KRas Inhibitor for Cancer Treatment

Abstract

The present invention provides compounds having the structure of formula (I) that can be used as KRas inhibitors, pharmaceutical compositions containing such compounds, methods for preparing such compounds, and the use of these compounds in the treatment of cancer.

Description

Technical Field

[0001] The present invention relates to the field of medicinal chemistry. More specifically, the present invention relates to a class of compounds with novel structures that can be used as KRas inhibitors, pharmaceutical compositions containing such compounds, methods for preparing such compounds, and uses of these compounds in treating cancer. Background Art

[0002] Ras, or rat sarcoma oncogene homolog, represents a group of closely related monomeric globular proteins that belong to the GTPase family. Specifically, under normal physiological conditions, Ras is activated by growth factors and various other extracellular signals, regulating functions such as cell growth, survival, migration, and differentiation. These regulatory functions of Ras are achieved through a "molecular switch" that switches between GDP-bound and GTP-bound states (Alamgeer et al., Current Opin Pharmacol. 2013, 13:394-401). Ras bound to GDP is in an inactive form and is in a dormant or closed state. At this time, the signaling system is shut down. When it is exposed to some pro-growth stimuli, it will be activated. For example, it can be induced by guanine nucleotide exchange factors (GEFs) to release GDP and bind to GTP. As a result, Ras is "turned on" and converted into the active form of Ras, which recruits and activates various downstream effectors to carry out signal transduction. It can transmit signals on the cell surface to the cytoplasm, thereby controlling many key cellular processes such as differentiation, survival and proliferation (Zhi Tan et al., Mini-Reviews in Medicinal Chemistry, 2016, 16, 345-357).

[0003] Ras has GTPase activity, which cleaves the terminal phosphate of GTP and converts it to GDP, essentially converting it to an inactive state. However, Ras' endogenous GTPase activity is very low, and the conversion of GTP-Ras into GDP-Ras requires the exogenous protein GAP (GTPase-activating protein). GAP interacts with Ras and promotes the conversion of GTP to GDP. Therefore, any Ras gene mutation that affects the interaction between Ras and GAP or the conversion of GTP to GDP will cause Ras to remain in an activated state for a long time, thereby continuously transmitting growth and division signals to cells, stimulating continuous cell proliferation, and ultimately leading to tumor formation and progression.

[0004] Among human tumor-associated genes, there are three ubiquitously expressed Ras genes: H-RAS, K-RAS, and N-RAS, which encode highly homologous, approximately 21KDa HRas, NRas, and KRas proteins, respectively. In 1982, researchers first discovered that Ras was activated by mutation in cancer cell lines (Chang, EH et al., Proceedings of the National Academy of Sciences of the United States of America, 1982, 79(16), 4848-4852). Subsequent large-scale genome sequencing studies in different cancer types revealed that Ras proteins were mutated in more than 30% of cancer types, with the highest mutation rates in pancreatic cancer (>90%), colon cancer (45%), and lung cancer (35%). Transgenic and genetically engineered mouse models have also revealed that mutated Ras proteins are sufficient to drive and trigger various types of cancer, and that Ras oncogenes are crucial for the maintenance and progression of tumors in various cancer types. For example, in Ras mutant cancer cell lines and animal models, RNAi has been shown to slow tumor growth. These studies have made Ras oncoproteins widely accepted and attractive anti-cancer drug targets in the pharmaceutical field.

[0005] Studies have shown that Ras mutations are most common in KRas, and KRas mutations can be observed in about 85% of Ras mutation-driven cancers; the vast majority of Ras mutations occur at codons G12, G13, and Q61, of which about 80% of KRas mutations occur at glycine at codon 12, such as G12C mutation, G12D mutation, G13D mutation, etc. KRas mutations are common in pancreatic cancer, lung adenocarcinoma, colorectal cancer, gallbladder cancer, thyroid cancer, and bile duct cancer, and can also be seen in 25% of non-small cell lung cancer patients (McCormick, F. et al., Clinical Cancer Research 21(8), 1797-1801, 2015). Therefore, KRas mutant proteins have become the most important branch of Ras drug target research, and the development of its inhibitors is also considered a very promising research and development direction in the development of anti-cancer / tumor drugs.

[0006] However, over the past three decades, drug development for Ras has shown that due to the smooth surface of the Ras protein, the lack of obvious grooves or pockets for binding small molecule inhibitors, and its very high affinity for guanine substrates (picomolar), the development of its small molecule inhibitors has been in an intractable dilemma. As a result, Ras has long been considered an "undruggable" target in the industry. Despite this, continued efforts to target Ras mutant proteins have achieved some encouraging results. A series of Ras inhibitors have been developed that inhibit Ras, especially KRas mutations, through various pathways, including direct targeting of Ras, suppressing Ras expression levels, disrupting the localization of Ras proteins, targeting synthetic lethal components, targeting Ras-GEF interactions, targeting Ras and effector interactions, and targeting Ras dimerization (Zhi Tan et al., Mini-Reviews in Medicinal Chemistry, 2016, 16, 345-357).

[0007] Ras inhibitors that have been developed, such as inhibitors targeting the most common mutant protein KRas-G12C, include allosteric covalent inhibitors such as the 6H05 series, quinazoline series, ARS series, and tetrahydropyridopyrimidine series, as well as positive binding covalent inhibitors, which are reviewed in the literature (Duan Ni et al., Pharmacology & Therapeutics, https: / / doi.org / 10.1016 / j.pharmthera.2019.06.007). Various structural types of KRas inhibitors are also described in several patent documents, such as CN10256421, US2019 / 0144444A1, and WO2019 / 110751A1.

[0008] However, the above-mentioned KRas inhibitors still have problems that need to be solved, and their "drugability" is still unsatisfactory. For example, many KRas-dependent cancers are prone to drug resistance to such targeted therapeutics, have side effects such as off-target effects, produce chemically active metabolites, have poor metabolic stability, or produce immunogenic covalent adducts (John P. O'Bryan et al., Pharmacological Research 139 (2019) 503-511; Duan Ni et al., Pharmacology & Therapeutics, https: / / doi.org / 10.1016 / j.pharmthera.2019.06.007). Therefore, there is still a need for more alternative KRas inhibitors in the clinic, which are expected to have KRas inhibitory activity comparable to or improved than existing inhibitors, improved "drugability", better safety such as fewer drug interactions or metabolic properties, improved pharmacokinetic properties, and / or higher selectivity for different patient groups or specific tumor types.

[0009] The present invention provides novel structural inhibitor compounds with KRas mutant protein inhibitory activity. These compounds, particularly preferred compounds of the present invention, have improved structural patterns and, compared to existing KRas mutant protein inhibitors, exhibit the following technical benefits:

[0010] (1) Retaining comparable, enhanced, or even significantly enhanced KRas mutant protein and related cancer cell proliferation inhibitory activity;

[0011] (2) Different bioactivity profiles for different disease types or patient groups;

[0012] (3) Improved metabolic stability, leading to better pharmacokinetic properties;

[0013] (4) They have improved physicochemical properties, thus having good drugability and safety, such as being more easily absorbed in the body. Summary of the Invention

[0014] Through research, the present inventors have discovered that the compounds of formula (I) defined herein, their isomers, or pharmaceutically acceptable salts or solvates thereof are effective inhibitors of Ras mutations, especially KRas mutant proteins, and are capable of inhibiting Ras mutations, especially KRas activity, in cells. They can be used to treat or prevent diseases or conditions mediated by or benefiting from the inhibition of Ras mutations, especially KRas mutant proteins. In particular, they can inhibit abnormal cell proliferation by inhibiting Ras mutations, especially KRas mutant proteins, thereby treating or preventing tumors or cancer.

[0015] The first aspect of the present invention provides a compound of formula (I), an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof,

[0016]

[0017] in,

[0018] A is selected from CR a or N, where R a Selected from halogen, CN, nitro, C 3-6 Cycloalkyl or C optionally substituted by halogen 1-6 alkyl;

[0019] R 1 、R 2 and R 3 are each independently selected from H, halogen or C 1-6 Alkyl, wherein the alkyl group is optionally independently selected from halogen, -N(R c )2、-OR c or substituted with a 3-8 membered heterocycloalkyl group;

[0020] Rb is independently selected at each occurrence from H, halogen, CN or C optionally substituted with halogen or CN. 1-6 alkyl;

[0021] X is selected from a bond, -O- or -NH-;

[0022] R 4 Selected from H, halogen, C 1-6 Alkyl, -C 0-6 Alkyl-C 6-10 Aryl, -C 0-6 Alkyl-C 3-8 Cycloalkyl, -C 0-6 Alkyl-C 3-8 Cycloalkenyl, -C 0-6 Alkyl-3-8 membered heterocycloalkyl, -C 0-6 Alkyl-3-8 membered heterocycloalkenyl, -C 0-6 Alkyl-5-10 membered heteroaryl, wherein said alkyl, aryl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl and heteroaryl are optionally substituted by one or more groups independently selected from the group consisting of halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -SR c 、-(CR c R c ) 0-6 -(CO)0-1-OR c 、-(CRc R c ) 0-6 -(CO) 0-1 -N(R c )2, wherein -(CR c R c ) 0-6 -(CO) 0-1 -N(R c ) 2 optionally through the group connected to N, together with the atoms on the cyclic group to which it is connected and the adjacent atoms to form a 4-7 membered nitrogen-containing cyclic group;

[0023] R c is independently selected at each occurrence from H or C optionally substituted by halogen 1-6 Alkyl, or two R c Each independently forms a 3-6 membered cyclic group together with the carbon atom or nitrogen atom to which they are attached;

[0024] E is selected from halogen, -OR d or -N(R d )2-, where R d are each independently H or C optionally substituted by halogen 1-6 alkyl;

[0025] R 5 for

[0026] m is 0 or 1;

[0027] represents an aromatic ring;

[0028] B and D are each independently selected from N or C;

[0029] Z, G and Y are each independently selected from C, N, O or S;

[0030] R 6 、R 7 and R 8 are each independently selected from H, halogen and C optionally substituted by halogen 1-6 alkyl;

[0031] The condition is that B and D are not N at the same time; and at most three of Z, G, Y, D, and B are not C.

[0032] The present invention further provides a compound of formula (II), an isomer, a pharmaceutically acceptable salt or a solvate thereof.

[0033]

[0034] where R 1 、R 2 、R 3 、R 4 、R 6 、R 7 、R 8 ,A,R b each has the meaning given above for the compounds of formula (I).

[0035] The second aspect of the present invention provides a pharmaceutical composition comprising a compound of formula (I) or formula (II) of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof.

[0036] In a third aspect, the present invention provides a compound of formula (I) or formula (II), an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof for use as a medicament.

[0037] The fourth aspect of the present invention provides a compound of formula (I) or formula (II), an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof for use in treating and / or preventing diseases mediated by Ras mutations, especially KRas mutations, preferably KRasG12C, KRasG12D or KRasG13D mutations, most preferably KRasG12C mutations.

[0038] In a fifth aspect, the present invention provides the use of a compound of formula (I) or formula (II), an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition comprising the same, in the preparation of a medicament for treating and / or preventing diseases mediated by Ras mutations, particularly KRas mutations, preferably KRas G12C, KRas G12D or KRas G13D mutations, most preferably KRas G12C mutations.

[0039] In a sixth aspect, the present invention provides a method for treating and / or preventing diseases mediated by Ras mutations, particularly KRas mutations, preferably KRasG12C, KRasG12D or KRasG13D mutations, and most preferably KRasG12C mutations, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of formula (I) or formula (II) of the present invention, its isomers or pharmaceutically acceptable salts or solvates thereof, or a pharmaceutical composition comprising the same.

[0040] In a seventh aspect, the present invention provides a method for preparing the compound of formula (I) or formula (II), its isomers, or pharmaceutically acceptable salts or solvates thereof.

[0041] In an eighth aspect, the present invention provides a pharmaceutical combination comprising a compound of formula (I) or (II) of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof, and one or more other pharmaceutically active agents. Detailed Description of the Invention

[0042] definition

[0043] Unless otherwise indicated, the various terms used in the specification and claims have the meanings shown below. In the absence of a particular definition of a particular term or phrase, it should not be construed as undefined or unclear, but rather should be understood according to its ordinary meaning in the art. Many of the groups defined herein are optionally substituted, and the list of substituents provided in this definition section is merely illustrative and is not intended to limit the substituents defined elsewhere in the specification and claims.

[0044] As used herein, the term "Ras mutant" or "Ras mutant protein" refers to a protein encoded and expressed by a Ras gene in which one or more codons are mutated, typically including but not limited to a Ras protein in which a glycine residue at codon 12, a glycine residue at codon 13, or a glutamine residue at codon 61 of Ras is mutated, such as a mutant HRas, NRas, or KRas. These residues are located in the active site of Ras, and their mutation can impair the intrinsic or GAP-catalyzed GTPase activity of Ras, resulting in the persistence of GTP-bound Ras.

[0045] For the purposes of the present invention, "Ras mutation" or "Ras mutant protein" are used interchangeably and generally refer to mutant HRas, NRas or KRas, such as but not limited to HRas-G12C (glycine to cysteine mutation at codon G12), NRas-G12C, KRas-G12C, KRas-G12D (glycine to aspartic acid mutation at codon G12), KRas-G13D (glycine to aspartic acid mutation at codon G13); particularly to KRas mutant proteins, more particularly to KRas-G12C mutant protein, KRas-G12D mutant protein, KRas-G13D mutant protein, and most particularly to KRas-G12C mutant protein.

[0046] The term "treatment" as used herein refers to administering one or more compounds of formula (I), isomers thereof, or pharmaceutically acceptable salts or solvates thereof to a subject, such as a mammal, such as a human, suffering from the disease or having symptoms of the disease, to cure, alleviate, mitigate or affect the disease or the symptoms of the disease. In a specific embodiment of the present invention, the disease is a Ras mutation-mediated disease as defined below, in particular a tumor or cancer.

[0047] The term "prevention" as used herein is well known in the art and refers to administering one or more compounds of formula (I), isomers thereof, or pharmaceutically acceptable salts or solvates thereof, as described herein to a subject, such as a mammal, such as a human, suspected of suffering from or susceptible to a Ras mutation-mediated disease as defined herein, especially cancer or tumor, so that the risk of suffering from the defined disease is reduced. The term "prevention" includes the use of the compounds of the present invention before the diagnosis or determination of any clinical and / or pathological symptoms.

[0048] As used herein, the terms "inhibit" and "reduce" or any variations of these terms refer to the ability of a bioactive agent to reduce the signaling activity of a target of interest by interacting directly or indirectly with the target, and refers to any measurable reduction or complete inhibition of the activity of the target of interest. For example, compared to normal, the activity (e.g., KRas activity) can be reduced by about, up to about, or at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99% or more, or any range derivable therein.

[0049] As used herein, the term "selective inhibition" refers to the ability of a bioactive agent to preferentially reduce the signaling activity of a target over off-target signaling activity by directly or indirectly interacting with the target. With respect to compounds of Formula (I) of the present invention, the compounds have the ability to selectively inhibit G12 or G13 mutations in KRas, HRas, or NRas proteins, such as G12C mutations, G12D mutations, and G13D mutations, relative to various mutations occurring in one or more codons of the Ras protein. The ability to selectively inhibit the G12C mutation in the KRas protein is preferred. For example, the present invention has at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99% or more, or any range derivable therein, inhibition of better activity against a specific Ras mutation as compared to another specific Ras mutation, or at least 1-, 2-, 3-, 4-, 5-, 10-, 25-, 50-, 100-, 250-, or 500-fold better activity against a specific Ras mutation (e.g., KRas-G12C) as compared to its activity against another specific Ras mutation.

[0050] As used herein, the term "Ras mutation-mediated disease" refers to a disease in which Ras mutations contribute to the onset and progression of the disease, or in which inhibition of Ras mutations reduces the incidence, ameliorate, or eliminates disease symptoms. For purposes of the present invention, "Ras mutation-mediated disease" preferably refers to a disease mediated by KRas mutations, most preferably a disease mediated by KRas-G12C, and even more preferably a cancer or tumor.

[0051] As used herein, the term "cancer" or "tumor" refers to abnormal cell growth and proliferation, whether malignant or benign, and all precancerous and cancerous cells and tissues. For purposes of the various aspects of the present invention, the cancer or tumor includes, but is not limited to, lung adenocarcinoma, lung cancer, bone cancer, pancreatic cancer, skin cancer, head and neck cancer, cutaneous or intraocular melanoma, uterine cancer, ovarian cancer, rectal cancer, cancer of the anal region, stomach cancer, colon cancer, breast cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, esophageal cancer, small intestine cancer, cancer of the endocrine system, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal cell carcinoma, renal pelvis cancer, central nervous system tumor (CNS), primary CNS lymphoma, spinal tumor, brain stem glioma, or pituitary adenoma.

[0052] For various aspects of the present invention, preferably, the cancer or tumor is associated with a Ras mutation, particularly a KRas mutation, preferably a KRas G12C, KRas G12D or KRas G13D mutation, most preferably a KRas G12C mutation, including but not limited to the above-mentioned tumor types and their preferred ranges. Particularly preferred tumors of the present invention include lung cancer, lung adenocarcinoma, colon cancer, rectal cancer, pancreatic cancer, endometrial cancer, bile duct cancer, leukemia and ovarian cancer.

[0053] As used herein, the terms "subject," "individual," or "patient" refer to a vertebrate. In certain embodiments, the vertebrate is a mammal. Mammals include, but are not limited to, farm animals (e.g., cattle), sports animals, pets (e.g., guinea pigs, cats, dogs, rabbits, and horses), primates, mice, and rats. In certain embodiments, the mammal is a human.

[0054] The term "therapeutically effective amount" as used herein refers to an amount or dosage that is generally sufficient to produce a beneficial therapeutic effect on a cancer or tumor patient in need of treatment. Those skilled in the art can determine the effective amount or dosage of the active ingredient of the present invention by conventional methods and in combination with conventional influencing factors.

[0055] The term "drug combination" as used herein means that the compounds of the present invention can be combined with other active agents to achieve the purpose of the present invention. The other active agents may be one or more additional compounds of the present invention, or may be a second or additional (e.g., a third) compound that is compatible with the compounds of the present invention, i.e., does not adversely affect each other, or has complementary activity, such as these active agents are known to regulate other biologically active pathways, or regulate different components in the biologically active pathways involved in the compounds of the present invention, or even overlap with the biological targets of the compounds of the present invention. Such active agents are suitably combined in an effective amount to achieve the intended purpose. The other active agents may be co-administered with the compounds of the present invention in a single pharmaceutical composition, or administered separately from the compounds of the present invention in different discrete units, and when administered separately, may be performed simultaneously or sequentially. The sequential administration may be close or distant in time.

[0056] In one aspect, other active agents that can be used in combination with the compounds of the invention include, but are not limited to, chemotherapeutic agents, therapeutic antibodies, and radiation therapy, such as alkylating agents, antimetabolites, cell cycle inhibitors, mitotic inhibitors, topoisomerase inhibitors, antihormonal drugs, angiogenesis inhibitors, or cytotoxic agents.

[0057] As used herein, the term "pharmaceutically acceptable" refers to molecular entities and compositions that do not produce adverse, allergic or other untoward reactions when administered in appropriate amounts to animals, such as humans.

[0058] As used herein, the term "pharmaceutically acceptable salt" refers to salts that retain the biological effectiveness and properties of the parent compound and are not biologically or otherwise undesirable, including acid addition salts and base addition salts. "Pharmaceutically acceptable acid addition salts" can be formed by compounds having free bases with inorganic acids or organic acids, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, carbonic acid, phosphoric acid, etc., and organic acids can be selected from aliphatic, alicyclic, aromatic, araliphatic, heterocyclic, carboxylic and sulfonic 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, benzenesulfonic acid, p-toluenesulfonic acid, salicylic acid, etc. "Pharmaceutically acceptable base addition salts" include those derived from inorganic bases such as sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum salts and the like, as well as salts derived from pharmaceutically acceptable organic non-toxic bases including, but not limited to, primary, secondary, and tertiary amines, substituted ammoniums including naturally occurring substituted amines, cyclic amines, and basic ion exchange resins, such as ammonia, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, diethanolamine, 2-dimethylaminoethanol, 2-diethylaminoethanol, tromethamine, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, hydrazine, choline, betaine, ethylenediamine, glucosamine, methylglucamine, triethanolamine, theobromine, purines, piperazine, piperidine, N-ethylpiperidine, polyamine resins, and the like.

[0059] The term "isomer" as used herein refers to any stereoisomer, enantiomeric mixture, including racemates, diastereomeric mixtures, geometric isomers, atropisomers and / or tautomers that may exist in the structure of a compound. The determination and separation methods of the stereochemistry of the isomers are well known to those skilled in the art (S. P. Parker, Ed., McGraw-Hill Dictionary of Chemical Terms (1984) McGraw-Hill Book Company, New York; and Eliel, E. and Wilen, S., "Stereochemistry of Organic Compounds", John Wiley & Sons, Inc., New York, 1994). Therefore, the present invention covers all possible isomeric forms of the compounds of formula (I) as defined above, and pharmaceutically acceptable salts or solvates thereof.

[0060] The compound structural formula or structural fragments used herein or Indicates the absolute configuration of a stereocenter, i.e., a chiral center. Accordingly, R and S are used to indicate the absolute configuration of the chiral center in the names of the compounds or intermediates provided by the present invention. The determination of the absolute configuration is well known to those skilled in the art.

[0061] The structural fragments used in this article The bonds indicated to be cross-linked are the bonds connecting the structural fragment to the rest of the molecule.

[0062] As used herein, the term "solvate" refers to a solvent addition form containing a stoichiometric or non-stoichiometric amount of a solvent, including any solvated form of a compound of the invention, including, for example, solvates with water, such as a hydrate, or with an organic solvent, such as methanol, ethanol, or acetonitrile, i.e., as a methanolate, ethanolate, or acetonitrile, respectively; or in the form of any polymorph. It should be understood that such solvates of the compounds of the invention also include solvates of the pharmaceutically acceptable salts of the compounds of the invention.

[0063] As used herein, the term "isotopic variant" refers to a compound that contains unnatural ratios of isotopes on one or more atoms constituting the compound. The compounds of the present invention may contain unnatural ratios of atomic isotopes on one or more atoms constituting the compound, thereby forming isotopic variations of the compounds of the present invention or their pharmaceutically acceptable salts, whether or not radioactive, are intended to be encompassed within the scope of the present invention. Examples of isotopes that can be incorporated into the compounds of the present invention and their pharmaceutically acceptable salts include, for example 2 H. 3 H. 13 C. 14 C. 15 N. 17 O. 18 O. 31 p、 32 p、 35 S. 18 F and 36 It will be appreciated that isotopic variations of the compounds of the invention and their pharmaceutically acceptable salts can generally be prepared by conventional methods using appropriate isotopic variations of suitable reagents. For example, radioactive isotopes (e.g. 3 H or 14 C) Certain isotopic variations of the compounds of the present invention and their pharmaceutically acceptable salts are useful in drug and / or substrate tissue distribution studies. 3 H and carbon-14 14 C isotopes are particularly preferred due to their ease of preparation and detectability. 2H substitution may offer certain therapeutic advantages resulting from greater metabolic stability, such as increased in vivo half-life or reduced dosage requirements, and thus may be preferred in some circumstances. In addition, it is possible to prepare 11 C. 18 F. 15 O and 13 The compounds of the present invention are substituted with N) and can be used in positron emission tomography (PET) studies for substrate receptor occupancy detection.

[0064] As used herein, the term "metabolite" refers to a product produced by the metabolism of a particular compound in vivo. Such products may, for example, result from oxidation, reduction, hydrolysis, amidation, deamidation, esterification, deesterification, enzymatic cleavage, etc. of the administered compound. Identification and analysis of metabolite products are performed in a manner well known to those skilled in the art.

[0065] The term "prodrug" as used herein refers to a compound that can be converted into a bioactive compound as described herein, such as a compound of formula (I) under physiological conditions or by solvolysis. Therefore, the term "prodrug" refers to a precursor of a pharmaceutically acceptable bioactive compound. In some aspects, a prodrug is inactive when administered to a subject, but is converted into an active compound in vivo, for example, by hydrolysis. Prodrug compounds generally provide advantages of solubility, tissue compatibility, or delayed release in mammalian organisms (see, for example, Bundgard, H., Design of Prodrugs (1985), pp. 7-9, pp. 21-24 (Elsevier, Amsterdam)). Discussions of prodrugs can be found in Higuchi, T. et al., ACS Symposium Series, Vol. 14, and "Bioreversible Carriers in Drug Design," Edward B. Roche, U.S. Pharmaceutical Association & Pergamon Press, 1987, all of which are incorporated herein by reference. The term "prodrug" is also intended to include any covalently bonded carriers that release the active compound in vivo when such prodrugs are administered to a mammalian subject. Prodrugs of active compounds as described herein are typically prepared by modifying functional groups present in the active compound such that the modifications can be cleaved into the parent active compound during routine manipulation or in vivo. Prodrugs include compounds wherein a hydroxyl, amino, or sulfhydryl group is bonded to any group that cleaves to form a free hydroxyl, free amino, or free sulfhydryl group when the prodrug is administered to a mammal. Examples of prodrugs include, but are not limited to, acetate, formate, and benzoate derivatives of hydroxyl functional groups, or acetamide, formamide, and benzamide derivatives of amine functional groups in the active compound. In some embodiments, prodrugs include phosphate-containing prodrugs, boronate-containing prodrugs, thiophosphate-containing prodrugs, sulfate-containing prodrugs, peptide-containing prodrugs, D-amino acid-modified prodrugs, glycosylated prodrugs, β-lactam-containing prodrugs, optionally substituted phenoxyacetamide-containing prodrugs, or optionally substituted phenylacetamide-containing prodrugs, as well as 5-fluorocytosine and 5-fluorouridine prodrugs.

