Benzene ring derivative, and preparation method therefor and use thereof in medicine

Abstract

Disclosed are a compound as represented by formula (I) and / or a stereoisomer, tautomer or pharmaceutically acceptable salt or solvate thereof, a preparation method therefor, a pharmaceutical composition comprising the compound of formula (I), and a use thereof as a therapeutic agent in medicine, in particular as a KIF18A inhibitor in the treatment of disorders and / or diseases mediated by KIF18A.

Description

A benzene ring derivative, its preparation method and its application in medicine Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to a benzene ring derivative, a preparation method thereof and its application in medicine. Background Art

[0002] Tumors are multifactorial diseases characterized by the presence of cells with complex nuclear heterogeneity. In 1914, German scientist Theodor Boveri proposed that genomic instability is the cause of tumors. One of the most common forms of genomic instability in tumors is chromosome instability (CIN), a persistent state of mitotic dysfunction that leads to karyotype abnormalities and aneuploidy (Bielski et al., NATURE COMMUNICATIONS, 2021, 12:3663). At the cellular level, CIN promotes heterogeneity within tumors and allows tumors to explore a larger evolutionary space to obtain more adaptable subclones that can continue to expand under selective pressure. CIN also causes the release of genomic DNA into the cytoplasm, triggering an immune response and activating inflammatory pathways that allow tumor cells to spread metastatically (Bakhoum et al., Nature, 2018, 553, 467–472).

[0003] KIF18A is a member of the kinesin-8 family of motor proteins that plays an important role in various cellular processes, including motility, cell division, microtubule dynamics, and organelle transport (Jordan et al., Nat Rev Cancer, 2004(4):253-265; Gardner et al., Trends Cell Biol, 2008(18):307-310; Du et al. Curr Biol 2021(20),374-380; Weaver et al., Curr Biol 2011(21),1500-1506). KIF18A can rely on the energy released by hydrolyzing ATP in the cell to move toward the positive pole using microtubules as a track. At the same time, during mitosis, KIF18A is localized at the positive end of microtubules, which can regulate the dynamic instability of microtubules and promote microtubule depolymerization (Mayr et al., Curr Biol, 2007(17),488-498). KIF18A weakens chromosome oscillations by depolymerizing microtubules during cell division, thereby enhancing chromosome aggregation. Therefore, during mitosis, KIF18A can regulate spindle microtubule dynamics and chromosome amplitude, and has a key role in timely completing the whole array, maintaining genome stability and successfully completing mitosis in the mitotic phase chromosome. The abnormality of KIF18A function can affect chromosome separation and cause CIN, thereby leading to the occurrence of tumors (Hartwell et al., Science 1994 (266), 1821-1828). Existing studies have shown that KIF18A is a non-essential gene for normal diploid cells, such as KIF18A knockout mice that can survive to adulthood. However, in tumor cells, KIF18A can be overexpressed and its overexpression is associated with poor prognosis in tumor patients. Therefore, knocking out KIF18A can reduce the viability of tumor cells without affecting normal cell viability (Quinton et al., Nature, 2021, 590 (7846): 492-497). Cancer types in which KIF18A is overexpressed include, but are not limited to, esophageal cancer, liver cancer, colorectal cancer, breast cancer, lung cancer, pancreatic cancer, prostate cancer, bladder cancer, head cancer, neck cancer, cervical cancer, and ovarian cancer. These studies have fully confirmed that the KIF18A gene can be an important anti-tumor drug target.

[0004] However, there are currently no approved drugs targeting KIF18A inhibitors, which remains an unmet clinical need in the field of oncology. Therefore, there is still a need to develop safe and effective KIF18A inhibitors to better meet the clinical needs of patients.

[0005] Summary of the Invention

[0006] The first aspect of the present invention provides a novel benzene ring derivative represented by general formula (I), or a stereoisomer, tautomer, pharmaceutically acceptable salt, or deuterated derivative thereof. The compound of general formula (I) can be used as a KIF18A inhibitor for treating KIF18A-mediated conditions and / or diseases, including cancer, inflammation, or ciliary pathology.

[0007] The compounds provided by the present invention have microtubule-based KIF18A regulatory activity, in particular, KIF18A inhibitory activity. To this end, the second aspect of the present invention provides pharmaceutical compositions comprising compounds of Formula I and stereoisomers, tautomers, pharmaceutically acceptable salts, or deuterated derivatives thereof for the preparation and manufacture of drugs for therapeutic, preventive, acute or chronic treatment of KIF18A-mediated diseases and disorders (including but not limited to cancer). Therefore, the compounds of the present invention can be used to manufacture anticancer drugs. The present invention also provides methods for preparing compounds of Formula I, and intermediates useful in such methods.

[0008] The present invention discloses a compound represented by general formula (I) or its stereoisomers, tautomers, pharmaceutically acceptable salts or deuterated derivatives:

[0009] in:

[0010] X 1 C or N;

[0011] X 3 C or N;

[0012] X 2 、X 4 and X 5 are each independently selected from N, O, NH or CH;

[0013] W 1 N or CR 2 ;

[0014] R 1 is a 6-membered heterocycloalkyl group containing 0, 1 or 2 N atoms and 0 or 1 O atoms, and further, the 6-membered heterocycloalkyl group is substituted by 0, 1, 2 or 3 groups selected from the following: F, Cl, Br, C 1-3 alkyl;

[0015] R 2 For hydrogen, halogen, CN, C 1-8 Alkyl, C 1-4 Halogenated alkyl, C 0-4 Alkyl-OR a 、C 0-4 Alkyl-NR a Ra ;

[0016] R 3 For hydrogen, C 1-4 Alkyl, C 1-4 Halogenated alkyl, C 0-4 Alkyl-OR a 、C 0-4 Alkyl-NR a R a ;

[0017] R 4 and R 5 are independently selected from hydrogen or C 1-3 Alkyl; or R 4 、R 5 Connected to form a cyclopropyl group; or R 4 、R 5 The carbon atom to which it is connected forms C=CR j R k ; Among them, R j and R k are each independently selected from the group consisting of: hydrogen, halogen;

[0018] When W 1 When N, R 6 is -OCH2OP(=O)(OH)2, -OP(=O)(OH)2, -OP(=O)(OH)C 1-5 Alkyl, -OC(=O)C 1-5 Alkyl, wherein the C 1-5 The alkyl group is substituted with 0, 1, 2 or 3 substituents selected from the group consisting of OH, NH2, OCH3, or a 5-, 6- or 7-membered heterocycloalkyl group containing 0, 1 or 2 N atoms and 0 or 1 O atom;

[0019] When W 1 CR 2 When R 6 is -OH, -OCH2OP(=O)(OH)2, -OP(=O)(OH)2, -OP(=O)(OH)C 1-5 Alkyl, -OC(=O)C 1-5 Alkyl, wherein the C 1-5 The alkyl group is substituted with 0, 1, 2 or 3 substituents selected from the group consisting of OH, NH2, OCH3, or a 5-, 6- or 7-membered heterocycloalkyl group containing 0, 1 or 2 N atoms and 0 or 1 O atom;

[0020] R a is independently H or R b ; and R b In each case, independently C1-6 Alkyl, wherein the C 1-6 The alkyl group is substituted with 0, 1, 2 or 3 substituents selected from the group consisting of halogen, -OH, -OC 1-4 Alkyl, -NH2, -NHC 1-4 Alkyl and -N(C 1-4 Alkyl)C 1-4 alkyl.

[0021] The present invention also provides preferred embodiments related to the compounds described by general formula (I).

[0022] In one or more embodiments, the W 1 N, R 6 -OP(=O)(OH)2 or

[0023] In one or more embodiments, the W 1 CR 2 , where R 2 -OCH3; R 6 -OH, -OP(=O)(OH)2 or

[0024] In one or more embodiments, the R 6 It is -OP(=O)(OH)2.

[0025] In one or more embodiments, the group Selected from Among them, the side with * is the side connected to the benzene ring.

[0026] In one or more embodiments, the group Selected from Among them, the side with * is the side connected to the benzene ring.

[0027] In one or more embodiments, the group for

[0028] In one or more embodiments, the R 1 Selected from

[0029] In one or more embodiments, the R 1 for

[0030] In one or more embodiments, the R 3 Selected from hydrogen,

[0031] In one or more embodiments, the R 3 For hydrogen.

[0032] Typical compounds of the present invention include, but are not limited to:

[0033] or its stereoisomers, tautomers, any pharmaceutically acceptable salts or deuterated derivatives.

[0034] One or more embodiments of the present invention provide a pharmaceutical composition comprising an effective dose of a compound of Formula I or its stereoisomers, tautomers, pharmaceutically acceptable salts or deuterated derivatives, and a pharmaceutically acceptable carrier, excipient or a combination thereof.

