阅读说明：本技术 用于治疗囊肿状纤维化的化合物及其治疗方法 (Compounds and methods for treating cystic fibrosis ) 是由 M·P·扎维斯托斯基 C·奥尔曼 F·李 A·科尔德齐 M·莫宁斯塔 于 2020-01-28 设计创作，主要内容包括：本发明涉及式I化合物,包括式I化合物的药物组合物,及其药学上可接受的盐,包括这种化合物的药物组合物以及治疗囊肿性纤维化的方法,所述方法包括向需要的患者给药治疗有效量的式I化合物的步骤。(The present invention relates to compounds of formula (I),)

1. A compound of formula (I)

Or a pharmaceutically acceptable salt thereof, wherein:

r and R₁Independently selected from hydrogen, optionally substituted alkyl; optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl or optionally substituted heteroarylalkyl;

or R and R₁Together with the nitrogen atom to which they are attached, form an optionally substituted 3-to 7-membered heterocyclyl;

R₂is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted aralkyl, optionally substituted heteroaryl or optionally substituted heteroaralkyl;

R₃is hydrogen, optionally substituted alkyl, R₇C(O)-、R₇SO₂-or R₇NHC(O)-；

Or R₂And R₃Together with the atoms to which they are attached, form an optionally substituted 3-to 7-membered heterocyclyl;

each R₄Independently halogen, optionally substituted alkyl, CN, optionally substituted alkoxy, NR₁₂R₁₃Or a hydroxyl group;

R₅is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl or optionally substituted cycloalkyl;

R₆is OR₈、SR₈Or NR₉R₁₀；

R₇Is optionally substituted alkyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted aralkyl;

R₈is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted heteroaryl;

R₉is hydrogen, OR₁₁Optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl, optionally substituted heteroaryl, heterocyclyl, SO₂-R₈、SO₂NR_aR_bOr N (R)_a)R_b；R₁₀Is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted heteroaryl;

or R₉And R₁₀Together with the nitrogen atom to which they are attached, form an optionally substituted heterocyclyl;

R_aand R_bIndependently selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted heteroaryl;

R₁₁is hydrogen or optionally substituted alkyl;

R₁₂and R₁₃Each independently selected from hydrogen, optionally substituted alkyl, R₇C(O)-、R₇SO₂-or R₇NHC(O)-；

Or R₁₂And R₁₃Together with the nitrogen atom to which they are attached, form an optionally substituted heterocyclic group, and

n is 0, 1,2,3 or 4.

2. The compound of claim 1, wherein R₁Is optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aralkyl or optionally substituted heteroaralkyl; preferred are optionally substituted phenyl or optionally substituted 6-membered heteroaryl.

3. A compound according to claim 1 or 2, wherein R is hydrogen, optionally substituted C₁-C₆-an alkyl group; optionally substituted C₃-C₈-a cycloalkyl group.

4. A compound according to claim 3, wherein R is hydrogen, methyl, ethyl, n-propyl, isopropyl, C₃-C₆-cycloalkyl, C₃-C₆-cycloalkylmethyl, 2-dimethylaminoethyl or 3-hydroxycyclobutyl.

5. The compound according to any one of claims 1-4, wherein R₂Is hydrogen, optionally substituted C₁-C₆Alkyl, optionally substituted aryl-C₁-C₆-alkyl or optionally substituted heteroaryl-C₁-C₆-an alkyl group.

6. The compound of claim 5, wherein R₂Is hydrogen, C₁-C₄-alkyl, halo-C₁-C₄-alkyl, optionally substituted arylmethyl or optionally substituted heteroarylmethyl.

7. The compound of claim 6, wherein R₂Is hydrogen, C₁-C₄-alkyl, halo-C₁-C₄-alkyl, aryl optionally substituted with 1 to 5 halogens or aryl optionally substituted with 1 to 5 halogens-C₁-C₂-an alkyl group.

8. The compound of claim 7, wherein R₂Is hydrogen, benzyl, optionallySubstituted phenyl-CF₂-, optionally substituted heteroaryl-CF₂-、CF₃、CF₃CH₂-or isopropyl.

9. A compound according to any one of claims 1 to 8, wherein R₃Is hydrogen, C₁-C₄-alkyl, halo-C₁-C₄-alkyl radical, C₁-C₄Alkyl C (O) -, aryl C₁-C₄Alkyl C (O) -, aryl C₁-C₄Alkyl S (O)₂-, aryl-C₁-C₄Alkyl NHC (O) -, or aryl NHC (O) -.

10. The compound of claim 9, wherein R₃Is hydrogen, methyl, CF₃CH₂-, acetyl, propionyl, phenethyl C (O) -, phenethyl SO₂-, phenylmethylNHC (O) -or phenylNHC (O) -.

11. A compound according to any one of claims 1 to 10, wherein R₂And R₃At least one of which is hydrogen.

12. A compound according to any one of claims 1 to 4, wherein R₂And R₃Together with the atoms to which they are attached, form an optionally substituted saturated 4-to 6-membered heterocyclyl.

13. The compound of claim 12, wherein saturated 4-to 6-membered heterocyclyl is unsubstituted or optionally substituted with one or more substituents independently selected from halogen, CN, hydroxy, C₁-C₃-alkoxy, halo-C₁-C₃-alkoxy, C₁-C₃-alkyl, halo-C₁-C₃-alkyl, spirocyclic cycloalkyl, spiroheterocyclic group or optionally substituted C₁-C₃-an alkylene group.

14. The compound of claim 1, represented by formula II,

or a pharmaceutically acceptable salt thereof, wherein

m is 0, 1,2 or 3; and is

Each R₁₄Independently is hydroxy, protected hydroxy, cyano, amino, protected amino, halogen, optionally substituted alkoxy, optionally substituted alkyl SO₂-, optionally substituted alkyl C (O) -or optionally substituted alkyl C (O) NH-; or

Two adjacent R₁₄A group which, together with the carbon atom to which they are attached, forms an optionally substituted 3-to 7-membered carbocyclic or heterocyclic group; or

Two paired R14 groups, together with the carbon atom to which they are attached, form an optionally substituted spiro 3-to 7-membered carbocyclic or heterocyclic group; or

Two paired R₁₄The radicals together form (R)₁₃)₂Wherein each R is₁₃Independently is halogen, C₁-C₄-alkyl or halo-C₁-C₄-an alkyl group.

15. The compound of claim 1, represented by formula III,

wherein X is O or C (R)_a)₂；

Each R₁₄Independently is hydroxy, protected hydroxy, cyano, amino, protected amino, halogen, optionally substituted alkoxy, optionally substituted alkyl SO₂-, optionally substituted alkyl C (O) -or optionally substituted alkyl C (O) NH-; or

Two adjacent R₁₄A group which, together with the carbon atom to which they are attached, forms an optionally substituted 3-to 7-membered carbocyclic or heterocyclic group; or

Two paired R₁₄A group which, together with the carbon atom to which they are attached, forms an optionally substituted spiro 3-to 7-membered carbocyclic or heterocyclic group; or

Two paired R₁₄The radicals together form (R)₁₃)₂Wherein each R is₁₃Independently of one another are halogen, C₁-C₄-alkyl or halo-C₁-C₄-an alkyl group; and

each R_aIndependently is hydrogen, hydroxy, protected hydroxy, cyano, amino, protected amino, halogen, optionally substituted alkoxy, or optionally substituted alkyl.

16. The compound according to any one of claims 1-15, wherein n is 0 to 2.

17. The compound of any one of claims 1-16, wherein each R is₄Is a halogen.

18. The compound of claim 17, wherein n is 2 and each R₄Is chlorine.

19. The compound of claim 16, wherein

Is thatOr

20. The method of claim 19A compound of (1), whereinIs that

21. The compound of claim 14, represented by formula IV,

or a pharmaceutically acceptable salt thereof, wherein p is 0, 1 or 2.

22. The compound of claim 21, wherein p is 1 or 2, and each R is₁₄Independently selected from amino, protected amino, cyano, hydroxy, fluoro, chloro, C₁-C₄-alkoxy, halo-C₁-C₄-alkoxy, C₁-C₄-alkyl SO₂、C₁-C₄Alkyl radicals C (O), C₁-C₄Alkyl C (O) NH-and C₁-C₄-an alkyl group optionally substituted with one or more substituents independently selected from the group consisting of hydroxy, fluoro, chloro and amino; or

p is 2, and two R₁₄The radicals together with the adjacent carbon atoms forming a spiro ring C₃-C₆-cycloalkyl or spiro-3 to 6-membered heterocycloalkyl; or

p is 2, and two R₁₄The radicals together form (R)₁₃)₂Wherein each R is₁₃Independently of one another are hydrogen, fluorine, chlorine, methyl, CF₃Or CHF₂。

23. The compound of claim 22, wherein p is 2.

24. The compound of claim 21, wherein each R₁₄Independently selected from hydroxy, methyl, fluorine, chlorine, amino, CH₃SO₂-、CF₃、CHF₂、CH₂F. Cyano, acyl oxide or amide.

25. The compound of claim 21, wherein

Two R₁₄The groups together with their adjacent carbon atoms form a spiro-cyclopropyl group; or

Two R₁₄The radicals together forming F₂C＝。

26. A compound according to any one of claims 1 to 25, wherein R₁Is optionally substituted phenyl or optionally substituted 5-or 6-membered heteroaryl.

27. The compound of claim 26, wherein R₁Is phenyl, pyridyl, pyrazinyl, pyrimidinyl, pyrazolyl, oxazolyl, oxadiazolyl, thiazolyl, thiadiazolyl, or triazolyl, each of which may be optionally substituted.

28. A compound of claim 26 or 27, wherein R₁Is substituted by 1 to 3 substituents independently selected from C₁-C₄-alkyl, halo-C₁-C₄Alkyl, halogen, C₁-C₄-alkoxy and halo-C₁-C₄-alkoxy groups.

29. The compound of claim 28, wherein the substituents are selected from methyl, methoxy, fluorine, chlorine, CHF₂、CF₃、CHF₂O-and CF₃O-。

30. The compound of claim 27, wherein R₁Selected from the following groups:

31. a compound of claim 26, wherein R₁Is shown as

Wherein X₁-X₄Each independently is N or CR₁₇Wherein R is₁₇Is hydrogen, optionally substituted alkyl, optionally substituted alkoxy or halogen.

32. The compound of claim 31, wherein R1 is selected from the group consisting of:

33. the compound of claim 26, wherein R₁Is shown as

Wherein, Y₁，Y₂，Y₃And Y₄Is O, S or NR₁₆And others independently of each other are N or CR₁₇Wherein R is₁₆Is hydrogen, optionally substituted alkyl, R₇C(O)-，R₇SO₂-or R₇NHC(O)-；R₁₇Is hydrogen, optionally substitutedAlkyl, optionally substituted alkoxy or halogen; and is

R7 is optionally substituted alkyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted aralkyl.

34. The compound of claim 33, wherein Y₃Is C-CF₃，Y₁，Y₂And Y₄Is O, S or NR₁₆And others independently of each other are N or CR₁₇。

35. The compound of claim 33, wherein R₁Selected from the following groups:

36. a compound selected from the group of compounds listed in the following table, or a pharmaceutically acceptable salt thereof,

37. a pharmaceutical composition comprising a compound of any one of claims 1-36 and a pharmaceutically acceptable carrier or excipient.

38. A method of treating a disease or disorder mediated by cystic fibrosis transmembrane conductance regulator (CFTR) in a patient in need thereof, comprising administering to the patient a compound of any one of claims 1 to 36.

39. The method of claim 38, wherein the disease or disorder is selected from the group consisting of: cystic fibrosis, hereditary emphysema, hereditary hemochromatosis, coagulation-fibrinolysis deficiency, hereditary angioedema type 1, lipid processing deficiencies, e.g. hyperlipoproteinemia, chylomicronemia type 1, chylomicronemia, lysosomal storage diseases, e.g. mucolipidosis type II/mucopolysaccharidosis, gangliosidosis/tay-saxose, crigler-najal type II, polyendocrinosis/hyperinsulinemia, diabetes, larch dwarfism, myeloperoxidase deficiency, primary hypoparathyroidism, melanoma, glucocorticoid disease type CDG 1, congenital hyperthyroidism, osteogenesis imperfecta, hereditary hypofibrinogenemia, ACT deficiency, Diabetes Insipidus (DI), neurovegetative diabetes insipidus (Neurosporaldi), Nephrogenic diabetes insipidus (neprogenicicdi), progressive neuromuscular atrophy (Charcot-MarieTooth) syndrome, pelizaeus-merzbacher disease, neurodegenerative diseases such as alzheimer's disease, parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, Pick's disease, various polyradical amine nervous system diseases such as Huntington's chorea, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentatorubral pallidoluysian atrophy (dentorubal pallidoluysian) and myotonic dystrophy, and spongiform encephalopathies (spongiform encyclopaedies), such as hereditary Creutzfeldt-jakob disease, jafarebriesis, schwerner's syndrome (Straussler-Straussler syndrome), Chronic Obstructive Pulmonary Disease (COPD), chronic obstructive pulmonary disease such as COPD, Chronic Obstructive Pulmonary Disease (COPD), or congenital ductal disease, Disseminated bronchiectasis, allergic pulmonary aspergillosis, chronic sinusitis, protein C deficiency, a-lipoproteinemia, mild pulmonary disease, lipid management deficiency, thrombofibrinolysis, CFTR-associated metabolic syndrome, chronic bronchitis, constipation, pancreatic insufficiency, melanoma, glycosylated CDG type 1, ACT deficiency, and cholestatic liver disease.

40. A method of treating a disease or disorder mediated by cystic fibrosis transmembrane conductance regulator (CFTR) in a patient in need thereof, comprising administering to the patient a therapeutically effective amount of a compound of any one of claims 1 to 36.

41. The method according to any one of claims 38 to 40, further comprising administering to a patient in need thereof a therapeutically effective amount of a compound that is a CFTR modulator, a viscous solution, or an antibiotic.

Background

Cystic Fibrosis (CF) is a fatal, progressive, invisible genetic disease that affects every 2500 individuals among caucasians. (Cohen-Cymberknoh, M.et al., am.J.Respir.Crit.Care Med.1463-1471, 2011; Boat et al., The Metabolic Basis of Inherited diseases, 6th ed., pp 2649-. Approximately 1 out of 25 individuals are vectors for genetic defects associated with this disease. The main symptoms of cystic fibrosis include chronic lung disease, insufficient pancreatic exocrine secretion, loss of male hair and increased sweat electrolyte levels. The symptoms are consistent with cystic fibrosis, an exocrine abnormality. (Hantashf: U.S. patent application No. 20060057593).

The Cystic Fibrosis (CF) gene encoding the cAMP/PKA-dependent, ATP-requiring, membrane-bound chloride pathway is called CFTR (cystic fibrosis transmembrane conductance regulator) and is usually localized to the apical membrane of many secretory organs. Over 1900 mutations are currently known to affect cystic fibrosis transmembrane conductance regulator (CFTR), many of which contribute to the disease phenotype. Approximately 75% of Cystic Fibrosis (CF) alleles contain the Δ F508 mutation, in which the triplet codon is lost, resulting in a deletion of phenylalanine at position 508 of the protein. Such altered proteins are not trafficked in the correct location in the cell and are usually destroyed by the proteasome. The small amount of the above proteins that can reach the correct site is clearly insufficient in function. (Cuthbert AW, British Journal of Pharmacology, 163(1), 173-.

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene result in the loss of or cause dysfunction of proteins that regulate ion transport across certain epithelial surface apical membranes. While the cystic fibrosis transmembrane conductance regulator (CFTR) functions primarily as a chloride pathway, there are other effects, including inhibiting sodium transport through the epithelial sodium channel, regulating the externally corrected chloride pathway, the ATP pathway, intracellular vesicle transport, and inhibiting endogenous calcium-activated chloride pathways. Cystic fibrosis transmembrane conductance regulator (CFTR) is also involved in the bicarbonate-chloride exchange process. The lack of bicarbonate secretion results in poor solubility and aggregation of luminal mucins. Blockage of the pancreatic endothelium by thickened secretions results in autolysis of the pancreatic tissue, replacement of the pancreas by fat, leading to pancreatic insufficiency and subsequent malnutrition. In the lung, cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction leads to depletion of airway surface fluid (ASL) and thickening of mucus attached to the airway surface. This results in decreased mucociliary clearance (MCC) and decreased host resistance. The dried viscous secretions cause infections of the bronchial interior with specific bacteria, mainly staphylococcus aureus and pseudomonas aeruginosa, and loss of bicarbonate secretion due to loss of cystic fibrosis transmembrane conductance regulator (CFTR) function results in a decrease in airway surface pH, thereby impairing the anti-bacterial lethal activity and increasing susceptibility to infection. The severe inflammatory response resulting from chronic lung infection can lead to bronchodilation and progressive obstructive airway disease. Pulmonary insufficiency causes the vast majority of Cystic Fibrosis (CF) -related deaths. (Cohen-Cymberknoh, m.et al., am.j.respir.crit.care med.1463-1471, 2011).

In the last 40 years, the prognosis for the treatment of Cystic Fibrosis (CF) has improved. This is achieved by improving pancreatic enzyme supplementation, designing drugs to treat pulmonary infections, reducing inflammation, and increasing mucociliary clearance. Current therapeutic agents challenge the biochemical defect for correcting Cystic Fibrosis (CF) and identify an effective treatment for chronic respiratory infections. (Frericchs C.et al, Expert Opin Pharmacother.10(7), 1191-.

Disclosure of Invention

In one embodiment, the present invention relates to a compound of formula (I)

Or a pharmaceutically acceptable salt thereof, wherein,

r and R₁Independently selected from hydrogen, optionally substituted alkyl; optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl, optionally substituted aralkyl, optionally substituted heteroaryl, or optionally substituted heteroaralkyl;

or, R and R₁Together with the nitrogen atom to which they are attached, form an optionally substituted 3-to 7-membered heterocyclyl;

R₂is hydrogen, optionally substituted alkyl, optionally substituted aryl, optionally substituted aralkyl, optionally substituted heteroaryl or optionally substituted heteroaralkyl; in certain embodiments, R₂Is hydrogen, optionally substituted alkyl, optionally substituted aryl or optionally substituted aralkyl;

R₃is hydrogen, optionally substituted alkyl, R₇C(O)-、R₇SO₂-or R₇NHC(O)-；

Or R₂And R₃Together with the atoms to which they are attached, form an optionally substituted 3-to 7-membered heterocyclyl;

each R₄Independently halogen, optionally substituted alkyl, CN, optionally substituted alkoxy, NR₁₂R₁₃Or a hydroxyl group;

R₅is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, or optionally substituted ringAn alkyl group;

R₆is OR₈Or NR₉R₁₀(ii) a Or R₆is-SR₈；

R₇Is optionally substituted alkyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted aralkyl;

R₈is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted heteroaryl;

R₉is hydrogen, OR₁₁Optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted heteroaryl; or R₉Is optionally substituted heterocyclyl, SO₂R₈、SO₂NR_aR_bOr N (R)_a)R_b；

R_aAnd R_bEach independently is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted heteroaryl;

R₁₀is hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted aryl or optionally substituted heteroaryl;

or R₉And R₁₀Together with the nitrogen atom to which they are attached, form an optionally substituted heterocyclyl;

R₁₁is hydrogen or optionally substituted alkyl;

R₁₂and R₁₃Independently of one another, from hydrogen, optionally substituted alkyl, R₇C(O)-、R₇SO₂-or R₇NHC(O)-；

Or R₁₂And R₁₃Together with the nitrogen atom to which they are attached, form an optionally substituted heterocyclyl; and is

n is 0, 1,2,3 or 4; preferably, n is 1 or 2.

In other embodiments, the present invention relates to a pharmaceutical composition comprising a compound of formula (I) and pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable carrier or excipient.

In another embodiment, the invention relates to a method of treating a CFTR mediated disease or disorder (e.g., cystic fibrosis) in a subject in need thereof comprising administering to the subject a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof.

Detailed description of the preferred embodiments

The present invention relates to compounds of formula (I) and pharmaceutically acceptable salts thereof, pharmaceutical compositions comprising the compounds, and methods of using the compounds to treat CFTR-mediated diseases or disorders in a subject in need thereof.

In certain embodiments, the compounds of the present invention have the absolute stereochemistry shown in formula (Ia) or formula (Ib).

In certain embodiments of the compounds of the present invention, R₁Is optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aralkyl, e.g. optionally substituted aryl-C₁-C₆-an alkyl group; or optionally substituted heteroaralkyl, e.g. heteroaryl-C₁-C₆-an alkyl group; preferred are optionally substituted phenyl or optionally substituted 6-membered heteroaryl.

In certain embodiments of the compounds of the present invention, R is hydrogen, optionally substituted C₁-C₆-an alkyl group; optionally substituted C₃-C₈-a cycloalkyl group; in certain embodiments, R is hydrogen, methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, sec-butyl, n-pentyl, neopentyl, optionally substitutedSubstituted C₃-C₆Cycloalkyl, optionally substituted C₃-C₆-cycloalkylmethyl, 2-dimethylaminoethyl, or 3-hydroxycyclobutyl. In certain embodiments, R is optionally substituted C₃-C₁₂-cycloalkyl-C₁-C₆Alkyl, preferably optionally substituted C₃-C₁₂-cycloalkyl-methyl. In certain embodiments, R is hydrogen or C₁-C₆Alkyl, for example, hydrogen or methyl. In certain embodiments, R is a branched chain C₃-C₁₀Alkyl, preferably branched C₃-C₈-an alkyl group. In certain embodiments, R is a beta-branched C₄-C₁₀Alkyl radicals, such as the 2,2,3,3, -tetramethylbutyl radical or the 2,2, -dimethylpropyl radical.

In certain embodiments of the compounds of the present invention, R₂Is hydrogen, optionally substituted C₁-C₆Alkyl, optionally substituted aryl-C₁-C₆-alkyl, or optionally substituted heteroaryl-C₁-C₆-an alkyl group. In certain embodiments, R₂Is hydrogen, C₁-C₄-alkyl, halo-C₁-C₄-alkyl, optionally substituted arylmethyl, or optionally substituted heteroarylmethyl. In certain embodiments, R₂Is hydrogen, benzyl, optionally substituted phenyl-CF₂-, optionally substituted heteroaryl-CF₂-, benzyl-O-CH₂-、CF₃、CF₃CH₂-or isopropyl. In certain embodiments, R₂Is hydrogen, C₁-C₄-alkyl, halo-C₁-C₄-alkyl, aryl optionally substituted with 1-5 halogens or aryl optionally substituted with 1-5 halogens-C₁-C₂-an alkyl group. In certain embodiments, R₂Is hydrogen, CF₃Isopropyl, benzyl-O-CH₂-, 3-hydroxy-n-propyl, or α, α -difluorobenzyl.

In certain embodiments of the compounds of the present invention, R₃Is hydrogen, C₁-C₄-alkyl, halo-C₁-C₄-alkanesBase, C₁-C₄Alkyl C (O) -, aryl C₁-C₄Alkyl C (O) -, aryl C₁-C₄Alkyl S (O)₂-, aryl-C₁-C₄Alkyl NHC (O) -, or aryl NHC (O) -. In certain embodiments, R₃Is hydrogen, methyl, CF₃CH₂-, acetyl, propionyl, phenethyl C (O) -, phenethyl SO₂-, phenylmethylNHC (O) -or phenylNHC (O) -.

In certain embodiments of the compounds of the present invention, R₂And R₃Is hydrogen.

In certain embodiments, R₂And R₃Together with the atoms to which they are attached, form an optionally substituted saturated 4 to 6-membered heterocyclic group, preferably an optionally substituted saturated 5-membered heterocyclic group, and more preferably an optionally substituted pyrrolidine. In certain embodiments, R₂And R₃Together with the atom to which they are attached, form an optionally substituted saturated 6-membered heterocyclyl group, for example, an optionally substituted piperidinyl group or an optionally substituted morpholinyl group. In certain embodiments, a saturated 4-to 6-membered heterocyclyl is unsubstituted or substituted with one or more substituents independently selected from halogen, CN, hydroxy, C₁-C₃-alkoxy, halo-C₁-C₃-alkoxy, C₁-C₃-alkyl, halo-C₁-C₃-alkyl, spirocycloalkyl, spiroheterocyclyl or optionally substituted C₁-C₃-an alkylene group.

In certain embodiments of the compounds of the present invention, each R is₄Are independently of one another halo, for example chloro or fluoro.

In certain embodiments of the compounds of the present invention, R₅Is hydrogen or C₁-C₆-an alkyl group; preferably hydrogen or methyl;

in certain embodiments of the compounds of the present invention, R₆Is OR₈And R is₈Is hydrogen, optionally substituted C₁-C₁₀-alkyl or optionally substitutedC of (A)₂-C₁₀-alkenyl. In certain embodiments, R₈Is hydrogen or optionally substituted C₁-C₁₀-an alkyl group. In certain embodiments, R₈Is hydrogen, C₁-C₄-alkyl or allyl. In certain embodiments, R₈is-CH₂-O-R_cWherein R is_cis-C (O) -C₁-C₈-alkyl or

In certain embodiments of the compounds of the present invention, R₆Is NR₉R₁₀. In certain embodiments, R₉And R₁₀Are all C₁-C₄-alkyl, preferably methyl. In certain embodiments, R₉Is OH or O-C₁-C₂-alkyl, preferably methyl, and R₁₀Is hydrogen or C₁-C₃-alkyl, preferably hydrogen or methyl. In certain embodiments, R₉Is SO₂R₈Or SO₂NR_aR_b. In certain embodiments, R₉is-SO₂-C₁-C₄-alkyl, -SO₂-phenyl, -SO₂NH₂or-SO₂N(CH₃)₂。

In certain embodiments of the compounds of the present invention, R₁Is optionally substituted aryl or heteroaryl, preferably optionally substituted phenyl or optionally substituted 6-membered heteroaryl; r is hydrogen, C₁-C₈-alkyl or C₁-C₆-an alkyl group; preference is given to hydrogen, methyl or beta-branched C₄-C₁₀-an alkyl group; r₅Is hydrogen or C₁-C₆-an alkyl group; preferably hydrogen or methyl; and R₆Is OR₈And R is₈Is hydrogen, or optionally substituted C₁-C₁₀-an alkyl group; or R₈Is hydrogen, optionally substituted C₁-C₁₀-an alkyl group; or optionally substitutedC of (A)₂-C₆-alkenyl.

In certain embodiments, the compounds of formula (I) may be represented by formula (II),

wherein m is 0, 1,2,3, 4,5 or 6; and

wherein R is₁₄Independently of one another, is hydroxy, protected hydroxy, cyano, amino, protected amino, halogen, optionally substituted alkoxy, or optionally substituted alkyl; or

Two adjacent R₁₄A group which, together with the carbon atom to which they are attached, forms an optionally substituted 3-to 7-membered carbocyclic or heterocyclic group; or

Two paired R₁₄A group which, together with the carbon atom to which they are attached, forms an optionally substituted spiro 3-to 7-membered carbocyclic or heterocyclic group; or

Two paired R₁₄The radicals together form (R)₁₅)₂Wherein each R is₁₅Independently of one another are hydrogen, halogen, C₁-C₄-alkyl or halo-C₁-C₄-an alkyl group. In certain embodiments of the compounds of formula II, m is 0 or 2. In certain embodiments, m is 2 and two R are₁₄The groups are attached to the same carbon atom.

In certain embodiments, the compound of formula (II) has the absolute stereochemistry shown in formula (IIa) or formula (IIb).

In certain embodiments, the compounds of formula (I) may be represented by formula (III),

wherein X is O or C (R)_a)₂And each R is_aEach independently is hydrogen, hydroxy, protected hydroxy, cyano, amino, protected amino, halogen, optionally substituted alkoxy, or optionally substituted alkyl.

In certain embodiments, the compound of formula (III) has the absolute stereochemistry shown in formula (IIIa) or formula (IIIb).

In certain embodiments, the compounds of formula (I) may be represented by formula (IV),

or a pharmaceutically acceptable salt thereof, wherein R₁₄Is as defined above and p is 0, 1 or 2.

In certain embodiments, the compound of formula (IV) has the absolute stereochemistry shown in formula (IVa) or formula (IVb).

In certain embodiments, the compounds of formula (I) may be represented by formula (V),

or a pharmaceutically acceptable salt thereof, wherein R₁₄And p is as defined above.

In certain embodiments, the compound of formula (V) has the absolute stereochemistry shown in formula (Va) or formula (Vb).

In certain embodiments of the compounds of the present invention, R₁Is an optionally substituted aryl group, or an optionally substituted 5-or 6-membered heteroaryl group, for example, phenyl, naphthyl, pyridyl, pyrazinyl, pyrimidinyl, pyrazolyl, oxazolyl, oxadiazolyl, thiazolyl, thiadiazolyl, triazolyl or pyrrolyl. In certain embodiments, R₁Is an optionally substituted fused bicyclic heteroaryl group, for example, quinolinyl, quinazolinyl, naphthyl, benzimidazolyl, isoquinolinyl, pyrazolyl, benzothiazolyl, naphthol, indolyl or indolyloxyl. In certain embodiments, R₁Is optionally substituted phenyl-C₁-C₆-alkyl, optionally substituted heteroaryl-C₁-C₆-alkyl, or optionally substituted diaryl groups, for example, optionally substituted diphenyl, phenylheteroaryl or heteroarylphenyl, including phenylpyrazine.

Preferably, R₁Is unsubstituted or substituted by 1,2 or 3 members independently selected from C₁-C₄-alkyl, halo-C₁-C₄Alkyl, halogen, C₁-C₄-alkoxy and halo-C₁-C₄-substituent substitution of alkoxy. More preferably, the substituents are independently selected from methyl, methoxy, fluoro, chloro, methoxy, CHF₂、CF₃、CHF₂O-and CF₃O-。

In certain embodiments, R₁Selected from the group consisting of:

in certain embodiments, R₁Is shown as

Wherein, X₁-X₄Each independently is N or CR₁₇Wherein each R is₁₇Are independent of each otherAnd is hydrogen, optionally substituted alkyl, optionally substituted alkoxy or halogen. In certain embodiments, each R is₁₇Independent of each other H, CF₃、CH₃、OCH₃、OCF₃Or a halogen. Preferably, X₁，X₂，X₃And X₄No more than two of which are N. More preferably, X₁，X₂，X₃And X₄No more than one of which is N.

In certain embodiments, R₁Selected from the group as follows:

in certain embodiments of the compounds of the present invention, R₁Is shown as

Wherein, Y₁，Y₂，Y₃And Y₄Is O, S or NR₁₆And others independently of each other are N or CR₁₇Wherein R is₁₆Is hydrogen, optionally substituted alkyl, R₇C(O)-、R₇SO₂-or R₇NHC (O) -and R₁₇Is hydrogen, optionally substituted alkyl, optionally substituted alkoxy, CN or halogen. Preferably, R₁₆Is hydrogen or methyl. Preferred R₁₇Is H; CF (compact flash)₃；CN；C₁-C₄Alkyl radicals, e.g. CH₃；OCH₃；OCF₃Or a halogen. Preferably, Y is₁To Y₄Is at least one of CR₁₇. In certain embodiments, Y₃Is C-CF₃，Y₁，Y₂And Y₄Is O, S or NR₁₆And others independently of each other are N or CR₁₇. In certain embodiments, R₁Selected from the group as follows:

in certain embodiments of the compounds of formula II,

selected from the group shown below:

in other embodiments of the compounds of formula II,

selected from the group shown below:

in a preferred embodiment of the compound of formula II,

selected from the group shown below:

in certain embodiments of the compounds of the present invention,

selected from the group shown below:

representative compounds of the invention include the compounds listed in the following table and pharmaceutically acceptable salts thereof.

In the compounds shown here, stereoisomers are not shown, and preferably the compounds are stereoisomers, having the absolute stereochemistry shown in formulae (Ia), (IIa), (IIIa), (IVa) and (Va) or formulae (Ib), (IIb), (IIIb), (IVb) and (Vb). In certain embodiments, preferred stereoisomers have the absolute stereochemistry shown in formulae (Ia), (IIa), (IIIa), (IVa), and (Va).

The compounds of the invention may be useful as modulators of cystic fibrosis transmembrane conductance regulator (CFTR) and for treating diseases or disorders modulated by cystic fibrosis transmembrane conductance regulator (CFTR). Accordingly, the present invention provides a method of treating a cystic fibrosis transmembrane conductance regulator (CFTR) modulated disease or disorder in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound of the invention. Diseases or disorders modulated by cystic fibrosis transmembrane conductance regulator (CFTR) include cystic fibrosis, hereditary emphysema, hereditary hemochromatosis, coagulation-fibrinolysis deficiency, hereditary angioedema type 1, lipid processing deficiencies, e.g., hyperlipoproteinemia, chylomicronemia type 1, abetalipoproteinemia, lysosomal storage diseases, e.g., mucolipidosis type II/mucopolysaccharidosis, gangliosidosis/Tay-saxopathy, Crigler-Najal type II disease, polyendocrinopathy/hyperinsulinemia, diabetes, Raynaud's dwarfism, myeloperoxidase deficiency, primary hypoparathyroidism, melanoma, Gl-type glycosylation disease, congenital hyperthyroidism, osteogenesis imperfecta, hereditary hypofibrinogenemia, diabetes, Crohn's disease, chronic myelofibrosis, chronic myelogenous leukemia, ACT deficiency, Diabetes Insipidus (DI), neurophysiologic diabetes insipidus (NeprongicDI), progressive neuromuscular atrophy (Charcot-Marie Tooth) syndrome, Papy-Mei disease (Perlizeus-Merzbacher disease), neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, Pick's disease, several polyglutamine neurological disorders such as Huntington's chorea, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentatorubral erythronuclear pallidoluysian atrophy (Dentalpallidoluysian) and myotonic dystrophy, and spongiform encephalic diseases such as hereditary Ochropathy-Creutzfeldt-Jatbakolkulare syndrome, Chronic obstructive pulmonary disease (COPD-Chronic obstructive pulmonary disease), Chronic obstructive pulmonary disease, Parkinson's syndrome, and other, Dry eye or sjogren's syndrome, congenital bilateral vasectomy (CBAVD), disseminated bronchiectasis, allergic pulmonary aspergillosis, chronic sinusitis, protein C deficiency, a-lipoproteinemia, mild pulmonary disease, lipid processing deficiencies, thrombofibrinolysis, CFTR-related metabolic syndrome, chronic bronchitis, constipation, pancreatic insufficiency, melanoma, glycosylated CDG type 1, ACT deficiency, allergic pulmonary aspergillosis; abdominal cavity diseases; vascular inflammation atherosclerotic disease, increased glucagon production, cholestatic liver disease (such as Primary Biliary Cirrhosis (PBC) and Primary Sclerosing Cholangitis (PSC)).

In certain embodiments, the CFTR-mediated disease is selected from congenital bilateral vas deferens defect; acute, recurrent or chronic pancreatitis; disseminated bronchiectasis; asthma; allergic pulmonary aspergillosis; smoking-related lung diseases (e.g. chronic obstructive pulmonary disease, COPD); dry eye syndrome; sicca syndrome; chronic sinusitis; cholestatic liver diseases such as primary biliary cirrhosis and primary sclerosing cholangitis; polycystic kidney disease (autosomal dominant inheritance).

In certain embodiments, the CFTR mediated disease or disorder is selected from celiac disease; atherosclerotic disease; dry eye (keratoconjunctivitis sicca) with or without autoimmune disease; polycystic kidney disease; cystic fibrosis-associated diabetes; increased glucagon production; non-atopic asthma; non-CF type bronchiectasis; and constipation.

The compounds of the invention may be administered in combination with one or more additional therapeutic agents, such as antibiotics, anti-inflammatory agents, bronchodilators or mucolytics. In particular, antibiotics for the treatment of the bacterium Pseudomonas mucosae may be used in combination with the compounds of the invention. Inhalation antibiotics such as tobramycin, colistin and aztreonam may be used in combination with the treatment of the compounds of the present invention. Anti-inflammatory agents may also be used in combination with the compounds of the present invention to treat CFTR-related diseases. Bronchodilators may be used in combination with the compounds of the present invention to treat CFTR related diseases. In certain embodiments, a therapeutic agent of the invention is administered in combination with a second compound (CFTR modulator).

In one embodiment, the present invention provides a method of treating cystic fibrosis, or a symptom thereof, in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound of the invention. The compounds of the invention are optionally administered in combination with one or more additional agents useful in the treatment of cystic fibrosis, e.g., compounds that are modulators of CFTR expression, activity and/or function. Suitable other agents include, but are not limited to, gentamicin, evakavut (KALYDECO)^TM) Lummatou, Tezakipoto, VX-445PTI-428, PTI-801, PTI-808, GLPG1837, GLPG2222, GLPG2737, FDL169 and FDL 176. In certain embodiments, a compound of the invention is administered in combination with two or more additional CFTR modulators. For example, in one embodiment, the compounds of the invention are administered in combination with FDL169 and/or FDL 176. In one embodiment, the compounds of the invention are administered in combination with both FDL169 and FDL 176. In one embodiment, the invention relates to a pharmaceutical composition comprising a compound of the invention and a pharmaceutically acceptable excipient or carrier. The compositions may include one or more compounds of the invention and a pharmaceutically acceptable carrier, adjuvant, or vehicle. In certain embodiments, these compositions further comprise one or more additional therapeutic agents for treating a CFTR-mediated disease or disorder.

Pharmaceutical composition

The pharmaceutical compositions of the present invention comprise a therapeutically effective amount of a compound of the present invention formulated with one or more pharmaceutically acceptable carriers or excipients.

As used herein, the term "pharmaceutically acceptable carrier or excipient" refers to a non-toxic inert solid, semi-solid, gel or liquid filler, diluent, coating material, or formulation adjuvant of any type. Some examples of carrier materials which may be pharmaceutically acceptable are sugars, such as lactose, glucose and sucrose; cyclodextrins, such as alpha cyclodextrin, beta cyclodextrin, and gamma cyclodextrin; starches such as corn starch and potato starch; cellulose and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelling; talc; excipients, such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols (glycols), such as propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; water without pyrogen; isotonic saline; a Roger's solution; alcohols, and phosphate buffers, as well as other non-toxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the composition, as judged by the formulator.

The pharmaceutical composition of the present invention may be administered by: oral, parenteral, by inhalation spray, topical, rectal, intranasal, buccal, vaginal or via an implanted pump. In a preferred embodiment, administration is by oral administration. Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1, 3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitol, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

The pharmaceutical compositions of the present invention may contain any conventional non-toxic pharmaceutically acceptable carrier, adjuvant or vehicle. In some cases, the pH of the formulation may be adjusted using pharmaceutically acceptable acids, bases or buffers to enhance the stability of the formulated compound or its delivery form. The term parenteral as used herein includes subcutaneous, intradermal, intravenous, intramuscular, intraarticular, intraarterial, intrasynovial, intrasternal, intrathecal, intralesional and intracranial injection techniques or fusion techniques.

In another embodiment, the administration is parenteral administration by injection. Injectable preparations, for example sterile, injectable aqueous or oleaginous suspensions, may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable suspension or emulsion, for example OrOr in a non-toxic parenterally acceptable diluent or solvent, for example, as a solution in 1, 3-butanediol.Is an intravenous fat emulsion, and contains soybean oil 10-30%, yolk phospholipid 1-10%, glycerol 1-10% and water.Also an intravenous fat emulsion contains 2-15% of safflower oil, 2-15% of soybean oil, 0.5-5% of lecithin, 1-10% of glycerin and water.Is an emulsion for transfusion, and contains fish oil 5-25%, lecithin 0.5-10%, glycerol 1-10% and water. Acceptable carriers and solvents include water, ringer's solution, USP, and isotonic sodium chloride solution. In addition, sterile fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil may be employed including synthetic mono-or di-triglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.

The injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by introducing a sterilizing agent in the form of a sterile solid composition which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.

Compositions for rectal or vaginal administration are preferably suppositories which can be prepared by mixing the compounds of the invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectal or vaginal cavity and release the active compound.

Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or: A) fillers or extenders such as starch, lactose, sucrose, glucose, mannitol, and silicic acid, B) binders such as carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and gum arabic, C) wetting agents such as glycerol, D) disintegrating agents such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, E) solution retarding agents such as paraffin, F) absorption promoters such as quaternary ammonium compounds, G) wetting agents such as cetyl alcohol and glycerol monostearate, H) absorbents such as kaolin and bentonite, and I) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures of the foregoing. In the case of capsules, tablets and pills, the dosage form may also include buffering agents.

Solid compositions of a similar type may also be used as fillers in soft-filled as well as hard-filled gel capsules, wherein such excipients are used: lactose and high molecular weight polyethylene glycols and similar agents.

Solid dosage forms such as the tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and may also be of a composition that releases the active ingredient only, or preferentially, in a certain portion of the intestinal tract, optionally in a delayed manner. Examples of embedding compositions that may be used include polymeric substances as well as waxes.

The dosage forms of the compounds of the present invention for topical or transdermal administration include ointments, patches, creams, lotions, gels, powders, solutions, sprays, inhalants or Patches (PATCH). The active ingredient is mixed under sterile conditions with a pharmaceutically acceptable carrier and any necessary preservatives or buffers as may be required. It is also contemplated that ophthalmic formulations, ear drops, eye ointments, powders and solutions are also within the scope of the present invention.

In addition to the active compounds described in the present invention, the ointments, patches, creams and gels may contain excipients such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or mixtures of these substances.

In addition to the compounds described in the present invention, powders and sprays may contain excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicate and polyamide powder, or mixtures of these substances. The spray may additionally contain conventional propellants such as chlorofluorohydrocarbons.

Transdermal patches have the additional advantage of: providing the compound to the body in a controlled delivery manner. Such dosage forms may be prepared by dissolving or dispersing the compound in the appropriate medium. Absorption enhancers may also be used to increase the flux of the compound across the skin. The rate may be controlled by providing a rate controlling membrane or by dispersing the compound in a polymer matrix or gel.

For pulmonary delivery, the therapeutic compositions of the present invention are formulated and administered to the patient in solid or liquid particulate form by direct administration, such as inhalation, into the respiratory system. Solid or liquid particulate forms of the active compounds prepared for the practice of the present invention include particles having respirable sizes: that is, the particles are small enough to pass through the mouth and throat and into the bronchi and alveoli via inhalation. The delivery of aerosolized therapeutic agents, particularly aerosolized antibiotics, is known in the art (see, e.g., U.S. patent No.5767068 to VANDEVANTER et al, U.S. patent No. 5508269 to SMITH et al, and WO98/43650 to MONTGOMERY).

