8,163.4,161.2,160.4,154.8,128.4(C×2),121.4,116.0(C×2),103.4,102.6,100.2,98.8,62.5,52.4(C×2),52.0(C×2),51.4,8.1,3.6(C×2)
example 178- ((4- (cyclohexylmethyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 17)
Compound 17 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ12.98(bs,1H),7.93(d,J＝8.4Hz,2H),6.93(d,J＝8.4Hz,2H),6.76(s,1H),6.16(s,1H),3.91(s,2H),2.60(bs,4H),2.36(bs,4H),2.06(d,J＝6.8Hz,2H),1.78–1.52(m,5H),1.44(bs,1H),1.23–1.07(m,3H),0.89–0.69(m,2H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.7,163.3,161.1,160.3,154.7,128.4(C×2),121.3,115.9(C×2),103.3,102.6,100.1,98.7,64.5,52.9(C×2),52.1(C×2),51.4,34.3,31.1(C×2),26.3,25.4(C×2).
example 185, 7-dihydroxy-2- (4-hydroxyphenyl) -8- (((4-isopentylpiperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 18)
Compound 18 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.02(s,1H),8.11–7.76(m,2H),7.08–6.84(m,2H),6.73(s,1H),6.14(s,1H),3.88(s,2H),2.78–2.50(m,4H),2.51–2.29(m,4H),2.25(s,2H),1.69–1.42(m,1H),1.40–1.19(m,2H),1.04–0.64(m,6H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.8,163.3,161.2,160.4,154.7,128.3(C×2),121.3,115.9(C×2),103.3,102.6,100.1,98.8,55.7,52.6(C×2),52.0(C×2),51.4,35.2,25.7,22.5(C×2).
example 195, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (4-methylbenzyl) piperazin-1-yl) methyl-yl) -4H-benzopyran-4-one (Compound 19)
Compound 20 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.00(bs,1H),7.91(d,J＝8.7Hz,2H),7.22–7.07(m,4H),6.94(d,J＝8.6Hz,2H),6.74(s,1H),6.16(s,1H),3.90(s,2H),3.41(s,2H),2.61(bs,4H),2.40(bs,4H),2.25(s,3H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.7,163.3,161.1,160.3,154.7,135.9,134.8,128.7(C×2),128.6(C×2),128.3(C×2),121.3,115.9(C×2),103.4,102.6,100.1,98.7,61.5,52.2(C×2),52.0(C×2),51.4,20.6.
example 208- ((4- (4-chlorobenzyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 20)
Compound 20 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.00(bs,1H),7.93(d,J＝8.6Hz,2H),7.41–7.26(m,4H),6.94(d,J＝8.7Hz,2H),6.77(s,1H),6.17(s,1H),3.90(s,2H),3.46(s,2H),2.61(bs,4H),2.41(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.5,163.4,161.1,160.3,154.7,137.1,131.4,130.5(C×2),128.4(C×2),128.1(C×2),121.3,115.9(C×2),103.4,102.6,100.2,98.7,60.8,52.3(C×2),52.0(C×2),51.2.
example 215, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (4-methoxybenzyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 21)
Compound 21 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ12.98(bs,1H),7.91(d,J＝5.8Hz,2H),7.17(d,J＝7.6Hz,2H),6.89(dd,J＝26.2,7.9Hz,4H),6.74(s,1H),6.16(s,1H),3.89(s,2H),3.71(s,3H),3.38(s,2H),2.59(bs,4H),2.38(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.7,163.3,161.1,160.3,158.3,154.7,129.9(C×2),129.7,128.3(C×2),121.3,115.9(C×2),113.5(C×2),103.4,102.6,100.1,98.7,61.1,54.9,52.2(C×2),52.0(C×2),51.4.
