Macrocyclic inhibitors of KRAS for treatment of cancer

Abstract

The present invention relates to: compounds of formula (I) # imgabs0 # wherein R1 to R7, A1 and A2 are as described herein, and pharmaceutically acceptable salts thereof; as well as compositions comprising the compounds and methods of using the compounds.

Description

[0001] The present invention relates to organic compounds useful for treatment and / or prevention in mammals, and in particular to the inhibition of KRAS G12C useful for the treatment of cancer. Technical Field

[0002] RAS is one of the best-known oncogenes. About 30% of human cancers contain mutations in the three most significant members (KRAS, HRAS, and NRAS), making them the most common oncogenic drivers. KRAS mutations are often associated with poor prognosis, especially in colorectal cancer, pancreatic cancer, and lung cancer. As the most frequently mutated RAS subtype, KRAS has been extensively studied in the past few years. Among the most common KRAS alleles (including G12D, G12V, G12C, G13D, G12R, G12A, G12S, Q61H, etc.), G12C, G12D, and G12V account for more than half of all K-RAS-driven cancers (including colorectal cancer (CRC), pancreatic ductal adenocarcinoma (PDAC), and lung adenocarcinoma (LUAD)). Notably, among all KRAS-altered cancers (ovarian, esophageal and gastric, uterine), KRAS wild-type amplification was also found in approximately 7%, ranking among the top alterations.

[0003] All RAS proteins belong to the protein family of small GTPases that hydrolyze GTP to GDP. KRAS is structurally divided into an effector-binding lobe, followed by an allosteric lobe and a carboxyl-terminal region responsible for membrane anchoring. The effector lobe includes the P-loop, switch I, and switch II regions. The switch I / II loop plays a key role in KRAS downstream signaling by mediating protein-protein interactions with effector proteins, such as RAF in the mitogen-activated protein kinase (MAPK) pathway or PI3K in the phosphatidylinositol 3-kinase (PI3K) / protein kinase B (AKT) pathway.

[0004] The KRAS protein switches between an inactive form and an active form via binding to GTP and GDP, respectively. Under physiological conditions, the transition between these two states is regulated by guanine nucleotide exchange factors (GEFs), such as seven son homolog 1 (SOS1) or GTPase activating proteins (GAPs) involved in catalyzing the exchange of GDP for GTP, thereby enhancing intrinsic GTPase activity or accelerating RAS-mediated GTP hydrolysis. In response to extracellular stimuli, inactive RAS-GDP is converted to active RAS-GTP, which directly binds to the RAF RAS binding domain (RAF RBD) binds to the RAF family of kinases, thereby recruiting the RAF kinase family from the cytoplasm to the cell membrane, where they dimerize and become active. The activated RAF then undergoes a series of phosphorylation reactions on its downstream mitogen-activated protein kinase (MEK) and extracellular signal-regulated kinase (ERK), and propagates growth signals. Among the RAF protein kinase family (three known subtypes ARAF, BRAF, CRAF / RAF1), BRAF is the most frequently mutated and remains the most potent activator of MEK. Although individual RAS and RAF family members exhibit different binding preferences, all RAFs have a conserved RBD for forward transmission of MAPK signaling, which is frequently used to characterize KRAS inhibition (such as KRAS-BRAF in this article). RBD For KRAS, mutations at positions 12, 13, 61, and 146 lead to a shift to the active KRAS form by impairing nucleotide hydrolysis or activating nucleotide exchange, thereby causing overactivation of the MAPK pathway that causes tumorigenesis.

[0005] Despite its widely recognized importance in cancer malignancies, sustained efforts in the past have failed to develop approved therapies for KRAS mutant cancers until recently, when the first selective drug, AMG510, was rapidly approved as a second-line treatment for KRAS G12C-driven non-small cell lung cancer (NSCLC). However, clinically acquired resistance to KRAS G12C inhibitors emerges dramatically as the disease progresses after approximately 6 months of treatment. All mutations converge to reactivate RAS-MAPK signaling, with secondary RAS mutants observed at oncogenic hotspots (e.g., G12 / G13 / Q61) and within the switch II pocket (e.g., H95, R68, and Y96); moreover, more than 85% of all cancers driven by KRAS mutant or wild-type amplification still lack novel agents. In summary, both the myriad escape mechanisms and the variety of oncogenic alleles highlight the urgent medical need for additional KRAS therapies. Therefore, we have developed oral compounds that target and inhibit KRAS alleles for the treatment of KRAS mutant-driven cancers.

[0006] First-generation KRAS G12C inhibitors such as sotorosib and adagrasib, which target the 'GDP-bound off' form (RASOFF) of the KRAS G12C mutation, have demonstrated promising efficacy. Although this treatment benefits many patients with activating KRAS mutations, almost all of the patients who initially benefit eventually acquire resistance through various mechanisms. An increasing number of KRAS G12C secondary mutations that demonstrate resistance to KRAS (OFF) G12C inhibitors have been identified from patient samples (Tanaka et al., Cancer Discovery (2021), Awad et al., NEJM (2021), Ho et al., EJC (2021), Zhao et al., Nature (2021), Tsai et al., JCI (2022)), or from saturation mutagenesis (Siyu et al., PNAS (2022)) and ENU mutagenesis (Takamasa et al., J Thorac Oncol (2021)), such as Y96D, R68S, H95D, H95Q, H95R, V8L. Therefore, there is an unmet need to prevent the acquisition of one or more mutations in RAS that confer resistance to RAS (OFF) inhibitors. Summary of the invention

[0007] The present invention relates to novel compounds having formula (I),

[0008]

[0009] in

[0010] R 1 for

[0011] Where R 8 C 1-6 alkyl;

[0012] R 9 C 3-7 Cycloalkyl, azetidinyl or phenyl, the C 3-7 Cycloalkyl, azetidinyl and phenyl are halogenated 3-6 Alkynyl, (halogenated C 3-6 Alkylpyrimidinyl)C 2-6 Alkynyl or pyrimidinyl C 2-6 Alkynyl substitution;

[0013] R 2 C 1-6 alkyl;

[0014] R 3 is H or halogen;

[0015] R 4 is H or halogen;

[0016] R 5 C 1-6 Alkyl or halogenated C 1-6 alkyl;

[0017] R 6 C 1-6 Alkoxy C 1-6 alkyl;

[0018] R 7 For morpholinyl, (halogenated C 1-6 alkyl) piperazinyl or C 1-6 Alkylpiperazinyl;

[0019] A 1 is thiazolylidene;

[0020] A 2 C 1-6 Alkylene;

[0021] The condition is R 3 and R 4 Not at the same time H;

[0022] or a pharmaceutically acceptable salt thereof.

[0023] The invention also relates to their manufacture, to medicaments based on the compounds according to the invention and their production and to the use of the compounds of formula (I) or (Ia) as KRAS inhibitors.

[0024] Compared with the reference compound, the compounds of the present invention solve the GSH toxicity problem. The compounds of formula (I) or (Ia) show good KRAS inhibition on G12C, G12D and G12V. In another embodiment, the compounds of the present invention show excellent cancer cell inhibition and human hepatocyte stability. In addition, the compounds of formula (I) or (Ia) also show good or improved cytotoxicity, solubility and single dose pharmacokinetic (SDPK) characteristics. In addition, the compounds of the present invention demonstrate good efficacy against the secondary mutations mentioned in this application. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 .X-ray crystallographic analysis of compound G5. DETAILED DESCRIPTION

[0026] definition

[0027] The term "C 1-6 "Alkyl" means a saturated, straight-chain or branched alkyl group containing 1 to 6, especially 1 to 4 carbon atoms, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, etc. In particular, "C1-6 "Alkyl" groups are methyl, ethyl and n-propyl.

[0028] The term "C 1-6 "Alkoxy" means C 1-6 Alkyl-O-.

[0029] The term “(C 1-6 ")Alkylene" means a straight or branched saturated divalent hydrocarbon group of 1 to 6 carbon atoms or a divalent branched saturated divalent hydrocarbon group of 3 to 6 carbon atoms. 1-6 ) Examples of the alkylene group include methylene, ethylene, propylene, 2-methylpropylene, butylene, 2-ethylbutylene, pentylene, hexylene.

[0030] The terms "halogen" and "halo" are used interchangeably herein to refer to fluorine, chlorine, bromine or iodine.

[0031] The term "C 2-6 "Alkynyl" refers to a monovalent straight or branched hydrocarbon group of 2 to 6 carbon atoms with at least one triple bond. In particular embodiments, the alkynyl group has 2 to 4 carbon atoms and at least one triple bond. 2-6 Examples of alkynyl groups include ethynyl (-C≡CH), prop-1-ynyl (-C≡CCH 3 ), prop-2-ynyl (propargyl, –CH 2 C≡CH), but-1-ynyl, but-2-ynyl and but-3-ynyl.

[0032] The terms "halogen" and "halo" are used interchangeably herein to refer to fluorine, chlorine, bromine or iodine.

[0033] The term "C 3-6 "Alkynyl" refers to a monovalent straight or branched hydrocarbon group of 3 to 6 carbon atoms with at least one triple bond. In certain embodiments, the alkynyl group has 3 to 4 carbon atoms and at least one triple bond. 3-6 Examples of alkynyl groups include prop-1-ynyl (-C≡CCH 3 ), prop-2-ynyl (propargyl, –CH 2 C≡CH), but-1-ynyl, but-2-ynyl and but-3-ynyl.

[0034] The term "halogenated C 1-6 "Alkyl" means C 1-6 Alkyl groups, where C 1-6 At least one hydrogen atom of the alkyl group has been replaced by identical or different halogen atoms, in particular fluorine atoms. Examples of haloalkyl groups include monofluoro-, difluoro- or trifluoro-methyl, -ethyl or -propyl, for example 3,3,3-trifluoropropyl, 2-fluoroethyl, 2,2,2-trifluoroethyl, fluoromethyl or trifluoromethyl.

[0035] The term "halogenated C 3-6 "Alkynyl" means C 3-6 Alkynyl group, wherein the C 3-6 At least one of the hydrogen atoms of the alkynyl group has been replaced by identical or different halogen atoms. 3-6 Examples of the alkynyl group include 3,3,3-trifluoroprop-1-ynyl.

[0036] The term "C 3-7 "Cycloalkyl" means a monovalent saturated monocyclic or bicyclic hydrocarbon radical of 3 to 7 ring carbon atoms. Bicyclic means consisting of two saturated carbocyclic rings having one or more common carbon atoms. Examples of monocyclic cycloalkyl are cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl or cycloheptyl. Examples of bicyclic cycloalkyl are bicyclo[1.1.0]butyl, bicyclo[2.2.1]heptyl, bicyclo[1.1.1]pentanyl or bicyclo[2.2.2]octyl.

[0037] The term "thiazolylene" refers to a divalent thiazolyl group.

[0038] The term "oxo" refers to a divalent oxygen atom =0.

[0039] The term "2,2-propylene" means

[0040] The term "blocking group" means a group that selectively blocks a reactive site in a multifunctional compound so that a chemical reaction occurs selectively on another unprotected reactive site that is usually associated therewith in synthetic chemistry. The blocking group can be removed at an appropriate time point. Exemplary blocking groups are amino-protecting groups, carboxyl-protecting groups or hydroxyl-protecting groups.

[0041] It will be appreciated by those skilled in the art that the following structures of compounds of formula (Ia) and (Ia') are equivalent, especially with respect to the chiral center:

[0042]

[0043] The term "pharmaceutically acceptable salts" refers to salts that are not biologically or otherwise undesirable. "Pharmaceutically acceptable salts" include both acid addition salts and base addition salts.

[0044] "Pharmaceutically acceptable acid addition salts" refer to those pharmaceutically acceptable salts formed with inorganic acids, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, carbonic acid, phosphoric acid, and the like, and organic acids, which may be selected from aliphatic, alicyclic, aromatic, aromatic aliphatic, heterocyclic, formic and sulfonic acid organic acids, such as formic acid, acetic acid, propionic acid, glycolic acid, gluconic acid, lactic acid, pyruvic acid, oxalic acid, malic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, aspartic acid, ascorbic acid, glutamic acid, anthranilic acid, benzoic acid, cinnamic acid, mandelic acid, pamoic acid, phenylacetic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid, and the like.

[0045] The term "pharmaceutical base addition salt" refers to those pharmaceutical salts formed with organic or inorganic bases. Examples of acceptable inorganic bases include sodium, potassium, ammonium, calcium, magnesium, iron, zinc, copper, manganese and aluminum salts. Salts derived from pharmaceutically acceptable organic non-toxic bases include primary amines, secondary amines, tertiary amines, substituted amines (including naturally occurring substituted amines), cyclic amines and basic ion exchange resins (such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, 2-diethylaminoethanol, tromethamine, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, ethylenediamine, glucosamine, methylglucamine, theobromine, purines, piperazine, piperidine, N-ethylpiperidine and polyamine resins).

[0046] The term "pharmaceutically active metabolite" refers to a pharmacologically active product produced by the metabolism of a specific compound or its salt in the body. After entering the human body, most drugs are substrates for chemical reactions that may change their physical properties and biological effects. These metabolic transformations, which usually affect the polarity of the compounds of the present invention, change the way the drug is distributed in the body and excreted from the body. However, in some cases, drug metabolism is necessary for therapeutic effects.

[0047] The term "therapeutically effective amount" refers to an amount of a compound or molecule of the invention that, when administered to a subject, (i) treats or prevents a particular disease, disorder or condition, (ii) reduces, ameliorates or eliminates one or more symptoms of a particular disease, disorder or condition, or (iii) prevents or delays the onset of one or more symptoms of a particular disease, disorder or condition as described herein. The therapeutically effective amount depends on the compound, the disease state being treated, the severity of the disease being treated, the age and relative health of the subject, the route and form of administration, the judgment of the attending medical or veterinary doctor and other factors.

[0048] The term "pharmaceutical composition" means a mixture or solution comprising a therapeutically effective amount of an active drug ingredient together with a pharmaceutically acceptable excipient for administration to a mammal (eg, a human) in need thereof.

[0049] The terms "pharmaceutically acceptable excipient," "pharmaceutically acceptable carrier," and "therapeutically inert excipient" are used interchangeably and refer to any pharmaceutical ingredient in a pharmaceutical composition that is not therapeutically active and is non-toxic to the subject to which it is administered, such as a disintegrant, binder, filler, solvent, buffer, tonicity agent, stabilizer, antioxidant, surfactant, carrier, diluent, or lubricant used to formulate a drug product.

[0050] KRAS inhibitors

[0051] The present invention relates to (i'), which is a compound of formula (I),

[0052]

[0053] in

[0054] R 1 for

[0055] Where R 8 C 1-6 alkyl;

[0056] R 9 C 3-7 Cycloalkyl, azetidinyl or phenyl, the C 3-7 Cycloalkyl, azetidinyl and phenyl are halogenated 3-6 Alkynyl, (halogenated C 3-6 Alkylpyrimidinyl)C 2-6 Alkynyl or pyrimidinyl C 2-6 Alkynyl substitution;

[0057] R 2 C 1-6 alkyl;

[0058] R 3 is H or halogen;

[0059] R 4 is H or halogen;

[0060] R 5 C 1-6 Alkyl or halogenated C 1-6 alkyl;

[0061] R 6 C 1-6 Alkoxy C 1-6 alkyl;

[0062] R 7 For morpholinyl, (halogenated C 1-6 alkyl) piperazinyl or C 1-6 Alkylpiperazinyl;

[0063] A1 is thiazolylidene;

[0064] A 2 C 1-6 Alkylene;

[0065] The condition is R 3 and R 4 Not at the same time H;

[0066] or a pharmaceutically acceptable salt thereof.

[0067] Another embodiment of the present invention is (ii') a compound of formula (Ia),

[0068]

[0069] in

[0070] R 1 for

[0071] Where R 8 C 1-6 alkyl;

[0072] R 9 C 3-7 Cycloalkyl, azetidinyl or phenyl, the C 3-7 Cycloalkyl, azetidinyl and phenyl are halogenated 3-6 Alkynyl, (halogenated C 3-6 Alkylpyrimidinyl)C 2-6 Alkynyl or pyrimidinyl C 2-6 Alkynyl substitution;

[0073] R 2 C 1-6 alkyl;

[0074] R 3 is H or halogen;

[0075] R 4 is H or halogen;

[0076] R 5 C 1-6 Alkyl or halogenated C 1-6 alkyl;

[0077] R 6 C 1-6 Alkoxy C 1-6 alkyl;

[0078] R 7 For morpholinyl, (halogenated C 1-6 alkyl) piperazinyl or C 1-6 Alkylpiperazinyl;

[0079] A 1 is thiazolylidene;

[0080] A 2 C 1-6 Alkylene;

[0081] The condition is R 3 and R 4 Not at the same time H;

[0082] or a pharmaceutically acceptable salt thereof.

[0083] The present invention relates to (i) a compound of formula (I),

[0084]

[0085] in

[0086] R 1 for

[0087] Where R 8 C 1-6 alkyl;

[0088] R 9 C 3-7 Cycloalkyl, azetidinyl or phenyl, the C 3-7 Cycloalkyl, azetidinyl and phenyl are halogenated 3-6 Alkynyl or pyrimidinyl C 2-6 Alkynyl substitution;

[0089] R 2 C 1-6 alkyl;

[0090] R 3 is H or halogen;

[0091] R 4 is H or halogen;

[0092] R 5 C 1-6 Alkyl or halogenated C 1-6 alkyl;

[0093] R 6 C 1-6 Alkoxy C 1-6 alkyl;

[0094] R 7 For morpholinyl, (halogenated C 1-6 alkyl) piperazinyl or C 1-6 Alkylpiperazinyl;

[0095] A 1 is thiazolylidene;

[0096] A 2 C 1-6 Alkylene;

[0097] The condition is R 3 and R 4 Not at the same time H;

[0098] or a pharmaceutically acceptable salt thereof.

[0099] Another embodiment of the present invention is (ii) a compound of formula (Ia),

[0100]

[0101] in

[0102] R 1 for

[0103] Where R 8 C 1-6 alkyl;

[0104] R 9 C 3-7 Cycloalkyl, azetidinyl or phenyl, the C 3-7 Cycloalkyl, azetidinyl and phenyl are halogenated 3-6 Alkynyl or pyrimidinyl C 2-6 Alkynyl substitution;

[0105] R 2 C 1-6 alkyl;

[0106] R 3 is H or halogen;

[0107] R 4 is H or halogen;

[0108] R 5 C 1-6 Alkyl or halogenated C 1-6 alkyl;

[0109] R 6 C 1-6 Alkoxy C 1-6 alkyl;

[0110] R 7 For morpholinyl, (halogenated C 1-6 alkyl) piperazinyl or C 1-6 Alkylpiperazinyl;

[0111] A 1 is thiazolylidene;

[0112] A 2 C 1-6 Alkylene;

[0113] The condition is R 3 and R 4 Not at the same time H;

[0114] or a pharmaceutically acceptable salt thereof.

[0115] A further embodiment of the present invention is (iii) a compound of formula (I) or (Ia) according to (i), (ii), (i') or (ii'), or a pharmaceutically acceptable salt thereof, wherein R 1 for Where R 8 C 1-6 Alkyl; R 9 For halogenated C 3-6 Alkynyl substituted C 3-7 Cycloalkyl.

[0116] A further embodiment of the present invention is (iv) a compound of formula (I) or (Ia) according to any one of (i) to (iii), (i') and (ii'), or a pharmaceutically acceptable salt thereof, wherein R 1 for Where R 8 is methyl; R 9 is cyclobutyl substituted by 3,3,3-trifluoroprop-1-ynyl.

[0117] A further embodiment of the present invention is (v), a compound of formula (I) or (Ia) according to any one of (i) to (iv), (i') and (ii'), wherein R 9 It is 3-(3,3,3-trifluoroprop-1-ynyl)cyclobutyl.

[0118] A further embodiment of the present invention is (vi), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (v), (i') and (ii'), wherein R 2 It is isopropyl.

[0119] A further embodiment of the present invention is (vii), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (vi), (i') and (ii'), wherein R 3 It is a halogen.

[0120] A further embodiment of the present invention is (viii), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (vii), (i') and (ii'), wherein R 3 For fluorine.

[0121] A further embodiment of the present invention is (ix), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (viii), (i') and (ii'), wherein R 4 is H or fluorine.

[0122] A further embodiment of the present invention is (x), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (ix), (i') and (ii'), wherein R 4 For H.

[0123] A further embodiment of the present invention is (xi), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (x), (i') and (ii'), wherein R 5 It is ethyl or 2,2,2-trifluoroethyl.

[0124] A further embodiment of the present invention is (xii), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (xi), (i') and (ii'), wherein R 6 It is 1-methoxyethyl.

[0125] A further embodiment of the present invention is (xiii), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (xii), (i') and (ii'), wherein R 7 is morpholinyl, 4-(2,2,2-trifluoroethyl)piperazin-1-yl or 4-methylpiperazin-1-yl.

[0126] A further embodiment of the present invention is (xiv), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (xiii), (i') and (ii'), wherein A 1 for Where bond "a" is attached to the indole ring.

[0127] A further embodiment of the present invention is (xv), a compound of formula (I) or (Ia) or a pharmaceutically acceptable salt thereof according to any one of (i) to (xiv), (i') and (ii'), wherein A 2 It is 2,2-propylene.

[0128] A further embodiment of the present invention is (xvi), a compound of formula (I) or (Ia) according to (i) or (ii), (i') or (ii'), wherein

[0129] R 1 for Where R 8 C 1-6Alkyl; R 9 For halogenated C 3-6 Alkynyl substituted C 3-7 Cycloalkyl;

[0130] R 2 C 1-6 alkyl;

[0131] R 3 is a halogen;

[0132] R 4 is H;

[0133] R 5 C 1-6 Alkyl or halogenated C 1-6 alkyl;

[0134] R 6 C 1-6 Alkoxy C 1-6 alkyl;

[0135] R 7 For morpholinyl, (halogenated C 1-6 alkyl) piperazinyl or C 1-6 Alkylpiperazinyl;

[0136] A 1 for wherein bond "a" is attached to the indole ring;

[0137] A 2 C 1-6 Alkylene;

[0138] or a pharmaceutically acceptable salt thereof.