[0035] One or more embodiments of the present invention provide a method for treating a condition treatable with a KIF18A inhibitor, comprising administering to a patient in need thereof a therapeutically effective amount of a compound of Formula I or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a pharmaceutical combination comprising a compound of Formula I.

[0036] In one or more embodiments, the condition is a cancer selected from the group consisting of: (a) a solid tumor or a hematogenic tumor selected from the group consisting of bladder cancer, endometrial cancer, squamous cell lung cancer, breast cancer, colon cancer, kidney cancer, liver cancer, lung cancer, small cell lung cancer, esophageal cancer, gallbladder cancer, brain cancer, head and neck cancer, ovarian cancer, pancreatic cancer, gastric cancer, cervical cancer, thyroid cancer, prostate cancer, and skin cancer; (b) a hematopoietic tumor of the lymphoid lineage selected from the group consisting of leukemia, acute lymphoblastic leukemia, acute lymphoblastic leukemia, B-cell lymphoma, T-cell lymphoma, Hodgkin lymphoma, non-Hodgkin lymphoma, hairy cell lymphoma, and Burkett lymphoma; (c) a hematopoietic neoplasm of the myeloid lineage selected from the group consisting of acute and chronic myeloid leukemias, myelodysplastic syndromes, and promyelocytic leukemias; (d) a tumor of mesenchymal origin selected from the group consisting of fibrosarcoma and rhabdomyosarcoma; (e) a tumor of the central and peripheral nervous system selected from the group consisting of astrocytoma, neuroblastoma, glioma, and schwannoma; or (f) a melanoma, a seminoma, a teratoma, an osteosarcoma, xeroderma pigmentosum, a keratoacanthoma, a follicular thyroid carcinoma, or a Kaposi's sarcoma.

[0037] One or more embodiments of the present invention provide a method for reducing the size of a solid tumor in a subject, the method comprising administering to a subject in need thereof a therapeutically effective amount of a compound of Formula I or its stereoisomers, tautomers, pharmaceutically acceptable salts or deuterated derivatives, or a pharmaceutical composition comprising a compound of Formula I.

[0038] One or more embodiments of the present invention provide a method for treating a cell proliferative disorder in a subject, the method comprising administering to a subject in need thereof a therapeutically effective amount of a compound of Formula I or its stereoisomers, tautomers, pharmaceutically acceptable salts or deuterated derivatives, or a pharmaceutical composition comprising a compound of Formula I.

[0039] One or more embodiments of the present invention provide a method for inhibiting KIF18A in a cell, comprising contacting the cell with a compound of Formula I or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a pharmaceutical composition comprising a compound of Formula I.

[0040] One or more embodiments of the present invention provide the use of a compound of Formula I or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a pharmaceutical composition containing a compound of Formula I in the preparation of a medicament for a condition treatable with a KIF18A inhibitor.

[0041] One or more embodiments of the present invention provide a use of a compound of Formula I or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a pharmaceutical composition containing a compound of Formula I in the preparation of a medicament for reducing the size of a solid tumor in a subject.

[0042] One or more embodiments of the present invention provide the use of a compound of Formula I or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a pharmaceutical composition containing a compound of Formula I in the preparation of a medicament for treating a cell proliferative disorder in a subject.

[0043] One or more embodiments of the present invention provide the use of a compound of Formula I or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a pharmaceutical composition containing the compound of Formula I in the preparation of a medicament for inhibiting KIF18A in a cell. DETAILED DESCRIPTION

[0044] definition

[0045] Unless otherwise stated, some of the terms used in the specification and claims of the present invention are defined as follows:

[0046] A dash ("-") that is not between two letters or symbols is used to indicate the point of attachment of a substituent. For example, -CONR a R b Attached through carbon atoms.

[0047] The carbon, hydrogen, oxygen, sulfur, nitrogen or F, Cl, Br, I involved in the groups and compounds of the present invention include their isotopes, and the carbon, hydrogen, oxygen, sulfur or nitrogen involved in the groups and compounds of the present invention are optionally further replaced by one or more of their corresponding isotopes, wherein the isotopes of carbon include 12 C. 13 C and 14 C, hydrogen isotopes include protium (H), deuterium (D, also called heavy hydrogen), tritium (T, also called super tritium), oxygen isotopes include 16 O. 17 O and 18 O, sulfur isotopes include 32 S. 33 S. 34 S and 36 S, nitrogen isotopes include 14 N and 15 N, fluorine isotopes include 17 F and 19 F, chlorine isotopes include 35 Cl and 37 Isotopes of Cl, bromine include 79 Br and 81 Br.

[0048] As used herein, the term "stereoisomer" refers to any of the various stereoisomeric configurations that may exist for a given compound of the present disclosure, and includes enantiomers, diastereomers, conformers (rotamers), geometric isomers (cis / trans) isomers, atropisomers, and the like. "Enantiomers" refers to two non-superimposable isomers of a compound that are mirror images of each other. "Diastereomers" refers to stereoisomers that have two or more chiral centers and whose molecules are not mirror images of each other. Diastereomers have different physical properties, such as melting points, boiling points, spectral properties, and reactivity. Mixtures of diastereoisomers can be separated by high-resolution analytical procedures such as electrophoresis and chromatography, for example, HPLC. The stereochemical definitions and rules used herein generally follow those of SP 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. Any asymmetric atom (e.g., carbon, etc.) in the compounds disclosed herein may exist in racemic or enantiomerically enriched form, such as (R)-, (S)-, or (R,S)-configuration. In certain embodiments, each asymmetric atom has at least 50% enantiomeric excess, at least 60% enantiomeric excess, at least 70% enantiomeric excess, at least 80% enantiomeric excess, at least 90% enantiomeric excess, at least 95% enantiomeric excess, or at least 99% enantiomeric excess in terms of the (R)- or (S)-configuration. Any resulting mixture of stereoisomers can be separated into the pure or substantially pure geometric isomers, enantiomers, and diastereomers on the basis of the differences in the constituent physicochemical properties, for example, by chromatography and / or fractional crystallization.

[0049] The term "tautomer" herein refers to structural isomers with different energies that can be converted to each other through a low energy barrier. If tautomerism is possible (such as in a solution), the chemical equilibrium of the tautomer can be reached. For example, proton tautomers (protontautomers) (also referred to as prototropic tautomers) include interconversions carried out by proton migration, such as keto-enol isomerization and imine-enamine isomerization. Valence tautomers (valence tautomers) include interconversions carried out by the reorganization of some bonding electrons. The specific example of keto-enol tautomerism is the interconversion of pentane-2,4-dione and 4-hydroxypent-3-ene-2-one tautomers. Another example of tautomerism is phenol-ketone tautomerism. A specific example of phenol-ketone tautomerism is the interconversion of pyridine-4-ol and pyridine-4 (1H)-one tautomers. Unless otherwise indicated, all tautomeric forms of the compounds of the invention are within the scope of the invention.

[0050] "Alkyl" refers to a linear or branched saturated aliphatic hydrocarbon group of 1 to 20 carbon atoms, preferably an alkyl group of 1 to 8 (e.g., 1, 2, 3, 4, 5, 6, 7, 8) carbon atoms, more preferably an alkyl group of 1 to 6 carbon atoms, and even more preferably an alkyl group of 1 to 4 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, neobutyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, and various branched chain isomers thereof.

[0051] "Heterocyclyl," "heterocycle," or "heterocyclic" are used interchangeably herein and refer to non-aromatic heterocyclic groups in which one or more of the ring atoms is a heteroatom, such as oxygen, nitrogen, or sulfur, and include monocyclic, fused, bridged, and spirocyclic rings. Preferably, the heterocyclic group has a 5- to 7-membered monocyclic ring or a 7- to 10-membered bi- or tricyclic ring, which may contain 1, 2, or 3 atoms selected from nitrogen, oxygen, and / or sulfur. Examples of "heterocyclyl" include, but are not limited to, morpholinyl, oxetanyl, thiomorpholinyl, tetrahydropyranyl, 1,1-dioxo-thiomorpholinyl, piperidinyl, 2-oxo-piperidinyl, pyrrolidinyl, 2-oxo-pyrrolidinyl, piperazin-2-one, 8-oxa-3-aza-bicyclo[3.2.1]octyl, and piperazinyl. Heterocyclyl groups may be substituted or unsubstituted.