EXAMPLE 228- (((4- (3, 5-Dimethoxybenzyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 22)
Prepared by the method of reference Compound 1An object 22;1H NMR(300MHz,DMSO–d6)δ13.00(bs,1H),7.94(d,J＝8.8Hz,2H),6.94(d,J＝8.8Hz,2H),6.77(s,1H),6.46(d,J＝2.2Hz,2H),6.36(d,J＝2.2Hz,1H),6.17(s,1H),3.92(s,2H),3.71(s,6H),3.41(s,2H),2.63(bs,4H),2.42(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.6,163.4,161.1,160.3(2C),160.3,154.7,140.5,128.4(2C),121.3,115.9(2C),106.5(2C),106.5,103.4,102.6,100.1,98.7,61.8,55.0(2C),52.3(2C),52.0(2C),51.4.
example 235, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (2,3, 4-trimethoxybenzyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 23)
Compound 23 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.00(bs,1H),7.93(d,J＝8.0Hz,2H),7.02–6.87(m,3H),6.83–6.67(m,2H),6.17(s,1H),3.91(s,2H),3.77(s,6H),3.73(s,3H),3.40(s,2H),2.61(bs,4H),2.43(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.6,163.4,161.1,160.3,154.7,152.5,151.9,141.8,128.3(2C),124.6,123.4,121.3,115.9(2C),107.5,103.3,102.6,100.1,98.7,60.8,60.2,55.7,55.6,52.2(2C),52.1(2C),51.3
example 245, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (4- (trifluoromethyl) benzyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 24)
Compound 24 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ12.99(s,1H),7.93(d,J＝8.3Hz,2H),7.66(d,J＝7.8Hz,2H),7.51(d,J＝7.6Hz,2H),6.94(d,J＝8.4Hz,2H),6.77(s,1H),6.17(s,1H),3.90(s,2H),3.56(s,2H),2.62(bs,4H),2.43(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.5,163.4,161.1,160.3,154.8,143.2,130.1(q,JCF＝270.5Hz),129.2(2C),128.4(q,JCF＝31.2Hz,2C),128.2,124.9(q,JCF＝3.1Hz,2C),121.3,116.0(2C),103.4,102.6,100.3,98.7,61.0,52.4(2C),52.0(2C),51.2；19F NMR(376MHz,DMSO–d6)δ-62.4.
example 258- (4- (4-fluorobenzyl) phenylazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl-yl) -4H-benzopyran-4-one (Compound 25)
Compound 25 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.00(s,1H),7.93(d,J＝8.6Hz,2H),7.37–7.23(m,2H),7.18–7.02(m,2H),6.94(d,J＝8.6Hz,2H),6.77(s,1H),6.17(s,1H),3.90(s,2H),3.45(s,2H),2.60(bs,4H),2.40(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,164.8(d,JCF＝249.5Hz),164.5,163.4,161.1,160.3,154.7,134.2,130.6(d,JCF＝7.7Hz,2C),128.4(2C),121.3,116.0(2C),115.0(d,JCF＝20.8Hz,2C),103.4,102.6,100.2,98.7,60.8,52.2(2C),52.0(2C),51.3；19F NMR(376MHz,DMSO–d6)δ-115.93.
example 265, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- ((tetrahydrofuran-2-yl) methyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 26)
Compound 26 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.02(s,1H),8.20–7.70(m,2H),7.09–6.82(m,2H),6.75(s,1H),6.22(s,1H),4.00(bs,3H),3.80(s,2H),3.63(s,2H),3.37(bs,4H),2.50(bs,4H),1.70(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,163.7,161.2,160.4,155.1,154.4,128.4(2C),121.4,116.0(2C),103.6,102.6,101.0,98.6,64.1,51.9(2C),50.5,45.0,43.3,28.7,26.0；
example 275, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (thien-2-ylmethyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 27)
Compound 27 was prepared according to compound 1;1H NMR(300MHz,DMSO-d6)δ12.98(bs,1H),9.16(bs,2H),7.92(d,J＝8.7Hz,2H),7.47–7.31(m,1H),7.03–6.85(m,4H),6.74(s,1H),6.16(s,1H),3.90(s,2H),3.68(s,2H),2.62(bs,4H),2.46(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.5,163.4,161.1,160.3,154.7,141.4,128.4(2C),126.4,126.0,125.4,121.3,116.0(2C),103.4,102.6,100.2,98.7,56.0,52.0(2C),51.2.