[0139] Another embodiment of the present invention is (xvii), a compound of formula (I) or (Ia) according to (xvi), wherein

[0140] R 1 for Where R 8 is methyl; R 9 is 3-(3,3,3-trifluoroprop-1-ynyl)cyclobutyl;

[0141] R 2 is isopropyl;

[0142] R 3 For fluorine;

[0143] R 4 is H;

[0144] R 5 is ethyl or 2,2,2-trifluoroethyl;

[0145] R6 is (1S)-1-methoxyethyl;

[0146] R 7 is morpholinyl, 4-(2,2,2-trifluoroethyl)piperazin-1-yl or 4-methylpiperazin-1-yl;

[0147] A 1 for wherein bond "a" is attached to the indole ring;

[0148] A 2 is 2,2-propylene;

[0149] or a pharmaceutically acceptable salt thereof.

[0150] Another embodiment of the present invention is (xviii), a compound of formula (I) or (Ia) selected from the following compounds:

[0151] trans-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide;

[0152] N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide;

[0153] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide;

[0154] N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)benzamide;

[0155] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(2-pyrimidin-2-ylethynyl)cyclobutanecarboxamide;

[0156] cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide;

[0157] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide;

[0158] cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide;

[0159] trans-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxamide;

[0160] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxamide;

[0161] cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide;

[0162] cis-N-[(1S)-1-[[(7S,13S)-25-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide;

[0163] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-[2-[5-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxamide;

[0164] N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide;

[0165] N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide;

[0166] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-[2-[4-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxamide;

[0167] N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide;

[0168] (2S)-N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]-2-isopropyl-4-oxo-4-[3-(3,3,3-trifluoroprop-1-ynyl)azetidin-1-yl]butanamide;

[0169] N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide;

[0170] cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-[2-[4-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxamide; and

[0171] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide;

[0172] or a pharmaceutically acceptable salt thereof.

[0173] Another embodiment of the present invention relates to (xix), a method for preparing a compound according to any one of (i) to (xviii), comprising the following steps:

[0174] a) reacting a compound of formula (II) in the presence of a coupling agent and a base,

[0175] With acid (III),

[0176] A coupling reaction is carried out between them to form a compound of formula (I);

[0177] Where R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , A 1 and A 2 As defined in any one of (i) to (xvii); the coupling reagent is T 3 P, HATU, PyBOP or EDCI / HOBt; the base is TEA, DIEPA or DMAP.

[0178] Another embodiment of the present invention is (xx), a compound or a pharmaceutically acceptable salt according to any one of (i) to (xviii), (i') and (ii') for use as a therapeutically active substance.

[0179] Another embodiment of the present invention is (xxi), a pharmaceutical composition comprising: a compound according to any one of (i) to (xviii), (i') and (ii'); and a pharmaceutically acceptable excipient.

[0180] Another embodiment of the present invention is (xxii), use of the compound according to any one of (i) to (xviii), (i') and (ii') for treating a KRAS G12C protein-related disease.

[0181] Another embodiment of the present invention is (xxiii), use of the compound according to any one of (i) to (xviii) for treating diseases associated with KRAS G12C, G12D and G12V proteins.

[0182] Another embodiment of the present invention is (xxiv), use of the compound according to any one of (i) to (xviii), (i') and (ii') for inhibiting the interaction of RAS with downstream effectors, wherein the downstream effectors are RAF and PI3K.

[0183] Another embodiment of the present invention is (xxv), use of a compound according to any one of (i) to (xviii), (i') and (ii') for inhibiting propagated oncogenic MAPK and PI3K signaling.

[0184] Another embodiment of the present invention is (xxvi), use of a compound according to any one of (i) to (xviii), (i') and (ii') for treating or preventing a KRAS mutation-driven cancer, wherein the cancer is selected from pancreatic cancer, colorectal cancer, lung cancer, esophageal cancer, gallbladder cancer, melanoma, ovarian cancer and endometrial cancer.

[0185] Another embodiment of the present invention is (xxvii), use of a compound according to any one of (i) to (xviii), (i') and (ii') for treating or preventing a KRAS mutation-driven cancer, wherein the cancer is selected from pancreatic adenocarcinoma, colorectal cancer and non-small cell lung cancer.

[0186] Another embodiment of the present invention is (xxviii), a compound or a pharmaceutically acceptable salt according to any one of (i) to (xviii), (i') and (ii'), for use in treating or preventing a KRAS mutation-driven cancer, wherein the cancer is selected from pancreatic adenocarcinoma, colorectal cancer and non-small cell lung cancer.

[0187] Another embodiment of the present invention is (xxix), the use of a compound according to any one of (i) to (xviii), (i') and (ii') for treating or preventing a KRAS mutation-driven cancer, wherein the cancer comprises: a first mutation of G12C; and a second mutation at a position selected from V8A, V9Y, S17E, T58I, A59T, S65W, R68S, D69P, M72I, D92R, H95N, Y96D, Q99F, Q99W, Y96H and F156L.

[0188] Another embodiment of the present invention is (xxx), the use of a compound according to any one of (i) to (xviii), (i') and (ii') for the preparation of a medicament for treating or preventing a KRAS mutation-driven cancer, wherein the cancer is selected from pancreatic adenocarcinoma, colorectal cancer and non-small cell lung cancer.

[0189] Another embodiment of the present invention is (xxxi), the use of a compound according to any one of (i) to (xviii), (i') and (ii') for the preparation of a medicament for treating or preventing a KRAS mutation-driven cancer, wherein the cancer comprises: a first mutation of G12C; and a second mutation at a position selected from V8A, V9Y, S17E, T58I, A59T, S65W, R68S, D69P, M72I, D92R, H95N, Y96D, Q99F, Q99W, Y96H and F156L.

[0190] Another embodiment of the present invention is (xxxii), a method for treating or preventing a KRAS mutation-driven cancer, wherein the cancer is selected from pancreatic adenocarcinoma, colorectal cancer and non-small cell lung cancer, the method comprising administering a therapeutically effective amount of a compound as defined in any one of (i) to (xviii), (i') and (ii').

[0191] Another embodiment of the present invention is (xxxiii), a method for treating or preventing a KRAS mutation-driven cancer, wherein the cancer comprises: a first mutation of G12C; and a second mutation at a position selected from V8A, V9Y, S17E, T58I, A59T, S65W, R68S, D69P, M72I, D92R, H95N, Y96D, Q99F, Q99W, Y96H and F156L.

[0192] Another embodiment of the present invention is (xxxiv), a compound or a pharmaceutically acceptable salt according to any one of (i) to (xviii), (i') and (ii'), which is prepared according to the method described in (xix).

[0193] Pharmaceutical compositions and administration

[0194] Another embodiment provides a pharmaceutical composition or medicine containing the compounds of this invention and a therapeutic inert carrier, diluent or excipient, and methods for preparing such compositions and medicines using the compounds of this invention. In one example, the compound of formula (I) can be formulated as a galenical administration form by mixing with a physiologically acceptable carrier (i.e., a carrier that is nontoxic to the recipient at the dosage and concentration used) at ambient temperature under appropriate pH and desired purity. The pH of the preparation depends primarily on the specific use and concentration of the compound, but is preferably in the range of about 3 to about 8. In one example, the compound of formula (I) is prepared in an acetate buffer at pH 5. In another embodiment, the compound of formula (I) is sterile. The compound can be stored, for example, as a solid or amorphous composition, as a lyophilized preparation, or as an aqueous solution.

[0195] The composition is formulated, metered, and administered in a manner consistent with good medical practice. Factors to be considered in this case include the specific disorder being treated, the specific mammal being treated, the clinical condition of the individual patient, the cause of the disorder, the delivery site of the agent, the method of administration, the timing of administration, and other factors known to practitioners. The "effective amount" of the compound to be administered will be affected by such considerations and is the minimum amount necessary to inhibit the interaction of mutant RAS (e.g., KRAS G12C) with RAF, thereby blocking oncogenic MAPK signaling. For example, the amount may be lower than an amount that is toxic to normal cells or mammals as a whole.

[0196] In one example, the pharmaceutically effective amount of the compound of the present invention administered parenterally per dose will be in the range of about 0.1 to 1000 mg / kg patient body weight per day, alternatively about 0.1 to 1000 mg / kg patient body weight, typically an initial range of 0.3 to 15 mg / kg / day of the compound used. In another embodiment, oral unit dosage forms such as tablets and capsules preferably contain about 1 to about 1000 mg of the compound of the present invention.

[0197] The compounds of the present invention can be administered by any suitable means, including oral, topical (including buccal and sublingual), rectal, vaginal, transdermal, parenteral, subcutaneous, intraperitoneal, intrapulmonary, intradermal, intrathecal and epidural and intranasal, and (if necessary for local treatment) intralesional administration. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal or subcutaneous administration.

[0198] The compounds of the present invention can be administered in any convenient administration form, for example, tablets, powders, capsules, solutions, dispersions, suspensions, syrups, sprays, suppositories, gels, emulsions, patches, etc. Such compositions may contain conventional components of pharmaceutical preparations, for example, diluents, carriers, pH adjusters, sweeteners, fillers and other active agents.

[0199] Typical formulations are prepared by mixing the compounds of the invention with carriers or excipients. Suitable carriers and excipients are well known to those skilled in the art and are described in, for example, Ansel, Howard C. et al., Ansel's Pharmaceutical Dosage Forms and Drug Delivery Systems. Philadelphia: Lippincott, Williams & Wilkins, 2004; Gennaro, Alfonso R. et al. Remington:The Science and Practice of Pharmacy .Philadelphia: Lippincott, Williams & Wilkins, 2000; and Rowe, Raymond C. Handbook of Pharmaceutical Excipients .Chicago, Pharmaceutical Press, 2005. The formulation may also contain one or more buffers, stabilizers, surfactants, wetting agents, lubricants, emulsifiers, suspending agents, preservatives, antioxidants, opacifiers, glidants, processing aids, colorants, sweeteners, flavoring agents, flavoring agents, diluents and other known additives to provide an aesthetically pleasing display of a drug (e.g., a compound of the present invention or a pharmaceutical composition thereof) or to facilitate the preparation of a pharmaceutical product (e.g., a drug).

[0200] An example of a suitable oral dosage form is a tablet containing about 1 to 1000 mg of a compound of the present invention and about 1 to 1000 mg of anhydrous lactose, about 1 to 1000 mg of cross-linked sodium carboxymethylcellulose, about 1 to 1000 mg of polyvinyl pyrrolidone (PVP) K30 and about 1 to 1000 mg of magnesium stearate. The powdered ingredients are first mixed together and then mixed with the PVP solution. The resulting composition can be dried, granulated, mixed with magnesium stearate and compressed into tablet form using conventional equipment. An example of an aerosol formulation can be prepared by dissolving the compound of the present invention (e.g., 5 mg to 400 mg) in a suitable buffer solution (e.g., phosphate buffer) and adding a penetration enhancer (e.g., a salt such as sodium chloride) if necessary. The solution can be filtered, for example, using a 0.2 micron filter to remove impurities and contaminants.

[0201] Thus, embodiments include a pharmaceutical composition comprising a compound of formula (I) or a stereoisomer or a pharmaceutically acceptable salt thereof. Further embodiments include a pharmaceutical composition comprising a compound of formula (I) or a stereoisomer or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier or excipient.

[0202] Another embodiment includes a pharmaceutical composition comprising a compound of formula (I) for use in treating a mutant KRAS driven cancer. Another embodiment includes a pharmaceutical composition comprising a compound of formula (I) for use in treating a mutant KRAS driven cancer.

[0203] The following compositions A and B illustrate typical compositions of the present invention but are intended to be representative thereof only.

[0204] Composition A

[0205] The compounds of the invention can be used as active ingredients in a manner known per se to produce tablets of the following composition:

[0206]

[0207] Composition B

[0208] The compounds of the invention can be used as active ingredients in a manner known per se to produce capsules of the following composition:

[0209]

[0210] Indications and treatments

[0211] The compounds of the present invention induce new binding pockets in KRAS by driving the formation of a high-affinity tri-complex between KRAS protein and widely expressed cyclophilin A (CYPA), and these compounds inhibit the interaction of KRAS with downstream effectors (such as RAF and PI3K). Therefore, the compounds of the present invention can be used to inhibit the propagation of oncogenic MAPK and PI3K signal transduction, reducing cell (especially cancer cell) proliferation. The compounds of the present invention can be used to terminate RAS signal transduction in cells of pancreatic cancer, colorectal cancer, lung cancer, esophageal cancer, gallbladder cancer, melanoma, ovarian cancer, endometrial cancer, etc. driven by RAS mutants (especially KRAS mutations). Alternatively, the compounds of the present invention can be used to terminate RAS signal transduction in malignant solid tumors, wherein the carcinogenic effect of KRAS mutations is enhanced by the dysregulation or mutation of effector pathways such as signal transduction driven by MAPK, PI3K-AKT-mTOR (mammalian rapamycin target), and these compounds are used for targeted therapies of pancreatic adenocarcinoma, colorectal cancer, non-small cell lung cancer, etc.

[0212] Another embodiment includes a method of treating or preventing cancer in a mammal in need of such treatment, wherein the method comprises administering to said mammal a therapeutically effective amount of a compound of formula (I), a stereoisomer, tautomer, prodrug or pharmaceutically acceptable salt thereof.

[0213] synthesis

[0214] The compounds of the present invention may be prepared by any conventional method. Suitable methods for synthesizing these compounds and their starting materials are provided in the following schemes and examples. Unless otherwise indicated, all substituents, especially R 1 To R 7 , A 1 and A 2 As defined above. Furthermore, unless otherwise specifically indicated, all reactions, reaction conditions, abbreviations and symbols have the meanings well known to one of ordinary skill in the art of organic chemistry.

[0215] The general synthetic route for preparing compounds of formula (I) is shown below.

[0216] Solution 1

[0217]

[0218] Compounds of formula II can be prepared according to the procedures described in Intermediates A to K. Compounds of formula (I) are prepared by reacting the compounds with a coupling reagent such as TEA, DIEPA and DMAP in the presence of a base such as TEA, DIEPA and DMAP. 3 The obtained product is obtained by coupling reaction between the acid (III) and the compound of formula (II) using P, HATU, PyBOP and EDCI / HOBt.

[0219] The compounds of the present invention can be obtained as diastereomers or mixtures of enantiomers, which can be separated by methods well known in the art, for example, (chiral) HPLC or SFC. In another embodiment, compounds of formula (I) can be obtained according to the above scheme using the corresponding chiral starting materials.

[0220] The present invention also relates to a method for preparing a compound of formula (I), the method comprising the following steps:

[0221] a) reacting a compound of formula (II) in the presence of a coupling agent and a base,

[0222] With acid (III),

[0223] A coupling reaction is carried out between them to form a compound of formula (I);

[0224] in

[0225] In step a), the coupling reagent may be, for example, T3 P, HATU, PyBOP or EDCI / HOBt; the base can be, for example, TEA, DIEPA or DMAP.

[0226] The compounds of formula (I) or (Ia) produced according to the above-described process are also an object of the present invention.

[0227] Examples

[0228] The present invention will be more fully understood by reference to the following examples. However, they should not be construed as limiting the scope of the present invention.

[0229] abbreviation

[0230] The present invention will be more fully understood by reference to the following examples. However, they should not be construed as limiting the scope of the present invention.

[0231] The abbreviations used in this article are as follows:

[0232] CAN Acetonitrile

[0233] aq. water-based

[0234] Boc-N-Me-Val-OH N-(tert-Butoxycarbonyl)-N-methyl-L-valine

[0235] (Boc) 2 O Di-tert-butyl dicarbonate

[0236] (R)-binap (R)-(+)-2,2′-bis(diphenylphosphino)-1,1'-binaphthyl

[0237] CDCl 3 : Deuterated chloroform

[0238] CD 3 OD: deuterated methanol

[0239] COMU (1-cyano-2-ethoxy-2-oxoethylideneaminooxy)dimethylamino-morpholino-carbonium hexafluorophosphate

[0240] DIEPA: N,N-diethylpropylamine

[0241] DIBAL-H Diisobutylaluminum hydride

[0242] DMAP: 4-dimethylaminopyridine

[0243] DMF: Dimethylformamide

[0244] DMP 1,1,1-tri(acetoxy)-1,1-dihydro-1,2-benzimidoyl-3-(1H)-one

[0245] DMSO: dimethyl sulfoxide

[0246] EDCI: N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride

[0247] EtOAc or EA: Ethyl acetate

[0248] FRET Fluorescence Resonance Energy Transfer

[0249] HATU: (1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate)

[0250] hr: hours

[0251] HPLC: High Performance Liquid Chromatography

[0252] HOBt: N-Hydroxybenzotriazole

[0253] H-VAL-OTBU HCl (S)-tert-Butyl 2-amino-3-methylbutyrate hydrochloride

[0254] [Ir(OMe)(COD)] 2 (1,5-cyclooctadiene)(methoxy)iridium(I) dimer

[0255] LDA Lithium diisopropylamide

[0256] MS: (ESI): Mass spectrometry (electrospray ionization)

[0257] min(s) minutes

[0258] MTBE Methyl tert-butyl ether

[0259] NMM N-Methylmorpholine

[0260] NMR: Nuclear Magnetic Resonance

[0261] NMO 4-Methylmorpholine N-oxide

[0262] obsd. Observed value

[0263] Pd(dppf)Cl 2 [1,1'-Bis(diphenylphosphino)ferrocene]palladium(II) chloride

[0264] Pd(dtbpf)Cl 2 [1,1'-Bis(di-tert-butylphosphino)ferrocene]palladium(II) dichloride

[0265] prep-HPLC Preparative High Performance Liquid Chromatography

[0266] PyBOP: Benzotriazol-1-yloxytripyrrolidinophosphonium hexafluorophosphate

[0267] RT or rt: room temperature

[0268] sat. saturated

[0269] SFC Supercritical Fluid Chromatography

[0270] TEA: Triethylamine

[0271] TFA: trifluoroacetic acid

[0272] THF: Tetrahydrofuran

[0273] TEA: Trimethylamine

[0274] TMEDA Tetramethylethylenediamine

[0275] TMSCF 3 Trifluoromethyltrimethylsilane

[0276] T 3 P: Propylphosphonic anhydride

[0277] General experimental conditions

[0278] Intermediates and final compounds were purified by flash chromatography using one of the following instruments: i) Biotage SP1 system and Quad 12 / 25 Cartridge module. ii) ISCO combi-flash chromatograph. Silica gel brand and pore size: i) KP-SIL Particle size: 40-60 μm; ii) CAS registration number: silica gel: 63231-67-4, particle size: 47-60 micron silica gel; iii) ZCX from Qingdao Ocean Chemical Co., Ltd., pore size: 200-300 or 300-400.

[0279] Intermediates and final compounds were purified by preparative HPLC on a reverse phase column using XBridge TM Prep-C18 (5μm, OBDTM 30×100mm) column, SunFire TM Prep-C18(5μm,OBD TM30×100mm) chromatographic column, Phenomenex Synergi-C18 (10μm, 25×150mm) or Phenomenex Gemini-C18 (10μm, 25×150mm). Waters AutoP purification system (sample manager 2767, pump 2525, detector: Micromass ZQ and UV 2487, solvent system: acetonitrile and 0.1% ammonium hydroxide in water; acetonitrile and 0.1% FA in water, or acetonitrile and 0.1% TFA in water). Or Gilson-281 purification system (pump 322, detector: UV 156, solvent system: acetonitrile and 0.05% ammonium hydroxide in water; acetonitrile and 0.225% FA in water; acetonitrile and 0.05% HCl in water; acetonitrile and 0.075% TFA in water; or acetonitrile and water).

[0280] For SFC chiral separation, the intermediates were separated by chiral columns (Daicel chiralpak IC, 5 μm, 30 × 250 mm), AS (10 μm, 30 × 250 mm) or AD (10 μm, 30 × 250 mm) using Mettler Toledo Multigram III system SFC, Waters 80Q preparative SFC or Thar 80 preparative SFC, solvent system: CO 2 and IPA (0.5% TEA in IPA) or CO 2 and MeOH (0.1% NH 3 ·H 2 O in MeOH), back pressure 100 bar, UV detection at 254 or 220 nm.

[0281] Using LC / MS (Waters TM The LC / MS spectra of the compounds were obtained using an Agilent Alliance 2795-Micromass ZQ, Shimadzu Alliance2020-Micromass ZQ or Agilent Alliance 6110-Micromass ZQ. The LC / MS conditions were as follows (run time 3 or 1.5 min):

[0282] Acidic conditions I: A: 0.1% TFA in HO 2 O solution; B: 0.1% TFA solution in acetonitrile;

[0283] Acidic conditions II: A: 0.0375% TFA in HO 2O solution; B: 0.01875% TFA solution in acetonitrile;

[0284] Alkaline conditions I: A: 0.1% NH 3 ·H 2 O in H 2 O solution; B: acetonitrile;

[0285] Alkaline conditions II: A: 0.025% NH 3 ·H 2 O in H 2 O solution; B: acetonitrile;

[0286] Neutral conditions: A:H 2 O; B: acetonitrile.

[0287] Mass spectra (MS): Usually only ions representing the parent mass are reported, and unless otherwise stated, the mass ion quoted is the positive mass ion (MH) + .

[0288] NMR spectra were obtained using a Bruker Avance 400 MHz.

[0289] Microwave-assisted reactions were performed in a Biotage Initiator Sixty microwave synthesizer. All reactions involving air-sensitive reagents were performed under argon or nitrogen atmosphere. Unless otherwise stated, reagents were purchased from commercial suppliers as received without further purification.

[0290] Preparation Example

[0291] Preparation of intermediates

[0292] Intermediate A1-[6-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-pyridinyl]-4-methyl-piperazine

[0293]

[0294] The title intermediate A was prepared according to the following scheme:

[0295]

[0296] Step 1: Preparation of 3-bromo-2-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine (Compound A2)

[0297] To a solution of 3-bromo-2-[(1S)-1-methoxyethyl]pyridine (Compound A1, 2.0 g, 9.26 mmol) and bis(pinacolato)diboron (3.5 g, 13.9 mmol) in THF (30 mL) was added 4,4'-di-tert-butyl-2,2'-bipyridine (372.7 mg, 1.39 mmol) and [Ir(OMe)(COD)] 2 (306.3 mg, 0.460 mmol). The mixture was stirred at N 2 The mixture was stirred at 75 ° C for 16 hours. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was purified by silica gel column chromatography (EA / PE: 0-20%) to give 3-bromo-2-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine (Compound A2, 2.4 g) as a yellow oil. 1 H NMR (400MHz, CDCl3) δppm 8.91 (d, J = 1.4Hz, 1H), 8.21 (d, J = 1.4Hz, 1H), 4.95 (q, J = 6.5Hz, 1H), 3.30 (s, 3H), 1.49 (d, J = 6.5Hz, 3H), 1.35 (s, 12H).