[0052] "Aryl" refers to a substituted or unsubstituted aromatic ring, which can be a 5-8 membered (e.g., 5, 6, 7, 8 membered) monocyclic ring, a 5-12 membered (e.g., 5, 6, 7, 8, 9, 10, 11, 12 membered) bicyclic ring, or a 10-15 membered (e.g., 10, 11, 12, 13, 14, 15 membered) tricyclic ring system, which can be a bridged ring or a spirocyclic ring, non-limiting examples of which include phenyl and naphthyl. The aryl group may optionally be further substituted with one or more substituents.

[0053] "Heteroaryl" refers to a substituted or unsubstituted aromatic ring, which can be a 3-8 membered (e.g., 3, 4, 5, 6, 7, 8 membered) monocyclic ring, a 5-12 membered (e.g., 5, 6, 7, 8, 9, 10, 11, 12 membered) bicyclic ring or a 10-15 membered (e.g., 10, 11, 12, 13, 14, 15 membered) tricyclic ring system, and contains 1 to 6 (e.g., 1, 2, 3, 4, 5, 6) heteroatoms selected from N, O or S, preferably a 5- to 8-membered heteroaryl group, and the 1 to 4 (e.g., 1, 2, 3, 4) N and S optionally substituted in the heteroaryl ring can be oxidized to various oxidation states. The heteroaryl group may be attached to a heteroatom or a carbon atom, and may be a bridged ring or a spirocyclic ring. Non-limiting examples include cyclic pyridyl, furyl, thienyl, pyranyl, pyrrolyl, pyrimidinyl, pyrazinyl, pyridazinyl, imidazolyl, piperidinylbenzimidazolyl, benzopyridinyl, and pyrrolopyridinyl. The heteroaryl group may be further substituted with one or more substituents.

[0054] "Bicyclic" refers to a group having two connected rings. Bicyclic can be a carbocyclic ring (all ring atoms are carbon atoms) or a heterocyclic ring (in addition to carbon atoms, the ring atoms include, for example, 1, 2 or 3 heteroatoms, such as N, O or S). Both rings can be aliphatic (e.g., decalin and norbornane), or can be aromatic (e.g., naphthalene), or a combination of aliphatic and aromatic (e.g., tetralin). Bicyclic includes (a) spirocyclic compounds, in which the two rings share only one single atom (spiro atom, which is typically a quaternary carbon). Examples of spirocyclic compounds include, but are not limited to:

[0055] (b) Fused bicyclic compounds, where the two rings share two adjacent atoms. In other words, the rings share a covalent bond, i.e., the bridgehead atoms are directly connected (e.g., α-thujene and decalin). Examples of fused bicyclic rings include, but are not limited to:

[0056] and (c) bridged bicyclic compounds in which the two rings share three or more atoms and the two bridgehead atoms are separated by a bridge comprising at least one atom. For example, norbornane, also known as bicyclo[2.2.1]heptane, can be considered a pair of cyclopentane rings, each sharing three of their five carbon atoms. Examples of bridged bicyclic rings include, but are not limited to:

[0057] "Monocyclic" refers to a single carbon ring contained by itself or in combination with other terms. "Carbocyclic" refers to a saturated or unsaturated aromatic or non-aromatic ring. When it is an aromatic ring, its definition is the same as the definition of "aryl" above; when it is a non-aromatic ring, it can be a 3-10 membered (e.g., 3, 4, 5, 6, 7, 8, 9, 10 membered) monocyclic ring, a 4-12 membered (e.g., 4, 5, 6, 7, 8, 9, 10, 11, 12 membered) bicyclic ring or a 10-15 membered (e.g., 10, 11, 12, 13, 14, 15 membered) tricyclic ring system, which can be a bridged ring or a spirocyclic ring, without limitation. Examples include cyclopropyl, cyclobutyl, cyclopentyl, 1-cyclopentyl-1-enyl, 1-cyclopentyl-2-enyl, 1-cyclopentyl-3-enyl, cyclohexyl, 1-cyclohexyl-2-enyl, 1-cyclohexyl-3-enyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cyclooctyl, cyclononyl, cyclodecyl, cycloundecyl, cyclododecyl, and the "carbocyclyl" or "carbocycle" is optionally further substituted by one or more substituents.

[0058] "Cycloalkyl" refers to a saturated cyclic hydrocarbon group, which can be a 3- to 10-membered (e.g., 3, 4, 5, 6, 7, 8, 9, 10-membered) monocyclic ring, a 4- to 12-membered (e.g., 4, 5, 6, 7, 8, 9, 10, 11, 12-membered) bicyclic ring, or a 10- to 20-membered (e.g., 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20-membered) polycyclic ring system, preferably having 3 to 10 carbon atoms, more preferably 3 to 8 carbon atoms. Non-limiting examples of "cycloalkyl" include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, 1,5-cyclooctadienyl, 1,4-cyclohexadienyl, and cycloheptatrienyl. When a cycloalkyl group is substituted, it may be optionally further substituted with one or more substituents.

[0059] "Heterocycloalkyl" refers to a substituted or unsubstituted saturated non-aromatic ring group, which can be a 3-8 membered (e.g., 3, 4, 5, 6, 7, 8 membered) monocyclic ring, a 4-12 membered (e.g., 4, 5, 6, 7, 8, 9, 10, 11, 12 membered) bicyclic ring, or a 10-15 membered (e.g., 10, 11, 12, 13, 14, 15 membered) tricyclic ring system, and contains 1, 2, or 3 heteroatoms selected from N, O, or S, preferably a 3- to 8-membered heterocyclic ring. The 1, 2, or 3 N or S atoms optionally substituted in the "heterocycloalkyl" ring can be oxidized to various oxidation states; the "heterocycloalkyl" can be attached to a heteroatom or a carbon atom; the "heterocycloalkyl" can be a bridged ring or a spirocyclic ring. Non-limiting examples of “heterocycloalkyl” include oxiranyl, aziridinyl, oxetanyl, azetidinyl, 1,3-dioxolanyl, 1,4-dioxolanyl, 1,3-dioxanyl, azepanyl, piperidinyl, piperidinyl, morpholinyl, thiomorpholinyl, 1,3-dithianyl, tetrahydrofuranyl, tetrahydropyrrolyl, tetrahydroimidazolyl, tetrahydrothiazolyl, tetrahydropyranyl, azabicyclo[3.2.1]octanyl, azabicyclo[5.2.0]nonanyl, oxatricyclo[5.3.1.1]dodecyl, azaadamantyl, and oxaspiro[3.3]heptanyl.

[0060] " alkoxy " refers to that at least 1 carbon atom in the alkyl is replaced by the formed group of oxygen atom. Non-limiting examples include methoxyl group, ethoxyl group, n-propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy, n-pentyloxy, n-hexyloxy, cyclopropyloxy and cyclobutyloxy. Described alkyl definition is identical with " alkyl " definition mentioned above. Unless clearly stated, otherwise alkoxy group can alternatively be substituted by one or more substituents (as one, two or three substituents, or 1-4 substituents, or up to the quantity of the hydrogen present in the unsubstituted alkoxy group) substituted alkoxy hydrogen atom part. Unless otherwise specified, otherwise applicable substituent is selected from the substituent for alkyl group as listed above, except that hydroxyl and amino are usually not present on the carbon directly connected with the oxygen of substituted alkyl-O group.

[0061] "Haloalkyl" refers to a hydrocarbon group as defined herein that is substituted by one or more halogen groups as defined herein. Unless otherwise specified, the hydrocarbon moiety of a haloalkyl group includes 1-4 carbon atoms. A haloalkyl group can be a monohaloalkyl, a dihaloalkyl, a trihaloalkyl, or a polyhaloalkyl (including a perhaloalkyl group). A monohaloalkyl group can have one iodine, bromine, chlorine, or fluorine within the hydrocarbon group. A dihaloalkyl group and a polyhaloalkyl group can have two or more identical halogen atoms or a combination of different halogen groups within the hydrocarbon group. Polyhaloalkyl groups include, for example, up to 6, or 4, or 3, or 2 halogen groups. Examples of haloalkyl groups include, but are not limited to, fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl, pentafluoroethyl, heptafluoropropyl, difluorochloromethyl, dichlorofluoromethyl, difluoroethyl, difluoropropyl, dichloroethyl, and dichloropropyl. A perhaloalkyl group refers to a hydrocarbon group (e.g., trifluoromethyl) in which all hydrogen atoms are replaced by halogen atoms. In some embodiments, unless otherwise specified, haloalkyl group comprises monofluoro-, difluoro- and trifluoro-substituted methyl and ethyl groups (for example,-CF ,-CF H,-CFH , and-CH CF ).Unless clearly stated, haloalkyl group can alternatively be substituted by one or more substituents (such as one, two or three substituents, or 1-4 substituents, or up to the quantity of the hydrogen present in the unsubstituted haloalkyl group) replacing the unsubstituted hydrogen atom of haloalkyl.Unless otherwise specified, otherwise applicable substituent is selected from, for example, the substituent for alkyl group as listed above.