[0066] The term "pharmaceutically acceptable excipient or carrier" as used herein refers to one or more compatible solid or liquid fillers or gel substances that are suitable for human use and have sufficient purity and sufficiently low toxicity. Examples include, but are not limited to, cellulose and its derivatives (such as sodium carboxymethyl cellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as magnesium stearate), calcium sulfate, vegetable oils, polyols (such as propylene glycol, glycerol, mannitol, sorbitol, etc.), emulsifiers (such as Tweens), wetting agents (such as sodium lauryl sulfate), colorants, flavorings, stabilizers, antioxidants, preservatives, etc.

[0067] As used herein, the term "halogen" or "halo" refers to F, Cl, Br or I. Furthermore, the term "halogen-substituted" group is intended to include mono- or poly-halogenated groups in which one or more same or different halogens replace one or more hydrogens in the group.

[0068] As used herein, term " alkyl " refers to the straight or branched saturated hydrocarbon group consisting of carbon atoms and hydrogen atoms. Specifically, alkyl has 1-10, such as 1 to 6, 1 to 5, 1 to 4, 1 to 3 or 1 to 2 carbon atoms. For example, as used herein, term " C1-C6 alkyl " refers to the straight or branched saturated hydrocarbon group with 1 to 6 carbon atoms, examples of which are methyl, ethyl, propyl (including n-propyl and isopropyl), butyl (including n-butyl, isobutyl, sec-butyl or tert-butyl), pentyl (including n-pentyl, isopentyl, neopentyl), n-hexyl, 2-methylpentyl etc. Specific alkyl has 1 to 3 carbon atoms.

[0069] The term "alkoxy" as used herein refers to the group -O-alkyl, wherein alkyl has the meaning given herein. Specifically, the term includes the group -OC 1-6 Alkyl, more specifically -OC 1-3 Alkyl. Representative examples of alkoxy include, but are not limited to, methoxy, ethoxy, propoxy (including n-propoxy, isopropoxy), butoxy (including n-butoxy, isobutoxy, tert-butoxy), pentoxy (including n-pentoxy, isopentoxy, neopentoxy), hexoxy (including n-hexoxy, isohexoxy), and the like. Specific alkoxy groups have 1 to 3 carbon atoms.

[0070] As used herein, the term "alkylthio" refers to an -S-alkyl group, wherein the alkyl group is as defined above for "alkyl". Specifically, the term includes the group -SC 1-6 Alkyl, more specifically -SC 1-3Representative examples of alkylthio groups include, but are not limited to, methylthio, ethylthio, propylthio (including n-propylthio and isopropylthio), butylthio (including n-butylthio, isobutylthio and tert-butylthio), pentylthio (including n-pentylthio, isopentylthio and neopentylthio), hexylthio (including n-hexylthio and isohexylthio), and the like. Specific alkylthio groups have 1 to 3 carbon atoms.

[0071] As used herein, the term "halogen-substituted C1-C6 alkyl" refers to the C1-C6 alkyl described above, in which one or more (e.g., 1, 2, 3, 4, or 5) hydrogen atoms are replaced by halogen. It will be understood by those skilled in the art that when the halogen substituent is more than one, the halogens may be the same or different and may be located on the same or different C atoms. Examples of "halogen-substituted C1-C6 alkyl" include, for example, -CH2F, -CHF2, -CF3, -CCl3, -C2F5, -C2Cl5, -CH2CF3, -CH2Cl, -CH2CH2CF3, or -CF(CF3)2.

[0072] As used herein, the term "cycloalkyl" refers to a monocyclic, fused polycyclic, bridged polycyclic, or spirocyclic non-aromatic saturated monovalent hydrocarbon ring structure having a specified number of ring atoms. A cycloalkyl group may have 3 to 12 carbon atoms (i.e., C3-C 12 The cycloalkyl group may be any of a plurality of cycloalkyl groups, for example, 3 to 10, 3 to 8, 3 to 7, 3 to 6, or 5 to 6 carbon atoms. Examples of suitable cycloalkyl groups include, but are not limited to, monocyclic structures such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, or cyclooctyl; or polycyclic (e.g., bicyclic) structures, including spiro, fused, or bridged systems such as bicyclo[1.1.1]pentyl, bicyclo[2.2.1]heptyl, spiro[3.4]octanyl, bicyclo[3.1.1]hexyl, bicyclo[3.1.1]heptyl, or bicyclo[3.2.1]octanyl.

[0073] The term "cycloalkenyl" as used herein means a monocyclic, fused polycyclic, bridged polycyclic, or spirocyclic non-aromatic unsaturated hydrocarbon ring structure having the specified number of ring atoms, containing at least one (e.g., 1, 2, or 3) carbon-carbon double bonds. A cycloalkenyl group may have 3 to 12 carbon atoms (i.e., C3-C 12 The term "cycloalkenyl" refers to a cycloalkyl group (e.g., a cycloalkyl group) having 3 to 10, 3 to 8, 3 to 7, 3 to 6, or 5 to 6 carbon atoms. Suitable cycloalkenyl groups include, but are not limited to, monocyclic structures such as cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclopentadienyl, cyclohexenyl, cyclohexadienyl, cycloheptenyl, cycloheptadienyl, cycloheptatrienyl, or cyclooctenyl.

[0074] As used herein, the term "heterocycloalkyl" refers to a monocyclic, fused polycyclic, spirocyclic, or bridged polycyclic non-aromatic saturated ring structure comprising one or more (e.g., 1, 2, 3, or 4) heteroatoms independently selected from O, N, and S and the specified number of ring atoms, or an N-oxide thereof, or an S-oxide or S-dioxide thereof. A heterocycloalkyl group may have 3 to 12 ring members (which may be referred to as a 3-12 membered heterocycloalkyl group), for example, 3 to 10 ring members, 3 to 8 ring members, 3 to 7 ring members, 4 to 7 ring members, 4 to 6 ring members, or 5 to 6 ring members. A heterocycloalkyl group typically contains up to 4 (e.g., 1, 2, 3, or 4) heteroatoms. Examples of suitable heterocycloalkyl groups include, but are not limited to, azetidinyl, oxetanyl, thietanyl, pyrrolidinyl (e.g., 1-pyrrolidinyl, 2-pyrrolidinyl, and 3-pyrrolidinyl), tetrahydrofuranyl (e.g., 1-tetrahydrofuranyl, 2-tetrahydrofuranyl, and 3-tetrahydrofuranyl), tetrahydrothiophenyl (e.g., 1-tetrahydrothiophenyl, 2-tetrahydrothiophenyl, and 3-tetrahydrothiophenyl), piperidinyl (e.g., 1-piperidinyl, 2-piperidinyl, 3-piperidinyl, and 4-piperidinyl), tetrahydropyranyl (e.g., 4-tetrahydropyranyl), tetrahydrothiopyranyl (e.g., 4-tetrahydrothiopyranyl), morpholinyl (e.g., morpholino), thiomorpholinyl, dioxanyl, piperazinyl, or azepanyl, diazepanyl, such as 1,4-diazepanyl, 3,6-diaza-bicyclo[3.1.1] The atom in the heterocycloalkyl group that is attached to the rest of the compound may be a carbon atom or a heteroatom, as long as it is chemically feasible.

[0075] Preferred heterocycloalkyl groups are, for example It is understood that structures having asymmetric centers encompass racemic and / or single enantiomeric forms thereof, e.g. Can represent and / or

[0076] The term "heterocycloalkenyl" as used herein means a "heterocycloalkyl" as defined herein that contains at least one (e.g., 1, 2, or 3) double bond. Examples of suitable heterocycloalkenyl groups include, but are not limited to:

[0077]

[0078] wherein each W is selected from CH2, NH, O and S; each Y is selected from NH, O, C(=O), SO2 and S; and each Z is selected from N and CH, provided that each ring contains at least one atom selected from N, O or S; for example, pyrrolinyl (e.g., 1-pyrrolinyl, 2-pyrrolidinyl, 3-pyrrolinyl, 4-pyrrolinyl or 5-pyrrolinyl), dihydrofuranyl (e.g., 1-dihydrofuranyl, 2-dihydrofuranyl, 3-dihydrofuranyl, 4-dihydrofuranyl or 5-dihydrofuranyl), dihydrothienyl (e.g., 1-dihydrothienyl, 2-dihydrothienyl, 3-dihydrothienyl or 4-dihydrothienyl), tetrahydropyridinyl (e.g., 1-, 2-, 3-, 4-, 5- or 6-tetrahydropyridinyl), tetrahydropyranyl (e.g., 4-tetrahydropyranyl) or tetrahydrothiopyranyl (e.g., 4-tetrahydrothiopyranyl).

[0079] As used herein, the term "aryl" refers to a monovalent aromatic hydrocarbon radical derived by removing one hydrogen atom from a single carbon atom in an aromatic ring system. Specifically, aryl refers to a monocyclic or fused polycyclic aromatic ring structure having the specified number of ring atoms. Specifically, the term includes radicals containing 6 to 14, e.g., 6 to 10, preferably 6, ring members. Specific aryl radicals include phenyl and naphthyl, with phenyl being the most specific.

[0080] As used herein, the term "heteroaryl" refers to a monocyclic or fused polycyclic aromatic ring structure comprising one or more (e.g., 1, 2, 3, or 4) heteroatoms independently selected from O, N, and S and the specified number of ring atoms, or an N-oxide thereof, or an S-oxide or S-dioxide thereof. Specifically, the aromatic ring structure may have 5 to 10 ring members. A heteroaryl group may be, for example, a 5-6 membered monocyclic ring, or a fused bicyclic structure formed by condensing two 6-membered rings, condensing two 5-membered rings, condensing a 6-membered ring and a 5-membered ring, or condensing a 5-membered ring and a 4-membered ring. The heteroaryl ring will typically contain up to 4 heteroatoms, more typically up to 3 heteroatoms, more typically up to 2 heteroatoms, for example, a single heteroatom independently selected from O, N, and S, wherein N and S may be in an oxidation state such as N-oxide, S=O, or S(O)2. In one embodiment, the heteroaryl ring contains at least one ring nitrogen atom, at least one ring sulfur atom, or at least one ring oxygen atom. For example, the heteroaryl group can be a fused ring containing 1, 2, 3 or 4 heteroatoms independently selected from N, O or S, such as benzofuran, benzothiophene, indole, benzimidazole, indazole, benzotriazole, pyrrolo[2,3-b]pyridine, pyrrolo[2,3-c]pyridine, pyrrolo[3,2-c]pyridine, pyrrolo[3,2-b]pyridine, imidazo[4,5-b]pyridine, imidazo[4,5-c]pyridine, pyrazolo[4,3-d]pyridine, pyrazolo[4,3-c]pyridine, pyrazolo[3,4-c]pyridine, pyrazolo[3,4-b]pyridine, isoindole, purine, indolizine, imidazo[1,2-a]pyridine, imidazo[1,5-a]pyridine, pyrazolo[1,5-a]pyridazine, pyrrolo[1,2-b]pyrimidine, imidazo[1,2-c]pyrimidine, 5H-pyrrolo[3,2-b]pyrazine, 1H-pyrazolo[4,3-b]pyrazine, 1H-pyrazolo[3,4-d]pyrimidine, 7H-pyrrolo[2,3-d]pyrimidine, quinoline, isoquinoline, cinnoline, quinazoline, quinoxaline, phthalazine, 1,6-

[0015] The heteroaryl group may be a 5- to 6-membered heteroaryl group comprising 1 or 2 heteroatoms independently selected from N, O, or S. Examples of suitable 5-membered monocyclic heteroaryl groups include, but are not limited to, pyrrolyl, furanyl, thienyl, imidazolyl, furazanyl, oxazolyl, oxadiazolyl, oxatriazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, triazolyl, and tetrazolyl; examples of suitable 6-membered monocyclic heteroaryl groups include, but are not limited to, pyridinyl, pyrazinyl, pyridazinyl, pyrimidinyl, and triazinyl.The atom in a heteroaryl group that is attached to the rest of the compound can be a carbon atom or a heteroatom, as long as it is chemically feasible.

[0081] Substituents are described as "optionally substituted" meaning that the group can be unsubstituted or substituted by one or more (e.g., 0, 1, 2, 3, 4, or 5 or more, or any range of derivatives thereof) of the substituents listed for the group, wherein the substituents can be the same or different. In one embodiment, the optionally substituted group is substituted by 1 substituent. In another embodiment, the optionally substituted group is substituted by 2 substituents. In another embodiment, the optionally substituted group is substituted by 3 substituents. In another embodiment, the optionally substituted group is substituted by 4 substituents.

[0082] Those skilled in the art of organic synthesis understand that a stable, chemically feasible heterocycle, whether aromatic or non-aromatic, wherein the maximum number of heteroatoms or the type of heteroatoms present is determined by the ring size, the degree of unsaturation, and the valence of the heteroatoms. Generally speaking, a heterocycle may have from 1 to 4 heteroatoms, provided that the heterocycle or heteroaromatic ring is chemically feasible and stable.

[0083] As used herein, the term "hydroxy" refers to an -OH group.

[0084] As used herein, the term "mercapto" refers to a -SH group.

[0085] As used herein, the term "nitro" refers to a -NO2 group.

[0086] As used herein, the term "optionally substituted" means, unless otherwise indicated, that a group may be unsubstituted or substituted with one or more (e.g., 0, 1, 2, 3, 4, or 5 or more, or any range derivatizable therein) of the substituents listed for the group, wherein the substituents may be the same or different. In one embodiment, the optionally substituted group has 1 substituent. In another embodiment, the optionally substituted group has 2 substituents. In another embodiment, the optionally substituted group has 3 substituents. In another embodiment, the optionally substituted group has 4 substituents. In another embodiment, the optionally substituted group has 5 substituents.

[0087] Unless otherwise specified, C in the definition of compounds of the present invention is n-n+m or C n -C m Including various cases from n to n+m carbons, such as C 1-6 Including C1, C2, C3, C4, C5 and C6, and also including any range from n to n+m, such as C 0-6 Including C1, C2, C3, C4, C5, C6, C 0-1 、C0-2 、C 0-3 、C 0-4 、C 0-5 、C 1-2 、C 1-3 、C 1- 4. C 2-3 etc., C. 1-6 Including C 1-2 、C 1-3 、C 1-4 、C 2-6 、C 3-6 Similarly, the n-membered to n+m-membered in the definition of the compounds of the present invention means that the number of ring atoms is n to n+m, for example, 3-12-membered ring includes 3-membered ring, 4-membered ring, 5-membered ring, 6-membered ring, 12-membered ring, etc., and also includes any range of n to n+m-membered, for example, 3-12-membered ring includes 3-6-membered ring, 3-8-membered ring, 3-9-membered ring, 4-7-membered ring, 4-5-membered ring, 5-6-membered ring, 5-7-membered ring, 5-8-membered ring, 5-9-membered ring, 6-7-membered ring, 6-8-membered ring and 6-10-membered ring, etc.

[0088] As used in this specification and the claims that follow, the word "comprise" and variations of the word such as "include" and "comprising" mean "including but not limited to," and are not intended to exclude, for example, other additives, ingredients, integers, or steps. When an element is described as comprising a plurality of ingredients, steps, or conditions, it should be understood that the element may also be described as comprising any combination of the plurality of ingredients, steps, or conditions, or "consisting of" or "consisting essentially of" the plurality or combination of ingredients, steps, or conditions.

[0089] It should be understood that the dosages referred to herein when describing the compounds of the present invention, pharmaceutical compositions, pharmaceutical combinations, kits containing the same, and related uses and methods are based on the weight of the free form and do not include any salts, hydrates or solvates thereof, unless the description indicates that the dosage is based on the weight of the salt, hydrate or solvate.

[0090] Compounds of the present invention

[0091] The terms "inventive compound" and "compound of the present invention" and the like as used throughout this application, unless otherwise indicated, encompass compounds of formula (I) or formula (II) as defined in the various embodiments herein and preferred embodiments thereof, including isomers thereof, including atropisomers, enantiomeric mixtures, in particular racemates, diastereomeric mixtures, geometric isomers, tautomers, solvates, metabolites, isotopic variants, salts (e.g., pharmaceutically acceptable salts) and prodrugs.

[0092] Therefore, the above-mentioned various isomers and derivatives of the compound of formula I are thus encompassed within the scope of the present invention, and their respective meanings, preparations and specific examples are as defined in the "Definitions" section above, or are well known in the art. In some embodiments, metabolites, isotopic variants or prodrugs and any combination thereof are excluded as appropriate. However, preferably, it is a substantially pure enantiomer (enantiomerically pure) or diastereomer of a compound of formula (I) or formula (II) and / or a pharmaceutically acceptable salt thereof.

[0093] The present invention also encompasses N-oxides of compounds of formula (I) or (II) where these compounds contain a basic nitrogen atom, such as a nitrogen atom present in a nitrogen-containing heterocycle. Certain compounds of the present invention may exist in polymorphic or amorphous forms and therefore also fall within the scope of the present invention.

[0094] In one aspect, the present invention provides a compound of formula (I), an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof,

[0095]

[0096] in,

[0097] A is selected from CR a or N, where R a Selected from halogen, CN, nitro, C 3-6 Cycloalkyl or C optionally substituted by halogen 1-6 alkyl;

[0098] R 1 、R 2 and R 3 are each independently selected from H, halogen or C 1-6 Alkyl, wherein the alkyl group is optionally independently selected from halogen, -N(R c )2、-OR c or substituted with a 3-8 membered heterocycloalkyl group;

[0099] R b is independently selected at each occurrence from H, halogen, CN, or C optionally substituted with halogen or CN 1-6 alkyl;

[0100] X is selected from a bond, -O- or -NH-;

[0101] R 4 Selected from H, halogen, C 1-6 Alkyl, -C 0-6 Alkyl-C 6-10 Aryl, -C 0-6 Alkyl-C 3-8 Cycloalkyl, -C 0-6 Alkyl-C 3-8Cycloalkenyl, -C 0-6 Alkyl-3-8 membered heterocycloalkyl, -C 0-6 Alkyl-3-8 membered heterocycloalkenyl, -C 0-6 Alkyl-5-10 membered heteroaryl, wherein said alkyl, aryl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl and heteroaryl are optionally substituted by one or more groups independently selected from the group consisting of halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -SR c 、-(CR c R c ) 0-6 -(CO) 0-1 -OR c 、-(CR c R c ) 0-6 -(CO) 0-1 -N(R c )2, wherein -(CR c R c ) 0-6 -(CO) 0-1 -N(R c ) 2 optionally through the group connected to N, together with the atoms on the cyclic group to which it is connected and the adjacent atoms to form a 4-7 membered nitrogen-containing heterocyclic ring;

[0102] R c is independently selected at each occurrence from H or C optionally substituted by halogen 1-6 Alkyl, or two R c Each independently forms a 3-6 membered cyclic group together with the carbon atom or nitrogen atom to which they are attached;

[0103] E is selected from halogen, -OR d or -N(R d )2-, where R d are each independently H or C optionally substituted by halogen 1-6 alkyl;

[0104] R 5 for

[0105] m is 0 or 1;

[0106] represents an aromatic ring;

[0107] B and D are each independently selected from N or C;

[0108] Z, G and Y are each independently selected from C, N, O or S;

[0109] R 6 、R 7 and R 8 are each independently selected from H, halogen and C optionally substituted by halogen 1-6 alkyl;

[0110] The condition is that B and D are not N at the same time; and at most three of Z, G, Y, D, and B are not C;

[0111] or an isomer, pharmaceutically acceptable salt or solvate thereof.

[0112] In one embodiment of the compound of formula (I), A is N.

[0113] In one embodiment of the compound of formula (I), A is CR a , where R a Selected from halogen.

[0114] In one embodiment of the compound of formula (I), A is CR a , where R a is F or Cl, preferably Cl.

[0115] In one embodiment of the compound of formula (I), R 1 is selected from H or halogen.

[0116] In one embodiment of the compound of formula (I), R 1 H or F.

[0117] In one embodiment of the compound of formula (I), R 2 and R 3 Both are H.

[0118] In one embodiment of the compound of formula (I), R b For H.

[0119] In one embodiment of the compound of formula (I), R b Not H, and there is 1 R b Or there are 2 R b .

[0120] In one embodiment of the compounds of formula (I), there is one R b And R b C 1-6 alkyl.

[0121] In a specific embodiment, there is an R b And R b C1-6 Alkyl, preferably C 1-3 Alkyl; in more specific embodiments, there is one R b And R b is -CH3; in a more specific embodiment, there is one R b And R b for In a more specific embodiment, there is one R b And R b for And the connection mode on the piperazine ring is In a more specific embodiment, there is one R b And R b for And the connection mode on the piperazine ring is

[0122] In one embodiment of the compounds of formula (I), there is one R b And R b CN; in a specific embodiment, there is one R b And R b is CN and is attached to the carbon atom ortho to the amide nitrogen.

[0123] In one embodiment of the compounds of formula (I), there is one R b And R b C 1-6 Alkyl, preferably C 1-3 Alkyl, substituted with CN; in a specific embodiment, there is one R b And R b is cyanomethyl; in a more specific embodiment, there is one R b And R b is a cyanomethyl group, and the connection mode on the piperazine ring is

[0124] In one embodiment of the compounds of formula (I), there are two R b And R b C 1-6 Alkyl, preferably C 1-3 Alkyl; in a specific embodiment, there are two R b And R b is -CH3; in a more specific embodiment, there are two R b And R b is -CH3, and the connection mode on the piperazine ring is

[0125] In one embodiment of the compounds of formula (I), X is a bond.

[0126] In one embodiment of the compounds of formula (I), X is -O- or -NH-.

[0127] In one embodiment of the compounds of formula (I), X is -O-.

[0128] In one embodiment of the compounds of formula (I), X is -NH-.

[0129] In one embodiment of the compound of formula (I), R 4 Selected from H, halogen, C 1-6 Alkyl, -C 0-3 Alkyl-phenyl, -C 0-3 Alkyl-C 3-6 Cycloalkyl, -C 0-3 Alkyl-C 3-6 Cycloalkenyl, -C 0-3 Alkyl-4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, -C 0-3 Alkyl-5-6 membered heterocycloalkenyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S and -C 0-3 Alkyl-5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, wherein said alkyl, phenyl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl and heteroaryl are optionally substituted by 1, 2 or 3 groups independently selected from halogen, C optionally substituted by halogen 1-6 Alkyl, -(CR c R c ) 0-6 -SR c 、-(CR c R c ) 0-6 -OR c 、-(CR c R c ) 0-6 -N(R c )2, wherein -(CR c R c ) 0-6 -N(R c )2 optionally forms a 4-7 membered nitrogen-containing heterocyclic ring through the group connected to N, together with the atoms on the cyclic group to which it is connected and the adjacent atoms.

[0130] In one embodiment of the compound of formula (I), R 4 Selected from H, halogen, C 1-6 Alkyl, -C 0-3 Alkyl-phenyl, -C 0-3Alkyl-C 3-6 Cycloalkyl, -C 0-3 Alkyl-4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S and -C 0-3 Alkyl-5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, wherein said alkyl, phenyl, cycloalkyl, heterocycloalkyl and heteroaryl are optionally substituted by 1, 2 or 3 groups independently selected from halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -SR c 、-(CR c R c ) 0-6 -OR c 、-(CR c R c ) 0-6 -N(R c )2, wherein -(CR c R c ) 0-6 -N(R c )2 optionally forms a 4-7 membered nitrogen-containing heterocyclic ring through the group connected to N, together with the atoms on the cyclic group to which it is connected and the adjacent atoms.

[0131] In one embodiment of the compound of formula (I), R 4 Selected from H, halogen, C 1-6 Alkyl, phenyl, C 3-6 cycloalkyl, 4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, and 5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, wherein said alkyl, phenyl, cycloalkyl, heterocycloalkyl and heteroaryl are optionally substituted by 1, 2 or 3 groups independently selected from halogen, C 2-3 optionally substituted by halogen, C 3-4 optionally substituted by halogen, C 5-6 optionally substituted by halogen, C 6-7 optionally substituted by halogen, C 7-8 optionally substituted by halogen, C 8-9 optionally substituted by halogen, C 9-10 optionally substituted by halogen, C 10-11 optionally substituted by halogen, C 11-12 optionally substituted by halogen, C 12-13 optionally substituted by halogen, C 13-14 optionally substituted by hal 1-6 Alkyl, -(CR c R c ) 0-6 - OR c 、-(CR c R c ) 0-6 -N(R c )2, wherein -(CR c R c ) 0-6 -N(R c)2 optionally forms a 4-7 membered nitrogen-containing heterocyclic ring through the group connected to N, together with the atoms on the cyclic group to which it is connected and the adjacent atoms.

[0132] In one embodiment of the compound of Formula (I), -XR 4 For H.

[0133] In one embodiment of the compound of Formula (I), -XR 4 Not for H.

[0134] In one embodiment of the compound of Formula (I), -XR 4 It is -OH.

[0135] In one embodiment of the compound of Formula (I), -XR 4 is -NH2.