[0298] Step 2: Preparation of 3-bromo-5-iodo-2-[(1S)-1-methoxyethyl]pyridine (Compound A3)

[0299] To a solution of 3-bromo-2-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)pyridine (Compound A2, 2.5 g, 7.3 mmol) in ACN (40 mL) was added N-iodosuccinimide (4.1 g, 18.27 mmol). The mixture was stirred under N 2 The reaction was stirred at 90°C for 40 hours. 2 SO 3 The solution (40mL) was quenched, and the reaction mixture was extracted with EtOAc (30mL, twice). The combined organic layer was washed with brine (50mL), filtered, and the filtrate was concentrated under vacuum. The residue was purified by silica gel column chromatography (EA / PE: 0-20%) to obtain 3-bromo-5-iodo-2-[(1S)-1-methoxyethyl]pyridine (compound A3, 660mg) as a yellow oil. MS calculated value 342 (MH + ); measured value 341.8 (MH + ).

[0300] Step 3: Preparation of benzyl 4-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylate (Compound A5)

[0301] To a solution of 3-bromo-5-iodo-2-[(1S)-1-methoxyethyl]pyridine (Compound A3, 660 mg, 1.9 mmol) and 1-Cbz-piperazine (Compound A4, 425.1 mg, 1.9 mmol) in toluene (10 mL) was added cesium carbonate (1.6 g, 4.83 mmol), (R)-BINAP (60.1 mg, 0.1 mmol) and palladium (II) acetate (43.3 mg, 0.19 mmol). The mixture was stirred under N 2 The mixture was stirred at 100 ° C for 12 hours. The mixture was filtered and the filtrate was concentrated under vacuum. The residue was purified by silica gel column chromatography (EA / PE: 0-50%) to give 4-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylic acid benzyl ester (Compound A5, 740 mg) as a yellow solid. MS calculated value 434.1 (MH + ); measured value 434.1 (MH + ).

[0302] Step 4: Preparation of 1-[6-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-pyridinyl]-4-methyl-piperazine (Intermediate A)

[0303] To a solution of benzyl 4-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylate (Compound A5, 740 mg, 1.7 mmol) and bis(pinacolato)diboron (519.2 mg, 2.04 mmol) in toluene (12 mL) was added KOAc (418.0 mg, 4.26 mmol) and Pd(dppf)Cl 2 (124.7 mg, 0.170 mmol). The reaction mixture was stirred at N 2 The mixture was stirred at 90 ° C for 12 hours under protection. The mixture was filtered and the filtrate was concentrated in vacuo. The residue was purified by silica gel column to give 1- [6- [(1S) -1- methoxyethyl] -5- (4,4,5,5- tetramethyl -1,3,2- dioxaborolan -2-yl) -3-pyridinyl] -4-methyl-piperazine (Intermediate A, 470 mg) as a brown solid. MS calculated value 482.3 (MH + ); measured value 482.2 (MH + ).

[0304] Intermediate B

[0305] (3S)-1-[(2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)-propionyl]hexahydropyridazine-3-carboxylic acid methyl ester

[0306]

[0307] Intermediate B was prepared according to the following scheme:

[0308]

[0309] Step 1: Preparation of (4-bromothiazol-2-yl)methanol (Compound B2)

[0310] To a solution of 4-bromothiazole-2-carboxaldehyde (compound B1, 6.0 g, 31.25 mmol) in methanol (70 mL) was added sodium borohydride (1.7 g, 46.87 mmol) at 0 ° C. The mixture was stirred at 25 ° C for 1 hour. The reaction was quenched with water (300 mL) at 0 ° C, and the reaction mixture was extracted with ethyl acetate (200 mL, three times). The combined organic phases were washed with brine (150 mL, twice), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under vacuum to obtain (4-bromothiazol-2-yl)methanol (compound B2, 6 g) as a colorless oil.

[0311] Step 2: Preparation of 4-bromo-2-(bromomethyl)thiazole (Compound B3)

[0312] To a solution of (4-bromothiazol-2-yl)methanol (Compound B2, 6.0 g, 30.92 mmol) in DCM (80 mL) was added CBr at 0 °C. 4 (15.4 g, 46.38 mmol) and triphenylphosphine (12.1 g, 46.38 mmol). After stirring at 25 °C for 1 hour, the mixture was filtered and the filtrate was concentrated under vacuum. The residue was purified by silica gel column eluting with ethyl acetate (0-10%) in petroleum ether to give (4-bromothiazol-2-yl)methanol (Compound B3, 6.0 g) as a yellow oil. MS calculated value 255.9 (MH + ); measured value 255.9 (MH + ).

[0313] Step 3: Preparation of 4-bromo-2-[[(2S,5R)-5-isopropyl-3,6-dimethoxy-2,5-dihydropyrazin-2-yl]methyl]thiazole (Compound B5)

[0314] To a mixture of (R)-2,5-dihydro-3,6-dimethoxy-2-isopropylpyrazine (Compound B4, 4.3 g, 23.45 mmol) in THF (60 mL) was slowly added n-butyl lithium (10 mL, 25.22 mmol, 2.5 M) at -78 °C. After addition, the mixture was stirred at -78 °C for 0.5 h. 4-Bromo-2-(bromomethyl)thiazole (Compound B3, 5.4 g, 21.02 mmol) was added to the above mixture at -78 °C and the mixture was stirred for another 1 h. The reaction was heated with saturated NH 4 Cl solution (100mL) is quenched, and the reaction mixture is extracted with EtOAc (100mL, twice). The combined organic layer is washed with brine (150mL), dried over anhydrous sodium sulfate, filtered, and the filtrate is concentrated under vacuum. The residue is purified by reverse phase chromatography to obtain 4-bromo-2-[[(2S, 5R) -5- isopropyl -3,6- dimethoxy -2,5- dihydropyrazine -2- bases] methyl] thiazole (compound B5, 3.6g) as a yellow oil. MS calculated value 360 ​​(MH + ); measured value 359.9 (MH + ).

[0315] Step 4: Preparation of methyl (2S)-2-amino-3-(4-bromothiazol-2-yl)propanoate (Compound B6)

[0316] To a solution of 4-bromo-2-[[(2S,5R)-5-isopropyl-3,6-dimethoxy-2,5-dihydropyrazin-2-yl]methyl]thiazole (Compound B5, 3.6 g, 10 mmol) in ACN (20 mL) was added hydrochloric acid (66.6 mL, 0.3 M). The mixture was stirred at 25 °C for 2 h. The mixture was washed with saturated NaHCO 3 The solution was basified until pH = 8. The mixture was extracted twice with EtOAc (80 mL). The combined organic layers were dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under vacuum to give (2S)-2-amino-3-(4-bromothiazol-2-yl)propanoic acid methyl ester (Compound B6, 3.1 g) as a yellow oil. MS calculated value 264.9 (MH + ); measured value 264.9 (MH + ).

[0317] Step 5: Preparation of methyl (2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propanoate (Compound B7)

[0318] To a solution of methyl (2S)-2-amino-3-(4-bromothiazol-2-yl)propanoate (compound B6, 3.1 g, 11.69 mmol) in DCM (40 mL) was added triethylamine (2.9 g, 29.23 mmol) and (Boc 2 O (3.8 g, 17.54 mmol). After stirring at 30 ° C for 12 hours, the mixture was concentrated under vacuum. The residue was purified by silica gel column eluted with ethyl acetate (0-30%) in petroleum ether to give (2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propanoic acid methyl ester (Compound B7, 3.2 g) as a yellow oil. MS calculated value 387 (MNa + ); measured value 386.9 (MNa + ).

[0319] Step 6: Preparation of (2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)-propionic acid (Compound B8)

[0320] To a solution of (2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propanoic acid methyl ester (Compound B7, 3.2 g, 8.76 mmol) in THF (30 mL), methanol (2 mL) and water (10 mL) was added lithium hydroxide (0.4 mL, 43.81 mmol). After stirring at 25 °C for 1 hour, the reaction mixture was acidified with 1M HCl solution until pH = 5. The mixture was extracted twice with EtOAc (40 mL). The combined organic layers were washed with brine (100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under vacuum to give (2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propanoic acid (Compound B8, 3.1 g) as a yellow oil. MS calculated value 373 (MNa + ); measured value 372.9 (MNa + ).

[0321] Step 7: Preparation of methyl (3S)-1-[(2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propionyl]hexahydropyridazine-3-carboxylate (Intermediate B)

[0322] To a solution of (2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propionic acid (compound B8, 3.1 g, 8.83 mmol) in DCM (50 mL) was added (3S)-methyl hexahydropyridazine-3-carboxylate hydrochloride (compound B9, 2.4 g, 13.24 mmol), EDCI (3.4 g, 17.65 mmol), 1-hydroxybenzotriazole (238.5 mg, 1.77 mmol) and NMM (9.92 mL, 88.26 mmol) at 0°C. After stirring at 25°C for 1 hour, the reaction mixture was diluted with water (60 mL) and extracted with EtOAc (60 mL, three times). The combined organic layers were washed with brine (100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under vacuum. The residue was purified by silica gel column and eluted with ethyl acetate (10-30%) in petroleum ether to give (3S)-1-[(2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propanoyl]hexahydropyridazine-3-carboxylic acid methyl ester (Intermediate B, 2.4 g). MS calculated value 477 (MH + ), measured value 476.9 (MH + ).

[0323] Intermediate C

[0324] (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]Octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione

[0325]

[0326] The title intermediate C was prepared according to the following scheme:

[0327]

[0328]

[0329] Step 1: Preparation of 1-(5-bromo-6-fluoro-1H-indol-3-yl)-3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropan-1-one (Compound C3)

[0330] To a mixture of 3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropanoyl chloride (Compound C1, 35.0 g, 116.8 mmol) in DCM (400 mL) was slowly added SnCl 4 Solution (97.2 mL, 121.5 mmol). After stirring at -40 °C for 0.5 h, 5-bromo-6-fluoro-1H-indole (compound C2, 25.0 g, 116.8 mmol) in DCM (200 mL) was added dropwise to the mixture stirred at -40 °C for 15 min. After the reaction was complete, saturated NaHCO 3 The aqueous solution (800 mL) was quenched and the reaction mixture was extracted with EtOAc (900 mL, twice). The combined organic layers were washed with brine (700 mL) and purified by Na 2 SO 4 The residue was dried, filtered and concentrated in vacuo. The residue was triturated with a solution (100 mL, petroleum ether: ethyl acetate = 8: 1) and filtered. The filter cake was dried in vacuo to give 1-(5-bromo-6-fluoro-1H-indol-3-yl)-3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropan-1-one (Compound C3, 50.0 g) as a yellow solid. MS calculated value 552.1 (MH + ); measured value 552.1 (MH + ).

[0331] Step 2: Preparation of [3-(5-bromo-6-fluoro-1H-indol-3-yl)-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound C4)

[0332] To a mixture of 1-(5-bromo-6-fluoro-1H-indol-3-yl)-3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropan-1-one (Compound C3, 50.0 g, 90.49 mmol) in THF (600 mL) was added dropwise LiBH 4 (48.4 mL, 193.49 mmol, 4 M in THF). The mixture was stirred at 70 ° C for 24 hours under a nitrogen atmosphere. After the reaction was completed, it was quenched by slowly adding water (600 mL) at 0 ° C, and the reaction mixture was extracted with EtOAc (600 mL, twice). The combined organic layer was washed with brine (600 mL) and washed with Na 2 SO 4Dry, filter and concentrate in vacuo. The residue was purified by silica column chromatography (EtOAc in PE = 20% to 33%) to give [3-(5-bromo-6-fluoro-1H-indol-3-yl)-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound C4, 46.0 g) as a white solid. MS calculated value 538.1 (MH + ); measured value 538.2 (MH + ).

[0333] Step 3: Preparation of [3-(5-bromo-6-fluoro-2-iodo-1H-indol-3-yl)-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound C5)

[0334] To a mixture of [3-(5-bromo-6-fluoro-1H-indol-3-yl)-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound C4, 35.4 g, 65.73 mmol) and iodine (18.4 g, 72.3 mmol) in THF (400 mL) at 0°C was added silver trifluoromethanesulfonate (20.3 g, 78.88 mmol). The mixture was stirred at 0°C for 10 minutes. After the reaction was complete, saturated Na 2 SO 3 The mixture was quenched with aqueous solution (400 mL) and EtOAc (400 mL), and the reaction mixture was filtered. The organic layer was washed with brine (100 mL) and purified by Na 2 SO 4 Dry, filter and concentrate in vacuo. The residue was purified by silica column chromatography (EtOAc in PE = 0% to 2.5%) to give [3-(5-bromo-6-fluoro-2-iodo-1H-indol-3-yl)-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound C5, 43.0 g) as a yellow solid. MS calculated value 664.0 (MH + ); measured value 664.1 (MH + ).

[0335] Step 4: Preparation of 4-[5-[5-bromo-3-[3-[tert-butyl(diphenyl)silyl]oxy-2,2-dimethyl-propyl]-6-fluoro-1H-indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylic acid benzyl ester (Compound C6)

[0336] To a mixture of [3-(5-bromo-6-fluoro-2-iodo-1H-indol-3-yl)-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound C5, 16.7 g, 25.13 mmol) and 4-[6-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-pyridyl]piperazine-1-carboxylic acid benzyl ester (Intermediate A, 16.7 g, 34.69 mmol) in a mixed solution of 1,4-dioxane (270 mL) / toluene (90 mL) / water (90 mL) were added potassium phosphate (15.7 g, 73.92 mmol) and Pd(dppf)Cl 2 (920mg, 1.26mmol). The mixture was stirred at 70°C for 12 hours under a nitrogen atmosphere. After completion of the reaction, the mixture was filtered and concentrated in vacuo. The residue was purified by silica column chromatography (EtOAc in PE = 20% to 50%) to give 4-[5-[5-bromo-3-[3-[tert-butyl(diphenyl)silyl]oxy-2,2-dimethyl-propyl]-6-fluoro-1H-indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridyl]piperazine-1-carboxylate (Compound C6, 19.5g) as a white solid. MS calculated value 891.3 (MH + ); measured value 891.3 (MH + ).

[0337] Step 5: Preparation of 4-[(5M)-5-[5-bromo-3-[3-[tert-butyl(diphenyl)silyl]oxy-2,2-dimethyl-propyl]-6-fluoro-1-(2,2,2-trifluoroethyl)indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylic acid benzyl ester (Compound C7)

[0338] At 0°C, 4-[5-[5-bromo-3-[3-[tert-butyl(diphenyl)silyl]oxy-2,2-dimethyl-propyl]-6-fluoro-1H-indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylate (Compound C6, 14.5 g, 16.26 mmol) and Cs 2 CO 3To a solution of 2,2,2-trifluoroethyl trifluoromethanesulfonate (15.9 g, 48.77 mmol) in DMF (200 mL) was added dropwise 2,2,2-trifluoroethyl trifluoromethanesulfonate (37.7 g, 162.56 mmol), and the mixture was stirred at 20 °C for 12 hours. After the reaction was complete, EtOAc (70 mL) and water (100 mL) were added and the layers were separated. The aqueous phase was extracted with EtOAc (70 mL, twice). The combined organic layers were washed with brine (100 mL, four times) and washed with Na 2 SO 4 The mixture was dried, filtered, and concentrated in vacuo to give a residue. The residue was purified by silica column chromatography to give 4-[(5M)-5-[5-bromo-3-[3-[tert-butyl(diphenyl)silyl]oxy-2,2-dimethyl-propyl]-6-fluoro-1-(2,2,2-trifluoroethyl)indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridyl]piperazine-1-carboxylic acid benzyl ester (Compound C7, 8.0 g, peak 1, faster elution) as a yellow oil. MS calculated value 973.3 (MH + ); measured value 973.2 (MH + ).

[0339] Step 6: Preparation of 4-[(5M)-5-[5-bromo-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-1-(2,2,2-trifluoroethyl)indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylic acid benzyl ester (Compound C8)

[0340] To a solution of 4-[(5M)-5-[5-bromo-3-[3-[tert-butyl(diphenyl)silyl]oxy-2,2-dimethyl-propyl]-6-fluoro-1-(2,2,2-trifluoroethyl)indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylic acid benzyl ester (Compound C7, 10.5 g, 10.78 mmol) in DMF (130 mL) was added cesium fluoride (8.2 g, 53.9 mmol) and the mixture was stirred at 60 °C for 24 hours. After the reaction was complete, EtOAc (100 mL) and water (100 mL) were added and the layers were separated. The aqueous phase was extracted with EtOAc (100 mL, twice). The combined organic layers were washed with brine (80 mL, three times) and purified by Na 2 SO 4Dry, filter, and concentrate under vacuum to give a residue. The residue was purified by silica column chromatography (EtOAc in PE = 25% to 66%) to give 4-[(5M)-5-[5-bromo-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-1-(2,2,2-trifluoroethyl)indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylic acid benzyl ester (Compound C8, 6.5 g) as a yellow solid. MS calculated value 735.2 (MH + ); measured value 735.1 (MH + ).

[0341] Step 7: Preparation of 4-[(5M)-5-[6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1-(2,2,2-trifluoroethyl)indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylic acid benzyl ester (Compound C9)

[0342] To a solution of benzyl 4-[(5M)-5-[5-bromo-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-1-(2,2,2-trifluoroethyl)indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylate (compound C8, 5.4 g), bis(pinacolato)diboron (2.8 g, 11.01 mmol) and potassium acetate (1.2 mL, 18.35 mmol) in toluene (70 mL) was added Pd(dppf)Cl 2 (537.1mg, 0.73mmol). The mixture was degassed and purged with a nitrogen atmosphere three times, and the mixture was stirred at 90°C for 12 hours. After the reaction was completed, the mixture was cooled to room temperature. The reaction mixture was filtered, and the filtrate was concentrated in vacuo to obtain a residue. The residue was purified by silica column chromatography (EtOAc in PE = 25% to 66%) to obtain 4-[(5M)-5-[6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1-(2,2,2-trifluoroethyl)indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridyl]piperazine-1-carboxylic acid benzyl ester (Compound C9, 5.2g) as a yellow oil. MS calculated value 783.3 (MH + ); measured value 783.3 (MH + ).

[0343] Step 8: Preparation of (3S)-1-[(2S)-3-[4-[(2M)-2-[5-(4-benzyloxycarbonylpiperazin-1-yl)-2-[(1S)-1-methoxyethyl]-3-pyridyl]-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]-2-(tert-butoxycarbonylamino)-propionyl]hexahydropyridazine-3-carboxylic acid methyl ester (Compound C10)

[0344] To a mixture of (3S)-1-[(2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propanoyl]hexahydropyridazine-3-carboxylic acid methyl ester (Intermediate B, 2.7 g, 5.69 mmol), 4-[(5M)-5-[6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1-(2,2,2-trifluoroethyl)indol-2-yl]-6-[(1S)-1-methoxyethyl]-3-pyridyl]piperazine-1-carboxylic acid benzyl ester (Compound C9, 4.9 g, 6.32 mmol) in toluene (60 mL) / 1,4-dioxane (20 mL) / water (20 mL) was added K under nitrogen atmosphere. 3 PO 4 (3.4g,15.81mmol) and Pd(dtbpf)Cl 2 (412.2 mg, 0.63 mmol). The mixture was stirred at 70 ° C for 12 hours. After the reaction was completed, the mixture was concentrated in vacuo to give a residue. The residue was purified by silica column (EtOAc in PE = 10% to 75%) to give (3S)-1-[(2S)-3-[4-[(2M)-2-[5-(4-benzyloxycarbonylpiperazine-1-yl)-2-[(1S)-1-methoxyethyl]-3-pyridyl]-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]-2-(tert-butoxycarbonylamino)-propionyl]hexahydropyridazine-3-carboxylic acid methyl ester (Compound C10, 3.6 g) as a brown solid. MS calculated value 1053.4 (MH + ); measured value 1053.3 (MH + ).

[0345] Step 9: Preparation of (3S)-1-[(2S)-3-[4-[(2M)-2-[5-(4-benzyloxycarbonylpiperazin-1-yl)-2-[(1S)-1-methoxyethyl]-3-pyridyl]-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]-2-(tert-butoxycarbonylamino)propanoyl]hexahydropyridazine-3-carboxylic acid (Compound C11)

[0346] To a solution of (3S)-1-[(2S)-3-[4-[(2M)-2-[5-(4-benzyloxycarbonylpiperazin-1-yl)-2-[(1S)-1-methoxyethyl]-3-pyridinyl]-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]-2-(tert-butoxycarbonylamino)-propionyl]-hexahydropyridazine-3-carboxylic acid methyl ester (Compound C10, 3.6 g, 3.42 mmol) in DCE (50 mL) was added trimethyltin alcohol (2.4 g, 13.67 mmol) and the mixture was stirred at 60 ° C for 12 hours. After the reaction was complete, EtOAc (80 mL) and water (60 mL) were added and the layers were separated. The aqueous phase was extracted with EtOAc (80 mL, twice). The combined organic layers were washed with brine (100 mL) and purified by Na 2 SO 4 The mixture was dried, filtered and concentrated under vacuum to give (3S)-1-[(2S)-3-[4-[(2M)-2-[5-(4-benzyloxycarbonylpiperazin-1-yl)-2-[(1S)-1-methoxyethyl]-3-pyridinyl]-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]-2-(tert-butoxycarbonylamino)propanoyl]hexahydropyridazine-3-carboxylic acid (Compound C11, 4.3 g) as a brown solid. MS calculated value 1039.4 (MH + ); measured value 1039.2 (MH + ).