The compositions described herein may be formulated in unit dosage form. The term "unit dosage form" refers to physically discrete units suitable as unitary dosages for the subject being treated, wherein each unit contains a predetermined quantity of active material calculated to produce the desired therapeutic effect, optionally in association with a suitable pharmaceutical carrier. The unit dosage form can be employed in a single daily dose or in one of a plurality of daily doses (e.g., about 1 to 4 or more times per day). When multiple daily doses are employed, the unit dosage form for each dose may be the same or different. The amount of active compound in a unit dosage form will depend, for example, on the subject being treated and the particular mode of administration. In one embodiment, a unit dosage form may have about 10mg, 20mg, 30mg, 40mg, 50mg, 100mg, 150mg, 200mg, 250mg, 300mg, 400mg, 500mg, 600mg, 700mg, 750mg, 800mg, 900mg, 1000mg or 1,250mg of one of the compounds of the invention as an active ingredient.

In some embodiments, a compound of the invention may be administered at a dosage of at least about 10 mg/day to at least about 1500 mg/day. In some embodiments, a compound of the invention is administered at a dose of at least about 300mg (e.g., at least about 450mg, at least about 500mg, at least about 750mg, at least about 1000mg, at least about 1250mg, or at least about 1500 mg).

Dosage adjustments can be made in patients with mild, moderate or severe liver damage (Child-Pugh grade A). In addition, the dosage of patients taking one or more of the cytochrome P450 inhibitors and inducers (in particular CYP3a4, CYP2D6, CYP2C9, CYP2C19 and CYP2B6 inhibitors and inducers) may be adjusted. Dosage adjustments may also be made in patients with impaired cytochrome P450 function, such as in patients with hypometabolism, moderate, extensive and ultra-rapid.

Definition of

Listed below are definitions of various terms used to describe the present invention. Unless otherwise defined in a specific context, such definitions shall apply to any term used throughout this specification and claims, whether such term is used alone or as part of a larger phrase.

The term "alkyl" refers to a saturated aliphatic hydroxyl radical/group having a specified number of carbon atoms, including branched and straight chain, substituted or unsubstituted. Preferred alkyl groups include from about 1 to about 24 carbon atoms ("C)₁-C₂₄"). Other preferred alkyl groups include from about 1 to about 8 carbon atoms ("C)₁-C₈"), for example, from about 1 to about 6 carbon atoms (" C)₁-C₆"), or alternatively, from about 1 to about 3 carbon atoms (" C ")₁-C₃”)。C₁-C₆Examples of alkyl radicals include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, hexyl radicals.

The term "alkenyl" embraces linear or branched radicals having at least one carbon-carbon double bond, with such radicals preferably having from about two to about twenty-four carbon atoms ("C₂-C₂₄"). Other preferred alkenyl radicals are "lower alkenyl" radicals having from two to about ten carbon atoms ("C)₂-C₁₀") for example, vinyl, allyl, propenyl, butenyl and 4-methylbutenyl. The term lower alkenyl radical as referred to herein includes from 2 to about 6 carbon atoms ("C)₂-C₆"). The terms "alkenyl" and "lower alkenyl" as used herein encompass radicals having "cis" and "trans" directions, or "E" and "Z" directions.

The term "alkynyl" embraces linear or branched radicals having at least one carbon-carbon triple bond, preferably two to about twenty-four carbon atoms ("C)₂-C₂₄"). Other preferred alkynyl radicals are "lower alkynyl" radicals having from two to about ten carbonsAtoms such as propargyl, 1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl and 1-pentynyl. Lower alkynyl radicals mentioned here include from 2 to about 6 carbon atoms ("C₂-C₆”)。

The term "aryl" as used herein refers to a monocyclic or polycyclic carbocyclic ring system comprising at least one aromatic ring, including, but not limited to, phenyl, naphthyl, tetrahydronaphthyl, indandione. Polycyclic aryl groups are polycyclic ring systems containing at least one aromatic ring. The polycyclic aryl group can comprise fused rings, covalently linked rings, or combinations thereof.

The term "heteroaryl" as used herein refers to a monocyclic or polycyclic aromatic group having one or more ring atoms selected from S, O and N; the remaining ring atoms are carbon, wherein any N or S in the ring may be optionally oxidized. Heteroaryl groups include, but are not limited to, pyridine, pyrazine, pyrrolyl, pyrimidinyl, pyrrolyl, pyrazolyl, imidazole, thiazole, oxazolyl, isoxazolyl, thiadiazolyl, oxadiazolyl, thiophenyl, furanyl, quinoline, isoquinoline, benzimidazolyl, phenylmethyloxazolyl, quinoxalinyl. The polycyclic heteroaryl group can comprise a fused ring, a covalently attached ring, or a combination thereof.

The term "aralkyl" as used herein refers to a functional group in which an alkylene chain is attached to an aryl group, e.g., -CH₂CH₂-phenyl. The term "substituted aralkyl" refers to an aralkyl functional group in which the aryl group is substituted. Similarly, the term "heteroaralkyl" refers to a functional group in which an alkylene chain is attached to a heteroaryl group. The term "substituted heteroaralkyl" refers to a heteroaralkyl functional group in which the heteroaryl group is substituted.

The term "alkoxy" as used herein, alone or in combination with other terms, means, unless otherwise stated, an alkyl group having the stated number of carbon atoms attached to the remainder of the molecule through an oxygen atom, e.g., methoxy, ethoxy, 1-propoxy, 2-propoxy (isopropoxy), and higher homologs and isomers. HeadIs selected from alkoxy radicals of (C)₁-C₃) An alkoxy group.

The term "cycloalkyl" refers to a saturated carbocyclic radical ("C") having three to about twelve carbon atoms₃-C₁₂"). The term "cycloalkyl" embraces saturated carbocyclic radicals having from three to about twelve carbon atoms. Examples of such radicals include cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

The term "alkoxy" means an alkyl-O-radical.

The term "cycloalkenyl" refers to a partially unsaturated carbocyclic radical having three to twelve carbon atoms. Partially unsaturated carbocyclic radicals containing two double bonds (which may or may not be conjugated), such cycloalkenyl radicals may be referred to as "cycloalkylene". More preferred cycloalkenyl radicals are "lower cycloalkenyl" radicals having from four to about eight carbon atoms. Examples of such radicals include cyclobutenyl, cyclopentenyl and cyclohexenyl.

The term "heterocycle" encompasses saturated, partially unsaturated, and unsaturated cyclic radicals containing heteroatoms, which may be selected from nitrogen, sulfur, and oxygen, and may be correspondingly referred to as "heterocycloalkyl", "heterocycloalkenyl", and "heteroaryl". Examples of saturated heterocycloalkyl radicals include saturated heteromonocyclic groups containing 3 to 6 members containing 1 to 4 nitrogen atoms (e.g., pyrrolidinyl, imidazolidinyl, piperidinyl, piperazinyl, etc.); saturated heteromonocyclic groups containing 3 to 6 members containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms (e.g., morpholinyl, etc.); saturated heteromonocyclic groups containing 3 to 6 members having 1 to 2 sulfur atoms and 1 to 3 nitrogen atoms (e.g., thiazolyl, etc.). Examples of partially unsaturated heterocycloalkyl radicals include dihydrothiophene, dihydropyran, dihydrofuran, and dihydrothiazole. The heterocycloalkyl radical may include a pentavalent nitrogen, as in tetrazole and pyridine radicals. The term "heterocycle" also includes those heterocycloalkyl radicals which are fused to an aryl radical or a cycloalkyl radical. Examples of such fused bicyclic radicals include benzofuran, benzothiophene, and the like.

The term "halogen" or "halo", as used herein, refers to an atom selected from fluorine, chlorine, bromine and iodine. Preferred halogens are fluorine and chlorine.

The term "haloalkyl" refers to an alkyl group that includes one or more halo substituents.

The term "haloalkoxy" refers to an alkoxy group that includes one or more halogen substituents.

The term "substituted" refers to the independent substitution of one, two, or three or more hydrogen atoms with substituents including, but not limited to, -F, -Cl, -Br, -I, -OH, C₁-C₁₂-an alkyl group; c₂-C₁₂-alkenyl, C₂-C₁₂-alkynyl, -C₃-C₁₂-cycloalkyl, protected hydroxy, -NO₂，-N₃，-CN，-NH₂Protected amino, oxy, thio, -NH-C₁-C₁₂-alkyl, -NH-C₂-C₈-alkenyl, -NH-C₂-C₈-alkynyl, -NH-C₃-C₁₂-cycloalkyl, -NH-aryl, -NH-heteroaryl, -NH-heterocycloalkyl, -dialkylamino, -diarylamino, -diheteroarylamino, -O-C₁-C₁₂-alkyl, -O-C₂-C₈-alkenyl, -O-C₂-C₈-alkynyl, -O-C₃-C₁₂-cycloalkyl, -O-aryl, -O-heteroaryl, -O-heterocycloalkyl, -C (O) -C₁-C₁₂-alkyl, -C (O) -C₂-C₈-alkenyl, -C (O) -C₂-C₈-alkynyl, -C (O) -C₃-C₁₂-cycloalkyl, -c (o) -aryl, -c (o) -heteroaryl, -c (o) -heterocycloalkyl, -CONH₂，-CONH-C₁-C₁₂-alkyl, -CONH-C₂-C₈-alkenyl, -CONH-C₂-C₈-alkynyl, -CONH-C₃-C₁₂-cycloalkyl, -CONH-aryl, -CONH-heteroaryl, -CONH-heterocycloalkyl, -OCO₂-C₁-C₁₂-alkyl, -OCO₂-C₂-C₈-alkenyl, -OCO₂-C₂-C₈-alkynyl, -OCO₂-C₃-C₁₂-cycloalkyl, -OCO₂-aryl, -OCO₂-heteroaryl, -OCO₂-heterocycloalkyl, -CO₂-C₁-C₁₂Alkyl, -CO₂-C₂-C₈Alkenyl, -CO₂-C₂-C₈Alkynyl radical, CO₂-C₃-C₁₂-cycloalkyl, -CO₂Aryl radical, CO₂-heteroaryl, CO₂-heterocycloalkyl, -OCONH₂，-OCONH-C₁-C₁₂-alkyl, -OCONH-C₂-C₈-alkenyl, -OCONH-C₂-C₈-alkynyl, -OCONH-C₃-C₁₂-cycloalkyl, -OCONH-aryl, -OCONH-heteroaryl, -OCONH-heterocycloalkyl, -NHC (O) H, -NHC (O) -C₁-C₁₂-alkyl, -NHC (O) -C₂-C₈-alkenyl, -NHC (O) -C₂-C₈-alkynyl, -NHC (O) -C₃-C₁₂-cycloalkyl, -nhc (o) -aryl, -nhc (o) -heteroaryl, -nhc (o) -heterocycloalkyl, -NHCO₂-C₁-C₁₂-alkyl, -NHCO₂-C₂-C₈-alkenyl, -NHCO₂-C₂-C₈-alkynyl, -NHCO₂-C₃-C₁₂-cycloalkyl, -NHCO₂-aryl, -NHCO₂-heteroaryl, -NHCO₂-heterocycloalkyl, -NHC (O) NH₂，-NHC(O)NH-C₁-C₁₂-alkyl, -NHC (O) NH-C₂-C₈-alkenyl, -NHC (O) NH-C₂-C₈-alkynyl, -NHC (O) NH-C₃-C₁₂-cycloalkyl, -NHC (O) NH-aryl, -NHC (O) NH-heteroaryl, -NHC (O) NH-heterocycloalkyl, NHC (S) NH₂，-NHC(S)NH-C₁-C₁₂-alkyl, -NHC (S) NH-C₂-C₈-alkenyl, -NHC (S) NH-C₂-C₈-alkynyl radical，-NHC(S)NH-C₃-C₁₂-cycloalkyl, -NHC (S) NH-aryl, -NHC (S) NH-heteroaryl, -NHC (S) NH-heterocycloalkyl, -NHC (NH) NH₂，-NHC(NH)NH-C₁-C₁₂-alkyl, -NHC (NH) NH-C₂-C₈-alkenyl, -NHC (NH) NH-C₂-C₈-alkynyl, -NHC (NH) NH-C₃-C₁₂-cycloalkyl, -NHC (NH) NH-aryl, -NHC (NH) NH-heteroaryl, -NHC (NH) NH-heterocycloalkyl, -NHC (NH) -C₁-C₁₂-alkyl, -NHC (NH) -C₂-C₈-alkenyl, -NHC (NH) -C₂-C₈-alkynyl, -NHC (NH) -C₃-C₁₂-cycloalkyl, -NHC (NH) -aryl, -NHC (NH) -heteroaryl, -NHC (NH) -heterocycloalkyl, -C (NH) NH-C₁-C₁₂-alkyl, -C (NH) NH-C₂-C₈-alkenyl, -C (NH) NH-C₂-C₈-alkynyl, -C (NH) NH-C₃-C₁₂-cycloalkyl, -C (NH) NH-aryl, -C (NH) NH-heteroaryl, -C (NH) NH-heterocycloalkyl, -S (O) -C₁-C₁₂-alkyl, -S (O) -C₂-C₈-alkenyl, -S (O) -C₂-C₈-alkynyl, -S (O) -C₃-C₁₂-cycloalkyl, -S (O) -aryl, -S (O) -heteroaryl, -S (O) -heterocycloalkyl, -SO₂NH₂，-SO₂NH-C₁-C₁₂-alkyl, -SO₂NH-C₂-C₈-alkenyl, -SO₂NH-C₂-C₈-alkynyl, -SO₂NH-C₃-C₁₂-cycloalkyl, -SO₂NH-aryl, -SO₂NH-heteroaryl, -SO₂NH-heterocycloalkyl, -NHSO₂-C₁-C₁₂-alkyl, -NHSO₂-C₂-C₈-alkenyl, -NHSO₂-C₂-C₈-alkynyl, -NHSO₂-C₃-C₁₂-cycloalkyl, -NHSO₂-aryl, -NHSO₂-heteroaryl, -NHSO₂-heterocycloalkyl, -CH₂NH₂，-CH₂SO₂CH₃-aryl, -arylalkyl, -heteroaryl, -heteroarylalkyl, -heterocycloalkyl, -C₃-C₁₂Cycloalkyl, polyalkoxyalkyl, polyalkoxy, -methoxymethylOxy, -methoxyethoxy, -SH, -S-C₁-C₁₂-alkyl, -S-C₂-C₈-alkenyl, -S-C₂-C₈-alkynyl, -S-C₃-C₁₂-cycloalkyl, -S-aryl, -S-heteroaryl, -S-heterocycloalkyl, or methylthio-methyl. In certain embodiments, the substituents are independently selected from halogen, preferably Cl and F; c_1-C₄-alkyl, preferably methyl and ethyl; halo-C_1-C₄Alkyl groups, such as fluoromethyl, difluoromethyl, and trifluoromethyl; c₂-C₄-an alkenyl group; halo-C₂-C₄-an alkenyl group; c₃-C₆Cycloalkyl, for example, cyclopropyl; c_1-C₄Alkoxy, for example, methoxy and ethoxy; halo-C₁-C₄Alkoxy radicals, such as fluoromethoxy, difluoromethoxy and trifluoromethoxy; -CN; -OH; NH (NH)₂；C₁-C₄-an alkylamino group; two (C)_1-C₄-alkyl) amino; and NO₂. It is to be understood that the aryl, heteroaryl, alkyl, cycloalkyl, heterocyclyl, and the like may also be further substituted. In some cases, where possible, the individual substituents in the substituent groups may additionally also be optionally substituted by one or more radicals, each radical independently of the others, selected from C_1-C₄-an alkyl group; -CF₃，-OCH₃，-OCF₃，-F，-Cl，-Br，-I，-OH，-NO₂-CN, and-NH₂. Preferably, a substituted alkyl group, such as a substituted methyl group, is substituted with one or more halogen molecules, more preferably, one or more fluorine or chlorine molecules.

The term "optionally substituted" as used herein means that the group in question may be substituted or unsubstituted. In one embodiment, the group referred to is optionally substituted with a zero substituent, i.e. the group referred to is unsubstituted. In another embodiment, the groups referred to are optionally substituted with one or more other groups individually and independently selected from the groups described herein.

The compounds of the present invention may occur in various forms, including salt forms, particularly pharmaceutically acceptable salts, co-crystals, solvates, hydrates, polymorphs, enantiomers, diastereomers, racemates and the like, having the formula as described herein. In certain embodiments, the compounds of the present invention occur as racemic mixtures, e.g., stereoisomers having the stereochemistry of formulae (Ia), (IIa), (IIIa), and (IVa), and formulae (Ib), (IIb), (IIIb), and (IVb). In other embodiments, the compound is present as a mixture of two enantiomers, with an enantiomeric excess of one enantiomer. In other embodiments, the compound is present as a substantially pure single enantiomer, e.g., having an enantiomeric excess of at least 90%, 95%, 98%, or 99%.

As used herein, the term "pharmaceutically acceptable salt" refers to salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without excessive toxicity, irritation, allergic response, and the like, commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. Such as S. Pharmaceutically acceptable salts are described by MBerge et al in j.pharmaceutical Sciences, 66:1-19 (1977). Salts may be prepared in situ during the final isolation and purification of the compounds of the invention or separately by reacting the free base function with a suitable organic acid. Examples of pharmaceutically acceptable salts include, but are not limited to, non-toxic acid addition salts, which are salts formed from an amino group and an inorganic acid (e.g., hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, and perchloric acid) or an organic acid (e.g., acetic acid, maleic acid, tartaric acid, citric acid, succinic acid, or malonic acid) or by using other methods used in the art (e.g., ion exchange). Other pharmaceutically acceptable salts include, but are not limited to, adipates, alginates, ascorbates, aspartates, benzenesulfonates, benzylsulfates, sodium bisulfates, borates, butyrates, camphorates, camphorsulfonates, citrates, cyclopentanepropionates, digluconates, dodecylsulfates, ethanesulfonates, formates, fumarates, glucoheptonates, glycerophosphates, gluconates, hemisulfates, heptanoates, hexanoates, hydroiodides, 2-hydroxy-ethanesulfonates, lactobionates, lactates, laurates, dodecylsulfates, malates, maleates, malonates, methanesulfonates, 2-naphthalenesulfonates, nicotinates, nitrates, oleates, oxalates, palmitates, pamonates, pectinates, persulfates, 3-phenylpropionates, salts of citric acid, salts of acetic acid, salts of citric acid, salts of acetic acid, salts of benzoic acid, salts of citric acid, salts of benzoic acid, salts of acetic acid, salts of benzoic acid, salts of benzoic acid, salts of acids, salts of esters of acids, salts of acids of salts of acids, salts of acids of salts of acids, salts of acids of salts of acids, salts of acids of salts of acids of salts of acids of salts of acids of salts of acids, Phosphates, picrates, pivalates, propionates, stearates, succinates, sulfates, tartrates, thiocyanates, p-toluenesulfonates, undecanoates, pentanoates, and the like. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Other pharmaceutically acceptable salts include acidic drug salts with non-toxic ammonium, quaternary ammonium, amine cations and salts.

The term "hydroxy protecting group" as used herein means that the labile hydroxy group is protected from undesirable reactions during synthesis using protecting groups known in the art. Following the above synthetic steps, the hydroxyl protecting groups as described herein may be selectively removed. Hydroxy protecting Groups are known in the art, e.g., T.H.Greene and P.G.M.Wuts in Protective Groups in Organic Synthesis, 3rd edition, John Wiley & Sons, New York (1999). As described in (1). Examples of the hydroxyl-protecting group include benzyloxycarbonyl, 4-methoxybenzyloxycarbonyl, t-butoxycarbonyl, isopropyloxycarbonyl, diphenylmethoxy 2,2, 2-trichlorocarbonylethoxycarbonyl, allyloxycarbonyl, acetyl, formyl, chloroacetyl, trifluoroacetyl, methoxyacetyl, phenoxyacetyl, benzyl, methyl, t-butyl, 2,2, 2-trichloroethyl, 2-trimethylsilylethyl, allyl, benzyltriphenyl-methyl (trimethylbenzene), methoxymethyl, methylmercaptan methyl, benzyloxymethyl, 2- (tri) methylsilyl) -ethoxymethyl, methanesulfonyl, trimethylsilane, triisopropyl and the like.

The term "protected hydroxyl group" as used herein refers to a hydroxyl group protected with a hydroxyl protecting group, as described above, including, for example, benzyl, acetyl, trimethylsilyl, triethylsilyl, methoxymethyl groups.

The term "amino protecting group" as used herein means that the labile amino group is protected from undesirable reactions during synthesis using protecting groups known in the art. Following the above synthetic steps, the amino protecting groups as described herein may be selectively removed. Amino protecting Groups are known in the art, e.g., T.H.Greene and P.G.M.Wuts in Protective Groups in Organic Synthesis, 3rd edition, John Wiley & Sons, New York (1999). As described in (1). Examples of amino protecting groups include, but are not limited to, methoxycarbonyl, t-butoxycarbonyl, 9-fluoren-methoxycarbonyl, benzyloxycarbonyl, and the like.

The term "protected amino" as used herein refers to an amino group that is protected with an amino protecting group as described above.

The present invention includes all pharmaceutically acceptable isotopically-labeled or enriched compounds of the present invention. These compounds contain an isotopic abundance or indicator element at one or more positions that is different from the natural isotopic distribution of the element. For example, where a hydrogen atom is depicted, deuterium may be included in a higher abundance than the natural abundance of deuterium.

Examples of isotopes suitable for inclusion within the scope of compounds of the invention include isotopes of hydrogen, for example,²h and³h; isotopes of carbon, for example,¹¹C，¹³c and¹⁴c; isotopes of nitrogen, for example,¹³n and¹⁵n; isotopes of oxygen¹⁵O，¹⁷O and¹⁸o; isotopes of chlorine, for example,³⁶cl; isotopes of fluorine, for example,¹⁸f; isotopes of iodine, for example,¹²³i and¹²⁵i; isotopes of phosphorus, for example,³²p; and isotopes of sulfur, such as, for example,³⁵S。

substituents attached through a variable point of attachment may be attached at any available position on the ring structure.

As used herein, the term "effective amount of a host compound," with respect to a method of treatment, refers to an amount of the host compound that, when delivered as part of a desired dosing regimen, is capable of controlling a disease or disorder to a clinically allowable standard.

"treatment" or "treating" refers to a means of achieving a beneficial or desired clinical result in a patient. Beneficial or desired clinical results for the present invention include, but are not limited to, one or more of the following: alleviating symptoms, diminishing the extent of disease, stabilizing (i.e., not worsening) the disease state, preventing the spread of disease (i.e., metastasis), preventing the disease from occurring or reoccurring, delaying or slowing the disease process, improving the disease state and relieving symptoms (whether partial or total).

The compounds and methods of the present invention will be better understood in conjunction with the following examples, which are included merely for purposes of illustration and are not intended to limit the scope of the invention. Various changes and modifications to the disclosed embodiments will be apparent to those skilled in the art, and such changes and modifications, including but not limited to those relating to chemical structures, substituents, derivatives, may be made in the formulations and/or methods of the present invention without departing from the spirit of the invention and the scope of the appended claims.

Examples

List of abbreviations

All temperatures are in degrees Celsius

BF₃.Et₂O-boron trifluoride acetate

CDCl₃Deuterated chloroform

CF-cystic fibrosis

CFTR-cystic fibrosis transmembrane conductance regulator

CH₃CN-acetonitrile

CH₃NO₂-nitromethane

CH₂Cl₂-methylene chloride

DIPEA-N, N-diisopropylethylamine

DMF-dimethylformamide

DMSO-d₆Deuterated dimethyl sulfoxide

ENaC-epithelial sodium channel

Et₂O-diethyl ethyl ester

EtOAc-ethyl acetate

H₂O-water

HATU- (1- [ bis (dimethylamine) methylene ] -1H-1,2, 3-triazolo [4,5-b ] pyridine 3-oxidohexafluorophosphate)

HBS-Hepes-buffer saline

HCl-hydrochloric acid

HCOOH-carboxylic acid

HPLC-high pressure liquid chromatography

hr-hour

HTS-high throughput screening

ms-microsecond

Na₂SO₄-sodium sulphate

NaBH₄Sodium borate

NaOH-sodium hydroxide

NaHCO₃Sodium bicarbonate

Normalized area under NAUC-curve

NH₄OAc-ammonium acetate

NMR-nuclear magnetic resonance

Ether-petroleum ether

PBS-phosphate buffer saline

Pd(PPh₃)₄-tetravalent palladium

s-second

rt–RT

TFA-trifluoroacetic acid

THF-tetrahydrofuran

YFP-yellow fluorescent protein

Example 1: synthesis of Compounds

The compounds of the invention were synthesized by method a or method B below. Process A is carried out by ring opening of the corresponding succinimide compound. These succinimide compounds are prepared as described in WO 2017/117239, which is incorporated herein by reference in its entirety.

Method A

The present method is an exemplary method for preparing 5, 7-dichloro-1 ' - ((3, 5-dichloro-4- (difluoromethoxy) phenyl) carbamoyl) -6', 6' -dimethyl-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -2' -carboxylic acid (compound 8), as shown in the following schematic diagram.

To a stirred solution of Compound A (100mg, 0.165mmol) in THF (33mL) at room temperature was added 1% saturated NaHCO₃(66mL) and the reaction mixture was stirred at room temperature for 24 h. The reaction mixture was cooled to 0 ℃ and acidified with 1N HCl (pH-4) then extracted with ethyl acetate EtOAc (2X30 mL). The combined organic layers were dried over anhydrous sodium sulfate, filtered and evaporated under vacuum to give the crude product. By preparative high performance liquid chromatography HPLC (0.05% HCOOH/CH)₃CN/H₂O) purification, yielding 40mg (39% yield) of compound 8 as a solid. LCMS 621.9[ M + H ]]⁺(ii) a (97.2% purity).¹H NMR(500MHz，DMSO-d₆)δ＝12.28(bs，1H)，10.89(s，1H)，10.51(s，1H)，8.16(d，J＝2.0Hz，1H)，7.82(s，2H)，7.45(d，J＝2.0Hz，1H)，7.11(t，J＝72.5Hz，1H)，4.20-4.13(m，2H)，3.47(t，J＝7.0Hz，1H)，2.49(d，J＝8.0Hz，1H)，1.93(d，J＝7.5Hz，1H)，1.61-1.58(m，1H)，1.29-1.23(m，1H)，1.00(s，3H)，0.91(s，3H)；¹⁹F NMR(470.59MHz，DMSO-d₆): -80.26, -80.10. Chiral SFC purification (97.9% ee).

Preparative HPLC conditions

A chromatographic column: symmerry-C8 (300 × 19), 7 u; mobile phase 0.1% formic acid in H₂O:CH₃Gradient solution in CN (T% B) 0/30, 8/80, 8.1/98, 10/98, 10.1/30, 13/30 flow rate 20 ml/min; diluent CH₃CN+H₂O+THF。

Method B

The present method is an exemplary method for preparing (1'R, 2' S, 3R, 7a 'R) -1' - ((3- (tert-butyl) phenethyl) carbamoyl) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid (Compound 44)

(Z) -4- (allyloxy) -4-oxybut-2-enoic acid (A)

A solution of maleic anhydride (15.0g, 152mmol) in allyl alcohol (200mL) was stirred at room temperature for 16 h. The reaction mixture was concentrated in vacuo to a residue, which was dissolved in Et₂In O (1.0L), with H₂O (3X1L) and brine (500mL) and then dried over anhydrous Na₂SO₄Dried and concentrated in vacuo to give 12.0g (50% yield) of a as a colorless liquid. LCMS M/z157.0[ M + H ]]⁺(ii) a (99.5% purity).

2' - ((allyloxy) carbonyl) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazin-cyclo ] -1' -carboxylic acid (E).

To a stirred solution of B (5g, 32.0mmol) in THF (70mL) at room temperature was added 5, 7-dichloroisatin (C) (6.9g, 32.0mmol) and 4, 4-difluoro-L-proline-TFA salt (D) (7.9g, 32.0 mmol). The reaction mixture was stirred at 80 ° for 3 hours, then cooled to room temperature and evaporated in vacuo to give the crude product as a mixture of diastereomers. Purification by column chromatography, with Pet ether: EtOAc (80:20) eluted to give the crude product as a brown solid. The brown material was triturated with CH2Cl2 and filtered to give, after drying in vacuo, 1.3g (9% yield) of E as a white solid (single isomer). LCMS M/z461.0[ M + H ]]⁺(ii) a (99.2% purity).¹HNMR (500MHz, acetone-d)₆)11.8–10.9(brs，1H)，9.96(s，1H)，7.85(d, J ═ 2Hz, 1H), 7.39(d, J ═ 2Hz, 1H), 5.56-5.53 (m, 1H), 5.15-5.07 (m, 2H), 4.33-4.32 (m, 2H), 4.22-4.20 (m, 1H), 4.10-4.09 (m, 1H), 3.82-3.79(m, 1H), 3.30-3.25 (m, 1H), 2.53-2.45 (m, 1H), 2.34-2.20 (m, 1H), the desired diastereomer (i.e. having the relative mr stereochemistry described in formulae (Ia), (IIa) and (IIIa)) was confirmed by 2 dnd.

(E) -1- (tert-butyl) -3- (2-nitrovinyl) benzene (F).

To NH4OAc (4.2g, 55.1mmol) in CH at 90 deg.C₃NO₂To the solution in (150mL) was added 3- (tert-butyl) benzaldehyde (F) (3g, 18.3mmol), and the resulting mixture was stirred at 120 ℃ for 16 hours. The reaction mixture was cooled to room temperature and concentrated in vacuo to afford the crude product as a brown liquid. Purification by column chromatography, with Pet ether: EtOAc (80:20) elution afforded 2.9g (65% yield) of product g as a light brown solid.¹H NMR(500MHz，CDCl₃)8.03(d，J＝13.5Hz，1H)，7.60(d，J＝14Hz，1H)，7.55–7.53(m，2H)，7.39–7.38(m，2H)。

2- (3- (tert-butyl) phenyl) ethan-1-amine (I).

To NaBH₄(847mg, 22.4mmol) to a stirred solution in anhydrous THF (20mL) BF was added₃.Et₂O (3.3mL, 26.8mmol), temperature 0 ℃. After stirring at 0 ℃ for 15 min, the reaction mixture was warmed to room temperature and then a solution of (H) (920mg, 4.48mmol) in anhydrous THF (10mL) was added. The resulting mixture was stirred at 85 ℃ for 5 hours and then cooled to 0 ℃. H is added over 20 minutes₂O (40mL) and 1N HCl (40mL) were added to the reaction mixture, and the resulting mixture was stirred at 85 ℃ for 2 hours. The reaction mixture was cooled to 0 ℃ and 5N Na was usedThe OH solution is alkalized (pH value is 12). The resulting mixture was extracted with EtOAc (2 × 100mL) and the combined organic layers were washed with brine, over anhydrous Na₂SO₄Dried above and concentrated in vacuo to give 700mg (88% yield) of product (I) as a liquid, which was used in the next step without purification. LCMS M/z 178.1[ M + H ]]⁺；(99.5％purity)。

Allyl 1' - ((3- (tert-butyl) phenethyl) carbamoyl) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -2' -carboxylate (J).

To a stirred solution of E (200mg, 0.43mmol) in DMF (10mL) at room temperature were added DIPEA (0.15mL, 0.86mmol), HATU (196mg, 0.51mmol) and I (115mg, 0.65 mmol). The resulting reaction mixture was stirred at room temperature for 30min, then poured into ice-cold H₂In O and stirred for 10 min. Collecting the precipitate, cooling with cold H₂O washes and dries in vacuo to give 250mg (94% yield) of product J as a white solid, which is used without purification in the next step LCMS: M/z 620.1[ M + H ]]⁺(ii) a (10.9% + 83.3% purity).

1' - ((3- (tert-butyl) phenethyl) carbamoyl) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -2' -carboxylic acid (44).

Aniline (30mg, 0.32mmol) and Pd (PPh)₃)₄(37mg, 0.032mmol) was added to a stirred solution of J (200mg, 0.32mmol) in dry THF (5mL) at room temperature. The resulting mixture was stirred at room temperature for 1 hour, then diluted with EtOAc (100mL) and washed with brine, over anhydrous Na₂SO₄Dried above, and concentrated in vacuo to give the crude product as a liquid. The crude product is passed throughPreparative high performance liquid chromatography purification using the following conditions gave 95mg (50% yield) of the product 44: INERTSIL-ODS column (250 mm. times.20 20 mm. times.5 5 uM); mobile phase A-0.1% formic acid water solution, B-0.05% formic acid CH3CN solution; time (min)/% B0/60, 8/85, 9/85, 12/95, 12.1/60, 15/60; flow rate-20 mL/min.

LCMS:m/z 580.1[M+H]⁺；(98.9％de)。¹H NMR(500MHz，DMSO-d₆)：12.40(s，1H)，10.95(s，1H)，8.25(br s，2H)，7.43(d，J＝1.5Hz，1H)，7.26(s，1H)，7.24-7.20(m，2H)，7.07-7.05(m，1H)，3.96-3.90(m，2H)，3.46-3.44(m，2H)，3.17-3.13(m，2H)，2.77-2.71(m，2H)，2.58-2.51(m，1H)，2.10-2.07(m，1H)，1.99-1.85(m，1H)，1.28(s，9H)。

Compounds 1-65 were prepared and characterized by LCMS as described herein. For each compound, the synthesis method used is shown in the last column.

EXAMPLE 66 Synthesis of 5, 7-dichloro-4 '- ((3, 5-dichlorophenyl) carbamoyl) -5' - (difluoro (pyridin-4-yl) methyl) -2-oxospiro [ indole-3, 2 '-pyrrolidine ] -3' -carboxylic acid, 66.9a, 66.9b1, 66.9b2, 66.9b3

66.2 Synthesis

To a stirred suspension of activated copper powder (3.8g, 60.9mmol), CuI (2.3g, 12.2mmol) in DMSO (30mL) at room temperature was added 4-iodinePyridine, 66.1(5.0g, 24.4mmol) and ethyl 2-bromo-2, 2-difluoroacetate (12.4g, 60.9 mmol). The resulting reaction mixture was stirred at room temperature for 16 h. The reaction mixture was cooled to room temperature, poured into ice water (50mL) and diethyl ether (2 × 50mL) extracted with deionized water. The combined organic layers were washed with anhydrous Na₂SO₄Dried, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography (80g silica gel column, 20% ethyl acetate in petroleum ether) to give 66.2(4g, 81%) as a liquid.¹H NMR(500MHz，CDCl₃):8.81(br s，2H)，7.53(d，J＝5.0Hz，2H)，4.32(q，J＝7.0Hz，2H)，1.32(t，J＝7.0Hz，3H)。

66.3 Synthesis

To a stirred solution of 66.2(2.2g, 10.9mmol) in MeOH (50mL) at-60 deg.C was added NaBH in portions₄(289mg, 7.6 mmol). The resulting reaction mixture was stirred at-60 ℃. By NH at-60 deg.C₄The reaction mixture was quenched with Cl and then extracted with EtOAc (2 × 30 mL). The combined organic phases were washed with water (30ml), brine (30ml) and anhydrous Na₂SO₄Drying, filtration and concentration under reduced pressure gave 66.3(1.8g) as a solid which was used directly in the next step without purification.¹H NMR(400MHz，DMSO-d₆):8.90(br s，2H)，7.56(br s，2H)，7.33(d，J＝6.4Hz，1H)，4.89(q，J＝6.0Hz，1H)，3.29(s，3H)。

66.4 Synthesis

To a stirred solution of 66.3(1.8g, 9.49mmol) in toluene (30mL) was added (S) -2-amino 2-phenyleth-1-ol (1.3g, 9.49mmol), PTSA (180mg, 0.94mmol) at room temperature and the resulting mixture was refluxed for 2 hours using a Dean-Stark apparatus. The reaction mixture was concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 10% EtOAc in petroleum ether) to give diastereomer mixture 66.4(2.1g, 80%) as a colorless liquid. LCMS (37.2+ 52.1%, M/z [ M + H ]]⁺＝277.0。

Synthesis of 66.5

TMSCN (1.4g, 14.5mmol) and BF3.Et2O (2g, 14.5mmol) were added to a stirred solution of 66.4(2.0g, 7.23mmol) in CH2Cl2(50mL)In solution, at-78 ℃ for 10 minutes. The resulting reaction mixture was stirred at-78 ℃ for 8 hours. The reaction mixture was poured into saturated NaHCO3 solution (80mL) and extracted with CH2Cl2(2 × 30 mL). The combined organic layers were washed with water (50mL), brine (50mL), dried over anhydrous Na2SO4, filtered and concentrated under vacuum to afford a residue. The residue was purified by flash column chromatography (40g silica gel column, 30% EtOAc in petroleum ether) to give diastereomer mixture 66.5(1.8g, 81%) as a light brown solid. LCMS (30.5+ 66.2%), M/z [ M + H ]]⁺＝304.3。

66.6 Synthesis

66.5(2.4g, 7.91mmol) in MeOH (50mL) and CH at 0 deg.C₂Cl₂(80mL) to the stirred solution was added Pb (OAc)₄(5.2g, 11.9 mmol). The resulting reaction mixture was stirred at 0 ℃ for 2 hours. The reaction mixture was poured into 0.2M phosphate buffer (50mL) at room temperature and then filtered through a celite bed. CH for aqueous layer₂Cl₂(2 × 20mL). By H₂The combined organic layers were washed with O (2X50mL), brine (50mL) and dried over anhydrous Na₂SO₄Dried, filtered and concentrated under vacuum to afford a mixture of diastereomer 66.6(1.8g) which exists as a brown viscous state, which was used in the next step without purification.

66.7 Synthesis

A stirred solution of 66.6(1.7g, 6.26mmol) in concentrated hydrochloric acid (60mL) was heated at 100 ℃ for 6 hours. The resulting reaction mixture was cooled to room temperature and concentrated under vacuum at 50 ℃. By CH₂Cl₂The resulting residue was extracted to give diastereomer 66.7(550mg) as a light grey solid. LCMS 87.5%, M/z [ M + H ]]⁺＝203.2。

Synthesis of 66.8a &66.8b

To a stirred solution of 66.7(500mg, 2.47mmol) in EtOH (40mL) was added 1- (3.5- [ chloro ] phenyl) -1H-pyrrole-2, 5-dione (598mg, 2.47mmol), 5, 7- [ chloro ] indole-2, 3-dione (534mg, 2.47mmol) at room temperature. The reaction mixture was stirred at 90 ℃ for 2 h. Volatile components were removed under vacuum. The resulting residue was purified by flash chromatography (silica gel 100-200mesh, 50% EtOAc/petroleum ether) to afford 66.8a (120mg, 8%) and 66.8b (240mg, 16%) as solids.

66.8a:¹H NMR(500MHz，DMSO-d₆):11.18(s，1H)，8.76(d，J＝5.0Hz，2H)，7.78(d，J＝1.5Hz，1H)，7.58(d，J＝6.0Hz，2H)，7.53(d，J＝2.0Hz，1H)，7.33(d，J＝2.0Hz，1H)，7.25(d，J＝2.0Hz，2H)，4.60-4.49(m，2H)，3.93(s，2H)；LCMS:97.5％，m/z[M+H]⁺＝597.1。

66.8b:¹H NMR(500MHz，DMSO-d₆):11.07(br s，1H)，8.73(d，J＝5.5Hz，2H)，7.78(t，J＝2.0Hz，1H)，7.61(d，J＝6.0Hz，2H)，7.49(d，J＝1.5Hz，1H)，7.33(d，J＝1.5Hz，2H)，7.05(d，J＝1.5Hz，1H)，5.02-4.94(m，1H)，4.35(d，J＝5.0Hz，1H)，3.90(t，J＝8.5Hz，1H)，3.67(d，J＝8.0Hz，1H)；LCMS:99.4％，m/z[M+H]⁺＝597.1。

66.9a Synthesis

To a stirred solution of 66.8a (80mg, 0.13mmol) in THF (30mL) at room temperature was added 1% NaHCO₃Solution (60 mL). The resulting reaction mixture was stirred at room temperature for 96 h. After completion of the reaction, the pH of the solution was adjusted to 6-7 with 1N HCl solution and extracted with EtOAc (2 × 20mL). The combined organic layers were washed with brine (30mL) over anhydrous Na₂SO₄Dried, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative HPLC [ X-bridge C18(150X30) mm, 5 μ; a is 0.1 percent HCOOH aqueous solution, B is acetonitrile; gradient: (time/% B):0/40, 8/80, 9/80, 9.1/98, 14/98, 14.1/40, 17/40 and 25mL/min]66.9a (32mg, 39%) was obtained as a solid.¹H NMR(500MHz，DMSO-d₆):12.91(br s，1H)，11.21(br s，1H)，11.03(br s，1H)，8.71(d，J＝5.5Hz，2H)，7.54-7.52(m，4H)，7.44(br s，1H)，7.28(s，1H)，7.25-7.16(m，1H)，4.80-4.69(m，1H)，4.30-4.15(m，2H)，3.54-3.51(m，1H)；LCMS:97.0％，m/z[M+H]⁺＝615.1。

Synthesis of 66.9b.1, 66.9b.2&66.9b.3

To a stirred solution of 66.8b (180mg, 0.3mmol) in THF (60mL) at room temperature was added 1% NaHCO₃Solution (120 mL). The resulting reaction mixture was stirred at room temperature for 96 h. After completion of the reaction, the pH of the solution was adjusted to 6-7 with 1N HCl solution and extracted with EtOAc (2 × 20mL). The combined organic layers were washed with brine (30mL) over anhydrous Na₂SO₄Dried, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative HPLC [ X-bridge C18(150X30) mm, 5 μ; a is 0.1 percent HCOOH aqueous solution, B is acetonitrile; gradient: (time/% B):0/40, 8/80, 9/80, 9.1/98, 14/98, 14.1/40, 17/40 and 25mL/min]66.9b.1(7mg, 4%) as solid, 66.9b.2(7mg, 4%) as solid and 66.9b.3(64mg, 35%) as solid were obtained.