[0136] In one embodiment of the compound of Formula (I), -XR 4 is halogen; in a specific embodiment, -XR 4 is Cl or F.

[0137] In one embodiment of the compound of formula (I), R 4 C 1-6 Alkyl, optionally substituted by 1, 2 or 3 independently selected from halogen, C 1-6 Alkyl, hydroxyl, C 1-6 Alkoxy and -(CR c R c ) 0-6 -N(R c ) 2 groups are substituted. In a specific embodiment, exemplary R 4Including but not limited to -CH3, -CH2CH3, -CH2CH2CH3, -CH(CH3)CH3, -CH2CH(CH3)CH3, -CH2CH2CH2CH3, -C(CH3)(CH3)(CH3), -CH2F, -CHF2, -CF3, -CH2CF3, -CH2CH2CF3, - CH(CF3)CH3, -CH2CH(CF3)CH3, -CH2CH2CH2CF3, -C(CF3)(CH3)(CH3), -CH2OH, -CH2CH2OH, -CH2CH2CH2OH, -CH2CH(OH)CH3, -CH2CH(CH2OH)CH3, -CH2CH2CH2CH2OH, - C(CH2OH)(CH3)(CH3)、-CH2OCH3、-CH2OCH2CH3、-CH2CH2OCH3、-CH2CH2OCH2CH3、- CH2CH2CH2OCH3, -CH2CH(OCH3)CH3, -CH2CH(CH2OCH3)CH3, -CH2CH2CH2CH2OCH3, -NH2, -NHCH3, -NHCH2CH3, -N(CH3)2, -N(CH3)(CH2CH3), -N(CH2CH3)2, -CH2NH2, -CH2NHCH3, -CH2N(CH3)2, -CH2NHCH2CH3, -CH2N(CH3)(CH2CH3), -CH2N(CH2CH3)2, -CH2CH2NH2, - CH2CH2NHCH3, -CH2CH2N(CH3)2, -CH(CH3)N(CH3)2, CH2CH2NHCH2CH3, -CH2CH2N(CH2CH3)2, -CH2CH2CH2NH2, -CH2CH2CH2NHCH3, -CH2CH2CH2N(CH3)2, - CH2CH(NH2)CH3, -CH2CH(CH2NH2)CH3, -CH2CH2CH2CH2NH2, -CH2CH2CH2CH2NHCH3, and -CH2CH2CH2CH2N(CH3)2.

[0138] In a preferred embodiment, R 4 C 1-6 Alkyl, optionally substituted by 1, 2 or 3 independently selected from halogen, C 1-6 Alkyl, C 1-6 More preferably, R 4 Selected from -CH3, -CH2CH3, -CH2CH2CH3, -CH(CH3)CH3, -CH2OCH3, -CH2CH2OCH3, -CH2CH2OCH2CH3 or -CF3.

[0139] In one embodiment of the compound of Formula (I), -XR 4 C 1-6 Alkyl, where C 1-6 Alkyl groups have the above pairs as R 4 C 1-6 Alkyl is given the meaning of the embodiment, preferred or more preferred embodiment. In one embodiment of the compound of formula (I), -XR 4 For-OC 1-6 Alkyl, where C 1-6 Alkyl groups have the above pairs as R 4 C 1-6 In one embodiment of the compound of formula (I), -XR 4 -NH-C 1-6 Alkyl, where C 1-6 Alkyl groups have the above pairs as R 4 C 1-6 Alkyl means an embodiment, a preferred or a more preferred embodiment.

[0140] In one embodiment of the compound of formula (I), R 4 -C 0-3 Alkyl-phenyl, preferably phenyl, said phenyl being optionally substituted by 1, 2 or 3 independently selected from halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c and-(CR c R c ) 0-6 -N(R c)2. In specific embodiments, exemplary substituents include, but are not limited to, F, Cl, Br, I, -CH3, -CH2CH3, -CH2CH2CH3, -CH(CH3)CH3, -CH2CH(CH3)CH3, -CH2CH2CH2CH3, -C(CH3)(CH3)(CH3), -CH2F, -CHF2, -CF3, -CH2CF3, -CH2CH2CF3, -CH(CF3)CH3, -CH2CH(CF3)CH3, -CH2CH2CH2CF3, -C(CF3)(CH3)(CH3), -CH2OH, -CH2CH2OH, -CH2CH2CH2OH, -CH2CH(OH)CH3, - CH2CH(CH2OH)CH3, -CH2CH2CH2CH2OH, -C(CH2OH)(CH3)(CH3), -OH, -OCH3, -OCH2CH3, -OCH2CH2CH3, -OCH(CH3)(CH3), -CH2OCH3, -CH2OCH2CH3, -CH2CH2OCH3, - CH2CH2OCH2CH3, -CH2CH2CH2OCH3, -CH2CH(OCH3)CH3, -CH2CH(CH2OCH3)CH3, - CH2CH2CH2CH2OCH3, -NH2, -NHCH3, -NHCH2CH3, -N(CH3)2, -N(CH3)(CH2CH3), -N(CH2CH3)2, -CH2NH2, -CH2NHCH3, -CH2N(CH3)2, -CH2NHCH2CH3, -CH2N(CH3)(CH2CH3), - CH2N(CH2CH3)2, -CH2CH2NH2, -CH2CH2NHCH3, -CH2CH2N(CH3)2, -CH(CH3)N(CH3)2, CH2CH2NHCH2CH3, -CH2CH2N(CH2CH3)2, -CH2CH2CH2NH2, -CH2CH2CH2NHCH3, - CH2CH2CH2N(CH3)2, -CH2CH(NH2)CH3, -CH2CH(CH2NH2)CH3, -CH2CH2CH2CH2NH2, - CH2CH2CH2CH2NHCH3, and -CH2CH2CH2CH2N(CH3)2.

[0141] In a preferred embodiment, R 4 is phenyl, substituted by 1, 2 or 3 independently selected from halogen, optionally substituted by halogen, C 1-6 Alkyl and -(CR c R c ) 0-6 -N(Rc )2, exemplary substituents include, but are not limited to, F, Cl, Br, I, -CH3, -CH2CH3, -CH2CH2CH3, -CH(CH3)CH3, -CH2CH(CH3)CH3, -CH2CH2CH2CH3, -C(CH3)(CH3)(CH3), -CH2F, -CHF2, -CF3, -CH2CF3, -CH2CH2CF3, -CH(CF3)CH3, -CH2CH(CF3)CH3, -CH2CH2CH2CF3, -C(CF3)(CH3)(CH3), -NH2, -NHCH3, -NHCH2CH3, -N(CH3), -N(CH3)(CH2CH3), -N(CH2CH3), -CH2NH2, -CH2NHCH3, -CH2N(CH3), -CH2NHCH2CH3, -CH2N(CH3)(CH2CH3), -CH2N(CH2CH3)2, -CH2CH2NH2, -CH2CH2NHCH3, -CH2CH2N(CH3)2, -CH(CH3)N(CH3)2, CH2CH2NHCH2CH3, -CH2CH2N(CH2CH3)2, -CH2CH2CH2NH2, - More preferably, R 4 Selected from

[0142] In one embodiment of the compound of formula (I), R 4 -C 0-3 Alkyl-phenyl, preferably phenyl, is -(CR c R c ) 0-6 -N(R c )2 substituted, wherein R c Each independently is H or C 1-6 Alkyl, and wherein R on N c One of them forms a 4-7 membered nitrogen-containing heterocyclic ring together with the atoms on the benzene ring to which it is connected and the adjacent atoms. 4 Selected from Each R cIndependently selected from H, -CH3, -CH2CH3, -CH2CH2CH3, -CH(CH3)CH3, -CH2CH(CH3)CH3, -CH2CH2CH2CH3 and -C(CH3)(CH3)(CH3).

[0143] In a preferred embodiment, R 4 for

[0144] In one embodiment of the compound of Formula (I), -XR 4 -C 0-6 Alkyl-phenyl, preferably -C 0-3 Alkyl-phenyl, more preferably phenyl, having the above pair as R 4 In one embodiment of the compound of formula (I), -XR 4 For-OC 0-6 Alkyl-phenyl, preferably -OC 0-3 Alkyl-phenyl, more preferably -O-phenyl, wherein the phenyl group has the above pair as R 4 In one embodiment of the compound of formula (I), -XR 4 -NH-C 0-6 Alkyl-phenyl, preferably C 0-3 Alkyl-phenyl, more preferably -NH-phenyl, wherein the phenyl group has the above pair as R 4 The phenyl group has the meaning given in the embodiment, preferred or more preferred embodiment.

[0145] In one embodiment of the compound of formula (I), R 4 -C 0-3 Alkyl-C 3-6 Cycloalkyl, preferably -C 3-6 Cycloalkyl, said cycloalkyl being optionally substituted by 1, 2 or 3 independently selected from halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c and- (CR c R c ) 0-6 -N(R c)2. In specific embodiments, exemplary substituents include, but are not limited to, F, Cl, Br, I, -CH3, -CH2CH3, -CH2CH2CH3, -CH(CH3)CH3, -CH2CH(CH3)CH3, -CH2CH2CH2CH3, - C(CH3)(CH3)(CH3), -CH2F, -CHF2, -CF3, -CH2CF3, -CH2CH2CF3, -CH(CF3)CH3, - CH2CH(CF3)CH3, -CH2CH2CH2CF3, -C(CF3)(CH3)(CH3), -CH2OH, -CH2CH2OH, - CH2CH2CH2OH, -CH2CH(OH)CH3, -CH2CH(CH2OH)CH3, -CH2CH2CH2CH2OH, - C(CH2OH)(CH3)(CH3), -OH, -OCH3, -OCH2CH3, -OCH2CH2CH3, -OCH(CH3)(CH3), -CH2OCH3, -CH2OCH2CH3, -CH2CH2OCH3, -CH2CH2OCH2CH3, -CH2CH2CH2OCH3, -CH2CH(OCH3)CH3, -CH2CH(CH2OCH3)CH3, -CH2CH2CH2CH2OCH3, -NH2, -NHCH3, -NHCH2CH3, -N(CH3)2, -N(CH3)(CH2CH3), -N(CH2CH3)2, -CH2NH2, -CH2NHCH3, -CH2N(CH3)2, -CH2NHCH2CH3, -CH2N(CH3)(CH2CH3), -CH2N(CH2CH3)2, -CH2CH2NH2, -CH2CH2NHCH3, -CH2CH2N(CH3)2, -CH(CH3)N(CH3)2, CH2CH2NHCH2CH3, -CH2CH2N(CH2CH3)2, -CH2CH2CH2NH2, - CH2CH2CH2NHCH3, -CH2CH2CH2N(CH3)2, -CH2CH(NH2)CH3, -CH2CH(CH2NH2)CH3, - CH2CH2CH2CH2NH2, -CH2CH2CH2CH2NHCH3, and -CH2CH2CH2CH2N(CH3)2.

[0146] In a preferred embodiment, R 4 is selected from cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl; more preferably cyclopropyl.

[0147] In one embodiment of the compound of Formula (I), -XR 4 -C0-6 Alkyl-C 3-6 Cycloalkyl, preferably -C 0-3 Alkyl-C 3-6 Cycloalkyl, more preferably C 3-6 Cycloalkyl, said cycloalkyl having the meanings given above for the embodiments, preferred or more preferred embodiments of cycloalkyl in R4. In one embodiment of the compounds of formula (I), -XR 4 For-OC 0-6 Alkyl-C 3-6 Cycloalkyl, preferably -OC 0-3 Alkyl-C 3-6 Cycloalkyl, more preferably -OC 3-6 Cycloalkyl, the cycloalkyl having the above pair as R 4 In one embodiment of the compound of formula (I), -XR 4 -NH-C 0-6 Alkyl-C 3-6 Cycloalkyl, preferably -NH-C 0-3 Alkyl-C 3-6 Cycloalkyl, more preferably -NH-C 3-6 Cycloalkyl, the cycloalkyl having the above pair as R 4 The cycloalkyl group in the embodiment, preferred or more preferred embodiment has the meaning given in the preceding sentence.

[0148] In one embodiment of the compound of formula (I), R 4 -C 0-3 Alkyl-containing 4-6 membered heterocycloalkyl groups comprising 1, 2 or 3 heteroatoms independently selected from N, O or S, preferably containing 4-6 membered heterocycloalkyl groups comprising 1, 2 or 3 heteroatoms independently selected from N, O or S, wherein the heterocycloalkyl groups are optionally substituted by 1, 2 or 3 groups independently selected from halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c and-(CR c R c ) 0-6 -N(R c)2. In specific embodiments, exemplary substituents include, but are not limited to, F, Cl, Br, I, -CH3, -CH2CH3, -CH2CH2CH3, -CH(CH3)CH3, -CH2CH(CH3)CH3, -CH2CH2CH2CH3, -C(CH3)(CH3)(CH3), -CH2F, -CHF2, -CF3, -CH2CF3, -CH2CH2CF3, - CH(CF3)CH3, -CH2CH(CF3)CH3, -CH2CH2CH2CF3, -C(CF3)(CH3)(CH3), -CH2OH, -CH2CH2OH, -CH2CH2CH2OH, -CH2CH(OH)CH3, -CH2CH(CH2OH)CH3, -CH2CH2CH2CH2OH, - C(CH2OH)(CH3)(CH3), -OH, -OCH3, -OCH2CH3, -OCH2CH2CH3, -OCH(CH3)(CH3), -CH2OCH3, -CH2OCH2CH3, -CH2CH2OCH3, -CH2CH2OCH2CH3, -CH2CH2CH2OCH3, -CH2CH(OCH3)CH3, -CH2CH(CH2OCH3)CH3, -CH2CH2CH2CH2OCH3, -NH2, -NHCH3, -NHCH2CH3, -N(CH3)2, -N(CH3)(CH2CH3), -N(CH2CH3)2, -CH2NH2, -CH2NHCH3, -CH2N(CH3)2, -CH2NHCH2CH3, -CH2N(CH3)(CH2CH3), -CH2N(CH2CH3)2, -CH2CH2NH2, -CH2CH2NHCH3, -CH2CH2N(CH3)2, -CH(CH3)N(CH3)2, CH2CH2NHCH2CH3, -CH2CH2N(CH2CH3)2, -CH2CH2CH2NH2, -CH2CH2CH2NHCH3, -CH2CH2CH2N(CH 3)2, -CH2CH(NH2)CH3, -CH2CH(CH2NH2)CH3, -CH2CH2CH2CH2NH2, -CH2CH2CH2CH2NHCH3 and -CH2CH2CH2CH2N(CH3)2.

[0149] In one embodiment of the compound of formula (I), R 4The 4-6 membered heterocycloalkyl group in the group is selected from azetidinyl, oxetanyl, thietanyl, pyrrolidinyl (e.g., 1-pyrrolidinyl, 2-pyrrolidinyl, and 3-pyrrolidinyl), tetrahydrofuranyl (e.g., 1-tetrahydrofuranyl, 2-tetrahydrofuranyl, and 3-tetrahydrofuranyl), tetrahydrothiophenyl (e.g., 1-tetrahydrothiophenyl, 2-tetrahydrothiophenyl, and 3-tetrahydrothiophenyl), piperidinyl (e.g., 1-piperidinyl, 2-piperidinyl, 3-piperidinyl, and 4-piperidinyl), tetrahydropyranyl (e.g., 4-tetrahydropyranyl), tetrahydrothiopyranyl (e.g., 4-tetrahydrothiopyranyl), morpholinyl (e.g., morpholino), thiomorpholinyl, dioxanyl, or piperazinyl.

[0150] In a preferred embodiment, R 4 Selected from The most preferred 4 Selected from

[0151] In one embodiment of the compound of Formula (I), -XR 4 -C 0-6 Alkyl-containing 4-6 membered heterocycloalkyl groups containing 1, 2 or 3 heteroatoms independently selected from N, O or S, preferably -C 0-3 Alkyl-containing 4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, more preferably containing 1, 2 or 3 heteroatoms independently selected from N, O or S, the heterocycloalkyl having the above as R 4 In one embodiment of the compound of formula (I), -XR 4 For-OC 0-6 Alkyl-4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, preferably -OC 0-3 Alkyl-5-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, more preferably 4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, said heterocycloalkyl having the above as R 4 In one embodiment of the compound of formula (I), -XR 4 -NH-C 0-6 Alkyl-4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, preferably -NH-C 0-3Alkyl-containing 5-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, more preferably containing 4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, said heterocycloalkyl having the above as R 4 The heterocycloalkyl group in the embodiment, preferred or more preferred embodiment has the meaning given above.

[0152] In one embodiment of the compound of formula (I), R 4 -C 0-3 Alkyl-5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, preferably 5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, wherein the heteroaryl is optionally substituted by 1, 2 or 3 groups independently selected from halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c and-(CR c R c ) 0-6 -N(R c)2. In specific embodiments, exemplary substituents include, but are not limited to, F, Cl, Br, I, -CH3, -CH2CH3, -CH2CH2CH3, -CH(CH3)CH3, -CH2CH(CH3)CH3, -CH2CH2CH2CH3, -C(CH3)(CH3)(CH3), -CH2F, -CHF2, -CF3, -CH2CF3, -CH2CH2CF3, - CH(CF3)CH3, -CH2CH(CF3)CH3, -CH2CH2CH2CF3, -C(CF3)(CH3)(CH3), -CH2OH, -CH2CH2OH, -CH2CH2CH2OH, -CH2CH(OH)CH3, -CH2CH(CH2OH)CH3, -CH2CH2CH2CH2OH, - C(CH2OH)(CH3)(CH3), -OH, -OCH3, -OCH2CH3, -OCH2CH2CH3, -OCH(CH3)(CH3), -CH2OCH3, -CH2OCH2CH3, -CH2CH2OCH3, -CH2CH2OCH2CH3, -CH2CH2CH2OCH3, -CH2CH(OCH3)CH3, -CH2CH(CH2OCH3)CH3, -CH2CH2CH2CH2OCH3, -NH2, -NHCH3, -NHCH2CH3, -N(CH3)2, -N(CH3)(CH2CH3), -N(CH2CH3)2, -CH2NH2, -CH2NHCH3, -CH2N(CH3)2, -CH2NHCH2CH3, -CH2N(CH3)(CH2CH3), -CH2N(CH2CH3)2, -CH2CH2NH2, -CH2CH2NHCH3, -CH2CH2N(CH3)2, -CH(CH3)N(CH3)2, CH2CH2NHCH2CH3, -CH2CH2N(CH2CH3)2, -CH2CH2CH2NH2, -CH2CH2CH2NHCH3, -CH2CH2CH2N(CH 3)2, -CH2CH(NH2)CH3, -CH2CH(CH2NH2)CH3, -CH2CH2CH2CH2NH2, -CH2CH2CH2CH2NHCH3 and -CH2CH2CH2CH2N(CH3)2.

[0153] In one embodiment of the compound of formula (I), R 4 The 5-6 membered heteroaryl group is selected from pyrrolyl, furanyl, thienyl, imidazolyl, furazanyl, oxazolyl, oxadiazolyl, oxatriazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, triazolyl and tetrazolyl, pyridyl, pyrazinyl, pyridazinyl, pyrimidinyl and triazinyl.

[0154] In a preferred embodiment, R 4 Selected from Most preferably, R 4 Selected from

[0155] In one embodiment of the compound of Formula (I), -XR 4 -C 0-6 Alkyl-5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, preferably -C 0-3 Alkyl-containing 5-6 membered heteroaryl groups containing 1, 2 or 3 heteroatoms independently selected from N, O or S, more preferably containing 5-6 membered heteroaryl groups containing 1, 2 or 3 heteroatoms independently selected from N, O or S, said heteroaryl groups having the above conditions for R 4 In one embodiment of the compound of formula (I), -XR 4 For-OC 0-6 Alkyl-5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, preferably -OC 0-3 Alkyl-containing 5-6 membered heteroaryl groups containing 1, 2 or 3 heteroatoms independently selected from N, O or S, more preferably containing 5-6 membered heteroaryl groups containing 1, 2 or 3 heteroatoms independently selected from N, O or S, said heteroaryl groups having the above conditions for R 4 In one embodiment of the compound of formula (I), -XR 4 -NH-C 0-6 Alkyl-5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, preferably -NH-C 0-3 Alkyl-containing 5-6 membered heteroaryl groups containing 1, 2 or 3 heteroatoms independently selected from N, O or S, more preferably containing 5-6 membered heteroaryl groups containing 1, 2 or 3 heteroatoms independently selected from N, O or S, said heteroaryl groups having the above conditions for R 4 The heteroaryl group of the present invention has the meaning given in the embodiment, preferred or more preferred embodiment.

[0156] In one embodiment of the compounds of formula (I), as R 4 Substituent -(CR c R c ) 0-6 -SR c 、-(CR c R c ) 0-6 -(CO)0-1 - OR c 、-(CR c R c ) 0-6 -(CO) 0-1 -N(R c )2 are each preferably -(CR c R c ) 0-6 -SR c 、-(CR c R c ) 0-6 -OR c 、-(CR c R c ) 0-6 -N(R c )2; More preferably -(CR c R c ) 0-3 -SR c 、-(CR c R c ) 0-3 -OR c 、-(CR c R c ) 0-3 -N(R c )2.

[0157] In one embodiment of the compound of formula (I), E is halogen, preferably F.

[0158] In one embodiment of the compounds of formula (I), E is -OR d , R d is C optionally substituted by halogen 1-6 Alkyl, preferably C 1-6 Alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, trifluoromethyl, difluoromethyl, trifluoroethyl, and the like.

[0159] In one embodiment of the compound of formula (I), R 5 for Among them, at most two of Z, G, Y, B, and D are not C. Exemplary R 5 Including but not limited to

[0160] In a preferred embodiment, R 5 for Among them, at most two of Z, Y, B, and D are not C. Exemplary R 5 Including but not limited to

[0161] In a more preferred embodiment, R 5 for The most preferred 5 for

[0162] In a preferred embodiment, R 5 for More preferred

[0163] In one embodiment of the compound of formula (I), R 6 、R 7 and R 8 At least one of them is halogen, preferably F. In a more preferred embodiment, R 6 is F, and R 7 and R 8 For H.

[0164] In one embodiment of the compound of formula (I), R 6 、R 7 and R 8 At least one of the C 1-6 Alkyl, preferably -CF3.

[0165] In one embodiment of the compound of formula (I), R c H or C 1-6 alkyl.

[0166] The present invention also provides a compound of formula (II), an isomer, a pharmaceutically acceptable salt or a solvate thereof.

[0167]

[0168] where R 1 、R 2 、R 3 、R 4 、R 6 、R 7 、R 8 ,A,E,R b each has the meaning given above for the compounds of formula (I).

[0169] In a preferred embodiment of the compound of formula (II),

[0170] A is selected from CR a or N, where R a selected from halogen;

[0171] R 1 、R 2 and R 3are each independently selected from H, halogen or C 1-6 alkyl;

[0172] R b is independently selected at each occurrence from H, halogen, CN, or C optionally substituted with halogen or CN 1-6 alkyl;

[0173] X is selected from a bond, -O- or -NH-;

[0174] R 4 Selected from H, halogen, C 1-6 Alkyl, -C 0-3 Alkyl-C 6-10 Aryl, -C 0-3 Alkyl-C 3-8 Cycloalkyl, -C 0-3 Alkyl-C 3-8 Cycloalkenyl, -C 0-3 Alkyl-3-8 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, -C 0-3 Alkyl-3-8 membered heterocycloalkenyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, -C 0-3 Alkyl-5-10 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, wherein said alkyl, aryl, cycloalkyl, cycloalkenyl, heterocycloalkyl, heterocycloalkenyl and heteroaryl are optionally substituted by 1, 2 or 3 groups independently selected from the following: halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c 、-(CR c R c ) 0-6 -N(R c )2, wherein -(CR c R c ) 0-6 -N(R c ) 2 optionally through the group connected to N, together with the atoms on the cyclic group to which it is connected and the adjacent atoms to form a 4-7 membered nitrogen-containing heterocyclic ring;

[0175] R c is independently selected at each occurrence from H or C optionally substituted by halogen 1-6 alkyl;

[0176] E is selected from halogen, -OR d or -N(R d )2-, where R dare each independently H or C optionally substituted by halogen 1-6 alkyl;

[0177] R 6 、R 7 and R 8 are each independently selected from H, halogen and C optionally substituted by halogen 1-6 alkyl;

[0178] or an isomer, pharmaceutically acceptable salt or solvate thereof.

[0179] In a preferred embodiment of the compound of formula (II), A is CR a , where R a Selected from F or Cl, preferably Cl.

[0180] In a preferred embodiment of the compound of formula (II), R b For H.

[0181] In a preferred embodiment of the compound of formula (II), R b Not H, and there is 1 R b Or there are 2 R b .

[0182] In a preferred embodiment of the compound of formula (II), -XR 4 For H.

[0183] In a preferred embodiment of the compound of formula (II), -XR 4 Not for H.

[0184] In a preferred embodiment of the compound of formula (II), R 6 、R 7 and R 8 At least one of them is halogen, preferably F; more preferably R 6 is F, and R 7 and R 8 For H.

[0185] In a preferred embodiment of the compound of formula (II), E is halogen, preferably F.

[0186] In a preferred embodiment of the compound of formula (II), E is -OR d , R d is C optionally substituted by halogen 1-6 Alkyl, preferably C 1-6 Alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, trifluoromethyl, difluoromethyl, trifluoroethyl, and the like.