[0347] Step 10: 4-[5-[(7S,13S)-7-(tert-butoxycarbonylamino)-24-fluoro-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9 ,13 .0 22,26Preparation of 1-(25),2,5(28),19,22(26),23-hexaen-(20M)-20-yl)-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylic acid benzyl ester (Compound C12)

[0348] To a mixture of (3S)-1-[(2S)-3-[4-[(2M)-2-[5-(4-benzyloxycarbonylpiperazin-1-yl)-2-[(1S)-1-methoxyethyl]-3-pyridinyl]-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]-2-(tert-butoxycarbonylamino)propanoyl]hexahydropyridazine-3-carboxylic acid (Compound C11, 4.3 g, 4.14 mmol) in DCM (430 mL) was added DIEA (14.4 mL, 82.76 mmol), EDCI (11.9 g, 62.07 mmol) and 1-hydroxybenzotriazole (1.4 g, 10.35 mmol) at 0° C. The mixture was stirred at 15° C. for 12 hours. After the reaction was complete, the mixture was concentrated in vacuo, then diluted with water (80 mL) and extracted with EtOAc (80 mL, twice). The combined organic layers were washed with brine (80 mL) and purified by Na 2 SO 4 The residue was purified by silica column chromatography (EtOAc in PE = 25% to 66%) to give 4-[5-[(7S,13S)-7-(tert-butoxycarbonylamino)-24-fluoro-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1]] as a yellow gum. 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaen-(20M)-20-yl]-6-[(1S)-1-methoxyethyl]-3-pyridinyl]piperazine-1-carboxylic acid benzyl ester (Compound C12, 3.1 g). MS calculated value 1021.4 (MH + ); measured value 1021.2 (MH + ).

[0349] Step 11: N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 Preparation of tert-butyl 1(25),2,5(28),19,22(26),23-hexaen-7-yl)carbamate (Compound C13)

[0350] To 4-[5-[(7S,13S)-7-(tert-butoxycarbonylamino)-24-fluoro-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22 ,26 To a mixture of octacosyl-1(25),2,5(28),19,22(26),23-hexaen-(20M)-20-yl]-6-[(1S)-1-methoxyethyl]-3-pyridyl]piperazine-1-carboxylic acid benzyl ester (Compound C12, 3.1 g, 3.04 mmol) and aqueous formaldehyde solution (775.0 mg, 9.55 mmol) in methanol (150 mL) was added Pd(OH) on activated carbon 2 (2.79 g, 3.97 mmol). The mixture was degassed and washed with H 2 The mixture was purged three times. The mixture was hydrogenated at 30° C. for 18 hours. After the reaction was complete, the mixture was filtered and the filtrate was concentrated in vacuo to give N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1] as a brown solid. 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamic acid tert-butyl ester (Compound C13, 2.6 g). MS calculated value 901.3 (MH + ); measured value 901.3 (MH + ).

[0351] Step 12: (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 Preparation of octadecane-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C)

[0352] To N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 To a mixture of tert-butyl]octadecanoate-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamate (Compound C13, 2.6 g, 2.89 mmol) in DCM (18 mL) was added TFA (14.0 mL, 181.72 mmol). The mixture was stirred at 15 °C for 0.5 h. After the reaction was complete, the mixture was concentrated in vacuo and washed with saturated NaHCO 3 (30 mL) and extracted with EtOAc (30 mL, three times). The combined organic layers were washed with brine (50 mL) and purified by Na 2 SO 4 The mixture was dried, filtered and concentrated in vacuo to give (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1] as a yellow solid. 2,5 .1 9 ,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C, 2.0 g), which was used directly in the next step. MS: calculated value 801.3 (MH +); measured value 801.2 (MH + )

[0353] Intermediate D

[0354] (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]Octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione

[0355]

[0356] The title compound was prepared analogously to the preparation of intermediate C by using iodoethane instead of 2,2,2-trifluoroethyl trifluoromethanesulfonate.

[0357] Intermediate E

[0358] (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]Octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione

[0359]

[0360] The compounds were prepared according to the following scheme:

[0361]

[0362]

[0363] Step 1: Preparation of 1-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridinyl]-4-(2,2,2-trifluoroethyl)piperazine (Compound E2).

[0364] To a mixture of 3-bromo-5-iodo-2-[(1S)-1-methoxyethyl]pyridine (Compound A3, 2.03 g, 5.95 mmol) and 1-(2,2,2-trifluoroethyl)piperazine (Compound E1, 1.0 g, 5.95 mmol) in toluene (15 mL) was added Cs 2 CO 3 (4.85 g, 14.88 mmol), (R)-binap (92.6 mg, 0.15 mmol) and Pd(OAc) 2 (66.8 mg, 0.3 mmol). The reaction mixture was degassed and purged with nitrogen 3 times, and the mixture was stirred at 100 ° C for 12 hours under a nitrogen atmosphere. After cooling to room temperature, the reaction mixture was filtered, and the filtrate was concentrated in vacuo to give a residue. The residue was purified by column chromatography to 1- [5-bromo-6- [(1S) -1-methoxyethyl] -3-pyridinyl] -4- (2,2,2-trifluoroethyl) piperazine (Compound E2, 2.0 g) as a yellow oil. MS: Calculated value 382.2 (MH + ); measured value 382.1 (MH + )

[0365] Step 2: 1-[6-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-pyridinyl]-4-(2,2,2-trifluoroethyl)piperazine (Compound E3).

[0366] To a solution of 1-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridinyl]-4-(2,2,2-trifluoroethyl)piperazine (Compound E2, 3.2 g, 8.37 mmol), bis(pinacolato)diboron (3.19 g, 12.56 mmol) and KOAc (2.1 g, 20.93 mmol) in toluene (50 mL) was added Pd(dppf)Cl 2 (306.3 mg, 0.42 mmol). The mixture was degassed and purged with nitrogen three times and the mixture was stirred at 90 ° C for 12 hours under a nitrogen atmosphere. After cooling to room temperature, the reaction mixture was filtered and the filtrate was concentrated in vacuo to give a residue, which was purified by reverse phase column to give 1-[6-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-pyridinyl]-4-(2,2,2-trifluoroethyl)piperazine (Compound E3, 1.9 g) as a yellow gum. MS calculated value 430.2 (MH + ), measured value 348.4 (MC 6 H 10 +H +).

[0367] Step 3: Preparation of [3-[5-bromo-6-fluoro-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1H-indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound E4).

[0368] To a solution of 1-[6-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-pyridinyl]-4-(2,2,2-trifluoroethyl)piperazine (Compound E3, 1.9 g, 4.41 mmol), [3-(5-bromo-6-fluoro-2-iodo-1H-indol-3-yl)-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound C5, 2.1 g, 3.15 mmol) in 1,4-dioxane (24 mL), water (8 mL) and toluene (8 mL) was added K 3 PO 4 (2.1 g, 9.5 mmol) and Pd(dppf)Cl 2 (231 mg, 0.37 mmol). The mixture was degassed by bubbling nitrogen for 2 minutes, and the reaction mixture was stirred at 70 ° C for 12 hours. After cooling to room temperature, the reaction mixture was filtered. The filtrate was concentrated in vacuo to give a residue. The residue was purified by column chromatography (EtOAc in PE: 30%-60%) to give [3-[5-bromo-6-fluoro-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazine-1-yl]-3-pyridyl]-1H-indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound E4, 960.0 mg) as a yellow gum. MS: Calculated value 839.3 (MH + ); measured value 839.3 (MH + )

[0369] Step 4: Preparation of [3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound E5).

[0370] To a solution of [3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1H-indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound E4, 1 g, 1.14 mmol) in DMF (35 mL) was added Cs 2 CO 3 (1.1 g, 3.44 mmol) and 2,2,2-trifluoroethyl trifluoromethanesulfonate (2.7 g, 11.63 mmol). After stirring at 20 °C for 15 hours, the reaction mixture was poured into water (100 mL) and extracted with EtOAc (50 mL, three times). The combined organic phases were washed with brine (50 mL, three times) and purified by Na 2 SO 4 Dry, filter and concentrate under vacuum to give a residue, which was purified by column chromatography (EtOAc in PE: 30%-40%) to give [3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound E5, 640.0 mg, faster elution) as a white solid. MS calculated value 921.3 (MH + ); measured value 921.4 (MH + ).

[0371] Step 5: Preparation of 3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound E6).

[0372] To a solution of [3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazine-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound E5, 640.0 mg, 0.69 mmol) in DMF (7 mL) was added cesium fluoride (421.8 mg, 2.78 mmol). The mixture was stirred at 60 ° C for 16 hours. After cooling to room temperature, the reaction mixture was filtered and the filtrate was concentrated in vacuo to give a residue. The residue was purified by column chromatography (EtOAc in PE: 30%-60%) to give 3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound E6, 360.0 mg) as a yellow oil. MS calculated 683.2 (MH + ); measured value 683.1 (MH + ).

[0373] Step 6: Preparation of 3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound E7).

[0374] To a solution of 3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound E6, 360.0 mg, 0.53 mmol), bis(pinacolato)diboron (200.6 mg, 0.79 mmol) in toluene (6 mL) were added potassium acetate (0.08 mL, 1.32 mmol) and Pd(dppf)Cl 2(40mg, 0.1mmol). The reaction mixture was degassed by bubbling nitrogen for 5 minutes and then stirred at 80°C for 15 hours. After cooling to room temperature, the reaction mixture was filtered and the filtrate was concentrated in vacuo to give a residue. The residue was purified by column chromatography (EtOAc in PE: 30%-50%) to give 3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazine-1-yl]-3-pyridyl]-1-(2,2,2-trifluoroethyl)indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound E7, 300.0mg) as a yellow jelly. MS calculated value 731.4 (MH + ); measured value 731.4 (MH + ).

[0375] Step 7: Preparation of (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid methyl ester (Compound E8).

[0376] To a mixture of 3-[5-bromo-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound E7, 0.3 g, 0.41 mmol) and (3S)-1-[(2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propanoyl]hexahydropyridazine-3-carboxylic acid methyl ester (Intermediate B, 196.7 mg, 0.41 mmol) in toluene (3 mL), 1,4-dioxane (1 mL) and water (1 mL) was added K 3 PO 4 (221.3 mg, 1.04 mmol) and Pd(dtbpf)Cl 2(27.05mg, 0.04mmol). The mixture was stirred at 70 ° C for 12 hours under a nitrogen atmosphere. After cooling to room temperature, the reaction mixture was filtered, and the filtrate was concentrated in vacuo to obtain a residue. The residue was purified by column chromatography (EtOAc in PE: 60%-80%) to obtain (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazine-1-yl]-3-pyridyl]-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]propionyl]hexahydropyridazine-3-carboxylic acid methyl ester (Compound E8, 200.0mg). MS calculated value 1001.4 (MH + ); measured value 1001.4 (MH + ).

[0377] Step 8: Preparation of (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid (Compound E9).

[0378] To a mixture of (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid methyl ester (Compound E8, 200.0 mg, 0.2 mmol) in DCE (5 mL) was added MeOH. 3 SnOH (200.0 mg, 1.11 mmol). The mixture was stirred at 60 °C for 12 hours. The reaction mixture was concentrated under vacuum to give a residue. EtOAc (10 mL) and water (10 mL) were added to the residue and the layers were separated. The aqueous phase was extracted with EtOAc (15 mL, twice). The combined organic layers were washed with brine (20 mL) and purified by Na 2 SO 4The mixture was dried, filtered, and concentrated in vacuo to give (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid (Compound E9, 188.0 mg) as a brown solid. MS calculated value 987.4 (MH + ); measured value 987.4 (MH + ).

[0379] Step 9: N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 Preparation of tert-butyl]octacosahedral-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamate (Compound E10).

[0380] To a mixture of (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-1-(2,2,2-trifluoroethyl)indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid (Compound E9, 188.0 mg, 0.19 mmol) in DCM (20 mL) at 0°C were added DIEA (0.7 mL, 3.81 mmol), EDCI (550.0 mg, 2.87 mmol) and HOBt (65.0 mg, 0.48 mmol). After stirring at 20°C for 12 hours, the reaction mixture was poured into water (20 mL) and extracted with EtOAc (20 mL, three times). The combined organic layers were washed with brine (30 mL) and purified by Na 2 SO 4The mixture was dried, filtered and concentrated under vacuum to give a residue which was purified by column chromatography (EtOAc in PE: 50%-70%) to give N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1] as a yellow solid. 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamic acid tert-butyl ester (Compound E10, 110.0 mg). MS calculated value 969.4 (MH + ); measured value 969.5 (MH + ).

[0381] Step 10: (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] Preparation of octadeca-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate E).

[0382] To N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26To a solution of tert-butyl]octadecanoyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamate (compound E10, 110.0 mg, 0.11 mmol) in DCM (1 mL) was added TFA (1.0 mL, 12.98 mmol). The mixture was stirred at 20 °C for 1 hour. After the reaction was complete, the reaction mixture was concentrated under vacuum to give a residue. Saturated NaHCO 3 The mixture was added with 4% paraformaldehyde (20 mL) and the mixture was extracted with EtOAc (15 mL, three times). The combined organic layers were washed with brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated in vacuo to afford (7S, 13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1] as a yellow solid. 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate E, 98.0 mg). MS calculated value 869.4 (MH + ); measured value 869.2 (MH + ).

[0383] Intermediate F

[0384] (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]Octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione

[0385]

[0386] The title compound was prepared analogously to the preparation of intermediate E by using iodoethane instead of 2,2,2-trifluoroethyl trifluoromethanesulfonate.

[0387] Intermediate G

[0388] (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo-[17.5.2.1 2,5 .1 9 ,13 .0 22,26 ]Octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione

[0389]

[0390] The compounds were prepared according to the following scheme:

[0391]

[0392]

[0393] Step 1: Preparation of 4-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridinyl]morpholine (Compound G1)

[0394] To a mixture of 3-bromo-5-iodo-2-[(1S)-1-methoxyethyl]pyridine (Compound A3, 30 g, 87.73 mmol) and morpholine (7.6 g, 87.73 mmol) in toluene (450 mL) was added Cs 2 CO 3 (57.2 g, 175.45 mmol), (R)-binap (2.7 g, 4.39 mmol) and Pd(OAc) 2 To 4-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridyl]morpholine (compound G1, 21 g) was added to the mixture (0.98 g, 4.39 mmol). The reaction mixture was degassed and purged with nitrogen 3 times, and the mixture was stirred at 90 ° C for 12 hours under a nitrogen atmosphere. After cooling to room temperature, the reaction mixture was filtered, and the filtrate was concentrated in vacuo to obtain a residue. The residue was purified by column chromatography to obtain 4-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridyl]morpholine (compound G1, 21 g) as a yellow oil. MS calculated value 301.1 (MH + ); measured value 301.1 (MH + ).

[0395] Step 2: Preparation of 4-[6-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-pyridyl]morpholine (Compound G2)

[0396] To a solution of 4-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridyl]morpholine (Compound G1, 21 g, 63.3 mmol), bis(pinacolato)diboron (24.0 g, 94.63 mmol) and KOAc (13.6 g, 138.79 mmol) in toluene (500 mL) was added Pd(dppf)Cl 2 (4.4 g, 6.31 mmol). The mixture was degassed and purged with nitrogen three times and the mixture was stirred at 90 ° C for 12 hours under a nitrogen atmosphere. After cooling to room temperature, the reaction mixture was filtered and the filtrate was concentrated in vacuo to give the crude product 4-[6-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-pyridyl]morpholine (Compound G2, 45 g) as a yellow gum, which was used in the next step. MS calculated value 349.2 (MH + ); measured value 349.2 (MH + ).

[0397] Step 3: Preparation of [3-[5-bromo-6-fluoro-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]-1H-indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound G3)

[0398] To a solution of 4-[6-[(1S)-1-methoxyethyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3-pyridyl]morpholine (Compound G2, 40.6 g, 46.65 mmol), [3-(5-bromo-6-fluoro-2-iodo-1H-indol-3-yl)-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound C5, 31 g, 46.65 mmol) in 1,4-dioxane (420 mL) and water (80 mL) was added K 3 PO 4 (29.7 g, 2.33 mmol) and Pd(dppf)Cl 2 (1.7 g, 0.29 mmol). The mixture was degassed by bubbling nitrogen for 2 minutes, and the reaction mixture was stirred at 90°C for 18 hours. After cooling to room temperature, the reaction mixture was extracted with EA (200 mL, three times). The combined organic layers were washed with brine (200 mL) and purified by Na 2 SO 4The mixture was dried, filtered, and the filtrate was concentrated in vacuo to give a residue. The residue was purified by column chromatography to give [3-[5-bromo-6-fluoro-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-1H-indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound G3, 17.2 g) as a yellow oil. MS calculated value 758.3 (MH + ); measured value 758.3 (MH + ).

[0399] Step 4: Preparation of [3-[5-bromo-1-ethyl-6-fluoro-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound G4)

[0400] To a solution of [3-[5-bromo-6-fluoro-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]-1H-indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound G3, 15 g, 19.77 mmol) in DMF (300 mL) was added Cs 2 CO 3 (19.3 g, 59.3 mmol) and iodoethane (6.16 g, 39.53 mmol). After stirring at 20 °C for 16 hours, the reaction mixture was poured into water (200 mL) and extracted with EtOAc (200 mL, three times). The combined organic layers were washed with brine (10 mL, three times) and purified by Na 2 SO 4 The mixture was dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography to give [3-[5-bromo-1-ethyl-6-fluoro-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (Compound G4, 14.7 g) as a yellow oil. MS calculated value 786.3 (MH + ); measured value 786.4 (MH + ).

[0401] Step 5: Preparation of 3-[5-bromo-1-ethyl-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound G5) and 3-[5-bromo-1-ethyl-6-fluoro-(2P)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound G6)

[0402] To a solution of [3-[5-bromo-1-ethyl-6-fluoro-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-3-yl]-2,2-dimethyl-propoxy]-tert-butyl-diphenyl-silane (compound G4, 14.7 g, 18.68 mmol) in DMF (160 mL) was added cesium fluoride (14.2 g, 93.41 mmol). The mixture was stirred at 60 ° C for 48 hours. After cooling to room temperature, EtOAc (300 mL) and water (300 mL) were added to the reaction mixture, and the layers were separated. The aqueous phase was extracted three times with EtOAc (200 mL). The combined organic layers were washed with brine (200 mL, four times) and washed with Na 2 SO 4 The mixture was dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography to give 3-[5-bromo-1-ethyl-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound G5, 6 g, faster elution) and 3-[5-bromo-1-ethyl-6-fluoro-(2P)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound G6, 4.5 g, slower elution) as a colorless foam. Compound G5: MS calculated value 548.2 (MH + ); measured value 548.2 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.41(d,J=2.4Hz,1H),7.92(d,J=6.8Hz,1H),7.37-7.33(m,2H),4.58(s,1H),4.05-3.98(m,2H),3.87-3.82(m,5H),3.27-3.23(m,4H),3 .15-3.13(m,1H),3.00(s,3H),2.75-2.71(m,1H),2.24-2.22(m,1H),1 .42(d,J=6.4Hz,3H),1.22(t,J=7.2Hz,3H),0.76(s,3H),0.76(s,3H).

[0403] X-ray crystallographic analysis of compound G5

[0404] The absolute configuration structure of compound G5 was confirmed by X-ray crystallography of its single crystal. Figure 1 ).

[0405] Step 6: Preparation of 3-[1-ethyl-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound G7)

[0406] To a solution of 3-[5-bromo-1-ethyl-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-3-yl]-2,2-dimethyl-propan-1-ol (compound G5, 6 g, 10.94 mmol), bis(pinacolato)diboron (4.2 g, 16.41 mmol) in toluene (60 mL) were added potassium acetate (2.7 g, 27.35 mmol) and Pd(dppf)Cl 2 (0.8 g, 1.09 mmol). The reaction mixture was degassed by bubbling nitrogen for 5 minutes and then stirred at 90 ° C for 15 hours. After cooling to room temperature, the reaction mixture was filtered and the filtrate was concentrated in vacuo to obtain a residue. The residue was purified by column chromatography to obtain 3- [1- ethyl -6- fluoro - (2M) -2- [2- [(1S) -1- methoxyethyl] -5- morpholino -3- pyridyl] -5- (4,4,5,5- tetramethyl -1,3,2- dioxaborolan -2-yl) indol -3-yl] -2,2- dimethyl -propan-1-ol (compound G7, 4.5 g) as a colorless jelly. MS calculated value 596.4 (MH + ); measured value 596.4 (MH + ).

[0407] Step 7: Preparation of (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[1-ethyl-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid methyl ester (Compound G8)

[0408] To a mixture of 3-[1-ethyl-6-fluoro-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)indol-3-yl]-2,2-dimethyl-propan-1-ol (Compound G7, 4.5 g, 7.56 mmol) and (3S)-1-[(2S)-3-(4-bromothiazol-2-yl)-2-(tert-butoxycarbonylamino)propanoyl]hexahydropyridazine-3-carboxylic acid methyl ester (Intermediate B, 3.6 g, 7.56 mmol) in toluene (45 mL), 1,4-dioxane (15 mL) and water (15 mL) was added K 3 PO 4 (4.0g,18.89mmol) and Pd(dtbpf)Cl 2 (492.5mg, 0.75mmol). The mixture was stirred at 70 ° C for 12 hours under a nitrogen atmosphere. After cooling to room temperature, the reaction mixture was filtered, and the filtrate was concentrated in vacuo to obtain a residue. The residue was purified by column chromatography to obtain (3S) -1- [(2S) -2- (tert-butoxycarbonylamino) -3- [4- [1-ethyl -6-fluoro-3- (3-hydroxy-2,2-dimethyl-propyl) - (2M) -2- [2- [(1S) -1-methoxyethyl] -5-morpholino -3-pyridyl] indol -5-yl] thiazol-2-yl] propionyl] hexahydropyridazine -3- carboxylic acid methyl ester (compound G8, 3.8g) as a colorless jelly. MS calculated value 866.4 (MH + ); measured value 866.4 (MH + ).