66.9b.1:LCMS:93.8％，m/z[M+H]⁺＝615.1；

66.9b.2:LCMS:87.0％，m/z[M+H]⁺＝615.1；

66.9b.3:¹H NMR(500MHz，DMSO-d₆):12.46(br s，1H)，10.95(s，1H)，10.57(s，1H)，8.66(d，J＝5.0Hz，2H)，8.19(d，J＝1.5Hz，1H)，7.67(d，J＝1.5Hz，2H)，7.51(d，J＝6.0Hz，2H)，7.46(s，1H)，7.32(s，1H)，4.34-4.28(m，1H)，4.00-3.90(m，1H)，3.76-3.73(m，1H)，3.60-3.59(m，1H)；LCMS:99.4％，m/z[M+H]⁺＝615.1。

EXAMPLE 67 Synthesis of 5, 7-dichloro-5 '- ((4-chlorophenyl) difluoromethyl) -4' - ((3, 5-dichlorophenyl) carbamoyl) -2-oxaspiro [ indole-3, 2 '-pyrrolidine ] -3' -carboxylic acid, 67.9a &67.9b

67.2 Synthesis

To a stirred solution of ethyl 2- (4-chlorophenyl) -2-oxoacetic acid, 67.1(5.0g, 23.6mmol) in DCM (50mL) was added DAST (11mL, 82.5mmol) at room temperature. The resulting reaction mixture was stirred at room temperature for 16 h. The reaction mixture was cooled to 0 ℃ and quenched with saturated NaHCO3 solution. The organic layer was separated and washed with water (2 × 50mL) and brine (15 mL). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to provide 67.2(5.0g) as a light brown liquid which was used directly in the next step without purification.

¹H NMR(400MHz，CDCl₃):7.55(d，J＝8.8Hz，2H)，7.44(d，J＝8.8Hz，2H)，4.30(q，J＝7.2Hz，2H)，1.31(t，J＝7.2Hz，3H)。

67.3 Synthesis

To a stirred solution of 67.2(5g, 21.3mmol) in MeOH (25mL) at-60 deg.C was added NaBH in portions₄(800mg, 21.3 mmol). The resulting reaction mixture was stirred at-60 ℃ for 1 hour. Quench the reaction with 1N HCl solution (50mL) at 0 deg.C and Et₂O (2 × 30 mL). The combined organic phases were washed with water (30mL) and brine (30mL), dried over anhydrous Na₂SO was dried, filtered and concentrated under reduced pressure to yield 67.3(5.0g) as a pale yellow solid which was used in the next step without further purification and analysis.

67.4 Synthesis

To a stirred solution of 67.3(5.0g, 22.4mmol) in toluene (50mL) was added (S) -2-amino-2-phenyleth-1-ol (2.99g, 22.4mmol), PTSA (112mg, 0.44mmol) at room temperature. The reaction mixture was refluxed for 1 hour using a Dean-Stark apparatus. The reaction mixture was then concentrated under reduced pressure. Flash column chromatography (40g silica gel column, 20% EtOAc in petroleum ether)]The residue was purified to give a mixture of diastereomer 67.4(6.5g, 94%) as a colourless liquid. LCMS (47.7+ 40.0)%, M/z [ M + H]⁺＝310。

67.5 Synthesis

67.4(6.5g, 21.0mmol) in CH at-78 deg.C over 10min₂Cl₂(100mL) to the stirred solution were added TMSCN (5.2mL, 42.0mmol) and BF₃.Et₂O (5.1mL, 42.0 mmol). The resulting reaction mixture was stirred at room temperature for 16 hours. The reaction mixture was cooled to 0 ℃ and saturated NaHCO was used₃Solution (80mL) quenched and CH₂Cl₂(2 × 30 mL). The combined organic phases were washed with water (50mL) and brine (50mL), dried over anhydrous Na₂SO was dried, filtered and concentrated under vacuum. By flash column chromatography (80g silica gel bucket, 20 g)% EtOAc in petroleum ether) the residue was purified to give diastereomer 67.5(5.8g, 82%) as a solid.¹H NMR(400MHz，CDCl₃):7.52-7.22(m，7H)，7.02(d，J＝7.6Hz，2H)，4.13-3.96(m，1H)，3.81-3.59(m，2H)，3.53-3.46(m，1H)，2.61-2.59(m，1H)，1.73-1.65(m，1H)；LCMS:(31.1+68.3)％，m/z[M+H]⁺＝337.1。

67.6 Synthesis

67.5(5.8g, 17.2mmol) in MeOH (100mL) and CH at 0 deg.C₂Cl₂(200mL) to the stirred solution was added Pb (OAc)₄(11.5g, 25.8 mmol). The resulting reaction mixture was stirred at 0 ℃ for 30 minutes. The reaction mixture was poured into 0.2M phosphate buffer (50mL) at room temperature and then filtered through a celite bed. By CH₂Cl₂(2x40mL) extract the aqueous layer. By H₂The combined organic phases were washed with O (2X50mL) and brine (50mL) and anhydrous Na₂SO₄Drying, filtration and concentration in vacuo yielded 67.6(6.0g) as a liquid which was used in the next step without purification.¹H NMR(500MHz，CDCl₃):8.45(s，1H)，7.74(d，J＝1.5Hz，2H)，7.54-7.42(m，7H)，5.22-5.19(m，1H)。

67.7 Synthesis

A solution of 67.6(1.5g, 6.36mmol) in concentrated hydrochloric acid (20mL) was heated at 110 ℃ for 16 h. The reaction mixture was cooled to room temperature and then concentrated in vacuo at 50 ℃. The residue obtained is substituted by CH₂Cl₂Extraction yielded 67.7(550mg) as a light gray solid. LCMS 89.0%, M/z [ M + H ]]⁺＝236.1。

Synthesis of 67.8a &67.8b

To a solution of 66.7(1.0g, 4.25mmol) in EtOH (20mL) was added 1- (3, 5-dichlorophenyl) -1H-pyrrole-2, 5-dione (1.09g, 4.25mmol), 5, 7-dichloroindole-2, 3-dione (0.91g, 4.25mmol) at room temperature. The reaction mixture was heated at 80 ℃ for 2 hours. The reaction mixture was then concentrated under vacuum to yield a residue. Flash chromatography (80g silica gel column, elution gradient 50% EtOAc/petroleum ether) afforded diastereomer 67.8a (170mg, 6%) as a slightly white solid and 678b (600mg, 22%) as a bulk solid.

67.8a:¹H NMR(500MHz，DMSO-d₆):11.18(br s，1H)，7.78(t，J＝2.0Hz，1H)，7.64-7.55(m，4H)，7.53(d，J＝2.0Hz，1H)，7.30(d，J＝1.5Hz，1H)，7.25(d，J＝1.5Hz，2H)，4.52-4.49(m，2H)，3.91(s，2H)；LCMS:95.7％，m/z[M-H]^-＝628.1。

67.8b:¹H NMR(500MHz，DMSO-d₆):11.07(br s，1H)，7.79(t，J＝2.0Hz，1H)，7.63(d，J＝9.0Hz，2H)，7.56(d，J＝9.0Hz，2H)，7.50(d，J＝2.0Hz，1H)，7.30(d，J＝2.0Hz，2H)，7.03(d，J＝2.0Hz，1H)，5.00-4.92(m，1H)，4.32(d，J＝4.5Hz，1H)，3.86(t，J＝8.5Hz，1H)，3.66(d，J＝6.5Hz，1H)；LCMS:99.1％，m/z[M-H]^-＝628.1。

67.9a Synthesis

To a solution of 67.8a (170mg, 0.269mmol) in THF (75mL) was added 1% NaHCO at room temperature₃Solution (150 mL). The resulting reaction mixture was stirred at room temperature for 2 days. After completion of the reaction, the pH of the solution was adjusted to 6-7 with 1N HCl solution and extracted with EtOAc (2 × 40 mL). The combined organic layers were washed with brine (30mL) over anhydrous Na₂SO₄Dried, filtered and concentrated under reduced pressure to give a residue. The resulting residue was purified by preparative HPLC [ X-bridge C18(150X30) mm, 5 μ; a is 0.1% formic acid water solution, B is acetonitrile; gradient: (time/% B):0/50, 8/80, 10/80, 13/98, 14/98, 14.1/50, 17/50 at a flow rate of 25mL/min]To give 67.9a (12mg, 7%) as a white solid.¹H NMR(500MHz，DMSO-d₆):12.91(br s，1H)，11.23(br s，1H)，10.92(br s，1H)，7.55-7.27(m，9H)，4.85-4.68(m，1H)，4.25-4.05(m，1H)，3.55-3.40(m，1H)；LCMS:96.9％，m/z[M-H]^-＝646.1。

67.9b Synthesis

To a stirred solution of 67.8b (400mg, 0.633mmol) in THF (150mL) at room temperature was added 1% NaHCO₃Solution (300 mL). The resulting reaction mixture was stirred at room temperature for 2 days. After the reaction was complete, the pH of the solution was adjusted to 6-7 with 1N HCl solution and EtOAc (2 × 60 mL). The combined organic layers were washed with brine (60mL) over anhydrous Na₂SO₄Dried, filtered and concentrated under reduced pressure to give a residue. The resulting residue was purified by preparative HPLC [ X-bridge C18(150X30) mm, 5 μ; a is 0.1% formic acid water solution, B is acetonitrile; gradient: (time/% B):0/65, 8/90, 10/90, 10.1/98, 12/98, 12.1/65, 15/65at a flow rate of 25mL/min]67.9b (40mg, 10%) was produced as a white solid.¹H NMR(500MHz，DMSO-d₆):12.48(br s，1H)，10.94(br s，1H)，10.52(br s，1H)，8.18(d，J＝1.5Hz，1H)，7.66(d，J＝2.0Hz，2H)，7.54-7.45(m，5H)，7.30(br s，1H)，4.35-4.22(m，1H)，3.95-3.86(m，1H)，3.72-3.62(m，1H)，3.60-3.50(m，1H)；LCMS:96.1％，m/z[M-H]^-＝646.1。

Table 1:

following examples 66 and 67, the following diastereomeric examples are prepared. Depending on the substituent at the corresponding position on analogue intermediate 66.8a or 66.8b, it may be possible to find the epimerized product or, in the case of the corresponding analogue intermediate 67.8a or 67.8b, to isolate the desired product without epimerization.

Example 80: 5, 7-dichloro-6 ', 6' -difluoro-1 ' - ((6-methoxy-4- (trifluoromethyl) pyridin-2-yl) carbamoyl) -2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -2' -carboxylic acid, 80.7a.1, 80.7a.2, 80.7b.1&80.7b.2

80.2 Synthesis:

80.1(15g, 69.4mmol) in NH was stirred in a stainless steel bowl at 180 deg.C₄Solution in OH (150mL) for 16 h. The reaction mixture was cooled to room temperature, diluted with water and extracted with ethyl acetate (3 × 200 mL). The combined organic layers were concentrated under reduced pressure. The residue was purified by column chromatography (silica gel 100-.

¹HNMR(400MHz，CDCl₃):6.85(s，1H)，6.58(s，1H)，4.96(br s，2H)；LCMS:76.1％，m/z[M+H]⁺＝197.0。

80.3 Synthesis:

80.2(12g, 61.0mmol) was dissolved in a solution of 25% NaOH in MeOH (120 mL). Stir in a stainless steel bowl for 16 hours at 100 ℃, then cool the reaction mixture to room temperature, dilute with water and extract with ethyl acetate (3x200 mL). The combined organic phases were concentrated in vacuo. The residue was purified by column chromatography (silica gel 100-200mesh, 20% EtOAc in petroleum ether) to yield 80.3(9.8g, 83%) as a solid.

¹H NMR(400MHz，CDCl₃):6.28(s，1H)，6.23(s，1H)，4.56(br s，2H)，3.87(s，3H)；LCMS:88.1％，m/z[M+H]⁺＝193.1。

80.4 Synthesis:

to a solution of furan-2, 5-dione (4.49g, 45.8mmol) in MTBE (100mL) was added 80.3(8.8g, 45.8mmol) at room temperature. After stirring at room temperature for 16 hours, the precipitated solid was filtered and dried in vacuo. The residue was washed with MTBE to give 80.4(13.0g, 66%) as a solid.

¹H NMR(500MHz，CDCl₃):8.46(d，J＝6.5Hz，1H)，8.03(s，1H)，6.87(s，1H)，6.54(d，J＝12.5Hz，1H)，6.41(d，J＝12.5Hz，1H)，3.93(s，3H)；LCMS:98.4％，m/z[M+H]⁺＝291.0。

80.5 Synthesis:

80.4(5.0g, 19.2mmol) in Ac at room temperature₂To the stirred solution of O (50mL) was added NaOAc (1.57g, 19.2 mmol). Stirring at 80 ℃ for 2 hours, cooling the reaction mixture to room temperature and then evaporating excess Ac under reduced pressure₂And O. The resulting residue was diluted with DCM (50mL) and washed with water (2 × 25 mL). With Na₂SO₄The combined organic layers were dried, filtered and concentrated. Purification of the residue by flash column chromatography (80g silica gel column, 20% EtOAc in petroleum ether) afforded 80.5(3.0g, 64%) as a solid.

¹H NMR(400MHz，CDCl₃):7.14(s，1H)，6.99(s，1H)，6.90(s，2H)，3.97(s，3H)；LCMS:98.1％，m/z[M+H]⁺＝273.0。

80.6 Synthesis:

to a solution of 80.5(2.5g, 9.19mmol) in EtOH (30mL) was added 5, 7-dichloroindole-2, 3-dione (1.98g, 9.19mmol), (S) -4, 4-difluoropyrrolidine-2-carboxylic acid (2.27g, 9.19mmol) at room temperature. The reaction mixture was stirred at 80 ℃ for 2 hours and concentrated under reduced pressure. Purification of the resulting residue by flash chromatography (120g silica gel column, 30% EtOAc/petroleum ether) gave the major diastereomer 80.6(4.4g, 84%) as a yellow solid.

¹H NMR(500MHz，DMSO-d₆):11.40(s，1H)，7.59(d，J＝2.0Hz，1H)，7.46(s，1H)，7.39(d，J＝1.0Hz，1H)，7.14(d，J＝1.5Hz，1H)，4.44-4.39(m，1H)，4.02(s，3H)，3.99-3.91(m，1H)，3.90(t，J＝7.5Hz，1H)，3.09-3.05(m，1H)，2.69-2.63(m，1H)，2.57-2.51(m，1H)，2.36-2.30(m，1H)；LCMS:96.9％，m/z[M-H]^-574.8; chiral purity (46.6+ 47.9)%.

80.6a &80.6b preparation (absolute stereochemistry of enantiomers 1 and 2 was not determined):

compound 80.6(4.4g) was purified by chiral SFC using chiral package IG (250x30) mm, 5 μ; solution of 0.2% TFA in n-hexane: isopropanol (85:15), RT (isocratic 42.0mL/min, 13min run time, detection wavelength 254 nm). The pure fractions were concentrated under reduced pressure to give 480mg of 80.6a (enantiomer-1) and 470mg of 80.6b (enantiomer-2) as yellow solids.

80.6a:¹H NMR(500MHz，DMSO-d₆):11.40(s，1H)，7.59(d，J＝2.0Hz，1H)，7.46(s，1H)，7.39(d，J＝0.5Hz，1H)，7.14(d，J＝2.0Hz，1H)，4.44-4.39(m，1H)，4.02(s，3H)，3.98(d，J＝7.5Hz，1H)，3.90(t，J＝7.5Hz，1H)，3.09-3.07(m，1H)，2.71-2.65(m，1H)，2.60-2.51(m，1H)，2.36-2.29(m，1H)；LCMS:99.5％，m/z[M+H]⁺576.9; the chiral purity is 98.0 percent.

80.6b:¹H NMR(500MHz，DMSO-d₆):11.04(s，1H)，7.59(d，J＝2.0Hz，1H)，7.46(s，1H)，7.39(s，1H)，7.14(d，J＝1.5Hz，1H)，4.44-4.39(m，1H)，4.02(s，3H)，3.98(d，J＝7.5Hz，1H)，3.90(t，J＝7.5Hz，1H)，3.11-3.05(m，1H)，2.69-2.63(m，1H)，2.57-2.51(m，1H)，2.35-2.30(m，1H)；LCMS:98.6％，m/z[M+H]⁺576.9; the chiral purity is 99.9 percent.

Synthesis of 80.7a.1&80.7 a.2:

to a stirred solution of 80.6a (300mg, 0.52mmol) in THF (100mL) at room temperature was added 1% NaHCO₃Solution (200 mL). The resulting reaction mixture was stirred at room temperature for 16 hours. After completion of the reaction, the pH of the solution obtained from the reaction was adjusted to about 6-7 with 1N HCl solution and extracted with ethyl acetate (2 × 50 mL). The combined organic phases were washed with brine (60ml) and anhydrous Na₂SO₄Dried, filtered and concentrated under reduced pressure to yield a residue. Preparative HPLC [ X-bridge C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/40, 8/80, 9/80, 9.1/98, 14/98, 14.1/40, 17/40 flow rate 25mL/min]The residue was purified to give 80.7a.1(11mg) and 80.7a.2(208mg) as white solids.

80.7a.1:¹H NMR(500MHz，DMSO-d₆):12.74(br s，1H)，11.11(br s，1H)，7.83(s，1H)，7.49(s，1H)，7.05(br s，1H)，6.88(s，1H)，4.25-4.15(m，2H)，3.83(s，3H)，3.78(d，J＝7.0Hz，1H)，3.55-3.50(m，1H)，3.24-3.15(m，1H)，2.85-7.73(m，1H)，2.40-2.30(m，1H)；LCMS:98.6％，m/z[M+H]⁺＝595.2。

80.7a.2:¹H NMR(500MHz，DMSO-d₆):12.49(br s，1H)，11.08(br s，1H)，11.04(br s，1H)，8.05(s，1H)，8.02(s，1H)，7.48(d，J＝1.5Hz，1H)，6.92(s，1H)，4.08-4.04(m，2H)，3.94(s，3H)，3.85-3.78(m，1H)，3.17-3.14(m，1H)，2.67-2.63(m，1H)，2.50-2.44(m，1H)，2.08-1.90(m，1H)；LCMS:98.2％，m/z[M+H]⁺＝595.2。

Synthesis of 80.7b.1&80.7 b.2:

to a stirred solution of 80.6b (300mg, 0.519mmol) in THF (100mL) at room temperature was added 1% NaHCO₃Solution (200 mL). The resulting reaction mixture was stirred at room temperature for 16 hours. After completion of the reaction, the pH of the solution obtained from the reaction was adjusted to about 6-7 with 1N HCl solution and extracted with ethyl acetate (2 × 50 mL). The combined organic phases were washed with brine (60ml) and anhydrous Na₂SO₄Dried, filtered and concentrated under reduced pressure to yield a residue. Preparative HPLC [ X-bridge C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/50, 9/80, 11/80, 11.1/98, 12/98, 12.1/50, 15/50 flow rate 25mL/min]The residue was purified to give 80.7b.1 as a white solid (13mg) and 80.7b.2 as a white solid (135 mg).

80.7b.1:¹H NMR(500MHz，DMSO-d₆):12.76(br s，1H)，11.12(br s，1H)，7.83(s，1H)，7.49(s，1H)，7.06(br s，1H)，6.89(s，1H)，4.26-4.05(m，2H)，3.83(s，3H)，3.78(d，J＝8.0Hz，1H)，3.55-3.49(m，1H)，3.28-3.11(m，1H)，2.82-2.77(m，1H)，2.38-2.36(m，1H)；LCMS:98.6％，m/z[M+H]⁺＝595.2。

80.7b.2:¹H NMR(500MHz，DMSO-d₆):12.50(br s，1H)，11.08(br s，1H)，11.02(br s，1H)，8.06(br s，1H)，8.02(s，1H)，7.49(d，J＝1.5Hz，1H)，6.92(s，1H)，4.09-4.04(m，2H)，3.94(s，3H)，3.85-3.78(m，1H)，3.19-3.12(m，1H)，2.69-2.62(m，1H)，2.50-2.40(m，1H)，2.08-1.90(m，1H)；LCMS:98.0％，m/z[M+H]⁺＝595.2。

Example 81: synthesis of 5, 7-dichloro-4 ' - ((3, 5-dichlorophenyl) carbamoyl) -1' -methyl-2-oxo-5 ' - (trifluoromethyl) spiro [ indole-3, 2' -pyrrolidine ] -3' -carboxylic acid

Synthesis of 81.2_1 and 81.2_ 2:

to a stirred solution of 81.1(1g, 6.98mmol) in EtOH (40mL) were added 1- (3, 5-dichlorophenyl) -1H-pyrrole-2, 5-dione (1.69g, 6.98mmol) and 5, 7-dichloroindole-2, 3-dione (1.50g, 6.98 mmol). After stirring at 80 ℃ for 3 hours, the reaction mixture was cooled to room temperature and concentrated. The residue was purified by flash chromatography (40g silica gel column, 10% ethyl acetate in petroleum ether) to yield a small amount of diastereomer 81.2_1(400mg, 10%) as a solid and a large amount of diastereomer 81.2_2(2.2g, 59%) as a solid.

81.2_1:¹H NMR(400MHz，DMSO-d₆):11.28(br s，1H)，7.78(t，J＝2.0Hz，1H)，7.54(d，J＝2.0Hz，1H)，7.48(d，J＝2.0Hz，1H)，7.26(d，J＝2.0Hz，2H)，4.88(d，J＝5.2Hz，1H)，4.50-4.46(m，1H)，4.12(d，J＝10.0Hz，1H)，3.95-3.92(m，1H)；LCMS:98.8％，m/z[M-H]^-＝535.9。

81.2_2:¹H NMR(400MHz，DMSO-d₆):11.17(br s，1H)，7.79(t，J＝2.0Hz，1H)，7.53(d，J＝2.0Hz，1H)，7.38(d，J＝2.0Hz，2H)，7.07(d，J＝2.0Hz，1H)，4.88-4.83(m，2H)，4.06-4.01(m，1H)，3.71(d，J＝8.4Hz，1H)；LCMS:98.3％，m/z[M-H]^-＝535.9。

81.2_2a &81.2_2b separation:

at room temperature, 5 μ using (R, R) Whelk-01(30X250 mm); 80% CO₂Chiral SFC purification of 81.2_2(2.2g) with 20% acetonitrile (isocratic 90g/min, detected at 214 nm) yielded 81.2_2a (enantiomer-1, 900mg, 82%) as a solid and 81.2_2b (enantiomer-2, 850mg, 77%) as a solid. (Absolute stereology of enantiomers 1 and 2)Bulk chemistry not detected).

81.2_2a:¹H NMR(400MHz，DMSO-d₆):11.17(br s，1H)，7.80(t，J＝2.0Hz，1H)，7.54(d，J＝2.0Hz，1H)，7.38(d，J＝2.0Hz，2H)，7.07(d，J＝2.0Hz，1H)，4.91-4.84(m，2H)，4.05-4.01(m，1H)，3.71(d，J＝8.0Hz，1H)；LCMS:99.4％，m/z[M-H]^-＝535.9。

81.2_2b:¹H NMR(400MHz，DMSO-d₆):11.18(br s，1H)，7.80(t，J＝2.0Hz，1H)，7.53(d，J＝2.0Hz，1H)，7.38(d，J＝2.0Hz，2H)，7.07(d，J＝1.6Hz，1H)，4.89-4.84(m，2H)，4.05-4.01(m，1H)，3.71(d，J＝8.0Hz，1H)；LCMS:99.3％，m/z[M-H]^-＝535.9。

81.3 Synthesis:

to a stirred solution of 81.2_2b (200mg, 0.37mmol) in DCM (5mL) at 0 deg.C was added TEA (0.3mL, 2.14mmol) followed by trifluoromethyl (0.3mL, 2.74 mmol). After stirring at room temperature for 16h, the reaction mixture was quenched with ice-water and extracted with ethyl acetate (2 × 20mL). With Na₂SO₄The combined organic phases were dried, filtered and concentrated under vacuum. The resulting residue was purified by preparative HPLC [ column: INERTSIL-ODS 2(250X19mm), 5. mu.M; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B) 0/55, 8/80, 11/90, 11.1/98, 13/98, 13.1/55, 17/55 flow rate is 18mL/min]Purification to give 81.3 as a white solid (14mg, 7%).

¹H NMR(500MHz，DMSO-d₆):11.51(br s，1H)，7.80(t，J＝2.0Hz，1H)，7.59(d，J＝1.5Hz，1H)，7.40(d，J＝1.5Hz，2H)，7.10(d，J＝1.5Hz，1H)，4.57-4.54(m，1H)，4.11(t，J＝9.0Hz，1H)，3.78(d，J＝8.5Hz，1H)，2.13(s，3H)；LCMS:98.7％，m/z[M-H]^-＝549.9。

81 Synthesis:

to a stirred solution of 81.3(30mg, 0.054mmol) in THF (20mL) at room temperature was added 1% NaHCO₃Solution (41 mL). Stirring at room temperature for 16 hours, adjusting the pH of the reaction solution to about 6-7 with 1N HCl solution, and adding ethyl acetate (2X)10mL) was extracted. The combined organic phases were washed with brine (10ml), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: symmerry-C8 (300x19mm), 7 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B) 0/65, 8/90, 10/90, 10.1/98, 12/98, 12.1/65, 16/65 at a flow rate of 18mL/min]Yield compound 81(6mg, 19%) as a solid.

¹H NMR(500MHz，DMSO-d₆):12.61(br s，1H)，11.02(br s，1H)，10.70(s，1H)，8.22(s，1H)，7.67(d，J＝1.5Hz，2H)，7.39(s，1H)，7.27(s，1H)，4.13-4.03(m，1H)，3.81-3.70(m，1H)，3.62-3.55(m，1H)，2.05(s，3H)；LCMS:98.7％，m/z[M-H]^-＝567.9。

Example 82: (1'R, 2' S, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino-cyclo ] -2' -carboxylic acid (82.6a) and

synthesis of (1'S, 2' R, 7a 'S) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino-cyclo ] -2' -carboxylic acid, (82.6b)

82.4_1&82.4_2 Synthesis

To a stirred solution of (S) -pyrrolidine-2-carboxylic acid, 82.1, (10g, 66.2mmol) in MeCN (100mL) was added (Z) -4- (allyloxy) -4-oxybutan-2-dioic acid, 82.2, (10.3g, 66.2mmol) and 5, 7-dichloroindole-2, 3-dione, 82.3, (14.3g, 66.2mmol) at room temperature. After refluxing for 2 hours, the reaction mixture was cooled to room temperature. The resulting precipitate was filtered and washed with MeCN (2 × 20mL) and then dried under high vacuum to yield 82.4_1&82.4_2(LCMS ratio: 26: 35).

82.4_1: rac- (1'R, 2' S, 3R, 7a 'R) -2' - ((allyloxy) carboxy) -5, 7-dichloro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexa-kis-phenylHydrospiro [ indole-3, 3' -pterin ring]-1' -carboxylic acid.¹H NMR(500MHz，DMSO-d₆):12.56(br s，1H)，11.00(s，1H)，7.77(d，J＝2.0Hz，1H)，7.46(d，J＝2.0Hz，1H)，5.50-5.45(m，1H)，5.10-5.05(m，2H)，4.26(d，J＝5.5Hz，2H)，4.06(d，J＝7.5Hz，1H)，3.97-3.96(m，1H)，3.55-3.52(m，1H)，2.64-2.62(m，1H)，2.26-2.23(m，1H)，1.91-1.78(m，3H)，1.53-1.49(m，1H)；LCMS:94.0％，m/z[M+H]⁺425.0; chiral purity (49.7+ 50.2)%. Regiochemistry and relative stereochemistry were confirmed by 2d nmr studies.

82.4_2:¹H NMR(500MHz，DMSO-d₆):12.40(br s，1H)，10.95(s，1H)，7.56(d，J＝2.0Hz，1H)，7.51(d，J＝2.0Hz，1H)，5.86-5.80(m，1H)，5.27-5.15(m，2H)，4.45-4.39(m，2H)，4.27-4.25(m，1H)，3.64(d，J＝8.5Hz，1H)，3.47-3.44(m，1H)，2.51-2.48(m，1H)，2.42-2.35(m，1H)，2.10-2.00(m，1H)，1.90-1.80(m，1H)，1.73-1.71(m，2H)；LCMS:80.7％，m/z[M+H]⁺425.0. The chemistry of the region of unknown interest.

82.4_1a &82.4_1b separation

Separation of 82.4-1 (10g) with chiral pack-IG (4.6X250) mm, 5. mu. chiral SFC; a concentration of 0.5% TFA in isopropanol at room temperature (isocratic 42.0mL/min, 16min run time detected at 214 nm) gave 1.8g 82.4. mu.1 a (peak-1) as a white solid and 3.8g 82.4-1 b (peak-2) as a solid (absolute stereochemistry of enantiomers 1 and 2 was not determined).

82.4_1a:¹H NMR(500MHz，DMSO-d₆):12.55(br s，1H)，11.00(s，1H)，7.77(d，J＝2.0Hz，1H)，7.46(d，J＝2.0Hz，1H)，5.50-5.45(m，1H)，5.10-5.05(m，2H)，4.26(d，J＝5.5Hz，2H)，4.06(d，J＝8.0Hz，1H)，3.97-3.96(m，1H)，3.54-3.51(m，1H)，2.64-2.62(m，1H)，2.26-2.23(m，1H)，1.91-1.80(m，3H)，1.53-1.49(m，1H)；LCMS:99.0％，m/z[M+H]⁺425.0; the chiral purity is 99.9 percent.

82.4_1b:¹H NMR(400MHz，DMSO-d₆):12.58(br s，1H)，11.04(s，1H)，7.76(s，1H)，7.47(d，J＝1.6Hz，1H)，5.53-5.43(m，1H)，5.11-5.05(m，2H)，4.27(d，J＝5.2Hz，2H)，4.07(d，J＝7.6Hz，1H)，4.04-3.97(m，1H)，3.58-3.54(m，1H)，2.70-2.65(m，1H)，2.30-2.27(m，1H)，1.93-1.75(m，3H)，1.59-1.52(m，1H)；LCMS:97.2％，m/z[M+H]⁺425.0; the chiral purity is 97.7%.

82.5a Synthesis

Thionyl chloride (6mL) was added to 82.4_1a (300mg, 0.70mmol) at room temperature. The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. To the resulting acid chloride in CH₂Cl₂₍3mL) was added 3, 5-dichloro-N-neopentylphenylamine (245mg, 1.05mmol) in CH₂Cl₂(2 mL). Stirred at 55 ℃ for 16h, the reaction mixture was quenched with water (20ml) and CH₂Cl₂(2 × 20mL). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by column chromatography (40g silica gel column, 20% EtOAc/petroleum ether) to yield compound 82.5a (200mg, 50%) as a solid.

¹H NMR(400MHz，DMSO-d₆) 10.97/10.92(s, 1H), 8.22/7.70(d, J-2.0 Hz, 1H), 7.64(d, J-1.2 Hz, 1H), 7.54-7.40(m, 3H), 5.52-5.43(m, 1H), 5.21-5.08(m, 2H), 4.34-4.28(m, 1H), 4.22-4.15(m, 2H), 3.83-3.77(m, 1H), 3.71-3.58(m, 3H), 2.70-2.64(m, 1H), 2.20-2.14(m, 1H), 1.97-1.83(m, 1H), 1.81-1.73(m, 2H), 1.57-1.50(m, 1H), 0.89-0.78(m, 9H); LCMS 98.8%, M/z [ M + H ]]⁺＝638.0。

82.6a Synthesis

To a stirred solution of 82.5a (200mg, 0.31mmol) in THF (4mL) at room temperature was added aniline (30mg, 0.31mmol) and Pd (PPh)₃)₄(72mg, 0.06 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ KROMOSIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time min/% B) 0/70, 8/90, 11/95, 11.1/98, 13/98,13.1/70，15/70，20mL/min]this gave compound 82.6a (58mg, 37%) as a solid.¹H NMR(500MHz，DMSO-d₆) 12.44(br s, 1H), 10.90/10.84(s, 1H), 8.37/7.86(d, J ═ 2.0Hz, 1H), 7.62-7.61(m, 1H), 7.53-7.38(m, 3H), 4.08(d, J ═ 8.0Hz, 1H), 3.99-3.97(m, 1H), 3.69-3.65(m, 1H), 3.52-3.45(m, 2H), 2.72-2.68(m, 1H), 2.13-2.12(m, 1H), 1.91-1.87(m, 1H), 1.77-1.70(m, 2H), 1.52-1.48(m, 1H), 0.83(s, 9H); LCMS 98.7%, M/z [ M + H ]]⁺598.0; the chiral purity is 98.0 percent.

82.5b Synthesis:

thionyl chloride (6mL) was added to 82.4_1b (350mg, 0.82mmol) at room temperature. The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. To the resulting acid chloride in CH₂Cl₂(3mL) to a solution of 3, 5-dichloro-N-neopentylphenylamine (286mg, 1.23mmol) in CH was added₂Cl₂(2 mL). Stirred at 55 ℃ for 16h, the reaction mixture was quenched with water (20ml) and CH₂Cl₂(2 × 20mL). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by column chromatography (40g silica gel column, 20% EtOAc/petroleum ether) to yield 82.5b (155mg, 30%) as a solid.

¹H NMR(400MHz，CDCl₃):8.33(d，J＝2.0Hz，1H)，7.38(br s，1H)，7.33(br s，1H)，7.26-7.18(m，2H)，5.49-5.44(m，1H)，5.14-5.06(m，2H)，4.36-4.31(m，1H)，4.24-4.19(m，1H)，4.12(d，J＝8.0Hz，1H)，3.91-3.86(m，1H)，3.75-3.70(m，2H)，3.48-3.44(m，1H)，2.85-2.79(m，1H)，2.38-2.33(m，1H)，2.06-2.01(m，1H)，1.96-1.91(m，1H)，1.88-1.81(m，1H)，1.72-1.65(m，1H)，0.91(m，9H)；LCMS:98.1％，m/z[M+H]⁺＝638.0。

Synthesis of 82.6b

To a stirred solution of 82.5b (155mg, 0.24mmol) in THF (4mL) at room temperature was added aniline (22mg, 0.24mmol) and Pd (PPh)₃)₄(56mg，0.05mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ KROMOSIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time in minutes/% B) 0/50, 8/90, 10/90, 10.1/98, 12/98, 12.1/50, 14/50, 22mL/min to yield compound 82.6B (88mg, 61%) as a solid.¹H NMR(500MHz，DMSO-d₆) 12.50(br s, 1H), 11.00/10.99(s, 1H), 8.37/7.87(d, J ═ 2.0Hz, 1H), 7.63-7.62(m, 1H), 7.53-7.40(m, 3H), 4.13(d, J ═ 8.0Hz, 1H), 4.01-3.98(m, 1H), 3.76-3.60(m, 2H), 3.47-3.45(m, 1H), 2.80-2.69(m, 1H), 2.27-2.20(m, 1H), 1.93-1.90(m, 1H), 1.81-1.75(m, 2H), 1.57-1.54(m, 1H), 0.84(s, 9H); LCMS 97.4%, M/z [ M + H ]]⁺598.0; the chiral purity is 96.0 percent.

Table 2:

the following compounds were prepared according to the procedure described in example 82 using the anilines listed below. The relative stereochemistry was determined by 2d nuclear magnetic studies. The absolute stereochemistry of the enantiomeric pair is unknown (a and b).

Example 85 a: synthesis of (1'S, 2' R, 7a 'S) -5, 7-dichloro-1' - ((cyclopentylmethyl) (3, 5-dichlorophenyl) carbamoyl) -1-methyl-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino-cyclo ] -2' -carboxylic acid:

synthesis 85.1:

at room temperature, 84a (110mg, 0.17mmol) in CH₃Adding Cs to CN (5mL)₂CO₃(66mg, 0.20mmol), followed by the addition of MeI (72mg, 0.50mmol) and stirring at room temperature for 6 h. The reaction mixture was quenched with water (10ml) and the combined organic phases were dried over anhydrous sodium sulfate extracted with ethyl acetate (2 × 20ml), filtered and concentrated under reduced pressure to give 85.1(110mg) as a solid which was used directly in the next step without further purification. LCMS 88.2%, M/z [ M + H ]]⁺＝666.0。

Synthesis 85:

to a stirred solution of 85.1(110mg, 0.16mmol) in THF (3mL) at room temperature were added aniline (15mg, 0.16mmol) and Pd (PPh)₃)₄(38mg, 0.03 mmol). The resulting reaction mixture was stirred at room temperature for 2 hours. After completion of the reaction, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-BRIDGE-C18(150X30), 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/75, 8/90, 10/95, 12/98, 12.1/75, 14/75at 25mL/min]Thus yielding 85(39mg, 38%) as a solid. The absolute stereochemistry of the enantiomeric pair is unknown (a and b).

¹H NMR(500MHz，DMSO-d₆) (in wheel form): 12.40(br s, 1H), 8.48/8.03(d, J ═ 2.0Hz, 1H), 7.68/7.56(t, J ═ 2.0Hz, 1H), 7.45/7.41(d, J ═ 2.5Hz, 1H), 7.38/7.34(d, J ═ 1.5Hz, 2H), 4.08(d, J ═ 8.0Hz, 1H), 3.90-3.88(m, 1H), 3.67-3.65(m, 1H), 3.55-3.51(m, 1H), 3.45/3.43(s, 3H), 3.40-3.37(m, 1H), 2.68-2.60(m, 1H), 2.12-2.05(m, 1H), 1.92-1.87(m, 2.69(m, 1H), 1.80-1.63 (m, 1H), 1.18-1H), 1.46-1H (m, 1H), 1H), 1.46-1H, 1H); LCMS 98.1%, M/z [ M + H ]]⁺624.0; the chiral purity is 97.4%.

Example 85 b: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-1' - ((cyclopentylmethyl) (3, 5-dichlorophenyl) carbamoyl) -1-methyl-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid:

compound 85b was prepared from 84b following the procedure described in example 85 a.

¹H NMR(500MHz，DMSO-d₆) 12.39(br s, 1H), 8.48/8.03(d, J ═ 2.5Hz, 1H), 7.68/7.56(t, J ═ 2.0Hz, 1H), 7.45/7.41(d, J ═ 2.0Hz, 1H), 7.38-7.34(m, 2H), 4.08(d, J ═ 8.0Hz, 1H), 3.92-3.87(m, 1H), 3.67-3.65(m, 1H), 3.55-3.51(m, 1H), 3.45/3.42(s, 3H), 3.40-3.37(m, 1H), 2.68-2.60(m, 1H), 2.10-2.04(m, 1H), 1.92-1.87(m, 2H), 1.80 (m, 1H), 1.63-1H), 1.27.55-1H (m, 1H), 1.27.27-1H (m, 1H), 1.27.55-3.42 (m, 1H); LCMS 99.6%, M/z [ M + H ]]⁺624.0; the chiral purity is 97.6%. The absolute stereochemistry of the enantiomeric pair is unknown (a and b).

Examples 90 and 91: synthesis of (1' R, 2' S, 3R, 6' S, 7a ' R) -6' - (benzyloxy) -5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -6' -methyl-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrido-cyclo ] -2' -carboxylic acid (90) and (1' R, 2' S, 3R, 6' S, 7a ' R) -5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -6' -hydroxy-6 ' -methyl-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazin cyclo ] -2' -carboxylic acid (91):

synthesis of 90.2:

addition funnel was used to funnel to CH at-40 deg.C₃MgBr(3M in Et₂O, 728mL, 2.18mol) in dry THF (6L) was added dropwise to a solution of 90.1(200g, 872mmol) in dry THF (2L) over 2 hours. Slowly raising the temperature to room temperatureAfter stirring at room temperature for 16h, the reaction mixture was cooled to-5 ℃ and quenched with 1N HCl until the reaction mixture became a clear solution, then extracted with ethyl acetate EtOAc (3 × 3L). The combined organic phases were washed with water and dried over anhydrous sodium sulfate, filtered and concentrated. The resulting residue was purified by column chromatography (silica gel 100-200mesh, 2% MeOH in DCM) to yield 90.2(85g, 40%) as an off-white solid.

¹H NMR(500MHz，DMSO-d₆) 12.50-12.30(br s, 1H), 4.95-4.75(br s, 1H), 4.14-4.08(m, 1H), 3.26-3.18(m, 2H), 2.15-2.08(m, 1H), 1.95-1.91(m, 1H), 1.39/1.34(s, 9H), 1.21(s, 3H); LCMS 99.3%, M/z [ M-H ]]^-＝244.1。

Synthesis of 90.3:

to a stirred solution of 90.2(15g, 61.1mmol) in THF (150ml) at 0 deg.C was added NaH (14.6g, 367 mmol). After stirring at 65 ℃ for 30 minutes, benzyl bromide (10.9mL, 91.7mmol) was added at 65 ℃. After stirring at 65 ℃ for 16h, the reaction mixture was cooled to 0 ℃, quenched with 10% citric acid solution and extracted with ethyl acetate (3 × 100 mL). The combined organic phases were washed with water and dried over anhydrous sodium sulfate, filtered and concentrated. The resulting residue was purified by column chromatography (silica gel 100-200mesh, 30% ethyl acetate in petroleum ether) to yield 90.3 as a light brown solid (12g, 58%).

¹H NMR(400MHz，DMSO-d₆):12.44(br s，1H)，7.33-7.22(m，5H)，4.42-4.34(m，2H)，4.24-4.17(m，1H)，3.47(d，J＝11.2Hz，1H)，3.32-3.27(m，1H)，2.28-2.16(m，2H)，1.04-1.36(m，12H)；LCMS:94.8％，m/z[M-H]^-＝334.1。

Synthesis of 90.4:

90.3(12g, 35.7mmol) in CH at room temperature₂Cl₂To the stirred solution (240mL) was added TFA (12mL), stirred at room temperature for 16h, and the reaction mixture was concentrated under reduced pressure to give 90.4(12g) as a brown solid. LCMS 86.2%, [ M-TFA + H%]⁺＝236.1。

Synthesis of 90.7:

to a stirred solution of 90.4(3.2g, 13.6mmol) in MTBE (100mL) at room temperature was added Et₃N (1.9mL, 13.6mmol), 90.5(2.12g, 13.6mmol) and 90.6(2.93g, 13.6 mmol). After stirring at 80 ℃ for 16h, the reaction mixture was cooled to room temperature and concentrated under reduced pressure. The resulting residue was purified by column chromatography (silica gel 100-200mesh, 30% ethyl acetate in petroleum ether) to yield 90.7(2.1g, 30%) of a solid. The region and relative stereochemistry of 90.7 was determined by 2D nuclear magnetic analysis.