[0062] "Haloalkoxy" refers to a haloalkyl-O-, wherein haloalkyl is as defined above. Examples of haloalkoxy include, but are not limited to, fluoromethoxy, difluoromethoxy, trifluoromethoxy, trichloromethoxy, 2-chloroethoxy, 2,2,2-trifluoroethoxy, 1,1,1,3,3,3-hexafluoro-2-propoxy, and the like. In some embodiments, the haloalkoxy group comprises 1-4 carbon atoms and up to three halogen elements, for example, monofluoro, difluoro, and trifluoro substituted methoxy and ethoxy groups. Unless expressly stated, the haloalkoxy group may optionally be substituted by replacing the hydrogen atoms of the unsubstituted alkyl portion of the haloalkoxy group with one or more substituents (such as one, two, or three substituents, or 1-4 substituents, or up to the number of hydrogens present on the unsubstituted haloalkoxy group). Unless otherwise specified, suitable substituents are selected, for example, from the substituents listed above for alkyl groups, except that hydroxy and amino groups are generally not present on the carbon directly attached to the oxygen of a substituted haloalkyl-O group.

[0063] "Hydroxy" refers to an -OH group.

[0064] "Halo" refers to fluorine, chlorine, bromine, and iodine. Groups and moieties substituted with halogen elements, such as alkyl groups substituted with halogen elements (haloalkyl), can be monohalogenated, polyhalogenated, or perhalogenated. In some embodiments, unless otherwise specified, chlorine and fluorine are examples of halogen substituents on alkyl groups or cycloalkyl groups; unless otherwise specified, fluorine, chlorine, and bromine are used, for example, on aryl groups or heteroaryl groups.

[0065] "Substituted" means that one or more hydrogen atoms, preferably up to 5, more preferably 1 to 3 hydrogen atoms, in a group are replaced independently of one another by a corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and a person skilled in the art can determine (by experiment or theory) which substitutions are possible or impossible without undue effort. For example, an amino or hydroxyl group with free hydrogen may be unstable when combined with a carbon atom with an unsaturated (e.g., olefinic) bond.

[0066] When the above-mentioned "alkyl", "haloalkyl", "alkoxy", "bicyclic", "monocyclic", "aryl", "heteroaryl", "heterocyclyl", "heterocycle", "cycloalkyl", "heterocycloalkyl" or "heterocyclyl" is substituted, it may be further substituted by 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 groups selected from F, Cl, Br, I, hydroxyl, mercapto, nitro, cyano, amino, C 1-6 Alkylamino, =O, C 1-6 Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, -NR q4 R q5 、=NR q6 、-C(=O)OC 1-6 Alkyl, -OC(=O)C 1-6 Alkyl, -C(=O)NR q4 R q5 、C 3-8 Cycloalkyl, C 3-8 Heterocycloalkyl, C 6-10 Aryl, C 5-10 Heteroaryl, -C(=O)OC 6-10 Aryl, -OC(=O)C 6-10 Aryl, -OC(=O)C 5-10 Heteroaryl, -C(=O)OC 5-10 Heteroaryl, -OC(=O)C 3-8 Heterocycloalkyl, -C(=O)OC 3-8 Heterocycloalkyl, -OC(=O)C 3-8 Cycloalkyl, -C(=O)OC 3-8 Cycloalkyl, -NHC(=O)C3-8 Heterocycloalkyl, -NHC(=O)C 6-10 Aryl, -NHC(=O)C 5-10 Heteroaryl, -NHC(=O)C 3-8 Cycloalkyl, -NHC(=O)C 3-8 Heterocycloalkyl, -NHC(=O)C 2-6 Alkenyl or -NHC(=O)C 2-6 substituted by a substituent of an alkynyl group, wherein the substituent C 1-6 Alkyl, C 1-6 Alkoxy, C 2-6 Alkenyl, C 2-6 Alkynyl, C 3-8 Cycloalkyl, C 3-8 Heterocycloalkyl, C 6-10 Aryl, C 5-10 Heteroaryl, -NHC(=O)C 6-10 Aryl, -NHC(=O)C 5-10 Heteroaryl, -NHC(=O)C 3-8 Heterocycloalkyl or -NHC(=O)C 3-8 The cycloalkyl group is optionally further substituted by 1 to 3 groups selected from OH, F, Cl, Br, I, C 1-6 Alkyl, C 1-6 Alkoxy, -NR q4 R q5 Or substituted by a substituent of =O; R q1 Selected from C 1-6 Alkyl, C 1-6 Alkoxy or C 6-10 Aryl; R q2 、R q3 Selected from H or C 1-6 Alkyl; wherein R q4 、R q5 Selected from H, C 1-6 Alkyl, -NH(C=NR q1 )NR q2 R q3 、-S(=O)2NR q2 R q3 、-C(=O)R q1 or -C(=O)NR q2 R q3 , wherein the C 1-6 The alkyl group is optionally further substituted by one or more groups selected from OH, F, Cl, Br, I, C 1-6 Alkyl, C 1-6 Alkoxy, C 6-10 Aryl, C 5-10 Heteroaryl, C 3-8 Cycloalkyl or C3-8 is substituted by a substituent of a heterocycloalkyl group; or R q4 With R q5 and the N atom form a 3- to 8-membered heterocyclic ring, which may contain one or more heteroatoms selected from N, O or S.

[0067] "Pharmaceutically acceptable salts" refer to salts of the above compounds that retain their original biological activity and are suitable for pharmaceutical use. Pharmaceutically acceptable salts of the compounds represented by formula (I) may be metal salts or amine salts formed with suitable acids.

[0068] A "pharmaceutical composition" refers to a mixture containing one or more compounds described herein, or their physiologically and pharmaceutically acceptable salts or prodrugs, together with other chemical components, and other components such as physiologically and pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to an organism, promote absorption of the active ingredient, and thereby exert its biological activity.

[0069] "Carrier" refers to a material that does not cause significant irritation to an organism and does not abrogate the biological activity and properties of the administered compound.

[0070] "Excipient" refers to an inert substance added to a pharmaceutical composition to facilitate administration of a compound. Non-limiting examples include calcium carbonate, calcium phosphate, sugars, starches, cellulose derivatives (including microcrystalline cellulose), gelatin, vegetable oils, polyethylene glycols, diluents, granulating agents, lubricants, binders, and disintegrants.

[0071] "Prodrugs" refer to compounds of the present invention that can be converted into biologically active compounds through in vivo metabolism. Prodrugs of the present invention are prepared by modifying amino or carboxyl groups in compounds of the present invention. These modifications can be removed by conventional manipulation or in vivo to yield the parent compound. When the prodrugs of the present invention are administered to a mammalian subject, the prodrugs are cleaved to form free amino or carboxyl groups.

[0072] "Optional" or "optionally" or "selectively" or "optionally" means that the subsequently described event or circumstance may but need not occur, and that the description includes instances where the event or circumstance occurs and instances where it does not. For example, "heterocyclyl optionally substituted with alkyl" means that the alkyl group may but need not be present, and that the description includes instances where the heterocyclyl group is substituted with alkyl and instances where the heterocyclyl group is not substituted with alkyl.

[0073] "Treating," "treat," "treatment," or "alleviation" refers to administering at least one compound disclosed herein, and / or at least one stereoisomer thereof (if any), at least one stable isotope thereof, or at least one pharmaceutically acceptable salt thereof, to a subject suffering from, for example, cancer, identified in need thereof.

[0074] The term "effective amount" refers to an amount of at least one compound disclosed herein, and / or at least one stereoisomer thereof (if any), at least one stable isotope thereof, or at least one pharmaceutically acceptable salt thereof, effective for "treating" (as defined above) a disease or disorder in a subject.

[0075] Various embodiments are disclosed herein. It will be appreciated that the features specified in each embodiment can be combined with the features of other specifications to provide additional embodiments of the present disclosure. The following non-limiting examples are representative of the present disclosure.

[0076] Scheme 1 illustrates a general method for preparing compounds of the present disclosure and intermediates. Detailed descriptions and syntheses are disclosed in the following examples. Those skilled in the art will be able to find other synthetic methods or modify the following methods using conventional chemistry to prepare suitable compounds encompassed by Formula I. Therefore, these methods are equally applicable to the preparation of compounds of other embodiments. Although specific starting materials and reagents are depicted and discussed below in the scheme, other starting materials and reagents can be easily replaced to provide various compounds and / or reaction conditions.