[0409] Step 8: Preparation of (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[1-ethyl-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid (Compound G9)

[0410] To a mixture of (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[1-ethyl-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid methyl ester (Compound G8, 3.8 g, 4.39 mmol) in DCE (76 mL) was added MeOH. 3 SnOH (3.2 g, 17.55 mmol). The mixture was stirred at 60 °C for 48 hours. The reaction mixture was concentrated under vacuum to give a residue. EtOAc (200 mL) and water (100 mL) were added to the residue and the layers were separated. The aqueous phase was extracted with EtOAc (150 mL, twice). The combined organic layers were washed with brine (200 mL) and washed with Na 2 SO 4 The mixture was dried, filtered, and concentrated in vacuo to give (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[1-ethyl-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid (Compound G9, 3.7 g) as a brown solid. MS calculated value 852.4 (MH + ); measured value 852.4 (MH + ).

[0411] Step 9: N-[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 Preparation of tert-butyl 1(25),2,5(28),19,22(26),23-hexaen-7-yl)carbamate (Compound G10)

[0412] To a mixture of (3S)-1-[(2S)-2-(tert-butoxycarbonylamino)-3-[4-[1-ethyl-6-fluoro-3-(3-hydroxy-2,2-dimethyl-propyl)-(2M)-2-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]indol-5-yl]thiazol-2-yl]propanoyl]hexahydropyridazine-3-carboxylic acid (compound G9, 2.5 g, 2.93 mmol) in DCM (250 mL) was added DIEA (7.58 mL, 58.68 mmol), EDCI (8.4 g, 44.01 mmol) and HOBt (991.2 mg, 0.91 mmol) at 0° C. After stirring at 20° C. for 12 hours, the reaction mixture was poured into water (100 mL) and extracted with EtOAc (100 mL, three times). The combined organic layers were washed with brine (30 mL) and purified by Na 2 SO 4 The mixture was dried, filtered and concentrated under vacuum to give a residue which was purified by column chromatography to give N-[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1] as a yellow oil. 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamic acid tert-butyl ester (Compound G10, 1.2 g). MS calculated value 834.4 (MH + ); measured value 834.4 (MH + ).

[0413] Step 10: (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridinyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 Preparation of octacosa-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate G)

[0414] To N-[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 To a solution of tert-butyl]octadecanoyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamate (compound G10, 1.2 g, 1.44 mmol) in DCM (12 mL) was added TFA (6.0 mL). The mixture was stirred at 20 °C for 3 hours. After the reaction was complete, the reaction mixture was concentrated under vacuum to obtain a residue. Saturated NaHCO 3 Aqueous solution (60 mL), and the mixture was extracted with EtOAc (80 mL, three times). The combined organic layers were washed with brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated in vacuo to afford (7S, 13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1] as a yellow solid. 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate G, 1 g). MS calculated value 734.3 (MH + ); measured value 734.3 (MH + ).

[0415] Intermediate H

[0416] (7S,13S)-7-amino-25-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]Octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione

[0417]

[0418] The title compound was prepared in analogy to the preparation of intermediate C by using 5-bromo-4-fluoro-1H-indole instead of 5-bromo-6-fluoro-1H-indole (compound C2).

[0419] Intermediate I

[0420] tert-Butyl cis-3-ethynylcyclobutanecarboxylate

[0421]

[0422] The compounds were prepared according to the following scheme:

[0423]

[0424] Step 1: cis-Cyclobutane-1,3-dicarboxylic acid O 1 -tert-Butyl Ester O 3 - Preparation of methyl ester (Compound I2)

[0425] To a mixture of cis-3-methoxycarbonylcyclobutanecarboxylic acid (Compound I1, 25.0 g, 158.08 mmol) and DMAP (38.6 g, 316.16 mmol) in tert-butanol (450 mL) was added dropwise Boc in tert-butanol (50 mL). 2 O (37.9 g, 173.89 mmol). After stirring at 25 ° C for 0.5 hours, the reaction mixture was diluted with water (150 mL) and extracted with EtOAc (150 mL, three times). The combined organic layer was washed with brine (300 mL) and purified by Na 2 SO 4 Dry, filter and concentrate in vacuo. The obtained residue was purified by silica column (EtOAc in PE = 5% to 10%) to give cis-cyclobutane-1,3-dicarboxylic acid O as a colorless liquid. 1 -tert-Butyl Ester O 3 -methyl ester (Compound I2, 29.0 g). MS calculated value 215.1 (MH + ); measured value 215.1 (MH + ).

[0426] Step 2: Preparation of cis-3-(hydroxymethyl)-cyclobutanecarboxylic acid tert-butyl ester (Compound I3) In a nitrogen atmosphere at 0°C, add cis-cyclobutane-1,3-dicarboxylic acid O 1 -tert-Butyl Ester O 3To a mixture of -methyl ester (Compound I2, 29.0 g, 140.02 mmol) in THF (290 mL) was added lithium borohydride (9.2 g, 420.05 mmol). After stirring at 25 °C for 2 hours, EtOAc (800 mL) and water (200 mL) were added to the reaction mixture and the layers were separated. The aqueous phase was extracted three times with EtOAc (300 mL). The combined organic layers were washed with brine (500 mL) and purified by Na 2 SO 4 Dry, filter, and concentrate under vacuum to give a residue. The residue was purified by silica column (EtOAc in PE = 10% to 30%) to give cis-3-(hydroxymethyl)-cyclobutanecarboxylic acid tert-butyl ester (Compound I3, 19 g) as a colorless liquid. MS calculated value 186.1 (MH + ); measured value 186.1 (MH + ).

[0427] Step 3: Preparation of tert-butyl cis-3-formylcyclobutanecarboxylate (Compound I4)

[0428] To a mixture of cis-3-(hydroxymethyl)-cyclobutanecarboxylic acid tert-butyl ester (Compound I3, 19.0 g, 102.01 mmol) in DCM (230 mL) was added DMAP (51.9 g, 122.42 mmol) at 0°C. After stirring at 25°C for 1 hour, the reaction mixture was filtered. The filtrate was concentrated in vacuo and purified by a silica column (EtOAc in PE = 5% to 20%) to give cis-3-formylcyclobutanecarboxylic acid tert-butyl ester (Compound I4, 15.0 g) as a colorless oil. MS calculated value 184.1 (MH + ); measured value 184.1 (MH + ).

[0429] Step 4: Preparation of tert-butyl cis-3-ethynylcyclobutanecarboxylate (Intermediate I)

[0430] To a mixture of cis-3-formylcyclobutanecarboxylic acid tert-butyl ester (compound I4, 15.0 g, 81.42 mmol) and potassium carbonate (22.5 g, 162.84 mmol) in methanol (200 mL) was added (1-diazo-2-oxopropyl)-phosphonic acid dimethyl ester (compound I5, 23.5 g, 122.13 mmol) at 0 ° C. The mixture was stirred at 25 ° C for 3 hours. The reaction mixture was concentrated in vacuo to remove the solvent and diluted with water (100 mL) and extracted with ethyl acetate (100 mL, twice). The combined organic layer was washed with brine (100 mL), dried over anhydrous sodium sulfate, filtered and concentrated under vacuum to obtain a residue. The resulting residue was purified by silica gel chromatography to obtain cis-3-ethynylcyclobutanecarboxylic acid tert-butyl ester (intermediate I, 7 g) as a colorless oil. MS calculated value 180.1 (MH + ); measured value 180.1(MH + ).

[0431] Intermediate J

[0432] (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-3-pyridinyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]Octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione

[0433]

[0434] The title compound was prepared in analogy to the preparation of intermediate G by using 3-bromo-2-[(1S)-1-methoxyethyl]pyridine (compound A1) instead of 4-[5-bromo-6-[(1S)-1-methoxyethyl]-3-pyridyl]morpholine (compound G1).

[0435] Intermediate K

[0436] (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26]Octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione

[0437]

[0438] The title intermediate was prepared analogously to the preparation of intermediate G by using 2,2,2-trifluoroethyl trifluoromethanesulfonate instead of iodoethane.

[0439] Example 1

[0440] trans-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide

[0441]

[0442] The compounds were prepared according to the following scheme:

[0443]

[0444] Step 1: trans-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 Preparation of octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide

[0445] At 0°C, trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butanoic acid (Compound II, 70.0 mg, 0.23 mmol), (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] To a solution of octadecacarbon-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C, 100.0 mg, 0.12 mmol), EDCI (50.0 mg, 0.26 mmol) and DIEA (0.1 mL, 0.62 mmol) in DMF (2 mL) was added HOBT (34.0 mg, 0.25 mmol). After stirring at 16 ° C for 1 hour, the reaction mixture was poured into water (20 mL) and extracted with EtOAc (20 mL, three times). The combined organic layer was washed with brine (30 mL) and purified by Na 2 SO 4 The mixture was dried, filtered, and concentrated under vacuum to give a residue. The resulting residue was purified by preparative HPLC to give N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1] as an off-white solid. 2 ,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide (Example 1, 34.8 mg). MS calculated value 1088.5 (MH + ); measured value 1088.5 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.68(d,J=7.2Hz,1H),8.50(d,J=3.2Hz,1H),7.69(d,J=2.4Hz,1H),7.54-7.43(m,2H),5.72-5.65(m,1H),5.26-5. 10(m,1H),4.82-4.75(m,1H),4.46-4.38(m,1H),4.27-4.18(m,2H),4.12-3.86(m,2H),3.82-3.56(m,5H),3.54-3.41( m,2H),3.37-3.33(m,4H),3.30-3.23(m,2H),3.21-3.06(m,2H),3.02-2.91(m,6H),2.87-2.68(m,3H),2.65-2.16(m,6 H),2.05-1.55(m,4H),1.45(d,J=6.0Hz,3H),1.35-1.25(m,1H),1.03-0.94(m,5H),0.91-0.82(m,3H),0.44(s,3H)ppm.

[0446] Compound 1I was prepared according to the following scheme:

[0447]

[0448] Step 1: Preparation of methyl 3-(methoxymethylene)cyclobutanecarboxylate (Compound 1B)

[0449] To a solution of (methoxymethyl)triphenylphosphonium chloride (267.5 g, 780.46 mmol) in THF (1.6 L) was slowly added potassium tert-butoxide (87.6 g, 780.46 mmol) at 0 ° C and warmed to 20 ° C. After 1.5 hours, methyl 3-oxocyclobutanecarboxylate (Compound 1A, 50.0 g, 390.23 mmol) was added to the reaction mixture. After stirring at 70 ° C for 3 hours, the reaction mixture was concentrated under vacuum to obtain a residue. A mixed solution of PE in EtOAc (10: 1, 1.1 L) was added to the residue. After stirring at 20 ° C for 0.5 hours, the suspension was filtered and the filtrate was concentrated under vacuum to obtain a residue. The residue was purified by column chromatography (EtOAc in PE: 0%-10%) to give methyl 3-(methoxymethylene)cyclobutanecarboxylate (Compound 1B, 18.0 g) as a yellow oil. 1 H NMR (400 MHz, chloroform-d) δ: 5.85-5.79 (m, 1H), 3.70 (s, 3H), 3.60 (s, 3H), 3.29-3.09 (m, 1H), 3.01-2.90 (m, 2H), 2.89-2.71 (m, 2H) ppm.

[0450] Step 2: Preparation of methyl 3-formylcyclobutanecarboxylate (Compound 1C)

[0451] To a solution of methyl 3-(methoxymethylene)cyclobutanecarboxylate (Compound 1B, 26.0 g, 166.47 mmol) in DCM (300 mL) and water (30 mL) was added TFA (26.0 mL). The reaction mixture was stirred at 20 °C for 3 hours. After the reaction was complete, H 2 O (600 mL), then extracted with DCM (100 mL, three times). The organic layer was washed with brine (500 mL) and purified by Na 2 SO 4 Dry, filter, and concentrate under vacuum to give methyl 3-formylcyclobutanecarboxylate (Compound 1C, 18.0 g, 126.63 mmol) as a yellow oil. 1 H NMR (400 MHz, chloroform-d) δ: 9.84-9.52 (m, 1H), 3.75-3.63 (m, 3H), 3.32-3.20 (m, 1H), 3.18-3.07 (m, 1H), 2.67-2.38 (m, 4H) ppm.

[0452] Step 3: Preparation of methyl 3-ethynylcyclobutanecarboxylate (Compound 1D)

[0453] A solution of methyl 3-formylcyclobutanecarboxylate (Compound 1C, 10.0 g, 70.35 mmol) in methanol (120 mL) was cooled to 0°C, and then dimethyl (1-diazo-2-oxopropyl) phosphonate (21.0 g, 109.31 mmol) and potassium carbonate (20.0 g, 144.71 mmol) were added to the reaction mixture. After stirring at 20°C for 3 hours, H 2 O (150 mL), and then extracted with PE (60 mL, twice). The combined organic layers were washed with brine (80 mL), 2 SO 4 Dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography (EtOAc in PE: 0% to 25%) to give methyl 3-ethynylcyclobutanecarboxylate (Compound ID, 6.0 g) as a colorless oil. 1 HNMR (400 MHz, chloroform-d) δ: 3.75-3.62 (m, 3H), 3.42-3.21 (m, 1H), 3.07-2.89 (m, 1H), 2.65-2.33 (m, 4H), 2.23-2.17 (m, 1H) ppm.

[0454] Step 4: Preparation of 3-ethynylcyclobutanecarboxylic acid (Compound 1E)

[0455] To a solution of methyl 3-ethynylcyclobutanecarboxylate (Compound 1D, 6.0 g, 43.43 mmol) in THF (10 mL) and water (30 mL) was added lithium hydroxide (3.6 g, 86.86 mmol) at 0° C., and the solution was then stirred at 20° C. for 3 hours. After the reaction was complete, the reaction mixture was concentrated under vacuum to remove THF, and then H 2 O (60 mL) and extracted with MTBE (30 mL). The MTBE phase was discarded and the pH of the aqueous phase was acidified to pH = 5 with aqueous HCl (1N, 60 mL) and extracted with EtOAc (60 mL, three times). The combined organic layers were washed with brine (80 mL) and purified by Na 2 SO 4 Dried, filtered, and concentrated in vacuo to give 3-ethynylcyclobutanecarboxylic acid (Compound IE, 3.8 g) as a colorless oil. 1 H NMR (400 MHz, chloroform-d) δ: 12.14-9.87 (m, 1H), 3.36-3.15 (m, 1H), 3.10-2.95 (m, 1H), 2.68-2.53 (m, 2H), 2.51-2.35 (m, 2H), 2.22 (dd, J = 15.2, 2.4 Hz, 1H) ppm.

[0456] Step 5: Preparation of trans-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1F) and cis-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1G)

[0457] To a solution of 3-ethynylcyclobutanecarboxylic acid (Compound 1E, 3.8 g, 30.61 mmol) in DMF (50 mL) was added DIEA (19.0 mL, 114.96 mmol), HATU (14.3 g, 37.48 mmol). After stirring at 0°C for 10 minutes, tert-butyl (2S)-3-methyl-2-(methylamino)butanoate (5.7 g, 30.44 mmol) was added to the reaction mixture. The reaction mixture was stirred at 0°C for another 1 hour. After the reaction was complete, H was added to the reaction mixture. 2 O (120 mL), then extracted with EtOAc (40 mL, three times). The combined organic layers were washed with brine (60 mL) and purified by Na 2 SO 4The residue was dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography (EtOAc in PE: 9%-16%) and preparative HPLC (column: Welch Ultimate XB-CN 250×50×10 μm; mobile phase: hexane-EtOH (0.1% FA); B%: 1%-20%, 15 minutes) to give cis-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1G, faster elution, 3g) and trans-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1F, slower elution, 2g) as a yellow oil.

[0458] Cis-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1G, peak 1). MS calculated value 294.2 (MH + ); measured value 294.1 (MH + ). 1 HNMR (400 MHz, chloroform-d) δ = 4.80 (d, J = 10.4 Hz, 0.5H), 3.58 (d, J = 10.8 Hz, 0.5H), 3.27-3.11 (m, 1H), 3.01-2.91 (m, 1H), 2.87 (d, J = 6.8 Hz, 3H), 2.59-2.40 (m, 4H), 2.25-2.12 (m, 2H), 1.45 (s, 9H), 1.00 (dd, J = 14.4, 6.4 Hz, 3H), 0.84 (dd, J = 6.8, 1.2 Hz, 3H) ppm. The stereochemistry of compound 1G was confirmed by 2D-NMR.

[0459] trans-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1F, peak 2). MS calculated value 294.2 (MH + ); measured value 294.1 (MH + ). 1HNMR (400 MHz, chloroform-d) δ = 4.80 (d, J = 10.4 Hz, 0.5H), 3.69-3.46 (m, 1.5H), 3.11-3.01 (m, 1H), 2.88 (d, J = 3.2 Hz, 3H), 2.73-2.60 (m, 2H), 2.40-2.27 (m, 2H), 2.25-2.11 (m, 2H), 1.45 (d, J = 2.8 Hz, 9H), 1.01 (dd, J = 12.0, 6.8 Hz, 3H), 0.84 (dd, J = 6.8, 1.6 Hz, 3H) ppm. The stereochemistry of compound 1F was confirmed by 2D-NMR.

[0460] Step 6: Preparation of trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid tert-butyl ester (Compound 1H)

[0461] CuI (408.9 mg, 2.15 mmol), K 2 CO 3 A suspension of 1,4-dimethoxy-2-nitropropene (593.5 mg, 4.29 mmol) and TMEDA (249.5 mg, 2.15 mmol) in DMF (10 mL) was stirred at 25 °C for 20 minutes under argon atmosphere. 3 (407.1 mg, 2.86 mmol) was added to the reaction and the reaction mixture was stirred for 10 minutes under argon atmosphere. 3 (407.09 mg, 2.86 mmol) and a solution of trans-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1F, 420.0 mg, 1.43 mmol) in DMF (10 mL) were added to the reaction. The reaction mixture was stirred at 0 ° C for 30 minutes and allowed to warm to 25 ° C. After stirring at 25 ° C for another 12 hours, H was added to the reaction mixture. 2 O (30 mL), then extracted with EtOAc (10 mL, three times). The combined organic layers were washed with brine (50 mL) and purified by Na 2 SO 4 The mixture was dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by reverse phase chromatography and preparative HPLC to give trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butanoic acid tert-butyl ester (Compound 1H, 80.0 mg) as a yellow oil. MS calculated value 362.2 (MH + ); measured value 362.1 (MH + ). 1H NMR (400 MHz, chloroform-d) δ = 4.80 (d, J = 10.0 Hz, 0.5 H), 3.62-3.46 (m, 1.5 H), 3.28-3.13 (m, 1 H), 2.89 (d, J = 4.4 Hz, 3 H), 2.82-2.67 (m, 2 H), 2.48-2.38 (m, 2 H), 2.29-2.15 (m, 1 H), 1.46 (d, J = 2.8 Hz, 9 H), 1.05-0.98 (m, 3 H), 0.85 (d, J = 6.8 Hz, 3 H) ppm.

[0462] Step 7: Preparation of trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I)

[0463] To a solution of trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid tert-butyl ester (Compound 1H, 80.0 mg, 0.22 mmol) in DCM (1 mL) was added TFA (1.0 mL), and the mixture was stirred at 20 ° C for 1 hour. After the reaction was completed, the reaction mixture was concentrated under vacuum to give trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I, 80.0 mg) as a yellow oil, which was used directly in the next step. MS calculated value 306.0 (MH + ); measured value 306.0 (MH + ).

[0464] Example 2

[0465] N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide

[0466]

[0467] The title compound was prepared similarly to the preparation of Example 1 by using (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid (Compound 2E) instead of trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I). Example 2 (78.9 mg) was obtained as a yellow solid. MS calculated value 1089.5 (MH + ), measured value 1089.7 (MH + ). 1 H NMR (400 MHz, methanol-d 4 )δ=8.68(d,J=7.2Hz,1H),8.50(d,J=2.8Hz,1H),7.70(d,J=2.0Hz,1H),7.51(d,J=2.4Hz,1H),7.48(d,J=12.8Hz,1H),5.71(t,J=8.8Hz,1H ),5.23-5.13(m,1H),4.47-4.38(m,2H),4.37-4.11(m,6H),4.10-3.93(m,3H),3.81-3.77(m,1H),3.76-3.59(m,4H),3.47(d,J=14.8Hz,1H ),3.35(s,3H),3.29-3.23(m,2H),3.19-3.10(m,2H),2.99(s,3H),2.87(s,3H),2.84-2.79(m,1H),2.57(d,J=14.4Hz,1H),2.25-2.16(m,2 H),2.01-1.92(m,1H),1.87-1.77(m,1H),1.70-1.58(m,1H),1.45(d,J=6.0Hz,3H),1.39-1.25(m,1H),1.00-0.85(m,10H),0.44(s,3H)ppm.

[0468] Compound 2E was prepared according to the following scheme:

[0469]

[0470] Step 1: Preparation of 3-ethynylazetidine (Compound 2B)

[0471] To a solution of tert-butyl 3-ethynylazetidine-1-formate (compound 2A, 3.5 g, 19.31 mmol) in DCM (36 mL) was added TFA (17.7 g, 155.76 mmol). The reaction mixture was stirred at 20 ° C for 1 hour. After the reaction was completed, the reaction mixture was evaporated and co-evaporated with DCM (20 mL) three times to obtain 3-ethynylazetidine (compound 2B, 3.5 g, crude product, TFA salt) as a yellow oil, which was used directly in the next step without purification.

[0472] Step 2: Preparation of (2S)-2-[(3-ethynylazetidine-1-carbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 2C)

[0473] To a mixture of (2S)-3-methyl-2-(methylamino)butyric acid tert-butyl ester (3.7 g, 19.76 mmol) in DCM (50 mL) was added DIEA (8.5 mL, 48.8 mmol) and triphosgene (2.1 g, 7.08 mmol). After stirring at 0 °C for 10 minutes, a mixture of 3-ethynylazetidine; 2,2,2-trifluoroacetic acid (compound 2B, 3.5 g, 17.94 mmol) and DIEA (13.0 mL, 74.64 mmol) in DCM (50 mL) was added to the reaction. The resulting mixture was stirred at 20 °C for another hour. After the reaction was complete, saturated NaHCO was added to the reaction mixture. 3 The solution was stirred for 2 hours and then extracted with EtOAc (100 mL, twice). The combined organic layers were washed with brine (600 mL) and purified by Na 2 SO 4 The mixture was dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography (EtOAc in PE: 11%-25%) to give (2S)-2-[(3-ethynylazetidine-1-carbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 2C, 3.2 g) as a yellow oil. MS calculated value 239.2 (MC 4 H 9 +H + ), measured value 239.0 (MC 4 H 9 +H + ).