¹H NMR(400MHz，DMSO-d₆):12.65(br s，1H)，11.00(s，1H)，7.68(s，1H)，7.46(s，1H)，7.27-7.22(m，5H)，5.55-5.42(m，1H)，5.10-5.04(m，2H)，4.38-4.26(m，5H)，4.00(d，J＝7.6Hz，1H)，3.64-3.60(m，1H)，2.67(d，J＝9.6Hz，1H)，2.54-2.48(m，1H)，2.16-2.09(m，1H)，1.61-1.53(m，1H)，1.26(s，3H)；LCMS:80.2％，m/z[M+H]⁺＝545.1。

Synthesis of 90.9:

SOCl₂(5mL) was added to 90.7(1.0g, 1.83 mmol). Stirring at room temperature for 2 hours, evaporating SOCl at 40 ℃ under reduced pressure₂Thereby producing an acid chloride intermediate. 90.8(638mg, 2.75mmol) in CH at room temperature₂Cl₂To the solution in (10mL) was added the acid chloride intermediate. The reaction mixture was stirred at 50 ℃ for 16 hours and concentrated under reduced pressure. The resulting residue was purified by flash chromatography (40g silica gel column, 10% ethyl acetate in petroleum ether) to yield 90.9(950mg, 70%) as a yellow solid.

¹H NMR(500MHz，DMSO-d₆):10.95/10.91(s，1H)，8.14/7.69(d，J＝2.0Hz，1H)，7.64-7.63(m，1H)，7.54-7.42(m，3H)，7.33-7.21(m，5H)，5.67-5.45(m，1H)，5.12-5.10(m，2H)，4.37-4.30(m，4H)，4.24-4.23(m，1H)，4.11(d，J＝8.0Hz，1H)，3.95-3.88(m，1H)，3.74-3.66(m，2H)，2.73(d，J＝9.5Hz，1H)，2.50-2.46(m，1H)，2.15-2.05(m，1H)，1.69-1.59(m，1H)，1.28(s，3H)，0.84/0.82(s，9H)；LCMS:96.0％，m/z[M+H]⁺＝760.6。

Synthesis of 90:

to a stirred solution of 90.9(200mg, 0.26mmol) in THF (5mL) at room temperature were added aniline (29mg, 0.32mmol) and Pd (PPh)₃)₄(61mg, 0.05 mmol). After stirring at room temperature for 2 hours, the resulting reaction mixture was diluted with ethyl acetate (15 ml). The organic solvents were collected, washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated. The resulting residue was purified by flash chromatography (24g silica gel column, 20% ethyl acetate in petroleum ether) followed by trituration with diethyl ether to yield 90(45mg, 24%) solid.

¹H NMR(500MHz，DMSO-d₆) 12.49(br s, 1H), 10.90/10.88(s, 1H), 8.31/7.85(d, J ═ 2.0Hz, 1H), 7.61-7.40(m, 4H), 7.32-7.20(m, 5H), 4.39-4.29(m, 2H), 4.05(d, J ═ 8.0Hz, 1H), 3.96-3.89(m, 2H), 3.61-3.59(m, 1H), 3.51(d, J ═ 13.5Hz, 1H), 2.74(d, J ═ 8.5Hz, 1H), 2.43(d, J ═ 9.0Hz, 1H), 2.03-2.00(m, 1H), 1.62-1.58(m, 1H), 1.26(s, 3.26, 0H), 9.85 (s, 85H); LCMS 99.0%, M/z [ M + H ]]⁺718.0; the chiral purity is 98.8 percent.

91 synthesis:

90(150mg, 0.21mmol) in CH at room temperature₂Cl₂(3mL) to the stirred solution were added TFA (0.3mL) and CF₃SO₃H (0.3 mL). After stirring for 16 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was diluted with ethyl acetate (25ml), washed with water (15ml), brine (15ml), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by preparative HPLC [ column: X-SELECT-C18(150X19), 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/50, 8/90, 10/90, 10.1/98, 11/98, 11.1/50, 14/50at20mL/minute]Thus, 91(45mg, 38%) solid was produced.

¹H NMR(500MHz，DMSO-d₆) 12.38/12.32(br s, 1H), 10.90/10.82(br s, 1H), 8.30/7.79(s, 1H), 7.62-7.39(m, 4H), 4.65/4.54 (in wheel form)(s，1H)，4.17-3.80(m，3H)，3.62-3.49(m，2H)，2.63(d，J＝8.0Hz，1H)，2.20/2.10(d，J＝8.0Hz，1H)，1.71-1.69(m，1H)，1.55-1.51(m，1H)，1.15(s，3H)，0.83(s，9H)；LCMS:99.1％，m/z[M+H]⁺628.0; the chiral purity is 99.6 percent.

Table 3:

the following compounds were prepared using the following anilines, as shown in example 90 or 91, using intermediate 90.7 and the listed anilines.

Example 97: synthesis of (1' R, 2' S, 3R, 6' S, 7a ' R) -6' - (phenylmethyloxy) -5, 7-dichloro-1 ' - ((cyclopentylmethyl) (3, 5-dichlorophenyl) carbamoyl) -1, 6' -dimethyl-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -2' -carboxylic acid:

97.2 Synthesis:

SOCl₂(10mL) was added to 90.7(1.2g, 2.20 mmol). Stirring at room temperature for 2 hours, evaporating SOCl under reduced pressure₂Thereby producing an acid chloride intermediate. 97.1(805mg, 3.30mmol) in CH at room temperature₂Cl₂₍15mL) was added to the stirred solutionPreparing an acid chloride intermediate. The reaction mixture was stirred at room temperature for 16 hours and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 15% ethyl acetate in petroleum ether) to yield 97.2(800mg, 50%) solid.

¹H NMR(400MHz，CDCl₃):8.35(d，J＝1.6Hz，1H)，7.42(s，1H)，7.36(t，J＝1.6Hz，1H)，7.33-7.17(m，7H)，5.51-5.44(m，1H)，5.14-5.06(m，2H)，4.40-4.32(m，3H)，4.27-4.17(m，2H)，4.11(d，J＝8.0Hz，1H)，3.89-3.84(m，1H)，3.67-3.61(m，1H)，3.36(t，J＝8.0Hz，1H)，2.86(d，J＝9.2Hz，1H)，2.64(d，J＝8.8Hz，1H)，2.10-2.00(m，2H)，1.80-1.66(m，5H)，1.39(s，3H)，1.29-1.24(m，3H)，0.93-0.83(m，1H)；LCMS:97.3％，m/z[M+H]⁺＝772.49。

97.3 Synthesis:

to 97.2(300mg, 0.39mmol) in CH at room temperature₃CN (5mL) and K was added to the stirred solution₂CO₃(80mg, 0.58mmol) and CH₃I (0.05mL, 0.78 mmol). After stirring at room temperature for 5 hours, the resulting reaction mixture was diluted with ethyl acetate (15 ml). The organic solution was washed with water (20ml), brine (20ml), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (24g silica gel column, 5% ethyl acetate in petroleum ether) to yield 97.3(220mg, 72%) as a solid.

¹H NMR(400MHz，CDCl₃):8.37(d，J＝2.0Hz，1H)，7.37-7.16(m，9H)，5.52-5.48(m，1H)，5.17-5.07(m，2H)，4.41-4.15(m，5H)，4.06(d，J＝8.0Hz，1H)，3.89-3.84(m，1H)，3.65-3.60(m，1H)，3.51(s，3H)，3.39-3.35(m，1H)，2.78(d，J＝8.8Hz，1H)，2.56(d，J＝8.8 4Hz，1H)，2.06-2.02(m，2H)，1.76-1.53(m，6H)，1.39(s，3H)，1.28-1.24(m，3H)；LCMS:94.4％，m/z[M+H]⁺＝786.4。

97 Synthesis:

to a stirred solution of 97.3(220mg, 0.28mmol) in THF (3mL) was added aniline (26mg, 0.28 m) at room temperaturemol) and Pd (PPh)₃)₄(64mg, 0.06 mmol). After stirring for 2 hours, the reaction mixture was concentrated under pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMOSIL-C18(150X25) mm, 10. mu.l, A: 0.1% formic acid in water, B: acetonitrile; gradient: (time/% B) 0/80, 8/95, 11/95, 11/98, 12.1/98, 12.1/80, 15/80at 25mL/minute]This gave 97(58mg, 27%) of a solid.

¹H NMR(500MHz，DMSO-d₆) 12.43(s, 1H), 8.42/8.00(d, J-2.5 Hz, 1H), 7.67/7.56(t, J-2.0 Hz, 1H), 7.47(d, J-2.0 Hz, 1H), 7.43-7.41(m, 2H), 7.37-7.23(m, 5H), 4.36-4.29(m, 2H), 4.04(d, J-8.0 Hz, 1H), 3.92-3.88(m, 2H), 3.57-3.53(m, 1H), 3.50-3.45(m, 1H), 3.45/3.41(s, 3H), 2.67(d, J-9.0 Hz, 1H), 2.37(d, J-9.0 Hz, 1H), 1.95-1H, 1.65 (m, 1H), 1.95-1H), 1.35-1H, 6-6.65 (m, 1H), 1H, 6-6H, 6(m, 1H), 1.19-1.12(m, 1H); LCMS 99.1%, M/z [ M + H ]]⁺743.9; the chiral purity is 99.8 percent.

Example 100: synthesis of rac- (1'R, 2' S, 7a 'R) -6, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid (100.6b)

Synthesis of 100.4a &100.4b

To a stirred solution of 100.2(500mg, 3.18mmol) in EtOH (10mL) was added 100.1(789mg, 3.18mmol) and 100.3(680mg, 3.18mmol) at room temperature. After 2 hours of reflux, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by flash chromatography (80g silica gel column, 30% -35% EtOAc/petroleum ether) to yield diastereomer 100.4a (150mg, 10%) as a white solid in trace amount and diastereomer 100.4b (250mg, 17%) as a white solid in bulk amount.

100.4a:¹H NMR(500MHz，DMSO-d₆):12.65(s，1H)，11.13(s，1H)，7.54(d，J＝8.0Hz，1H)，7.28(d，J＝8.0Hz，1H)，5.86-5.80(m，1H)，5.27-5.15(m，2H)，4.47-4.39(m，3H)，3.73-3.68(m，2H)，3.10-2.95(m，1H)，2.88-2.76(m，1H)，2.62-2.40(m，2H)；LCMS:88.6％，m/z[M+H]⁺461.0. The regional chemistry and the relative stereochemistry were confirmed by 2d NMR studies

100.4b:¹H NMR(400MHz，DMSO-d₆):12.80(s，1H)，11.17(s，1H)，7.55(d，J＝8.4Hz，1H)，7.25(d，J＝8.4Hz，1H)，5.53-5.44(m，1H)，5.11-5.04(m，2H)，4.34-4.22(m，2H)，4.15-4.09(m，1H)，3.92(d，J＝7.6Hz，1H)，3.76-3.72(m，1H)，3.32-3.24(m，1H)，2.75-2.67(m，1H)，2.51-2.40(m，1H)，2.35-2.15(m，1H)；LCMS:89.1％，m/z[M-H]^-459.0. The regional chemistry and the relative stereochemistry were confirmed by 2d NMR studies

100.5b Synthesis

Thionyl chloride (3mL) was added to 100.4b (250mg, 0.54mmol) at room temperature. After stirring for 2 hours, the reaction mixture was concentrated under reduced pressure to yield an acidic chloride residue. To this acid chloride is in CH₂Cl₂(5mL) to a solution of 3, 5-dichloro-N-methylaniline (144mg, 0.81mmol) in CH was added₂Cl₂(5 mL). After stirring at room temperature for 16 hours, the reaction was quenched with water (10ml). Separating the organic layer with CH₂Cl₂Extract the aqueous layer (2x15 mL). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography (X-BRIDGE-C18(150X30) mm, 5 u; A: 0.1% formic acid in water, B: acetonitrile; gradient (T% B):0/70, 8/85, 11/85, 11.1/98, 12/98, 12.1/70, 15/70at 20mL/min) to yield 100.5B (80mg, 24%) solid, LCMS: 96.6%, M/z [ M + H ] 96.6%, M/z]⁺＝618.1。

100.6b Synthesis

To a stirred solution of 100.5b (80mg, 0.13mmol) in THF (2mL) at room temperature were added aniline (10mg, 0.13mmol) and Pd (PPh)₃)₄(29mg, 0.03 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative HPLC [ atlantins-T3 (250x20) mm, 5 μ; a is a solution of 10mM ammonium bicarbonate in water, B is acetonitrile; gradient: (time/% B) 0/55, 8/80, 11/90, 11.1/98, 13/98, 13.1/55, 16/55at 18mL/min]This gave 100.6b (25mg, 34%) as a solid.

¹H NMR(500MHz，DMSO-d₆) 12.44(br s, 1H), 11.10/11.01(s, 1H), 8.20/7.80(d, J ═ 8.0Hz, 1H), 7.66-7.20(m, 4H), 4.25-4.07(m, 1H), 3.99(d, J ═ 7.5Hz, 1H), 3.85-3.82(m, 1H), 3.55(t, J ═ 7.5Hz, 1H), 3.39/3.23(s, 3H), 2.60-2.50(m, 1H), 2.42-2.35(m, 1H), 2.25-2.05(m, 1H); LCMS 95.2%, M/z [ M + H ]]⁺578.0; chiral purity (49.1+ 48.6)%.

Table 4:

the following compounds were prepared as shown in example 100 using various isatins instead of 6, 7-dichloroindole-2, 3-dione, 100.3.

Example 110: synthesis of (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid (110)

Synthesis 110.4_1&110.4_2&110.4_3&110.4_4

To a solution of (S) -4, 4-difluoropyrrolidine-2-carboxylic acid, 110.1, (10g, 66.2mmol) in MTBE (200mL) was added (Z) -4- (allyloxy) -4-oxybutan-2-oic acid, 110.2, (10.3g, 66.2mmol) and 5, 7-dichloroindole-2, 3-dione, 110.3, (14.3g, 66.2mmol) at room temperature. After stirring at reflux for 16 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by column chromatography (silica gel, 100-200mesh, gradient 20% -25% EtOAc/petroleum ether) to yield 110.4_1, 110.4_2, 110.4_3&110.4_4 (LCMS ratio: 25:15:6: 5).

110.4_1 rac- (1' R, 2' S, 3R, 7a ' R) -2' - ((allyloxy) carboxy) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pipecolin ring ]]-1' -carboxylic acid¹H NMR(500MHz，DMSO-d₆):12.89(br s，1H)，11.15(s，1H)，7.69(d，J＝2.0Hz，1H)，7.50(d，J＝2.0Hz，1H)，5.47-5.44(m，1H)，5.11-5.06(m，2H)，4.28-4.27(m，2H)，4.07-4.02(m，2H)，3.64(t，J＝7.0Hz，1H)，3.23-3.15(m，1H)，2.75-2.65(m，1H)，2.51-2.49(m，1H)，2.19-2.08(m，1H)，¹⁹F NMR(376.49MHz，DMSO-d₆):-89.57(d，J＝226Hz)，-94.10(d，J＝226Hz)；LCMS 98.0％，m/z[M+H]⁺461.0. The relative regiochemistry was determined by 2D NMR studies.

110.4_2 rac- (1' R, 2' R, 3R, 7a ' R) -2' - ((allyloxy) carboxy) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pipecolin ring ]]-1' -carboxylic acid¹H NMR(400MHz，DMSO-d₆):8.96(br s，1H)，7.62(d，J＝2.0Hz，1H)，7.54(d，J＝2.0Hz，1H)，5.85-5.79(m，1H)，5.30-5.13(m，2H)，4.48-4.35(m，3H)，3.56-3.53(m，2H)，3.00-2.79(m，2H)，2.51-2.40(m，2H)；¹⁹F NMR(376.49MHz，DMSO-d₆):-92.55(d，J＝228Hz)，-100.21(d，J＝228Hz)；LCMS 97.8％，m/z[M+H]⁺461.0. Determination of relative regional chemistry by 2D NMR studies

110.4_3:¹H NMR(400MHz，DMSO-d₆):12.75(br s，1H)，11.09(s，1H)，7.77(d，J＝2.0Hz，1H)，7.48(d，J＝2.0Hz，1H)，6.01-5.91(m，1H)，5.42-5.37(m，1H)，5.26(dd，J＝10.4Hz，1.6Hz，1H)，4.71-4.61(m，2H)，4.06-3.94(m，2H)，3.62(t，J＝6.4Hz，1H)，3.18-3.09(m，1H)，2.70-2.61(m，1H)，2.49-2.32(m，1H)，2.09-1.98(m，1H)；¹⁹F NMR(376.49MHz，DMSO-d₆):-89.13(d，J＝231Hz)，-92.96(d，J＝231Hz)；LCMS99.6％，m/z[M+H]⁺460.9. The chemistry of the region of unknown interest.

110.4_4:¹H NMR(400MHz，DMSO-d₆):12.87(br s，1H)，11.42(s，1H)，7.54(d，J＝2.0Hz，1H)，6.87(d，J＝2.0Hz，1H)，5.76-5.66(m，1H)，5.23-5.16(m，2H)，4.51-4.35(m，3H)，3.87(t，J＝8.0Hz，1H)，3.58(d，J＝7.2Hz，1H)，3.08-3.01(m，1H)，2.89-2.67(m，1H)，2.58-2.32(m，2H)；¹⁹F NMR(376.49MHz，DMSO-d₆):-92.33(d，J＝230Hz)，-101.91(d，J＝230Hz)；LCMS 95.2％，m/z[M+H]⁺461.0. The chemistry of the region of unknown interest.

Separation of 110.4_1a &110.4_1b

At room temperature with chiral package-IG (250x30) mm, 5 μ; 0.2% TFA in n-hexane: chiral SFC purification in solution in isopropanol (85:15) 110.4-1 (45g) (isocratic 42.0mL/min, 13min run time detection wavelength 254 nm). The pure fraction was concentrated under reduced pressure to give 20g of 110.4-1 a (peak-1) and 14.7g of 110.4-1 b (peak-2) as a white solid

110.4_1a (1' R, 2' S, 3R, 7a ' R) -2' - ((allyloxy) carboxy) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino-cyclo]-1' -carboxylic acid. [ alpha ] to]²⁵ _D+78.8(c 1.0，MeOH)；¹H NMR(500MHz，DMSO-d₆):12.86(s，1H)，11.16(s，1H)，7.68(d，J＝2.0Hz，1H)，7.51(d，J＝2.0Hz，1H)，5.49-5.43(m，1H)，5.11-5.06(m，2H)，4.28-4.27(m，2H)，4.07-4.02(m，2H)，3.64(t，J＝6.5Hz，1H)，3.21-3.18(m，1H)，2.69-2.52(m，1H)，2.51-2.50(m，1H)，2.15-2.05(m，1H)；¹⁹F NMR(376.49MHz，DMSO-d₆):-89.56(d，J＝226Hz)，-94.08(d，J＝226Hz)；LCMS 98.9％，m/z[M+H]⁺461.2; the chiral purity is 99.8 percent. The absolute stereochemistry was determined by single crystal x-ray diffraction.

110.4_1b (1' S, 2' R, 3S, 7a ' S) -2' - ((allyloxy) carboxy) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino-ring)]-1' -carboxylic acid [ alpha ]]²⁵ _D-73.2(c 1.0，MeOH)；¹H NMR(400MHz，DMSO-d₆):12.88(br s，1H)，11.16(s，1H)，7.69(d，J＝2.0Hz，1H)，7.50(d，J＝2.0Hz，1H)，5.50-5.42(m，1H)，5.11-5.06(m，2H)，4.28-4.26(m，2H)，4.09-4.01(m，2H)，3.64(t，J＝7.2Hz，1H)，3.23-3.14(m，1H)，2.74-2.65(m，1H)，2.53-2.44(m，1H)，2.16-2.08(m，1H)；¹⁹F NMR(376.49MHz，DMSO-d₆):-89.56(d，J＝226Hz)，-94.08(d，J＝226Hz)；LCMS 98.7％，m/z[M+H]⁺461.0; the chiral purity is 99.9 percent.

Synthesis of 110.5a

Thionyl chloride (8mL) was added to 110.4_1a (500mg, 1.08mmol) at room temperature. The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The residue obtained is dissolved in CH₂Cl₂(10mL), to which was added 3, 5-dichloro-N-neopentylphenylaniline (500mg, 2.16mmol) in CH₂Cl₂(5 mL). Stirred at 55 ℃ for 16h, the reaction mixture was quenched with water (20ml) and CH₂Cl₂(2 × 20mL). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by column chromatography (40g silica gel column, gradient 10% EtOAc/petroleum ether) to yield 110.5a (410mg, 53%) as a solid.

¹H NMR(400MHz，CDCl₃):8.28(d，J＝2.0Hz，1H)，7.43(s，1H)，7.34(t，J＝1.6Hz，1H)，7.27-7.26(m，2H)，5.48-5.42(m，1H)，5.14-5.06(m，2H)，4.36-4.31(m，1H)，4.24-4.19(m，1H)，4.04-3.97(m，2H)，3.81-3.68(m，2H)，3.51(t，J＝7.6Hz，1H)，3.33-3.26(m，1H)，2.77-2.69(m，1H)，2.31-2.17(m，2H)，0.91(s，9H)；LCMS 99.3％，m/z[M+H]⁺＝674.0。

110 Synthesis

To a solution of 110.5a (400mg, 0.59mmol) in THF (10mL) at room temperature was added aniline (55mg, 0.59mmol) and Pd (PPh)₃)₄(136mg, 0.11 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The residue was purified by preparative HPLC (KROMOSIL-C18(150X30) mm, 10. mu.; A: 0.1% formic acid in water; B: acetonitrile; gradient (time in minutes/% B):0/70, 8/90, 10/95, 24mL/min) to yield 110(161mg, 42%) solid.

[α]²⁵ _D-6.41(c 0.5，MeOH)；¹H NMR(500MHz，DMSO-d₆):12.65(br s，1H)，10.98(s，1H)，8.26(d，J＝2.0Hz，1H)，7.81-7.43(m，4H)，4.05(d，J＝7.5Hz，1H)，4.01(d，J＝14Hz，1H)，3.84-3.69(m，1H)，3.60(dd，J＝7.5Hz，J＝6.5Hz，1H)，3.44(d，J＝14.0Hz，1H)，3.32-3.24(m，1H)，2.63-2.52(m，1H)，2.40-2.32(m，1H)，2.11-2.04(m，1H)，0.84/0.82(s，9H)；¹⁹F NMR(470.59MHz，DMSO-d₆):-89.21(d，J＝226Hz)，-97.17(d，J＝226Hz)；LCMS 99.3％，m/z[M+H]⁺633.9; the chiral purity is 98.9 percent.

Example 111: synthesis of (1'S, 2' R, 3S, 7a 'S) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid

Synthesis of 111.1b

Thionyl chloride (5mL) was added to 110.4_1b (300mg, 0.65mmol) at room temperature. After stirring for 2 hours, the reaction mixture was concentrated under reduced pressure. The residue obtained is dissolved in CH₂Cl₂(3mL), then 3, 5-dichloro-N-neopentylphenylaniline (217mg, 0.93mmol) in CH₂Cl₂(2 mL). Stirring at 55 deg.C for 16 hours, adding water (C)10ml) quench the reaction mixture and use CH₂Cl₂(2 × 20mL). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by column chromatography (40g silica gel column, gradient 15% EtOAc/petroleum ether) to yield 111.1b (125mg, 30%) as a solid.

¹H NMR(400MHz，DMSO-d₆):11.10/11.04(s，1H)，8.13(d，J＝2.0Hz，1H)，7.71-7.44(m，4H)，5.46-5.36(m，1H)，5.18-5.05(m，2H)，4.28-4.19(m，2H)，4.14(d，J＝7.6Hz，1H)，3.87-3.78(m，2H)，3.70(t，J＝7.2Hz，1H)，3.61(d，J＝14.0Hz，1H)，3.28-3.26(m，1H)，2.67-2.57(m，1H)，2.44-2.38(m，1H)，2.16-2.07(m，1H)，0.84(s，9H)；LCMS 94.5％，m/z[M+H]⁺＝674.0。

Synthesis 111

To a stirred solution of 111.1b (125mg, 0.18mmol) in THF (3mL) at room temperature was added aniline (17mg, 0.18mmol) and Pd (PPh)₃)₄(42mg, 0.03 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The residue was purified by preparative HPLC (XSELECT-C18(150X30), 5. mu.; A: 0.1% formic acid in water; B: acetonitrile; gradient (time in minutes/% B):0/70, 8/95, 12/98, 24mL/min) to yield 111(40mg, 34%) solid.

[α]²⁵ _D+9.70(c 0.5，MeOH)；¹H NMR(500MHz，DMSO-d₆):12.69(br s，1H)，11.05/10.99(s，1H)，8.27(d，J＝2.0Hz，1H)，7.82-7.43(m，4H)，4.05(d，J＝7.5Hz，1H)，4.01(d，J＝14Hz，1H)，3.84-3.80(m，1H)，3.60(dd，J＝7.5Hz，J＝6.5Hz，1H)，3.45(d，J＝14.0Hz，1H)，3.29-3.27(m，1H)，2.60-2.54(m，1H)，2.38-2.34(m，1H)，2.10-2.05(m，1H)，0.85/0.82(s，9H)；¹⁹F NMR(376.49MHz，DMSO-d₆):-89.21(d，J＝226Hz)，-97.18(d，J＝226Hz)；LCMS 95.9％，m/z[M+H]⁺634.0; the chiral purity is 97.6%.

Table 5:

the following examples were prepared using 110.4_1a or 110.4_1b and the amine-based coupling procedure (as used in the preparation of 110.5a or 111.1 b) and deprotection procedure (110.6a or 111.2b) described below.

Example 200: synthesis of (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-1-methyl-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid:

synthesis of 200.1:

thionyl chloride (6mL) was added to 110.4_1a (300mg, 0.65mmol) at room temperature and stirred for 2 h. The excess thionyl chloride is concentrated under reduced pressure to give the acid chloride derivative. To this acid chloride is in CH₂Cl₂(5mL) to the solution was added 3, 5-dichloro-methylaniline (228mg, 1.3mmol) in CH₂Cl₂(5 mL). After stirring at room temperature for 16 hours, the reaction mixture was quenched with water (10ml) and the organic layer was separated. By CH₂Cl₂The aqueous layer was extracted (2x10mL). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash chromatography (40g silica gel column, 20% ethyl acetate in petroleum ether) to yield 200.1(280mg, 69%) solid LCMS: 62.1%, M/z [ M + H ]]⁺＝620.0。

Synthesis of 200.2:

to a stirred solution of 200.1(280mg, 0.45mmol) in MeCN (10mL) at room temperature was added K₂CO₃(62mg, 0.45mmol) followed by the addition of MeI (69mg, 0.48 mmol). After stirring at room temperature for 16h, the reaction mixture was quenched with water (10mL) and extracted with ethyl acetate (15 mL). The combined organic phases were washed with water (15ml), brine (15ml), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. This gave 200.2(270mg) as a light brown solid. The residue obtained was used in the next step without purification. LCMS 58.1%, M/z [ M + H ]]⁺＝634.2。

Synthesis of 200:

to a stirred solution of 200.2(260mg, 0.41mmol) in THF (10mL) was added aniline (38mg, 0.41mmol) and Pd (PPh) at room temperature₃)₄(95mg, 0.08 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. Trituration was carried out with diethyl ether n-pentane. The resulting residue was purified by preparative HPLC [ column: x-bridge C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/60, 8/85, 10/90, 10.1/98, 13/98, 13.1/60, 16/60, 18mL/min]This gave 200(45mg, 17%) of a solid.

¹H NMR(500MHz，DMSO-d₆) 12.53(br s, 1H), 8.40/7.98(d, J ═ 2.0Hz, 1H), 7.67-7.42(m, 4H), 4.11-4.02(m, 1H), 3.82-3.78(m, 1H), 3.58-3.55(m, 1H), 3.43(s, 3H), 3.23(s, 3H), 3.22-3.16(m, 1H), 2.57-2.50(m, 1H), 2.38-2.30(m,1H)，2.11-2.04(m，1H)；LCMS:95.0％，[M+H]⁺＝592.0。

table 6:

the following compounds were prepared according to the procedure for example 200 using 110.4_1a and the aniline listed herein.

Example 208: synthesis of (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1-cyclopropyl-1' - ((cyclopropylmethyl) (3, 5-dichlorophenyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino-cyclo ] -2' -carboxylic acid:

synthesis 208.1:

208.1 is synthesized from 110.4_1a following the steps described in synthesis 200.1.

Synthesis 208.2:

to a stirred solution of 208.1(300mg, 0.45mmol) in DCM (10mL) was added cyclopropylboronic acid (78mg, 0.91mmol) and TEA (0.12mL, 0.91 mmol). Ten minutes after flushing in oxygen, Cu (OAc) was added₂(82mg, 0.45mmol) and flushed again with oxygen for 5 minutes. After stirring at room temperature for 16h, the reaction mixture was diluted with DCM (20ml) and filtered through a small pad of celite, then washed with DCM (50ml)A polyester diatomite pad. The filtrate was washed with water (20ml) and brine (20ml). The combined organic phases were dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash chromatography (40g silica gel column, 20% ethyl acetate in petroleum ether) to yield 208.2(120mg, 37%) as a solid.

¹H NMR(400MHz，CDCl₃):8.32(d，J＝2.4Hz，1H)，7.41-7.40(m，1H)，7.30-7.26(m，1H)，7.24-7.17(br s，2H)，5.48-5.40(m，1H)，5.16-5.07(m，2H)，4.28-4.23(m，1H)，3.97-3.93(m，2H)，3.72-3.67(m，1H)，3.60-3.55(m，1H)，3.51-3.45(m，1H)，3.35-3.32(m，1H)，3.22-3.13(m，1H)，2.94-2.92(m，1H)，2.65-2.57(m，1H)，2.29-2.15(m，2H)，1.12-1.10(m，2H)，0.98-0.86(m，3H)，0.52-0.50(m，2H)，0.21-0.19(m，2H)；LCMS:98.6％，m/z[M+H]⁺＝700.0。

Synthesis 208:

to a stirred solution of 208.2(100mg, 0.14mmol) in THF (10mL) at room temperature were added aniline (13mg, 0.14mmol) and Pd (PPh)₃)₄(33mg, 0.02 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMOSIL-C18(150X25mm), 10 u; a is 0.1% formic acid solution in water, acetonitrile; gradient: (time/% B):0/60, 8/85, 12/95, 12.1/98, 14/98, 14.1/60, 16/60, 22mL/min]Thus, 208(80mg, 85%) of a solid was produced.

¹H NMR(500MHz，DMSO-d₆) 12.53(br s, 1H), 8.36/7.98(d, J ═ 2.5Hz, 1H), 7.68/7.60(t, J ═ 2.0Hz, 1H), 7.51-7.38(m, 3H), 4.12-3.98(m, 1H), 3.79-3.75(m, 2H), 3.44-3.40(m, 2H), 3.18(m, 1H), 2.96-2.94(m, 1H), 2.51-2.50(m, 1H), 2.41-2.32(m, 1H), 2.15-1.93(m, 1H), 1.13-0.96(m, 2H), 0.88-0.85(m, 2H), 0.80-0.65(m, 1H), 0.42-0.40(m, 0.02-0.02 (m, 1H), 0.16-1H); LCMS 98.2%, M/z [ M + H ]]⁺＝657.9。

Example 216: synthesis of (1'S, 2' R, 3S, 7a 'S) -5-chloro-1' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -6', 6', 7-trifluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid and (1'R, 2' S, 3R, 7a 'R) -5-chloro-1' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -6', 6', 7-trifluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid (216.8a and 216.8b):

synthesis 216.2:

at 0 ℃ 216.1(3.0g, 18.2mmol) in concentrated H₂SO₄(15mL) and CH₃SO₃To the stirred solution of H (15mL) was added trichloroisocyanic acid (2.1g, 9.08 mmol). After stirring at room temperature for 4 hours, the reaction was cooled to 0 ℃ and quenched with ice-cooled water (100 ml). The resulting precipitate was filtered, washed with water (200ml), collected and dried under vacuum to give 216.2(3.5g, 97%) as an orange solid.

¹H NMR(400MHz，DMSO-d₆):11.66(s，1H)，7.77(dd，J＝2.0Hz，10.0Hz，1H)，7.47(d，J＝2.0Hz，1H)；GCMS:97.7％，m/z[M+H]⁺＝200.8。

Synthesis 216.5a and 216.5 b:

to a stirred solution of 216.3(3g, 12.1mmol) in MTBE (100mL) at room temperature were added 216.2(2.4g, 12.1mmol) and 216.4(1.88g, 12.1 mmol). After stirring at 80 ℃ for 16h, the reaction mixture was cooled to room temperature and concentrated under reduced pressure. The diastereomeric mixture (dr 7%: 26%: 27%: 5% LCMS) was purified by column chromatography (silica gel 100-. 2D NMR studies of defined regions and associated stereochemistry.

¹H NMR(500MHz，DMSO-d₆):12.86(br s，1H)，11.23(s，1H)，7.58(d，J＝2.0Hz，1H)，7.42(dd，J＝1.5Hz，9.5Hz，1H)，5.50-5.45(m，1H)，5.11-5.06(m，2H)，4.28-4.27(m，2H)，4.09-4.05(m，1H)，4.01(d，J＝7.5Hz，1H)，3.65(t，J＝7.0Hz，1H)，3.21-3.18(m，1H)，2.69-2.68(m，1H)，2.50-2.46(m，1H)，2.19-2.09(m，1H)；LCMS:97.9％，m/z[M+H]⁺445.0; chiral purity (50.5+ 49.4)%.

216.5a and 216.5b separation

216.5(1.7g) was separated with chiral SFC using chiral OX-H (30X250) mm, 5 μ; a is 75% CO₂% B25% (0.5% DEA in methanol at room temperature (isocratic 90g/min, detection at 214 nm.) the pure fraction was concentrated under reduced pressure to give 216.5a (enantiomer-1, 760mg, 89%) and 216.5B (enantiomer-2, 670mg, 79%) as an off-white solid.

216.5a:¹H NMR(500MHz，DMSO-d₆):12.88(br s，1H)，11.23(s，1H)，7.58(d，J＝1.5Hz，1H)，7.42(dd，J＝2.0Hz，10.0Hz，1H)，5.50-5.45(m，1H)，5.12-5.06(m，2H)，4.28-4.27(m，2H)，4.10-4.05(m，1H)，4.01(d，J＝7.5Hz，1H)，3.65(t，J＝7.0Hz，1H)，3.21-3.18(m，1H)，2.74-2.65(m，1H)，2.50-2.45(m，1H)，2.20-2.09(m，1H)；LCMS:97.5％，m/z[M+H]⁺445.0; the chiral purity is 99.7 percent.

216.5b:¹H NMR(500MHz，DMSO-d₆):12.86(br s，1H)，11.23(s，1H)，7.58(d，J＝2.0Hz，1H)，7.42(dd，J＝2.0Hz，10.0Hz，1H)，5.50-5.45(m，1H)，5.12-5.06(m，2H)，4.29-4.27(m，2H)，4.08-4.06(m，1H)，4.01(d，J＝7.5Hz，1H)，3.65(t，J＝7.0Hz，1H)，3.21-3.18(m，1H)，2.74-2.65(m，1H)，2.50-2.44(m，1H)，2.20-2.09(m，1H)；LCMS:98.7％，m/z[M+H]⁺445.0; the chiral purity is 99.8 percent.

Synthesis 216.7 a:

to 216.5a (200mg, 0.44mmol) was added SOCl₂(3 mL). Stirring at room temperature for 2 hours, evaporating SOCl under reduced pressure₂Thereby producing an intermediate acid chloride. To the intermediate acid chloride prepared above was added 216.6(150mg, 0.64mmol) in CH at room temperature₂Cl₂The solution of (1). Stirring at 50 deg.C for 16 hr, and steamingThe reaction mixture was purified by flash chromatography of the resulting residue (24g silica gel column, 15% ethyl acetate in petroleum ether) to give 216.7a as an off-white solid (150mg, 53%).

¹H NMR(400MHz，CDCl₃):8.18(d，J＝2.0Hz，1H)，7.60(br s，1H)，7.34(t，J＝1.6Hz，1H)，7.26(s，1H)，7.08(dd，J＝2.0Hz，9.6Hz，1H)，5.50-5.42(m，1H)，5.15-5.06(m，2H)，4.37-4.32(m，1H)，4.25-4.20(m，1H)，4.05-3.98(m，2H)，3.81-3.68(m，2H)，3.54-3.49(m，1H)，3.34-3.29(m，1H)，2.75-2.70(m，1H)，2.31-2.10(m，2H)，0.92(s，9H)；LCMS:96.1％，m/z[M+H]⁺＝660.0。

Synthesis 216.8 a:

to a stirred solution of 216.7a (130mg, 0.19mmol) in THF (3mL) at room temperature were added aniline (18mg, 0.19mmol) and Pd (PPh)₃)₄(46mg, 0.04 mmol). After stirring for one hour, the resulting reaction mixture was diluted with ethyl acetate (15 ml). The organic solvent was collected, washed with brine (10mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-BRIDGE-C18(150X30) mm, 5. mu.g, A: 0.1% formic acid in water, B: acetonitrile; gradient: (time/% B) 0/70, 8/90, 9/90, 9.1/98, 11/98, 11.1/70, 14/70, 20mL/min]This gave 216.8a (47mg, 38%) of a solid.

¹H NMR(500MHz，DMSO-d₆) 12.59/12.45(s, 1H), 11.10/11.06(s, 1H), 8.16/7.71(d, J ═ 1.5Hz, 1H), 7.61-7.34(m, 4H), 4.06-4.00(m, 2H), 3.84-3.80(m, 1H), 3.61-3.58(m, 1H), 3.44(d, J ═ 14Hz, 1H), 3.36-3.29(m, 1H), 2.59-2.50(m, 1H), 2.38-2.34(m, 1H), 2.13-2.06(m, 1H), 0.85/0.82(s, 9H); LCMS 98.1%, M/z [ M + H ]]⁺618.0; the chiral purity is 99.9 percent.

Synthesis of 216.7 b:

to 216.5b (200mg, 0.44mmol) was added SOCl₂(3 mL). Stirring at room temperature for 2 hours, evaporating SOCl under reduced pressure₂Thereby producing an intermediate acidA chlorinated compound. To the intermediate acid chloride prepared above was added compound 216.6(150mg, 0.64mmol) in CH at room temperature₂Cl₂The solution of (1). After stirring rapidly for 16h at 50 ℃, the resulting reaction mixture was evaporated and the resulting residue was purified by flash column chromatography (24g silica gel column, 15% ethyl acetate in petroleum ether) to give 216.7b (180mg, 63%) as an off-white solid.

¹H NMR(400MHz，CDCl₃):8.18(d，J＝2.0Hz，1H)，7.65(br s，1H)，7.34(t，J＝1.6Hz，1H)，7.26(s，1H)，7.08(dd，J＝1.6Hz，9.2Hz，1H)，5.50-5.42(m，1H)，5.14-5.06(m，2H)，4.37-4.32(m，1H)，4.25-4.20(m，1H)，4.05-3.98(m，2H)，3.81-3.68(m，2H)，3.54-3.50(m，1H)，3.36-3.27(m，1H)，2.76-2.68(m，1H)，2.31-2.05(m，2H)，0.92(s，9H)；LCMS:90.2％，m/z[M+H]⁺＝660.0。

Synthesis 216.8 b:

to a solution of 216.7b (160mg, 0.24mmol) in THF (3mL) at room temperature was added aniline (23mg, 0.24mmol) and Pd (PPh)₃)₄(56mg, 0.05 mmol). After stirring for one hour, the resulting reaction mixture was diluted with ethyl acetate (15 ml). The organic solvent was collected, washed with brine (10mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-bridge-C18(150X30) mm, 5. mu.g, A: 0.1% formic acid in water, B: acetonitrile; gradient: (time/% B) 0/60, 8/90, 10/90, 10.1/98, 11/98, 11.1/60, 14/60, 20mL/min]This gave 216.8b (93mg, 62%) as a solid.

¹H NMR(500MHz，DMSO-d₆) 12.59/12.45(s, 1H), 11.10/11.06(s, 1H), 8.16/7.71(d, J ═ 1.5Hz, 1H), 7.61-7.34(m, 4H), 4.06-4.00(m, 2H), 3.85-3.80(m, 1H), 3.60(t, J ═ 7.0Hz, 1H), 3.44(d, J ═ 14Hz, 1H), 3.36-3.28(m, 1H), 2.59-2.50(m, 1H), 2.38-2.34(m, 1H), 2.11-2.06(m, 1H), 0.85/0.82(s, 9H); LCMS 97.4%, M/z [ M + H ]]⁺618.0; the chiral purity is 99.6 percent.

Table 7:

the following compounds were prepared as shown in example 216 using the following isatin instead of 5-chloro-7-fluoroindole-2, 3-dione, 216.2, the regiostereochemistry and relative stereochemistry being determined by 2d nuclear magnetic resonance studies. The absolute stereochemistry of the enantiomeric pair is unknown (a and b).

Example 219: synthesis of rac- (1' R, 2' S, 3R, 8a ' R) -5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -2-oxo-1 ', 5', 6', 7', 8', 8a ' -hexahydro-2 ' H-spiro [ indole-3, 3' -indole ring ] -2' -carboxylic acid:

synthesis 219.4:

to a stirred solution of 219.1(3.0g, 23.2mmol) in acetonitrile (50mL) at room temperature was added 219.2(5.0g, 23.2mmol) and 219.3(3.62g, 23.2 mmol). The reaction mixture was stirred at 90 ℃ for 16h and concentrated. The resulting residue was purified by column chromatography (silica gel 100-. The resulting diastereomer mixture was purified by preparative HPLC [ column: X-SELECT-C18(150X30) mm, 5 mu, A: 0.1% formic acid in water, B: acetonitrile; gradient: (time/% B) 0/30, 8/70, 11/70, 12/98, 12.1/98, 15/9, 22mL/min to yield 219.4 as an off-white solid (200mg, 2%). 2D NMR analysis to determine regional chemistry and related stereochemistry.

¹H NMR(500MHz，DMSO-d₆，at 100℃):11.93(br s，1H)，10.62(br s，1H)，7.85(d，J＝2.0Hz，1H)，7.28(d，J＝2.0Hz，1H)，5.51-5.45(m，1H)，5.08-5.03(m，2H)，4.23-4.21(m，2H)，3.72(d，J＝7.5Hz，1H)，3.36-3.33(m，1H)，3.27-3.24(m，1H)，2.29-2.20(m，2H)，1.83-1.72(m，2H)，1.48-1.46(m，1H)，1.22-1.17(m，3H)；LCMS:95.5％，m/z[M+H]⁺439.0; chiral purity (50.8+ 49.2)%.