[0187] In a preferred embodiment of the compound of formula (II), R 4 Selected from H, halogen, C 1-6 Alkyl, phenyl, C 3-6 cycloalkyl, 4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, and 5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, wherein said alkyl, aryl, cycloalkyl, heterocycloalkyl and heteroaryl are optionally substituted by 1, 2 or 3 groups independently selected from halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c and-(CR c R c ) 0-6 -N(R c )2, wherein -(CR c R c ) 0-6 -N(R c )2 optionally forms a 4-7 membered nitrogen-containing heterocyclic ring through the group connected to N, together with the atoms on the cyclic group to which it is connected and the adjacent atoms.

[0188] In a preferred embodiment of the compound of formula (II), R 4 is selected from halogen, preferably F or Cl.

[0189] In a preferred embodiment of the compound of formula (II), R 4 Selected from C 1-6 Alkyl, such as methyl, ethyl, propyl, isopropyl, etc., optionally substituted by 1, 2 or 3 groups independently selected from the group consisting of halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c and-(CR c R c ) 0-6 -N(R c )2.

[0190] In a preferred embodiment of the compound of formula (II), R 4 is selected from phenyl, optionally substituted by halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR cand-(CR c R c ) 0-6 -N(R c )2 replaced.

[0191] In a preferred embodiment of the compound of formula (II), R 4 Selected from C 3-6 Cycloalkyl, optionally substituted by 1, 2 or 3 groups independently selected from the group consisting of halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c and-(CR c R c ) 0-6 -N(R c )2.

[0192] In a preferred embodiment of the compound of formula (II), R 4 is selected from 4-6 membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, optionally substituted by 1, 2 or 3 groups independently selected from halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c and-(CR c R c ) 0-6 -N(R c )2.

[0193] In a preferred embodiment of the compound of formula (II), R 4 is selected from 5-6 membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, optionally substituted by 1, 2 or 3 groups independently selected from halogen, C 1-6 Alkyl, -(CR c R c ) 0-6 -OR c and-(CR c R c ) 0-6 -N(R c )2.

[0194] In each of the above embodiments, wherein the -(CR c R c ) 0-6 - N(R c)2 optionally forms a 4-7 membered nitrogen-containing heterocyclic ring through the group connected to N, together with the atoms on the cyclic group to which it is connected and the adjacent atoms.

[0195] In a preferred embodiment of the compound of formula (II), R c independently selected at each occurrence from H or C 1-6 alkyl.

[0196] The present invention also provides an embodiment of the compound of formula (II) wherein R 1 、R 2 、R 3 、R 4 、R 6 、R 7 、R 8 , A, E, R b Each has the meaning given above for the compounds of formula (I) in one of the embodiments, preferred, more preferred or most preferred embodiments.

[0197] It should be noted that the compounds of the present invention encompass the above independent embodiments or specific embodiments, and also encompass embodiments consisting of any combination or sub-combination of the above embodiments or specific embodiments, and also encompass embodiments consisting of any combination of any preferred or exemplified embodiments.

[0198] Specific embodiments of the compounds of the present invention include the following specific compounds or their isomers, pharmaceutically acceptable salts or solvates,

[0199]

[0200]

[0201]

[0202] The compounds defined herein above and various embodiments thereof are inhibitors of Ras mutations, particularly KRas mutations, including mutations at codons G12, G13, and Q61, such as G12C mutations, G12D mutations, and G13D mutations. The compounds of the present invention, particularly the compounds specifically exemplified herein, have been shown to have proliferation inhibitory activity against cells with Ras mutations, particularly KRas G12C mutations, in cell assays, as shown in the Activity Examples section below. Therefore, the compounds of the present invention are useful for treating or preventing diseases mediated by Ras mutations, preferably KRas mutations, and most preferably KRas G12C mutations, such as diseases or conditions that can be treated by inhibiting Ras mutations, preferably KRas mutations, and most preferably KRas G12C mutations, or diseases or conditions in which Ras mutations, preferably KRas mutations, and most preferably KRas G12C mutations play a role or are implicated in their activity, particularly for treating or preventing tumors or cancers by inhibiting Ras mutations, preferably KRas mutations, and most preferably KRas G12C mutations.

[0203] In addition to exhibiting KRas G12C mutation inhibitory activity, some of the compounds of the present invention also exhibit inhibitory activity against KRas G12D mutation, and other compounds of the present invention exhibit inhibitory activity against KRas G13D mutation.

[0204] In addition to exhibiting Ras mutation, preferably KRas mutation inhibitory activity, the compounds defined herein and their various specific embodiments, especially the example compounds, have improved structural patterns and, compared to existing KRas mutant protein inhibitors in the prior art, retain comparable, enhanced, or even significantly enhanced KRas mutant protein and related cancer cell proliferation inhibitory activity; have different biological activity spectra and can be used for new indications; have improved metabolic stability, resulting in better pharmacokinetic properties; and have improved physicochemical properties, resulting in good drugability, such as easier absorption in the body.

[0205] Based on the above, the present invention also provides technical solutions in the following aspects.

[0206] In one aspect, the present invention provides a compound of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof for use as a medicament.

[0207] In another aspect, the present invention provides a compound of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof for use in treating and / or preventing a disease mediated by Ras mutation, preferably KRas mutation.

[0208] Pharmaceutical compositions and their administration

[0209] In another aspect, the present invention provides a pharmaceutical composition comprising a compound of formula (I) or formula (II) as defined above, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier, diluent, or excipient. The pharmaceutical composition of the present invention can be used to treat or prevent diseases mediated by Ras mutations, particularly KRas mutations, such as diseases mediated by KRas G12C, KRas G12D, or KRas G13D mutations, such as tumors or cancers.

[0210] The pharmaceutical composition of the present invention can be formulated by techniques known to those skilled in the art, such as those disclosed in Remington's Pharmaceutical Sciences 20th edition.

[0211] The administration and use of the pharmaceutical composition of the present invention are in accordance with good medical practice. Factors to be considered in this context include the specific disorder being treated, the specific mammal being treated, the clinical condition of the individual patient, the cause of the disorder, the position of the drug delivery, the method of administration, the arrangement of administration, and other factors well known to physician practitioners. The optimal dose level and the frequency of administration of the pharmaceutical composition of the present invention will be determined by clinical trials required by the pharmaceutical field. Typically, for example, the daily dose range for oral administration is between about 0.001 mg and about 100 mg per kg patient weight, often 0.01 mg to about 50 mg per kg body weight, for example 0.1 to 10 mg per kg body weight, preferably about 0.01 to about 35 mg per kg body weight, taken in single dose or divided dose. For a 70 kg human subject, a suitable dosage range is about 0.07 to about 7000 mg / day, preferably about 0.7 to about 2500 mg / day. It should be understood that it may be necessary to use dosages exceeding these limits in some cases.

[0212] The compositions of the present invention can be administered in any suitable manner, including oral, topical (including buccal and sublingual), rectal, vaginal, transdermal, parenteral, subcutaneous, intraperitoneal, intrapulmonary, intradermal, intrathecal, inhalation, epidural, and intranasal, and, if desired for local treatment, intralesional administration can also be employed. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. In some embodiments, oral administration is employed.

[0213] The compositions of the present invention can be administered in any convenient administration form, such as tablets, powders, capsules, lozenges, granules, solutions, dispersions, suspensions, syrups, sprays, suppositories, gels, emulsions, patches, and the like. The compositions may contain conventional components of pharmaceutical preparations, such as diluents (e.g., glucose, lactose, or mannitol), carriers, pH regulators, buffers, sweeteners, fillers, stabilizers, surfactants, wetting agents, lubricants, emulsifiers, suspending agents, preservatives, antioxidants, opacifiers, glidants, processing aids, colorants, flavorings, flavorings, other known additives, and other active agents. Suitable carriers and excipients are well known to those skilled in the art and are described in detail in, for example, Ansel, Howard C., et al., Ansel's Pharmaceutical Dosage Forms and Drug Delivery Systems. Philadelphia: Lippincott, Williams & Wilkins, 2004.

[0214] Treatment methods and uses

[0215] As described above, the compounds of formula (I) or formula (II) of the present invention and the compounds of various specific embodiments thereof, especially the compounds specifically prepared and characterized in the examples, show inhibitory effects on Ras mutations, especially KRas mutations, such as KRasG12C, KRasG12D or KRasG13D mutations.

[0216] Therefore, on the other hand, the present invention provides a method for inhibiting Ras mutation, especially KRas mutation, preferably KRasG12C, KRas G12D or KRas G13D mutation, most preferably KRas G12C mutation in a cell, comprising contacting the cell with a compound of formula (I) or formula (II) of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof to inhibit the activity of Ras mutation, especially KRas mutation, preferably KRas G12C, KRas G12D or KRas G13D mutation, most preferably KRasG12C mutation in the cell.

[0217] Based on the same properties, the present invention also provides a method for inhibiting abnormal cell growth in a mammal, comprising administering to the mammal a therapeutically effective amount of a compound of formula (I) or (II) of the present invention, its isomers, or pharmaceutically acceptable salts or solvates thereof, or a pharmaceutical composition comprising a compound of formula (I) or (II) of the present invention, its isomers, or pharmaceutically acceptable salts or solvates thereof.

[0218] On the other hand, the present invention provides a method for treating and / or preventing diseases mediated by Ras mutations, particularly KRas mutations, preferably KRasG12C, KRasG12D or KRasG13D mutations, most preferably KRasG12C mutations, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of formula (I) or formula (II) of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition comprising a compound of formula (I) or formula (II) of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof.

[0219] On the other hand, the present invention provides the use of a compound of formula (I) or (II) of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition comprising a compound of formula (I) or (II) of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof, for inhibiting Ras mutations in cells, in particular KRas mutations, preferably KRas G12C, KRas G12D or KRas G13D mutations, most preferably KRas G12C mutations, or for inhibiting abnormal cell growth in mammals, or for treating and / or preventing diseases mediated by Ras mutations, in particular KRas mutations, preferably KRas G12C, KRasG12D or KRas G13D, most preferably KRas G12C mutations.

[0220] On the other hand, the present invention provides the use of a compound of formula (I) or (II) of the present invention, its isomers or pharmaceutically acceptable salts or solvates thereof, or a pharmaceutical composition comprising a compound of formula (I) or (II) of the present invention, its isomers or pharmaceutically acceptable salts or solvates thereof, in the preparation of a medicament for treating and / or preventing diseases mediated by Ras mutations, especially KRas mutations, preferably KRas G12C, KRas G12D or KRas G13D mutations, most preferably KRas G12C mutations.

[0221] For the various methods and application solutions provided by the present invention, the abnormal cell growth or the disease mediated by Ras mutation, especially KRas mutation, preferably KRas G12C, KRas G12D or KRas G13D, most preferably KRas G12C mutation, especially refers to cancer or tumor. Exemplary such cancers or tumors include, but are not limited to, lung cancer, lung adenocarcinoma, bone cancer, pancreatic cancer, skin cancer, head and neck cancer, cutaneous or intraocular melanoma, uterine cancer, ovarian cancer, rectal cancer, cancer of the anal region, stomach cancer, colon cancer, breast cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, esophageal cancer, small intestine cancer, cancer of the endocrine system, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal cell carcinoma, renal pelvis cancer, central nervous system tumor (CNS), primary CNS lymphoma, spinal tumor, brain stem glioma, or pituitary adenoma.

[0222] For the various methods and use technical solutions provided by the present invention, the abnormal cell growth or the disease mediated by Ras mutation, especially KRas mutation, preferably KRas G12C, KRas G12D or KRas G13D is preferably selected from lung adenocarcinoma, lung cancer, colon cancer, rectal cancer, pancreatic cancer, bile duct cancer, endometrial cancer, ovarian cancer, and leukemia; most preferably selected from lung adenocarcinoma, colon cancer, rectal cancer, pancreatic cancer, and bile duct cancer.

[0223] Therefore, in preferred embodiments of this aspect, the present invention provides the above-mentioned methods and uses for treating or preventing cancer or tumors by inhibiting the KRas-G12C mutation. In further preferred embodiments, the present invention provides the above-mentioned methods and uses for treating or preventing lung adenocarcinoma, colon cancer, rectal cancer, pancreatic cancer, and bile duct cancer by inhibiting the KRas-G12C mutation.

[0224] Drug combinations

[0225] The compounds of the present invention may be administered as the sole active ingredient or in combination with another drug or therapy.

[0226] Therefore, in another aspect, the present invention provides a pharmaceutical combination comprising a compound of formula (I) or formula (II), an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof, and another active agent, or consisting of both. The pharmaceutical combination is used to inhibit abnormal cell growth in mammals, or to treat and / or prevent diseases mediated by Ras mutations, preferably KRas mutations.

[0227] The other active agent may be one or more additional compounds of the present invention, or may be a second or additional (e.g., a third) compound that is compatible with the compounds of the present invention, i.e., does not adversely affect each other, or has complementary activities. For example, these active agents may be compounds that are known to regulate other biological activity pathways, or may be compounds that regulate different components in the biological activity pathways involved in the compounds of the present invention, or even compounds that overlap with the biological targets of the compounds of the present invention.

[0228] In a specific embodiment, other active agents that can be used in combination with the compounds of the present invention include, but are not limited to, chemotherapeutic agents, therapeutic antibodies, and radiotherapy, such as alkylating agents, antimetabolites, cell cycle inhibitors, mitotic inhibitors, topoisomerase inhibitors, antihormonal drugs, angiogenesis inhibitors, and cytotoxic agents.

[0229] Other active agents used in combination with the present invention can be administered simultaneously, separately or sequentially with the compounds of the present invention by the same or different routes of administration. The other active agents can be co-administered with the compounds of the present invention in a single pharmaceutical composition, or administered separately in different discrete units from the compounds of the present invention, such as a combination product, preferably in the form of a kit, which can be administered simultaneously or sequentially when administered separately, and the sequential administration can be close or distant in time. They can be prepared and / or formulated by the same or different manufacturers. Moreover, the compounds of the present invention and the other active agents can be (i) administered before the combination product is sent to the physician (e.g., in the case of a kit comprising the compounds of the present invention and another drug); (ii) administered by the physician himself (or under the guidance of a physician) before administration; (iii) administered by the patient himself, e.g., during the sequential administration of the compounds of the present invention and the other active agents, in a combination therapy.

[0230] The compounds of the present invention may also be combined with anti-tumor therapies including, but not limited to, surgery, radiation therapy, transplantation (eg, stem cell transplantation, bone marrow transplantation), tumor immunotherapy, chemotherapy, and the like.

[0231] Therefore, in another aspect, the present invention also provides a kit comprising two or more separate pharmaceutical compositions, at least one of which comprises a compound of formula (I) or formula (II) of the present invention, an isomer thereof, or a pharmaceutically acceptable salt or solvate thereof, and a device for separately containing the compositions, such as a container, a sub-bottle, or a separate foil package, for example, a blister package for packaging tablets, capsules, etc. The kit of the present invention is particularly suitable for administering different dosage forms, such as an oral dosage form and a parenteral dosage form, or for administering different compositions at different dosage intervals.

[0232] For the technical solutions of the pharmaceutical composition, drug combination or drug kit of the present invention, the abnormal cell growth or the disease mediated by Ras mutation, especially KRas mutation, preferably KRas G12C, KRas G12D or KRas G13D, most preferably KRas G12C mutation is as defined above for the method and use of the present invention.

[0233] For the above-mentioned compounds, pharmaceutical compositions, methods, uses, pharmaceutical combinations and kits of the present invention, the compounds of formula (II) described above, their isomers or pharmaceutically acceptable salts or solvates thereof are preferred, and the compounds defined in the specific embodiments of formula (II) and the specific compounds listed above, i.e., compounds 1-25, are more preferred.

[0234] When dosages of a drug or a pharmaceutically acceptable salt thereof are described herein, it is understood that the dosage is based on the weight of the free base and does not include any hydrate or solvate thereof unless the specification indicates that the dosage is based on the weight of the salt, hydrate or solvate. BRIEF DESCRIPTION OF THE DRAWINGS

[0235] Attachment Figure 1 The anti-tumor activity of the compound of Example 9 in the human non-small cell lung cancer NCI-H358 xenograft mouse model and its effect on body weight are shown.

[0236] Attachment Figure 2 The antitumor activity and effect on body weight of the compound of Example 9 in a BALB / c nude mouse model of subcutaneous xenograft tumors of human pancreatic cancer Mia PaCa-2 cells are shown.

[0237] Preparation method of the compound of the present invention

[0238] In another aspect, the present invention also provides a method for preparing the compound defined in the present invention.

[0239] The compounds of formula (I) or formula (II) of the present invention, their isomers or their pharmaceutically acceptable salts or solvates can be prepared by a variety of methods, including the methods given below, the methods given in the examples or methods analogous thereto. The following illustrates a general synthetic scheme for synthesizing the compounds of the present invention.

[0240] For each reaction step of each general synthesis scheme, appropriate reaction conditions are known to those skilled in the art or can be routinely determined. The process steps for synthesizing the compounds of the invention can be carried out under reaction conditions known per se (including those specifically mentioned), in the absence or generally in the presence of a solvent or diluent (including, for example, a solvent or diluent that is inert to the reagents used and in which the reagents are soluble), in the absence or presence of a catalyst, a condensing agent or a neutralizing agent (for example, an ion exchanger, such as a cation exchanger, for example H + The reaction is carried out at reduced, normal or elevated temperature (e.g., from about -100°C to about 190°C, including, for example, from about -78°C to about 150°C, such as from about 0°C to about 125°C, room temperature, -20 to 40°C or reflux temperature), under atmospheric pressure or in a closed vessel, under pressure when appropriate, and / or under an inert atmosphere, such as an argon or nitrogen atmosphere, depending on the nature of the reaction and / or the reactants.

[0241] The above reaction is usually carried out at a temperature between room temperature and the boiling temperature of the solvent used, depending on the reactivity of the compounds used.

[0242] The starting materials and reagents used in preparing these compounds are generally commercially available or can be prepared by the methods described below, methods analogous to the methods given below, or methods known in the art.

[0243] Unless otherwise indicated in the description of the process, suitable solvents for any particular reaction include those specifically mentioned, or for example, water; esters, such as lower alkyl esters of lower alkanoic acids, such as ethyl acetate; ethers, such as aliphatic ethers, such as diethyl ether, or cyclic ethers, such as tetrahydrofuran or dioxane; liquid aromatic hydrocarbons, such as benzene or toluene; alcohols, such as methanol, ethanol, or 1- or 2-propanol; nitriles, such as acetonitrile; halogenated hydrocarbons, such as dichloromethane or chloroform; amides, such as dimethylformamide, N-methylpyrrolidin-2-one, or dimethylacetamide; bases, such as heterocyclic nitrogen bases, such as pyridine or triethylamine; carboxylic anhydrides, such as lower alkanoic acid anhydrides, such as acetic anhydride; cyclic, linear, or branched hydrocarbons, such as cyclohexane, hexane, or isopentane; or mixtures of these solvents, such as aqueous solutions. Such solvent mixtures can also be used for work-up, such as by chromatography or partitioning.

[0244] If necessary, the starting materials and intermediates in the synthetic reaction schemes can be separated and purified using conventional techniques, including but not limited to filtration, distillation, crystallization, chromatography, and the like. If the intermediates and final products are obtained in solid form, purification can also be performed by recrystallization or aging. The materials can be characterized using conventional methods, including physical constants and spectral data.

[0245] The reaction mixture is worked up in a customary manner, for example by mixing with water, separating the phases and, if appropriate, purifying the crude product by chromatography.

[0246] Those skilled in the art will recognize whether a stereocenter is present in a compound of formula (I) or (II). At all stages of the reaction, the resulting mixture of isomers can be separated into individual isomers, such as diastereomers or enantiomers, or into any desired mixture of isomers, such as racemates or mixtures of diastereomers, see, for example, E.L. Eliel, S.H. Wilen and L.N. Mander, "Stereochemistry of Organic Compounds" (Wiley-Interscience, 1994).

[0247] In some specific cases, it may be necessary to use appropriate blocking groups to protect specific reactive groups to avoid side reactions with other reactive groups, and the group may be present in formula (I) or formula (II) compounds and may compete or interfere with reactions. As an example only, if one or more groups in formula (I) or formula (II) compounds are or comprise groups C (O) OH, NH or OH and the group has similar or even stronger reactivity than the desired reaction position, it is advantageous to protect these groups before the desired reaction occurs. In these cases, it may be necessary to carry out additional deprotection steps to remove these blocking groups after the desired reaction is complete. Suitable blocking groups and the method for adopting such suitable blocking groups to protect and deprotect different substituents are well known to those skilled in the art; Examples thereof can be found in T.Greene and P.Wuts, Protective Groups in Organic Synthesis (3rd edition), John Wiley & Sons, NY (1999).

[0248] The present invention also relates to the following preparation methods: a compound which can be obtained as an intermediate in any step of the individual preparation methods and processes described below is used as a starting material and the remaining process steps are carried out, or a starting material is formed in situ under the reaction conditions or used in the form of a derivative, for example in a protected form or salt form, or a compound obtainable according to the process of the present invention is generated under the process conditions and is further processed in situ.

[0249] Synthesis Scheme I:

[0250] The compounds of the present invention can be prepared according to the following illustrative schemes or prepared analogously, wherein the variables are as defined above unless otherwise stated.

[0251]

[0252] The present invention prepares the compound of formula (I) or formula (II) through synthetic scheme I. In step A, compound 2 is obtained by chlorination reaction of an aromatic compound. Then, compound 3 is obtained through methyl esterification reaction in step B, and compound 4 is obtained through amino acid condensation in step C. Compound 4 is cyclized under alkaline conditions to obtain compound 5, and then intermediate Int A is obtained through chlorination reaction. The key intermediate Int A is reacted with piperazine or a piperazine derivative with a single protective group under alkaline conditions to obtain intermediate Int B. The latter is reacted through aromatic nucleophilic substitution reaction (when X=O) or metal-catalyzed coupling reaction (when X=directly connected) to obtain compound 6. In step H, compound 6 is introduced into R with a protective group through catalytic coupling reaction. 5 Compound 7 is deprotected to give compound 8, which is then acylated with an acyl chloride or carboxylic acid to give the compound of formula I shown.

[0253] The typical reaction conditions and reagents used for the chlorination, esterification, condensation, cyclization, nucleophilic substitution, catalytic coupling and acylation reactions involved in Synthesis Scheme 1 are well known in the art and fall within the routine experience of those skilled in the art. Alternatively, they can be determined by those skilled in the art by making appropriate changes based on the typical conditions of such reactions in the art, the characteristics of the raw materials used, and the target products.

[0254] Synthesis Scheme II:

[0255] Among them -XR 4 Compounds of the present invention where H is H can be synthesized according to the following illustrative general schemes.

[0256]

[0257] In Synthesis Scheme II, intermediate Int B is dechlorinated by catalytic reduction to obtain intermediate Int C, which is then subjected to the same reaction steps H, I, and J as in Scheme I to obtain -XR 4 The compound of the present invention is H.

[0258] Synthesis Scheme III:

[0259] Compounds of the present invention wherein A is N can be synthesized according to the following illustrative general schemes.

[0260]

[0261] In this synthesis scheme, compound 11 undergoes iodination to give compound 12, which then undergoes a metal-catalyzed carbonyl insertion to give compound 13. Starting from compound 13, the same steps C to J as in Scheme I are followed to give the compound of the present invention wherein A is N.

[0262] Synthesis Scheme IV:

[0263] Where A is N and -XR 4 Compounds of the present invention where H is H can be synthesized according to the following illustrative general schemes.

[0264]

[0265] In this synthesis scheme, starting from the intermediate IntE, the same method as in Scheme II is used to synthesize wherein A is N and -XR 4 The compound of the present invention is H.

[0266] In the above synthesis schemes II, III and IV, Represents a piperazine ring, wherein the typical reaction conditions and reagents used in each reaction involved are well known in the art and fall within the routine experience of those skilled in the art, or can be determined by those skilled in the art by making appropriate changes based on the typical conditions of such reactions in the art, based on the characteristics of the raw materials used and the target product.

[0267] Synthesis Example

[0268] The present invention will be further described below with reference to the following examples. It should be noted that the following examples are illustrative only and should not be considered as limiting the scope of protection of the present invention.