[0077] The following examples provide the preparation of representative compounds represented by formula (I) and related structural identification data. It must be noted that the following examples are used to illustrate the present invention rather than to limit the present invention. 1 H NMR spectra were obtained using a Bruker instrument (400 MHz), and chemical shifts are expressed in ppm using tetramethylsilane as an internal standard (0.00 ppm). 1 H NMR notation: s = singlet, d = doublet, t = triplet, m = multiplet, br = broadened, dd = doublet of a doublet, dt = doublet of a triplet. Coupling constants, when given, are given in Hz.

[0078] Mass spectra were obtained using LC / MS, and the ionization method could be ESI or APCI.

[0079] The thin layer chromatography silica gel plate uses Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plate. The specification of the silica gel plate used in thin layer chromatography (TLC) is 0.15mm~0.2mm, and the specification used for thin layer chromatography separation and purification products is 0.4mm~0.5mm.

[0080] Column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh silica gel as the carrier.

[0081] In the following examples, unless otherwise indicated, all temperatures are in degrees Celsius. Unless otherwise indicated, various starting materials and reagents are commercially available or synthesized according to known methods. Commercially available raw materials and reagents are used directly without further purification, unless otherwise indicated. Commercial manufacturers include but are not limited to Aldrich Chemical Company, ABCR GmbH & Co. KG, Acros Organics, Guangzan Chemical Technology Co., Ltd. and Jingyan Chemical Technology Co., Ltd.

[0082] CD3OD: deuterated methanol.

[0083] CDCl3: deuterated chloroform.

[0084] DMSO-d6: deuterated dimethyl sulfoxide.

[0085] Argon atmosphere means that the reaction bottle is connected to an argon balloon with a capacity of about 1 L.

[0086] Unless otherwise specified in the examples, the solution in the reaction refers to an aqueous solution.

[0087] The compound is purified using a silica gel column chromatography eluent system and thin layer chromatography, wherein the eluent system is selected from: A: petroleum ether and ethyl acetate system; B: dichloromethane and methanol system; C: dichloromethane:ethyl acetate; the volume ratio of the solvent varies according to the polarity of the compound and can also be adjusted by adding a small amount of acidic or alkaline reagents, such as acetic acid or triethylamine.

[0088] The following examples illustrate certain embodiments of the present disclosure and how to make and use them. Therefore, the following examples are not intended to limit the scope of the present invention. Those skilled in the art will readily recognize that various non-critical parameters and conditions can be changed or modified to produce substantially the same results. According to one or more of the assays described herein, the following exemplified compounds were found to be inhibitors of the KIF18A protein.

[0089] The present invention is further described below with reference to the following examples, but these examples are not intended to limit the scope of the present invention.

[0090] TCFH:N,N,N',N'-Tetramethylchloroformamidine hexafluorophosphate

[0091] NMI:N-Methylimidazole

[0092] K2CO3: Potassium carbonate

[0093] Cs2CO3: Cesium carbonate

[0094] DMF:N,N-dimethylformamide

[0095] NMP:N-Methylpyrrolidone

[0096] THF:Tetrahydrofuran

[0097] MeOH: methanol

[0098] DCM: dichloromethane

[0099] EDCI:1-Ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride

[0100] DMAP:4,4-dimethylaminopyridine

[0101] DIPEA:N,N-diisopropylethylamine

[0102] HATU:2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate

[0103] DIBAL-H:Diisobutylaluminum hydride

[0104] T3P:1-propylphosphoric anhydride

[0105] Pd2(dba)3: Tris(dibenzylideneacetone)dipalladium

[0106] XantPhos: 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene

[0107] Oxone: Potassium persulfate

[0108] Intermediate 1: 4-iodo-2-(6-azaspiro[2.5]octyl-6-yl)benzoic acid (Intermediate 1)

[0109] Under nitrogen protection, 2-fluoro-4-iodobenzoic acid (1a) (10 g, 37.59 mmol), 6-azaspiro [2.5] octane hydrochloride (1b) (7.21 g, 48.87 mmol) and anhydrous potassium carbonate (15.59 g, 112.77 mmol) were added to DMSO (100 mL). The reaction solution was stirred at 140 ° C for 8 hours. After cooling to room temperature, the reaction solution was slowly poured into ice water (150 mL) and extracted with petroleum ether. The pH of the system was adjusted to 6 with HCl aqueous solution (6 M). A large amount of solid precipitated. The liquid was filtered, the solid was collected, and dried to obtain 4-iodo-2- (6-azaspiro [2.5] octyl-6-yl) benzoic acid (intermediate 1) (pale yellow solid, 9.4 g, 70% yield).

[0110] 1H NMR (400MHz, DMSO-d6) δ8.06(s,1H),7.74(d,J=8.0Hz,1H),7.69(d,J=8.0Hz,1H),3.31(s,1H),3.15-3.05(m,4H),1.59-1.49(m,4H),0.41(s,4H).

[0111] LC-MS m / z(ESI)=358.0[M+H].

[0112] Intermediate 2: 6-(5-bromo-2-ethynylphenyl)-6-azaspiro[2.5]octane (Intermediate 2)

[0113] Step 1: 2-(4,4-difluoropiperidin-1-yl)pyrimidine-4-carboxylic acid methyl ester (2b)

[0114] To a microwave reaction vial, methyl 2-bromopyrimidine-4-carboxylate (2a) (1 g, 4.61 mmol), 4,4-difluoropiperidine hydrochloride (796 mg, 5.07 mmol), DIPEA (1.78 g, 13.83 mmol), and NMP (10 mL) were added. The reaction mixture was heated to 150°C in a microwave reactor and stirred for 30 minutes. After cooling to room temperature, water (30 mL) was added to the reaction mixture, and the mixture was extracted with ethyl acetate (30 mL x 2). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 4 / 1) to afford methyl 2-(4,4-difluoropiperidin-1-yl)pyrimidine-4-carboxylate (2b) (yellow solid, 1 g, 84% yield).

[0115] LCMS m / z (ESI) = 258.1 [M+H].

[0116] Step 2: 2-(4,4-difluoropiperidin-1-yl)pyrimidine-4-carbohydrazide (2c)

[0117] Methyl 2-(4,4-difluoropiperidin-1-yl)pyrimidine-4-carboxylate (2b) (1 g, 3.89 mmol) was added to methanol (10 mL). Hydrazine hydrate (973 mg, 19.45 mmol) was slowly added dropwise under ice-water cooling. The mixture was stirred at room temperature for 2 hours. The mixture was cooled under ice-water cooling, filtered, and the filter cake was washed with a small amount of methanol and water, and dried to obtain 2-(4,4-difluoropiperidin-1-yl)pyrimidine-4-carbohydrazide (2c) (white solid, 800 mg, 80% yield).

[0118] LCMS m / z (ESI) = 258.2 [M+H].

[0119] Step 3: 2-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)pyrimidine-4-carbohydrazide (2d)

[0120] 2-(4,4-difluoropiperidin-1-yl)pyrimidine-4-carbohydrazide (2c) (800 mg, 3.11 mmol), 4-iodo-2-(6-azaspiro[2.5]octyl-6-yl)benzoic acid (Intermediate 1) (1.22 g, 3.42 mmol), and NMI (3.55 g, 9.33 mmol) were added to acetonitrile (15 mL). TCFH (481 mg, 3.73 mmol) was slowly added under ice-water cooling. The mixture was stirred at room temperature for 4 hours. The mixture was cooled in an ice-water bath, filtered, and the filter cake was washed with a small amount of acetonitrile and dried to obtain 2-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octyl-6-yl)benzoyl)pyrimidine-4-carbohydrazide (2d) (yellow solid, 1.3 g, 71% yield).

[0121] LCMS m / z (ESI) = 597.2 [M+H].

[0122] Step 4: 2-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (2e)

[0123] To a microwave reaction vial was added 2-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)pyrimidine-4-carbohydrazide (2d) (300 mg, 0.5 mmol), T3P (477 mg, 1.5 mmol), TEA (153 mg, 1.5 mmol), and 1,4-dioxane (5 mL). The reaction mixture was heated to 150°C in a microwave reactor and stirred for 20 minutes. After cooling to room temperature, water (100 mL) was added and the mixture was extracted with ethyl acetate (30 mL x 2). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by silica gel column chromatography (petroleum ether / ethyl acetate = 2 / 1) to give 2-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (2e) (yellow solid, 250 mg, 86% yield).

[0124] LCMS m / z (ESI) = 279.2 [M+H].