Synthesis 219.6:

thionyl chloride (2mL) was added to 219.4(110mg, 0.25mmol) at room temperature. After stirring for 2 hours, thionyl chloride was evaporated under reduced pressure to give an acid chloride. 219.5(66mg, 0.38mmol) in CH was added to the intermediate acid chloride at room temperature₂Cl₂(3 mL). The reaction mixture was concentrated under reduced pressure with rapid stirring at 50 ℃ for 16 h. The resulting residue was purified by column chromatography (silica gel 100-200mesh, 20% EtOAc/petroleum ether) to yield 219.6(30mg, 20%) as a light brown solid. LCMS 69.9%, M/z [ M + H ]]⁺＝598.0。

Synthesis 219:

to a stirred solution of 219.6(30mg, 0.05mmol) in THF (2mL) at room temperature were added aniline (5mg, 0.05mmol) and Pd (PPh)₃)₄(11mg, 0.01 mmol). After stirring at room temperature for 2 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMOSIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/60, 8/80, 9/80, 9.1/98, 12/98, 12.1/60, 14/60, 22mL/min]This gave 219(3mg, 10%) as an off-white solid.

¹H NMR(400MHz，DMSO-d₆) 12.40/12.30(s, 1H), 10.94/10.90(s, 1H), 8.32/8.01(d, J ═ 2.0Hz, 1H), 7.63/7.53(t, J ═ 1.6Hz, 1H), 7.46-7.35(m, 3H), 3.64(d, J ═ 7.6Hz, 1H), 3.53(t, J ═ 6.0Hz, 1H), 3.43/3.22(s, 3H), 3.01-2.99(m, 1H), 2.20-2.13(m, 2H), 1.68-1.66(m, 1H), 1.54-1.51(m, 1H), 1.43-1.40(m, 1H), 1.19-1.05(m, 3H); LCMS 90.1%, M/z [ M + H ]]⁺556.1; chiral purity (46.7+ 45.2)%.

Example 220: synthesis of 5, 7-dichloro-8 ' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -2-oxo-1 ', 3', 4', 7', 8', 8a ' -hexahydrospiro [ indole-3, 6' -pyrrolo [2, 1-c ] [1, 4] oxazine ] -7' -carboxylic acid 220.7a and 220.7c:

synthesis 220.4a, 220.4b &220.4 c:

to a stirred solution of 220.1(3.0g, 22.9mmol) in THF (100mL) at room temperature was added a solution of 220.2(4.94g, 22.9mmol), 220.3(3.57g, 22.9mmol) in DIPEA (2.95g, 22.9 mmol). Stirred at 80 ℃ for 16h, the reaction mixture was diluted with water (30ml) and extracted with ethyl acetate (2 × 50 mL). The combined organic phases were washed with brine (25mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to yield a crude mixture of diastereomers (dr ═ 5+3+ 6%). The crude mixture of diastereomers was purified by column chromatography (silica gel 100-200mesh, 30% ethyl acetate in petroleum ether) to yield 220.4a (250mg), 220.4b (60mg), 220.4c (450mg) solids.

220.4a:¹H NMR(500MHz，Acetone-d₆):11.11(s，1H)，9.90(s，1H)，7.98(d，J＝2.0Hz，1H)，7.33(d，J＝2.0Hz，1H)，5.61-5.52(m，1H)，5.14-5.05(m，2H)，4.31-4.28(m，2H)，4.09-4.06(m，1H)，3.85(d，J＝7.0Hz，1H)，3.71-3.68(m，1H)，3.62-3.55(m，2H)，3.29-3.24(m，2H)，2.60-2.59(m，1H)，2.28–2.26(m，1H)；LCMS:93.0％，m/z[M+H]⁺＝441.0。

220.4b:LCMS:80％，m/z[M+H]⁺＝441.1

220.4c:LCMS:68％，m/z[M+H]⁺＝441.1

Synthesis of 220.6 a:

adding SOCl at room temperature₂(5mL) was added to 220.4a (250mg, 0.59 mmol). After stirring for 2 hours, the reaction mixture was concentrated under reduced pressure to yield the intermediate acid chloride. To a solution of the above intermediate acid chloride was added a solution of 220.5(158mg, 0.68mmol) in DCM (10mL). The reaction mixture was stirred at 55 ℃ for 16 hours and concentrated under reduced pressure. The resulting residue was purified by column chromatography (silica gel 100-200mesh, 30% ethyl acetate in petroleum ether) to yield 220.6-1 (40mg, 11%) as a solid.

¹H NMR (500MHz, acetone-d₆):9.82(s，1H)，8.43(d，J＝1.5Hz，1H)，7.58-7.54(m，2H)，7.48(d，J＝2Hz，1H)，7.33(d，J＝2.0Hz，1H)，5.63-5.50(m，1H)，5.15-5.11(m，2H)，4.42-4.40(m，1H)4.29-4.21(m，1H)，3.89-3.83(m，4H)，3.62-3.60(m，2H)，3.31-3.24(m，3H)，2.52-2.49(m，1H)，2.25-2.23(m，1H)，0.91(s，9H)；LCMS:93.5％，m/z[M+H]⁺＝656。

Synthesis 220.7 a:

to a stirred solution of 220.6a (40mg, 0.06mmol) in THF (2mL) at room temperature were added aniline (6mg, 0.06mmol) and Pd (PPh)₃)₄(14mg, 0.01 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative HPLC [ column: KROMOSIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/60, 8/70, 8.1/98, 9/98, 9.1/60, 12/60, 25mL/min]This gave 220.7 — 1(3.0mg, 8%) as a solid.

LCMS:95.1％，m/z[M+H]⁺＝614.1

Synthesis 220.6 c:

adding SOCl at room temperature₂(10mL) was added to 220.4c (450mg, 1.02 mmol). After stirring under nitrogen for 2 hours, the reaction mixture was concentrated under reduced pressure to yield the intermediate acid chloride. To the acid chloride prepared previously was added a solution of 220.5(273mg, 1.18mmol) in DCM (15 mL). The reaction mixture was stirred at 55 ℃ for 16 hours and concentrated under reduced pressure. The resulting residue was purified using silica gel column chromatography (100-.

¹H NMR(400MHz，DMSO-d₆):11.24(s，1H)，7.81-7.68(m，3H)，7.49(d，J＝2.0Hz，1H)，6.65(s，1H)，5.34-5.26(m，1H)，5.06-5.00(m，2H)，4.26-4.21(m，1H)，4.15-3.97(m，3H)，3.87(d，J＝10.0Hz，1H)，3.57(d，J＝10.0Hz，1H)，3.27-3.03(m，4H)，2.50-2.45(m，2H)，2.15-2.11(m，1H)，0.80(s，9H)。LCMS:85.3％，m/z[M+H]⁺＝656。

Synthesis 220.7c:

to a solution of 220.6a (250mg, 0.38mmol) in THF (10mL) was added aniline (35mg, 0.38mmol) and Pd (PPh) at room temperature₃)₄(88mg, 0.08 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-SELECT-C18(150X19) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/10, 8/80, 11/89, 11.1/98, 13.98, 15/98, 18mL/min]This gave 220.7c (35mg, 15%) of solid.

¹H NMR(500MHz，DMSO-d₆):12.80(s，1H)，11.18(s，1H)，7.79(s，2H)，7.74(s，1H)，7.48(s，1H)，6.68(s，1H)，4.14(d，J＝14.0Hz，1H)，3.94(m，J＝10.0Hz，1H)，3.79(d，J＝10.5Hz，1H)，3.56(d，J＝9.5Hz，1H)，3.30-3.20(m，2H)，3.09-3.01(m，2H)，2.50-2.46(m，1H)，2.42-2.36(m，1H)，2.10-2.07(m，1H)，0.77(s，9H)。LCMS:97.7％，m/z[M+H]⁺614.0; chiral purity (51.5+ 48.4)%.

Example 225: synthesis of 5, 7-dichloro-N1 '- (3, 5-dichlorophenyl) -6', 6 '-difluoro-N2' -hydroxy-N2 '-methyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino-cyclo ] -1', 2' -dicarboxamide (225a and 225b)

To a stirred solution of 225.1(350mg, 0.64mmol) in DMF (15mL) at 0 deg.C in a sealed tube was added TEA (1.33mL, 9.58mmol) and N-methylhydroxylamine hydrochloride (534mg, 6.39 mmol). After allowing to warm slowly to room temperature and stirring for 16h, the reaction mixture was diluted with ethyl acetate (20ml) and water (2)x20 mL). The organic layer was dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by column chromatography followed by SFC purification [ column: chiral package IC (4.6x250) mm, 5 μ; 75% CO₂25% methanol, room temperature (isocratic 20mL/min, detection at 214 nm)]This gave 225a as an off-white solid (20mg, 5%) and 225b as an off-white solid (45mg, 12%). The absolute stereochemistry of 225a and 225b was not established.

225a:¹H NMR(500MHz，DMSO-d₆):11.20(br s，1H)，10.06(br s，1H)，10.03(br s，1H)，7.55(d，J＝2.0Hz，1H)，7.41(d，J＝1.5Hz，2H)，7.29(t，J＝2.0Hz，1H)，7.10(br s，1H)，4.58-4.52(m，1H)，4.47-4.42(m，1H)，3.83-3.74(m，1H)，3.34-3.32(m，1H)，3.08(s，3H)，3.01-2.90(m，1H)，2.83-2.78(m，1H)，2.37-2.31(m，1H)；LCMS:94.8％，m/z[M+H]⁺＝593.1。

225b:¹H NMR(500MHz，DMSO-d₆):10.85(br s，1H)，10.32(s，1H)，9.86(br s，1H)，7.68(d，J＝2.0Hz，3H)，7.49(d，J＝2.0Hz，1H)，7.30(t，J＝2.0Hz，1H)，4.25(d，J＝7.5Hz，1H)，4.13-4.09(m，1H)，3.49-3.38(m，2H)，2.69(s，3H)，2.60-2.50(m，1H)，2.47-2.39(m，1H)，2.26-2.13(m，1H)；LCMS:95.8％，m/z[M+H]⁺＝593.1。

Example 226: synthesis of 5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N2 ' -hydroxy-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide (226a and 226b)

To a solution of 225.1(200mg, 0.37mmol) in DMF (3mL) at room temperature was added NH₂OH.HCl (127mg, 1.83mmol) and TEA (0.25mL, 1.83 mmol). After stirring at room temperature for 4 hours, the reaction mixture was poured into ice water (15mL) and stirring was continued for 10 minutes. The resulting precipitate was filtered, washed with cold water (10mL) and dried under high vacuum. Preparation form for the obtained residueHigh performance liquid chromatography purification [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/45, 8/70, 9.4/70, 9.5/98, 11/98, 11.1/45, 13/45, 25mL/min]This gave 226a as a white solid (30mg, 14%) and 226b as a white solid (17mg, 8%). The absolute stereochemistry of 226a and 226b was not established.

226a:¹H NMR(500MHz，DMSO-d₆):11.14(br s，1H)，10.53(br s，1H)，10.06(s，1H)，8.88(s，1H)，7.53(d，J＝2.0Hz，1H)，7.44-7.35(m，2H)，7.30-7.29(m，2H)，4.29-4.24(m，1H)，4.00-3.97(m，1H)，3.65-3.60(m，1H)，3.50(d，J＝7.5Hz，1H)，3.00-2.81(m，1H)，2.79-2.76(m，1H)，2.34-2.27(m，1H)；LCMS:97.5％，m/z[M-H]^-＝576.9。

226b:¹H NMR(500MHz，DMSO-d₆):11.08(br s，1H)，10.40(br s，1H)，10.17(br s，1H)，8.92(s，1H)，7.70-7.65(m，2H)，7.57(d，J＝2.0Hz，1H)，7.42(d，J＝1.5Hz，1H)，7.29(t，J＝2.0Hz，1H)，4.40-4.35(m，1H)，4.24-4.21(m，1H)，3.79-3.64(m，1H)，3.26(d，J＝8.0Hz，1H)，3.10-2.95(m，1H)，2.75-2.70(m，1H)，2.37-2.28(m，1H)；LCMS:95.6％，m/z[M-H]^-＝576.9。

Example 227: synthesis of (1' R, 2' S, 3R, 7a ' R) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N2 ' -methoxy-N2 ' -methyl-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino-yl ] -1', 2' -dicarboxamide

To a stirred solution of 227.1(300mg, 0.53mmol) in THF (10mL) at 0 deg.C was added N-methylmorpholine (87. mu.L, 0.79mmol) and isobutyl chloroformate (62. mu.L, 0.64 mmol). After stirring for 5 min, N, O-dimethylhydroxylamine hydrochloride (103mg, 1.06mmol) was added at 0 ℃. After stirring at room temperature for 16 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: SYMMETRY-C8(300x19) mm, 7 u; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B) 0/50, 8/80, 8.1/98, 10/98, 10.1/50, 13/50, 20mL/min followed by purification by conventional preparative HPLC [ column: chiral OX-H (250X30) mm, 5u, mobile phase acetonitrile, room temperature (isocratic 42.0mL/min, detection at 215 nm) ] to yield 227(59mg, 18%) as a white solid.

¹H NMR(400MHz，DMSO-d₆):10.89(s，1H)，10.38(s，1H)，7.70-7.68(m，3H)，7.51(d，J＝2.0Hz，1H)，7.31(t，J＝2.0Hz，1H)，4.26(d，J＝7.2Hz，1H)，4.13(dd，J＝7.2Hz，J＝6.8Hz，1H)，3.54-3.40(m，2H)，3.44(s，3H)，2.67(s，3H)，2.61-2.54(m，1H)，2.49-2.46(m，1H)，2.25-2.16(m，1H)；LCMS:98.3％，m/z[M+H]⁺＝606.9。

Example 228: synthesis of (1' R, 2' S, 3R, 7a ' R) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

To a solution of 227.1(200mg, 0.35mmol) in THF (20mL) at 0 deg.C was added N-methylmorpholine (58. mu.L, 0.53mmol) and isobutyl chloroformate (41. mu.L, 0.42 mmol). After stirring for five minutes, NH was passed into the reaction mixture at 0 deg.C₃Qi for ten minutes. After stirring at room temperature for 16 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: x-bridge C18(250X19) mm, 5; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/50, 8/80, 8.1/98, 10/98, 10.1/50, 13/50, 20mL/min]Thus, 228(96mg, 47%) was produced as a white solid.

¹H NMR(500MHz，DMSO-d₆):10.89(s，1H)，10.74(s，1H)，7.71-7.70(m，3H)，7.53(d，J＝2.0Hz，1H)，7.33(t，J＝2.0Hz，1H)，7.00(br s，1H)，6.66(br s，1H)，4.55-4.50(m，1H)，4.25(t，J＝10.5Hz，1H)，3.99(d，J＝11.0Hz，1H)，3.49-3.41(m，1H)，2.84(t，J＝11.5Hz，1H)，2.41-2.35(m，1H)，2.16-2.06(m，1H)；LCMS:96.9％，m/z[M+H]⁺＝562.9。

Example 229: synthesis of rel- (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N2 '-hydroxy-N1', N2 '-dimethyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyridoxine ] -1', 2' -dicarboxamide and 5, 7-dichloro-N1 '- (3, 5-dichlorophenyl) -6', 6 '-difluoro-N2' -hydroxy-N1 ', N2' -dimethyl-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3 ], 3' -pyrazino ] -1', 2' -dicarboxamide (229a and 229b):

229a and 229b are synthesized from 250.2 as described in the synthesis of 260a and 260 b. The absolute stereochemistry of 229a and 229b is not established.

229a:¹H NMR(500MHz，DMSO-d₆) 10.88/10.79(s, 1H), 10.07/9.95(s, 1H), 7.72/7.54(s, 1H), 7.51-7.30(m, 4H), 4.55/4.42(d, J ═ 8.0Hz, 1H), 4.17-4.05(m, 1H), 3.51-3.36(m, 2H), 3.36/3.22(s, 3H), 2.99/2.97(s, 3H), 2.74-2.50(m, 2H), 2.23-2.10(m, 1H); LCMS 96.0%, M/z [ M + H ]]⁺＝607.0。

229b:¹H NMR(500MHz，DMSO-d₆) 10.75/10.69(s, 1H), 9.90/9.82(s, 1H), 7.95/7.82(s, 1H), 7.77/7.63(s, 1H), 7.60-7.45(m, 3H), 4.22/4.07(d, J ═ 7.5Hz, 1H), 3.85-3.77(m, 1H), 3.56-3.35(m, 2H), 3.24(s, 3H), 2.72(s, 3H), 2.50-2.32(m, 2H), 2.22-2.10(m, 1H); LCMS 95.1%, M/z [ M + H ]]⁺＝607.0。

Example 230: synthesis of rel- (1' R, 2' S, 3R, 7a ' R) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N1 ' -methyl-2-oxo-N2 ' - (phenylsulfonyl) -1', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino-cyclo ] -1', 2' -dicarboxamide

To a stirred solution of 250.2(200mg, 0.34mmol) in DMF (5mL) at room temperature were added DIPEA (0.12mL, 0.69mmol) and HATU (196mg, 0.51 mmol). After stirring for 15 min, benzenesulfonamide (81mg, 0.51mmol) was added. After stirring at room temperature for 12h, the reaction mixture was quenched with ice water (5ml) and extracted with ethyl acetate (2 × 10mL). The combined organic phases were washed with brine (10ml), dried over anhydrous sodium sulfate, filtered and concentrated. The resulting residue was purified by preparative HPLC [ column: X-bridge-C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/40, 8/80, 11/90, 11.1/98, 12/98, 12.1/40, 15/40at 23mL/min to yield 230 as an off-white solid (24mg, 10%).

¹H NMR(500MHz，DMSO-d₆) 12.00(br s, 1H), 10.86/10.66(s, 1H), 7.74-7.39(m, 10H), 4.60/4.21(m, 1H), 4.00-3.92(m, 1H), 3.67-3.63(m, 1H), 3.27-3.23(m, 1H), 3.20(s, 3H), 2.96-2.87(m, 1H), 2.64-2.50(m, 1H), 2.14-2.02(m, 1H); LCMS 93.0%, M/z [ M-H ]]^-＝715.0。

Example 231: synthesis of rel- (1' R, 2' S, 3R, 7a ' R) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N1 ' -methyl-N2 ' - (methylsulfonyl) -2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide.

To a stirred solution of 252.2(1.0g, 1.72mmol) in DMF (15mL) at room temperature were added DIPEA (0.47mL, 2.59mmol) and HATU (0.98g, 2.59 mmol). After stirring for 30min, methylsulfonamide (0.27g, 2.59mmol) was added. Stir at room temperature for 16h, quench the reaction mixture with water (20mL) and extract the combined organic phases with ethyl acetate (2 × 20mL), wash with brine (20mL), dry over anhydrous sodium sulfate, filter and concentrate under reduced pressure. The residue obtained was purified by flash column chromatography (40g silica gel column, 30% ethyl acetate in petroleum ether) followed by preparative HPLC [ column: X-bridge-C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/50, 8/80, 9/80, 9.1/98, 10/98, 10.1/50, 12/50, 23mL/min to yield 231(71mg, 6%) as an off-white solid.

¹H NMR(500MHz，DMSO-d₆) 11.32(br s, 1H), 11.10/11.93(s, 1H), 7.75-7.48(m, 5H), 4.58/4.25(m, 1H), 4.00(d, J ═ 10.5Hz, 1H), 3.75-3.71(m, 1H), 3.39-3.33(m, 1H), 3.22(s, 3H), 3.01/2.98(s, 3H), 2.92-2.86(m, 1H), 2.50-2.43(m, 1H), 2.15-2.07(m, 1H); LCMS 99.0, M/z [ M + H ]]⁺655.0; the chiral purity is 96.4 percent.

Example 232: synthesis of (1' S, 2' R, 3S, 7a ' S) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N1 ' -methyl-2-oxo-N2 ' - (2, 2, 2-trifluoroethyl) -1', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino-cyclo ] -1', 2' -dicarboxamide (232.1a) and (1' R, 2' S, 3R, 7a ' R) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N1 ' -methyl-2-oxo-N2 ' - (2), 2, 2-trifluoroethyl) -1', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino-cyclo ] -1', 2' -dicarboxamide (232.1b)

To a stirred solution of 250.2(200mg, 0.34mmol) in DMF (5mL) at room temperature was added HATU (196mg, 0.52mmol) and Et₃N (0.14mL, 1.04 mmol). After stirring for 15 min 2,2, 2-trifluoroethylamine hydrochloride (92mg, 0.69mmol) was added. After stirring at room temperature for 3h, the reaction mixture was diluted with ice water (50ml) and extracted with ethyl acetate (2 × 60 mL). The combined organic phases were washed with brine (50mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The obtained residue is used for preparing high-efficiencyLiquid chromatography purification [ KROMOSIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/30, 8/80, 11/85, 11.1/98, 13/98, 13.1/30, 15/30, 22mL/min]This gave 232 as an off-white solid (110mg, 50%).

¹H NMR(500MHz，DMSO-d₆) 10.95/10.74(s, 1H), 7.82-7.45(m, 6H), 4.26(t, J ═ 10.5Hz, 1H), 4.00-3.97(m, 1H), 3.88-3.84(m, 1H), 3.77-3.72(m, 1H), 3.65-3.60(m, 1H), 3.46-3.39(m, 1H), 3.39/3.21(s, 3H), 2.89-2.85(m, 1H), 2.60-2.50(m, 1H), 2.25-2.10(m, 1H); LCMS 96.4%, M/z [ M-H ]]^-657.0; chiral purity (49.9% + 50.0%).

Separation 232a and 232 b:

232(100mg) was separated by palm SFC [ column (R, R) Whelk-01(30X250mm), 5. mu.l; 90% CO₂10% acetonitrile, at room temperature (isocratic 70g/min, detection at 214 nm)]This gave 232a (enantiomer-1, 17mg, 34%) as an off-white solid and 232b (enantiomer-2, 20mg, 40%) as an off-white solid. The absolute stereoisomer was not determined.

232a:¹H NMR(500MHz，DMSO-d₆) 10.95/10.74(s, 1H), 7.82-7.44(m, 6H), 4.26(t, J ═ 10Hz, 1H), 3.99-3.97(m, 1H), 3.90-3.84(m, 1H), 3.77-3.72(m, 1H), 3.67-3.62(m, 1H), 3.46-3.39(m, 1H), 3.39/3.21(s, 3H), 2.89-2.85(m, 1H), 2.60-2.50(m, 1H), 2.25-2.10(m, 1H); LCMS 99.1%, M/z [ M-H ]]^-657.0; the chiral purity is 98.65 percent.

232b:¹H NMR(500MHz，DMSO-d₆) 10.95/10.74(s, 1H), 7.76-7.44(m, 6H), 4.26(t, J ═ 10Hz, 1H), 3.99-3.97(m, 1H), 3.90-3.84(m, 1H), 3.77-3.72(m, 1H), 3.65-3.62(m, 1H), 3.48-3.37(m, 1H), 3.41/3.21(s, 3H), 2.89-2.85(m, 1H), 2.60-2.50(m, 1H), 2.25-2.10(m, 1H); LCMS 99.1%, M/z [ M-H ]]^-657.0; the chiral purity is 99.7 percent.

Example 233: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -N2 '-methoxy-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

To a stirred solution of 233.1(WO2017117239) (200mg, 0.39mmol) in DMF (15mL) was added methoxyamine hydrochloride (326mg, 3.9mmol) and TEA (1.6mL, 11.7mmol) at room temperature. Stirred at room temperature for 16h, the reaction mixture was quenched with ice water and extracted with ethyl acetate (2 × 30 mL). The combined organic layers were dried over sodium sulfate. Filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-bridge-C8 (150X19) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/40, 8/80, 9/80, 9.1/98, 11/98, 11.1/40, 14/40, 25mL/min ] to give 233(7mg, 3%) as an off-white solid.

¹H NMR(400MHz，DMSO-d₆):10.96(br s，1H)，10.79(br s，1H)，10.12(br s，1H)，7.67(s，2H)，7.57-7.49(m，2H)，7.26(t，J＝2.0Hz，1H)，4.22-4.10(m，1H)，4.10-4.01(m，1H)3.40(s，3H)，3.25-3.12(m，2H)，2.07-1.98(m，1H)，1.97-1.65(m，4H)；LCMS:94.6％，m/z[M+H]⁺557.0. Regional chemistry is unknown.

Example 234: synthesis of 5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -N2' -hydroxy-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide (234a and 234b)

To a stirred solution of 233.1(200mg, 0.39mmol) in DMF (5mL) at room temperature was added NH₂OH (137mg, 1.96mmol) and NEt₃(0.27mL, 1.96 mmol). After stirring at room temperature for 16 hours, the reaction mixture was poured into ice water (15mL) and stirred well. The resulting precipitate was filtered, washed with water (10mL) and washed with waterDrying under high vacuum. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/10, 9/80, 9.1/98, 11/98, 11.1/10, 13/10, 25mL/min]234a (40mg, 19%) and 234b (150mg, 71%) were thus obtained as off-white solids.

234a:¹H NMR(500MHz，DMSO-d₆):10.99(br s，1H)，10.44(br s，1H)，9.88(s，1H)，8.81(s，1H)，7.47(d，J＝2.0Hz，2H)，7.38(d，J＝2.0Hz，2H)，7.25(t，J＝2.0Hz，1H)，4.07-4.02(m，1H)，3.91-3.88(m，1H)，3.55-3.51(m，1H)，3.09-3.05(m，1H)，2.37-2.30(m，1H)，2.09-2.05(m，1H)，1.92-1.85(m，1H)，1.84-1.77(m，1H)，1.70-1.62(m，1H)；LCMS:84.6％，m/z[M+H]⁺542.9. Regional chemistry is unknown.

234b:¹H NMR(500MHz，DMSO-d₆):10.93(br s，1H)，10.22(br s，1H)，10.13(s，1H)，8.71(s，1H)，7.68(d，J＝1.5Hz，2H)，7.60(s，1H)，7.49(d，J＝1.5Hz，1H)，7.27(t，J＝2.0Hz，1H)，4.16-4.12(m，1H)，4.02-3.99(m，1H)，3.16-3.11(m，1H)，2.27-2.23(m，1H)，2.02-1.99(m，1H)，1.92-1.85(m，1H)，1.81-1.65(m，3H)；LCMS:85.1％，m/z[M+H]⁺542.9. Regional chemistry is unknown.

Example 235: synthesis of 5, 7-dichloro-N1 '- (3, 5-dichlorophenyl) -N2' -hydroxy-N2 '-methyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

To a stirred solution of 233.1(200mg, 0.39mmol) in DMF (10mL) at room temperature was added NEt₃(0.27mL, 1.96mmol) and N-methylhydroxylamine hydrochloride (164mg, 1.96 mmol). After stirring at room temperature for 16 hours, the reaction mixture was poured into ice water (15mL) and stirred for 15 minutes. The resulting precipitate was filtered, washed with water (10mL) and dried under high vacuum.The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/30, 8/80, 10/90, 10.1/98, 12/98, 12.1/30, 14/30, 25mL/min]Thus, 235a (15mg, 7%) and 235b (30mg, 14%) were obtained as white solids.

235a:¹H NMR(400MHz，DMSO-d₆):11.01(br s，1H)，9.85(br s，2H)，7.48(d，J＝2.0Hz，1H)，7.42(d，J＝2.0Hz，2H)，7.25-7.24(m，2H)，4.54-4.50(m，1H)，4.27-4.18(m，1H)，3.35-3.25(m，2H)，3.08(s，3H)，2.37-2.33(m，1H)，2.12-2.05(m，1H)，1.92-1.83(m，2H)，1.69-1.60(m，1H)；LCMS:86.30％，m/z[M+H]⁺557.0. Regional chemistry is unknown.

235b:¹H NMR(400MHz，DMSO-d₆):10.67(s，1H)，10.20(s，1H)，9.72(s，1H)，7.78(d，J＝1.6Hz，1H)，7.69(d，J＝2.0Hz，2H)，7.41(d，J＝2.0Hz，1H)，7.26(t，J＝2.0Hz，1H)，4.26(d，J＝7.6Hz，1H)，3.96-3.94(m，1H)，3.35-3.30(m，1H)，2.82-2.80(m，1H)，2.69(s，3H)，2.17-2.09(m，1H)，1.88-1.77(m，3H)，1.58-1.48(m，1H)；LCMS:99.7％，m/z[M+H]⁺557.0. Regional chemistry is unknown.

Example 236: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-N- (3, 5-dichlorophenyl) -2' - (hydrazinecarboxy) -2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino-cyclo ] -1' -carboxamide

To a stirred solution of 233.1(500mg, 0.97mmol) in THF (5mL) was added hydrazine monohydrate (98mg, 1.85mmol) at room temperature. Stirred at room temperature for 4 hours, the reaction mixture was quenched with water and extracted with ethyl acetate (2 × 10mL). The combined organic phases were washed with brine (10ml), dried over sodium sulfate, filtered and concentrated under reduced pressure. The residue is reacted with CH₂Cl₂(5mL) were all ground together to give a white solid236(320mg，60％)。

¹H NMR(400MHz，DMSO-d₆):10.92(s，1H)，10.13(s，1H)，8.81(s，1H)，7.68(d，J＝1.6Hz，2H)，7.52(s，1H)，7.48(s，1H)，7.26(s，1H)，4.19-4.13(m，1H)，4.08-4.06(m，3H)，3.40(d，J＝7.6Hz，1H)，3.16-3.10(m，1H)，2.30-2.26(m，1H)，1.99-1.88(m，2H)，1.78-1.68(m，2H)；LCMS:92.3％，m/z[M+H]⁺542.3; the chiral purity is 98.7 percent. Regional chemistry is unknown.

Example 237: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -N2'- (1-methyl-1H-imidazol-4-yl) -2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

Synthesis 237.3:

to a stirred solution of 237.2(1g, 8.84mmol) in DMF (20mL) at 0 ℃ was added NaH (0.7g, 17.7mmol) in portions over 10min, and MeI (1.5g, 10.6mmol) was added at 0 ℃ with stirring for one hour. After stirring at room temperature for 16h, the reaction mixture was quenched with ice water and extracted with ethyl acetate (3 × 200 mL). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. This gave 237.3(300mg, 27%) as a gum, which was used directly in the next step without purification. LCMS 95.7%, M/z [ M + H ]]⁺＝128.1。

Synthesis 237.4:

to a stirred solution of 237.3(1g, 7.86mmol) in EtOH (20mL) was added 10% Pd/C (0.2g) at room temperature. After hydrogenation for 16h at room temperature using balloon pressure, the reaction mixture was filtered through a pad of celite. The filtrate was concentrated under reduced pressure to give 237.4(800mg) as an off-white solid. LCMS 40.9%, M/z [ M + H ]]⁺＝98.1。

Synthesis 237:

to a stirred solution of 237.1(0.3g, 0.57mmol) in THF (10mL) at 0 deg.C was added N-methylmorpholine (86mg, 0.85mmol) and isobutyl chloroformate (93mg, 0.68 mmol). After stirring at 0 ℃ for 30min 237.4(69mg, 0.68mmol) was added. After stirring at 60 ℃ for 16h, the reaction mixture was diluted with water and extracted with ethyl acetate (3 × 60 mL). The combined organic phases were washed with brine and dried over anhydrous sodium sulfate and concentrated. The resulting residue was purified by column chromatography (silica gel 100-200mesh, 3% MeOH in DCM) to afford 237(18mg, 5%) as a brown solid.

¹H NMR(400MHz，DMSO-d₆):10.94(s，1H)，10.07(s，1H)，9.94(s，1H)，7.64(br s，2H)，7.45(s，1H)，7.30-7.20(m，4H)，4.34-4.22(m，2H)，3.62-3.57(m，1H)，3.61(s，3H)，3.26-3.20(m，1H)，2.35-2.33(m，1H)，2.18-2.14(m，1H)，1.90-1.80(m，2H)，1.72-1.62(m，1H)；LCMS:95.2％，m/z[M+H]⁺607.0. The absolute stereochemistry is unknown.

Example 238: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -N2'- (1-methyl-1H-imidazol-2-yl) -2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

Synthesis of 238.2:

to a stirred solution of 238.1(0.2g, 1.76mmol) in DMF (10mL) at 0 deg.C was added 60% NaH (0.14g, 3.54mmol) in portions. After stirring for one hour, MeI (0.25g, 1.77mmol) was added at 0 ℃. After stirring at room temperature for 16h, the reaction mixture was quenched with ice water and extracted with ethyl acetate (3 × 50 mL). The combined organic phases were washed with brine and dried over anhydrous sodium sulfate and concentrated to give compound 238.2(0.14g) as a gum.

¹H NMR(500MHz，CDCl₃):7.14(d，J＝1.0Hz，1H)，7.06(s，1H)，4.08(s，3H)；LCMS:57.04％，m/z[M+H]⁺＝127.9。

Synthesis of 238.3:

to 238.2(0.1g, 0.79mmol)) To the stirred solution in 1, 4-dioxane (5mL) was added 10% Pd/C (50% wet, 20 mg). After hydrogenation for 16h at room temperature using balloon pressure, the reaction mixture was filtered through a pad of celite. The filtrate was concentrated under reduced pressure to give 238.3(70mg) as an off-white solid. LCMS 98.2%, M/z [ M + H ]]⁺＝98.2。

Synthesis 238:

to a stirred solution of 237.1(0.25g, 0.47mmol) in THF (10mL) at 0 deg.C was added N-methylmorpholine (72mg, 0.71mmol) and isobutyl chloroformate (77mg, 0.57 mmol). After stirring for 30min at 0 ℃ 238.3(69mg, 0.71mmol) was added after stirring for 24 h at 60 ℃ the reaction mixture was diluted with water and extracted with ethyl acetate (3 × 50 mL). The combined organic phases were washed with brine and dried over anhydrous sodium sulfate and concentrated. The resulting residue was purified by column chromatography (silica gel 100-200 sieve, 3% MeOH in DCM) followed by preparative HPLC [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/30, 7/70, 7.1/98, 9/98, 9.1/30, 11/30, 25mL/min ] gave 238 as a pale pink solid (11mg, 4%).

¹H NMR(400MHz，DMSO-d₆):11.82(br s，1H)，10.61(s，1H)，10.54(s，1H)，7.76(s，2H)，7.45(s，2H)，7.25(s，1H)，6.76(s，1H)，6.60(s，1H)，4.21-4.11(m，1H)，4.06(d，J＝3.2Hz，2H)，2.96(s，3H)，2.91-2.83(m，1H)，2.50-2.40(m，1H)，1.81-1.72(m，2H)，1.61-1.48(m，2H)；LCMS:96.2％，m/z[M-H]^-605.2. The absolute stereochemistry is unknown.

Example 239: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -N2'- (1-methyl-1H-pyrazolyl-5-yl) -2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

N-methylmorpholine (151mg, 1.50mmol) was added to a solution of 237.1(400mg, 0.75mmol) in THF (40mL) at-10 deg.C followed by isobutyl chloroformate (204mg, 1.50 mmol). After stirring for 20 min at-10 ℃ 1-methyl-1H-pyrazolyl-5-amine (220mg, 2.26mmol) was added and stirred at the same temperature for 1H. The reaction mixture was concentrated under reduced pressure to obtain a residue, which was purified by reverse phase high performance liquid chromatography [ column: buchi Reveleries C18(40 g); b is 0.05 percent formic acid solution in water, B is acetonitrile. Freeze drying the pure fractions yielded 239(40mg, 8%) as an off-white solid.

¹H NMR(400MHz，DMSO-d₆):11.07(s，1H)，10.16(s，1H)，9.63(s，1H)，7.66(d，J＝1.6Hz，2H)，7.56(d，J＝2.0Hz，1H)，7.35(d，J＝2.0Hz，1H)，7.31(d，J＝1.6Hz，1H)，7.27(t，J＝2.0Hz，1H)，5.90(d，J＝1.6Hz，1H)，4.31-4.26(m，2H)，3.63(d，J＝7.2Hz，1H)，3.51(s，3H)，3.20-3.18(m，1H)，2.36-2.33(m，1H)，2.12-2.02(m，1H)，1.92-1.79(m，2H)，1.71-1.63(m，1H)；LCMS:96.9％，m/z[M+H]⁺607.0. The absolute stereochemistry is unknown.

Example 240: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -N2'- (1-methyl-1H-pyrazolyl-3-yl) -2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

To a stirred solution of 237.1(100mg, 0.18mmol) in THF (10mL) at 0 deg.C was added N-methylmorpholine (38mg, 0.37mmol) and isobutyl chloroformate (51mg, 0.37 mmol). After stirring for 15 min, 1-methyl-1H-pyrazolyl-3-amine (36mg, 0.37mmol) was added. After stirring for one hour at 0 ℃, the reaction mixture was concentrated under reduced pressure. The residue was purified by reverse phase chromatography [ column: buchi Reveleries C18(40 g); b: 0.05% formic acid in water, B: acetonitrile ] to yield 240 as an off-white solid (20mg, 17%).

¹H NMR(400MHz，DMSO-d₆):10.97(s，1H)，10.10(s，1H)，10.05(s，1H)，7.65(d，J＝2.0Hz，2H)，7.50(d，J＝2.0Hz，1H)，7.47(d，J＝2.0Hz，1H)，7.33(d，J＝2.0Hz，1H)，7.25(t，J＝2.0Hz，1H)，6.38(d，J＝2.0Hz，1H)，4.35-4.25(m，2H)，3.70(s，3H)，3.57(d，J＝8.0Hz，1H)，3.26-3.20(m，1H)，2.36-2.32(m，1H)，2.15-2.09(m，1H)，1.92-1.88(m，1H)，1.83-1.80(m，1H)，1.68-1.66(m，1H)；LCMS:93.1％，m/z[M+H]⁺607.0. The absolute stereochemistry is unknown.

Example 241: synthesis of (1' R, 2' S, 7a ' R) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -N2' -methoxy-N2 ' -methyl-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

To a stirred solution of 237.1(300mg, 0.56mmol) in DMF (10mL) at 0 deg.C was added N-methylmorpholine (0.09mL, 0.85mmol) and isobutyl chloroformate (0.1mL, 0.73 mmol). After 15 min N, O-dimethylhydroxylamine hydrochloride (276mg, 2.83mmol) was added at 0 ℃. After stirring at room temperature for 16 hours, the reaction mixture was poured into ice-water (20mL) and stirred for 20 minutes. The resulting precipitate was filtered. The filtrate was collected, washed with water (20mL) and dried under high vacuum. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/50, 8/80, 10/80, 10.1/98, 11/98, 11.1/50, 13/50, 25mL/min ] gave 241 as a white solid (17mg, 5% yield).

¹H NMR(400MHz，DMSO-d₆):10.72(br s，1H)，10.24(br s，1H)，7.81(d，J＝2.0Hz，1H)，7.70(d，J＝1.6Hz，2H)，7.44(d，J＝2.0Hz，1H)，7.27(t，J＝2.0Hz，1H)，4.25(d，J＝7.2Hz，1H)，4.01-3.95(m，1H)，3.43(s，3H)，3.37-3.35(m，1H)，2.90-2.82(m，1H)，2.66(s，3H)，2.17-2.10(m，1H)，1.90-1.72(m，3H)，1.61-1.53(m，1H)；LCMS:95.2％，m/z[M+H]⁺571.0. The absolute stereochemistry isIs unknown.

Example 242: synthesis of (1'R, 2' S, 7a 'R) -2' - (2-acetylhydrazine-1-carboxy) -5, 7-dichloro-N- (3, 5-dichlorophenyl) -2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -1' -carboxamide

To a stirred solution of 237.1(100mg, 0.18mmol) in THF (3mL) was added triethylamine (0.1mL, 0.75mmol) and acetic acid hydrazide (28mg, 0.37mmol) at room temperature, and after ten minutes, a solution of propylphosphoric anhydride (50% wt in ethyl acetate, 0.24mL, 0.75mmol) was added at room temperature. After stirring at room temperature for 16 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/40, 8/90, 8.1/98, 10/98, 10.1/40, 12/40at 25mL/min ] to yield 242 as a white solid (27mg, 24%).

¹H NMR(400MHz，DMSO-d₆):10.89(s，1H)，10.22(s，1H)，9.83(s，1H)，9.71(s，1H)，7.71-7.68(m，2H)，7.59(d，J＝2.0Hz，1H)，7.42(d，J＝2.0Hz，1H)，7.27(t，J＝2.0Hz，1H)，4.18-4.12(m，1H)，4.08-4.04(m，1H)，3.68(d，J＝8.0Hz，1H)，2.96-2.90(m，1H)，2.28-2.24(m，1H)，1.88-1.68(m，4H)，1.77(s，3H)；LCMS:95.6％，m/z[M+H]⁺584.0. The absolute stereochemistry is unknown.

Example 243: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -N2 '-morpholinyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

A solution of 237.1(200mg, 0.37mmol) in DMF (10mL) was stirred at 0 deg.CN-methylmorpholine (0.05mL, 0.49mmol) and isobutyl chloroformate (0.07mL, 0.56mmol) were added. After 10min, morpholinyl-4-amine hydrochloride (0.07mL, 0.56mmol) was added at 0 ℃. After stirring at room temperature for 2 hours, the reaction mixture was poured into ice water (15mL) and stirred for 15 minutes. The resulting precipitate was filtered, washed with water (10mL) and dried under high vacuum. Compound material was purified by preparative HPLC [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a10 mM NH₄OAc at H₂Solution in O, B is acetonitrile; gradient: (T% B):0/50, 8/80, 9/80, 9.1/50, 11/50, 25mL/min]Yielding 243(35mg, 15%) as a white solid.

¹H NMR(400MHz，DMSO-d₆):10.81(br s，1H)，10.15(br s，1H)，8.18(s，1H)，7.90(d，J＝1.6Hz，1H)，7.62(d，J＝1.2Hz，2H)，7.37(d，J＝1.6Hz，1H)，7.18(s，1H)，4.12(d，J＝7.2Hz，1H)，3.79-3.74(m，1H)，3.57-3.47(m，2H)，3.39-3.25(m，3H)，2.71-2.60(m，2H)，2.26-2.10(m，2H)，2.02-2.00(m，1H)，1.75-1.69(m，3H)，1.41-1.37(m，1H)，1.02-1.00(m，1H)；LCMS:99.6％，m/z[M+H]⁺612.0. The absolute stereochemistry is unknown.

Example 244: synthesis of (1'R, 2' S, 6'S, 7a' R) -5, 7-dichloro-N1 '- (3, 5-dichlorophenyl) -6' -hydroxy-6 '-methyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

Methanolic ammonia (10mL) was added to 244.1(WO2017117239) (250mg, 0.46mmol) at room temperature and after stirring for 16h, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: YMC TRIART-C18(150X25) mm, 10 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/20, 8/60, 10/60, 10.1/20, 12/20, 25mL/min ] gave 244 as a white solid (32mg, 12%).