[0269] In describing the embodiments and the specific examples that follow, the following abbreviations are used herein:

[0270] Boc (tert-butoxycarbonyl); n-BuLi (n-butyllithium); t-BuOK (potassium tert-butoxide); CDCl3 (deuterated chloroform); DCM (dichloromethane); DIEA (N,N-diisopropylethylamine); DME (ethylene glycol dimethyl ether); DMF (N,N-dimethylformamide); DMSO (dimethyl sulfoxide); DMSO-d6 (hexadeuterated dimethyl sulfoxide); EA (ethyl acetate); EDTA-K2 (ethylenediaminetetraacetic acid dipotassium salt); EtOH (ethanol); FCC (flash column chromatography); g (gram); h (hour); HATU (2-(7-azabenzotriazole)-N,N,N'N'-tetramethyluronium hexafluorophosphate); HCl (hydrogen chloride); HLM (human liver microsomes); H2O (water); H2SO4 (sulfuric acid); IV (intravenous administration); K2CO3 (potassium carbonate); KOH (potassium hydroxide); LCMS (liquid chromatography-mass spectrometry); LC-MS / MS (liquid chromatography-mass spectrometry-mass spectrometry- Mass spectrometry); MeCN (acetonitrile); MeOH (methanol); Methanol-d4 (tetradeuterated methanol); mg (milligram); MHz (megahertz); min (minute); mL (milliliter); mmol (millimole); m / z (mass-to-charge ratio); N2 (nitrogen); NaCl (sodium chloride); NaH (sodium hydride); NaHCO3 (sodium bicarbonate); NaOH (sodium hydroxide); Na2SO3 (sodium sulfite); Na2SO4 (sodium sulfate); NCCH2CO2H (2- Cyanoacetic acid); NCS (chlorosuccinimide); NH4Cl (ammonium chloride); NIS (iodosuccinimide); NMI (N-methylimidazole); NMP (N-methylpyrrolidone); NMR (nuclear magnetic resonance); Pd / C (palladium on carbon); Pd2(dba)3 (tris(dibenzylideneacetone)dipalladium); Pd(dppf)Cl2 (1,1′-bis(diphenylphosphinoferrocenepalladium dichloride); P d(PPh3)4 (tetrakistriphenylphosphine palladium); PE (petroleum ether); PEG (polyethylene glycol); PO (oral administration); POCl3 (phosphorus oxychloride); rt (room temperature); SiO2 (silica gel); TCFH (N,N,N′,N′-tetramethylchloroformamidine hexafluorophosphate); TEA (triethylamine); TFA (trifluoroacetic acid); THF (tetrahydrofuran); TLC (thin layer chromatography); TsOH (p-toluenesulfonic acid); TsOH·H2O (p-toluenesulfonic acid monohydrate); μL (microliter); μM (micromolar concentration); μmol (micromolar); v / v (liquid volume ratio); w / w (weight ratio); Xantphos (4,5-bis(diphenylphosphino-9,9-dimethylxanthene).

[0271] In the following examples, the names and structures of the synthesized target compounds are given. Any discrepancies between the names and structures are unintentional; in this case, the structure is decisive.

[0272] In the following examples, experimental procedures, where specific conditions are not specified, generally follow conventional conditions for such reactions or those recommended by the manufacturer. Unless otherwise noted, percentages and parts are by weight. Liquid ratios are by volume unless otherwise noted.

[0273] Unless otherwise specified, the experimental materials and reagents used in the following examples can be obtained from commercial channels, prepared according to existing methods, or prepared according to methods similar to those disclosed in this application.

[0274] In the following examples, 1 H-NMR spectra were recorded using a Bruker (400 MHz), and chemical shifts are expressed as δ (ppm) relative to the deuterated solvent peak (CDCl3: δ = 7.26 ppm; CD3OD: δ = 3.31 ppm; DMSO-d6: δ = 2.50 ppm); mass spectra were recorded using an Aglient 1100 liquid chromatograph + Aglient G6100 mass spectrometer LCMS.

[0275] Synthesis of intermediate a

[0276]

[0277] (2-((tert-Butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid

[0278]

[0279] Step A: N-((2-bromo-5-fluorophenyl)aminomethanesulfonyl)benzamide

[0280] In a round-bottom flask equipped with a magnetic rod, dissolve benzoyl isothiocyanate (28 g, 210 mmol) in THF (300 mL). After cooling in an ice bath, add a solution of 2-bromo-5-fluoroaniline (40 g, 210 mmol) in THF (100 mL) dropwise with stirring. After the addition is complete, continue stirring and react for 1 hour. LCMS confirms the reaction is complete. Concentrate under reduced pressure to obtain the crude product, which is then used directly in the next step. LCMS (m / z): 353.1 (M+H).

[0281] Step B: 1-(2-Bromo-5-fluorophenyl)thiourea

[0282] Under stirring at room temperature, a solution of NaOH (16.8 g, 420 mmol) in water (150 mL) was added to a solution of N-((2-bromo-5-fluorophenyl)aminomethanesulfonyl)benzamide (crude product from Step A) in THF (300 mL). The mixture was heated to 80°C for 16 h. After concentration under reduced pressure, the mixture was diluted with EA (200 mL) and washed with water (150 mL). The aqueous and organic phases were separated, and the aqueous phase was extracted twice with EA (100 mL x 2). The organic phases were combined, washed with saturated sodium chloride solution, and dried over anhydrous sodium sulfate. The filtrate was filtered and concentrated under reduced pressure to obtain the crude product. 200 mL of petroleum ether was added to the slurry and filtered to afford 1-(2-bromo-5-fluorophenyl)thiourea (40 g, 77% yield) as a white solid. LCMS (m / z): 249.0, 251.0 (M+H).

[0283] Step C: 4-Bromo-7-fluorobenzo[d]thiazol-2-amine hydrobromide

[0284] Under stirring, a solution of liquid bromine (23 g, 141 mmol) in chloroform (70 mL) was slowly added dropwise to an ice-cooled solution of 1-(2-bromo-5-fluorophenyl)thiourea (35 g, 141 mmol) in chloroform (300 mL). After the addition was complete, the ice bath was removed and the system was heated to 70°C for 48 h. After completion of the reaction, the mixture was concentrated under reduced pressure, slurried with EA (100 mL), and filtered to afford 4-bromo-7-fluorobenzo[d]thiazol-2-amine hydrobromide (30.5 g, 66% yield) as a white solid. LCMS (m / z): 247.0, 249.0 (M+H).

[0285] Step D: tert-Butyl (4-bromo-7-fluorobenzo[d]thiazol-2-yl)carbamate

[0286] At room temperature, a solution of 4-bromo-7-fluorobenzo[d]thiazol-2-amine hydrobromide (30 g, 93 mmol) and 4-dimethylaminopyridine (14 g, 112 mmol) in DCM (300 mL) was slowly added with di-tert-butyl dicarbonate (61 g, 280 mmol) in DCM (60 mL). The reaction was allowed to react at room temperature for 3 h, and the reaction was complete by LCMS. The reaction solution was concentrated under reduced pressure, diluted with EA (200 mL), washed with water (150 mL), and the aqueous and organic phases were separated. The aqueous phase was then extracted twice with EA (200 mL x 2). The organic phases were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and slurried with petroleum ether and ethyl acetate (20:1). The mixture was filtered to obtain tert-butyl (4-bromo-7-fluorobenzo[d]thiazol-2-yl)carbamate (18 g, 56% yield) as a white solid. 1H NMR (400MHz, Chloroform-d) δ 9.29 (s, 1H), 7.54 (dd, J=8.6, 4.8Hz, 1H), 6.94-6.81 (m, 1H), 1.52 (s, 9H). LCMS (m / z): 291.0, 293.0 (M+H).

[0287] Step E: (2-((tert-Butyloxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid

[0288] Under nitrogen, NaH (60% w / w, 3 g, 75.6 mmol) was added to an ice-cooled solution of tert-butyl (4-bromo-7-fluorobenzo[d]thiazol-2-yl)carbamate (17.5 g, 50.4 mmol) in THF (200 mL). Stirring was continued at this temperature for 30 minutes, and the mixture was transferred to a -68°C dry ice / ethanol bath. Then, n-butyllithium (30.2 mL, 75.6 mmol, 2.5 M / THF) was slowly added. After stirring for 10 minutes, trimethyl borate (156 g, 151 mmol) was added, and the reaction was stirred for another 30 minutes. The mixture was quenched with saturated aqueous ammonium chloride (50 mL), diluted with EA (200 mL), and washed with water (150 mL). The aqueous and organic phases were separated, and the aqueous phase was extracted twice with EA (100 mL x 2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to yield the crude product. The obtained crude product was added to petroleum ether and slurried, and filtered to obtain a white solid (2-((tert-butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid (10.7 g, yield 68%). 1 H NMR (400MHz, DMSO-d6) δ 12.25 (s, 1H), 8.50 (s, 2H), 7.84 (dd, J=8.0, 6.4Hz, 1H), 7.22-7.13 (m, 1H), 1.54 (s, 9H). LCMS (m / z): 313.2 (M+H).

[0289] Synthesis of intermediate IntA

[0290]

[0291] 7-Bromo-2,4,6-trichloro-8-fluoroquinoline-3-carbonitrile

[0292]

[0293] Step A: 2-Amino-4-bromo-5-chloro-3-fluorobenzoic acid

[0294] NCS (13.7 g, 102.6 mmol) was added portionwise to 2-amino-4-bromo-3-fluorobenzoic acid (20 g, 85.5 mmol) dissolved in DMF (140 mL) at room temperature. The reaction solution was then heated to 75°C and stirred at this temperature for 20 h. After cooling to room temperature, the reaction solution was poured into 700 mL of ice-water mixture to precipitate. The solid was collected by filtration and washed with water (500 mL) and petroleum ether (150 mL), respectively. The resulting product was dried under vacuum to afford 2-amino-4-bromo-5-chloro-3-fluorobenzoic acid (20 g, 87% yield) as a light yellow solid. LCMS (m / z): 268.0 (M+H).

[0295] Step B: Methyl 2-amino-4-bromo-5-chloro-3-fluorobenzoate

[0296] Disperse 2-amino-4-bromo-5-chloro-3-fluorobenzoic acid (20 g, 74.5 mmol) in methanol (150 mL). Cool in an ice-water bath and, with stirring, add thionyl chloride (50 mL) dropwise to the reaction mixture. After the addition is complete, incubate the reaction mixture at 65°C for 20 hours. After completion of the reaction, monitor the reaction by TLC. Concentrate the solvent to dryness to obtain the desired product, methyl 2-amino-4-bromo-5-chloro-3-fluorobenzoate (15.8 g, 75% yield). LCMS (m / z): 282.0 (M+H).

[0297] Step C: Methyl 4-bromo-5-chloro-2-(2-cyanoacetamido)-3-fluorobenzoate

[0298] At room temperature, methyl 2-amino-4-bromo-5-chloro-3-fluorobenzoate (15.8 g, 55.9 mmol), TCFH (18.8 g, 67.1 mmol), and cyanoacetic acid (5.2 g, 61.3 mmol) were dissolved in acetonitrile (100 mL). N-methylimidazole (6.7 mL, 83.9 mmol) was added with stirring. The reaction mixture was heated to 60°C and stirred for 20 h. TLC confirmed the completion of the reaction. The reaction solution was cooled to room temperature and poured into water (500 mL), whereupon a white precipitate formed. The solid was collected by filtration and washed with water (500 mL) and ethyl acetate / petroleum ether (1:4, 150 mL), respectively. After vacuum drying, methyl 4-bromo-5-chloro-2-(2-cyanoacetamido)-3-fluorobenzoate (15 g, 76% yield) was obtained as a white solid. LCMS (m / z): 349.0 (M+H).

[0299] Step D: 7-Bromo-6-chloro-8-fluoro-2,4-dihydroxyquinoline-3-carbonitrile

[0300] The compound methyl 4-bromo-5-chloro-2-(2-cyanoacetamido)-3-fluorobenzoate (6 g, 17.2 mmol) was dispersed in THF (120 mL). Potassium tert-butoxide (3.7 g, 32.6 mmol) was added portionwise with stirring in an ice bath. After the addition was complete, the reaction mixture was stirred in an ice bath for 1 hour. LCMS monitored the reaction completion. After removing the solvent by distillation under reduced pressure, water (200 mL) was added to the system. The pH of the mixture was adjusted to 7 with 1 N hydrochloric acid, and the solid was filtered and collected. The resulting solid was washed with water (100 mL) and petroleum ether (150 mL) respectively, and dried in vacuo to obtain 7-bromo-6-chloro-8-fluoro-2,4-dihydroxyquinoline-3-carbonitrile as a white solid (5.4 g, 99% yield). LCMS (m / z): 317.0 (M+H).

[0301] Step E: 7-Bromo-2,4,6-trichloro-8-fluoroquinoline-3-carbonitrile

[0302] Under stirring at room temperature, the compound 7-bromo-6-chloro-8-fluoro-2,4-dihydroxyquinoline-3-carbonitrile (5.4 g, 17 mmol) was dispersed in phosphorus oxychloride (100 mL). The system was heated to 120°C and stirred for 24 hours. After cooling to room temperature, the mixture was concentrated in vacuo to remove most of the phosphorus oxychloride, and acetonitrile (20 mL) was added. The acetonitrile solution of the product was slowly poured into water (250 mL) to precipitate a yellow solid. The solid was filtered and collected, washed with water (200 mL) and petroleum ether (100 mL), and dried in vacuo to obtain 7-bromo-2,4,6-trichloro-8-fluoroquinoline-3-carbonitrile (5.4 g, 90% yield) as a yellow solid. LCMS (m / z): 353.0 (M+H).

[0303] Synthesis of intermediate Int B1

[0304]

[0305] (R)-tert-Butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate

[0306]

[0307] DIEA (7.3 mL, 44 mmol) and (R)-tert-butyl 2-methylpiperazine-1-carboxylate (4.4 g, 22 mmol) were added sequentially to a solution of 7-bromo-2,4,6-trichloro-8-fluoroquinoline-3-carbonitrile (Intermediate IntA, 7.8 g, 22 mmol) in THF (100 mL) with stirring at room temperature. The resulting mixture was stirred at room temperature for 1 h. After completion of the reaction, the sample was concentrated using a rotary evaporator, and the crude product was washed with a PE / THF mixture (PE / THF = 10:1, 500 mL) to afford (R)-tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (9.0 g, 79% yield) as a yellow solid. LCMS (m / z): 519.2 (M+H).

[0308] Synthesis of intermediate Int B2

[0309]

[0310] tert-Butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate

[0311] The synthesis of Intermediate Int B2 was carried out as described in Intermediate Int B1, except that tert-butyl piperazine-1-carboxylate was substituted for tert-butyl (R)-2-methylpiperazine-1-carboxylate. LCMS (m / z): 503.2 (M+H).

[0312] Synthesis of intermediate Int C1

[0313]

[0314] (R)-tert-Butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate

[0315]

[0316] To a round-bottom flask equipped with a magnetic bar was added (R)-tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate Int B1, 206 mg, 0.4 mmol), Pd(PPh3)4 (23 mg, 20 μmol), formic acid (55 mg, 1.2 mmol), and triethylamine (80.5 mg, 0.8 mmol). After nitrogen evacuation three times, DMF (4 mL) was added, and the reaction system was heated to 50°C and stirred for 6 h. After completion of the reaction, the sample was concentrated on a rotary evaporator, and the crude product was purified by FCC (SiO2, EA / PE = 0-50%) to obtain tert-butyl (R)-4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (140 mg, 73% yield). LCMS (m / z): 485.3 (M+H).

[0317] Synthesis of intermediate Int C2

[0318]

[0319] tert-Butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate

[0320] The synthesis of Intermediate IntC2 was carried out as described for Intermediate IntC1, substituting tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate B2) for tert-butyl (R)-4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate B1). LCMS (m / z): 459.2, 471.2 (M+H).

[0321] Synthesis of intermediate Int D

[0322]

[0323] 2,4,7-Trichloro-8-fluoro-1,6-naphthyridine-3-carbonitrile

[0324]

[0325] Step A: 2-Chloro-3-fluoro-5-iodopyridin-4-amine

[0326] In a round-bottom flask at room temperature, 2-chloro-3-fluoropyridin-4-amine (3 g, 20.47 mmol) and NIS (6.91 g, 30.71 mmol) were dissolved in 30 mL of acetonitrile. TsOH·H₂O (400 mg, 2 mmol) was added with stirring. The reaction solution was heated to 70°C for 16 h. After completion of the reaction, the solution was cooled to room temperature and poured into 100 mL of saturated brine. The mixture was extracted with ethyl acetate (25 mL x 3). The combined organic phases were washed with saturated aqueous Na₂SO₃ and saturated brine, dried over anhydrous Na₂SO₄, and the solvent was concentrated to yield 2-chloro-3-fluoro-5-iodopyridin-4-amine (5.4 g, 97% yield) as a pale yellow solid. LCMS (m / z): 273.0 (M+H).

[0327] Step B: Ethyl 4-amino-6-chloro-5-fluoronicotinate

[0328] 2-Chloro-3-fluoro-5-iodopyridin-4-amine (5.4 g, 19.82 mmol), Pd(dppf)Cl2 (695 mg, 0.991 mmol), and triethylamine (555 mg, 49.6 mmol) were dispersed in anhydrous ethanol and heated to 90°C under a carbon monoxide atmosphere for 40 h. After completion of the reaction, the reaction solution was filtered through celite, and the filtrate was concentrated and purified by FCC (SiO2, EA / DCM = 0-100%) to obtain ethyl 4-amino-6-chloro-5-fluoronicotinate (3.5 g, 81% yield) as a yellow solid. LCMS (m / z): 219.0 (M+H).

[0329] Step C: Ethyl 6-chloro-4-(2-cyanoacetamido)-5-fluoronicotinate

[0330] Cyanoacetic acid (778 mg, 9.15 mmol) and trifluoroacetic anhydride (1.44 g, 6.86 mmol) were dissolved in acetonitrile (5 mL) and stirred at 55°C for 2 h. The reaction mixture and ethyl 4-amino-6-chloro-5-fluoronicotinate (1.0 g, 4.57 mmol) were placed in a 20 mL microwave tube and microwave irradiated at 105°C for 2 h. After completion of the reaction, the reaction mixture was poured into 25 mL of water and ethyl acetate (15 mL) was added. The organic phase was washed with water, saturated aqueous sodium bicarbonate, and saturated brine, respectively, and dried over anhydrous sodium sulfate. The solvent was concentrated and purified by FCC (SiO2, THF / DCM = 0-40%) to yield ethyl 6-chloro-4-(2-cyanoacetamido)-5-fluoronicotinate (550 mg, 42% yield) as a pale yellow solid. LCMS (m / z): 286.0 (M+H).

[0331] Step D: 7-Chloro-8-fluoro-2,4-dihydroxy-1,6-naphthyridine-3-carbonitrile

[0332]

[0333] Ethyl 6-chloro-4-(2-cyanoacetamido)-5-fluoronicotinate (300 mg, 1.05 mmol) was dissolved in THF (10 mL) and stirred in an ice bath for 2 min. Potassium tert-butoxide (236 mg, 2.10 mmol) was then slowly added to the reaction mixture. The mixture was allowed to return to room temperature in an ice bath and stirred for 1 h. After completion of the reaction, the reaction solution was added dropwise to a stirred saturated NH4Cl solution (50 mL). The pH of the mixture was adjusted to 7 with 1N hydrochloric acid. During this time, a white flocculent precipitate formed. The solid was collected by filtration and washed with water (10 mL) and petroleum ether (10 mL). After vacuum drying, 7-chloro-8-fluoro-2,4-dihydroxy-1,6-naphthyridine-3-carbonitrile (200 mg, 79% yield) was obtained as a white solid. LCMS (m / z): 240.1 (M+H).

[0334] Step E: 2,4,7-Trichloro-8-fluoro-1,6-naphthyridine-3-carbonitrile

[0335]

[0336] 7-chloro-8-fluoro-2,4-dihydroxy-1,6-naphthyridine-3-carbonitrile (200mg, 0.83mmol) is dissolved in POCl (15mL), the reaction solution is heated to 120 ℃ and stirred overnight. TLC monitoring reaction is complete, cooled to room temperature, vacuum concentration removes most of phosphorus oxychloride, and then adds acetonitrile (1mL). The acetonitrile solution of product is poured into saturated NaHCO aqueous solution (30mL), and yellow solid is produced. Filter, collect solid, gained solid is washed with water (10mL) and petroleum ether (10mL) respectively, obtains yellow solid 2,4,7-trichloro-8-fluoro-1,6-naphthyridine-3-carbonitrile (190mg, yield 82%) after vacuum drying.

[0337] Synthesis of intermediate Int E1

[0338]

[0339] tert-Butyl 4-(2,7-dichloro-3-cyano-8-fluoro-1,6-naphthyridin-4-yl)piperazine-1-carboxylate

[0340] The synthesis of Intermediate Int E1 was carried out as described for Intermediate Int B1, substituting 2,4,7-trichloro-8-fluoro-1,6-naphthyridine-3-carbonitrile (Intermediate Int D) for 7-bromo-2,4,6-trichloro-8-fluoroquinoline-3-carbonitrile (Intermediate Int A), and substituting tert-butyl piperazine-1-carboxylate for tert-butyl (R)-2-methylpiperazine-1-carboxylate. LCMS (m / z): 426.3 (M+H).

[0341] Synthesis of intermediate Int F

[0342]

[0343] (S)-benzyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate

[0344] The synthesis of Intermediate Int F was carried out as described in Intermediate Int B1, using (S)-2-(cyanomethyl)piperazine-1-carboxylic acid benzyl ester salt instead of (R)-2-methylpiperazine-1-carboxylic acid tert-butyl ester. LCMS (m / z): 576.3, 578.3 (M+H).

[0345] Example 1

[0346]

[0347] 4-((R)-4-Acryloyl-3-methylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinoline-3-carbonitrile

[0348]

[0349] Step A: (R)-tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate

[0350] Under stirring at room temperature, NaH (60% w / w, 9.3 mg, 0.23 mmol) was added to a solution of (3R,4R)-4-methoxy-1-methylpyrrolidin-3-ol (27.8 mg, 0.21 mmol) in THF (2 mL). The resulting mixture was stirred at room temperature for an additional 10 min. (R)-tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate B1, 110 mg, 0.21 mmol) was added in one portion and stirred at room temperature for 1 h. LCMS monitored the presence of residual Intermediate B1. A solution of NaH (60% w / w, 5 mg, 0.12 mmol) and (3R,4R)-4-methoxy-1-methylpyrrolidin-3-ol (10 mg, 0.78 mmol) in THF (1 mL) was added again, pre-stirred for 5 min. The resulting mixture was stirred for an additional 30 min. LCMS monitored the reaction completion and the reaction was quenched with saturated aqueous NH4Cl (2 mL). EA (30 mL) was added for dilution, and the resulting organic phase was washed with saturated NaCl (20 mL x 3), dried over anhydrous Na2SO4, filtered, and concentrated to afford crude (R)-tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate (130 mg, crude), which was directly used in the next step. LCMS (m / z): 612.4 (M+H).

[0351] Step B: (2R)-tert-Butyl 4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate

[0352] At room temperature, a solution of (R)-tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate (170 mg, 0.28 mmol), (2-((tert-butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid (112 mg, 0.36 mmol), Pd(dppf)Cl2 (24 mg, 0.03 mmol), and K3PO4 (104 mg, 0.49 mmol) in 1,4-dioxane / H2O (v / v = 3:1, 1.6 mL) was added to a microwave reaction tube equipped with a magnetron. After purging with nitrogen for 10 minutes, the reaction tube was sealed and microwave-heated at 100°C for 1 hour. The reaction mixture was cooled to room temperature, diluted with EA (20 mL), washed with saturated aqueous NaCl (20 mL x 3), dried over anhydrous NaSO, concentrated under reduced pressure, and initially purified by FCC (SiO, MeOH / DCM = 0-20%) to give tert-butyl (2R)-4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate (150 mg, 62% purity), which was used directly in the next step. LCMS (m / z): 800.6 (M+H).

[0353] Step C: 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)-4-((R)-3-methylpiperazin-1-yl)quinoline-3-carbonitrile

[0354] With stirring at room temperature, TFA (1 mL) was added dropwise to a solution of tert-butyl (2R)-4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate (150 mg, 62% purity) in DCM (2 mL). The resulting mixture was stirred at room temperature for 30 min. LCMS monitored the reaction for completion. The solvent, TFA, was removed by concentration under reduced pressure. A small amount of DCM was added and the solution was concentrated under reduced pressure again. This process was repeated three times to remove the residual TFA. The crude product was then used directly in the next step without purification.

[0355] Step D: 4-((R)-4-Acryloyl-3-methylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinoline-3-carbonitrile

[0356] The crude 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)-4-((R)-3-methylpiperazin-1-yl)quinoline-3-carbonitrile obtained in step C was dissolved in DCM (2 mL) and cooled in an ice-water bath. DIEA (103 uL, 0.62 mmol) was added and stirred for 5 min. After that, a solution of acryloyl chloride (10 uL, 0.12 mmol) in DCM (0.1 mL) was slowly added dropwise. After the addition was complete, stirring was continued in an ice-water bath for 20 min. The reaction was completed under LCMS monitoring. H2O (5 mL) and EA (30 mL) were added to the system, the aqueous phase was separated and removed, and the organic phase was washed with saturated aqueous NaCl solution (20 mL×3), dried over anhydrous Na2SO4, and concentrated. The crude product was purified by preparative HPLC to give 4-((R)-4-acryloyl-3-methylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinoline-3-carbonitrile (15.2 mg, total yield over four steps 7.5%) as a white solid. 1 H NMR (400MHz, Methanol-d4) δ 8.12-7.99 (m, 1H), 7.33-7.22 (m, 1H), 7.07-6.97 (m, 1H), 6.87 (dd, J=16.7, 10.7Hz, 1H), 6.30 (d, J=16.8Hz, 1H), 5.83 (d, J=10.7Hz, 1H), 5.65 (s, 1H), 4.24-4.20 (m, 1H), 4.09-3.89 (m, 2H), 3.83-3.77 (m, 1H), 3.62-3.48(m, 5H), 3.43-3.38(m, 2H), 3.26-3.18(m, 2H), 3.09-3.03(m, 1H), 2.98-2.92(m, 1H), 2.60(s, 3H), 1.46(d, J=6.8Hz, 3H). 19 F NMR (376MHz, Methanol-d4) delta -114.94, -120.48. LCMS (m / z): 654.5 (M+H).