EXAMPLE 288- (((4- (furan-2-ylmethyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 28)
Reference Compound 1 formulationPreparing a compound 28;1H NMR(300MHz,DMSO–d6)δ12.99(bs,1H),10.00(bs,1H),7.88(d,J＝8.5Hz,2H),7.54(s,1H),6.92(d,J＝8.6Hz,2H),6.71(s,1H),6.37(d,J＝1.6Hz,1H),6.24(s,1H),6.15(s,1H),3.86(s,2H),3.48(s,2H),2.59(bs,4H),2.43(bs,4H)；13C NMR(300MHz,DMSO–d6)δ181.8,164.6,163.4,161.1,160.4,154.7,151.6,142.3,128.3(2C),121.4,116.0(2C),110.2,108.6,103.4,102.6,100.2,98.7,53.6,52.0(2C),51.9(2C),51.3.
example 295, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (pyridin-4-ylmethyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 29)
Compound 29 was prepared according to the procedure for compound 1;1H NMR(300MHz,DMSO–d6)δ12.99(s,1H),8.49(d,J＝5.4Hz,2H),7.93(d,J＝8.7Hz,2H),7.30(d,J＝5.4Hz,2H),6.94(d,J＝8.7Hz,2H),6.76(s,1H),6.17(s,1H),3.90(s,2H),3.50(s,2H),2.62(bs,4H),2.43(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,164.4,163.4,161.2,160.3,154.8,149.8(2C),147.2,128.4(2C),123.6(2C),121.4,116.0(2C),103.5,102.7,100.3,98.7,60.3,52.4(2C),52.0(2C),51.2.
example 305, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (thien-3-ylmethyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 30)
Compound 30 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ7.87(d,J＝8.6Hz,2H),7.46–7.37(m,1H),7.24(s,1H),7.01(d,J＝4.8Hz,1H),6.94(d,J＝8.6Hz,2H),6.67(s,1H),6.15(s,1H),3.89(s,2H),3.48(s,2H),2.61(bs,4H),2.42(bs,4H)；13C NMR(75MHz,DMSO–d6)δ182.2,165.1,163.8,161.6,160.8,155.1,139.3,128.8,128.7(2C),126.1,123.2,121.8,116.4(2C),103.9,103.1,100.5,99.2,56.9,52.6(2C),52.5(2C),51.9.
example 318- (((4- (furan-3-ylmethyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 31)
Compound 31 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.01(bs,1H),7.91(d,J＝8.4Hz,2H),7.55(d,J＝16.6Hz,2H),6.94(d,J＝8.3Hz,2H),6.74(s,1H),6.40(s,1H),6.16(s,1H),3.89(s,2H),3.32(s,2H),2.61(bs,4H),2.41(bs,4H)；13C NMR(300MHz,DMSO–d6)δ181.8,164.7,163.4,161.1,160.4,154.7,143.2,140.8,128.3(2C),121.4,121.3,116.0(2C),111.4,103.4,102.6,100.1,98.8,52.1(2C),52.0(2C),51.8,51.3.
example 325, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4-phenylpiperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 32)
Compound 32 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.03(bs,1H),8.52(bs,2H),7.94(d,J＝8.6Hz,2H),7.28–7.12(m,2H),7.01–6.83(m,4H),6.81–6.73(m,2H),6.23(s,1H),3.90(s,2H),3.15(s,4H),2.71(s,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,164.0,163.5,161.1,160.3,155.0,150.7,128.8(2C),128.4(2C),121.4,118.9,115.9(2C),115.4(2C),103.5,102.6,100.7,98.6,52.1(2C),50.8,48.2(2C).
example 335, 7-dihydroxy-2- (2-hydroxyphenyl) -8- ((4- (2-hydroxyphenyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 33)
Compound 33 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.02(bs,1H),7.96(d,J＝8.7Hz,2H),6.95(d,J＝8.6Hz,2H),6.93–6.86(m,1H),6.85–6.71(m,4H),6.21(s,1H),3.96(s,2H),2.98(s,4H),2.76(s,4H)；13C NMR(75MHz,DMSO–d6)δ182.3,164.8,163.9,161.6,160.8,155.3,150.5,139.9,128.8(2C),123.3,121.8,119.7,119.1,116.4(2C),115.9,103.9,103.1,100.9,99.1,52.7(2C),51.7,50.4(2C).
example 345, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (4-hydroxyphenyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 34)
Compound 34 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.02(bs,1H),7.94(d,J＝7.5Hz,2H),6.94(d,J＝8.6Hz,2H),6.82–6.69(m,3H),6.70–6.55(m,2H),6.21(s,1H),3.91(s,2H),2.98(s,4H),2.70(s,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,164.3,163.5,161.2,160.4,154.9,151.0,143.9,128.4(2C),121.4,117.9(2C),116.0(2C),115.5(2C),103.5,102.7,100.7,98.7,52.3(2C),51.0,50.0(2C).