[0474] Step 3: Preparation of (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid tert-butyl ester (Compound 2D)

[0475] A mixture of CuI (1.5 g, 8.15 mmol), potassium carbonate (2.3 g, 16.34 mmol) and TMEDA (947.3 mg, 8.15 mmol) in DMF (30 mL) was stirred at 20 °C for 20 minutes under argon atmosphere. 3 (1.5 g, 10.87 mmol) was added to the reaction mixture, and then stirred at 20° C. for 20 minutes. (2S)-2-[(3-ethynylazetidine-1-carbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 2C, 1.6 g, 5.43 mmol) and TMSCF 3 A mixture of (1.5 g, 10.87 mmol) in DMF (30 mL) was added to the reaction mixture. After stirring at 20 °C for another 12 h under argon atmosphere, H 2 O (100 mL), then extracted with EtOAc (30 mL, three times). The combined organic layers were washed with brine (150 mL) and purified by Na 2 SO 4 Dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography (EtOAc in PE: 11% to 25%) and concentrated under vacuum to give a residue. The residue was purified again by reverse phase HPLC to give (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid tert-butyl ester (Compound 2D, 600.0 mg) as a yellow oil. MS calculated value 307.2 (MC 4 H 9 +H + ), measured value 307.1(MC 4 H 9 +H + ).

[0476] Step 4: Preparation of (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid (Compound 2E)

[0477] To a solution of (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid tert-butyl ester (compound 2D, 300.0 mg, 0.83 mmol) in DCM (2 mL) was added TFA (2.6 g, 23.36 mmol). The reaction mixture was stirred at 20 ° C for 1 h. After the reaction was completed, the reaction mixture was concentrated under vacuum to obtain a residue. The residue was co-evaporated with DCM (6 mL, three times) to obtain (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid (compound 2E, 250.0 mg) as a yellow oil, which was used directly in the next step without purification. MS calculated value 307.1 (MH + ); measured value 307.0 (MH + ).

[0478] Example 3

[0479] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide

[0480]

[0481] The title compound was prepared similarly to the preparation of Example 1 by using cis-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 3B) instead of trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I). Example 3 (202.7 mg) was obtained as a yellow solid. MS calculated value 1088.5 (MH + ); measured value 1088.5 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.68(d,J=7.6Hz,1H),8.50(d,J=2.8Hz 1H),8.67(d,J=2.4Hz 1H),7.52-7.43(m,2H),5.74-5.61(m,1H),5.21-5.12(m,1H),4.94-4.86(m,2H),4.83-4.81(m,1H),4.78(d,J=11.2Hz,1H),4.46- 4.39(m,1H),4.26-4.19(m,2H),4.11-3.90(m,2H),3.84-3.64(m,4H),3.62-3.43(m,4H),3.38-3.33(m,4H),3.18-3.08(m,2H),3. 00(s,3H),2.98-2.93(m,3H),2.84-2.75(m,1H),2.74-2.63(m,2H),2.61-2.44(m,3H),2.27-2.16(m,2H),2.01-1.93(m,1H),1.86 -1.76(m,1H),1.70-1.59(m,1H),1.45(d,J=6.0Hz,3H),1.10-1.00(m,1H),1.00-0.95(m,5H),0.90-0.83(m,3H),0.44(s,3H)ppm.

[0482] Similar to the preparation of compound 1I, compound 3B was prepared by using cis-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (compound 1G) instead of trans-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (compound 1F).

[0483] Example 4

[0484] N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)benzamide

[0485]

[0486] The compounds were prepared according to the following scheme:

[0487]

[0488] Step 1: N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 Preparation of tert-butyl]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-carbamate (Compound 4A)

[0489] To a mixture of BOC-N-ME-VAL-OH (93.8 mg, 0.41 mmol) and DIEA (0.2 mL, 0.94 mmol) in DMF (1 mL) were added HATU (154.3 mg, 0.41 mmol) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[175.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C, 250.0 mg, 0.31 mmol). After stirring at 15 °C for 1 hour, the reaction mixture was purified by reverse phase chromatography to give N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1] as a yellow solid. 2,5 .1 9,13 .0 22,26]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-carbamic acid tert-butyl ester (Compound 4A, 300.0 mg). MS calculated value 1014.4 (MH + ); measured value 1014.3 (MH + ).

[0490] Step 2: (2S)-N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 Preparation of octadecanoic acid-1(25),2,5(28),19,22(26),23-hexaen-7-yl]-3-methyl-2-(methylamino)butanamide (Compound 4B)

[0491] To N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 To a mixture of 1-(2-(2-(2-(dioctadecyl)-1(25),2,5(28),19,22(26),23-hexaen-7-yl)carbamoyl]-2-methyl-propyl]-N-methyl-tert-butyl carbamate (compound 4A, 300.0 mg, 0.3 mmol) in DCM (6 mL) was added TFA (4.0 mL). After stirring at 15 °C for 0.5 hours, the reaction mixture was concentrated in vacuo to give a residue. The resulting residue was washed with saturated NaHCO 3 The solution was diluted with aqueous solution (30 mL) and extracted with EtOAc (20 mL, three times). The combined organic layers were washed with brine (30 mL) and purified by Na 2 SO 4The mixture was dried, filtered and concentrated in vacuo to give (2S)-N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1] as a yellow oil. 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]-3-methyl-2-(methylamino)butanamide (Compound 4B, 270.0 mg) was used directly in the next step. MS calculated value 914.4 (MH + ); measured value 914.3 (MH + ).

[0492] Step 3: N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 Preparation of ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)benzamide (Example 4)

[0493] To (2S)-N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26To a solution of 1-(25-hydroxyoctadecane), 1-(25), 2-(5-hydroxyoctadecane), 1-(28-hydroxyoctadecane), 1-(26-hydroxyoctadecane), 2-(23-hydroxyoctadecane)-3-methyl-2-(methylamino)butanamide (compound 4B, 210 mg, 0.23 mmol) and 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid (compound 4C, 49.2 mg, 0.23 mmol) in DMF (0.5 mL) was added DIEA (0.1 mL, 0.38 mmol) and T 3 P (146.2 mg, 0.23 mmol). After stirring at 20 ° C for 0.5 hours, the reaction mixture was added to water (40 mL) and extracted with EtOAc (30 mL, three times). The combined organic layers were washed with brine (30 mL, three times), dried over anhydrous sodium sulfate, filtered and concentrated under vacuum to give a residue, which was purified by preparative HPLC to give N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)benzamide (Example 4, 9.8 mg). MS calculated value 1110.4 (MH + ); measured value 1110.6 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.69(d,J=9.2Hz,1H),8.47-8.43(m,1H),7.79(d,J=8.4Hz,2H),7.73-7.69(m,1H),7.58(d,J=8.0Hz,2H),7.49-7.44 (m,1H),7.39-7.33(m,1H),5.78-5.71(m,1H),5.19-5.10(m,1H),4.60-4.41(m,1H),4.32-4.15(m,2H),3.80-3.70(m,1H) ,3.53-3.40(m,5H),3.23-3.08(m,4H),2.97(s,3H),2.95-2.90(m,3H),2.84-2.71(m,2H),2.62-2.52(m,4H),2.46-2.14 (m,3H),2.08-1.53(m,4H),1.43(d,J=6.0Hz,3H),1.29(s,3H),1.06(t,J=6.4Hz,5H),0.97(s,3H),0.48-0.41(m,3H)ppm.

[0494] Compound 4B was prepared according to the following scheme:

[0495]

[0496] Step 1: Preparation of methyl 4-(3,3,3-trifluoroprop-1-ynyl)benzoate (Compound 8B)

[0497] A suspension of copper (I) iodide (1.8 g, 9.37 mmol), potassium carbonate (2.6 g, 18.77 mmol), and TMEDA (1.1 g, 9.37 mmol) in DMF (10 mL) was stirred at 20° C. under an argon atmosphere. After stirring for 20 minutes, TMSCF was added to the reaction mixture. 3 (1.8 g, 12.49 mmol). Methyl 4-ethynylbenzoate (Compound 4D, 1.0 g, 6.24 mmol) and TMSCF 3 A mixture of (1.8 g, 12.49 mmol) in DMF (10 mL) was slowly added to the reaction mixture, and then it was stirred at 20 °C for another 12 hours under argon atmosphere. After the reaction was completed, H 2 O (200 mL), and extracted with EtOAc (80 mL, three times). The combined organic layers were washed with brine (80 mL), 2 SO 4 The product was dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography (SiO 2, EtOAc in PE: 0%-95%) to give methyl 4-(3,3,3-trifluoroprop-1-ynyl)benzoate (Compound 4D, 233.0 mg) as a colorless solid. 1 H NMR (400 MHz, chloroform-d) δ = 8.08 (d, J = 8.4 Hz, 2H), 7.64 (d, J = 8.4 Hz, 2H), 3.95 (s, 3H) ppm.

[0498] Step 2: Preparation of 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid (Compound 4C)

[0499] Lithium hydroxide monohydrate (69.5 mg, 1.66 mmol) was added to a solution of 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid methyl ester (compound 4D, 180.0 mg, 0.79 mmol) in THF (1 mL) and water (1 mL). The mixture was stirred for 1 hour at 20 ° C. After the reaction was completed, the pH of the reaction mixture was adjusted to 6 with 1M HCl aqueous solution and extracted with EtOAc (10 mL, three times). The combined organic layer was concentrated under vacuum to obtain 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid (compound 4C, 168.0 mg) as a white solid, which was used directly in the next step.

[0500] Example 5

[0501] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(2-pyrimidin-2-ylethynyl)cyclobutanecarboxamide

[0502]

[0503] The title compound was prepared similarly to the preparation of Example 1 by using (2S)-3-methyl-2-[methyl-[cis-3-(2-pyrimidin-2-ylethynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 5B) instead of trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I). Example 4 (1.3 mg) was obtained as a yellow solid. MS calculated value 1098.5 (MH + ); measured value 1098.5 (MH + ). 1 H NMR (400 MHz, chloroform-d) δ=8.74-8.64 (m, 2H), 8.52 (d, J=2.8 Hz, 1H), 7.72-7.61 (m, 1H), 7.55-7.46 (m, 1H), 7.25-7.20 (m, 1H), 7.18-7.09 (m, 2H), 5.74-5.62 (m, 1H), 5.13-4.88 (m, 1H), 4.79-4.68 (m, 1H), 4.63-4.41 (m, 2H), 4.35-4.16 (m, 2H), 4.13-3.93 (m, 1H), 3.89-3 .77(m,2H),3.77-3.69(m,2H),3.66-3.53(m,3H),3.52-3.46(m,1H),3.39-3.33(m,3H),3.32-3.21(m,2H),3.20-3.07(m,3H),2.92 -2.87(m,3H),2.87-2.82(m,2H),2.80(s,1H),2.73-2.61(m,3H),2.43-2.37(m,1H),2.36-2.29(m,1H),2.26-2.20(m,1H),2.14(br s,1H),2.05-2.00(m,1H),1.98-1.89(m,4H),1.60-1.57(m,1H),1.49-1.45(m,3H),1.38-1.35(m,1H),1.28-1.25(m,3H),1.17(br dd,J=6.9,19.4Hz,2H),1.06(br d,J=6.5Hz,2H),1.11-0.76(m,5H)ppm.

[0504] Compound 5B was prepared according to the following scheme:

[0505]

[0506] Step 1: Preparation of tert-butyl (2S)-3-methyl-2-[methyl-[cis-3-(2-pyrimidin-2-ylethynyl)cyclobutanecarbonyl]amino]butanoate (Compound 5A)

[0507] To a solution of cis-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1G, 100.0 mg, 0.34 mmol) in THF (1 mL) was added triethylamine (0.2 mL, 1.02 mmol), 2-iodopyrimidine (70.2 mg, 0.34 mmol), tetrakis(triphenylphosphine)palladium(0) (39.4 mg, 0.03 mmol) and CuI (6.5 mg, 0.03 mmol). The reaction mixture was degassed and purged with nitrogen three times and stirred at 50 °C for 1 hour. After the reaction was complete, H was added to the reaction mixture. 2 O (60 mL), then extracted with EtOAc (40 mL, twice). The combined organic layers were washed with brine (60 mL) and purified by Na 2 SO 4 The mixture was dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by preparative HPLC to give (2S)-3-methyl-2-[methyl-[cis-3-(2-pyrimidin-2-ylethynyl)cyclobutanecarbonyl]amino]butanoic acid tert-butyl ester (Compound 5A, 60 mg) as a yellow oil. MS calculated value 372 (MH + ), measured value 372(MH + ).

[0508] Step 2: Preparation of (2S)-3-methyl-2-[methyl-[cis-3-(2-pyrimidin-2-ylethynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 5B)

[0509] To a solution of tert-butyl (2S)-3-methyl-2-[methyl-[cis-3-(2-pyrimidin-2-ylethynyl)cyclobutanecarbonyl]amino]butanoate (Compound 5A, 60.0 mg, 0.16 mmol) in DCM (1 mL) was added TFA (0.2 mL). After stirring at 20°C for 1 hour, the reaction mixture was concentrated in vacuo to give (2S)-3-methyl-2-[methyl-[cis-3-(2-pyrimidin-2-ylethynyl)cyclobutanecarbonyl]amino]butanoic acid (Compound 5B, 50 mg). The crude product was used in the next step. MS calculated value 316 (MH + ), measured value 316 (MH + ).

[0510] Example 6

[0511] cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide

[0512]

[0513] Similar to the preparation of Example 1, by using cis-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 3B) and (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate D) was substituted for trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 6 (32.2 mg) was obtained as a yellow solid. MS calculated value 1034.5 (MH + ); measured value 1034.5 (MH + ). 1H NMR (400 MHz, methanol-d 4 )δ=8.66(d,J=7.6Hz,1H),8.48(d,J=2.8Hz,1H),7.62(d,J=2.4Hz,1H),7.52(d,J=2.8Hz,1H),7.33(d,J=12.8Hz,1H),5.78-5 .67(m,1H),4.78-4.74(m,2H),4.48-4.36(m,1H),4.33-4.26(m,1H),4.24-3.96(m,4H),3.81-3.64(m,3H),3.56-3.42(m,3H), 3.34(s,4H),3.29-3.24(m,3H),3.12-2.89(m,8H),2.89-2.80(m,1H),2.80-2.31(m,6H),2.31-2.12(m,2H),1.98-1.87(m,1H) ,1.87-1.75(m,1H),1.75-1.54(m,1H),1.43(d,J=6.0Hz,3H),1.08-0.88(m,9H),0.85(d,J=6.4Hz,3H),0.58-0.41(m,3H)ppm.

[0514] Example 7

[0515] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide

[0516]

[0517] Similar to the preparation of Example 1, by using cis-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butanoic acid (Compound 3B) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22 ,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate E) was substituted for trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 7 (30.8 mg) was obtained as a white solid. MS calculated value 1156.5 (MH + ); measured value 1157.1 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.71(d,J=7.6Hz,1H),8.41(d,J=2.8Hz,1H),7.67(d,J=2.4Hz,1H),7.58(s,1H),7.47(d,J=12.4Hz,1H),5.70(t,J=8.8Hz,1 H),5.24-5.06(m,2H),4.80-4.74(m,2H),4.48-4.35(m,1H),4.30-4.15(m,2H),3.80-3.69(m,2H),3.62-3.47(m,2H),3.47-3.39 (m,5H),3.38-3.35(m,3H),3.22-3.11(m,3H),2.98-2.82(m,8H),2.72-2.60(m,3H),2.51-2.46(m,1H),2.28-2.16(m,2H),2.01- 1.91(m,1H),1.89-1.74(m,1H),1.69-1.58(m,1H),1.46(d,J=6.0Hz,3H),1.10-1.01(m,1H),1.01-0.83(m,9H),0.49(s,3H)ppm.

[0518] Example 8

[0519] cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide

[0520]

[0521] Similar to the preparation of Example 1, by using cis-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 3B) and (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate F) was substituted for trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9 ,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 8 (23.3 mg) was obtained as a white solid. MS calculated value 1102.5 (MH + ); measured value 1102.7 (MH + ). 1 HNMR (400 MHz, methanol-d 4)δ=8.65(d,J=7.2Hz,1H),8.40(d,J=2.8Hz,1H),7.61(d,J=2.4Hz,1H),7.33-7.27(m,2H),5.90-5.68(m,1H),4.89(s,1H),4.83-4.8 2(m,1H),4.77(d,J=11.2Hz,1H),4.47-4.37(m,1H),4.26-4.12(m,4H),3.82-3.42(m,5H),3.36-3.33(m,4H),3.27(s,1H),3.17-3.11 (m,2H),3.02(d,J=14.4Hz,1H),2.97-2.91(m,3H),2.90-2.81(m,5H),2.74-2.60(m,3H),2.60-2.36(m,2H),2.29-2.12(m,2H),1.99- 1.91(m,1H),1.88-1.75(m,1H),1.69-1.57(m,1H),1.42(d,J=6.0Hz,3H),1.17-0.88(m,10H),0.85(d,J=6.4Hz,3H),0.50(s,3H)ppm.

[0522] Example 9

[0523] trans-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxamide

[0524]

[0525] The title compound was prepared similarly to the preparation of Example 4 by using trans-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxylic acid (Compound 9g) instead of 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid (Compound 4C). Example 9 (30.8 mg) was obtained as a white solid. MS calculated value 1116.5 (MH + ); measured value 1116.7 (MH +). 1 H NMR (400 MHz, methanol-d 4 )δ=8.68(d,J=7.2Hz,1H),8.53-8.48(m,1H),7.70(d,J=2.4Hz,1H),7.51-7.45(m,2H),5.68(t,J=8.4Hz,1H),5.21-5. 14(m,1H),4.80(d,J=10.8Hz,1H),4.46-4.37(m,1H),4.25-4.18(m,2H),4.15-3.87(m,2H),3.81-3.76(m,1H),3.73-3. 64(m,2H),3.45(d,J=14.8Hz,1H),3.35(s,3H),3.27-3.23(m,1H),3.16(s,1H),3.09(s,3H),3.00(s,3H),2.88-2.76(m ,2H),2.61-2.53(m,2H),2.26-1.84(m,12H),1.66-1.56(m,4H),1.50-1.43(m,6H),1.02-0.83(m,9H),0.44(s,3H)ppm.

[0526] Compound 9g was prepared according to the following scheme:

[0527]

[0528] Step 1: Preparation of trans-cyclohexane-1,4-dicarboxylic acid 4-tert-butyl 1-methyl ester (Compound 9b)

[0529] To a solution of trans-4-methoxycarbonylcyclohexanecarboxylic acid (compound 9a, 5.0 g, 26.85 mmol) in tert-butyl alcohol (100 mL) was added 4-dimethylaminopyridine (6.6 g, 53.7 mmol), and then di-tert-butyl dicarbonate (6.5 g, 29.54 mmol) was slowly added to the reaction mixture at 20°C. The reaction mixture was stirred at 20°C for 2 hours. After the reaction was completed, H 2 O (500 mL), and extracted with EtOAc (80 mL, three times). The combined organic layers were washed with brine (400 mL), 2 SO 4 Dry, filter, and concentrate under vacuum to give a residue. The residue was purified by column chromatography (EtOAc in PE: 0%-10%) to give 4-tert-butyl 1-methyl trans-cyclohexane-1,4-dicarboxylate (Compound 9b, 6.5 g) as a white solid. 1H NMR (400 MHz, chloroform-d) δ = 3.67 (s, 3H), 2.32-2.23 (m, 1H), 2.21-2.12 (m, 1H), 2.05-1.96 (m, 4H), 1.44-1.42 (m, 13H) ppm.

[0530] Step 2: Preparation of trans-4-(hydroxymethyl)cyclohexanecarboxylic acid tert-butyl ester (Compound 9c)

[0531] To a solution of 4-tert-butyl 1-methyl trans-cyclohexane-1,4-dicarboxylate (compound 9b, 5.0 g, 20.63 mmol) in THF (100 mL) was slowly added LiBH 4 (1.4 g, 64.28 mmol). The reaction mixture was stirred at 20 °C for another 12 h. After the reaction was complete, it was heated at 0 °C with H 2 O (500 mL) was slowly quenched, and the reaction mixture was extracted with EtOAc (100 mL, three times). The combined organic layers were washed with brine (600 mL) and purified by Na 2 SO 4 Dry, filter, and concentrate under vacuum to give a residue. The residue was purified by column chromatography (EtOAc in PE: 10%-30%) to give trans-tert-butyl 4-(hydroxymethyl)cyclohexanecarboxylate (Compound 9c, 3.8 g) as a yellow oil. 1 H NMR (400 MHz, chloroform-d) δ: 3.47 (d, J = 6.0 Hz, 2H), 2.18-2.08 (m, 1H), 2.03-1.94 (m, 2H), 1.91-1.79 (m, 2H), 1.58-1.26 (m, 14H) ppm.

[0532] Step 3: Preparation of trans-4-formylcyclohexanecarboxylic acid tert-butyl ester (Compound 9d)

[0533] DMP (11.28 g, 26.6 mmol) was slowly added to a solution of trans-4-(hydroxymethyl)cyclohexanecarboxylic acid tert-butyl ester (compound 9c, 3.8 g, 17.73 mmol) in DCM (100 mL) at 0 ° C. The reaction mixture was stirred for 1 hour at 20 ° C. After the reaction was completed, the reaction mixture was concentrated under vacuum to directly remove DCM to obtain a residue. The residue was purified by column chromatography (EtOAc in PE = 10% to 30%) and concentrated under vacuum to obtain trans-4-formylcyclohexanecarboxylic acid tert-butyl ester (compound 9d, 2.5 g) as a yellow oil. 1H NMR (400 MHz, chloroform-d) δ = 9.63 (d, J = 1.2 Hz, 1H), 2.33-1.93 (m, 6H), 1.59-1.20 (m, 13H) ppm.