[0357] Example 2

[0358]

[0359] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinoline-3-carbonitrile

[0360] The synthesis of Example 2 was carried out as described in Example 1, using tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate B2) in Step A instead of tert-butyl (R)-4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate B1). 1 H NMR (400MHz, DMSO-d6) δ8.02-7.87 (m, 3H), 7.24 (dt, J=8.4, 5.1Hz, 1H), 7.07 (t, J=8.8Hz, 1H), 6.90 (dd, J=16.7, 10.5Hz, 1H), 6.19 (dd, J= 16.7, 2.4Hz, 1H), 5.75 (dd, J=10.4, 2.4Hz, 1H), 5.39-5.31 (m, 1H), 4.06-3.96 (m, 1H), 3.94- 3.81(m, 4H), 3.75-3.61(m, 4H), 3.32(s, 3H), 3.08-3.00(m, 1H), 2.92(dd, J=11.0, 6.2Hz, 1H), 2.73-2.64(m, 1H), 2.33(dt, J=9.3, 4.3Hz, 1H), 2.26-2.21(m, 3H). 19 F NMR (376MHz, DMSO-d6) δ -112.27 (d, J=4.4Hz), -119.59 (d, J=22.4Hz). LCMS (m / z): 640.5 (M+H).

[0361] Example 3

[0362]

[0363] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-(3-((dimethylamino)methyl)phenyl)-8-fluoroquinoline-3-carbonitrile

[0364]

[0365] Step A: tert-Butyl 4-(7-bromo-6-chloro-3-cyano-2-(3-((dimethylamino)methyl)phenyl)-8-fluoroquinolin-4-yl)piperazine-1-carboxylate

[0366] To a microwave tube was added tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate B2, 300 mg, 0.60 mmol), (3-((dimethylamino)methyl)phenyl)boronic acid (108 mg, 0.60 mmol), Pd(PPh3)4 (35 mg, 0.03 mmol), K2CO3 (166 mg, 1.2 mmol), acetonitrile (3 mL), and water (1 mL). The atmosphere was purged with nitrogen three times and then heated to 40°C in a microwave oven for 1 h. After cooling to room temperature, the reaction solution was concentrated, and the crude product was purified by FCC (SiO2, EA / PE = 0-60%) to give tert-butyl 4-(7-bromo-6-chloro-3-cyano-2-(3-((dimethylamino)methyl)phenyl)-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (260 mg, 72% yield) as a yellow solid. LCMS (m / z): 602.4 (M+H).

[0367] Step B to Step D: 4-(4-acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-(3-((dimethylamino)methyl)phenyl)-8-fluoroquinoline-3-carbonitrile

[0368] The subsequent synthesis of Example 3 was carried out as described in the synthesis of Example 1, except that tert-butyl 4-(7-bromo-6-chloro-3-cyano-2-(3-((dimethylamino)methyl)phenyl)-8-fluoroquinolin-4-yl)piperazine-1-carboxylate was used in Step B instead of tert-butyl (R)-4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate. 1 H NMR (400MHz, DMSO-d6) δ8.11-8.10(m, 1H), 7.92(s, 2H), 7.81-7.70(m, 2H), 7.53-7.46(m, 2H), 7.31-7.27(m, 1H), 7.11-7.09(m, 1H), 6.91(dd, J=16.6, 10.4Hz, 1H), 6.22-6.17(m, 1H), 5.77-5.74(m, 1H), 3.98-9.84(m, 4H), 3.83-3.70(m, 4H), 3.51-3.46(m, 2H), 2.24-2.13(m, 6H). LCMS (m / z): 645.5 (M+H).

[0369] Example 4

[0370]

[0371] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-(1-(2-(dimethylamino)ethyl)-1H-pyrazol-5-yl)-8-fluoroquinoline-3-carbonitrile

[0372] The synthesis of Example 4 was carried out as described in Example 3, using (1-(2-(dimethylamino)ethyl)-1H-pyrazol-5-yl)boronic acid instead of (3-((dimethylamino)methyl)phenyl)boronic acid in Step A. 1 H NMR (400MHz, CDCl3) δ7.96 (s, 1H), 7.63 (s, 1H), 7.40-7.30 (m, 1H), 7.01-6.93 (m, 2H), 6.68-6.59 (m, 1H), 6.39 (d, J=16.8Hz, 1H), 6.26 (s, 2H), 5.80 (d, J=10.6Hz, 1H), 4.66 (t, J=7.5Hz, 2H), 4.04-3.71 (m, 8H), 3.39 (s, 1H), 2.98 (s, 1H), 2.38 (s, 6H). 19 F NMR (376MHz, CDCl3) δ -111.07, -115.38. LCMS (m / z): 648.5 (M+H).

[0373] Example 5

[0374]

[0375] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-(3-(1-(dimethylamino)ethyl)phenyl)-8-fluoroquinoline-3-carbonitrile

[0376] The synthesis of Example 5 was carried out as described in Example 3, using (3-(1-(dimethylamino)ethyl)phenyl)boronic acid in Step A instead of (3-((dimethylamino)methyl)phenyl)boronic acid. 1H NMR (400MHz, MeOH-d4) δ8.16 (d, J=1.6Hz, 1H), 7.87 (s, 1H), 7.84-7.78 (m, 1H), 7.61-7.52 (m, 2H), 7.30 (dd, J=8.5, 5.4Hz, 1H), 7.09- 6.99 (m, 1H), 6.89 (dd, J=16.8, 10.6Hz, 1H), 6.31 (dd, J=16.7, 1.9Hz, 1H), 5.84 (dd, J=10.6, 1.9 Hz, 1H), 4.12-4.00 (m, 4H), 3.97-3.84 (m, 4H), 2.30 (s, 6H), 1.57-1.45 (m, 3H). 19 F NMR (376 MHz, MeOH-d4) δ -114.82, -118.65 (d, J=9.3Hz). LCMS (m / z): 658.5 (M+H).

[0377] Example 6

[0378]

[0379] 4-((R)-4-Acryloyl-3-methylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-(3-((dimethylamino)methyl)phenyl)-8-fluoroquinoline-3-carbonitrile

[0380] The synthesis of Example 6 was carried out as described in Example 3, using (R)-tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate Int B1) instead of tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate Int B2) in Step A. 1H NMR (400MHz, DMSO-d6) δ8.14 (d, J=3.3 Hz, 1H), 7.92 (d, J=19.9Hz, 2H), 7.82 (s, 1H), 7.77 (d, J=7.6Hz, 1H), 7.56-7.46 (m, 2H), 7.33 -7.27(m, 1H), 7.13-7.06(m, 1H), 6.94-6.84(m, 1H), 6.23-6.14(m, 1H), 5.78-5.72(m, 1H), 4.95-4.23 (m, 4H), 4.08 (t, J=12.7Hz, 1H), 4.00-3.90 (m, 1H), 3.77 (t, J=12.7Hz, 1H), 3.53 (s, 2H), 2.21 (s, 6H), 1.36-1.27 (m, 3H). LCMS (m / z): 658.5 (M+H).

[0381] Example 7

[0382]

[0383] 4-((R)-4-Acryloyl-3-methylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-(1-(2-(dimethylamino)ethyl)-1H-pyrazol-5-yl)-8-fluoroquinoline-3-carbonitrile

[0384] The synthesis of Example 6 was carried out as described in Example 3, using (R)-tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate Int B1) instead of tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate Int B2) in Step A, and (1-(2-(dimethylamino)ethyl)-1H-pyrazol-5-yl)boronic acid instead of (3-((dimethylamino)methyl)phenyl)boronic acid. 1H NMR (400MHz, DMSO-d6) δ 8.14 (s, 1H), 7.90 (d, J = 11.8Hz, 2H), 7.62 (d, J = 1.9Hz, 1H), 7.29 (dd, J = 8.6, 5.3Hz, 1H), 7.09 (t, J = 8.8Hz, 1H), 6.94- 6.83(m, 2H), 6.17(d, J=16.7Hz, 1H), 5.79-5.70(m, 1H), 4.62-4.32(m, 4H), 4.11-3.92(m, 3H), 3.81-3.66 (m, 2H), 3.55 (s, 2H), 2.10-1.83 (m, 6H), 1.34-1.20 (m, 3H). LCMS (m / z): 662.5, (M+H).

[0385] Example 8

[0386]

[0387] 4-((R)-4-Acryloyl-3-methylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile

[0388]

[0389] Step A: (2R)-tert-Butyl 4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate

[0390] To a microwave tube were added tert-butyl (R)-4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate C1, 120 mg, 0.25 mmol), (2-((tert-butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid (232 mg, 0.75 mmol), Pd(dppf)Cl2 (18.3 mg, 25 μmol), and K3PO4 (211 mg, 1.0 mmol). After nitrogen was purged three times, 1,4-dioxane / water (v / v = 3:1, 4 ml) was added. The reaction mixture was placed in a microwave reactor and heated to 90°C for 1 h. After completion of the reaction, the system was cooled to room temperature, and the reaction solvent was dried using a rotary evaporator. The crude product was purified by FCC (SiO2, EA / PE = 0-80%) to give tert-butyl (2R)-4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (92 mg, 47% yield) as a yellow solid. LCMS (m / z): 671.5 (M+H).

[0391] Step B: 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-4-((R)-3-methylpiperazin-1-yl)quinoline-3-carbonitrile

[0392] Under stirring at room temperature, TFA (2 mL) was added dropwise to a solution of tert-butyl (2R)-4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (92 mg, 0.14 mmol) in DCM (1 ml). Stirring was continued for one hour, and the reaction was complete as monitored by LCMS. The reaction solvent was dried on a rotary evaporator, a small amount of DCM was added and concentrated again, and the TFA residue was removed by repeating three times to give the crude product of 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-4-((R)-3-methylpiperazin-1-yl)quinoline-3-carbonitrile, which was used directly in the next step. LCMS (m / z): 471.3 (M+H).

[0393] Step C: 4-((R)-4-Acryloyl-3-methylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile

[0394] 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-4-((R)-3-methylpiperazin-1-yl)quinoline-3-carbonitrile (crude product from Step B) was added to a round-bottom flask equipped with a magnetic separator, followed by DCM (3 ml). The mixture was cooled to 0°C in an ice-water bath, and DIPEA (2 ml) was added, followed by the dropwise addition of a solution of acryloyl chloride (11 mg, 0.12 mmol) in DCM (1.0 mL). Stirring was continued in an ice-water bath for 30 min. After completion of the reaction, water (5 mL) was added to quench the reaction, and the mixture was extracted with DCM (10 mL x 3). The organic phases were combined, washed with saturated NaCl (10 mL x 3), and dried over anhydrous Na2SO4. The residue was concentrated on a rotary evaporator and purified by preparative HPLC to give 4-((R)-4-acryloyl-3-methylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile as a white solid (26 mg, 35% yield). 1 H NMR (400MHz, DMSO-d6) δ 8.88 (s, 1H), 8.08 (dd, J = 4.3, 1.4Hz, 1H), 7.92 (s, 2H), 7.38-7.24 (m, 1H), 7.12-7.04 (m, 1H), 6.86 (dd, J = 16.6, 10.5Hz, 1H), 6.17 (dd, J=16.6, 2.4Hz, 1H), 5.81-5.68 (m, 1H), 4.47 (d, J=125.4Hz, 3H), 3.94 (t, J=12.8 Hz, 1H), 3.87-3.64 (m, 2H), 3.56-3.41 (m, 1H), 1.49-1.19 (m, 3H). LCMS (m / z): 525.3 (M+H).

[0395] Example 9

[0396]

[0397] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile

[0398] The synthesis of Example 9 was carried out as described in Example 8, using tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate Int C2) instead of tert-butyl (R)-4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate Int C1) in Step A. 1H NMR (400MHz, Methanol-d4) δ8.78 (s, 1H), 8.10 (d, J = 1.8Hz, 1H), 7.26 (dd, J = 8.4, 5.4Hz, 1H), 7.06-6.97 (m, 1H), 6.87 (dd, J = 16.8, 10.7Hz, 1H), 6.30 (dd, J=16.8, 1.9Hz, 1H), 5.83 (dd, J=10.6, 1.9Hz, 1H), 4.05-3.95 (m, 4H), 3.90-3.78 (m, 4H). 19 F NMR (376MHz, Methanol-d4) delta -114.73, -119.40. LCMS (m / z): 511.3 (M+H).

[0399] Example 10

[0400]

[0401] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(2-methyl-1,2,3,4-tetrahydroisoquinolin-5-yl)quinoline-3-carbonitrile

[0402]

[0403] Step A: tert-Butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(2-methyl-1,2,3,4-tetrahydroisoquinolin-5-yl)quinolin-4-yl)piperazine-1-carboxylate

[0404] In a microwave reaction tube equipped with a magnetron, tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate B2, 150 mg, 0.3 mmol), (2-methyl-1,2,3,4-tetrahydroisoquinolin-5-yl)boronic acid (243 mg, 0.9 mmol), and K3PO4 (189 mg, 0.9 mmol) in CH3CN / H2O (v / v = 3:1, 2 mL) were added and purged with nitrogen for 5 minutes. Pd(dppf)Cl2 (22 mg, 0.03 mmol) was added and the nitrogen purging continued for 5 minutes. The reaction was heated in a microwave oven at 45°C for 4 hours and then cooled to room temperature. EA (10 mL) and H2O (5 mL) were added for separation, and the aqueous phase was extracted with EA (10 mL x 2). The combined organic phases were washed with saturated NaCl (30 mL), dried over anhydrous NaSO, and filtered. The filtrate was concentrated under reduced pressure to give the crude product, which was purified by FCC (SiO, EA / PE = 0-100%) to give tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(2-methyl-1,2,3,4-tetrahydroisoquinolin-5-yl)quinolin-4-yl)piperazine-1-carboxylate (80 mg, 44% yield) as a brown solid. LCMS (m / z): 614.4 (M+H).

[0405] Step B to Step D: 4-(4-acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(2-methyl-1,2,3,4-tetrahydroisoquinolin-5-yl)quinoline-3-carbonitrile

[0406] The subsequent synthesis of Example 10 was carried out as described in the synthesis of Example 1, except that tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(2-methyl-1,2,3,4-tetrahydroisoquinolin-5-yl)quinolin-4-yl)piperazine-1-carboxylate was used in step B instead of tert-butyl (R)-4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate. 1H NMR (400MHz, Methanol-d4) δ8.15 (s, 1H), 7.36-7.22 (m, 4H), 7.01 (t, J=8.8Hz, 1H), 6.86 ( dd, J=16.8, 10.6Hz, 1H), 6.29 (dd, J=16.7, 1.9Hz, 1H), 5.82 (dd, J=10.5, 1.9Hz, 1H), 4.57 (s, 2H), 3.99 (s, 4H), 3.85 (s, 4H), 3.74 (s, 2H), 2.84 (d, J=5.7Hz, 1H), 2.73 (t, J=5.9Hz, 1H), 2.45 (s, 3H). LCMS (m / z): 654.5 (M+H).

[0407] Example 11

[0408]

[0409] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)-1,6-naphthyridine-3-carbonitrile

[0410] The synthesis of Example 11 was carried out as described in Example 1, using tert-butyl 4-(2,7-dichloro-3-cyano-8-fluoro-1,6-naphthyridin-4-yl)piperazine-1-carboxylate (Intermediate E1) instead of tert-butyl (R)-4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate B1) in Step A. 1 H NMR (400MHz, DMSO-d6) δ9.14 (s, 1H), 7.95 (s, 2H), 7.44 (dd, J=8.5, 5.8Hz, 1H), 7.13-7.04 (m, 1H), 6.91 (dd, J=16.6, 10.4Hz, 1H), 6.19 (dd, J =16.7, 2.4Hz, 1H), 5.75 (dd, J = 10.4, 2.4Hz, 1H), 5.47-5.39 (m, 1H), 4.08-3.99 (m, 1H), 3.89 (s, 2H), 3.83 (s, 6H), 3.34 (s, 3H), 3.08 (dd, J=10.0, 6.4Hz, 1H), 2.98 (dd, J=11.1, 6.2Hz, 1H), 2.75 (dd, J=11.3, 2.8Hz, 1H), 2.40 (dd, J=10.0, 4.9Hz, 1H), 2.29 (s, 3H). 19F NMR (376MHz, DMSO-d6) delta -111.71, -135.46. LCMS (m / z): 607.5 (M+H).

[0411] Example 12

[0412]

[0413] 4-((R)-4-propenyl-3-methylamphetamine-1-yl)-2′-amino-6-chloro-5′,8-difluoro-[7,8′-bisquinolinol]-3-carbocyanine

[0414] The synthesis of Example 12 was carried out as described in Example 8, using (5-fluoro-2-((4-methoxybenzyl)amino)quinolin-8-yl)boronic acid instead of (2-((tert-butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid in Step A. LCMS (m / z): 519.1 (M+H).

[0415] Example 13

[0416]

[0417] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-methoxyquinoline-3-carbonitrile

[0418] The synthesis of Example 13 was carried out as described in Example 1, using tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate Int B2) instead of tert-butyl (R)-4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-methylpiperazine-1-carboxylate (Intermediate Int B1) in Step A, and sodium methoxide instead of the NaH mixture of (3R,4R)-4-methoxy-1-methylpyrrolidin-3-ol. 1 H NMR (400MHz, DMSO-d6) δ7.94 (d, J=5.9Hz, 3H), 7.25 (dd, J= 8.4, 5.6Hz, 1H), 7.08 (t, J=8.8Hz, 1H), 6.91 (dd, J=16.7, 10.5Hz, 1H), 6.20 (dd, J=16.7, 2.2Hz, 1H), 5.76 (dd, J=10.4, 2.2Hz, 1H), 4.05 (s, 3H), 3.87 (d, J=15.7Hz, 4H), 3.68 (s, 4H). 19F NMR (376MHz, DMSO) delta -112.27, -119.27. LCMS (m / z): 541.3 (M+H).

[0419] Example 14

[0420]

[0421] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(1-methyl-1H-pyrazol-5-yl)quinoline-3-carbonitrile

[0422] The synthesis of Example 14 was carried out as described in Example 3, using (1-methyl-1H-pyrazol-5-yl)boronic acid instead of (3-((dimethylamino)methyl)phenyl)boronic acid in Step A. 1 H NMR (400MHz, DMSO-d6) δ8.12 (s, 1H), 7.94 (s, 2H), 7.61 (s, 1H), 7.29 (dd, J=8.3, 5.7Hz, 1H), 7.10 (t, J=8.8Hz, 1H), 7.00-6.86 (m, 2H), 6.20 (d, J =16.7Hz, 1H), 5.77 (d, J = 8.7Hz, 1H), 4.04 (s, 3H), 3.90 (d, J = 14.7Hz, 4H), 3.79 (s, 4H). 19 F NMR (376MHz, DMSO) delta -112.07, -118.03. LCMS (m / z): 591.4 (M+H).

[0423] Example 15

[0424]

[0425] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-(1,3-dimethyl-1H-pyrazol-4-yl)-8-fluoroquinoline-3-carbonitrile

[0426] The synthesis of Example 15 was carried out as described in Example 3, using (1,3-dimethyl-1H-pyrazol-4-yl)boronic acid instead of (3-((dimethylamino)methyl)phenyl)boronic acid in Step A. 1H NMR (400MHz, DMSO-d6) δ8.30 (s, 1H), 8.05 (d, J=1.5Hz, 1H), 7.94 (s, 2H), 7.28 (dd, J=8.5, 5.6Hz, 1H), 7.09 (t, J=8.8Hz, 1H), 6.93 (dd, J= 16.7, 10.4Hz, 1H), 6.20 (dd, J=16.6, 2.4Hz, 1H), 5.77 (dd, J=10.4, 2.4Hz, 1H), 3.89 (d, J=11.9 Hz, 7H), 3.73 (d, J=5.5Hz, 4H), 2.41 (s, 3H). 19 F NMR (376MHz, DMSO) delta -112.23, -118.42. LCMS (m / z): 605.4 (M+H).

[0427] Example 16

[0428]

[0429] 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-4-(4-(2-fluoroacryloyl)piperazin-1-yl)quinoline-3-carbonitrile

[0430]

[0431] 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-4-(piperazin-1-yl)quinoline-3-carbonitrile (88 mg, 0.13 mmol), 2-fluoroacrylic acid (13.3 mg, 0.15 mmol), and HATU (77 mg, 0.20 mmol) were dissolved in DCM (5 mL). DIPEA (52 mg, 0.40 mmol) was added dropwise to the reaction mixture with stirring at room temperature. After completion of the reaction, as monitored by LCMS, 20 mL of water was added to the reaction system. The organic phase was separated, collected, concentrated, and dried, and purified by preparative HPLC to afford 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-4-(4-(2-fluoroacryloyl)piperazin-1-yl)quinoline-3-carbonitrile as a white solid (9 mg, 13% yield). 1H NMR (400MHz, DMSO-d6) δ8.89 (s, 1H), 8.06 (s, 1H), 7.94 (s, 2H), 7.29 (dd, J= 8.4, 5.6Hz, 1H), 7.09 (t, J=8.8Hz, 1H), 5.38-5.33 (m, 1H), 5.32 (dd, J=64, 4.1Hz, 1H), 3.91- 3.84 (m 4H), 3.79-3.74 (m, 4H). 19 F NMR (376MHz, DMSO-d6) delta -105.25, -112.15, -117.60. LCMS: 529.3 (M+H).

[0432] Example 17

[0433]

[0434] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-methylquinoline-3-carbonitrile

[0435]

[0436] Step A: tert-Butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-methylquinolin-4-yl)piperazine-1-carboxylate

[0437] To a reaction flask, add tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate B2, 500 mg, 0.99 mmol), methylboronic acid (71 mg, 1.19 mmol), K2CO3 (411 mg, 2.98 mmol), and DME (3 mL). Purge with nitrogen for 3 minutes. Add Pd(dppf)Cl2 (72 mg, 0.099 mmol), continue purging with nitrogen for 3 minutes, then seal the flask and react at 90°C for 1 hour. Cool to room temperature. Add EA (10 mL) and H2O (5 mL) to separate the liquids, and extract the aqueous phase with EA (20 mL x 2). The combined organic phases were washed with saturated NaCl (30 mL), dried over anhydrous NaSO, and filtered. The filtrate was concentrated under reduced pressure to give the crude product, which was purified by FCC (SiO, EA / PE = 0-100%) to give tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-methylquinolin-4-yl)piperazine-1-carboxylate (120 mg, 25% yield) as a yellow solid. LCMS (m / z): 483.3 (M+H).

[0438] Step B to Step D: 4-(4-acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-methylquinoline-3-carbonitrile

[0439] The subsequent synthesis of Example 17 was carried out as described in the synthesis of Example 1, except that tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-methylquinolin-4-yl)piperazine-1-carboxylate was used in Step B instead of tert-butyl (R)-4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate. 1 H NMR (400MHz, DMSO-d6) δ8.04-8.01 (m, 1H), 7.94 (s, 2H), 7.27 (dd, J=8.4, 5.6Hz, 1H), 7.12-7.05 (m, 1H), 6.92 (dd, J=16.7, 10.5Hz, 1H), 6.20 (dd, J=16.6, 2.4Hz, 1H), 5.76 (dd, J=10.5, 2.4Hz, 1H), 3.93 -3.82 (m, 4H), 3.69 (s, 4H), 2.74 (s, 3H). 19 F NMR (376MHz, DMSO-d6) delta -112.20, -118.13. LCMS (m / z): 525.4 (M+H).

[0440] Example 18

[0441]

[0442] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-cyclopropyl-8-fluoroquinoline-3-carbonitrile

[0443] The synthesis of Example 18 was carried out as described in Example 3, using cyclopropylboronic acid in Step A instead of (3-((dimethylamino)methyl)phenyl)boronic acid. 1H NMR (400MHz, DMSO-d6) δ8.01 (d, J=1.5Hz, 1H), 7.96 (s, 2H), 7.27 (dd, J=8.5, 5.6Hz, 1H), 7.13-7.05 (m, 1H), 6.94 (dd, J=16.7, 10.4Hz, 1H), 6.22 (dd, J= 16.6, 2.4Hz, 1H), 5.78 (dd, J=10.4, 2.4Hz, 1H), 4.00-3.81 (m, 4H), 3.79-3.65 (m, 4H), 2.57- 2.54 (m, 1H), 1.25-1.12 (m, 4H). LCMS: 551.5 (M+H).

[0444] Example 19

[0445]

[0446] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-hydroxyquinoline-3-carbonitrile

[0447]

[0448] Step A: tert-Butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)piperazine-1-carboxylate

[0449] KOH (44 mg, 0.78 mmol) was added to a solution of tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (200 mg, 0.39 mmol) in THF and H₂O (v / v = 2:1, 4 mL) with stirring at room temperature. The resulting mixture was stirred at 80°C for 3 h. LCMS monitored the reaction completion. The reaction solution was concentrated under reduced pressure to remove THF and then poured into a saturated aqueous NH₄Cl solution (4 mL). 1N dilute hydrochloric acid was slowly added dropwise to adjust the pH to 5-6. The product precipitated and was filtered. The filter cake was washed twice with water and petroleum ether, respectively, and dried in vacuo to afford tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)piperazine-1-carboxylate (190 mg, crude), which was used directly in the next step. LCMS (m / z): 429.2, 431.1 (M-56).