EXAMPLE 355, 7-dihydroxy-2- (4-hydroxyphenyl) -8- (((4- (p-tolyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 35)
Compound 35 was prepared according to compound 1;1H NMR(500MHz,DMSO–d6)δ13.02(bs,1H),9.83(bs,1H),8.00–7.89(m,2H),6.99(d,J＝5.2Hz,2H),6.94(d,J＝8.7Hz,2H),6.85–6.72(m,3H),6.22(s,1H),3.90(s,2H),3.08(s,4H),2.70(s,4H),2.17(s,3H)；13C NMR(75MHz,DMSO–d6)δ181.9,164.1,163.5,161.1,160.4,154.9,148.7,129.3(2C),128.4(2C),127.7,121.4,115.9(2C),115.7(2C),103.5,102.6,100.7,98.6,52.1(2C),50.9,48.7(2C),19.9.
example 368- (((4- (4-chlorophenyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 36)
Compound 36 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.03(bs,1H),10.23(bs,1H),7.95(d,J＝8.7Hz,2H),7.21(d,J＝8.9Hz,2H),7.02–6.85(m,4H),6.79(s,1H),6.24(s,1H),3.89(s,2H),3.16(bs,4H),2.69(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,163.8,163.5,161.1,160.3,155.0,149.5,128.5(2C),128.4(2C),122.4,121.3,116.8(2C),115.9(2C),103.5,102.6,100.9,98.6,51.9(2C),50.6,48.0(2C).
example 375, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (4- (trifluoromethyl) phenyl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 37)
Compound 37 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.04(bs,1H),7.95(d,J＝8.5Hz,2H),7.47(d,J＝8.5Hz,2H),7.03(d,J＝8.5Hz,2H),6.94(d,J＝8.5Hz,2H),6.78(s,1H),6.25(s,1H),3.88(s,2H),3.29(bs,4H),2.69(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,163.8,163.5,161.1,160.4,155.1,153.0,128.4(2C),126.7(q,J＝268.0Hz),126.1(q,J＝3.1Hz,2C),121.4,118.1(q,J＝31.9Hz),116.0(2C),114.2(2C),103.6,102.6,100.9,98.6,51.8(2C),50.5,46.9(2C).
example 388- ((4- (4-fluorophenyl) piperazin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 38)
Compound 38 was prepared according to compound 1;1H NMR(500MHz,DMSO–d6)δ13.02(s,1H),7.95(d,J＝7.4Hz,2H),7.12–6.99(m,2H),6.99–6.88(m,4H),6.79(s,1H),6.24(s,1H),3.91(s,2H),3.10(s,4H),2.71(s,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,164.0,163.1(d,J＝272.2Hz),161.1,160.3,157.6,155.0,147.7,128.4(2C),121.4,117.3,117.2,116.0(2C),115.3(d,J＝21.7Hz,2C),103.5,102.6,100.8,98.6,52.1(2C),50.7,49.0(2C)；19F NMR(376MHz,DMSO–d6)δ-124.4.
example 395, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4- (pyrimidin-5-yl) piperazin-1-yl) methyl) -4H-benzopyran-4-one (Compound 39)
Compound 39 was prepared according to compound 1;1H NMR(300MHz,DMSO–d6)δ13.03(bs,1H),9.99(bs,2H),8.34(d,J＝4.3Hz,2H),7.94(d,J＝8.2Hz,2H),6.93(d,J＝8.2Hz,2H),6.79(s,1H),6.69–6.50(m,1H),6.23(s,1H),3.88(s,2H),3.75(bs,4H),2.61(bs,4H)；13C NMR(75MHz,DMSO–d6)δ181.9,163.9,163.5,161.1,161.1,160.4,157.8(2C),155.1,128.4(2C),121.4,116.0(2C),110.1,103.6,102.7,100.9,98.6,52.0(2C),50.8,43.2(2C).