[0534] Step 4: Preparation of trans-4-ethynylcyclohexanecarboxylic acid tert-butyl ester (Compound 9e)

[0535] To a solution of trans-4-formylcyclohexanecarboxylic acid tert-butyl ester (Compound 9d, 2.5 g, 11.78 mmol) in methanol (50 mL) was added potassium carbonate (3.5 g, 25.32 mmol). The reaction mixture was cooled to 0 ° C, and dimethyl (1-diazo-2-oxopropyl)phosphonate (3.5 g, 18.22 mmol) was slowly added thereto. The reaction mixture was stirred at 20 ° C for another 3 hours. After the reaction was completed, H was added to the reaction mixture. 2 O (200 mL), then extracted with PE (60 mL, twice). The combined organic layers were washed with brine (800 mL) and purified by Na 2 SO 4 Dried, filtered, and concentrated under vacuum to give a residue. The residue was purified by column chromatography (EtOAc in PE: 0%-16%) to give trans-4-ethynylcyclohexanecarboxylic acid tert-butyl ester (Compound 9e, 1.5 g) as a white solid. 1 H NMR (400 MHz, chloroform-d) δ = 2.29-1.94 (m, 7H), 1.49-1.34 (s, 13H) ppm.

[0536] Step 5: Preparation of trans-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxylic acid tert-butyl ester (Compound 9f)

[0537] To a solution of TMEDA (450 mg, 3.87 mmol) in DMF (15 mL) was added CuI (740 mg, 3.6 mmol) and potassium carbonate (1 g, 7.24 mmol) at 25° C. After vigorous stirring at 25° C. for 20 minutes, TMSCF was added to the reaction mixture under an argon atmosphere. 3 (700 mg, 4.92 mmol) was then stirred at 25°C for another 20 minutes. The reaction mixture was cooled to 0°C, and tert-butyl trans-4-ethynylcyclohexanecarboxylate (Compound 9e, 500.0 mg, 2.4 mmol) and TMSCF were added thereto. 3(700mg, 4.92mmol) in DMF (10mL). The reaction mixture was stirred at 0 ° C for 30 minutes and allowed to warm to 25 ° C for 12 hours. After the reaction was completed, the reaction mixture was poured into water (60mL) and extracted with ethyl acetate (50mL, three times). The combined organic layer was washed with brine (80mL, three times), dried over anhydrous sodium sulfate, filtered and concentrated under vacuum to obtain a residue, which was purified by column chromatography (EtOAc in PE: 0%-10%) to obtain trans-4-(3,3,3-trifluoroprop-1-ynyl) tert-butyl cyclohexanecarboxylate (compound 9f, 500mg, 1.48mmol) as a white solid. 1 H NMR (400 MHz, chloroform-d) δ = 2.17-1.94 (m, 6H), 1.48-1.34 (m, 13H) ppm.

[0538] Step 6: Preparation of trans-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxylic acid (Compound 9g)

[0539] To a solution of trans-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxylic acid tert-butyl ester (compound 9f, 100.0 mg, 0.36 mmol) in DCM (1 mL) was added TFA (1.0 mL). The reaction mixture was stirred at 20 ° C for 0.5 h. After the reaction was completed, the reaction mixture was concentrated under vacuum to obtain a residue. The residue was co-evaporated three times with DCM (5 mL) to obtain trans-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxylic acid (compound 9g, 50.0 mg) as a white solid, which was used directly in the next step without purification.

[0540] Example 10

[0541] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxamide

[0542]

[0543] The title compound was prepared similarly to the preparation of Example 4 by using cis-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxylic acid (Compound 10g) instead of 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid (Compound 4C). Example 10 (39.7 mg) was obtained as a white solid. MS calculated value 1116.5 (MH + ); measured value 1116.4 (MH + ). 1 H NMR (400 MHz, methanol-d 4 )δ=8.74-8.65(m,1H),8.52-8.49(m,1H),7.72-7.64(m,1H),7.53-7.4 2(m,2H),5.75-5.61(m,1H),5.26-5.12(m,1H),4.99-4.92(m,1H),4.50 -4.35(m,1H),4.28-4.17(m,2H),4.17-3.88(m,2H),3.82-3.74(m,1H) ,3.73-3.51(m,3H),3.48-3.43(m,1H),3.37-3.34(m,3H),3.29-3.25(m ,1H),3.17-3.11(m,1H),3.09-3.03(m,3H),3.01-2.97(m,3H),2.90-2 .78(m,2H),2.57(d,J=14.8Hz,1H),2.45-2.31(m,1H),2.29-2.11(m,2H ),2.05-1.89(m,4H),1.88-1.71(m,10H),1.70-1.57(m,2H),1.45(d,J=6.0Hz,3H),1.14-0.91(m,6H),0.86(d,J=6.8Hz,3H),0.44(s,3H)ppm.

[0544] Compound 10g was prepared similarly to the preparation of compound 9g by using cis-4-methoxycarbonylcyclohexanecarboxylic acid (compound 10a) instead of trans-4-methoxycarbonylcyclohexanecarboxylic acid (compound 9a).

[0545] Example 11

[0546] cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .19,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide

[0547]

[0548] Similar to the preparation of Example 1, by using cis-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 3B) and (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate G) was substituted for trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 11 (39.6 mg) was obtained as a yellow solid. MS calculated value 1021.5 (MH + ); measured value 1021.5 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.71(d,J=7.6Hz,1H),8.39(d,J=2.8Hz,1H),7.88-7.82(m,1H),7.62(d,J=2.4Hz,1H),7.40-7.32(m,1H),5.84-5.72(m,1H),4.84- 4.72(m,1H),4.47-4.34(m,2H),4.32-4.22(m,1H),4.20-4.12(m,1H),4.10-3.98(m,1H),3.87(t,J=4.8Hz,4H),3.81-3.69(m,2H),3.5 3-3.36(m,9H),3.29-3.18(m,2H),3.08-2.97(m,1H),2.95-2.87(m,3H),2.83-2.58(m,4H),2.55-2.36(m,2H),2.27-2.12(m,2H),2.00 -1.91(m,1H),1.86-1.74(m,1H),1.71-1.56(m,1H),1.46(d,J=6.4Hz,3H),1.07-0.94(m,9H),0.89-0.82(m,3H),0.68-0.49(m,3H)ppm.

[0549] Example 12

[0550] cis-N-[(1S)-1-[[(7S,13S)-25-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide

[0551]

[0552] Similar to the preparation of Example 1, by using cis-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 3B) and (7S,13S)-7-amino-25-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate H) was substituted with trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9 ,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 12 (7.7 mg) was obtained as a white solid. MS calculated value 1088.5 (MH + ); measured value 1088.5 (MH + ). 1 HNMR (400 MHz, methanol-d 4)δ=8.54-8.50(m,1H),7.54-7.42(m,4H),5.84-5.96(m,1H),5.20-5.10(m,1H),4.68-4.60(m,1H),4.39(d,J=12Hz,1H),4.1 2-3.96(m,2H),3.75-3.60(m,3H),3.57-3.46(m,2H),3.45-3.38(m,1H),3.29-3.23(m,5H),3.22-3.12(m,5H),3.03-3.07(m, 1H),3.00(s,3H),2.98-2.95(m,3H),2.93-2.75(m,2H),2.74-2.53(m,4H),2.50-2.34(m,2H),2.25-2.16(m,1H),1.71-1.59( m,1H),1.47-1.43(m,3H),1.38-1.14(m,3H),0.98-0.92(m,3H),0.85-0.78(m,6H),0.76-0.68(m,1H),0.65-0.55(m,2H)ppm.

[0553] Example 13

[0554] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-[2-[5-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxamide

[0555]

[0556] The title compound was prepared similarly to the preparation of Example 4 by using cis-3-[2-[5-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxylic acid (Compound 13B) instead of 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid (Compound 4C). Example 13 (10.8 mg) was obtained as a white solid. MS calculated value 1166.5 (MH + ); measured value 1166.4 (MH + ).1 H NMR (400 MHz, methanol-d 4 )δ=9.07(s,2H),8.66(d,J=7.3Hz,1H),8.49(d,J=2.9Hz,1H),7.67(s,1H),7.46-7.38(m,2H),5.65-5.61(m,1H),5.15(br d,J=7.8Hz,1H),4.68-4.56(m,1H),4.46-4.37(m,1H),4.26-4.19(m,2H),3.80-3.67(m,3H),3.61(q,J=7.2Hz,5H),3.48-3.39(m,6H),3.15(br d,J=14.7Hz,1H),2.96(d,J=7.3Hz,6H),2.83-2.76(m,2H),2.70-2.66(m,1H ),2.60-2.52(m,2H),2.22(dt,J=3.2,7.5Hz,2H),1.99-1.93(m,1H),1.81(br s,1H),1.68-1.60(m,1H),1.45-1.41(m,3H),1.32-1.27(m,1H),1.18(t,J= 6.8Hz,4H),1.01-0.93(m,6H),0.87(d,J=6.4Hz,3H),0.45-0.40(s,3H)ppm.

[0557] Analogously to the preparation of compound 5B, compound 13B was prepared by using cis-3-ethynylcyclobutanecarboxylic acid tert-butyl ester (Intermediate I) and 2-iodo-5-(trifluoromethyl)-pyrimidine instead of cis-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1G) and 2-iodopyrimidine.

[0558] Example 14

[0559] N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide

[0560]

[0561] Similar to the preparation of Example 1, by using (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid (Compound 2E) and (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate D) was substituted for trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22 ,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 14 (41.2 mg) was obtained as a yellow solid. MS calculated value 1035.5 (MH + ), measured value 1035.5(MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.66(d,J=7.6Hz,1H),8.48(d,J=2.8Hz,1H),7.65(d,J=2.4Hz,1H),7.50(d,J=2.8Hz,1H),7.33(d,J=12.8Hz,1H), 5.79-5.74(m,1H),4.46-4.13(m,12H),4.08-4.03(m,2H),3.77-3.63(m,6H),3.49-3.46(m,1H),3.45-3.43(m,1H),3. 09-3.01(m,2H),3.00(s,3H),2.95(d,J=2.4Hz,2H),2.85(s,3H),2.81-2.74(m,1H),2.64-2.57(m,1H),2.32-2.14(m, 3H),1.99-1.92(m,1H),1.86-1.76(m,1H),1.68-1.60(m,1H),1.43(d,J=6.4Hz,3H),0.98-0.92(m,12H),0.50(s,3H).

[0562] Example 15

[0563] N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide

[0564]

[0565] Similar to the preparation of Example 1, by using (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid (Compound 2E) and (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.12,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate F) was substituted for trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9 ,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 15 (43.2 mg) was obtained as a yellow solid. MS calculated value 1103.5 (MH + ), measured value 1103.5(MH + ). 1 H NMR (400 MHz, methanol-d 4 )δ=8.70(d,J=7.2Hz,1H),8.39-8.36(m,1H),7.80-7.76(m,1H),7.68-7.62(m,1H),7.38-7.32(m,1H),5.85-5.78( m,1H),4.40-4.14(m,8H),4.10-4.02(m,2H),3.78-3.68(m,3H),3.49-3.45(m,4H),3.45-3.42(s,1H),3.19-3.14( m,2H),2.94-2.85(m,6H),2.83(s,4H),2.75-2.69(m,1H),2.22-2.14(m,2H),2.01-1.92(m,1H),1.86-1.75(m,1H) ,1.69-1.57(m,1H),1.45(d,J=6.4Hz,3H),1.34-1.23(m,1H),1.05-0.99(m,3H),0.97-0.90(m,11H),0.58(s,3H).

[0566] Example 16

[0567] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-[2-[4-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxamide

[0568]

[0569] The title compound was prepared similarly to the preparation of Example 4 by using cis-3-[2-[4-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxylic acid (Compound 16B) instead of 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid (Compound 4C). Example 16 (98.2 mg) was obtained as a white solid. MS calculated value 1166.5 (MH + ); measured value 1166.4 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=9.06-8.99(m,1H),8.71-8.63(m,1H),8.47-8.39(m,1H),7.82-7.73(m,1H),7. 71-7.64(m,1H),7.46-7.26(m,2H),5.77-5.60(m,1H),5.20-5.02(m,3H),4.48-4. 09(m,2H),3.82-3.74(m,1H),3.72-3.66(m,1H),3.60-3.53(m,1H),3.52-3.46(m, 1H),3.46-3.42(m,1H),3.41-3.38(m,1H),3.38-3.34(m,4H),3.17-3.08(m,1H),3 .03-2.92(m,3H),2.91-2.84(m,1H),2.83-2.77(m,1H),2.76-2.69(m,2H),2.66-2 .60(m,5H),2.60-2.54(m,2H),2.36(s,3H),2.29-2.15(m,2H),2.01-1.90(m,1H), 1.86-1.70(m,1H),1.69-1.53(m,1H),1.46-1.38(m,3H),1.32-1.25(m,1H),1.20- 1.02(m,1H),1.00-0.94(m,5H),0.92-0.82(m,4H),0.81-0.69(m,1H),0.40(s,3H).

[0570] Similar to the preparation of compound 5B, compound 16B was prepared by using cis-3-ethynylcyclobutanecarboxylic acid tert-butyl ester (Intermediate I) and 2-bromo-4-(trifluoromethyl)pyrimidine instead of cis-(2S)-2-[(3-ethynylcyclobutanecarbonyl)-methyl-amino]-3-methyl-butyric acid tert-butyl ester (Compound 1G) and 2-iodopyrimidine.

[0571] Example 18

[0572] N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide

[0573]

[0574] Similar to the preparation of Example 1, by using (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid (Compound 2E) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22 ,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate E) was substituted for trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 18 (24.8 mg) was obtained as a yellow solid. MS calculated value 1157.5 (MH + ); measured value 1157.5 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.71(d,J=7.2Hz,1H),8.41(d,J=2.8Hz,1H),7.73-7.64(m,2H),7.48(d,J=12.4Hz,1H),5.79-5.72(m,1H ),5.23-5.16(m,1H),4.44-4.28(m,4H),4.25-3.99(m,4H),3.83-3.73(m,1H),3.72-3.69(m,1H),3.50-3.42 (m,5H),3.40-3.36(s,3H),3.25-3.14(m,3H),2.91-2.82(m,9H),2.66(d,J=14.4Hz,1H),2.24-2.16(m,2H), 1.99-1.91(m,1H),1.91-1.68(m,1H),1.68-1.55(m,1H),1.48-1.45(m,3H),0.99-0.91(m,11H),0.51(s,3H).

[0575] Example 19

[0576] (2S)-N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]-2-isopropyl-4-oxo-4-[3-(3,3,3-trifluoroprop-1-ynyl)azetidin-1-yl]butanamide

[0577]

[0578] The title compound was prepared similarly to the preparation of Example 4 by using (2S)-4-tert-butoxy-2-isopropyl-4-oxo-butyric acid and 3-(3,3,3-trifluoroprop-1-ynyl)azetidine (Compound 19B) instead of BOC-N-ME-VAL-OH and 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid (Compound 4C). Example 19 (30.6 mg) was obtained as a white solid. MS calculated value 1074.4 (MH + ); measured value 1074.4 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.67(d,J=7.6Hz,1H),8.50(d,J=2.8Hz 1H),7.76-7.68(m,1H),7.55(s,1H),7.50-7.44(m,1H),5.78-5.66(m, 1H),5.22-5.13(m,1H),4.98-4.89(m,2H),4.57-4.48(m,1H),4.46-4.3 9(m,1H),4.35-4.27(m,1H),4.26-4.16(m,3H),4.14-4.04(m,1H),4.0 4-3.99(m,1H),3.98-3.92(m,1H),3.82-3.76(m,1H),3.74-3.62(m,3H) ,3.59-3.52(m,1H),3.50-3.39(m,2H),3.38-3.33(m,4H),3.21-3.09( m,2H),3.00(s,3H),2.86-2.70(m,2H),2.64-2.44(m,2H),2.26-2.14(m ,2H),2.00-1.88(m,2H),1.86-1.73(m,1H),1.70-1.58(m,1H),1.45(d, J=6.0Hz,3H),1.26-1.06(m,1H),1.03-0.92(m,9H),0.53-0.41(m,3H).

[0579] Compound 19B was prepared similarly to the preparation of compound 9g by using tert-butyl 3-ethynylazetidine-1-carboxylate (compound 2A) instead of tert-butyl trans-4-ethynylcyclohexanecarboxylate (compound 9e).

[0580] Example 20

[0581] N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide

[0582]

[0583] Similar to the preparation of Example 1, by using (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid (Compound 2E) and (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate G) was substituted for trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 20 (8.8 mg) was obtained as a yellow solid. MS calculated value 1022.5 (MH + ); measured value 1022.5 (MH + ). 1 H NMR (400 MHz, DMSO-d 6 )δ=8.55(d,J=7.6Hz,1H),8.50-8.39(m,2H),7.67(d,J=2.4Hz,1H),7.56(d,J=12.8Hz,1H),7.30(d,J=1.6Hz,1H),5.53-5.41(m,1H),5.14(d, J=12.0Hz,1H),4.31-4.21(m,4H),4.21-4.05(m,5H),4.05-3.98(m,2H ),3.81-3.70(m,6H),3.57(s,2H),3.26-3.23(m,3H),3.20(s,3H),2.93

[0584] -2.84(m,1H),2.78-2.68(m,4H),2.11-2.03(m,2H),1.88-1.67(m,2H),1.58

[0585] -1.45(m,1H),1.33(d,J=6.0Hz,3H),1.23(s,1H),0.95-0.83(m,10H),0.79(d,J=6.8Hz,3H),0.37(s,3H).

[0586] Example 21

[0587] cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-[2-[4-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxamide

[0588]

[0589] Similar to the preparation of Example 4, by using cis-3-[2-[4-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxylic acid (Compound 16B) and (7S,13S)-7-amino-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo-[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate G) was substituted with 4-(3,3,3-trifluoroprop-1-ynyl)benzoic acid (Compound 4C) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .19,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 21 (22.8 mg) was obtained as a white solid. MS calculated value 1099.5 (MH + ); measured value 1099.5 (MH + ). 1 H NMR (400 MHz, methanol-d 4 )δ=9.03(d,J=5.2Hz,1H),8.66-8.61(m,1H),8.40(d,J=2.8Hz,1H),7.78(d,J=5.0H z,1H),7.62(d,J=2.4Hz,1H),7.32(d,J=2.8Hz,1H),7.25(d,J=12.8Hz,1H),5.71(br d,J=9.0Hz,1H),4.92-4.88(m,1H),4.80-4.76(m,1H),4.47-4.38(m,1H),4.27-4.22(m,1H),4.21-4.16( m,2H),3.88-3.84(m,4H),3.78-3.69(m,2H),3.56-3.48(m,1H),3.46-3.36(m,2H),3.28-3.26(m,3H),3.0 5-2.92(m,5H),2.86-2.52(m,8H),2.25-2.17(m,2H),1.99-1.92(m,1H),1.88-1.74(m,1H),1.68-1.57(m ,1H),1.42(d,J=6.4Hz,3H),1.34-1.28(m,1H),1.12-1.02(m,1H),0.99-0.84(m,12H),0.53-0.45(m,3H).

[0590] Example 22

[0591] cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide

[0592]

[0593] Similar to the preparation of Example 1, by using (2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carbonyl]amino]butyric acid (Compound 2E) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate K) was substituted for trans-(2S)-3-methyl-2-[methyl-[3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarbonyl]amino]butyric acid (Compound 1I) and (7S,13S)-7-amino-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridyl]-17,17-dimethyl-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ] octacosyl-1(25),2,5(28),19,22(26),23-hexaene-8,14-dione (Intermediate C) to prepare the title compound. Example 22 (41.5 mg) was obtained as a white solid. MS calculated value 1075.4 (MH + ); measured value 1075.4 (MH + ). 1 H NMR (400 MHz, methanol-d 4)δ=8.80-8.63(d,J=7.2Hz,1H),8.44-8.36(d,J=2.8Hz,1H),7.71-7.61(m,2H),7. 53-7.42(d,J=12.8Hz,1H),5.76-5.63(m,1H),5.27-5.11(m,2H),4.80-4.72(m,1H) ,4.49-4.36(m,1H),4.34-4.13(m,2H),3.94-3.84(m,4H),3.83-3.76(m,1H),3.75 -3.67(m,1H),3.58-3.43(m,2H),3.39-3.35(m,6H),3.29-3.23(m,1H),3.17-3.09( m,1H),2.98-2.90(m,3H),2.88-2.77(m,1H),2.75-2.60(m,3H),2.56-2.43(m,2H) ,2.30-2.15(m,2H),2.04-1.91(m,1H),1.90-1.74(m,1H),1.72-1.56(m,1H),1.51- 1.43(d,J=6.0Hz,3H),1.41-1.34(d,J=6.4Hz,1H),1.22-1.14(d,J=6.4Hz,1H),1. 09-1.03(d,J=6.4Hz,1H),1.01-0.95(m,5H),0.92-0.80(m,3H),0.55-0.45(m,3H).

[0594] Biological Examples

[0595] Compound A168 from WO2021091982 (page 81 of Table 1) is cited as a reference compound of the present invention.

[0596]

[0597] Example 23

[0598] GSH reaction rate

[0599] Glutathione (GSH) is a tripeptide present in most tissues, especially in high concentrations in the liver, and plays a key role in protecting cells from oxidative damage and toxicity of exogenous electrophiles and maintaining redox balance. More specifically, glutathione conjugation aids in detoxification by binding electrophiles that might otherwise bind to proteins or nucleic acids, leading to cell damage and genetic mutations.

[0600] Many potentially toxic electrophilic xenobiotics and some endogenous compounds are detoxified by conversion to the corresponding glutathione S-conjugates, which deplete the intrinsic GSH and then attenuate the detoxification effect. Some drugs and halogenated workplace / environmental pollutants are bioactivated by this mechanism.

[0601] On the other hand, conjugation of glutathione to drug molecules in extrahepatic organs as well as in the liver usually results in poor PK properties of the molecule (especially high clearance) and increases the potential for off-target reactions (which may cause various toxicities). Therefore, strategies to minimize GSH metabolism are critical. 1 / 2 indicates that the GSH reaction rate is high. Therefore, T in the intrinsic GSH reaction assay 1 / 2 The assay is used to screen drug candidates.

[0602] The reference compounds and compounds of the present invention can potentially form conjugates with GSH via halogenation moiety substitution reactions or direct Michael addition reactions. Therefore, this test was performed to examine the GSH reactivity of the listed compounds.