[0450] Step B to Step D: 4-(4-acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-hydroxyquinoline-3-carbonitrile

[0451] The subsequent synthesis of Example 19 was carried out as described in the synthesis of Example 1, except that tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)piperazine-1-carboxylate was used in Step B instead of tert-butyl (R)-4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(((3R,4R)-4-methoxy-1-methylpyrrolidin-3-yl)oxy)quinolin-4-yl)-2-methylpiperazine-1-carboxylate. 1 H NMR (400MHz, DMSO-d6) δ12.11 (s, 1H), 7.98 (s, 2H), 7.74 (s, 1H), 7.22 (dd, J=8.5, 5.6Hz, 1H), 7.11-7.04 (m 1H), 6.90 (dd, J=16.6, 10.4Hz, 1H), 6.19 (dd, J=16.7, 2.3Hz, 1H), 5.76 (dd, J=10.4, 2.3Hz, 1H), 3.85 (d, J=19.0Hz, 4H), 3.65 (s, 4H). 19 F NMR (376MHz, DMSO) delta -112.03, -123.05. LCMS (m / z): 527.3 (M+H).

[0452] Example 20

[0453]

[0454] 4-(4-Acryloylpiperazin-1-yl)-2-amino-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile

[0455]

[0456] Step A: tert-Butyl 4-(7-bromo-6-chloro-3-cyano-2-((benzhydryl)amino)-8-fluoroquinolin-4-yl)piperazine-1-carboxylate

[0457] 4-(4-Acryloylpiperazin-1-yl)-2-amino-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile (350 mg, 694 μmol), diphenylmethaneimine (126 mg, 694 μmol), Pd2(dba)3 (64 mg, 69.4 μmol), XantPhos (80 mg, 139 μmol), Cs2CO3 (679 mg, 2.08 mmol) and dioxane (5 mL) were added to a microwave tube. After nitrogen replacement for 1 minute, the tube was covered and heated to 80°C for 1 h. After the reaction was completed, the system was cooled to room temperature, and the reaction solution was poured into water (50 mL) and extracted with EA (50 mL x 3). The extracts were collected, concentrated, and further purified by FCC (SiO2, EA / PE = 0-50%) to give tert-butyl 4-(7-bromo-6-chloro-3-cyano-2-((benzhydryl)amino)-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (280 mg, 62% yield) as a yellow solid. LCMS (m / z): 650.4 (M+H).

[0458] Step B: tert-Butyl 4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-2-((benzhydryl)amino)-8-fluoroquinolin-4-yl)piperazine-1-carboxylate

[0459] At room temperature, a reaction tube equipped with a magnetic separator was charged with tert-butyl 4-(7-bromo-6-chloro-3-cyano-2-((benzhydryl)amino)-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (280 mg, 431 μmol), (2-((tert-butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid (135 mg, 431 μmol), Pd(dppf)Cl2 (32 mg, 43 μmol), Na2CO3 (137 mg, 1.29 mmol), and a 1,4-dioxane / H2O (v / v = 4:1, 8 mL) solution. After purging with nitrogen for 1 min, the reaction tube was sealed and microwave-heated to 115°C for 2 h. The mixture was cooled to room temperature, diluted with water (60 mL), extracted with EA (50 mL×3), dried over anhydrous NaSO, and concentrated under reduced pressure to give tert-butyl 4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-2-((benzhydryl)amino)-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (310 mg, yield 86%). LCMS (m / z): 836.6 (M+H).

[0460] Step C: 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-((diphenylmethylene)amino)-8-fluoro-4-(piperazin-1-yl)quinoline-3-carbonitrile

[0461] At room temperature, TFA:CH2Cl2=1:1 (15 mL) was added to tert-butyl 4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-2-((diphenylmethylene)amino)-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (310 mg, 371 μmol) and stirred for half an hour. After completion of the reaction, LCMS analysis showed that the reaction was diluted with EA (10 mL) and poured into saturated NaHCO3 (20 mL). The reaction mixture was extracted with EA (30 mL×3) and dried over anhydrous Na2SO4. The extracts were collected to give 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-((diphenylmethylene)amino)-8-fluoro-4-(piperazin-1-yl)quinoline-3-carbonitrile (140 mg, yield 59%). LCMS (m / z): 636.6 (M+H).

[0462] Step D: 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-((benzhydryl)amino)-8-fluoroquinoline-3-carbonitrile

[0463] Under ice bath conditions, a diluted solution of acryloyl chloride (24 mg, 264 μmol) in CH2Cl2 (0.5 mL) was added dropwise to a mixed solution of 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-((diphenylmethylene)amino)-8-fluoro-4-(piperazin-1-yl)quinoline-3-carbonitrile (140 mg, 220 μmol), DIPEA (85 mg, 660 μmol) and CH2Cl2 (5 mL) using a syringe and stirred for 10 minutes. After completion of the reaction, the reaction mixture was poured into water (50 mL) and extracted with CH2Cl2 (30 mL x 3). The extracts were collected and concentrated to afford 4-(4-acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-((benzhydryl)amino)-8-fluoroquinoline-3-carbonitrile (150 mg, crude) as a yellow solid. LCMS (m / z): 690.0 (M+H).

[0464] Step E: 4-(4-Acryloylpiperazin-1-yl)-2-amino-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile

[0465] At room temperature, 4-(4-acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-((benzhydryl)amino)-8-fluoroquinoline-3-carbonitrile (150 mg, 217 μmol) was dissolved in 30 mL of ethanol. 1.5 mL of saturated aqueous citric acid was added, and the mixture was heated to 60°C and stirred overnight. LCMS detected the formation of the product. The reaction mixture was concentrated, and then water (30 mL) was added. The reaction mixture was concentrated with EA (50 mL x 3). The resulting crude product was further separated by preparative HPLC to afford 4-(4-acryloylpiperazin-1-yl)-2-amino-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile (11 mg, 10% yield). 1 H NMR (400MHz, Chloroform-d) δ7.71 (d, J=1.6Hz, 1H), 7.25-7.15 (m, 1H), 7.05-6.91 (m, 1H), 6.69-6.55 (m, 1H), 6.38 (dd, J=16.7, 1.8Hz, 1H), 5.80 (dd, J=10.6, 1.8Hz, 1H), 5.68 (s, 2H), 4.06-3.80 (m, 4H), 3.76-3.56 (m, 4H). 19 F NMR (376MHz, Chloroform-d) delta -111.62, -119.98. LCMS (m / z): 526.4 (M+H).

[0466] Synthesis of compounds 21-2 and 22-2

[0467]

[0468] (2S)-4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylic acid benzyl ester (21-2) and (2S)-4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylic acid benzyl ester (22-2)

[0469]

[0470] Step A: (S)-benzyl 4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (21-1) and (S)-benzyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (22-1)

[0471] To a round-bottom flask equipped with a magnetic bar were added benzyl (S)-4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (Int F, 500 mg, 866.2 μmol), Pd(PPh3)4 (100 mg, 87 μmol), formic acid (159 mg, 3.46 mmol), and triethylamine (438 mg, 4.33 mmol). After nitrogen was purged three times, DMF (15 mL) was added, and the reaction system was heated to 55°C and stirred for 6 h. After the reaction was completed, the mixture was cooled to room temperature and poured into 150 mL of water to produce a yellow solid. The solid was filtered and the filter cake was washed with PE (15 mL) and H2O (15 mL). The solid was collected to give a mixture of (S)-benzyl 4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (21-1) and (S)-benzyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (22-1) (530 mg). LCMS (m / z): 544.3 (M+H) (21-1) and LCMS (m / z): 558.3 (M+H) (22-1).

[0472] Step B: (2S)-benzyl 4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (21-2) and (2S)-benzyl 4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (22-2)

[0473] At room temperature, a reaction tube equipped with a magnetic bar was charged with a mixture of (S)-benzyl 4-(7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (21-1) and benzyl (S)-4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (22-1) (530 mg, 976.43 μmol), (2-((tert-butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid (335.24 mg, 1.07 mmol), Pd(dppf)Cl2 (71.64 mg, 97.64 μmol), Na2CO3 (313 mg, 2.93 mmol) and 1,4-dimethylbenzyl boronic acid (21-1). Dioxane / H2O (4:1, 10 mL) solution was purged with nitrogen for 1 min, the reaction tube was sealed, and microwave-heated to 115°C for 2 h. The mixture was cooled to room temperature, diluted with water (60 mL), and extracted with EA (50 mL x 3). The combined extracts were dried over anhydrous Na2SO4, concentrated under reduced pressure, and purified by FCC (SiO2, EA / PE = 0-60%) to give (2S)-4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylic acid benzyl ester (21-2) (80 mg, yield 17%). LCMS (m / z): 730.5 (M+H); and (2S)-4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (21-2). Benzyl 2-(cyanomethyl)piperazine-1-carboxylate (22-2) (380 mg, yield 78%). LCMS (m / z): 746.5 (M+H), 646.4 (M-100+H).

[0474] Example 21

[0475]

[0476] 4-((S)-4-Acryloyl-3-(cyanomethyl)piperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile

[0477]

[0478] Step A: tert-Butyl (4-(6-chloro-3-cyano-4-((S)-3-(cyanomethyl)piperazin-1-yl)-8-fluoroquinolin-7-yl)-7-fluorobenzo[d]thiazol-2-yl)carbamate

[0479] TMSI (110 mg, 548 μmol) was added dropwise to a mixed solution of (2S)-benzyl 4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (80 mg, 110 μmol) and MeCN (3 mL) at room temperature, and the reaction was stirred at room temperature for 5 hours. Et3N (1 mL) was then added and stirring was continued for 10 minutes. After completion of the reaction, as determined by LCMS, the reaction solution was poured into H2O (30 mL) and extracted with EA (30 mL x 3). The extracts were collected to provide tert-butyl (4-(6-chloro-3-cyano-4-((S)-3-(cyanomethyl)piperazin-1-yl)-8-fluoroquinolin-7-yl)-7-fluorobenzo[d]thiazol-2-yl)carbamate (68 mg, crude). LCMS (m / z): 596.4 (M+H).

[0480] Step B: 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-4-((S)-3-(cyanomethyl)piperazin-1-yl)-8-fluoroquinoline-3-carbonitrile

[0481] TFA:DCM (v / v=1:1, 5 mL) was added to tert-butyl (4-(6-chloro-3-cyano-4-((S)-3-(cyanomethyl)piperazin-1-yl)-8-fluoroquinolin-7-yl)-7-fluorobenzo[d]thiazol-2-yl)carbamate (68 mg, crude) at room temperature and stirred at room temperature for half an hour. After the reaction was completed, LCMS analysis showed that the reaction solution was concentrated to remove most of the reaction solution. The reaction solution was then diluted with EA (5 mL) and poured into saturated NaHCO3 (20 mL). The product was extracted with EA (30 mL×3) and dried over anhydrous Na2SO4. The extracts were collected to give 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-4-((S)-3-(cyanomethyl)piperazin-1-yl)-8-fluoroquinoline-3-carbonitrile (60 mg, crude). LCMS (m / z): 496.3 (M+H).

[0482] Step C: 4-((S)-4-Acryloyl-3-(cyanomethyl)piperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile

[0483] Under ice bath conditions, a diluted solution of acryloyl chloride (11 mg, 121 μmol) in DCM (0.5 mL) was added dropwise to a mixed solution of 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-4-((S)-3-(cyanomethyl)piperazin-1-yl)-8-fluoroquinoline-3-carbonitrile (60 mg, crude), saturated NaHCO3 (1.5 mL) and DCM (3 mL) using a syringe and stirred for 10 minutes. After the reaction was completed, the reaction solution was poured into water (30 mL) and extracted with DCM (20 mL × 2). The extract was collected and concentrated, and the crude product was further separated by preparative high-performance liquid chromatography to give a white solid 4-((S)-4-acryloyl-3-(cyanomethyl)piperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile (7 mg, total yield of three steps 10%). 1 H NMR (400 MHz, Methanol-d4) δ8.77 (s, 1H), 8.07 (d, J=1.6Hz, 1H), 7.26-7.11 (m, 1H), 7.00-6.88 (m, 1H), 6.82 (s, 1H), 6.26 (dd, J=16.7, 1.9Hz, 1H), 5.80 (dd, J=10.7, 1.8Hz, 1H), 5.24 (d, J=24.8Hz, 1H), 4.21 (s, 1H), 4.10-3.71 (m, 4H), 3.43 (d, J=12.4Hz, 1H), 3.15 (s, 1H), 3.01-2.86 (m, 1H). 19 F NMR (376MHz, Methanol-d4) delta -114.73, -118.76. LCMS (m / z): 550.4 (M+H).

[0484] Example 22

[0485]

[0486] 4-((S)-4-Acryloyl-3-(cyanomethyl)piperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-hydroxyquinoline-3-carbonitrile

[0487]

[0488] Step A: 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-4-((S)-3-(cyanomethyl)piperazin-1-yl)-8-fluoro-2-hydroxyquinoline-3-carbonitrile

[0489] To a mixed solution of (2S)-benzyl 4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (380 mg, 510 μmol) and MeCN (10 mL) was added dropwise TMSI (510 mg, 2.55 mmol) at room temperature. The mixture was stirred at room temperature for 5 hours, and then EtN (3 mL) was added and stirring was continued for 10 minutes. After completion of the reaction, as determined by LCMS, the reaction solution was poured into H2O (30 mL) and extracted with EA (50 mL x 2). The extracts were collected to afford 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-4-((S)-3-(cyanomethyl)piperazin-1-yl)-8-fluoro-2-hydroxyquinoline-3-carbonitrile as a yellow solid (200 mg, 76% yield). LCMS (m / z): 512.3 (M+H).

[0490] Step B: 4-((S)-4-Acryloyl-3-(cyanomethyl)piperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-hydroxyquinoline-3-carbonitrile

[0491] Acryloyl chloride (38 mg, 430 μmol) diluted in CH2Cl2 (0.5 mL) was added dropwise to a mixed solution of 7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-4-((S)-3-(cyanomethyl)piperazin-1-yl)-8-fluoroquinoline-3-carbonitrile (200 mg, 391 μmol), DIPEA (151 mg, 1.17 mmol) and DCM (5 mL) in an ice bath using a syringe and stirred for 10 minutes. After the reaction was completed, the reaction solution was poured into water (50 mL) and extracted with DCM (30 mL × 3). The extracts were collected and concentrated. The crude product was further separated by preparative high performance liquid chromatography to give a yellow solid 4-((S)-4-acryloyl-3-(cyanomethyl)piperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-hydroxyquinoline-3-carbonitrile (52 mg, yield 24%). 1H NMR (400MHz, Methanol-d4) δ7.83 (s, 1H), 7.27-7.16 (m, 1H), 7.04-6.95 (m, 1H), 6.88 (s, 1H), 6.32 (dd, J=16.6, 1.9Hz, 1H), 5.86 (dd, J=10.6, 1.9Hz, 1H), 5.48-5.12 (m, 1H), 5.02 (s, 1H), 4.09-3.80(m, 4H), 3.53-3.39(m, 2H), 3.05-2.87(m, 1H). 19 F NMR (376MHz, DMSO-d6) δ-112.18, -123.14. LCMS (m / z): 566.4 (M+H).

[0492] Examples 23-31

[0493] The following compounds were prepared in a similar manner as described above:

[0494]

[0495]

[0496] Example 32

[0497]

[0498] 4-((S)-4-Acryloyl-3-(cyanomethyl)piperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-methylquinoline-3-carbonitrile

[0499] The synthesis of Example 32 was carried out as described in Example 17, except that (S)-benzyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)-2-(cyanomethyl)piperazine-1-carboxylate (Intermediate F) was used instead of tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (Intermediate B2) in Step A. LCMS (m / z): 564.2 (M+H).

[0500] Example 33

[0501]

[0502] 4-(4-Acryloylpiperazin-1-yl)-2-amino-7-(2-aminobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile

[0503] The synthesis of Example 33 was carried out as described in Example 20, using (2-((tert-butoxycarbonyl)amino)benzo[d]thiazol-4-yl)boronic acid instead of (2-((tert-butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid in Step B. 1 HNMR (400MHz, DMSO-d6) δ7.80-7.69 (m, 2H), 7.63 (s, 2H), 7.19-7.01 (m, 4H), 6.91 (dd, J=16.6, 10.5Hz, 1H), 6.19 (dd, J=16.6, 2.4Hz, 1H), 5.75 (dd, J=10.3, 2.4Hz, 1H), 3.97-3.76 (m, 4H), 3.66-3.49 (m, 4H). LCMS (m / z): 508.4 (M+H).

[0504] Example 34

[0505]

[0506] 4-(4-Acryloylpiperazin-1-yl)-2-amino-7-(2-amino-5-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoroquinoline-3-carbonitrile

[0507] The synthesis of Example 34 was carried out as described in Example 20, using (2-((tert-butoxycarbonyl)amino)-5-fluorobenzo[d]thiazol-4-yl)boronic acid instead of (2-((tert-butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid in Step B. 1 H NMR (400 MHz, DMSO-d6) δ8.00-7.67 (m, 4H), 7.24-7.08 (m, 2H), 7.08-6.99 (m, 1H), 6.91 (dd, J=16.6, 10.5Hz, 1H), 6.19 (d, J=16.6Hz, 1H), 5.76 (d, J=10.8Hz, 1H), 3.99-3.73 (m, 4H), 3.73-3.46 (m, 4H). LCMS (m / z): 526.4 (M+H).

[0508] Example 35

[0509]

[0510] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(methylamino)quinoline-3-carbonitrile

[0511]

[0512] Step A: tert-Butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(methylamino)quinolin-4-yl)piperazine-1-carboxylate

[0513] NaH (143 mg, 3.6 mmol) was added to a mixed solution of methylamine hydrochloride (121 mg, 1.8 mmol) and THF (8 mL) at room temperature and stirred for 10 minutes. Tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (300 mg, 0.60 mmol) was then added to the mixture and allowed to react at room temperature for 2 hours. After LCMS analysis of the product, the reaction solution was added to H₂O (30 mL) and extracted with EA (50 mL x 3). The combined organic phases were washed with saturated brine and dried over anhydrous Na₂SO₄ to yield tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(methylamino)quinolin-4-yl)piperazine-1-carboxylate (200 mg, 67% yield) as a yellow solid. LCMS (m / z): 499.6 (M+H).

[0514] Step B to Step D: 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-fluoro-2-(methylamino)quinoline-3-carbonitrile

[0515] The subsequent synthetic steps of Example 35 were carried out as described in Example 19, except that tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-(methylamino)quinolin-4-yl)piperazine-1-carboxylate was used instead of tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)piperazine-1-carboxylate in Step B. LCMS (m / z): 540.4 (M+H).

[0516] Example 36

[0517]

[0518] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-(dimethylamino)-8-fluoroquinoline-3-carbonitrile

[0519] The synthesis of Example 36 was carried out as described in Example 35, except that dimethylamine hydrochloride was used in place of methylamine hydrochloride in Step A. 1H NMR (400MHz, DMSO-d6) δ7.91 (s, 2H), 7.80 (s, 1H), 7.23-7.20 (m, 1H), 7.08-7.03 (m, 1H), 6.94-6.87 (m, 1H), 6.19 (d, J=1.6Hz, 1H), 5.76 (d, J=1.2Hz, 1H), 3.87 (d, J=1.6Hz, 4H), 3.64 (s, 4H), 3.16 (s, 6H). 19 F NMR (376MHz, DMSO-d6) delta -112.51, -120.42. LCMS (m / z): 554.5 (M+H).

[0520] Example 37

[0521]

[0522] N-(4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-2-yl)acetamide

[0523]

[0524] Step A: tert-Butyl 4-(2-acetylamino-7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate

[0525] To a round-bottom flask containing a magnetic rod at room temperature, tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (600 mg, 1.2 mmol), acetamide (77 mg, 1.3 mmol), Cs2CO3 (1.16 g, 3.6 mmol), and dioxane (10 mL) were added. After nitrogen was replaced, Pd2(dba)3 (109 mg, 0.12 mmol) and Xantphos (65 mg, 0.12 mmol) were added. After further nitrogen replacement, the system was heated to 90°C and stirred overnight. The reaction was monitored for completion by LCMS. The reaction solution was poured into water and extracted with ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure to give a crude product, which was purified by FCC (SiO2, EA / PE = 0-100%) to afford tert-butyl 4-(2-acetamido-7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (205 mg, 33% yield). LCMS (m / z): 526.1 (M+H).

[0526] Step B to Step D: N-(4-(4-acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-2-yl)acetamide

[0527] The subsequent synthetic steps of Example 37 were carried out with reference to Example 19, except that tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoro-2-hydroxyquinolin-4-yl)piperazine-1-carboxylate was used in Step B instead of tert-butyl 4-(2-acetylamino-7-bromo-6-chloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate. 1 H NMR (400MHz, DMSO-d6) δ11.01 (s, 1H), 8.03 (d, J=1.5Hz, 1H), 7.97-7.90 (m, 2H), 7.27 (dd, J=8.4, 5.6Hz, 1H), 7.13-7.04 (m, 1H), 6.91 (dd, J=16.6, 10.4 Hz, 1H), 6.19 (dd, J=16.7, 2.4Hz, 1H), 5.76 (dd, J=10.4, 2.4Hz, 1H), 3.97-3.81 (m, 4H), 3.74-3.60 (m, 4H), 2.13 (s, 3H). 19 F NMR (376MHz, DMSO-d6) delta -112.16, -118.37. LCMS (m / z): 568.1 (M+H).

[0528] Example 38

[0529]

[0530] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-ethynyl-8-fluoroquinoline-3-carbonitrile

[0531]

[0532] Step A: tert-Butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-((trimethylsilyl)ethynyl)quinolin-4-yl)piperazine-1-carboxylate

[0533] At room temperature, tert-butyl 4-(7-bromo-2,6-dichloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (500 mg, 0.99 mmol), ethynyltrimethylsilane (292 mg, 3.0 mmol), Pd(PPh3)4 (115 mg, 0.099 mmol), CuI (38 mg, 0.20 mmol), and TEA (301 mg, 3.0 mmol) were added to DMF (10 mL), replaced with nitrogen three times, and reacted at 70°C for 16 hours. After product formation was detected, the reaction solution was added to H2O (100 mL) and extracted with EA (100 mL x 2), washed with saturated brine, dried over anhydrous Na2SO4, concentrated, and purified by FCC (SiO2, EA / PE = 0-50%) to give tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-((trimethylsilyl)ethynyl)quinolin-4-yl)piperazine-1-carboxylate (350 mg, 62% yield) as a yellow solid. LCMS (m / z): 566.0 (M+H).

[0534] Step B: tert-Butyl 4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoro-2-((trimethylsilyl)ethynyl)quinolin-4-yl)piperazine-1-carboxylate

[0535] At room temperature, a reaction flask containing a magnetic bar was charged with tert-butyl 4-(7-bromo-6-chloro-3-cyano-8-fluoro-2-((trimethylsilyl)ethynyl)quinolin-4-yl)piperazine-1-carboxylate (210 mg, 0.37 mmol), (2-((tert-butoxycarbonyl)amino)-7-fluorobenzothiazol-4-yl)boronic acid (139 mg, 0.45 mmol), K2CO3 (154 mg, 1.1 mmol), and dioxane / water (v / v = 3:1, 5 mL). The atmosphere was purged with nitrogen, and Pd(dppf)Cl2 (27 mg, 0.037 mmol) was added. The atmosphere was purged with nitrogen again, and the reaction mixture was heated to 90°C with stirring for 1 hour. The reaction mixture was cooled to room temperature, poured into water, and extracted with ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure to give a crude product, which was purified by FCC (SiO2, EA / PE = 0-100%) to afford tert-butyl 4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoro-2-((trimethylsilyl)ethynyl)quinolin-4-yl)piperazine-1-carboxylate (90 mg, 32% yield). LCMS (m / z): 753.2 (M+H).

[0536] Step C: tert-Butyl 4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-2-ethynyl-8-fluoroquinolin-4-yl)piperazine-1-carboxylate

[0537] By 4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoro-2-((trimethylsilyl)ethynyl)quinoline-4-yl)piperazine-1-carboxylic acid tert-butyl ester (90mg, 0.12mmol), KCO (33mg, 0.24mmol) and acetonitrile (2mL) mixture stirred at room temperature for 1h.LCMS monitoring reaction completion, the system was poured into water, EA extraction.The combined organic phase was washed with salt water, dried over anhydrous sodium sulfate, filtered and concentrated to give 4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-2-ethynyl-8-fluoroquinoline-4-yl)piperazine-1-carboxylic acid tert-butyl ester (70mg, crude product), without purification, directly used in the next step reaction. LCMS (m / z): 681.2 (M+H).

[0538] Steps D and E: 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-2-ethynyl-8-fluoroquinoline-3-carbonitrile

[0539] The steps of removing the protecting group and introducing acryloyl chloride involved in the subsequent synthesis of Example 38 can be basically carried out by referring to the method described in Example 1. LCMS (m / z): 535.1 (M+H).