example 401- ((5, 7-dihydroxy-2- (4-hydroxyphenyl) -4-oxo-4H-chromen-8-yl) methyl) piperidine-4-carboxylic acid methyl ester (Compound 40)
VI-1(1.6g,0.01mol) was dissolved in absolute ethanol and potassium hydroxide solid (2.8g,0.05mol) was added with vigorous stirring at room temperature, and the reaction color gradually turned red. After 4h, the PH was adjusted to 5 with 2M hydrochloric acid, a yellow solid gradually precipitated in the process, diluted with water (200mL), filtered and dried to obtain a solid, which was used as it was. Dissolving the intermediate obtained in the previous step in anhydrous dimethyl sulfoxide (30mL), adding iodine simple substance (0.13g,0.5mmol) as catalystThe reaction was then allowed to react at 110 ℃ for 8h, after completion the reaction was cooled to room temperature, 10% sodium thiosulfate solution (50mL) was added and after 10min, the mixture was extracted with ethyl acetate/water system, the organic phase was taken, dried and spun dry and the residue was purified by silica gel chromatography to give VII-1(1.3g, 48% yield over two steps). And with reference to scheme 1 to produce compound 40;1H NMR(300MHz,DMSO–d6)δ12.99(bs,1H),7.92(d,J＝8.8Hz,2H),6.94(d,J＝8.8Hz,2H),6.77(s,1H),6.16(s,1H),3.89(s,2H),3.60(s,3H),2.92(d,J＝11.7Hz,2H),2.47–2.37(m,1H),2.31(t,J＝10.5Hz,2H),1.87(d,J＝11.1Hz,2H),1.70–1.48(m,2H)；13C NMR(75MHz,DMSO–d6)δ181.8,174.4,164.8,163.3,161.1,160.3,154.8,128.3(2C),121.4,115.9(2C),103.3,102.7,100.2,98.8,51.6,51.5(2C),51.4,51.4,27.7(2C).
EXAMPLE 411- (((5, 7-dihydroxy-2- (4-hydroxyphenyl) -4-oxo-4H-benzopyran-8-yl) methyl) piperidine-4-carboxylic acid ethyl ester (Compound 41)
Compound 41 was prepared according to compound 40;1H NMR(300MHz,DMSO–d6)δ13.00(bs,1H),8.96(bs,2H),7.91(d,J＝8.8Hz,2H),6.93(d,J＝8.7Hz,2H),6.75(s,1H),6.15(s,1H),4.06(q,J＝7.1Hz,2H),3.88(s,2H),2.92(d,J＝11.7Hz,2H),2.45–2.22(m,3H),1.86(d,J＝11.0Hz,2H),1.58(dd,J＝21.4,10.8Hz,2H),1.17(t,J＝7.1Hz,3H)；13C NMR(75MHz,DMSO–d6)δ181.8,173.9,164.8,163.3,161.1,160.3,154.7,128.3(2C),121.4,115.9(2C),103.3,102.6,100.1,98.8,59.8,51.6,51.5(2C),39.6,27.7(2C),13.9
EXAMPLE 425, 7-dihydroxy-8- (((4- (hydroxymethyl) piperidin-1-yl) methyl) -2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 42)
Compound 42 was prepared according to compound 40;1H NMR(300MHz,DMSO–d6)δ12.97(bs,1H),7.91(d,J＝8.8Hz,2H),6.94(d,J＝8.7Hz,2H),6.74(s,1H),6.09(s,1H),3.97(s,2H),3.27(d,J＝6.1Hz,2H),3.02(d,J＝11.4Hz,2H),2.30(t,J＝11.0Hz,2H),1.73(d,J＝11.9Hz,2H),1.46(s,1H),1.30–1.08(m,2H)；13C NMR(75MHz,DMSO–d6)δ181.6,166.1,163.1,161.1,160.4,154.5,128.3(2C),121.4,115.9(2C),102.9,102.6,99.41,99.0,65.4,52.3,52.1(2C),37.6,28.2(2C).
example 435, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4-phenylpiperidin-1-yl) methyl) -4H-benzopyran-4-one (Compound 43)
Compound 43 was prepared according to compound 40;1H NMR(300MHz,DMSO–d6)δ7.94(d,J＝8.8Hz,2H),7.36–7.12(m,5H),6.95(d,J＝8.8Hz,2H),6.77(s,1H),6.16(s,1H),3.98(s,2H),3.11(d,J＝11.4Hz,2H),2.60(t,J＝11.8Hz,1H),2.40(t,J＝11.2Hz,2H),1.83(d,J＝11.9Hz,2H),1.75–1.58(m,2H)；13C NMR(75MHz,DMSO–d6)δ181.9,165.4,163.3,161.2,160.4,154.7,145.6,128.3(2C),128.2(2C),126.6(2C),126.1,121.4,116.0(2C),103.2,102.7,100.0,98.9,52.9(2C),51.9,41.1,32.6(2C).