[0125] Step 5: N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 2)

[0126] Under nitrogen, 2-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (2e) (100 mg, 0.17 mmol), 2-hydroxyethane-1-sulfonamide (32 mg, 0.26 mmol), CuI (16 mg, 0.085 mmol), N,N-dimethylethylenediamine (7 mg, 0.085 mmol), and potassium carbonate (70 mg, 0.51 mmol) were added to acetonitrile (2 mL). The reaction mixture was stirred at 50°C for 3 hours. After cooling to room temperature, water (50 mL) was added, and the mixture was extracted with ethyl acetate (30 mL x 2). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, and concentrated. The residue was separated and purified by silica gel column chromatography (DCM / MeOH=20 / 1) to give N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]oct-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 2) (white solid, 40 mg, 40% yield).

[0127] 1 H NMR (400MHz, DMSO-d6) δ10.16(s,1H),8.67(d,J=4.9Hz,1H),7.92(d,J=8.5Hz,1H),7.42(d,J=4.9Hz,1H),7.07(s,1H),6.97(d,J=8.5Hz,1H),4. 92(s,1H),4.07-3.97(m,4H),3.77(t,J=6.4Hz,2H),3.37(t,J=6.4Hz,2H ),2.97-2.87(m,4H),2.15-2.00(m,4H),1.60-1.44(m,4H),0.32(s,4H).

[0128] LCMS m / z (ESI) = 576.2 [M+H].

[0129] Intermediate 3: 6-(5-bromo-2-ethynylphenyl)-6-azaspiro[2.5]octane (Intermediate 3)

[0130] Step 1: 4-Bromo-2-fluoro-N-methoxy-N-methylbenzamide (3b)

[0131] 4-Bromo-2-fluorobenzoic acid (3a) (112 g, 0.51 mol), N,O-dimethylhydroxylamine hydrochloride (55 g, 0.56 mol), and HATU (291 g, 0.77 mol) were dissolved in DMF (1 L). DIPEA (198 g, 1.53 mol) was added, and the reaction mixture was stirred at room temperature for 2 hours. Ethyl acetate (3 L) was added to the reaction mixture, and the mixture was washed sequentially with water (1 L x 3) and saturated brine (1 L). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 2 / 1) to obtain 4-bromo-2-fluoro-N-methoxy-N-methylbenzamide (3b) (colorless liquid, 127 g, 95% yield).

[0132] Step 2: 4-Bromo-N-methoxy-N-methyl-2-(6-azaspiro[2.5]octan-6-yl)benzamide (3c)

[0133] 4-Bromo-2-fluoro-N-methoxy-N-methylbenzamide (3b) (122 g, 0.47 mol), 6-azaspiro[2.5]octane hydrochloride (83 g, 0.56 mol) and potassium carbonate (161 g, 1.17 mol) were added to DMF (SO, 1.0 L). The reaction solution was stirred at 140 ° C for 48 hours. The reaction solution was cooled to room temperature and filtered. The filter cake was washed with ethyl acetate. Ethyl acetate (3 L) was added to the filtrate and washed with water (1 L x 3) and saturated brine (1 L) in sequence. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 7 / 1) to give 4-bromo-N-methoxy-N-methyl-2-(6-azaspiro[2.5]octan-6-yl)benzamide (3c) (yellow liquid, 167 g, 98% yield)

[0134] Step 3: 4-Bromo-2-(6-azaspiro[2.5]octan-6-yl)benzaldehyde (3d)

[0135] Under nitrogen, 4-bromo-N-methoxy-N-methyl-2-(6-azaspiro[2.5]octan-6-yl)benzamide (3c) (167 g, 0.47 mol) was dissolved in dry THF (1 L). The reaction solution was cooled to -78°C and DIBAL-H (620 mL, 0.62 mol) was slowly added dropwise with stirring. The reaction solution was stirred at -78°C for 2 hours. The temperature was raised to -25°C and water (25 mL) and sodium hydroxide solution (15% W, 25 mL) were added to the reaction solution with stirring. The solution was returned to room temperature, filtered, and concentrated to give 4-bromo-2-(6-azaspiro[2.5]octan-6-yl)benzaldehyde (3d) (yellow solid, 120 g, 86% yield). Step 4: 6-(5-Bromo-2-ethynylphenyl)-6-azaspiro[2.5]octane (Intermediate 3)

[0136] 4-Bromo-2-(6-azaspiro[2.5]octan-6-yl)benzaldehyde (3d) (35 g, 0.12 mol) was dissolved in methanol (300 mL), and dimethyl (1-diazo-2-oxopropyl)phosphonate (34 g, 0.18 mol) and potassium carbonate (25 g, 0.18 mmol) were added portionwise under ice-water cooling. The reaction solution was stirred at room temperature overnight, saturated aqueous sodium bicarbonate solution (300 mL) was added, and the mixture was extracted with DCM (1.0 L x 3). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 50 / 1) to give 6-(2-ethynyl-5-bromophenyl)-6-azaspiro[2.5]octane (Intermediate 3) (colorless liquid, 31 g, 90% yield).

[0137] Intermediate 4: N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyridin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 4)

[0138] Step 1: 4-azido-2-(methylthio)pyrimidine (4b)

[0139] Under nitrogen, 4-chloro-2-(methylthio)pyrimidine (4a) (50 g, 0.31 mol) and NaN3 (30.4 g, 0.47 mol) were added to DMF (300 mL), and the reaction mixture was stirred at room temperature for 24 hours. The reaction mixture was added to water (2 L) and filtered to obtain 4-azido-2-(methylthio)pyrimidine (4b) (yellow solid, 45 g, 87% yield).

[0140] Step 2: 4-azido-2-(methylsulfonyl)pyrimidine (4c)

[0141] 4-Azido-2-(methylthio)pyrimidine (4b) (45 g, 0.27 mol) was dissolved in ethyl acetate (1.8 L), and an aqueous solution of Oxone (1.8 L, 0.81 mol) was slowly added dropwise. The reaction mixture was stirred at room temperature for 2 hours. Water (2 L) was added, and the mixture was then extracted with ethyl acetate (1.5 L x 2). The combined organic phases were washed with saturated brine (1 L), dried over anhydrous sodium sulfate, filtered, and concentrated. 4-Azido-2-(methylsulfonyl)pyrimidine (4c) was obtained as a white solid, 46 g, 86% yield.

[0142] Step 3: 4-azido-2-(4,4-difluoropiperidin-1-yl)pyrimidine (4d)

[0143] 4-Azido-2-(methylsulfonyl)pyrimidine (4c) (110 g, 0.55 mol), 4,4-difluoropiperidine (87 g, 0.72 mol), and triethylamine (83 g, 0.83 mol) were dissolved in THF (1 L). The reaction mixture was stirred at 65°C for 1 hour. Water (1 L) was added, followed by extraction with ethyl acetate (2 L x 2). The combined organic phases were washed with saturated brine (1 L), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (PE / EA = 5 / 1) to give 4-azido-2-(4,4-difluoropiperidin-1-yl)pyrimidine (4d) (white solid, 56 g, 42% yield).

[0144] LCMS m / z (ESI) = 241.0 [M+H].

[0145] Step 4: 6-(5-bromo-2-(1-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)phenyl)-6-azaspiro[2.5]octane (4e)

[0146] Under nitrogen, 4-azido-2-(4,4-difluoropiperidin-1-yl)pyrimidine (4d) (76 g, 0.32 mol) was dissolved in tert-butanol (700 mL) and water (700 mL). 6-(5-bromo-2-ethynylphenyl)-6-azaspiro[2.5]octane (Intermediate 3) (83 g, 0.29 mmol), sodium ascorbate (57 g, 0.29 mol), and CuSO4.5H2O (72 g, 0.29 mol) were added. The reaction mixture was stirred at 80°C for 10 hours. After cooling to room temperature, water (1 L) was added, and the mixture was extracted with DCM (1.5 L x 3). The combined organic phases were washed with saturated brine (1 L), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by silica gel column chromatography (PE / EA=8 / 1) to give 6-(2-(1-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-5-bromophenyl)-6-azaspiro[2.5]octane (4e) (yellow solid, 71 g, 47% yield).

[0147] LCMS m / z (ESI) = 530.0 [M+H].

[0148] Step 5: N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyridin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 4)

[0149] Under nitrogen, 6-(5-bromo-2-(1-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)phenyl)-6-azaspiro[2.5]octane (4e) (60 g, 0.11 mol), 2-hydroxyethane-1-sulfonamide (22 g, 0.17 mol), CuI (22 g, 0.11 mmol), N,N'-dimethylethylenediamine (10 g, 0.11 mol), and potassium carbonate (47 g, 0.34 mol) were added to acetonitrile (1 L). The reaction mixture was stirred at 80°C for 10 hours. After cooling to room temperature, water (2 L) was added, and the mixture was extracted with DCM (2 L x 3). The combined organic phases were washed with saturated brine (2 L), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by silica gel column chromatography (DCM / MeOH=30 / 1) to give N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyridin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 4) (white solid, 33 g, 51% yield).