[0540] Example 39

[0541]

[0542] 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-methoxyquinoline-3-carbonitrile

[0543]

[0544] Step A: tert-Butyl 4-(7-(2-((tert-Butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-methoxyquinolin-4-yl)piperazine-1-carboxylate

[0545] To a reaction flask equipped with a magnetic bar, tert-butyl 4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-fluoroquinolin-4-yl)piperazine-1-carboxylate (70 mg, 0.11 mmol) and sodium methoxide (20 mg, 0.37 mmol) were added. Molecular sieves (100 mg) and dioxane (2 mL) were added, the cap was tightened, and the mixture was heated to 110°C and stirred for 8 h. After completion of the reaction, EA (20 mL) was added to dilute the reaction system, which was washed sequentially with water (15 mL) and saturated brine (15 mL), and dried over anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to afford the crude product, which was purified by FCC (SiO2, EA / PE = 0-80%) to afford tert-butyl 4-(7-(2-((tert-butoxycarbonyl)amino)-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-3-cyano-8-methoxyquinolin-4-yl)piperazine-1-carboxylate (42 mg, 59% yield). LCMS (m / z): 669.1 (M+H).

[0546] Steps B and C: 4-(4-Acryloylpiperazin-1-yl)-7-(2-amino-7-fluorobenzo[d]thiazol-4-yl)-6-chloro-8-methoxyquinoline-3-carbonitrile

[0547] The steps of removing the protecting group and introducing acryloyl chloride involved in the subsequent synthesis of Example 39 can basically be carried out by referring to the method described in Example 1. 1 H NMR (400MHz, DMSO-d6) δ8.86 (s, 1H), 7.96 (s, 1H), 7.89-7.82 (m, 2H), 7.16 (dd, J=8.4, 5.7Hz, 1H), 7.07-7.01 (m, 1H), 6.91 (dd, J=16.7, 10.5Hz, 1H), 6.19 (dd, J=16.7, 2.4Hz, 1H), 5.75 (dd, J=10.4, 2.4Hz, 1H), 3.94-3.82 (m, 4H), 3.78 (s, 3H), 3.70-3.64 (m, 4H). 19 F NMR (376MHz, DMSO-d6) delta-113.34. LCMS (m / z): 523.1 (M+H).

[0548] Active Examples

[0549] Example 1: Inhibitory effect of the compounds of the present invention on proliferation of KRas G12C mutant cells

[0550] This experiment used Promega Luminescent Cell Viability Assay kit was used to evaluate and validate the proliferation inhibitory activity of the compound of the present invention on KRas G12C mutated NCI-H358 human non-small cell lung cancer cells.

[0551] Experimental Materials: NCI-H358 cell line (Cell Resource Center, Institute of Basic Medicine, Chinese Academy of Medical Sciences, Resource Number: 3111C0001CCC000470), 96-well transparent flat-bottom black-walled cell culture plates (Greiner Bio-one, Catalog Number #655096), RPMI-1640 medium (GE, Catalog Number #SH30809.01), fetal bovine serum (FBS) (Thermo Fisher, Catalog Number #10099-141), Luminescent Cell Viability Assay Kit (Promega, Catalog No. G7573), PBS (Solarbio, Catalog No. P1020), Trypsin (Thermo Fisher, Catalog No. 25200072), DMSO (Sigma, Catalog No. D2650), Methylcellulose (SIGMA, Catalog No. 9004-67-5).

[0552] [Experimental Procedure]: Add 180 μL of cell suspension (RPMI1640 solution containing 1% methylcellulose and 10% FBS) to a 96-well cell culture plate to a cell density of 1500 viable cells / well. A control group containing only 3D complete medium (RPMI1640 solution containing 1% methylcellulose and 10% FBS) without cells or compound (i.e., culture medium control) was established. A control group containing cells without compound (i.e., cell control) was also established. During the assay, compound AMG510 or the following reference compounds were used as positive controls. The cell plate was cultured in a cell culture incubator overnight. A 10x drug solution (1% DMSO in 10% FBS in RPMI 1640) was prepared at a concentration of 10 μM. 20 μL of drug solution was added to each well of a 96-well plate seeded with cells, resulting in a final compound concentration of 1 μM per well. Three replicate wells were prepared for each compound, and the DMSO content was 0.1%. A solution of compound AMG510 or a reference compound was prepared and added to the positive control wells in the same manner. The cell plates were incubated in a cell incubator for an additional 120 h. For endpoint detection, CellTiter-Glo reagent was thawed and the cell plates were equilibrated at room temperature for 30 min. 100 μL of CellTiter-Glo was added to each well of the cell plates. The cells were shaken on an orbital shaker for 5 min to fully lyse the cells. The cell plates were left at room temperature for 20 min to stabilize the luminescence signal. Luminescence values for each well were scanned across the full wavelength range using a multi-function microplate reader (Molecular Devices, Spectramax M3).

[0553] [Test samples] Compounds of Examples 1-39, and reference compound A (prepared and characterized according to the method described in WO2020 / 081282 A1) and reference compound B (Prepared and characterized with reference to the method described in WO2015054572).

[0554] [Data Analysis] The cell inhibition rate under the action of each compound was calculated using the following formula and GraphPad Prism 7.0 software:

[0555] Inhibition rate % = [1-(Lum 待测药 -Lum 培养液对照 ) / (Lum 细胞对照 -Lum 培养液对照 )】×100%

[0556] IC 50 The data were fitted with nonlinear S-curve regression using GraphPad Prism 7.0 software to obtain the dose-effect curve, which was then calculated.

[0557] [Experimental Results] The compounds of the present invention showed satisfactory anti-cell proliferation activity against NCI-H358 human non-small cell lung cancer cells with KRas G12C mutation. Specifically, the compounds tested in the examples all showed anti-cell proliferation activity, IC 50 The value is generally <1 μM, such as <0.5 μM, <0.1 μM, preferably <50 nM, more preferably <20 nM, most preferably <10 nM. For example, the compounds of Examples 17, 20, 32, 33, 34, and 37 all show an IC value of <50 nM. 50 The compounds of Examples 1, 2, 3, 5, 6, 8, 9, 10, and 39 showed IC values < 20 nM. 50 The specific data of some representative example compounds are shown in Table 1.

[0558] Table 1. Inhibitory activity of representative compounds on NCI-H358 cell proliferation (inhibition rate at 1 μM concentration and IC 50 )

[0559] Compound Inhibition rate % (1 μM) <![CDATA[IC 50 (μM) <!-- 63 -->]]> Example 1 94.7 0.015 Example 2 97.6 0.003 Example 3 97.7 0.009 Example 4 94.8 0.053 Example 5 94.9 0.007 Example 6 95.0 0.017 Example 7 90.0 0.26 Example 8 94.0 0.018 Example 9 98.8 0.0086 Example 10 96.1 0.018 Reference compound A 96.9 0.011 Reference compound B / 0.35

[0560] Example 2: Pharmacokinetic properties of the compounds of the present invention in rats

[0561] 2.1 The pharmacokinetic characteristics of some compounds of the present invention were evaluated by rat cassette pharmacokinetic experiment.

[0562] [Experimental materials]: Male SD rats, age: 6-8 weeks, weight 220-250g, purchased from Zhaoyan (Suzhou) New Drug Research Center Co., Ltd.; Tolbutamide (Aladdin, product number H1401054); Sulfonbutyl β-cyclodextrin (Captisol, Shandong Binzhou Zhiyuan Biological, product number 20191013); Propylene glycol (15) stearate (Solutol, Meilun Biological, product number S0206A); DMSO (Vetec, product number WXBD0293V); Acetonitrile (Sigma-Aldrich, product number WXBD1744V); Methanol (Sigma-Aldrich, product number WXBD2831V).

[0563] [Experimental Procedure]: The compound combination was formulated in a solvent of 5% DMSO / 10% Solutol / 85% (20% Captisol) to a final concentration of 1 mg / mL for each compound. The drug preparation was injected into the tail vein of SD rats at an injection volume of 1 mL / kg. Blood was collected by external jugular vein puncture at 5 minutes, 15 minutes, 30 minutes, 1 hour, 2 hours, 4 hours, 8 hours, and 24 hours. The blood was centrifuged at low temperature for 20 minutes, and the plasma was collected and stored at -80°C until testing.

[0564]

Sample Analysis

[0565] Preparation of standard curve: For each compound, aspirate 20 μL of 1 mg / mL DMSO stock solution and transfer it to 900 μL of 50% methanol working solution. Dilute the solution serially to obtain a standard curve working solution with the concentrations of 20,000, 10,000, 5,000, 1,000, 500, 100, 50, 20, and 10 ng / mL. Then, aspirate 5 μL of the standard curve working solution and mix it with 45 μL of rat blank plasma to obtain a standard curve with the concentrations of 2,000, 1,000, 500, 100, 50, 10, 5, 2, and 1 ng / mL for quantification of unknown samples.

[0566] Sample pretreatment: 50 μL of unknown plasma sample and standard curve sample were added with 250 μL of acetonitrile containing internal standard as a precipitant to precipitate plasma proteins and extract the test compound in the plasma. The mixture was centrifuged at low temperature for 20 minutes, and the supernatant was collected. The supernatant was mixed with 0.1% formic acid in water, and 5 μL was injected for analysis of drug blood concentration.

[0567] Data Processing: Mass spectrometry software was used to draw a standard curve and quantify unknown samples. Pharmacokinetic parameters were calculated using Winnonlin 8.2 based on the drug concentrations at each time point in the unknown samples.

[0568] [Experimental Results]: The experimental results show that the compounds of the present invention exhibit good or even improved pharmacokinetic properties in the cassette administration pharmacokinetic evaluation.

[0569] Table 2.

[0570]

[0571]

[0572] Compound AMG510: Prepared according to the method described by Lanman B et al., J. Med. Chem. 2020, 63, 52-65.

[0573] 2.2 Pharmacokinetic properties of representative compounds of the present invention

[0574] [Test materials] Same as 2.1 above.

[0575] Experimental Procedure: Rats were intravenously administered the compound at a concentration of 3 mg / mL using a method similar to that described in 2.1 above. Drug formulation, administration, and sample collection were performed as described in 2.1. For the oral administration group, the drug was prepared as a 3 mg / mL suspension in 0.5% methylcellulose MC-400cp (Aladdin, M112866) and administered orally at 10 mL / kg. Blood was collected by external jugular vein puncture at 0.25, 0.5, 1, 2, 4, 6, 8, and 24 hours after administration. Plasma was collected by cryogenic centrifugation for 20 minutes and stored at -80°C until analysis.

[0576] Subsequent sample analysis and data processing methods are similar to those described in 2.1 and are carried out accordingly.

[0577] [Experimental results]: The results show that Example 9 has excellent pharmacokinetic properties, as shown in Table 3.

[0578] Table 3

[0579]

[0580] Example 3: Antitumor activity of the compounds of the present invention in a human non-small cell lung cancer NCI-H358 xenograft mouse model

[0581] This study evaluated and verified the proliferation inhibitory activity of the compounds of the present invention in a human non-small cell lung cancer NCI-H358 xenograft mouse model.

[0582] Experimental Materials: The NCI-H358 cell line, carrying the KRAS G12C mutation, was provided by Kangyuan Broad Biotechnology (Beijing) Co., Ltd. (from ATCC, catalog number CRL-5807). Female NPSG mice were provided by Beijing Phinok Biotechnology Co., Ltd.

[0583] [Experimental procedures]: 6-8 week old female NPSG mice were subcutaneously inoculated with 5×10 6 NCI-H358 cells (containing 50% matrigel) were seeded in a volume of 0.1 mL. When the tumor grew to an average volume of 160-220 mm 3The mice were randomly divided into groups according to the tumor size and body weight for drug administration. Drug administration began immediately after grouping, and the day of drug administration was considered as day 0. The drug was administered once a day by gavage at a dose of 10 mg / kg or solvent control (50 mM citrate buffer containing 10% cyclodextrin pH 5.0). During the experiment, tumor volume and body weight were measured twice a week. The formula for calculating tumor volume is V = D × d × d / 2, where D is the long diameter of the tumor and d is the short diameter of the tumor. Average tumor inhibition rate TGI% = [(C 平均值 -C 0平均值 )-(T 平均值 -T 0平均值 )] / (C 平均值 -C 0平均值 )*100%, where T is the tumor volume of the drug-treated group, T0 is the initial tumor volume of the drug-treated group, C is the tumor volume of the control group, and C0 is the initial tumor volume of the control group.

[0584]

Experimental results

[0585] Table 4 shows that representative compounds of the present invention significantly inhibited NCI-H358 tumor growth. At equivalent doses, the compounds demonstrated comparable or superior tumor inhibition compared to the control compound, AMG510. Furthermore, there was no significant change in the body weight of mice in any of the groups.

[0586] Table 4

[0587] Group <![CDATA[Tumor volume (mm 3 )(Day 14)]]> TGI (%) Solvent control group 1170 / Example 1 478.4 71 Example 2 497.3 69 Example 8 519.2 67 AMG510 525.1 66

[0588] In addition, the same materials and methods as above were used to investigate the tumor inhibitory activity of the compound of Example 9 at different doses in the above non-small cell lung cancer mouse model and its effect on body weight. The results are shown in the attached table. Figure 1 As shown, the representative example compounds showed excellent tumor inhibitory activity and had no obvious side effects on body weight.

[0589] Example 4: Cytochrome P450 inhibition test of the compounds of the present invention

[0590] This experiment evaluates the inhibitory effect of the inventive compounds on cytochrome P450.

[0591] Experimental Materials: Human liver microsomes (Corning, Catalog No. 452161); reduced nicotinamide adenine dinucleotide phosphate (NADPH, MCE, Catalog No. HY-F0003 / CS-4998); phenacetin, diclofenac, α-naphthoflavone, omeprazole, and ketoconazole were purchased from TCI; S-mephenytoin and testosterone were purchased from CAYMAN; midazolam was purchased from Bioreclamation IVT; quinidine was purchased from Damas-beta; sulfaphenazole was purchased from MCE; and bufuralol was purchased from TRC.

[0592]

Experimental steps

[0593] Prepare 0.1 M potassium phosphate buffer (K-buffer): Prepare 100 mM potassium phosphate buffer (K-buffer) with potassium dihydrogen phosphate and dipotassium hydrogen phosphate, and adjust the pH to 7.4.

[0594] Prepare 400× the test compound and reference inhibitor: Dissolve 8 μL of a 10 mM stock solution of the test compound in 12 μL of acetonitrile. Prepare a mixed solution of CYP1A2, CYP2C9, and CYP2D6 inhibitors: Combine 12 μL of 1 mM α-naphthoflavone, 10 μL of 40 mM sulfaphenazole, 10 μL of 10 mM quinidine, and 8 μL of DMSO. Prepare a solution of CYP3A4 and CYP2C19 inhibitors: Dissolve 8 μL of DMSO in 12 μL of acetonitrile.

[0595] Prepare 4× NADPH potassium phosphate solution: Add 66.7 mg NADPH to 10 mL 0.1 M K-buffer, pH 7.4. Prepare 4× substrate potassium phosphate solution: Add different substrates to the required concentration in 10 mL 0.1 M K-buffer to make a solution 4 times the required concentration for the assay.

[0596] Prepare 0.2 mg / mL human liver microsome (HLM) solution: add 10 μL of 20 mg / mL human liver microsome to 990 μL K-buffer and store on ice until use.

[0597] Add 600 μL of 0.2 mg / mL HLM to a 96-well plate, followed by 3 μL of a 400-fold dilution of the test compound solution. Add 200 μL of 0.2 mg / mL HLM to the 96-well plate, followed by 1 μL of the diluted positive control inhibitor solution. Aliquot 30 μL of the compound-human liver microsome mixture into a 96-well plate, then add 15 μL of substrate solution. Preheat the above solution and the prepared NADPH solution at 37°C for 5 minutes. Add 15 μL of the preheated NADPH solution to the reaction plate, mix thoroughly, and start the reaction. Incubate the reaction plate at 37°C. Allow 3A4 to react for 5 minutes; 1A2, 2C9, and 2D6 for 10 minutes; and 2C19 for 45 minutes. At the end of the reaction, terminate the reaction by adding 120 μL of acetonitrile containing an internal standard. Vortex the sample for 10 minutes, centrifuge at 5594 g for 15 minutes, and prepare the sample for LC-MS / MS analysis.

[0598]

Experimental results

[0599] The experimental results show (Table 5) that, at the tested concentrations, the representative compounds of the present invention had no significant inhibitory effect on key CYP isoforms of drug metabolism compared with AMG510 and reference compound A, demonstrating better drug-drug interaction safety.

[0600] Table 5

[0601]

[0602] Example 5: Proliferation inhibition effect of the compounds of the present invention on a series of KRas mutant cells

[0603] This experiment used the CellTiter-Glo (CTG) kit from Promega to evaluate the anti-proliferative activity of the representative compounds of the present invention against 12 KRas mutant tumor cell lines.

[0604] Experimental Materials: RPMI1640 culture medium (Hyclone, catalog number SH30809.01); fetal bovine serum (FBS) (Gibco, catalog number 10099-141); phosphate-buffered saline (PBS) (Solarbio, catalog number P1020-500); DMSO (Sigma, catalog number D8418-1L); CTG detection kit (Promega, catalog number G7573); 96-well cell culture plates (Thermo, catalog number 165305); plate shaker (QILINBEIER, catalog number QB-9001); cell culture incubator (Thermo Scientific, catalog number Model 3100 Series); microscope (OLYMPUS, catalog number CKX41SF); multi-function microplate reader (BMG LABTECH, catalog number Plus); biological safety cabinet (Thermo, Model 1300 Series A2). All cell lines used in the following experiments were purchased from Kangyuan Bochuang Biotechnology (Beijing) Co., Ltd.

[0605]

Experimental method

[0606] cell lines Mutation type Organization Source Growth characteristics Complete medium H23 KRas-G12C lung Adherence RPMI-1640 + 10% FBS H1373 KRas-G12C lung Adherence RPMI-1640 + 10% FBS MIA-Paca-2 KRas-G12C pancreas Adherence DMEM + 10% FBS SW837 KRas-G12C Colorectal Adherence RPMI-1640 + 10% FBS Kyse-410 KRas-G12C esophagus Adherence RPMI-1640 + 10% FBS HCT116 KRas-G13D Colorectal Adherence RPMI-1640 + 10% FBS T84 KRas-G13D Colorectal Adherence DMEM + 10% FBS A549 KRasG12S lung Adherence DMEM + 10% FBS

[0607] Each of the above cell lines was cultured in the indicated complete medium at 37°C, 5% CO2. Cells in the logarithmic growth phase were harvested and counted using a platelet counter. Cell viability was assessed by trypan blue exclusion to ensure viability was above 90%. Cell density was adjusted with complete medium and then seeded into 96-well cell culture plates, with 90 μL per well for a total of 3,000 cells. The cells in the 96-well plates were incubated at 37°C, 5% CO2.

[0608] Prepare a 10x solution of the test compound in culture medium, with the highest concentration tested being 10 μM. Nine concentrations were diluted 3.16-fold. Transfer 10 μL of each dilution to the corresponding wells of a 96-well plate, with triplicate wells for each concentration. Incubate the cells in the treated 96-well plate at 37°C, 5% CO₂ for 72 hours before performing CTG analysis.

[0609] Thaw the CTG reagent and equilibrate the cell plate to room temperature for 30 minutes. Add 100 μL of CTG solution to each well. Shake on an orbital shaker for 5 minutes to lyse the cells. Place the cell plate at room temperature for 20 minutes to stabilize the luminescence signal. Read the luminescence value and collect the data. Analyze the data using GraphPad Prism 7.0 software. Fit the data using nonlinear S-curve regression to generate dose-effect curves. Calculate the relative and absolute IC values according to conventional methods known to those skilled in the art. 50 The maximum inhibition rate was %.

[0610] Inhibition rate % = [1-(Lum 待测药 -Lum 培养液对照 ) / (Lum 细胞对照 -Lum 培养液对照 )】×100%

[0611]

Experimental results

[0612] Table 6.

[0613]

[0614]

[0615] The experimental results in Table 6 show that the representative compounds of the present invention exhibited superior anti-proliferative activity to that of the reference compound A in a series of KRas tumor cells and had good selectivity.

[0616] Example 6: Inhibitory effect of the compounds of the present invention on KRas G12C inhibitor-resistant model cell proliferation

[0617] Acquired drug resistance is one of the key factors affecting the therapeutic effect of KRas G12C inhibitors. This experiment evaluated the potential application value of the compounds of the present invention in known drug resistance models by measuring them in the acquired resistance model of the KRas G12C inhibitor Adagrasib (Engl J Med 2021; 384: 2382-93.).

[0618] This experiment used the CellTiter-Glo (CTG) kit provided by Promega to evaluate the effects of the test compounds on cell proliferation of five KRas cell lines (with acquired resistance to the KRas G12C inhibitor Adagrasib), using reference compound A and AMG510 as control compounds.

[0619] [Experimental Materials]: The experimental materials and instruments used are the same as those listed in Example 5. The various cell lines used in this experiment were obtained from Kangyuan Bochuang Biotechnology (Beijing) Co., Ltd.

[0620]

Experimental method

[0621] cell lines Cell line type Growth characteristics Complete medium BaF3-KRas-G12C-Y96C Mouse primary B cells Suspension RPMI-1640 + 10% FBS BaF3-KRas-G12C-Y96D Mouse primary B cells Suspension RPMI-1640 + 10% FBS BaF3-KRas-G12C-R68S Mouse primary B cells Suspension RPMI-1640 + 10% FBS BaF3-KRas-G12C-H95Q Mouse primary B cells Suspension RPMI-1640 + 10% FBS BaF3-KRas-G12C-H95D Mouse primary B cells Suspension RPMI-1640 + 10% FBS

[0622] The same method and conditions as in Example 5 were used to investigate the inhibitory activity of the test compounds on cell growth. The results are shown in Table 7.

[0623] Table 7.

[0624]

[0625]

[0626] The results showed that the representative compounds of the present invention exhibited superior inhibitory activity to reference compound A and AMG510 in the above-mentioned drug-resistant cell model.

[0627] Example 7: In vivo pharmacodynamic study of the compounds of the present invention on a BALB / c nude mouse model of subcutaneous xenografts of human pancreatic cancer Mia PaCa-2 cells

[0628] The in vivo efficacy of representative compounds of the present invention was evaluated in a subcutaneous xenograft tumor model of human pancreatic cancer Mia PaCa-2 cells.

[0629] Experimental Materials: BALB / c nude mice, 6-8 weeks old, female, purchased from Zhejiang Weitonglihua Laboratory Animal Technology Co., Ltd.; human pancreatic cancer MiaPaCa-2 cells (ATCC, Catalog No. CRL-1420); Matrigel (Corning, Catalog No. 356234).

[0630]

Experimental method

[0631] Tumor volume was calculated according to the following formula: V = 0.5a × b 2 , a and b represent the long diameter and short diameter of the tumor, respectively.

[0632] The tumor inhibition efficacy TGI (%) of the test compound was calculated according to the following formula: TGI (%) = [1-(average tumor volume of the treated group at the end of drug administration-average tumor volume of the treated group at the beginning of drug administration) / (average tumor volume of the solvent control group at the end of treatment-average tumor volume of the solvent control group at the beginning of treatment)] × 100%.

[0633]

Experimental results

[0634] Table 8.

[0635] Group <![CDATA[Tumor volume (mm 3 )(Day 21)]]> TGI (%) Solvent control group 1202 / Example 9 (30 mpk) 162 97.7％ Example 9 (10 mpk) 246 89.8％ Example 9 (3 mpk) 665 50.4％ AMG 510 (10 mpg) 432 72.3％

Claims

1. A compound of formula (I), wherein, A is selected from C-R a , where R a is selected from halogen; R 1 , R 2 and R 3 are each independently selected from H; R b independently selected from H at each occurrence; -X-R 4 selected from -O-C 1-6 alkyl -phenyl, substituted by -(CR c R c ) 0-6 -N(R c )2, where R c are each independently H or C 1-6 alkyl; and -O- A 4- to 6-membered heterocycloalkyl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, wherein said heterocycloalkyl is substituted by 1, 2 or 3 groups independently selected from -C 1-6 alkyl and -(CR c R c ) 0-6 -OR c , where R c is selected from H or -C 1-6 alkyl, and -A 5- or 6-membered heteroaryl containing 1, 2 or 3 heteroatoms independently selected from N, O or S, substituted by 1, 2 or 3 -C 1-6 alkyl groups; E is selected from halogen; R 5 For or a pharmaceutically acceptable salt thereof.

2. The compound of formula (I) according to claim 1 or a pharmaceutically acceptable salt thereof, wherein A is C-R a , wherein R a is Cl.

3. The compound of formula (I) or a pharmaceutically acceptable salt according to any one of claims 1-2, wherein -X-R 4 is -O-C 1-6 alkyl.

4. The compound of formula (I) or a pharmaceutically acceptable salt according to claim 3, wherein -X-R 4 is -O-CH3.

5. The compound of formula (I) or a pharmaceutically acceptable salt according to any one of claims 1-2, wherein -X-R 4 is selected from 6. The compound of formula (I) or a pharmaceutically acceptable salt according to any one of claims 1-2, wherein -X-R 4 The 4- to 6-membered heterocycloalkyl in 7. A compound of formula (I) or a pharmaceutically acceptable salt according to any one of claims 1-2, wherein -X-R 4 is selected from 8. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to any one of claims 1-2, wherein E is F.

9. A compound selected from or a pharmaceutically acceptable salt thereof.

10. A pharmaceutical composition comprising the compound according to any one of claims 1-9 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

11. Use of the compound according to any one of claims 1-9 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition according to claim 10 in the manufacture of a medicament for preventing or treating a disease mediated by KRas G12C mutation, wherein the disease is selected from lung cancer, colon cancer, rectal cancer, pancreatic cancer, cholangiocarcinoma, esophageal cancer.

Images