EXAMPLE 448- (((4- (2, 3-difluorobenzoyl) piperidin-1-yl) methyl) -5, 7-dihydroxy-2- (4-hydroxyphenyl) -4H-benzopyran-4-one (Compound 44)
Compound 44 was prepared according to compound 40;1H NMR(300MHz,DMSO–d6)δ13.01(bs,1H),8.11–7.78(m,3H),7.50–7.32(m,1H),7.32–7.13(m,1H),6.94(d,J＝7.7Hz,2H),6.76(s,1H),6.16(s,1H),3.92(s,2H),3.33–3.07(m,1H),3.01(d,J＝9.8Hz,2H),2.46–2.23(m,2H),1.87(d,J＝11.7Hz,2H),1.72–1.38(m,2H)；13C NMR(75MHz,DMSO–d6)δ199.3(d,JCF＝3.7Hz),181.8,167.7(dd,JCF＝258.8Hz,14.2Hz),166.4(dd,JCF＝254.2Hz,13.0Hz),164.8,163.3,162.9,161.1,160.3,132.6(m),128.4(2C),124.3(m),122.0(m),121.4,116.0(2C),112.6(dd,JCF＝22.8Hz,3.6Hz),103.3,102.7,100.2,98.8,51.8(2C),51.5,46.2(d,JCF＝3.8Hz),27.5(2C)；19F NMR(376MHz,DMSO–d6)δ-136.9,-137.0.
example 455, 7-dihydroxy-2- (4-hydroxyphenyl) -8- ((4-phenyl-3, 6-dihydropyridin-1 (2H) -yl) methyl) -4H-benzopyran-4-one (Compound 45)
Compound 45 was prepared according to compound 40;1H NMR(300MHz,DMSO–d6)δ12.99(s,1H),7.91(d,J＝8.3Hz,2H),7.41(d,J＝7.3Hz,2H),7.31(t,J＝7.3Hz,2H),7.24(d,J＝7.0Hz,1H),6.92(d,J＝8.3Hz,2H),6.75(s,1H),6.17(d,J＝10.7Hz,2H),4.02(s,2H),3.31(s,2H),2.88(s,2H),2.51(d,J＝7.8Hz,2H)；13C NMR(75MHz,DMSO–d6)δ181.8,164.7,161.1,160.4,159.1,154.8,136.4,133.9,128.3(2C),128.3(2C),127.0,124.5(2C),121.3,120.9,115.9(2C),103.3,102.6,100.4,98.8,51.7,50.5,48.9,26.7.
EXAMPLE 46 IC of test Compounds at the enzyme level for inhibiting PARP-1/250Value of
Experimental methods
Dissolving a compound sample by using anhydrous dimethyl sulfoxide to prepare 10mM mother liquor, adding the compound into a screening system, wherein the detection concentration range of the compound is 0.5 nM-10 mu M, diluting according to a 3-fold gradient, and making two multiple holes at each concentration. The results of the experiments were converted to percent activity, and the IC of the test compound for inhibiting PARP-1/2 enzyme was calculated using the drug concentration as the abscissa and the percent enzyme activity corresponding to each concentration as the ordinate50The value is obtained. The specific operation steps are as follows: the inhibitory activity of the compounds of interest on the PARP-1/2 enzyme was tested in 96-well plates. Each well was precoated with histone (20. mu.g/mL) diluted in 100. mu.L of PBS buffer (10mM sodium dihydrogenphosphate, 10mM disodium hydrogenphosphate, 150mM sodium chloride, pH 7.4) and incubated overnight at 4 ℃. Thereafter, 100. mu.M NAD +, 25. mu.M biotinylated NAD +, and 200nM siDNA diluted in 30. mu.L buffer (50mM Tris, 2mM magnesium chloride, pH 8.0) were added per well, followed by the addition of 5. mu.L of test compound or solvent control at various concentrations. mu.L (5ng) of PARP-1 was added to each well at 30 ℃ for 1 hour, then 50. mu.L of HRP was added, and after incubation for 30min, 100. mu.L of buffer (0.1M H) was added2O2Citrate buffer, pH 5.4) and the chemiluminescence was measured on a SpectraMax M5 instrument.