¹H NMR(500MHz，DMSO-d₆):10.74(s，1H)，10.61(s，1H)，7.71(d，J＝1.5Hz，2H)，7.65(d，J＝1.5Hz，1H)，7.48(d，J＝2.0Hz，1H)，7.30(t，J＝2.0Hz，1H)，6.91(s，1H)，6.57(s，1H)，4.50-4.47(m，1H)，4.43(s，1H)，4.32-4.28(m，1H)，3.81(d，J＝11.5Hz，1H)，2.92(d，J＝8.5Hz，1H)，2.30(d，J＝8.0Hz，1H)，1.69-1.65(m，1H)，1.50-1.45(m，1H)，1.21(s，3H)；LCMS:99.5％，m/z[M+H]⁺557.0; the chiral purity is 99.9 percent. Regional chemistry is unknown.

Example 245: synthesis of (1' R, 2' S, 6' S, 7a ' R) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -N2', 6' -dihydroxy-6 ' -methyl-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

To a stirred solution of 244.1(500mg, 0.92mmol) in DMF (15mL) at room temperature was added NH₂OH.HCl (320mg, 4.61mmol)) and Et₃N (0.64mL, 4.61mmol), and the resulting reaction mixture was stirred for 16 hours. The reaction mixture was diluted with ice water (20ml) and the resulting precipitate was filtered to give a residue, which was purified by preparative HPLC [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/20, 8/60, 8/98, 10/98, 10.1/20, 13/20, 25mL/min]Resulting in 245(30mg, 5%) as an off-white solid.

¹H NMR(400MHz，DMSO-d₆):10.91(br s，1H)，10.21(br s，1H)，10.08(br s，1H)，8.75(br s，1H)，7.67(d，J＝2.0Hz，2H)，7.50(s，1H)，7.47(s，1H)，7.25(t，J＝1.6Hz，1H)，4.51-4.42(m，1H)，4.00(s，1H)，4.22-4.18(m，1H)，3.23(d，J＝9.2Hz，1H)，3.17(d，J＝7.6Hz，1H)，2.20-2.14(m，2H)，1.69-1.65(m，1H)，1.20(s，3H)；LCMS:90.5％，m/z[M+H]⁺573.0. Regional chemistry is unknown.

Example 246: synthesis of (1' R, 2' S, 6' S, 7a ' R) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -6' -hydroxy-N2 ' -methoxy-6 ' -methyl-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

Et was added to a stirred solution of 244.1(300mg, 0.55mmol) in DMF (15mL) at room temperature₃N (1.1mL, 8.31mmol) and CH₃ONH₂HCl (462mg, 5.54 mmol). Stirred at room temperature for 48 h, the reaction mixture was diluted with water (50mL) and extracted with ethyl acetate (2 × 30 mL). The combined organic layers were dried over sodium sulfate, filtered and evaporated under nitrogen. The resulting residue was purified by preparative HPLC [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/40, 7/65, 7.1/98, 9/98, 9.1/40, 11/40, 25mL/min]246(25mg, 12%) was produced as an off-white solid.

¹H NMR(500MHz，DMSO-d₆):11.00(s，1H)，10.80(s，1H)，10.13(s，1H)，7.66(d，J＝1.5Hz，2H)，7.56(s，1H)，7.42(s，1H)，7.26(d，J＝2.0Hz，1H)，4.57-4.48(m，1H)，4.43(s，1H)，4.27-4.23(m，1H)，3.45(s，3H)，3.32-3.24(m，1H)，3.01(d，J＝8.0Hz，1H)，2.24-2.14(m，2H)，1.68-1.61(m，1H)；LCMS:96.4％，m/z[M+H]⁺587.0. Regional chemistry is unknown.

Example 247: synthesis of (6'S) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -6 '-hydroxy-6' -methyl-N2 '- (methylsulfonyl) -2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

Synthesis 247.2:

to a stirred solution of 247.1(1g, 4.13mmol) in DMF (5mL) at room temperature was added methanesulfonamide (393mg, 4.13mmol) and TEA (1.72mL, 12.4 mmol). In the roomAfter stirring at room temperature for 2 hours, the reaction mixture was concentrated under reduced pressure to give compound 247.2(1.39g) as a thick brown liquid which was used in the next step without any purification. LCMS 57.2%, M/z [ M-H ]]^-＝334.7。

Synthesis 247: \ u

To a solution of 247.2(1.2g, 3.55mmol) in THF (20mL) was added a TFA salt of (2S, 4S) -4-hydroxy-4-methyl-1- (2, 2, 2-trifluoroacetyl) -1l 4-pyrrolidine-2-carboxylic acid (860mg, 3.55mmol) and 5, 7-dichloroindole-2, 3-dione (767mg, 3.55mmol) at room temperature. After stirring at 80 ℃ for 2h, the reaction mixture was cooled to room temperature and diluted with EtOAc. The organic solvent was collected, washed with ice water, dried over sodium sulfate, filtered and concentrated using a stream of nitrogen. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/45, 7/55, 7.1/98, 10/98, 10.1/45, 12/45, 25mL/min ] gave 247(26mg) as an off-white solid. Regiochemistry was not determined.

¹H NMR(400MHz，DMSO-d₆):11.75(br s，1H)，11.02(br s，1H)，9.98(s，1H)，7.49(s，1H)，7.41(d，J＝2Hz，2H)，7.26(s，1H)，7.12(br s，1H)，4.56-4.42(m，2H)，4.28-4.19(m，1H)，3.39(d，J＝7.6Hz，1H)，3.22-3.10(m，4H)，2.25-2.10(m，2H)，1.80-1.75(m，1H)，1.24(s，3H)；LCMS:90.9％，m/z[M+H]⁺＝634.9。

Example 248: synthesis of (6'S) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -N2'- (N, N-dimethylsulfamoyl) -6' -hydroxy-6 '-methyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

Synthesis of 248.2:

to a stirred solution of 247.1(500mg, 2.06mmol) and 248.1(256mg, 2.06mmol) in DMF (3mL) was added Et at room temperature₃N (0.86mL, 6.19 mmol). After stirring at room temperature for 2 hours, the resulting reaction mixture was evaporated under reduced pressure to give 248.2, which was used directly in the next step. LCMS (13+ 39)%, M/z [ M-H%]^-＝364.0。

Synthesis 248:

to a stirred solution of 248.2(300mg, 0.81mmol) in THF (10mL) at room temperature was added (2S, 4S) -4-hydroxy-4-methyl-1- (2, 2, 2-trifluoroamido) -1l 4-pyrrolidine-2-carboxylic acid (197mg, 0.81mmol) as TFA salt and 5, 7-dichloroindole-2, 3-dione (176mg, 0.81 mmol). After stirring at 80 ℃ for 2h, the reaction mixture was cooled to room temperature and diluted with EtOAc (20mL). The organic solvent was collected, washed with ice water, dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMASIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/40, 8/70, 8.1/40, 10/40, 25mL/min]This gave 248(16mg, 3%) as an off-white solid. Regiochemistry was not determined. LCMS 92.6%, M/z [ M + H ]]⁺＝664.0。

Example 249: synthesis of (1'S, 2' R, 3S, 7a 'S) -5, 7-dichloro-1' - ((3-chloro-5-methoxyphenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyridoxine ] -2' -carboxylic acid (249a) & (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3-chloro-5-methoxyphenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3 ], 3 '-pyrazino ] -2' -carboxylic acid (249b):

249a and 249b are synthesized from 110.4_1 using the steps described in synthesizing 265a and 265 b.

249a:¹H NMR(500MHz，DMSO-d₆) 12.55(br s, 1H), 10.97(br s, 1H), 8.34(s, 1H), 7.47-7.44(m, 1H), 7.05-6.86(m, 3H), 4.15-3.95(m, 1H), 3.83-3.79(m, 1H), 3.79(s，3H)，3.60-3.50(m，1H)，3.32-3.25(m，1H)，3.23(s，3H)，2.60-2.49(m，1H)，2.45-2.30(m，1H)，2.14-2.07(m，1H)；LCMS:93.1％，m/z[M+H]⁺＝574.1。

249b:¹H NMR(500MHz，DMSO-d₆) 12.51(br s, 1H), 11.04/10.97(br s, 1H), 8.34(s, 1H), 7.47-7.44(m, 1H), 7.05-6.85(m, 3H), 4.15-3.95(m, 1H), 3.83-3.79(m, 1H), 3.79/3.68(s, 3H), 3.60-3.50(m, 1H), 3.37-3.24(m, 1H), 3.23(s, 3H), 2.60-2.50(m, 1H), 2.45-2.30(m, 1H), 2.14-2.07(m, 1H); LCMS 92.2%, M/z [ M + H ]]⁺＝574.1。

Example 250: synthesis of rac- (5-methyl-2-oxo-1, 3-dioxo-4-yl) methyl (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino-cyclo ] -2' -carboxylate (250.3a) & rac- (5-methyl-2-oxo-1, 3-dioxo-4-yl) methyl (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-1- ((5-methyl-2-oxo-1, 3-dioxo-4-yl) methyl) -2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylate (250.3b)

Synthesis of 250.1:

thionyl chloride (20mL) was added to 110.4_1(1.5g, 3.25mmol) at room temperature. After stirring for 2 hours, the filtered thionyl chloride was removed under reduced pressure to yield the acid chloride. At room temperature, adding the acid chloride in CH₂Cl₂(25mL) was added 3, 5-dichloro-N-methylaniline (1.24g, 7.08mmol) in CH₂Cl₂(5 mL). After stirring at room temperature for 16 hours, the reaction mixture was quenched with water (10ml). Separating the organic layer with CH₂Cl₂(2 × 20mL) extract the aqueous layer. The combined organic phases were washed with brine (20mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressureAnd (5) concentrating. The resulting residue was purified by flash column chromatography (40g silica gel column, 30% ethyl acetate in petroleum ether) to yield 250.1(1.4g, 73%) solid.

¹H NMR(400MHz，DMSO-d₆) 11.10/11.05(br s, 1H), 8.20/7.76(d, J ═ 2.0Hz, 1H), 7.68/7.57(t, J ═ 2.0Hz, 1H), 7.50-7.42(m, 3H), 5.45-5.36(m, 1H), 5.10-5.06(m, 2H), 4.29-4.18(m, 2H), 4.13(d, J ═ 7.6Hz, 1H), 3.84-3.80(m, 1H), 3.64-3.60(m, 1H), 3.41/3.24(s, 3H), 3.35-3.24(m, 1H), 2.67-2.55(m, 1H), 2.43-2.33(m, 1H), 2.17-2.09(m, 1H); LCMS 95.1%, M/z [ M + H ]]⁺＝620.2。

Synthesis of 250.2:

to a stirred solution of 250.1(1.4g, 2.26mmol) in THF (20mL) at room temperature were added aniline (210mg, 2.26mmol) and Pd (PPh)₃)₄(522mg, 0.45 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-SELECT-C18(150X30) mm, 5 μ; a is 0.1% formic acid water solution, B is acetonitrile; gradient: (time/% B) 0/45, 8/80, 10/80, 10.1/98, 13/98, 13.1/45, 15/45, 18ml/min]This gave 250.2(430mg, 33%) of a solid.

¹H NMR(500MHz，DMSO-d₆) 12.54(br s, 1H), 11.07/10.99(br s, 1H), 8.30/7.88(d, J ═ 2.0Hz, 1H), 7.66-7.37(m, 4H), 4.01(d, J ═ 7.5Hz, 1H), 3.84-3.80(m, 1H), 3.56-3.53(m, 1H), 3.40/3.24(s, 3H), 3.30-3.24(m, 1H), 2.64-2.55(m, 1H), 2.42-2.32(m, 1H), 2.19-2.03(m, 1H); LCMS 91.4%, M/z [ M-H ]]^-＝576.2。

Synthesis 250.3a & synthesis 250.3 b:

to a stirred solution of 250.2(300mg, 0.51mmol) and 4- (hydroxymethyl) -5-methyl-1, 3-dioxan-2-one (100mg, 0.77mmol) in THF (10mL) at room temperature was added triphenylphosphine (160mg, 0.62mmol) and DIAD (130mg, 0.62 mmol). Stirred at room temperature for 16 hours and the reaction mixture was concentrated under reduced pressure to give a residue. The resulting residue was purified by preparative high performance liquid chromatography [ X-selective-C18 (150X30) mm, 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/45, 8/80, 10/80, 10.1/98, 13/98, 13.1/45, 15/45at 18mL/min ] to yield 250.3a (81mg, 23%) solids and 250.3B (21mg, 5%) solids.

250.3a:¹H NMR(500MHz，DMSO-d₆) 11.14/11.06(s, 1H), 8.18/7.76(d, J ═ 2.0Hz, 1H), 7.69/7.57(t, J ═ 2.0Hz, 1H), 7.45-7.37(m, 3H), 4.91-4.87(m, 1H), 4.66/4.59(d, J ═ 14.5Hz, 1H), 4.15(d, J ═ 7.5Hz, 1H), 3.82-3.79(m, 1H), 3.66-3.63(m, 1H), 3.32-3.19(m, 1H), 3.21(s, 3H), 2.61-2.56(m, 1H), 2.38-2.36(m, 1H), 2.13-2.03(m, 1H), 2.03/2.02 (m, 1H), 2.03/3H); LCMS 98.7%, M/z [ M + H ]]⁺＝689.9。

250.3b:¹H NMR(500MHz，DMSO-d₆) 8.33/7.91(d, J ═ 2.0Hz, 1H), 7.69/7.57(t, J ═ 2.0Hz, 1H), 7.53-7.42(m, 3H), 5.12-5.03(m, 2H), 4.94-4.91(m, 1H), 4.54/4.42(d, J ═ 14.5Hz, 1H), 4.31/4.23(d, J ═ 7.5Hz, 1H), 3.82-3.80(m, 1H), 3.67-3.65(m, 1H), 3.41/3.24(s, 3H), 3.19-3.12(m, 1H), 2.58-2.50(m, 1H), 2.41-2.30(m, 1H), 2.17/2.17 (s, 2.04), 2.01-2H (m, 1H), 2.07(m, 1H); LCMS 95.5%, M/z [ M + H ]]⁺＝801.9。

Example 251: synthesis of rac- (trimethylacetoxy) methyl (1' R, 2' S, 3R, 7a ' R) -5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrido-cyclo ] -2' -carboxylate (251a), (trimethylacetoxy) methyl 5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3 ], 3 '-Pyrrolidinyl-ring ] -2' -carboxylate (251b) & rac- (trimethylacetoxy) methyl (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1- ((trimethylacetoxy) methyl) -1', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-Pyrrolidinyl-ring ] -2' -carboxylate (251c)

Synthesis 251a, 251b &251 c:

to a mixture of 250.2(200mg, 0.35mmol) and chloromethyltrimethylacetyl (80mg, 0.51mmol) in CH at room temperature₃CN (10mL) and K was added to the stirred solution₂CO₃(100mg, 0.69 mmol). After stirring at room temperature for 16 hours, the reaction mixture was quenched with water and extracted with ethyl acetate (10mL). The combined organic phases were washed with water (5mL), brine solution (5mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ X-bridge-C18(150X30) mm, 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/70, 8/90, 10/95, 14/98, 14.1/70, 17/70, 22mL/min]This gave 251a (19mg, 8%) solid, 251b (20mg, 8%) solid and 251c (49mg, 17%) solid.

251a:¹H NMR(500MHz，DMSO-d₆) 11.01/10.91(br s, 1H), 7.84-7.68(m, 2H), 7.57-7.30(m, 3H), 5.54(s, 2H), 4.34(d, J ═ 10.5Hz, 1H), 4.03-3.99(m, 1H), 3.75-3.73(m, 1H), 3.48-3.41(m, 1H), 3.23(s, 3H), 2.75-2.70(m, 1H), 2.64-2.50(m, 1H), 2.37-2.20(m, 1H), 1.07/1.05(s, 9H); LCMS 96.9%, M/z [ M + H ]]⁺＝692.0；

251b:¹H NMR(500MHz，DMSO-d₆) 11.10(br s, 1H), 8.20(br s, 1H), 7.71(t, J ═ 2.0Hz, 1H), 7.56-7.44(m, 3H), 5.54-5.47(m, 2H), 4.14(d, J ═ 7.5Hz, 1H), 3.79-3.76(m, 1H), 3.68-3.66(m, 1H), 3.40/3.21(s, 3H), 3.18-3.05(m, 1H), 2.64-2.51(m, 1H), 2.42-2.32(m, 1H), 2.13-2.00(m, 1H), 1.15-1.03(m, 9H); LCMS 95.5%, M/z [ M + H ]]⁺692.0; (Absolute stereochemistry of epimerized Material was not determined)

251c:¹H NMR(500MHz，DMSO-d₆) (in wheel form) 8.01(d, J ═ 8.012.0Hz，1H)，7.77(br s，1H)，7.68-7.65(m，1H)，7.55-7.45(m，2H)，5.78-5.71(m，2H)，5.56-5.43(m，2H)，4.45(d，J＝10.0Hz，1H)，3.96-3.92(m，1H)，3.76-3.71(m，1H)，3.48-3.37(m，1H)，3.24(s，3H)，2.72-2.67(m，1H)，2.55-2.50(m，1H)，2.38-2.23(m，1H)，1.10-1.06(m，18H)；LCMS:93.7％，m/z[M+H]⁺＝806.0。

Example 252: synthesis of (5-methyl-2-oxo-1, 3-dioxo-4-yl) methyl (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylate

Synthesis of 252.1:

thionyl chloride (10mL) was added to 110.4_1a (500mg, 1.08mmol) at room temperature. After stirring for 2 hours, the filtered thionyl chloride was removed under reduced pressure to yield the intermediate acid chloride. Reaction of this intermediate acid chloride in CH at room temperature₂Cl₂(5mL) to a solution of 3, 5-dichloro-N-methylaniline (380mg, 2.16mmol) in CH was added₂Cl₂(5mL) was stirred at room temperature for 16h, and the reaction mixture was quenched with water (10mL). Separating the organic layer with CH₂Cl₂(2x10mL) the combined organic phases were extracted. The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 30% ethyl acetate in petroleum ether) to yield 252.1(510mg, 74%) of a solid.

¹H NMR(500MHz，DMSO-d₆) 11.10/11.05(br s, 1H), 8.20/7.76(d, J-1.5 Hz, 1H), 7.68/7.57(t, J-2.0 Hz, 1H), 7.50-7.42(m, 3H), 5.44-5.38(m, 1H), 5.10-5.06(m, 2H), 4.26-4.19(m, 2H), 4.13(d, J-7.5 Hz, 1H), 3.84-3.81(m, 1H), 3.63-3.61(m, 1H), 3.41/3.24(s, 3H), 3.32-3.24(m，1H)，2.64-2.58(m，1H)，2.40-2.36(m，1H)，2.19-2.09(m，1H)；LCMS:88.4％，m/z[M+H]⁺＝620.2。

Synthesis 252.2:

to a stirred solution of 252.1(500mg, 0.80mmol) in THF (10mL) at room temperature was added aniline (46mg, 0.50mmol) and Pd (PPh)₃)₄(144mg, 0.12 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-Selectivity-C18 (150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/45, 8/80, 10/80, 10.1/98, 13/98, 13.1/45, 15/45at 18mL/min]This gave 252.2(165mg, 35%) as a solid.

¹H NMR(500MHz，DMSO-d₆) 12.54(br s, 1H), 11.07/10.99(br s, 1H), 8.30/7.88(d, J ═ 1.5Hz, 1H), 7.66-7.43(m, 4H), 4.01(d, J ═ 7.5Hz, 1H), 3.84-3.80(m, 1H), 3.55-3.53(m, 1H), 3.40/3.24(s, 3H), 3.32-3.24(m, 1H), 2.64-2.50(m, 1H), 2.42-2.32(m, 1H), 2.14-2.06(m, 1H); LCMS 94.7%, M/z [ M + H ]]⁺578.0; the chiral purity is 99.8 percent.

Synthesis of 252:

to a stirred solution of 252.2(300mg, 0.51mmol) in 4- (hydroxymethyl) -5-methyl-1, 3-dioxan-2-one (101mg, 0.77mmol) in THF (5mL) was added triphenylphosphine (163mg, 0.62mmol) and DIAD (125mg, 0.62mmol) at room temperature. After stirring at room temperature for 16 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by flash chromatography (40g silica gel column, 30% ethyl acetate in petroleum ether) followed by preparative HPLC [ X-SELECT-C18(150X25) mm, 5. mu.; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/65, 8/80, 10/90, 10.1/65, 13/65, 22mL/min ] to yield 252(120mg, 33%) of a solid.

¹H NMR(500MHz，DMSO-d₆) 11.14/11.06(s, 1H), 8.18(d, J ═ 2.0Hz, 1H), 7.76-7.69(m, 1H), 7.57-7.37(m, 3H), 4.91-4.87(m,1H)，4.68-4.65(m，1H)，4.23-4.14(m，1H)，3.82-3.81(m，1H)，3.66-3.63(m，1H)，3.32-3.19(m，1H)，3.24(s，3H)，2.61-2.54(m，1H)，2.41-2.36(m，1H)，2.09-2.03(m，1H)，2.03(s，3H)；LCMS:98.1％，m/z[M+H]⁺689.9; the chiral purity is 97.2%.

Example 253: synthesis of (5-methyl-2-oxo-1, 3-dioxo-4-yl) methyl (1'S, 2' R, 3S, 7a 'S) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylate

Synthesis 253.1:

thionyl chloride (8mL) was added to 110.4_1b (400mg, 0.86mmol) at room temperature. After stirring for 2 hours, the filtered thionyl chloride was removed under reduced pressure to yield the intermediate acid chloride. To this acid chloride in CH at room temperature₂Cl₂(5mL) to a solution of 3, 5-dichloro-N-methylaniline (308mg, 1.75mmol) in CH was added₂Cl₂(5 mL). After stirring at room temperature for 16 hours, the reaction was quenched with water (10ml). Separating the organic layer with CH₂Cl₂The aqueous layer was extracted (10mL x 2). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 30% ethyl acetate in petroleum ether) to yield 253.1(410mg, 74%) as a solid.

¹H NMR(500MHz，DMSO-d₆) 11.13/11.05(br s, 1H), 8.20/7.76(d, J ═ 2.0Hz, 1H), 7.68/7.57(t, J ═ 2.0Hz, 1H), 7.50-7.42(m, 3H), 5.42-5.38(m, 1H), 5.10-5.06(m, 2H), 4.26-4.19(m, 2H), 4.13(d, J ═ 7.5Hz, 1H), 3.84-3.81(m, 1H), 3.63-3.61(m, 1H), 3.41/3.24(s, 3H), 3.32-3.24(m, 1H), 2.64-2.58(m, 1H), 2.42-2.34(m, 1H), 2.20-2.09(m, 1H); LCMS 84.1%, M/z [ M + H ]]⁺＝620.2。

Synthesis 253.2:

to a stirred solution of 253.1(400mg, 0.64mmol) in THF (10mL) at room temperature were added aniline (60mg, 0.64mmol) and Pd (PPh)₃)₄(138mg, 0.12 mmol). Stirred at room temperature for 2 hours. After completion of the reaction, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-SELECT-C18(150X30) mm, 5 μ; a is 0.1% formic acid water solution, B is acetonitrile; gradient: (time/% B) 0/45, 8/80, 10/80, 10.1/98, 13/98, 13.1/45, 15/45, 18mL/min]Thereby yielding 253(140mg, 37%) as a solid.

¹H NMR(500MHz，DMSO-d₆) 12.55(br s, 1H), 11.07/11.00(s, 1H), 8.30/7.88(d, J ═ 2.0Hz, 1H), 7.66/7.56(t, J ═ 2.0Hz, 1H), 7.49-7.43(m, 2H), 4.02(d, J ═ 8.0Hz, 1H), 3.84-3.80(m, 1H), 3.56-3.53(m, 1H), 3.40/3.24(s, 3H), 3.32-3.24(m, 1H), 2.64-2.54(m, 1H), 2.41-2.31(m, 1H), 2.17-2.02(m, 1H); LCMS 90.8%, [ M + H%]⁺578.0; the chiral purity is 99.8 percent.

Synthesis 253:

to a stirred solution of 253.2(300mg, 0.51mmol) and 4- (hydroxymethyl) -5-methyl-1, 3-dioxan-2-one (101mg, 0.77mmol) in THF (5mL) was added triphenylphosphine (163mg, 0.62mmol) and DIAD (125mg, 0.62mmol) at room temperature. After stirring at room temperature for 16 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by flash chromatography (40g silica gel column, 30% ethyl acetate in petroleum ether) followed by preparative HPLC [ KROMOSIL-C18(150x25) mm, 10 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/50, 8/70, 12/98, 14/98, 14.1/50, 16/50, 22mL/min ] to yield 253(134mg, 37%) solids.

¹H NMR(500MHz，DMSO-d₆) 11.10/11.06(s, 1H), 8.18(d, J ═ 2.0Hz, 1H), 7.76-7.69(m, 1H), 7.57-7.37(m, 3H), 4.91-4.87(m, 1H), 4.68-4.65(m, 1H), 4.23-4.14(m, 1H), 3.82-3.81(m, 1H), 3.66-3.63(m, 1H), 3.31-3.21(m，1H)，3.24(s，3H)，2.60-2.54(m，1H)，2.40-2.36(m，1H)，2.11-2.02(m，1H)，2.02(s，3H)；LCMS:99.0％，m/z[M+H]⁺689.9; the chiral purity is 99.5 percent.

Example 254: synthesis of (pivaloyloxy) methyl (1' R, 2' S, 3R, 7a ' R) -5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyridone ring ] -2' -carboxylate (254a) and (pivaloyloxy) methyl (1' R, 2' R, 3R, 7a ' R) -5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazin-cyclo ] -2' -carboxylate (254b):

252.2(350mg, 0.60mmol) and chloromethyltrimethyl acetate (273mg, 1.81mmol) in CH at room temperature₂Cl₂:CH₃To the stirred solution of CN (4:1, 5mL) was added DBU (275mg, 1.81 mmol). Stir at rt for 16h, wash the reaction mixture with water (5mL), brine (5mL), dry over sodium sulfate, filter and concentrate under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ X BRIDGE-C18(150X25) mm, 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/65, 8/85, 10/90, 14/98, 17/98, 17.1/65, 20/65at 24mL/min]This gave 254a (95mg, 22%) as a solid and 254b (58mg, 14%) as a solid (epimeric material).

254a:¹H NMR(500MHz，DMSO-d₆) 11.15/11.06(br s, 1H), 8.20(d, J ═ 1.5Hz, 1H), 7.84-7.70(m, 1H), 7.55-7.44(m, 3H), 5.54-5.47(m, 2H), 4.15-4.13(m, 1H), 3.80-3.76(m, 1H), 3.68-3.65(m, 1H), 3.21(s, 3H), 3.20-3.10(m, 1H), 2.60-2.54(m, 1H), 2.38-2.33(m, 1H), 2.07-2.04(m, 1H), 1.07/1.03(m, 9H); LCMS 99.4%, M/z [ M + H ]]⁺691.9; the chiral purity is 94.6 percent. Stereochemistry was determined by 2D NMR studies.

254b:¹H NMR(500MHz，DMSO-d₆) 11.08/10.92(br s, 1H), 7.84(d, J ═ 1.5Hz, 1H), 7.77(s, 1H), 7.54-7.44(m, 3H), 5.60-5.50(m, 2H), 4.34(d, J ═ 10.5Hz, 1H), 4.11-3.97(m, 1H), 3.77-3.71(m, 1H), 3.50-3.41(m, 1H), 3.23(s, 3H), 2.75-2.70(m, 1H), 2.55-2.45(m, 1H), 2.31-2.20(m, 1H), 1.07(s, 9H); LCMS 97.8%, M/z [ M + H ]]⁺691.9; chiral purity 95.7%. Determination of stereochemistry from 2D NMR studies

Example 255: synthesis of (pivaloyloxy) methyl (1' S, 2' R, 3S, 7a ' S) -5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyridone ring ] -2' -carboxylate (255a) and (pivaloyloxy) methyl (1' S, 2' S, 3S, 7a ' S) -5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazin-cyclo ] -2' -carboxylate (255b):

253.2(350mg, 0.60mmol) and chloromethyl trimethyl acetate (273mg, 1.81mmol) in CH at room temperature₂Cl₂:CH₃To the stirred solution of CN (4:1, 5mL) was added DBU (275mg, 1.81 mmol). Stir at room temperature for 16h, wash the reaction mixture with water (5mL), brine solution (5mL), dry over sodium sulfate, filter and concentrate under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ KROMOSIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/70, 8/90, 10/95, 12/98, 14/98, 14.1/70, 16/70, 18mL/min]This gave 255a (65mg, 15%) and 255b (108mg, 25%) as a solid.

255a:¹H NMR(500MHz，DMSO-d₆) 11.07/10.91(s, 1H), 7.84(s, 1H), 7.77(s, 1H), 7.54-7.44(m, 3H), 5.56-5.54(m, 2H), 4.43-4.33(m，1H)，4.03-3.99(m，1H)，3.77-3.71(m，1H)，3.50-3.40(m，1H)，3.23(s，3H)，2.75-2.64(m，1H)，2.54-2.50(m，1H)，2.47-2.25(m，1H)，1.07(s，9H)；LCMS:99.2％，m/z[M+H]⁺692.0; the chiral purity is 99.6 percent.

255b:¹H NMR(500MHz，DMSO-d₆) 11.06/10.90(br s, 1H), 8.20(d, J ═ 1.5Hz, 1H), 7.84-7.70(m, 1H), 7.55-7.45(m, 3H), 5.54-5.47(m, 2H), 4.18-4.13(m, 1H), 3.80-3.76(m, 1H), 3.68-3.65(m, 1H), 3.21(s, 3H), 3.21-3.10(m, 1H), 2.65-2.50(m, 1H), 2.40-2.30(m, 1H), 2.11-1.98(m, 1H), 1.15-1.03(m, 9H); LCMS 99.6%, M/z [ M + H ]]⁺691.9; the chiral purity is 97.1%.

Example 256: synthesis of acyloxymethyl (1'R, 2' R, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyridoxine ] -2' -carboxylate (256a) and acyloxymethyl (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3 ], 3 '-Pyrazin ring ] -2' -carboxylate (256b):

252.2(300mg, 0.52mmol) in DCM (6mL) and CH at 0 deg.C₃CN (1.5mL) was added to the stirred solution DBU (87mg, 0.52mmol) and methyl bromoacetate (87mg, 0.52 mmol). Stirred at room temperature for 12 hours. The reaction mixture was diluted with water (30mL) and extracted with DCM (2 × 50 mL). The combined organic phases were washed with brine (50mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography (column: X-bridge-C18(150X30) mm, 5 μ; A: 0.1% formic acid in water, B: acetonitrile; gradient: 0/60, 8/80, 10/85, 13/98, 15/98, 15.1/60, 18/60, 23mL/min) to yield 256a (Peak-1, 30mg, 9%) and 256B (Peak-2, 60mg, 18%) as off-white solids。

256a:¹H NMR(500MHz，DMSO-d₆) 11.10/10.93(s, 1H), 7.81(d, J ═ 1.5Hz, 1H), 7.77(s, 1H), 7.56-7.44(m, 3H), 5.54(d, J ═ 6.0Hz, 1H), 5.45(d, J ═ 6.0Hz, 1H), 4.33(d, J ═ 10.5Hz, 1H), 4.01-3.97(m, 1H), 3.78-3.73(m, 1H), 3.51-3.44(m, 1H), 3.23(s, 3H), 2.77-2.72(m, 1H), 2.51-2.49(m, 1H), 2.37-2.36(m, 1H), 2.01(s, 3H); LCMS 96.7%, M/z [ M + H ]]⁺650.0; the chiral purity is 99.9 percent.

256b:¹H NMR(500MHz，DMSO-d₆) 11.10/11.05(s, 1H), 8.18/7.75(d, J ═ 2.0Hz, 1H), 7.70(t, J ═ 2.0Hz, 1H), 7.58-7.45(m, 3H), 5.52/5.48(d, J ═ 6.0Hz, 1H), 5.40-5.38(m, 1H), 4.14(d, J ═ 7.5Hz, 1H), 3.80-3.77(m, 1H), 3.68-3.65(m, 1H), 3.40/3.23(s, 3H), 3.21-3.17(m, 1H), 2.64-2.63(m, 1H), 2.37-2.35(m, 1H), 2.11-2.08(m, 1H), 1.97 (m, 1H, 89H); LCMS 98.0%, M/z [ M + H ]]⁺650.1; the chiral purity is 99.5 percent.

Example 257: synthesis of acyloxymethyl (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylate:

257 was synthesized from 250.2 following the procedure described for 256a and 256 b.

¹H NMR(500MHz，DMSO-d₆) 11.15/11.05(s, 1H), 8.18/7.75(d, J ═ 2.0Hz, 1H), 7.69/7.57(t, J ═ 2.0Hz, 1H), 7.51-7.45(m, 3H), 5.52/5.48(d, J ═ 6.0Hz, 1H), 5.40-5.38(m, 1H), 4.14(d, J ═ 7.5Hz, 1H), 3.82-3.77(m, 1H), 3.66(t, J ═ 7.0Hz, 1H), 3.40/3.23(s, 3H), 3.21-3.17(m, 1H), 2.61-2.51(m, 1H), 2.42-2.30(m, 1H), 2.15-2.00(m, 1H), 2.89 (m, 1H); LCMS 97.5%, M/z [ M + H ]]⁺＝650。

Example 258: synthesis of 2- (dimethylamine) -2-oxyethyl (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyridoxine ring ] -2' -carboxylate:

252.2(400mg, 0.69mmol) and 2-bromo-N, N-dimethylacetamide (230mg, 1.38mmol) in CH at room temperature₂Cl₂:CH₃To the stirred solution of CN (4:1, 8mL) was added DBU (210mg, 1.38 mmol). Stirred at room temperature for 16 hours with CH₂Cl₂The reaction mixture was diluted (10mL). The organic layer was washed with water (10mL), brine (10mL), dried over sodium sulfate and filtered. The organic layer was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-bridge-C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/60, 8/80, 11/90, 11.1/98, 13/98, 13.1/60, 16/60at 18mL/min]Thus 258(198mg, 43%) solid was produced.

¹H NMR(500MHz，DMSO-d₆) 11.13/11.05(s, 1H), 8.17/7.75(d, J ═ 2.0Hz, 1H), 7.68-7.67(m, 1H), 7.56-7.45(m, 3H), 4.69-4.44(m, 2H), 4.25-4.07(m, 1H), 3.84-3.81(m, 1H), 3.74-3.71(m, 1H), 3.22(s, 3H), 3.19-3.00(m, 1H), 2.83/2.82(s, 3H), 2.77(s, 3H), 2.63-2.57(m, 1H), 2.35-2.34(m, 1H), 2.13-2.10(m, 1H); LCMS 99.2%, M/z [ M + H ]]⁺663.2; the chiral purity is 97.3 percent.

Example 259: synthesis of S- (2-amidoethyl) (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino-cyclo ] -2' -carbosulphate:

to a stirred solution of 252.2(400mg, 0.69mmol) in THF (8mL) at 0 deg.C was added N-methylmorpholine (140mg, 1.38mmol) and isobutyl chloroformate (141mg, 1.38 mmol). After 30min, N- (2-mercaptoethyl) acetamide (246mg, 2.07mmol) was added at 0 ℃. Stir at room temperature for 16h, quench the reaction mixture with water and extract with ethyl acetate (2 × 10mL), wash the combined organic phases with water (20mL), brine (20mL), dry over sodium sulfate, filter and concentrate under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMASIL C18(150x25) mm, 10 μ; a is a solution of 10mM ammonium bicarbonate in water, B is acetonitrile; gradient: (time/% B) 0/50, 8/80, 10/80, 10.1/98, 11/98, 11.1/50, 14/50, 20mL/min to yield 259(38mg, 8%) of a solid.

¹H NMR(500MHz，DMSO-d₆) 11.00(br s, 1H), 7.93-7.76(m, 3H), 7.57-7.46(m, 3H), 4.68-4.35(m, 1H), 4.07-4.03(m, 1H), 3.80-3.79(m, 1H), 3.54-3.49(m, 1H), 3.27(s, 3H), 3.00-2.98(m, 2H), 2.78-2.73(m, 3H), 2.57-2.50(m, 1H), 2.36-2.23(m, 1H). 1.75(s, 3H); LCMS 98.6%, M/z [ M + H ]]⁺＝679.2。

Example 260: synthesis of (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N2 '-hydroxy-N1', N2 '-dimethyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino-cyclo ] -1', 2' -dicarboxamide (260a) and (1'R, 2' R, 3R, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N2 '-hydroxy-N1', N2 '-dimethyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide (260b):

to a stirred solution of 252.2(900mg, 1.55mmol) in DMF (20mL) at room temperature was added HATU (709mg, 1.86mmol) and DIPEA (0.5mL, 3.10 mmol). After stirring for 15 min, N-methylhydroxylamine (193mg, 2.33mmol) was added. Stir at room temperature for 16h, dilute the reaction mixture with ice water (20mL) and extract with ethyl acetate (2 × 20mL). The combined organic phases were washed with cold water (20ml), brine (20ml), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMASIL C18(150x25) mm, 10 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/50, 8/65, 11/65, 11.1/98, 12/98, 12.1/50, 14/50at 23mL/min to yield 260a as an off-white solid (83mg) and 260B as a white solid (55mg, 6%).

260a:¹H NMR(500MHz，DMSO-d₆) 10.88/10.78(br s, 1H), 10.07/9.95(br s, 1H), 7.72(s, 1H), 7.54-7.30(m, 4H), 4.63-4.41(m, 1H), 4.12(m, 1H), 3.51-3.39(m, 2H), 3.37/3.22(s, 3H), 2.99/2.97(s, 3H), 2.74-2.69(m, 1H), 2.60-2.50(m, 1H), 2.22-2.10(m, 1H); LCMS 98.3%, M/z [ M + H ]]⁺607.2; the chiral purity is 98.9 percent.

260b:¹H NMR(500MHz，DMSO-d₆) 10.69(s, 1H), 9.81(s, 1H), 7.93(s, 1H), 7.63(s, 1H), 7.51(s, 2H), 7.45(d, J ═ 2.0Hz, 1H), 4.07(d, J ═ 7.5Hz, 1H), 3.83-3.79(m, 1H), 3.50-3.44(m, 1H), 3.36(t, J ═ 6.5Hz, 1H), 3.24(s, 3H), 2.72(s, 3H), 2.50-2.33(m, 2H), 2.21-2.12(m, 1H); LCMS 95.4%, M/z [ M + H ]]⁺607.1; the chiral purity is 87.9 percent.

Example 261: synthesis of (1' R, 2' S, 3R, 7a ' R) -5, 7-dichloro-N1 ' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N1 ' -methyl-2-oxo-N2 ' - (phenylsulfonyl) -1', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino-cyclo ] -1', 2' -dicarboxamide:

to a stirred solution of 252.2(400mg, 0.69mmol) in DMF (6mL) at room temperature were added HATU (395mg, 1.04mmol) and DIPEA (0.23mL, 1.39 mmol). After stirring for 15 min benzenesulfonamide (163mg, 1.04mmol) was added. After stirring at room temperature for 48 h, the reaction mixture was diluted with ice water (80ml) and extracted with ethyl acetate (2 × 50 mL). The combined organic phases were washed with brine (50mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: x bridge C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: 0/60, 8/75, 10/75, 10.1/98, 12/98, 12.1/60, 15/60, 23mL/min ] to yield 261(102mg, 21%) solid.

¹H NMR(300MHz，DMSO-d₆) 12.00(br s, 1H), 10.85/10.65(br s, 1H), 7.73-7.40(m, 10H), 4.65-4.52(m, 1H), 4.28-4.10(m, 1H), 4.01-3.92(m, 1H), 3.70-3.60(m, 1H), 3.20(s, 3H), 3.00-2.80(m, 1H), 2.51-2.40(m, 1H), 2.15-2.00(m, 1H); LCMS 94.2%, M/z [ M + H ]]⁺716.9; the chiral purity is 94.5 percent.

Example 265: synthesis of (1'S, 2' R, 3S, 7a 'S) -5, 7-dichloro-1' - ((3, 5-dichloro-2-fluorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyridoxal ] -2' -carboxylic acid (265a) & (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichloro-2-fluorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3 ], 3 '-pyranin cyclo ] -2' -carboxylic acid (265b):

synthesis 265.2:

to a stirred solution of 265.1(500mg, 2.77mmol) in MeOH (10mL) at room temperature was added paraformaldehyde (0.083g, 2.77mmol) and AcOH (0.1mL, cat.). After stirring at room temperature for 3 hours, NaCNBH was added₃(523mg, 8.33 mmol). Stirring at 60 ℃ for 16h, evaporating the resulting reaction mixture under reduced pressure, diluting with water (25mL) and extracting with ethyl acetate (2)x15 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified (silica gel 100. sup. 200mesh, 2% EtOAc/n-hexane) to give 265.2(200mg, 37%) as a yellow liquid.

¹HNMR(400MHz，CDCl₃):6.69-6.63(m，1H)，6.55-6.49(m，1H)，4.12(br s，1H)，2.87(d，J＝5.2Hz，3H)；LCMS:94.7％，m/z[M+H]⁺＝194.0。

Synthesis 265.3:

thionyl chloride (5mL) was added to compound 110.4_1(350mg, 0.76mmol) at room temperature. After stirring for 2 hours, the reaction mixture was concentrated under reduced pressure. To the intermediate acid chloride in CH₂Cl₂₍3mL) was added 265.2(221mg, 1.13mmol) in CH₂Cl₂(2mL), stirred at room temperature for 16h, quenched with water (25mL) and CH₂Cl₂(2 × 15 mL). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The batch was purified by HPLC (silica gel 100-200mesh, 10% EtOAc/n-hexane) to give 265.3(200mg, 41%) as a solid. LCMS 57.1%, M/z [ M-H ]]^-＝636.2。

Synthesis 265a &265 b:

to a stirred solution of 265.3(200mg, 0.31mmol) in THF (5mL) at room temperature were added aniline (29.0mg, 0.31mmol) and Pd (PPh)₃)₄(72.0mg, 0.062 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. By chiral SFC [ chiral inclusion IE (30x250) mm, 5 μ; 80% CO₂The residue was purified at room temperature (isocratic 70g/min, detection at 214 nm) in 20% 0.5% isopropylamine in isopropanol to yield 265a (7mg) and 265b (7mg) as solids.