[0603] To determine intrinsic GSH reactivity, 1 μM of the compound was incubated at 37°C in 100 mM potassium phosphate buffer (pH 7.4) in the presence and absence of 5 mM GSH for 0, 0.5, 1, 2, 4, and 6 h. At the end of the specified time points, the samples were quenched with acetonitrile containing 10 mM N-ethylmaleimide and an internal standard. The quenched samples were centrifuged and the supernatants were analyzed by LC-MS / MS for compound quantification. If the percent consumption after 6 hours of incubation was less than 20%, the compound was reported as stable; if the percent consumption was greater than 20%, the half-life value was reported.

[0604] Table 1. GSH reaction rates of examples and compounds of the present invention

[0605]

[0606] The above results clearly show that the reference compound (A168) forms a conjugate with GSH, which causes its depletion within 6 hours, while the compounds of the present invention maintain stability with less or no conjugation with GSH.

[0607] Example 24

[0608] Single-dose pharmacokinetic (SDPK) study in female BALB / c mice

[0609] The purpose of this study is to determine the pharmacokinetics of selected compounds after a single intravenous bolus or oral gavage administration in female BALB / c mice. In brief, two groups of female BALB / c mice (available from Zhejiang Vital River Laboratory Animal Technology Co., Ltd. or Shanghai Lingchang Biotechnology Co., Ltd) (N=3 / group) were treated with a single dose of compound at 3 mg / kg intravenous (IV) or 30 mg / kg oral (PO). Blood samples were collected at 5 minutes (only for IV), 15 minutes, 30 minutes, 1 hour, 2 hours, 4 hours, 7 hours and 24 hours after administration. Blood samples were placed on ice until centrifugation to obtain plasma samples. The concentration of the compound in the plasma sample was determined using the LC-MS / MS method. Pharmacokinetic parameters were calculated by non-compartmental analysis.

[0610] Table 2. SDPK results

[0611]

[0612] As can be seen from Table 2, Example 11 has good pharmacokinetic properties in the mouse model. In particular, Example 11 has a C max , 1.5 times the AUC 0-last and a much lower clearance rate, which makes Example 11 more suitable for use as an oral therapeutic active ingredient in the clinic to treat cancers with KRAS mutations.

[0613] Example 25

[0614] Human hepatocyte stability assay

[0615] The hepatocyte stability assay measures the rate of compound disappearance when incubated with cryopreserved human suspension hepatocytes. Positive controls, including midazolam, raloxifene, and dextromethorphan, were included in each experiment. 1 μM of the test compound was incubated with a suspension of human hepatocytes (1×10 6 The hepatocyte suspension was incubated in 5% CO 2The cultures were incubated at 37°C in an incubator with intermittent shaking at 900 rpm. The reaction was terminated by adding methanol containing an internal standard (2 μM tolbutamide) at 2, 10, 20, 40, 60, and 120 minutes after compound addition, and the consumption of the parent compound was monitored by LC-MS / MS analysis. For human data, CL_hep (mL / min / kg) > 16.24 was considered high clearance, and CL_hep (mL / min / kg) < 6.96 was considered low clearance. 16.24<CL_hep(mL / min / kg)> 6.96 is a medium clearance rate.

[0616] Table 3. Human hepatocyte stability of examples and compounds of the present invention

[0617]

[0618]

[0619] Achieving low clearance is beneficial for improving the in vivo performance of the compound, such as dose reduction, exposure enhancement and half-life extension. The above results clearly show that in the human hepatocyte stability assay, the reference compound (A168) shows a moderate clearance, while the compounds of the present invention maintain a low clearance.

[0620] Example 26

[0621] Cell viability assay

[0622] The purpose of this cellular assay is to determine the effect of the tested compounds on the proliferation of human cancer cell lines NCI-H358 (ATCC-CRL5807) cells, AGS (ATCC-CRL-1739) cells, SW620 (ATCC-CCL-227) over a 3-day treatment period by quantifying the amount of NADPH present at the endpoint using the Cell Counting Kit-8.

[0623] Cells were seeded in 96-well assay plates (Corning-3699) at 5,000 cells / well (NCI-H358), 2,000 cells / well (AGS), and 2,000 cells / well (SW620) and incubated overnight. On the day of the assay, diluted compounds were added at a final concentration of 0.5% DMSO. After 72 hours of incubation, one-tenth the volume of Cell Counting Kit 8 (Dnjindo-CK04) was added to each well. After 2 hours of incubation, the signal was read using EnVision (OD450 minus OD650). IC 50 Determined by fitting a 4-parameter sigmoidal concentration response model.

[0624] Table 4. Activity of Examples and Compounds of the Invention in KRAS Cell Viability Assay

[0625]

[0626]

[0627] Example 27

[0628] KRAS G12C-BRAF NanoBit Assay

[0629] To measure the ability of the tested compounds to disrupt the KRAS G12C-BRAF complex at the cellular level, we established the NanoBit Cell Assay in mammalian HEK293 (ATCC) cells.

[0630] HEK293 cells were grown and maintained using DMEM medium (Thermo Fisher Scientific) containing 10% fetal bovine serum and 1% penicillin / streptomycin. Both KRAS G12C and BRAF RBD were cloned into NanoBit vectors (BiBiT vector system, Promega) with SmBit-KRASG12C and BRAF RBD-LgBit orientations and co-transfected into HEK293 cells. Cells were then selected for 4 weeks with 100 μg / mL hygromycin B (10687010, Thermo Fisher) and blasticidin (5 μg / mL) to obtain a stable cell pool.

[0631] On the day of the assay, starting from a final concentration of 30 μM in DMSO, 75 nL of compound solution was present in a 384-well assay plate in 16 3-fold dilutions. Cells were then seeded at 10,000 cells / 25 μL / well in a 384-well plate. After a 3-hour incubation, a 6 μL volume of Live cell substrate (Promega) was added to each well. Luminescence was monitored at 20 minutes using the ultra384 model in Envision. Compounds that promote disruption of the KRAS G12C-BRAF RBD complex were identified as those that caused a decrease in luminescence compared to DMSO control wells.

[0632] Table 5. Activity of Examples and Compounds of the Invention in the KRAS G12C-BRAF NanoBit Assay

[0633] Examples <![CDATA[IC 50 (μM)]]> A168 0.205 Example 1 0.081 Example 2 0.018 Example 3 0.036 Example 4 0.219 Example 5 0.106 Example 6 0.029 Example 7 0.335 Example 8 0.054 Example 9 0.418 Example 10 0.12 Example 11 0.012 Example 12 0.067 Example 13 0.220 Example 14 0.036 Example 16 0.209 Example 18 0.240 Example 19 0.075 Example 20 0.084 Example 21 0.085 Example 22 0.036

[0634] Example 28

[0635] Interaction assay of KRAS-BRAF and CYPA (500 nM)

[0636] In this example, TR-FRET was also used to measure compound or compound-CYPA-dependent disruption of the KRAS G12C-BRAF complex. This protocol was also used to measure the disruption of KRAS G12D or KRAS G12V binding to BRAF by the compounds of the invention, respectively. In a 25 mM HEPES PH = 7.4 (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid, Thermo, 15630080), 0.002% Tween20, 0.1% BSA, 100 mM NaCl, 5 mM MgCl 2 , 10 μM GMPPNP (guanosine 5'-[β,γ-imino] triphosphate trisodium salt hydrate, Sigma, G0635), untagged CYPA, GMPPNP loaded with 6His-KRAS protein, and GST-BRAF RBD Mixed in the wells of a 384-well assay plate at final concentrations of 50 nM, 6.25 nM, and 1 nM, respectively. The compounds were present in the wells of the plate in a 16-point 3-fold dilution series starting from a final concentration of 10 μM and incubated for 3 hours. A mixture of MAb anti-6His-XL665 (Cisbio, 61HISXLB) and MAb anti-GST-TB cryptate (Cisbio, 61GSTTLB) was then added at final concentrations of 6.67 nM and 0.21 nM, respectively, and the plate was incubated for another 1.5 hours. The TR-FRET signal was read on a PHERstar FSX microplate reader (Ex 320 nm, Em 665 / 615 nm). Compounds that promote the destruction of the KRAS-BRAF complex were identified as compounds that cause a decrease in the TR-FRET ratio compared to the DMSO control wells.

[0637] Table 6. Activity of Examples and Compounds of the Invention in the KRAS-BRAF and CYPA (500 nM) Interaction Assay

[0638]

[0639]

[0640] Example 29

[0641] pERK inhibition assay

[0642] The assay is designed to measure the ability of the tested compounds to inhibit ERK phosphorylation, KRAS G12C in NCI-H358 cells, KRAS G12D in AGS cells, and KRAS G12V downstream signaling in SW620. NCI-H358 (ATCC-CRL5807) cells, AGS (ATCC-CRL-1739) cells, and SW620 (ATCC-CCL-227) cells were grown and maintained in RPMI-1640 medium (Thermo Fisher Scientific) containing 10% fetal bovine serum and 1% penicillin / streptomycin. The day before the addition of the compound, the cells were plated at a density of 30,000 cells / well, 20,000 cells / well, and 30,000 cells / well (for NCI-H358, AGS, and SW620) in a tissue culture treated 96-well plate (Corning-3699) and allowed to attach overnight. The diluted compounds were then added at a final concentration of 0.5% DMSO. After 4 hours of incubation, the culture medium was removed, 100 μL 4% formaldehyde was added, and the assay plate was incubated at room temperature for 20 minutes. The plate was then washed once with phosphate buffered saline (PBS) and permeabilized with 100 μL frozen methanol for 10 minutes. Nonspecific antibodies bound to the plate were blocked at room temperature for at least 1 hour using 50 μL 1X BSA blocking buffer (Thermo-37520, diluted 10 times with phosphate buffered saline (PBST)).

[0643] The amount of phosphor-ERK was determined using an antibody specific for the phosphorylated form of ERK. The primary antibody (pERK, CST-4370, Cell Signaling Technology) was diluted 1:300 in blocking buffer, with 50 μL aliquoted to each well and incubated overnight at 4 ° C. The cells were washed five times with PBST for 5 minutes. The secondary antibody (HRP-linked anti-rabbit IgG, CST-7074, Cell Signaling Technology) was diluted 1:1000 in blocking buffer, and 50 μL was added to each well and incubated at room temperature for 1-2 hours. The cells were washed 5 times with PBST for 5 minutes, 100 μL TMBELISA substrate (abcam-ab171523) was added, and gently shaken for 20 minutes. 50 μL stop solution (abcam-ab171529) was added, and the signal (OD450) was then read with EnVision.

[0644] IC 50 Determined by fitting a 4-parameter sigmoidal concentration response model.

[0645] Table 7. Activity of Examples and Compounds of the Invention in the KRAS pERK Inhibition Assay

[0646]

[0647]

[0648] Example 30

[0649] Stable KRAS mutant cell lines and cell viability assays.

[0650] The purpose of this study is to use (CTG) luminescent cell viability assay (Promega Corp., Madison, WI) was used to determine the potency and efficacy of the compounds on cell proliferation. We compared 14 KRAS G12C Variant sequences (V8A, V9Y, S17E, T58I, A59T, S65W, R68S, D69P, M72I, D92R, H95N, Y96D, Q99F, Q99W, Y96H and F156L) were cloned into Miapaca-2. A total of 14 stable Miapaca-2 mutant cell lines were established by lentiviral infection. For cell viability assays, compounds were administered to cells at a maximum dose of 10 μM using a 4-fold dilution series in a 9-point dose-response. KRAS mutant cells were maintained in DMEM+10% FBS+2.5% HI horse serum+1% PS+1 μg / mL puromycin and seeded into 96-well plates at 800-1,500 cells per well 24 hours before adding the compound, and then incubated with the compound for 3 days before determining viability (CellTiter-Glo, Promega). The assay was performed in biological replicates. Nonlinear regression curves were fitted using Xfit. IC50 (absolute IC50) is the dose at which the activity is estimated to be 50% relative to untreated wells. The inhibition rate of the compound was calculated as follows: % inhibition = 100-100 x (luminescence value-HPE) / (ZPE-HPE).

[0651] HPE: Luminescence value from wells containing culture medium only

[0652] ZPE: Luminescence value from wells containing DMSO

[0653] Table 8. Cell viability (IC50 (μM)) in mutant cells with KRAS G12C and other mutations

[0654]

Claims

1. A compound of formula (I), in R 1 for Where R 8 C 1-6 alkyl; R 9 C 3-7 Cycloalkyl, azetidinyl or phenyl, the C 3-7 Cycloalkyl, azetidinyl and phenyl are halogenated 3-6 Alkynyl, (halogenated C 3-6 Alkylpyrimidinyl)C 2-6 Alkynyl or pyrimidinyl C 2-6 Alkynyl substitution; R 2 C 1-6 alkyl; R 3 is H or halogen; R 4 is H or halogen; R 5 C 1-6 Alkyl or halogenated C 1-6 alkyl; R 6 C 1-6 Alkoxy C 1-6 alkyl; R 7 For morpholinyl, (halogenated C 1-6 alkyl) piperazinyl or C 1-6 Alkylpiperazinyl; A 1 is thiazolylidene; A 2 C 1-6 Alkylene; The condition is R 3 and R 4 Not at the same time H; or a pharmaceutically acceptable salt thereof.

2. A compound of formula (Ia), in R 1 for Where R 8 C 1-6 alkyl; R 9 C 3-7 Cycloalkyl, azetidinyl or phenyl, the C 3-7 Cycloalkyl, azetidinyl and phenyl are halogenated 3-6 Alkynyl, (halogenated C 3-6 Alkylpyrimidinyl)C 2-6 Alkynyl or pyrimidinyl C 2-6 Alkynyl substitution; R 2 C 1-6 alkyl; R 3 is H or halogen; R 4 is H or halogen; R 5 C 1-6 Alkyl or halogenated C 1-6 alkyl; R 6 C 1-6 Alkoxy C 1-6 alkyl; R 7 For morpholinyl, (halogenated C 1-6 alkyl) piperazinyl or C 1-6 Alkylpiperazinyl; A 1 is thiazolylidene; A 2 C 1-6 Alkylene; The condition is R 3 and R 4 Not at the same time H; or a pharmaceutically acceptable salt thereof.

3. The compound according to claim 1 or 2, wherein R 1 for Where R 8 C 1-6 Alkyl; R 9 For halogenated C 3-6 Alkynyl substituted C 3-7 Cycloalkyl.

4. The compound according to claim 1 or 2, wherein R 1 for Where R 8 is methyl; R 9 is cyclobutyl substituted by 3,3,3-trifluoroprop-1-ynyl.

5. A compound according to any one of claims 1 to 4, wherein R 9 It is 3-(3,3,3-trifluoroprop-1-ynyl)cyclobutyl.

6. A compound according to any one of claims 1 to 5, wherein R 2 It is isopropyl.

7. A compound according to any one of claims 1 to 6, wherein R 3 It is a halogen.

8. A compound according to any one of claims 1 to 7, wherein R 3 For fluorine.

9. A compound according to any one of claims 1 to 8, wherein R 4 is H or fluorine.

10. A compound according to any one of claims 1 to 9, wherein R 4 For H.

11. A compound according to any one of claims 1 to 10, wherein R 5 It is ethyl or 2,2,2-trifluoroethyl.

12. A compound according to any one of claims 1 to 11, wherein R 6 It is 1-methoxyethyl.

13. A compound according to any one of claims 1 to 12, wherein R 7 is morpholinyl, 4-(2,2,2-trifluoroethyl)piperazin-1-yl or 4-methylpiperazin-1-yl.

14. A compound according to any one of claims 1 to 13, wherein A 1 for Where bond "a" is attached to the indole ring.

15. A compound according to any one of claims 1 to 14, wherein A 2 It is 2,2-propylene.

16. The compound according to claim 1 or 2, wherein R 1 for Where R 8 C 1-6 Alkyl; R 9 For halogenated C 3-6 Alkynyl substituted C 3-7 Cycloalkyl; R 2 C 1-6 alkyl; R 3 is a halogen; R 4 is H; R 5 C 1-6 Alkyl or halogenated C 1-6 alkyl; R 6 C 1-6 Alkoxy C 1-6 alkyl; R 7 For morpholinyl, (halogenated C 1-6 alkyl) piperazinyl or C 1-6 Alkylpiperazinyl; A 1 for wherein bond "a" is attached to the indole ring; A 2 C 1-6 Alkylene; or a pharmaceutically acceptable salt thereof.

17. The compound according to claim 16, wherein R 1 for Where R 8 is methyl; R 9 is 3-(3,3,3-trifluoroprop-1-ynyl)cyclobutyl; R 2 is isopropyl; R 3 For fluorine; R 4 is H; R 5 is ethyl or 2,2,2-trifluoroethyl; R 6 is (1S)-1-methoxyethyl; R 7 is morpholinyl, 4-(2,2,2-trifluoroethyl)piperazin-1-yl or 4-methylpiperazin-1-yl; A 1 for wherein bond "a" is attached to the indole ring; A 2 is 2,2-propylene; or a pharmaceutically acceptable salt thereof.

18. A compound selected from: trans-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide; N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide; cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide; N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)benzamide; cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(2-pyrimidin-2-ylethynyl)cyclobutanecarboxamide; cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide; cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide; cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide; trans-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxamide; cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-4-(3,3,3-trifluoroprop-1-ynyl)cyclohexanecarboxamide; cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide; cis-N-[(1S)-1-[[(7S,13S)-25-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide; cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-[2-[5-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxamide; N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]- N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide; N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide; cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-[2-[4-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxamide; N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-[4-(2,2,2-trifluoroethyl)piperazin-1-yl]-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide; (2S)-N-[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-(4-methylpiperazin-1-yl)-3-pyridinyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]-2-isopropyl-4-oxo-4-[3-(3,3,3-trifluoroprop-1-ynyl)azetidin-1-yl]butanamide; N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]- N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)azetidine-1-carboxamide; cis-N-[(1S)-1-[[(7S,13S)-21-ethyl-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-[2-[4-(trifluoromethyl)pyrimidin-2-yl]ethynyl]cyclobutanecarboxamide; and cis-N-[(1S)-1-[[(7S,13S)-24-fluoro-(20M)-20-[2-[(1S)-1-methoxyethyl]-5-morpholino-3-pyridyl]-17,17-dimethyl-8,14-dioxo-21-(2,2,2-trifluoroethyl)-15-oxa-4-thia-9,21,27,28-tetraazapentacyclo[17.5.2.1 2,5 .1 9,13 .0 22,26 ]octacosyl-1(25),2,5(28),19,22(26),23-hexaen-7-yl]carbamoyl]-2-methyl-propyl]-N-methyl-3-(3,3,3-trifluoroprop-1-ynyl)cyclobutanecarboxamide; or a pharmaceutically acceptable salt thereof.

19. A method for preparing a compound according to any one of claims 1 to 18, the method comprising any one of the following steps: a) reacting a compound of formula (II) in the presence of a coupling agent and a base, With acid (III), A coupling reaction is carried out between them to form a compound of formula (I); Where R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , A 1 and A 2 As defined in any one of claims 1 to 17; the coupling reagent is T3P, HATU, PyBOP or EDCI / HOBt; the base is TEA, DIEPA or DMAP.

20. A compound or a pharmaceutically acceptable salt according to any one of claims 1 to 18 for use as a therapeutically active substance.

21. A pharmaceutical composition comprising a compound according to any one of claims 1 to 18; and a pharmaceutically acceptable excipient.

22. Use of a compound according to any one of claims 1 to 18 for treating a KRAS G12C protein-related disease.

23. Use of a compound according to any one of claims 1 to 18 for treating KRASG12C, G12D and G12V protein related diseases.

24. Use of a compound according to any one of claims 1 to 18 for inhibiting the interaction of RAS with downstream effectors, wherein the downstream effectors are RAF and PI3K.

25. Use of a compound according to any one of claims 1 to 18 for inhibiting propagated oncogenic MAPK and PI3K signalling.

26. Use of a compound according to any one of claims 1 to 18 for treating or preventing a KRAS mutation driven cancer, wherein the cancer is selected from pancreatic cancer, colorectal cancer, lung cancer, esophageal cancer, gallbladder cancer, melanoma ovarian cancer and endometrial cancer.

27. Use of a compound according to any one of claims 1 to 18 for treating or preventing a KRAS mutation driven cancer, wherein the cancer is selected from pancreatic adenocarcinoma, colorectal cancer and non-small cell lung cancer.

28. A compound or pharmaceutically acceptable salt according to any one of claims 1 to 18 for use in treating or preventing a KRAS mutation driven cancer, wherein the cancer is selected from pancreatic adenocarcinoma, colorectal cancer and non-small cell lung cancer.

29. Use of a compound according to any one of claims 1 to 18 for treating or preventing a cancer driven by a KRAS mutation, wherein the cancer comprises: is a first mutation of G12C; and a second mutation at a position selected from the group consisting of V8A, V9Y, S17E, T58I, A59T, S65W, R68S, D69P, M72I, D92R, H95N, Y96D, Q99F, Q99W, Y96H, and F156L.

30. Use of a compound according to any one of claims 1 to 18 for the preparation of a medicament for treating or preventing a KRAS mutation driven cancer, wherein the cancer is selected from pancreatic adenocarcinoma, colorectal cancer and non-small cell lung cancer.

31. Use of a compound according to any one of claims 1 to 18 for the preparation of a medicament for treating or preventing a cancer driven by a KRAS mutation, wherein the cancer comprises: It is the first mutation of G12C; and a second mutation at a position selected from the group consisting of V8A, V9Y, S17E, T58I, A59T, S65W, R68S, D69P, M72I, D92R, H95N, Y96D, Q99F, Q99W, Y96H, and F156L.

32. A method of treating or preventing a KRAS mutation driven cancer, wherein the cancer is selected from pancreatic adenocarcinoma, colorectal cancer and non-small cell lung cancer, the method comprising administering a therapeutically effective amount of a compound as defined in any one of claims 1 to 18.

33. A method of treating or preventing a KRAS mutation-driven cancer, wherein the cancer comprises: is a first mutation of G12C; and a second mutation at a position selected from the group consisting of V8A, V9Y, S17E, T58I, A59T, S65W, R68S, D69P, M72I, D92R, H95N, Y96D, Q99F, Q99W, Y96H, and F156L.

34. A compound or pharmaceutically acceptable salt according to any one of claims 1 to 18, prepared according to the process of claim 19.

35. The invention as herein described.

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