The experimental results are as follows: the results of the experiments show that the PARP-1 inhibition rate IC of the compounds 11-13,27 and 2850A value of less than 25nM, wherein the PARP-1 inhibition IC of compound 27 is preferred50The value was 15nM, slightly less effective than the marketed Olaparib (IC)505.1 nM). But the selectivity is 60, which is far greater than Olaparib (selectivity 0.2) on the market, and the clinical significance is larger.
PARP-1 IC at the enzyme level of the Compounds of Table 150Value of
EXAMPLE 47 cellular anti-proliferation assay for detection of preferred Compound 27
All cells were cultured according to the ATCC recommendations with olararamib as the positive control drug. SK-OV-3 tumor cells in the logarithmic growth phase are inoculated in a 96-well plate, the inoculation density is 5000 cells/well, compounds with different concentrations (0.1,1 and 10 mu M) are used for treatment after 24 hours, a blank control group is arranged, and 3 multiple wells are arranged in parallel. After 48h, MTT solution (5 mg. mL) was added to each well-1)20 μ L. At 37 ℃ 5% CO2After 4h of incubation in the incubator, the supernatant was aspirated, 150 μ L DMSO was added per well, shaken on a shaker for 10min to complete the solubilization of the formazan crystals, absorbance was measured at 490nm, and cell viability was calculated.
The results of the experiment are shown in figure 1,*p<0.05,**p<0.01 or***p<0.001 and the blank group, the result shows that the preferred compound 27 can obviously inhibit SK-OV-3 cell proliferation at lower concentration, and the effect is better than that of the marketed drug olaparib.
EXAMPLE 48 in vivo antitumor assay for detection of preferred Compound 27
Recovering SK-OV-3 cells, and amplifying the cells according to a conventional cell culture method. When the cells have risen to the exponential phase, the cells are collected and prepared to 5X 108cell/mL, tumor cell suspension preparation in sterile conditions (clean bench). 0.3mL of the prepared tumor cell suspension is inoculated under the right axilla of the mouse, and the inoculation is completed within 60 min. When the tumor volume reaches 100mm3Grouping and administration are performed.
Animal grouping: blank group, Olaparib group (Olaparib,10mg/kg), Compound 27 Low dose group(25mg/kg) and Compound 27 high dose group (50 mg/kg). Measuring the tumor diameter every two days, measuring the tumor short diameter a and the tumor long diameter b (including the skin thickness) by using a vernier caliper, and calculating the tumor volume V as a2X b/2, and a tumor growth curve is plotted. On the last day of the experiment, the tumors were detached and recorded by photography.
The experimental result is shown in figure 2, and the result shows that the compound 27 can obviously inhibit the growth of SK-OV-3 tumor cells under the skin of a tumor-bearing mouse under a lower dose, and the effect is superior to that of the drug olapanil on the market under the equal dose.
EXAMPLE 49 in vivo chemosensitization potency assay for preferred Compound 27
After the A549 cells are recovered, the cells are expanded according to a conventional cell culture method. When the cells have risen to the exponential phase, the cells are collected and prepared to 5X 108cell/mL, tumor cell suspension preparation in sterile conditions (clean bench). 0.3mL of the prepared tumor cell suspension is inoculated under the right axilla of the mouse, and the inoculation is completed within 60 min. When the tumor volume reaches 100mm3Grouping and administration are performed.
Animal grouping: blank, carboplatin + Olaparib (Olaparib,10mg/kg), carboplatin + compound 27 low dose (25mg/kg), and carboplatin + compound 27 high dose (50 mg/kg). Measuring the tumor diameter every two days, measuring the tumor short diameter a and the tumor long diameter b (including the skin thickness) by using a vernier caliper, and calculating the tumor volume V as a2X b/2, and a tumor growth curve is plotted. On the last day of the experiment, the tumors were detached and recorded by photography.
The experimental result is shown in fig. 3, and the result shows that the compound 27 is preferable under a lower dosage, so that the inhibition effect of the carboplatin on the tumor can be obviously increased, and the effect under the equal dosage is superior to that of the marketed drug olaparib.
EXAMPLE 50 in vivo test of antitumor Effect and chemotherapeutic sensitivity enhancing Effect of Compound 25
Cell inoculation and animal groups were the same as in examples 48 and 49.
The experimental results are shown in fig. 4, and the results show that the compound 25 can also obviously increase the inhibition effect of carboplatin on the tumor growth at lower doses, but the effect is weaker than that of the preferred compound 27 and stronger than that of the marketed drug olaparib at equal doses.
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