265a:¹H NMR(300MHz，DMSO-d₆) 12.83-12.14(br s, 1H), 10.99(br s, 1H), 8.26/8.22(s, 1H), 7.88-7.82(m, 2H), 7.70-7.43(m, 1H), 4.15-3.88(m, 3H), 3.31-3.26(m, 1H), 3.20(s, 3H), 2.73-2.43(m, 2H), 2.10-1.95(m, 1H); LCMS 98.1％，m/z[M+H]⁺595.9, chiral purity 96.9%. The absolute stereochemistry was not determined.

265b:¹H NMR(300MHz，DMSO-d₆) 11.19-10.24(br s, 1H), 8.28/8.22(br s, 1H), 7.83-7.38(m, 3H), 4.13-3.83(m, 4H), 3.32-3.25(m, 1H), 3.18(s, 3H), 2.60-2.44(m, 1H), 2.30-2.25(m, 1H); LCMS 95.7%, M/z [ M + H ]]⁺595.9; the chiral purity is 90.9%. The absolute stereochemistry was not determined.

Example 266: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino-cyclo ] -2' -carboxylic acid

Synthesis 266.1:

thionyl chloride (10mL) was added to 82.4_1(500mg, 1.17 mmol). After stirring at room temperature for 2 hours, the reaction mixture was concentrated under reduced pressure to yield the acid chloride intermediate. To this acid chloride intermediate was added 3, 5-dichloro-N-methylaniline (395mg, 2.26mmol) in CH₂Cl₂(10mL). The reaction mixture was stirred at room temperature for 16 hours and concentrated under reduced pressure. The resulting residue was purified by flash chromatography (silica gel, 15% EtOAc/petroleum ether) to give 266.1(200mg, 29%) as a light gray solid. LCMS 96.1%, M/z [ M-H ]]^-＝581.8。

Synthesis 266:

to a stirred solution of 266.1(100mg, 0.17mmol) in THF (5mL) at room temperature were added aniline (16.1mg, 0.17mmol) and Pd (PPh)₃)₄(39.6mg, 0.03 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ Kromasil C18(150x25), 10 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/60, 8/90, 9/80, 9.1/80, 10/98, 10.1/60, 14/60, 25mL/min]Thereby generating 266(60mg, 9% yield) as a solid.

¹H NMR(500MHz，DMSO-d₆) 12.47(brs, 1H), 10.98 (brs, 1H), 8.41/7.91(d, J ═ 1.5Hz, 1H), 7.68-7.54(m, 1H), 7.47-7.40(m, 3H), 4.09(d, J ═ 8.0Hz, 1H), 3.80-3.77(m, 1H), 3.59-3.50(m, 1H), 3.41/3.23(s, 3H), 2.88-2.79(m, 1H), 2.30-2.19(m, 1H), 1.98-1.88(m, 1H), 1.75-1.70(m, 2H), 1.60-1.50(m, 1H); LCMS 91.4%, M/z [ M + H ]]⁺＝542.1。

Example 267: synthesis of 1' - ((3- (tert-butyl) phenethyl) carbamoyl) -5, 7-dichloro-7 ', 7' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -2' -carboxylic acid:

synthesis 267.2:

to a stirred solution of 267.1(5.0g, 19.4mmol) in DCM (50mL) was added DAST (7.8mL, 58.3mmol) dropwise at-78 deg.C. Warming to room temperature and stirring at room temperature for 16h, adding saturated NaHCO at 0 deg.C₃The reaction mixture was quenched by solution and extracted with DCM (2 × 20mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash chromatography (80g silica gel column, 20% -30% EtOAc/petroleum ether) to yield 267.2(5.0g, 92%) as a liquid.

¹H NMR(500MHz，CDCl₃) 4.44-4.40(m, 1H), 4.30-4.23(m, 2H), 3.78-3.75(m, 1H), 3.57-3.55(m, 1H), 2.60-2.30(m, 2H), 1.48-1.42(m, 9H), 1.33-1.29(m, 3H).

Synthesis 267.3:

to 267.2(8g, 28.6mmol) was added 6N HCl (80mL) at room temperature. The reaction mixture was stirred at 65 ℃ for 6 hours and concentrated under reduced pressure. The crude product was washed with EtOAc: DCM (1: 3; 40mL) for 3h and dried under high vacuum to give 267.3 as a brown solid (5.6g, 73%).

¹H NMR(400MHz，D₂O):4.64-4.58(m，1H)，3.66(t，J＝7.6Hz，2H)，2.74-2.62(m，2H)。

Synthesis 267.4:

to a solution of (Z) -4- (allyloxy) -4-oxybut-2-enoic acid (4.5g, 28.8mmol) in EtOH (50mL) was added 267.3(5.39g, 28.8mmol) and 5, 7-dichloroindole-2, 3-dione (6.23g, 28.8mmol) at room temperature. After stirring at reflux for 2 hours, the reaction mixture was concentrated. The resulting residue was purified by flash chromatography (80g silica gel column, 100% EtOAc) to give 267.4(4.2g, 32%) as a brown solid.

LCMS:(28+32+10+6)％，m/z[M+H]⁺＝461.0)。

Synthesis 267.5:

to a stirred solution of 267.4(1.5g, 3.25mmol) in DMF (10mL) at room temperature were added DIPEA (1.7mL, 9.75mmol) and HATU (1.48g, 3.90 mmol). After stirring for 10min, 2- (3- (tert-butyl) phenyl) ethan-1-amine (0.86g, 4.87mmol) was added. After stirring at room temperature for 3 hours, the reaction mixture was poured into ice water and stirred for 10 minutes. The resulting precipitate was filtered, washed with ice water (2 × 20mL) and dried under vacuum (1.3 g). The resulting residue was purified by preparative high performance liquid chromatography [ column: X-Selectivity-C18 (150X19) mm, 5 μ; a is a solution of 10mM ammonium bicarbonate in water, B is acetonitrile; gradient: (T% B):0/50, 8/80, 10/80, 14/98, 16/98, 16.1/50, 18/50, 18mL/min ] to yield the major diastereomer (600mg) which was purified by flash chromatography (80g silica gel column, 20% -30% EtOAc/petroleum ether) to yield 267.5(200mg, 10%) solid.

¹HNMR(400MHz，DMSO-d₆):11.20(s，1H)，8.57-8.54(m，1H)，7.58(d，J＝1.6Hz，1H)，7.23-7.14(m，3H)，7.07-7.01(m，2H)，5.33-5.24(m，1H)，5.06-5.02(m，2H)，4.21-4.16(m，2H)，4.01-3.96(m，3H)，3.67-3.61(m，2H)，2.72-2.50(m，4H)，2.50-2.33(m，2H)，1.27(s，9H)；LCMS:95.5％，m/z[M+H]⁺＝620.0)。

Synthesis 267:

at room temperature to 2675(200mg, 0.33mmol) to a stirred solution in THF (2mL) was added aniline (30mg, 0.32mmol) and Pd (PPh)₃)₄(74mg, 0.064 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ XBRIDGE-C18(150x25), 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/60, 8/90, 8.1/98, 10/98, 10.1/60, 13/60at 24mL/min]This gave 267(160mg, 85%) as a solid.

¹H NMR(500MHz，DMSO-d₆):12.65(br s，1H)，11.13(br s，1H)，8.49(br s，1H)，7.57(s，1H)，7.23-7.18(m，3H)，7.11(s，1H)，7.07-7.01(m，1H)，4.02-3.90(m，2H)，3.60-3.56(m，1H)，3.32-3.22(m，2H)，2.71-2.67(m，2H)，2.50-2.22(m，4H)，1.27(s，9H)；LCMS:97.8％；m/z[M+H]⁺580.1; the absolute stereochemistry is unknown. Testing as racemate.

Example 268: synthesis of 5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -7', 7' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -2' -carboxylic acid (268a &268b):

synthesis 268.1a &268.1b

To a stirred solution of 267.4(500mg, 1.08mmol) in DCM (8mL) was added 3, 5-dichloro-N-methylaniline (230mg, 1.30mmol), TEA (0.36mL, 2.61mmol) and DMAP (39mg, 0.32mmol) at room temperature. After stirring for 30min, POCl dissolved in DCM (1mL) was added₃(0.10mL, 1.084 mmol). Stir at rt for 16h, quench the reaction mixture with water (5mL) and extract with DCM (2 × 10mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ X-bridge C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/70, 8/90, 10/90, 10.1/98, 12/98, 12.1/70, 15/70at20mL/min]This gave 268.1a as a white solid (Peak-1, 390mg, 58%) and 268.1b as a white solid (Peak-2, 140mg, 21%).

268.1a:¹H NMR(400MHz，DMSO-d₆):11.05(br s，1H)，7.90-7.44(m，5H)，5.80-5.64(m，1H)，5.25-4.90(m，2H)，4.90-4.75(m，1H)，4.39(br s，2H)，3.90-3.50(m，2H)，3.40-3.10(m，4H)，2.57-2.50(m，1H)，2.32-2.25(m，2H)；LCMS:95.8％，m/z[M+H]⁺＝620.0。

268.1b:¹H NMR(400MHz，DMSO-d₆):11.24(s，1H)，7.78(s，1H)，7.58-7.56(m，3H)，6.42(s，1H)，5.35-5.27(m，1H)，5.07-5.03(m，2H)，4.31-4.19(m，3H)，3.93-3.88(m，1H)，3.78-3.73(m，1H)，3.40-3.20(m，4H)，2.63-2.50(m，1H)，2.30-2.21(m，2H)；LCMS:86.9％，m/z[M+H]⁺＝620.2。

Synthesis 268 a:

to a stirred solution of 268.1a (400mg, 0.65mmol) in THF (8mL) at room temperature was added aniline (60. mu.L, 0.65mmol) and Pd (PPh)₃)₄(149mg, 0.13 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ X-SELECT-C18(150X19), 5. mu.l; a10 mM NH₄HCO₃At H₂Solution in O, B is acetonitrile; gradient: (time/% B) 0/20, 8/70, 8.1/98, 11/98, 11.1/20, 13/20at 18mL/min]This gave 268a (86mg, 23%) as a solid.

¹H NMR(500MHz，DMSO-d₆) 12.90-12.70(br s, 1H), 11.00-10.80(br s, 1H), 7.61-6.50(m, 5H), 4.84-4.79(m, 1H), 4.60-4.30(m, 1H), 3.85-3.68(m, 1H), 3.32(s, 3H), 3.30-3.10(m, 1H), 2.60-2.50(m, 1H), 2.40-2.15(m, 2H); LCMS 86.1 percent; m/z [ M + H]⁺578.0; the absolute stereochemistry is unknown. Testing as racemate.

Synthesis 268 b:

to a solution of 268.1b (150mg, 0.24mmol) in THF (3mL) at room temperature was added aniline (20. mu.L, 0.24mmol) and Pd (PPh)₃)₄(55mg, 0.05 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ KROMOSIL-C18(150x25mm), 10 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/50, 8/75, 9/75, 9.1/98, 11/98, 11.1/50, 13/50at 23mL/min]This gave 268b (50mg, 35%) as a solid.

¹H NMR(500MHz，DMSO-d₆):13.01(br s，1H)，11.17(s，1H)，7.76-7.55(m，4H)，6.43(s，1H)，4.10(d，J＝11.0Hz，1H)，3.88-3.84(m，1H)，3.73-3.68(m，1H)，3.27(s，3H)，2.64-2.59(m，1H)，2.50-2.36(m，1H)，2.25-2.14(m，2H)；LCMS:99.3％，m/z[M+H]⁺578.1; the absolute stereochemistry is unknown. Testing as racemate.

Example 269: synthesis (269a &269b):

synthesis 269.2_1 and 269.2_ 2:

to a stirred solution of 269.1(2g, 13.9mmol) in EtOH (70mL) was added (Z) -4- (allyloxy) -4-oxybutyl-2-enoic acid (2.18g, 13.9mmol) and 5, 7-dichloroindole-2, 3-dione (3g, 13.9mmol) at room temperature. After stirring at 80 ℃ for 3 hours, the reaction mixture was cooled to room temperature and concentrated in vacuo. The resulting residue was purified by flash chromatography (80g silica gel column, 40% ethyl acetate in petroleum ether) to yield 269.2_1(750mg, 12%) as a major diastereomer solid and 269.2_2(36mg) as a minor diastereomer solid.

269.2_1:¹H NMR(500MHz，DMSO-d₆) 12.78(br s, 1H), 11.04(s, 1H), 7.89(d, J ═ 2.0Hz, 1H), 7.42(d, J ═ 2.5Hz, 1H), 5.47-5.39(m, 1H), 5.08-5.04(m, 2H), 4.40-4.35(m, 2H), 4.25(d, J ═ 5.5Hz, 2H), 3.94(d, J ═ 7.5Hz, 1H), 3.65(t, J ═ 7.0Hz, 1H). Determination of regional chemistry and relative stereochemistry from 2D NMR studies

269.2_2:¹H NMR(500MHz，DMSO-d6):12.65(br s，1H)，10.98(br s，1H)，7.98(d，J＝2.0Hz，1H)，7.42(d，J＝2.0Hz，1H)，5.96-5.89(m，1H)，5.42-5.37(m，2H)，4.65-4.57(m，2H)，4.42-4.38(m，2H)，3.88-3.71(m，1H)，3.69-3.65(m，1H)；LCMS:90.5％，m/z[M+H]⁺＝453.2。

Synthesis 269.3:

EDC-HCl (592mg, 3.09mmol) was added to a stirred solution of 269.2(700mg, 1.54mmol) in pyridine (10mL) at room temperature. After stirring for 10min, 3, 5-dichloro-N-methylaniline (408mg, 2.31mmol) was added. The mixture was stirred at room temperature for 2 hours and the resulting reaction mixture was evaporated under reduced pressure. The resulting residue was purified by column chromatography (silica gel 100. sup. 200. sup. mesh, 20% ethyl acetate in petroleum ether) to yield 269.3(180mg, 19%) as a solid.

Isolation 269.3a &269.3b (absolute stereochemistry of enantiomers 1&2 not determined):

269.3(280mg) by chiral SFC (Chiralcel OX-H (30X250) mm, 5 μ; 50% CO₂50% acetonitrile at room temperature (isocratic 90g/min, detection at 214 nm). Concentration of the pure fractions under reduced pressure yielded 269.3a (enantiomer-1, 90mg, 64%) as a solid and 269.3b (enantiomer-2, 90mg, 64%) as a solid.

269.3a:¹H NMR(500MHz，DMSO-d₆):11.24(s，1H)，7.78(br s，1H)，7.61-7.55(m，2H)，7.49(br s，1H)，6.51(br s，1H)，5.33-5.28(m，1H)，5.06-5.03(m，2H)，4.57(d，J＝6.0Hz，1H)，4.24-4.18(m，3H)，3.85(d，J＝11.0Hz，1H)，3.76-3.72(m，1H)，3.25(s，3H)；LCMS:90.5％，m/z[M+H]⁺611.9; the chiral purity is 99.8 percent.

269.3b:¹H NMR(500MHz，DMSO-d₆):11.24(s，1H)，7.78(br s，1H)，7.66-7.55(m，2H)，7.49(br s，1H)，6.51(br s，1H)，5.33-5.28(m，1H)，5.06-5.03(m，2H)，4.57(d，J＝5.5Hz，1H)，4.24-4.18(m，3H)，3.85(d，J＝11.0Hz，1H)，3.77-3.72(m，1H)，3.26(s，3H)；LCMS:90.2％，m/z[M+H]⁺＝611.9。

Synthesis of 269 a:

to a stirred solution of 269.3a (80mg, 0.13mmol) in THF (3mL) at room temperature were added aniline (18mg, 0.19mmol) and Pd (PPh)₃)₄(30mg, 0.026 mmol). After stirring at room temperature for 2 hours, the reaction mixture was diluted with water (10mL) and acidified with 1N HCl. The reaction mixture was extracted with ethyl acetate (20mL). The organic layer was washed with brine (10mL), dried over sodium sulfate, filtered and evaporated. The resulting residue was purified by preparative high performance liquid chromatography [ column: x bridge-C18(150X25), 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient (T% B):0/50, 8/80, 10/80, 10.1/98, 12/98, 12.1/50, 15/50, flow rate 22mL/min]Yielding 269a (40mg, 38%) as a solid.

¹H NMR(400MHz，DMSO-d₆) 13.08(br s, 1H), 11.20(br s, 1H), 7.76-7.32(m, 4H), 6.53(br s, 1H), 4.49-4.46(m, 1H), 4.14-4.11(m, 1H), 3.81-3.58(m, 2H), 3.27(s, 3H); LCMS 99.4%, [ M-H%]^-567.9. (absolute stereochemistry not determined).

Synthesis of 269 b:

to a solution of 269.3b (90mg, 0.15mmol) in THF (3mL) was added aniline (21mg, 0.22mmol) and Pd (PPh) at room temperature₃)₄(17mg, 0.015 mmol). After stirring at room temperature for 2 hours, the reaction mixture was diluted with water (10mL) and acidified with 1N HCl. The reaction mixture was extracted with ethyl acetate (20mL) and the combined organic phases were washed with brine (10mL). The combined organic phases were dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: x bridge-C18(150X25), 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient (T% B):0/50, 8/70, 10/70, 11.4/80, 11.15/98, 13/98, 13.1/50, 16/50, 22mL/min]Yielding 269b (40mg, 38%) as a solid.

¹H NMR(400MHz，DMSO-d₆) 13.04(br s, 1H), 11.20(br s, 1H), 7.76-7.33(m, 4H), 6.52(br s, 1H), 4.55-4.42(m, 1H), 4.20-4.10(m, 1H), 3.82-3.68(m, 2H), 3.27(s, 3H); LCMS 99.0% [ M-H ]]^-568.0. (absolute stereochemistry undetermined.)

Example 270: synthesis of (3'S, 4' R, 5'S) -5, 7-dichloro-4' - ((cyclopropylmethyl) (3, 5-dichlorophenyl) carbamoyl) -2-oxo-5 '- (trifluoromethyl) spiro [ indole-3, 2' -pyrrolidine ] -3 '-carboxylic acid and (3' R, 4'S, 5' R) -5, 7-dichloro-4 '- ((cyclopropylmethyl) (3, 5-dichlorophenyl) carbamoyl) -2-oxo-5' - (trifluoromethyl) spiro [ indole-3, 2 '-pyrrolidine ] -3' -carboxylic acid (270a &270b):

270a and 270b are synthesized from 269.2_1 using the procedure described in the synthesis of 269a and 269 b.

270a:¹H NMR(500MHz，DMSO-d₆) 12.69(br s, 1H), 11.04(br s, 1H), 8.23/7.93(d, J ═ 1.5Hz, 1H), 7.69-7.26(m, 4H), 4.06-3.98(m, 2H), 3.87(d, J ═ 7.0Hz, 1H), 3.80-3.76(m, 1H), 3.66(t, J ═ 6.5Hz, 1H), 3.44-3.40(m, 1H), 0.89-0.85(m, 1H), 0.42-0.36(m, 2H), 0.19-0.12(m, 1H), 0.02-0.00(m, 1H); LCMS 98.1% [ M + H ]]⁺609.9; the chiral purity is 99.0 percent. 270b, unknown absolute stereochemistry:

¹H NMR(500MHz，DMSO-d₆) 12.69(br s, 1H), 11.04(s, 1H), 8.23/7.93(d, J ═ 1.5Hz, 1H), 7.69-7.26(m, 4H), 4.06-3.98(m, 2H), 3.87(d, J ═ 7.5Hz, 1H), 3.80-3.76(m, 1H), 3.66(t, J ═ 6.5Hz, 1H), 3.44-3.40(m, 1H), 0.89-0.85(m, 1H), 0.42-0.37(m, 2H), 0.19-0.16(m, 1H), 0.03-0.00(m, 1H); LCMS 96.2% [ M + H ]]⁺609.9; the chiral purity is 97.6%. The absolute stereochemistry being unknown

Example 271: synthesis of rac- (3' S, 4' R, 5' S) -5, 7-dichloro-4 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -1-methyl-2-oxo-5 ' - (trifluoromethyl) spiro [ indole-3, 2' -pyrrolidine ] -3' -carboxylic acid (271):

synthesis 271.1:

to a stirred solution of 269.3(250mg, 0.41mmol) in acetonitrile (5mL) at room temperature was added K₂CO₃(169mg, 1.22 mmol). After stirring at room temperature for 30min, MeI (0.05mL, 0.81mmol) was added. After stirring at room temperature for 16h, the reaction mixture was quenched with water (25mL) and extracted with ethyl acetate (50mL), the organic layer was dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified using silica gel column chromatography (silica gel 100-200mesh, 10% ethyl acetate in petroleum ether) to give 271.1 as a brown solid (130mg, 51%).

¹H NMR(400MHz，DMSO-d₆):7.78(br s，1H)，7.66-7.58(m，2H)，7.52(d，J＝1.6Hz，1H)，7.36(br s，1H)，6.54(br s，1H)，5.39-5.32(m，1H)，5.13-5.05(m，2H)，4.53(d，J＝6.4Hz，1H)，4.27-4.15(m，3H)，3.90-3.80(m，1H)，3.78-3.69(m，1H)，3.44(s，3H)，3.26(s，3H)；LCMS:93.9％，m/z[M+H]⁺＝626.00。

Synthesis 271:

to a stirred solution of 271.1(120mg, 0.19mmol) in THF (3mL) at room temperature were added aniline (21mg, 0.23mmol) and Pd (PPh)₃)₄(22mg, 0.019 mmol). After stirring at room temperature for 2 hours, the reaction mixture was diluted with water (10mL) and acidified with 1N HCl. The reaction mixture was extracted with ethyl acetate (20mL), the organic phase was washed with brine (10mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-bridge-C18(150X30), 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/70, 8/90, 10/90, 10.1/98, 12/98, 12.1/70, 15/70, 20mL/min) to yield 271(40mg, 38%) solids.

¹H NMR(500MHz，DMSO-d₆) 13.11(br s, 1H), 7.82-7.33(m, 4H), 6.57(s, 1H), 4.55-4.45(m, 1H), 4.26-4.16(m, 1H), 3.80(d, J ═ 11.5Hz, 1H), 3.72-3.68(m, 1H), 3.45(s, 3H), 3.26(s, 3H); LCMS 96.6%, M/z [ M + H ]]⁺＝584.0；

Example 272: synthesis (272):

synthesis 272.2:

at 120 ℃ to 272.1(3.0g, 18.5mmol) in CH₃NO₂(150mL) to the stirred solution was added NH₄And OAc. After stirring at 120 ℃ for 16h, the reaction mixture was cooled to room temperature and concentrated under reduced pressure. The resulting residue was purified by column chromatography (silica gel 100-200mesh, 2% ethyl acetate in petroleum ether) to yield 272.2(3.4g, 89%) as a liquid.

¹H NMR(500MHz，CDCl₃):8.03(d，J＝13.5Hz，1H)，7.60(d，J＝13.5Hz，1H)，7.55-7.54(m，2H)，7.41-7.37(m，2H)，1.35(s，9H)；LCMS:95.2％，m/z[M+H]⁺＝206.2。

Synthesis 272.3:

TMSOI (8.03g, 36.5mmol) was added in portions to a stirred solution of t-BuOK (4.09g, 36.5mmol) in DMSO (50mL) at room temperature. After stirring for 10min 272.2(3.0g, 14.6mmol) was added dropwise at room temperature. After stirring for 30min, the reaction mixture was poured into ice water and extracted with ethyl acetate (2 × 50 mL). The combined organic phases were washed with brine (25mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by column chromatography (silica gel 100-200mesh, 20% ethyl acetate in petroleum ether) to give 272.3(500mg, 15%). LCMS 82.8%, M/z [ M + H ]]⁺＝220.2。

Synthesis 272.4:

to a stirred solution of 272.3(140mg, 0.63mmol) in isopropanol (15mL) was added 1N HCl (4.2mL) and Zn (823mg, 12.6mmol) at room temperature. After stirring for 2 hours, the reaction mixture was filtered through a small pad of celite and the filtrate was concentrated under reduced pressure to give 272.4(110mg) as a light brown liquid which was used in the next step without further purification.LCMS:70.1％，m/z[M+H]⁺＝190.3。

Synthesis 272.5:

to a stirred solution of 110.4_1(364mg, 0.79mmol) in DMF (5mL) at room temperature were added DIPEA (0.16mL, 0.94mmol), HATU (300mg, 0.79mmol) and 272.4(150mg, 0.79 mmol). After stirring at room temperature for 30 minutes. The reaction mixture was diluted with ice water. After stirring for 10min, the resulting precipitate was filtered, washed with ice water and dried under vacuum to give 272.5(450mg) which was used in the next step without filtration. LCMS 38.4%, M/z [ M + H ]]⁺＝632.1。

And (3) synthesizing 272:

to a stirred solution of 272.5(450mg, 0.71mmol) in THF (10mL) at room temperature were added aniline (79mg, 0.85mmol) and Pd (PPh)₃)₄(82mg, 0.038 mmol). After stirring at room temperature for 2 hours, the reaction mixture was diluted with water (10mL) and the pH was adjusted to about 6 with 1N HCl. The reaction mixture was extracted with ethyl acetate (2 × 20mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-bridge-C18(150X30), 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (T% B):0/50, 8/80, 10/80, 12/98, 14/98, 14.1/50, 17/50, 20mL/min]This gave a mixture of diastereomers 272 as an off-white solid (90mg, 21%).

¹H NMR(500MHz，DMSO-d₆):12.35(br s，1H)，10.98(s，1H)，8.51(br s，1H)，8.26(d，J＝6.5Hz，1H)，7.44-6.88(m，5H)，4.03-3.90(m，2H)，3.32-3.22(m，2H)，2.87-2.85(m，1H)，2.61-2.56(m，1H)，2.45-2.34(m，1H)，2.20-1.90(m，2H)，1.28(s，9H)，1.28-1.09(m，2H)；LCMS:95.48％，m/z[M+H]⁺＝592.2。

Example 273: synthesis of 1' - ((4, 6-bis (trifluoromethyl) pyridin-2-yl) (methyl) carbamoyl) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino-cyclo ] -2' -carboxylic acid (273a &273b):

synthesis 273.3:

to a stirred solution of 273.1(12.0g, 118mmol) in sulfolane (75mL) was added 273.2(24.4g, 58.4mmol) at room temperature. After stirring at 120 ℃ for 16h, the reaction mixture was cooled to room temperature, poured into ice-water and stirred well. The resulting precipitate was filtered, washed with ice water (2 × 20mL) and dried under vacuum to give 273.3(18g, 56%) as a white solid.

¹H NMR(500MHz，DMSO-d₆):13.03(s，1H)，8.00(s，1H)，7.83(s，1H)，7.62(s，1H)；LCMS:98.2％，m/z[M+H]⁺＝275.0。

Synthesis 273.4:

concentrated H was added to 273.3(36g, 131mmol) at room temperature₂SO₄₍80mL) and H₂O (54 mL). After stirring at 170 ℃ for 12h, the reaction mixture was cooled to room temperature, poured into ice-water and stirred well. The resulting precipitate was filtered, washed with ice water (2 × 50mL) and dried under vacuum to give 273.4 as a white solid (20g, 66%). LCMS 99.6%, M/z [ M + H ]]⁺＝232。

Synthesis 273.5:

to a stirred solution of 273.4(20g, 86.6mmol) in DCM (200mL) was added DIPEA (16mL, 95.2mmol) at 0 deg.C. After stirring for 20 min, trifluoromethanesulfonic anhydride (14.5mL, 86.6mmol) was added dropwise at 0 ℃. After stirring at room temperature for 10 minutes, the reaction mixture was quenched with water (50mL) and the organic layer was separated. The aqueous layer was extracted with DCM (50 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by column chromatography (silica gel 100-.

¹H NMR(500MHz，CDCl₃):7.98(s，1H)，7.64(s，1H)；¹⁹F NMR(470MHz，CDCl₃):-64.56，-68.26，-72.05。

Synthesis 273.7:

to a stirred solution of 273.5(10g, 27.5mmol) in THF (60mL) at 0 deg.C was added DIPEA (9.6mL, 55.1 mmol). After stirring for 30min, a solution of 273.6(4.15g, 30.3mmol) in THF (40mL) was added at room temperature. Stirred at room temperature for 16h, the reaction mixture was quenched with water (20ml) and extracted with ethyl acetate (2 × 60 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (80g silica gel column; 10% EtOAc/petroleum ether) to yield 273.7(8.5g, 88%) as a solid.

¹H NMR(500MHz，CDCl₃):7.29-7.26(m，2H)，7.09(s，1H)，6.90-6.87(m，2H)，6.71(s，1H)，4.50(d，J＝6.0Hz，2H)，3.81(s，3H)；¹⁹F NMR(470MHz，CDCl₃):-65.35。-69.07；LCMS:87.6％，m/z[M+H]⁺＝351.0。

Synthesis 273.8:

to a stirred solution of 273.7(1.0g, 2.85mmol) in DMF (15mL) at 0 deg.C was added dropwise NaHMDS (1M, 4.2mL, 4.28mmol) and MeI (0.18mL, 2.85 mmol). Stirred at room temperature for 2h, the reaction mixture was quenched with water (5mL) and extracted with ethyl acetate (2 × 15 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (24g silica gel column; 20% EtOAc/petroleum ether) to yield 273.8(0.85g, 85%) of a solid.

¹H NMR(400MHz，CDCl₃):7.21(d，J＝8.4Hz，2H)，7.05(s，1H)，6.87-6.85(m，2H)，6.80(s，1H)，4.79(s，2H)，3.79(s，3H)，3.11(s，3H)；LCMS:94.3％，m/z[M+H]⁺＝365.1。

Synthesis 273.9:

to a stirred solution of 273.8(0.700g, 1.92mmol) in DCM (5mL) was added TFA (3.5mL) at room temperature. After stirring at room temperature for one hour, the reaction mixture was diluted with DCM (20mL) and the organic phase was collected with saturated NaHCO₃(2 × 10mL) and water (10mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. This gave 273.9(0.35g, 74%) as a solid.

¹H NMR(500MHz，CDCl₃):7.08(s，1H)，6.71(s，1H)，5.07(br s，1H)，3.01(d，J＝5.5Hz，3H)。

Synthesis 273.10a &273.10 b:

thionyl chloride (5mL) was added at room temperature and 110.4_1a (0.500g, 1.08mmol) at RT was added. After stirring for 2 hours, the reaction mixture was concentrated under reduced pressure to yield the intermediate acid chloride. To this acid chloride is in CH₂Cl₂(5mL) to a solution of 273.9(0.45g, 1.87mmol) in CH was added₂Cl₂(5 mL). After stirring at room temperature for 16 hours, the reaction mixture was quenched with water (10ml). Separating the organic layer with CH₂Cl₂The organic phase was extracted (2 × 15 mL). The combined organic phases were washed with brine (10ml), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: x-bridge C18(150X30), 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/70, 8/80, 10/98, 14/98, 16/98, 16.1/70, 20/70, 23mL/min]This gave 273.10a as a white solid (peak-1, 35mg, 5%) and 273.10b as a white solid (peak-2, 25mg, 3%).

273.10a:¹H NMR(400MHz，DMSO-d₆):11.22(s，1H)，8.38(s，1H)，8.27(s，1H)，7.55(s，1H)，7.33(d，J＝2.0Hz，1H)，5.30-5.21(m，1H)，5.02-4.98(m，2H)，4.24-4.11(m，3H)，4.09-3.99(m，3H)，3.60(s，3H)，2.91-2.72(m，2H)，2.54-2.50(m，1H)；LCMS:98.9％，m/z[M+H]⁺687.2. The absolute stereochemistry was not determined.

273.10b:¹H NMR(400MHz，DMSO-d₆):11.20-11.10(br s，1H)，8.52(s，1H)，8.26(s，2H)，7.50(s，1H)，5.45-5.39(m，1H)，5.06-5.02(m，2H)，4.22-3.90(m，5H)，3.50(s，3H)，3.22-3.12(m，1H)，2.69-2.67(m，1H)，2.15-2.09(m，2H)；LCMS:97.1％，m/z[M+H]⁺687.2. The absolute stereochemistry was not determined.

Synthesis of 273 a:

273.10a (30mg, 0.043mmol) in THF (2mL) was stirred at room temperature to obtain a solutionAniline (4. mu.L, 0.043mmol) and Pd (PPh) were added to the solution₃)₄(9mg, 0.008 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMOSIL-C18(150x25mm), 10 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/20, 8/80, 10/80, 10.1/98, 12/98, 12.1/20, 14/20, 20mL/min]This gave 273a (14mg, 50%) solid.

¹H NMR(500MHz，DMSO-d₆):12.82(br s，1H)，11.15(br s，1H)，8.39(br s，1H)，8.28(s，1H)，7.55(s，1H)，7.27(br s，1H)，4.13-4.10(m，1H)，4.01-3.93(m，2H)，3.57(s，3H)，2.86-2.83(m，1H)，2.78-2.72(m，1H)，2.60-2.50(m，2H)。LCMS:99.3％，m/z[M+H]⁺647.0; the chiral purity is 97.1%.

Synthesis 273 b:

to a stirred solution of 273.10b (20mg, 0.029mmol) in THF (2mL) at room temperature was added aniline (2. mu.L, 0.029mmol) and Pd (PPh)₃)₄(6mg, 0.005 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMOSIL-C18(150x25mm), 10 μ; a is a 0.1% formic acid solution in water; b is acetonitrile; gradient: (time/% B) 0/20, 8/80, 10/80, 10.1/98, 11/98, 11.1/40, 13/40, flow rate 24mL/min]This gave 273b (5mg, 27%) as a solid. LCMS 94.1%, M/z [ M + H ]]⁺＝647.0。

Example 274: synthesis of (1'R, 2' S, 7a 'R) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -6', 6' -dimethyl-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid and (1'S, 2' R, 7a 'S) -5, 7-dichloro-1' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -6', 6' -dimethyl-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid (274.6a &274.6b):

composition 274.4_1&274.4_2

To a solution of 274.1(5g, 20.8mmol) in MTBE (50mL) was added 274.2(3.2g, 20.8mmol) and 274.3(4.4g, 20.8mmol) at room temperature, stirred at 80 ℃ for 2h and the reaction mixture cooled to RT. The precipitate was filtered and washed with ethyl acetate. The filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (silica gel 100-.

274.4_1:¹H NMR(500MHz，DMSO-d₆):12.53(s，1H)，10.96(s，1H)，7.78(d，J＝2.0Hz，1H)，7.46(d，J＝2.0Hz，1H)，5.51-5.45(m，1H)，5.10-5.05(m，2H)，4.27-4.26(m，2H)，4.17-4.10(m，1H)，4.01(d，J＝7.5Hz，1H)，3.55(t，J＝7.5Hz，1H)，2.57(d，J＝7.5Hz，1H)，1.98(d，J＝7.0Hz，1H)，1.63-1.61(m，1H)，1.47-1.45(m，1H)，1.01(s，3H)，0.97(s，3H)；LCMS:99.1％，m/z[M+H]⁺453.0; HPLC purity is 98.9%; chiral purity (49.8+ 50.1)%. The regional chemistry and the relative stereochemistry were confirmed by 2d NMR studies

274.4_2:¹H NMR(400MHz，DMSO-d₆):12.35(br s，1H)，10.90(s，1H)，7.55(d，J＝1.6Hz，1H)，7.50(d，J＝2.0Hz，1H)，5.87-5.80(m，1H)，5.27-5.14(m，2H)，4.45-4.37(m，3H)，3.74(d，J＝8.8Hz，1H)，3.46-3.42(m，1H)，2.51-2.48(m，1H)，2.08(d，J＝8.0Hz，1H)，1.92-1.87(m，1H)，1.60-1.55(m，1H)，1.02(s，6H)；LCMS:92.6％，m/z[M+H]⁺453.1. The regional chemistry and the relative stereochemistry were confirmed by 2d NMR studies

Separation 274.4_1a &274.4_1b

274.4_1(1.6g) was purified by chiral SFC using Lux Cellulose-4(30X250) mm, 5 μ; 70% CO₂30% isopropanol at room temperature (isocratic 100g/min, detection at 214 nm). HHh concentration of pure fractions under reduced pressure gave 274.4_1a (peak-1, 625mg, 78%) and 274.4_1b (peak-2, 650mg, 81%) solids as white solidsThe absolute stereochemistry of enantiomers 1 and 2 was not determined).

274.4_1a:¹H NMR(500MHz，DMSO-d₆):12.50(s，1H)，10.96(s，1H)，7.78(d，J＝2.0Hz，1H)，7.46(d，J＝2.0Hz，1H)，5.51-5.45(m，1H)，5.10-5.05(m，2H)，4.28-4.26(m，2H)，4.17-4.10(m，1H)，4.01(d，J＝8.0Hz，1H)，3.55(t，J＝7.5Hz，1H)，2.57(d，J＝7.5Hz，1H)，1.98(d，J＝7.5Hz，1H)，1.63-1.61(m，1H)，1.48-1.45(m，1H)，1.01(s，3H)，0.97(s，3H)；LCMS:99.2％，m/z[M+H]⁺453.1; the chiral purity is 99.7 percent.

274.4_1b:¹H NMR(500MHz，DMSO-d₆):12.53(s，1H)，10.96(s，1H)，7.79(d，J＝1.5Hz，1H)，7.46(d，J＝2.0Hz，1H)，5.49-5.46(m，1H)，5.11-5.05(m，2H)，4.28-4.27(m，2H)，4.14-4.13(m，1H)，4.00(d，J＝6.5Hz，1H)，3.55(t，J＝7.5Hz，1H)，2.57(d，J＝7.5Hz，1H)，1.97(d，J＝7.5Hz，1H)，1.64-1.61(m，1H)，1.48-1.44(m，1H)，1.01(s，3H)，0.97(s，3H)；LCMS:98.9％，m/z[M+H]⁺453.1; the chiral purity is 99.7 percent.

Synthesis 274.5 a:

thionyl chloride (6mL) was added to 274.4_1a (300mg, 0.66mmol) at room temperature. The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure to yield the intermediate acid chloride. To this acid chloride is in CH₂Cl₂(5mL) to a solution of 3, 5-dichloro-N-neopentylphenylamine (296mg, 1.27mmol) in CH was added₂Cl₂(5 mL). The reaction mixture was stirred at 55 ℃ for 16 hours and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 20% to 30% EtOAc/petroleum ether) to give 274.5a (230mg, 53%) as a solid.

¹H NMR(500MHz，DMSO-d₆) (in wheel form) 10.90/10.84(s, 1H), 8.26(s, 1H), 7.63(s, 1H), 7.44-7.39(m, 3H), 5.48-5.39(m, 1H), 5.09-5.05(m, 2H), 4.27-4.13(m, 3H), 3.84-3.68(m, 3H), 3.54(t, J ═ 7.5Hz, 1H), 2.68(d, J ═ 7.0Hz, 1H), 1.86(d, J ═ 7.0Hz, 1H), 1.0 (d, J ═ 7.0Hz, 1H)H)，1.54-1.49(m，2H)，0.97(s，6H)，0.84/0.82(s，9H)；LCMS:98.8％，m/z[M+H]⁺＝668.0。

Synthesis 274.6 a:

to a stirred solution of 274.5a (230mg, 0.34mmol) in THF (10mL) at room temperature was added aniline (48mg, 0.68mmol) and Pd (PPh)₃)₄(80mg, 0.06 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: X-bridge-C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/70, 8/90, 10/70, 10.1/98, 12/98, 12.1/70, 15/70at 20mL/min to yield 274.6a (118mg, 55%) of a solid.

¹H NMR(500MHz，DMSO-d₆) 12.40(br s, 1H), 10.85/10.78(s, 1H), 8.41/7.91(d, J ═ 2.0Hz, 1H), 7.61/7.52(t, J ═ 2.0Hz, 1H), 7.47-7.39(m, 3H), 4.07(d, J ═ 8.0Hz, 1H), 3.94(d, J ═ 14.0Hz, 1H), 3.84-3.78(m, 1H), 3.51(d, J ═ 14.0Hz, 1H), 3.45(t, J ═ 7.0Hz, 1H), 2.68(d, J ═ 7.0Hz, 1H), 1.82(d, J ═ 7.0Hz, 1H), 1.50-1.41(m, 2H), 2.96 (s, 0H), 6.82 (s, 0H), 9H, 84(s, 0H); LCMS 99.3%, M/z [ M + H ]]⁺626.0; the chiral purity is 99.8 percent.

Synthesis 274.5 b:

thionyl chloride (5mL) was added to 274.4_1b (100mg, 0.22mmol) at room temperature. The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure to yield the acid chloride intermediate. To this acid chloride is in CH₂Cl₂(5mL) to a solution of 3, 5-dichloro-N-neopentylphenylamine (98mg, 0.42mmol) in CH was added₂Cl₂(5 mL). The reaction mixture was stirred at 55 ℃ for 16 hours and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 20% to 30% EtOAc/petroleum ether) to give 274.5b as a white solid (100mg, 68%).

¹H NMR(400MHz，DMSO-d₆) (in wheel form) 10.90/10.84(s, 1H), 8.26/7.78(d，J＝2.0Hz，1H)，7.63/7.54(t，J＝2.0Hz，1H)，7.44-7.39(m，3H)，5.48-5.39(m，1H)，5.09-5.04(m，2H)，4.30-4.12(m，3H)，3.85-3.67(m，3H)，3.54(t，J＝7.6Hz，1H)，2.68-2.66(m，1H)，1.86(d，J＝6.8Hz，1H)，1.54-1.49(m，2H)，0.97(s，6H)，0.84/0.81(s，9H)；LCMS:96.0％，m/z[M+H]⁺＝667.9。

Synthesis 274.6 b:

to a stirred solution of 274.5b (100mg, 0.15mmol) in THF (5mL) at room temperature were added aniline (17mg, 0.18mmol) and Pd (PPh)₃)₄(34.6mg, 0.03 mmol). After stirring at room temperature for 3 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMOSIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/70, 8/95, 10/98, 12/98, 12.1/70, 14/70, 25mL/min]Thereby yielding 274.6b (65mg, 69%) as a solid.

¹H NMR(400MHz，DMSO-d₆) 12.38(br s, 1H), 10.80/10.77(s, 1H), 8.41/7.92(d, J ═ 2.0Hz, 1H), 7.60/7.52(t, J ═ 1.6Hz, 1H), 7.47-7.38(m, 3H), 4.06(d, J ═ 8.0Hz, 1H), 3.93(d, J ═ 14.0Hz, 1H), 3.84-3.77(m, 1H), 3.52(d, J ═ 14.0Hz, 1H), 3.45(t, J ═ 7.2Hz, 1H), 2.68(d, J ═ 6.8Hz, 1H), 1.82(d, J ═ 6.8Hz, 1H), 1.48-1.43(m, 2H), 2.96 (s, 0.84, 0H), 9.82(s, 0H); LCMS 99.0%, M/z [ M + H ]]⁺626.0; the chiral purity is 99.4%.