[0150] 1 H NMR (400MHz, DMSO-d6) δ9.89(s,1H),9.49(s,1H),8.67(d,J=5.2Hz,1H),8.10(d,J=8.5Hz,1H),7.38(d,J=5.2Hz,1H),7.20(d,J=1.9Hz ,1H),7.09(dd,J=8.5,1.9Hz,1H),4.94(s,1H),4.05-4.00(m,4H),3.78(t,J=6.5Hz,2H),3.32(t,J=6.5Hz,2H),2.90-2.82(m,4H),2.18 -1.98(m,4H),1.62-1.50(m,4H),0.38(s,4H);

[0151] LCMS m / z (ESI) = 575.3 [M+H].

[0152] Intermediate 5: (R)-2-Hydroxy-N-(4-(1-(2-(2-methylmorpholino)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)ethane-1-sulfonamide (Intermediate 5)

[0153] Step 1: (R)-4-(4-azidopyrimidin-2-yl)-2-methylmorpholine (5a)

[0154] 4-Azido-2-(methylsulfonyl)pyrimidine (4c) (3 g, 15.06 mmol), (R)-2-methylmorpholine hydrochloride (3.11 g, 22.59 mmol), and triethylamine (6.10 g, 60.24 mmol) were added to THF (10 mL). The reaction solution was stirred at 60°C for 1 hour. The reaction solution was cooled to room temperature, water (50 mL) was added, and extraction was performed with ethyl acetate (50 mL x 3). The organic phases were combined, washed with saturated brine (60 mL), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain crude (R)-4-(4-azidopyrimidin-2-yl)-2-methylmorpholine (5a) (yellow oil, 2.4 g, 72% yield). This compound was used in the next reaction without further purification.

[0155] LCMS m / z (ESI) = 221.3 [M+H].

[0156] Step 2: (R)-4-(4-(4-(-4-bromo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1H-1,2,3-triazol-1-yl)pyrimidin-2-yl)-2-methylmorpholine (5b)

[0157] Under nitrogen, 6-(5-bromo-2-ethynylphenyl)-6-azaspiro[2.5]octane (Intermediate 3) (1.1 g, 3.79 mmol), (R)-4-(4-azidopyrimidin-2-yl)-2-methylmorpholine (5a) (918 mg, 4.17 mmol), sodium ascorbate (750 mg, 3.79 mmol), and CuSO4.5H2O (946 mg, 3.79 mmol) were added to tert-butyl alcohol (5 mL) and water (5 mL). The reaction solution was stirred at 50°C for 3 hours. The reaction solution was cooled to room temperature, water (100 mL) was added, and the solution was extracted with DCM / MeOH = 10 / 1 (20 mL x 3). The organic phases were combined, washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by slurry separation (methanol) to give (R)-4-(4-(4-(-4-bromo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1H-1,2,3-triazol-1-yl)pyrimidin-2-yl)-2-methylmorpholine (5b) as a yellow solid, 600 mg, 31% yield. LCMS m / z (ESI) = 510.6 [M+H].

[0158] Step 3: (R)-2-Hydroxy-N-(4-(1-(2-(2-methylmorpholino)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)ethane-1-sulfonamide (Intermediate 5)

[0159] Under nitrogen, (R)-4-(4-(4-(-4-bromo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1H-1,2,3-triazol-1-yl)pyrimidin-2-yl)-2-methylmorpholine (5b) (600 mg, 1.18 mmol), 2-hydroxy-1-sulfonamide (221 mg, 1.77 mmol), N,N'-dimethylethylenediamine (52 mg, 0.59 mmol), CuI (112 mg, 0.59 mmol), and potassium carbonate (488 mg, 3.54 mmol) were added to acetonitrile (10 mL). The reaction solution was stirred at 80°C for 2 hours. The reaction solution was cooled to room temperature, water (50 mL) was added, and the solution was extracted with ethyl acetate (20 mL x 3). The organic phases were combined, washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by reverse phase chromatography (methanol / water = 3:1) to give (R)-2-hydroxy-N-(4-(1-(2-(2-methylmorpholino)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)ethane-1-sulfonamide (Intermediate 5) (yellow solid, 170 mg, 26% yield).

[0160] 1 H NMR (400MHz, DMSO-d6) δ9.43(s,1H),8.63(d,J=5.2Hz,1H),8.09(d,J=8.4Hz,1H),7.32(d ,J=5.2Hz,1H),7.17(s,1H),7.07(d,J=8.4Hz,1H),4.58-4.45(m,2H),3.96-3.89(m,1H), 3.79–3.72(m,2H),3.61–3.51(m,2H),3.32(t,J=6.5Hz,2H),3.13-3.03(ma,1H),2.87-2. 81(m,4H),2.80-2.73(m,1H),1.65-1.42(m,4H),1.19(d,J=6.2Hz,3H),0.35(s,4H); LCMS m / z(ESI)=555.9[M+H];

[0161] Intermediate 6: 4-ethynyl-2-(methylthio)pyrimidine (Intermediate 6)

[0162] Step 1: 2-(Methylthio)-4-((trimethylsilyl)ethyl)pyrimidine (6b)

[0163] Under nitrogen, 4-chloro-2-methylthiopyrimidine (6a) (3 g, 18.68 mmol), (trimethylsilyl)acetylene (1.83 g, 18.68 mmol), Pd(PPh3)2Cl2 (0.66 g, 0.93 mmol), triethylamine (7.8 mL, 56.04 mmol), and CuI (0.35 g, 1.87 mmol) were added to DMF (30 mL). The reaction mixture was stirred at 80°C for 4 hours. The reaction mixture was cooled to room temperature, water (150 mL) was added, and the mixture was extracted with ethyl acetate (50 mL x 3). The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 10 / 1) to give 2-(methylthio)-4-((trimethylsilyl)ethyl)pyrimidine (6b) (yellow oil, 2.75 g, 66% yield).

[0164] LC-MS m / z(ESI)=223.3[M+H].

[0165] Step 2: 4-ethynyl-2-(methylthio)pyrimidine (Intermediate 6)

[0166] 2-(Methylthio)-4-((trimethylsilyl)ethyl)pyrimidine (6b) (2.56 g, 11.51 mmol) was dissolved in MeOH (15 mL) and THF (15 mL), and anhydrous potassium carbonate (2.39 g, 17.27 mmol) was added. The mixture was stirred at room temperature for 30 minutes. Water (50 mL) was added, and the mixture was extracted with ethyl acetate (50 mL x 3). The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 10 / 1) to give 4-ethynyl-2-(methylthio)pyrimidine (Intermediate 6) (yellow solid, 830 mg, 48% yield).

[0167] LC-MS m / z(ESI)=151.2[M+H].

[0168] Intermediate 7: N-(4-(4-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 7)

[0169] Step 1: 6-(5-Bromo-2-nitrophenyl)-6-azaspiro[2.5]octane (7b)

[0170] 4-Bromo-2-fluoro-1-nitrobenzene (7a) (2.2 g, 10.0 mmol) and 6-azaspiro[2.5]octane hydrochloride (2.94 g, 20.0 mmol) were added to acetonitrile (25 mL), and DIPEA (3.87 g, 30.0 mmol) was added. The reaction mixture was stirred at room temperature for 4 hours. Water (100 mL) was added to the reaction mixture, and the mixture was extracted with ethyl acetate (50 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 10 / 1) to obtain 6-(5-bromo-2-nitrophenyl)-6-azaspiro[2.5]octane (7b) (yellow oil, 3.0 g, 97% yield).

[0171] Step 2: 4-Bromo-2-(6-azaspiro[2.5]octan-6-yl)aniline (7c)

[0172] 6-(5-Bromo-2-nitrophenyl)-6-azaspiro[2.5]octane (7b) (2.3 g, 9.64 mmol), iron powder (3.24 g, 57.84 mmol), and NH4Cl (3.12 g, 57.84 mmol) were added to ethanol (30 mL) and water (10 mL). The reaction solution was stirred at 80°C for 3 hours. The reaction solution was cooled to room temperature and quenched by adding saturated sodium bicarbonate solution (30 mL). The solution was filtered through celite, and the filtrate was extracted with ethyl acetate (50 mL x 3). The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated to afford 4-bromo-2-(6-azaspiro[2.5]octan-6-yl)aniline (7c) (yellow solid, 2.5 g, 92% yield).

[0173] LCMS m / z (ESI) = 281.2 [M+H].