Example 275: synthesis of (5'R, 6' S, 7'R, 7a' R) -5', 7' -dichloro-7 '- ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -2' -oxo-1 ', 6', 7', 7a' -tetrahydro-3 'H-disuccincyclo [ cyclopropane-1, 2' -pyrazino-ring-5 ', 3' -indole ] -6 '-carboxylic acid and (5' S, 6'R, 7' S, 7a 'S) -5', 7 '-dichloro-7' - ((3, 5-dichlorophenyl) (neopentyl) carbamoyl) -2 '-oxo-1', 6', 7', 7a '-tetrahydro-3' H-disuccino [ cyclopropane-1, 2' -pyrazino-ring-5 ', 3 "-indole ] -6' -carboxylic acid (275.7a &275.7 b):

synthesis 275.2:

to a stirred solution of 275.1(10g, 41.4mmol) in DCM (100mL) was added TFA (16mL, 207mmol) at 0 ℃. The reaction mixture was stirred at room temperature for 16 hours and concentrated under reduced pressure. The resulting residue was triturated with diethyl ether (200mL) to give 275.2(7.4g, 75%) as a solid.

¹H NMR(400MHz，DMSO-d₆):9.47(br s，1H)，4.43(dd，J＝8.4Hz，J＝6.8Hz，1H)，3.16(d，J＝11.2Hz，2H)，3.12(d，J＝11.2Hz，2H)，2.24-2.18(m，1H)，2.03-1.98(m，1H)，0.68-0.61(m，4H)；LCMS:87.6％，m/z[M+H]⁺＝142.2。

Synthesis 275.5_1&275.5_ 2:

to a solution of 275.2(4.2g, 17.6mmol) in MTBE (60mL) was added 275.3(2.75g, 17.6mmol) and 275.4(3.81g, 17.6mmol) at room temperature. After stirring at 80 ℃ for 2 hours, the reaction mixture was cooled to room temperature. The resulting precipitate was filtered and washed with ethyl acetate. The filtrate was concentrated under reduced pressure. The resulting residue was purified by column chromatography (silica gel 100. sup. 200. sup. mesh, 10% -30% EtOAc/petroleum ether). The major diastereomer was triturated with DCM (50mL) to give 275.5 — 1 (racemate, 3.0g, 38%) as a white solid.

¹H NMR(500MHz，DMSO-d6):12.60(br s，1H)，11.03(s，1H)，7.74(d，J＝2.0Hz，1H)，7.46(d，J＝2.0Hz，1H)，5.51-5.48(m，1H)，5.11-5.06(m，2H)，4.28(d，J＝5.5Hz，2H)，4.22-4.16(m，1H)，4.04(d，J＝7.5Hz，1H)，3.63-3.58(m，1H)，2.75(d，J＝7.5Hz，1H)，2.04(d，J＝8.0Hz，1H)，1.93-1.85(m，1H)，1.52-1.49(m，1H)，0.50-0.36(m，4H)；LCMS:96.8％，m/z[M+H]⁺451.1; chiral purity (49.9% + 50.0%). regional and relative stereochemistry was determined by 2DNMR analysis.

275.5_2:¹H NMR(400MHz，DMSO-d₆):12.45(br s，1H)，10.98(s，1H)，7.58(d，J＝1.6Hz，1H)，7.51(d，J＝2.0Hz，1H)，5.89-5.79(m，1H)，5.28-5.15(m，2H)，4.51-4.37(m，3H)，3.65-3.57(m，2H)，2.72(d，J＝8.8Hz，1H)，2.04-1.99(m，2H)，1.77-1.72(m，1H)，0.65-0.35(m，4H)；LCMS:91.9％，m/z[M+H]⁺451.0. Regional and relative stereochemistry were determined by 2D NMR analysis.

Separation 275.5_1a &275.5_1b

275.5 — 1(3g) was purified by chiral SFC using chiral package AD-H (30X250) mm, 5 μ; a is 80% CO₂% B20% of HHh0.5% TFA in isopropanol at room temperature (isocratic 100g/min, detection at 214 nm). Concentration of the pure fractions under reduced pressure yielded 275.5_1a (peak-1, 1.2g, 80%) solids and 275.5_1b (peak-2, 1.4g, 93%) solids. The absolute stereochemistry of enantiomers 1 and 2 was not determined.

275.5_1a:¹H NMR(500MHz，DMSO-d₆):12.60(br s，1H)，11.10(s，1H)，7.71(d，J＝1.5Hz，1H)，7.49(d，J＝2.0Hz，1H)，5.53-5.49(m，1H)，5.13-5.07(m，2H)，4.31(d，J＝5.5Hz，2H)，4.26-4.24(m，1H)，4.05(d，J＝7.5Hz，1H)，3.69-3.63(m，1H)，2.88-2.82(m，1H)，2.09(d，J＝8.0Hz，1H)，1.97-1.90(m，1H)，1.55-1.52(m，1H)，0.52-0.36(m，4H)；LCMS:98.3％，m/z[M+H]⁺451.0; the chiral purity is 99.1 percent.

275.5_1b:¹H NMR(500MHz，DMSO-d₆):12.65(br s，1H)，11.10(s，1H)，7.71(s，1H)，7.49(s，1H)，5.55-5.48(m，1H)，5.12-5.07(m，2H)，4.31(d，J＝5.5Hz，2H)，4.27-4.23(m，1H)，4.05(d，J＝8.0Hz，1H)，3.69-3.66(m，1H)，2.85(d，J＝7.0Hz，1H)，2.09(d，J＝8.0Hz，1H)，1.97-1.93(m，1H)，1.55-1.52(m，1H)，0.51-0.37(m，4H)；LCMS:98.0％，m/z[M+H]⁺451.0; the chiral purity is 95.4 percent;

synthesis 275.6 a:

thionyl chloride (5mL) was added to 275.5 — 1a (300mg, 0.66mmol) at room temperature and after stirring for 2 hours, the reaction mixture was concentrated under reduced pressure to give the acid chloride intermediate. To this acid chlorine compound in CH₂Cl₂(5mL) to the solution was added 3, 5-dichloro-N-neopentylbenzeneAmine (296mg, 1.27mmol) in CH₂Cl₂(5 mL). The reaction mixture was stirred at 55 ℃ for 16 hours and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 20% to 30% EtOAc/petroleum ether) to yield 275.6a (300mg, 71%) of a solid.

¹H NMR(400MHz，DMSO-d₆) 10.91/10.98(s, 1H), 8.21/7.71(d, J ═ 1.6Hz, 1H), 7.63/7.54(t, J ═ 1.6Hz, 1H), 7.43-7.39(m, 3H), 5.67-5.46(m, 1H), 5.23-5.08(m, 2H), 4.34-4.19(m, 3H), 4.14(d, J ═ 8.0Hz, 1H), 3.89-3.80(m, 2H), 3.68-3.60(m, 2H), 2.80(d, J ═ 7.6Hz, 1H), 1.93-1.88(m, 1H), 1.43-1.39(m, 1H), 0.83/0.81(s, 9H), 0.47-4H (m, 47H); LCMS 98.2%, M/z [ M + H ]]⁺＝665.9。

Synthesis 275.7 a:

to a stirred solution of 275.6a (300mg, 0.44mmol) in THF (10mL) at room temperature were added aniline (50mg, 0.53mmol) and Pd (PPh)₃)₄(104mg, 0.08 mmol). The reaction mixture was stirred at room temperature for 2 hours and concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMOSIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/70, 8/95, 10/98, 12/98, 12.1/70, 14/70at 25mL/min]This gave 275.7a (113mg, 40%) of a solid.

¹H NMR(400MHz，DMSO-d₆) 12.43/12.30(br s, 1H), 10.90/10.85(s, 1H), 8.37/7.87(d, J ═ 2.0Hz, 1H), 7.61/7.53(t, J ═ 2.0Hz, 1H), 7.48(d, J ═ 1.6Hz, 2H), 7.42/7.39(d, J ═ 2.0Hz, 1H), 4.07(d, J ═ 8.0Hz, 1H), 4.00(d, J ═ 14.0Hz, 1H), 3.88-3.86(m, 1H), 3.55(t, J ═ 7.2Hz, 1H), 3.44(d, J ═ 14.0, 1H), 2.87(d, J ═ 7.2, 1H), 1H, 2.82 (t, 1H), 1H, 4.82 (d, J ═ 14.0, 1H), 2.87(d, J ═ 7.2, 1H), 1H), 4.82 (m, 0, 33.82 (m, 0H); LCMS 99.0%, M/z [ M + H ]]⁺624.0; the chiral purity is 98.3 percent.

Synthesis 275.6 b:

sulfur dioxide at room temperatureAcid chloride (5mL) was added to 275.5_1b (300mg, 0.66 mmol). After stirring for 2 hours, the reaction mixture was concentrated under reduced pressure to yield the acid chloride intermediate. To this acid chloride is in CH₂Cl₂(5mL) to a solution of 3, 5-dichloro-N-neopentylphenylamine (296mg, 1.27mmol) in CH was added₂Cl₂(5 mL). The reaction mixture was stirred at 55 ℃ for 16 hours and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 20% to 30% EtOAc/petroleum ether) to give 275.6b (200mg, 45%) of a solid.

LCMS:92.9％，m/z[M+H]⁺＝666.1。

Synthesis 275.7 b:

to a stirred solution of 275.6b (200mg, 0.30mmol) in THF (10mL) at room temperature was added aniline (56mg, 0.60mmol) and Pd (PPh)₃)₄(104mg, 0.08 mmol). After stirring at room temperature for three hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ column: KROMOSIL-C18(150X25) mm, 10. mu.l, A: 0.1% formic acid in water, B: acetonitrile; gradient: (time/% B) 0/70, 8/95, 10/98, 12/98, 12.1/70, 14/70, 25mL/min]This gave 275.7b (30mg, 16%) as a solid.

¹H NMR(400MHz，DMSO-d₆) 12.42/12.30(br s, 1H), 10.90/10.85(s, 1H), 8.37/7.86(d, J ═ 2.0Hz, 1H), 7.61/7.53(t, J ═ 2.0Hz, 1H), 7.48(d, J ═ 1.6Hz, 2H), 7.42/7.39(d, J ═ 2.0Hz, 1H), 4.07(d, J ═ 7.6Hz, 1H), 4.00(d, J ═ 14.0Hz, 1H), 3.88-3.86(m, 1H), 3.55(t, J ═ 7.2Hz, 1H), 3.44(d, J ═ 14.0, 1H), 2.87(d, J ═ 7.6, 1H), 1H, 89.82 (t, 1H), 1H, 9.82 (m, 32H), 2H, 1H, 4.32H, 2.82 (m, 2H); LCMS 95.8%, M/z [ M + H ]]⁺624.0; the chiral purity is 95.8 percent.

Example 280: synthesis of (1'R, 2' R, 7a 'R) -5, 7-dichloro-1' - ((cyclopropylmethyl) (3, 5-dichlorophenyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid

280 was synthesized from 280.1b as described in synthesis 110. The absolute stereochemistry is unknown.

¹H NMR(500MHz，DMSO-d₆) 12.66(br s, 1H), 10.91(br s, 1H), 7.92-7.38(m, 5H), 4.62-4.55(m, 1H), 3.67-3.59(m, 1H), 3.48-3.38(m, 2H), 3.20-3.07(m, 2H), 2.74-2.64(m, 1H), 2.50-2.41(m, 1H), 2.30-2.15(m, 1H), 0.90-0.80(m, 1H), 0.38-0.36(m, 2H), 0.06-0.04(m, 2H); LCMS 92.6%, M/z [ M + H ]]⁺618.0; the chiral purity is 95.1%.

Example 281: synthesis of (1'S, 2' S, 7a 'S) -5, 7-dichloro-1' - ((cyclopropylmethyl) (3, 5-dichlorophenyl) carbamoyl) -6', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a' -hexahydrospiro [ indole-3, 3 '-pyrazino ] -2' -carboxylic acid

281 is synthesized from 110.4_2 as described in synthesis 111. The absolute stereochemistry is unknown.

¹H NMR(500MHz，DMSO-d₆) 12.66(br s, 1H), 10.91(br s, 1H), 7.92-7.38(m, 5H), 4.61-4.53(m, 1H), 3.68-3.58(m, 1H), 3.49-3.39(m, 2H), 3.22-3.07(m, 2H), 2.72-2.65(m, 1H), 2.50-2.40(m, 1H), 2.30-2.15(m, 1H), 0.90-0.81(m, 1H), 0.38-0.36(m, 2H), 0.06-0.04(m, 2H); LCMS 90.3%, M/z [ M + H ]]⁺＝618.0。

Example 282: synthesis of methyl (1' R, 2' S, 7a ' R) -5, 7-dichloro-1 ' - ((3, 5-dichlorophenyl) (methyl) carbamoyl) -6', 6' -difluoro-2 ' -methyl-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyridoxine ] -2' -carboxylate

Synthesis 282.4:

to a stirred solution of 282.1(2.5g, 22.3mmol) in THF (25mL) was added 282.2(5.5g, 22.3mmol) and 282.3(4.8g, 22.3mmol) at room temperature. After 8 hours stirring at room temperature, the resulting reaction mixture was evaporated under reduced pressure to give 282.4(8.5g) as a brown solid which was used in the next step without purification. LCMS 16%, M/z [ M + H ]]⁺＝416.9。

Synthesis 282.5a and 282.5 b:

a solution of 282.4(8.5g, 20.4mmol) in allyl alcohol (50mL) was heated at 90 ℃ for 16 h. The reaction mixture was concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (120g silica gel column, 30% EtOAc/petroleum ether) to yield a mixture of regioisomers. Purification of the regioisomers by preparative HPLC gave 282.5a (160mg, 2%) and 282.5b (1.1g, 11%) as off-white solids. 2D NMR analysis to determine regional chemistry and related stereochemistry.

282.5a data:¹H NMR(400MHz，MeOH-d₄):7.89(d，J＝1.6Hz，1H)，7.29(d，J＝1.6Hz，1H)，5.56-5.47(m，1H)，5.11-5.05(m，2H)，4.35-4.24(m，2H)，3.82-3.78(m，1H)，3.74(s，1H)，3.07-2.98(m，1H)，2.73-2.64(m，1H)，2.38-2.32(m，1H)，2.19-2.10(m，1H)，1.59(s，3H)；LCMS:95.5％，m/z[M+H]⁺＝475.0。

282.5b:¹H NMR(500MHz，DMSO-d₆):13.40(br s，1H)，11.10(br s，1H)，7.61(s，1H)，7.22(s，1H)，5.95-5.87(m，1H)，5.34(dd，J＝17.5Hz，J＝1.5Hz，1H)，5.23(dd，J＝10.5Hz，J＝1.5Hz，1H)，4.62-4.54(m，2H)，4.45-4.42(m，1H)，4.02(d，J＝10.5Hz，1H)，3.88-3.72(m，1H)，3.20-2.98(m，1H)，2.80-2.75(m，1H)，2.31-2.26(m，1H)，1.30(s，3H)；LCMS:91.5％，m/z[M+H]⁺＝475.0。

Synthesis of 282.6 b:

to a stirred solution of 282.5b (400mg, 0.84mmol) in methanol (8mL) at 0 deg.C was added (trimethylsilane) diazomethane (2M in hexane, 2.1mL, 4.2 mmol). After stirring at room temperature for 16 hours, the reaction mixture was cooled to 0 ℃ and extracted with acetic acid (0.5 mL). The reaction mixture was stirred at room temperature for one hour and concentrated at reduced pressure at low temperature. The resulting residue was purified by flash column chromatography (24g silica gel column, 10% to 25% EtOAc in petroleum ether) to afford 282.6b as an off-white solid (260mg, 63%).

¹H NMR(400MHz，DMSO-d₆):11.31(s，1H)，7.67(d，J＝2.0Hz，1H)，6.91(d，J＝2.0Hz，1H)，5.94-5.86(m，1H)，5.37-5.25(m，2H)，4.59(d，J＝4.8Hz，2H)，4.54-4.47(m，1H)，4.11(d，J＝10.4Hz，1H)，3.77-3.69(m，1H)，3.64(s，3H)，2.91-2.81(m，2H)，2.40-2.33(m，1H)，1.24(s，3H)；LCMS:91.0％，m/z[M-H]^-＝486.9。

Synthesis 282.7 b:

to a stirred solution of 282.6b (240mg, 0.49mmol) in THF (5mL) at room temperature was added aniline (46mg, 0.49mmol) and Pd (PPh)₃)₄(114mg, 0.09 mmol). After stirring at room temperature for 4 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by column chromatography (silica gel 100-.

¹H NMR(400MHz，DMSO-d₆):12.90(br s，1H)，11.28(s，1H)，7.66(d，J＝1.6Hz，1H)，6.90(d，J＝2.0Hz，1H)，4.49-4.42(m，1H)，4.06-3.97(m，2H)，3.82-3.71(m，1H)，3.64(s，3H)，2.97-2.84(m，1H)，2.39-2.30(m，1H)，1.19(s，3H)；LCMS:95.1％，m/z[M+H]⁺＝450.4。

Synthesis of 282:

thionyl chloride (1mL) was added to 282.7b (70mg, 0.15mmol) at room temperature and after stirring for 2 hours, the reaction mixture was concentrated under reduced pressure to give the intermediate acid chloride. To this acid chloride is in CH₂Cl₂(2mL) to a solution of 3, 5-dichloro-N-methylaniline (38mg, 0.21mmol) in CH was added₂Cl₂(3mL) and a catalytic amount of DMAP (2 mg). The reaction mixture was stirred at 55 ℃ for 16 hours and concentrated under reduced pressure. The residue was purified by flash column chromatography (12g silica gel column, 15% to 20% EtOAc/petroleum ether) to give 282(50mg, 57%) as an off-white solid.

¹H NMR(500MHz，DMSO-d₆) 11.31/11.13(s, 1H), 7.77-7.44(m, 4H), 6.99-6.91(m, 1H), 4.40/4.33(d, J ═ 10.0Hz, 1H), 3.96-3.72(m, 1H), 3.65/3.58(s, 3H), 3.46-3.39(m, 1H), 3.34/3.17(s, 3H), 2.88-2.77(m, 2H), 2.17-2.07(m, 1H), 1.10/1.04(s, 3H); LCMS 94.5%, M/z [ M + H ]]⁺＝605.9。

Example 284: synthesis of (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N1 '-neopentyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide:

to a stirred solution of 110(300mg, 0.47mmol) in DMF (6mL) at room temperature was added HATU (360mg, 0.94mmol) and DIPEA (0.51mL, 2.83 mmol). After stirring for 10 minutes, NH was added₄Cl (128mg, 2.36 mmol). After stirring at room temperature for 16 hours, the reaction mixture was diluted with ice water (50mL) and stirred for 10 minutes. The resulting precipitate was filtered, washed with water and dried under vacuum. The crude product was purified by preparative chromatography [ KROMOSIL-C18(150X25) mm, 10. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/55, 8/80, 8.1/98, 10/98, 10.1/55, 13/55, 20mL/min]This gave 284 as an off-white solid (70mg, 23%).

¹H NMR(400MHz，DMSO-d₆) (in wheel form) 10.80/10.70(s, 1H), 7.70(t, J ═ 1.6Hz, 1H), 7.67(d, J ═ 1.6Hz, 1H), 7.61-7.54(m, 2H), 7.49(d, J ═ 1.6Hz, 1H), 6.95(br s, 1H), 6.42(br s, 1H), 4.33-4.27(m, 1H), 3.92(d, J ═ 11.6Hz, 1H), 3.78(d, J ═ 14.0H), and so forthz，1H)，3.68-3.58(m，2H)，3.45-3.35(m，1H)，2.81-2.76(m，1H)，2.50-2.44(m，1H)，2.28-2.13(m，1H)，0.81(s，9H)；LCMS:99.1％，m/z[M+H]⁺633.0; chiral purity 94.7%.

Example 285: synthesis of (1'R, 2' S, 3R, 7a 'R) -5, 7-dichloro-N1' - (3, 5-dichlorophenyl) -6', 6' -difluoro-N2 '-hydroxy-N2' -methyl-N1 '-neopentyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino ] -1', 2' -dicarboxamide

To a stirred solution of 110(400mg, 0.62mmol) in DMF (5mL) at room temperature was added EDC.HCl (240mg, 1.25mmol), HOBt (255mg, 1.88mmol) and Et₃N (0.88mL, 6.29 mmol). After stirring for 5 min, (Me) NHOH. HCl (420mg, 5.03mmol) was added. After stirring at room temperature for 16 hours, the reaction mixture was diluted with ice water (50mL) and stirred for 10 minutes. The resulting precipitate was filtered, washed with water and dried under vacuum. Preparative HPLC [ X-SELECT-C18(250X19) mm, 5. mu.l; a is a solution of 10mM ammonium bicarbonate in water, B is acetonitrile; gradient: (time/% B) 0/65, 8/80, 10/80, 10.1/98, 12/98, 12.1/65, 15/65at 20mL/min]The crude compound was purified to give 285(53mg, 13%) as an off-white solid.

¹H NMR(500MHz，DMSO-d₆):10.77(s，1H)，10.10(s，1H)，7.67(s，1H)，7.59-7.44(m，3H)，7.28(s，1H)，4.40(d，J＝8.0Hz，1H)，4.15-4.08(m，1H)，3.78(d，J＝14.0Hz，1H)，3.63(d，J＝14.0Hz，1H)，3.52(t，J＝9.0Hz，1H)，3.44-3.40(m，1H)，2.98(s，3H)，2.71-2.66(m，1H)，2.64-2.55(m，1H)，2.21-2.10(m，1H)，0.82(s，9H)；LCMS:97.2％，m/z[M+H]⁺662.9; chiral purity 93.2%.

Example 286: synthesis of 5, 7-dichloro-N1 '- (3, 5-dichlorophenyl) -6', 6 '-difluoro-N2' -hydroxy-N1 '-neopentyl-2-oxo-1', 2', 5', 6', 7', 7a '-hexahydrospiro [ indole-3, 3' -pyrazino-cyclo ] -1', 2' -dicarboxamide (286a &286b)

To a stirred solution of 110(400mg, 0.62mmol) in DMF (5mL) at room temperature were added HATU (360mg, 0.94mmol) and DIPEA (1.2mL, 6.29mmol), and after stirring for 10min, NH was added₂OH.HCl (352mg, 5.03 mmol). After stirring at room temperature for 16 hours, the reaction mixture was diluted with ice water (50mL) and stirred for 15 minutes. The resulting precipitate was filtered, washed with water (10mL) and dried under vacuum. The resulting residue was purified by preparative high performance liquid chromatography [ X-SELECT-C18(150X30) mm, 5. mu.l; a is a solution of 10mM ammonium bicarbonate in water, B is acetonitrile; gradient: (time/% B) 0/60, 8/90, 11/90, 11.1/98, 15/98, 15.1/60, 19/60, 22mL/min]286a as an off-white solid (Peak-1, 37mg, 9%) and 286b as an off-white solid (Peak-2, 38mg, 9%) were produced.

286a:¹H NMR(500MHz，DMSO-d₆):10.74(br s，1H)，9.96(br s，1H)，8.83(s，1H)，7.71-7.52(m，5H)，4.32(t，J＝10.5Hz，1H)，3.80-3.75(m，1H)，3.71-3.63(m，3H)，3.38-3.32(m，1H)，2.87-2.83(m，1H)，2.64-2.50(m，1H)，2.23-2.16(m，1H)，0.80(s，9H)；LCMS:97.0％，m/z[M+H]⁺648.9; the chiral purity is 99.8 percent.

286b:LCMS:95.9％，m/z[M+H]⁺＝649.0。

Example 287: synthesis of 1' - (((3s, 5s, 7s) -tricyclodecan-1-yl) (methyl) carbamoyl) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino ] -2' -carboxylic acid

Synthesis 287.1_1 and 287.1_2:

thionyl chloride (3mL) was added to 110.4_1a (300mg, 0.65mmol) at room temperature and after stirring for 2h, the reaction mixture was concentrated under reduced pressure to give the intermediate acid chloride. To the acid chloride prepared above in CH₂Cl₂(5mL) to the solution was added (3s, 5s, 7s) -N-methyltricyclodecan-1-amine (161mg, 0.97mmol) in CH₂Cl₂(5 mL). Stirred at 50 ℃ for 16h, quenched with water (10ml) and CH₂Cl₂(2x15 mL). The combined organic phases were washed with brine, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 20% -25% EtOAc/petroleum ether) to give 287.1_1 (peak-1, 105mg, 26%) as a light brown solid and 287.1_2 (peak-2, 84mg, 21%) as a light brown solid.

287.1_1:LCMS:24.6％，m/z[M+H]⁺＝608.1。

287.1_2:LCMS:81.9％，m/z[M+H]⁺＝608.1。

Synthesis 287:

to a stirred solution of 287.1-1 (105mg, 0.17mmol) in THF (5mL) at room temperature was added aniline (16mg, 0.17mmol) and Pd (PPh)₃)₄(39mg, 0.03 mmol). After stirring for 2 hours, the reaction mixture was concentrated under reduced pressure. The resulting residue was purified by preparative high performance liquid chromatography [ X-bridge-C18(150X30) mm, 5. mu.l; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/65, 8/80, 8.1/98, 10/98, 10.1/98, 13/65, 22mL/min]This gave 287 as a solid (16mg, 15%).

¹H NMR(500MHz，DMSO-d₆):12.40(br s，1H)，11.06(br s，1H)，7.68(s，1H)，7.52(s，1H)，4.13(d，J＝11.0Hz，1H)，3.89-3.85(m，1H)，3.72-3.67(m，1H)，3.06(s，3H)，2.86-2.64(m，3H)，2.50-2.47(m，1H)，2.13-2.04(m，9H)，1.65-1.60(m，6H)；LCMS:94.1％，m/z[M+H]⁺568.1; the chiral purity is 90.5%.

Example 288: synthesis of 1' - ((2- (3- (tert-butyl) phenyl) -2-methylpropyl) carbamoyl) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3, 3' -pyrazino-cyclo ] -2' -carboxylic acid (288a) & rac- (1' R, 2' S, 3R, 7a ' R) -1' - ((2- (3- (tert-butyl) phenyl) -2-methylpropyl) carbamoyl) -5, 7-dichloro-6 ', 6' -difluoro-2-oxo-1 ', 2', 5', 6', 7', 7a ' -hexahydrospiro [ indole-3 ], 3 '-Pyrroline ring ] -2' -carboxylic acid (288b)

Synthesis 288.2:

288.1(5g, 33.2mmol) in CH at 0 deg.C₂Cl₂To a solution in (60mL) was added pyridine (2.9mL, 36.5mmol) and trifluoromethanesulfonic anhydride (6.1mL, 36.5 mmol). After stirring at room temperature for 16 hours, the reaction mixture was extracted with ice water, the organic layer was separated and washed with water (60mL), brine (60mL), dried over sodium sulfate, and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (40g silica gel column, 10% ethyl acetate in petroleum ether) to yield 288.2(3.9g, 42%) as a colorless oily solid.

¹H NMR(500MHz，CDCl₃):7.42-7.35(m，2H)，7.26-7.25(m，1H)，7.10-7.07(m，1H)，1.33(s，9H)。

Synthesis 288.4:

to a stirred solution of 288.3(3g, 21.2mmol) in EtOH (40mL) and water (401mg, 22.2mmol) was added potassium tert-butoxide (2.4g, 21.2mmol) in EtOH (20mL) at room temperature over 30min at 60 ℃. After stirring for 2 hours at 60 ℃, the reaction mixture was concentrated under reduced pressure. The resulting residue was triturated with diethyl ether and EtOH to give 288.4(1.8g, 56%) as a solid.

¹H NMR(500MHz，DMSO-d₆):1.29(s，6H)。

Synthesis 288.5:

to a suspension of 288.2(2g, 7.08mmol), 288.4(1.28g, 8.5mmol) and 4, 5-bis diphenylphosphine-9, 9-dimethylxanthene (205mg, 0.35mmol) in trimethylbenzene (20mL) was flushed argon for 15 minutes, followed by the addition of bis (allyl) dichloropalladium (78mg, 0.21mmol) and continued flushing with argon for five minutes, after stirring at 130 ℃ for 6 hours, the reaction mixture was cooled to room temperature and poured into ice water (30 mL). The organic layer was separated, washed with water (20mL), brine (20mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (80g silica gel column, 10% ethyl acetate in petroleum ether to give 288.5 as a colorless oil (1.2g, 72%).

¹H NMR(500MHz，CDCl₃):7.51(d，J＝1.5Hz，1H)，7.36-7.25(m，3H)，1.73(s，6H)，1.34(s，9H)；GCMS:99.4％，m/z[M+H]⁺＝202.2。

Synthesis 288.6:

to a stirred solution of 288.5(1g, 4.96mmol) in THF (15mL) at 0 deg.C was added LiAlH₄Solution (1M in THF, 7.5mL, 7.45 mmol). After stirring at 0 ℃ for 4 hours, the reaction mixture was annealed with 10% NaOH solution, extracted with ethyl acetate (20mL) at 0 ℃. The organic layer was separated, washed with water (20mL), brine (20mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The residue was triturated with n-pentane (10mL) to give 288.6(850mg, 84%) as a solid.

¹H NMR(500MHz，DMSO-d₆):7.34(d，J＝2.0Hz，1H)，7.34-7.13(m，3H)，3.62-3.59(m，1H)，2.67(s，2H)，1.78-1.76(m，1H)，1.28(s，9H)，1.23(s，6H)；LCMS:92.9％，m/z[M+H]⁺＝206.2。

Synthesis 288.7:

to a stirred solution of 110.4_1(300mg, 0.65mmol) in DMF (3mL) at room temperature were added DIPEA (252mg, 1.95mmol) and HATU (297mg, 0.77 mmol). After stirring for 15 min at room temperature 288.6(200mg, 0.97mmol) was added. After stirring at room temperature for 6 hours, the reaction mixture was quenched with water (10mL) and extracted with ethyl acetate (10 mL.) the combined organic phases were washed with water (20mL), brine (20mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. This gave 288.7(500mg) as a brown gum as a diastereoisomeric mixture. The residue obtained was used in the next step without purification.

Synthesis 288a &288 b:

288.7(500mg, 0.77mmol) in THF (20mL) at room temperatureAniline (70mg, 0.77mmol) and Pd (PPh) were added to the stirred solution₃)₄(180mg, 0.2 mmol). After stirring at room temperature for 2 hours, the reaction mixture was concentrated under reduced pressure and triturated with diethyl ether and pentane. The resulting residue was purified by preparative high performance liquid chromatography [ X-bridge C18(150X30) mm, 5 μ; a is 0.1% formic acid solution in water, B is acetonitrile; gradient: (time/% B) 0/65, 9/80, 11/80, 11.1/65, 14/65at 25mL/min]This gave a minor diastereomer 288a (20mg, 5%) as a solid and a major diastereomer 288b (57mg, 14%) as a solid.

288a:¹H NMR(500MHz，DMSO-d₆):12.50(br s，1H)，11.07(br s，1H)，7.54-7.39(m，2H)，7.31(s，1H)，7.24-7.17(m，3H)，7.10-7.07(m，1H)，4.15-4.05(m，2H)，3.57-3.47(m，2H)，3.32-3.20(m，2H)，2.76-2.64(m，2H)，2.31-2.24(m，1H)，1.39-1.30(m，9H)，1.28-1.26(m，6H)；LCMS:98.5％，m/z[M+H]⁺608.3. (absolute stereochemistry undetermined)

288b:¹H NMR(500MHz，DMSO-d₆):12.35(br s，1H)，10.93(br s，1H)，8.19(br s，1H)，7.88(br s，1H)，7.42(d，J＝1.5Hz，2H)，7.23-7.19(m，3H)，3.92-3.86(m，2H)，3.63-3.59(m，1H)，3.47-3.40(m，1H)，3.16-3.01(m，2H)，2.56-2.50(m，1H)，1.94-1.87(m，1H)，1.71-1.58(m，1H)，1.33-1.26(m，15H)；LCMS:98.5％，m/z[M+H]⁺＝608.3。

Table 8:

the following compounds were prepared according to the procedure described in example 288, using 110.4_1 and the following aniline.

Assays for detecting and measuring the effects of compounds on the F508del-CFTR channel

CFTR-YFP high throughput experiment:

the protocol was designed to selectively screen small molecule compounds with F508 delCFTR-correcting activity in HTSYFO (yellow fluorescent protein) flux assays. In this protocol, cells are incubated with the compound to be detected for 24 hours, washed with phosphate buffer, stimulated with forskolin and standard potentiators, and the fluorescence of each plate measured kinetically is read on a 384-well HTS plate reader (e.g., Hamamatsu FDDD-6000).

YFP fluorescence intensity values were obtained at high speed before and after iodine buffer was injected into the experimental cells. Iodine enters the cell through CFTR channels on the plasma membrane and quenches YFP fluorescence. The fluorescence quenching rate is proportional to the total CFTR activity on the cell membrane. The dF508-CFTR syndrome increases the number of CFTR molecules on the plasma membrane of the cell being detected, thus accelerating YFP quenching. .

This method was originally developed to leave a position for the ceiling reader (Galietta et al, 2001) and is applicable to HTS mode (Sui et al, Assay Drug dev.

Fischer Rat Thyroid (FRT) cells stably express human af 508-CFTR and halide sensitive yellow fluorescent protein (YFP-H148Q/I152L 25, 22) (Galietta et al am.j.physiol Cell Physiol 281(5), C1734, 2001) and are cultured on plastic surfaces with Coon's modified Ham's F12 medium supplemented with FBS 10%, L-glutamine 2mM, penicillin 100U/ml and streptomycin 100 μ g/ml. FRT cells expressing F508CFTR and YFP were screened with G418(0.75-1.0 mg/ml) and bleomycin (3.2. mu.g/ml). For primary screening, FRT cells were placed in 384-well black-wall, clear-bottom microtiter plates (Costar; Corning) at a cell density of 20,000-40,000 cells per well. The test compounds were applied to the cells at different concentrations. Cells were cultured in a cell incubator at 37 ℃ in 5% carbon dioxide for 24-26 hours. The plates were washed with DPBS medium (Thermo, cat # SH30028.02) to remove unbound cells and compounds. Stimulation medium (25. mu.l) containing 20. mu.M Forskolin (Forskolin) and 30. mu.M P3[6- (ethyl-phenyl-sulfonyl) -4-oxo-1, 4-dihydro-quinolinyl-3-carboxylic acid 2-methoxy-benzylamine in F-12 coon's modified medium was added to the plate wells and incubated at room temperature for 60-120 min. Then 25 microliters of HEPES-PBS-I buffer (10mM HEPES, 1mM MgCl2, 3mM KCl, 1mM CaCl2, 150mM Na I) was added and the fluorescence quenching curve (excitation 500 nm/emission 540 nm; exposure 136 ms) on the FDSS-6000 plate reader (Hamamatsu) was immediately recorded. The quench rate is from least squares fit data as described by Sui et al (2010).

Reference to the literature

Galietta，L.J.，Jayaraman，S.，and Verkman，A.S.Cell-based assay for high-throughput quantitative screening of CFTR chloride transport agonists.Am.J.Physiol Cell Physiol 281(5)，C1734，2001。

Sui J，Cotard S，Andersen J，Zhu P，Staunton J，Lee M，Lin S.(2010)Optimization of a Yellow fluorescent protein-based iodide influx high-throughput screening assay for cystic fibrosis transmembrane conductance regulator(CFTR)modulators.Assay Drug Dev Technol.2010Dec；8(6):656-68。

Alejandro A.Pezzulo，Xiao Xiao Tang，Mark J.Hoegger，Mahmoud H.Abou Alaiwa，Shyam Ramachandran，Thomas O.Moninger，Phillip H.Karp，Christine L.Wohlford-Lenane，Henk P.Haagsman，Martin van Eijk，Botond Ba′nfi，Alexander R.Horswill，David A.Stoltz，Paul B.McCray Jr，Michael J.Welsh，Joseph Zabner.Reduced airway surface pH impairs bacterial killing in the porcine cystic fibrosis lung.Nature 487，109-115(2012)。

Determination of the Activity of Primary CF cells:

cell culture:

primary CF airway epithelial cells were obtained from UNC cystic fibrosis tissue procurement and cell culture nuclei. Cells were cultured in Heracell 150i incubator at 37 ℃ using growth medium (BEGM, Fischer Co.). The cells were then transferred to differentiation medium (airway liquid interface medium (ALI) medium; Lechner JF and LaVeck MA, J.tissue Culture Methods 1985, 9:43-48) and cultured on coated ribs, snapwells, for at least 4 weeks. Two days prior to the uss test, mucus from the apical surface of the cells was aspirated after incubation with 200 μ L of differentiation medium for at least thirty (30) minutes. On the day before the uss test, test compounds at different test concentrations were added to the basolateral surface of the cells, dissolved in dimethyl sulfoxide. Duplicate wells were prepared according to the scheme n-3 or n-4.

Electrophysiological procedures

Cells were treated with different concentrations of experimental material, combinations and concentrations of reference standards (3 μ M VX809 or 3 μ MFDL169, positive control) for 24 hours. Compound stock solutions were prepared in DMSO and 1/1000 was diluted in ALI medium to its final assay concentration. Cells were treated with the combined solution (2ml per dilution) and incubated at 37 ℃ for 24 hours.

Filtration experiment in Europe

For the ews experiment, cells on four Snapwell (6 well) plates were treated 24 hours prior to the experiment. The next day, each Snapwell filter was detached from the plate and mounted vertically in a 37 ℃ pre-equilibrated ews chamber at 37 ℃ on top of and outside of the substrate of 5ml HBS (pH 7.4) and bubbled with room air to facilitate mixing when adding the compound. The resting current was recorded for 10 minutes to ensure baseline stabilization. Resting current was blocked by apical addition of 3uM benzalkonium, an ENaC inhibitor. After 10 minutes, 10 micromolar forskolin was added to the apical and basolateral side to stimulate CFTR. An increase in chloride current is detected as an upward deflection of the trace. After an additional 10 minutes, potentiometer VX770(1 micromolar) was added, further increasing chloride current. CFTR-172 (a CFTR inhibitor, 20 micromolar) and/or bumetanide (20 micromolar) was finally added to block CFTR mediated chloride current, resulting in the observed reduction in current.

Equivalent current analysis

For equivalent amperometry, cells on four Transwell (24-well) plates were processed. The top surface of each Transwell plate was filled with 200. mu.l HBS, and the outer surface of the base was filled with 2ml HBS. The steel plate was placed horizontally on a 37 ℃ heating rack and equilibrated for a few minutes. The resting current was measured for 15 minutes, then 5 micromoles of benzalkonium chloride was added through the tip. After 20 minutes, 10 micromolar forskolin and 1 micromolar VX770 were added to the apical and basolateral side to stimulate CFTR. An increase in chloride current is considered a trace of the upward deflection. After another 30 minutes, CFTR-172 (a CFTR inhibitor, 20 micromolar) and/or bumetanide (20 μ M) was added to block CFTR mediated chloride current.

Data collection and analysis method

The raw data of the current vs time (sampling interval: 10s) and voltage vs time of the ews cell and the raw data of the resistance vs time (sampling interval: 5 min) in the equivalent current analysis were transmitted to Excel (Microsoft Office Professional, version 14.0.7106.5003) for analysis. CFTR specific current was measured as the average magnitude of the increase in current induced upon addition of mao monkey hormone and ended after addition of CFTR channel specific blocker CFTR-172. This mean is equal to the sum of the mean of the activated mao monkey hormones and the mean of the VX770 boost current. The average current ISTD measured in the cells IV treated with the carrier (0.1% DMSO) was subtracted from the current of the test article ITA or of the calibrator reference VX809(3 μm). For duplicate measurements, the mean vehicle minus response of the test article was normalized to the mean vehicle minus inhibitor response of reference corrector VX809(3 μ M).

I_NSC＝(I_TA-I_V)_(ave)/(I_STD-I_V)_(ave)(formula 1)

The second endpoint of the equivalent amperometry is NAUC, the normalized area under the curve (AUC), used to measure the response after addition of mao monkey hormone and VX770 to the assay, at a point prior to addition of the CFTR inhibitor. AUC is actually the average response times the duration of the response. The AUC of the experimental material, i.e. the AUCTA time, was then corrected by subtracting the average carrier reaction AUCV, ave over the same time period and normalizing the inhibitor sensitivity current according to the difference between the corrector reference standard VX809(3 μ M VX809r, ave) or FDL176(3 μ M FDL176r, ave) and the carrier reaction:

NAUC_TA＝[AUC_TA-AUC_V，ave]/[AUC_Vx809r，avs-AUC_V，ave](formula 2).

The normalized value for DMSO is 0.0 and the normalized value for VX-809 is 1.0. Combinations of compounds with VX-809 normalized to greater than 1.0 showed activity in combination assays. A value of 2 indicates that the test compound only doubles the effect on VX-809.

The experiment was repeated at least n-4 per concentration. Since the distribution of the ratio of two normal distributions is a cauchy distribution, the median value must be used for the mean value and the mean deviation must be used for the error of all normalized data. Potency (EC50) and efficacy (maximal response) were determined by fitting dose response data to a sigmoid colon dose response model (GraphPad Prism 5.04, manufacturer) using equation 3:

E＝E_min+(E_max-E_min)/(1+10^((LogEC50-S)*n_H) (formula 3)

Where E is the recorded reaction and S is the concentration of the test compound with VX-809. Since there are at most 8 points in the dose-response curve, only EC50 and the maximum (Emax) were allowed to change, whereas the minimum (Emin) was fixed to VX-809 response of 1.0, and the slope, nH, was fixed to 1.

Statistical comparisons (t-test and Mann-Whitney) were performed in Excel, and mean and error were calculated.

The following table provides the results of equivalent amperometric analysis in primary CF airway epithelium for 24 hours of cells treated with the combination solutions (2mL of each dilution: FDL169 (3. mu.M) or VX-809 (3. mu.M) (as described in column 5-1, "position corrector") and incubated at 37 ℃.

NAUC "+ + + + +" refers to a positive reference where NAUC > 170% was observed; NAUC "+ +" refers to a positive reference of 140% NAUC 170-; NAUC "+" refers to a positive reference with NAUC < 140%.

The patents and academic literature referred to herein establish the knowledge available to those of ordinary skill in the art. All U.S. patents or published or unpublished U.S. patent applications referred to herein are incorporated by reference in their entirety. All foreign patents or published or unpublished foreign patent applications mentioned herein are incorporated by reference in their entirety. All other published references, documents, manuscripts and scientific literature are incorporated herein by reference.