[0174] Step 3: 6-(2-azido-5-bromophenyl)-6-azaspiro[2.5]octane (7d)

[0175] 4-Bromo-2-(6-azaspiro[2.5]octan-6-yl)aniline (7c) (2.3 g, 8.18 mmol) and sodium nitrite (790 mg, 11.45 mmol) were dissolved in acetic acid (20 mL) and water (10 mL). The reaction mixture was cooled to 0°C, and sodium azide (800 mg, 12.27 mmol) was added portionwise. The reaction mixture was stirred in an ice bath for 1 hour. The reaction mixture was returned to room temperature and adjusted to pH 7 by adding saturated aqueous sodium carbonate. The mixture was extracted with ethyl acetate (20 mL x 3). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 10 / 1) to afford 6-(2-azido-5-bromophenyl)-6-azaspiro[2.5]octane (7d) as a yellow oil, 1.3 g, 52% yield.

[0176] LCMS m / z (ESI) = 307.6 [M+H].

[0177] Step 4: 6-(5-bromo-2-(4-(2-(methylthio)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)phenyl)-6-azaspiro[2.5]octane (7e)

[0178] Under nitrogen, 6-(2-azido-5-bromophenyl)-6-azaspiro[2.5]octane (7d) (1.2 g, 3.91 mmol), 4-ethynyl-2-(methylthio)pyrimidine (Intermediate 6) (646 mg, 4.30 mmol), sodium ascorbate (775 mg, 3.91 mmol), and copper sulfate pentahydrate (976 mg, 3.91 mmol) were added to tert-butyl alcohol (10 mL) and water (10 mL). The reaction mixture was stirred at 70°C for 1 hour. The reaction mixture was cooled to room temperature, water (100 mL) was added, and the mixture was extracted with ethyl acetate (30 mL x 3). The organic phases were combined, washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 10 / 1) to give 6-(5-bromo-2-(4-(2-(methylthio)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)phenyl)-6-azaspiro[2.5]octane (7e) (yellow solid, 1.42 g, 79% yield).

[0179] LCMS m / z (ESI) = 456.9 [M+H].

[0180] Step 5: 6-(5-Bromo-2-(4-(2-(methylsulfonyl)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)phenyl)-6-azaspiro[2.5]octane (7f)

[0181] 6-(5-Bromo-2-(4-(2-(methylthio)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)phenyl)-6-azaspiro[2.5]octane (7e) (1.32 g, 2.89 mmol) and Oxone (8.88 g, 14.45 mmol) were added to ethyl acetate (20 mL) and water (20 mL). The reaction mixture was stirred at 60°C for 8 hours. The reaction mixture was cooled to room temperature, water (100 mL) was added, and the mixture was extracted with ethyl acetate (30 mL x 3). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 5 / 1) to give 6-(5-bromo-2-(4-(2-(methylsulfonyl)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)phenyl)-6-azaspiro[2.5]octane (7f) (off-white solid, 620 mg, 45% yield).

[0182] LCMS m / z (ESI) = 489.1 [M+H].

[0183] Step 6: 6-(5-Bromo-2-(4-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)phenyl)-6-azaspiro[2.5]octane (7 g)

[0184] 5-Bromo-2-(4-(2-(methylsulfonyl)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)phenyl)-6-azaspiro[2.5]octane (7f) (550 mg, 1.12 mmol), 4,4-difluoropiperidine hydrochloride (265 mg, 1.68 mmol), and triethylamine (453 mg, 4.48 mmol) were added to NMP (8 mL). The mixture was heated to 180°C in a microwave reactor for 40 minutes. The reaction mixture was cooled to room temperature and directly purified by reverse phase chromatography to afford 6-(5-bromo-2-(4-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)phenyl)-6-azaspiro[2.5]octane (7 g) as a yellow solid, 460 mg, 77% yield.

[0185] LCMS m / z (ESI) = 530.1 [M+H].

[0186] Step 7: N-(4-(4-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 7)

[0187] Under nitrogen, 6-(5-bromo-2-(4-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)phenyl)-6-azaspiro[2.5]octane (7 g) (400 mg, 0.75 mmol), 2-hydroxyethane-1-sulfonamide (141 mg, 1.13 mmol), N,N'-dimethylethylenediamine (33 mg, 0.38 mmol), CuI (72 mg, 0.38 mmol), and potassium carbonate (311 mg, 2.25 mmol) were added to acetonitrile (6 mL). The reaction mixture was stirred at 80°C for 6 hours. The reaction mixture was cooled to room temperature, water (50 mL) was added, and the mixture was extracted with ethyl acetate (10 mL x 3). The organic phases were combined, washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by reverse phase chromatography (methanol / water = 3:1) to give N-(4-(4-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-1-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 7) (yellow solid, 200 mg, 46% yield).

[0188] 1 H NMR (400MHz, DMSO-d6) δ9.28 (s, 1H), 8.54 (d, J = 5.0Hz, 1H), 7.59 (d, J = 8.5Hz, 1H), 7. 31(d,J=5.0Hz,1H),7.15(d,J=2.3Hz,1H),7.05(dd,J=8.5,2.3Hz,1H),4.05-3.97(m, 4H),3.80(t,J=6.6Hz,2H),3.38-3.34(m,2H),2.72-2.67(m,4H),2.10-1.95(m,4H),1.35-1.25(m,4H),0.27(s,4H);

[0189] LCMS m / z (ESI) = 575.2 [M+H].

[0190] Example 1: N-(4-(5-(6-(4,4-difluoropiperidin-1-yl)-5-methoxypyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-)6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 1)

[0191] Step 1: 6-(4,4-difluoropiperidin-1-yl)-5-methoxypicolinic acid methyl ester (1B)

[0192] Under nitrogen, methyl 6-bromo-5-methoxypicolinate (1A) (800 mg, 3.25 mmol) and 4,4-difluoropiperidine hydrochloride (564 mg, 3.58 mmol) were added to 1,4-dioxane (10 mL). Cs2CO3 (3.18 g, 9.75 mmol), Pd2(dba)3 (149 mg, 0.16 mmol), and XantPhos (39 mg, 0.33 mmol) were also added. The reaction solution was stirred at 100°C for 10 hours. After cooling to room temperature, water (30 mL) was added to the reaction solution, and the solution was extracted with ethyl acetate (30 mL x 2). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by silica gel column chromatography (petroleum ether / ethyl acetate = 10 / 1-5 / 1) to give methyl 6-(4,4-difluoropiperidin-1-yl)-5-methoxypicolinate (1B) (yellow solid, 570 mg, 61% yield).

[0193] LCMS m / z (ESI) = 287.3 [M+H].

[0194] Step 2: 6-(4,4-difluoropiperidin-1-yl)-5-methoxypicolinohydrazide (1C)

[0195] Methyl 6-(4,4-difluoropiperidin-1-yl)-5-methoxypicolinate (1B) (570 mg, 1.99 mmol) was dissolved in MeOH (8 mL). Hydrazine hydrate (399 mg, 7.56 mmol) was slowly added dropwise under ice-water cooling and returned to room temperature. The reaction mixture was stirred at room temperature for 2 hours. The mixture was cooled under ice-water cooling and filtered. The filter cake was washed with a small amount of methanol and water and dried to obtain 6-(4,4-difluoropiperidin-1-yl)-5-methoxypicolinohydrazide (1C) (white solid, 350 mg, 61% yield).

[0196] LCMS m / z (ESI) = 287.1 [M+H].

[0197] Step 3: 6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)-5-methoxypicolinohydrazide (1D)

[0198] 6-(4,4-difluoropiperidin-1-yl)-5-methoxypicolinohydrazide (1C) (350 mg, 1.22 mmol), 4-iodo-2-(6-azaspiro[2.5]octyl-6-yl)benzoic acid (Intermediate 1) (524 mg, 1.46 mmol), and NMI (300 mg, 3.66 mmol) were added to acetonitrile (12 mL). TCFH (411 mg, 1.46 mmol) was slowly added under ice-water cooling. The reaction mixture was stirred at room temperature for 4 h. After cooling in an ice-water bath, the mixture was filtered. The filter cake was washed with a small amount of acetonitrile and dried to give 6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octyl-6-yl)benzoyl)-5-methoxypicolinohydrazide (1D) (white solid, 650 mg, 85% yield).

[0199] LCMS m / z (ESI) = 626.2 [M+H].

[0200] Step 4: 2-(6-(4,4-difluoropiperidin-1-yl)-5-methoxypyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (1E)

[0201] To a microwave reaction vial was added 6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)-5-methoxypicolinohydrazide (1D) (650 mg, 1.04 mmol), T3P (1.98 g, 3.12 mmol), triethylamine (315 mg, 3.12 mmol), and 1,4-dioxane (10 mL). The reaction was heated to 150°C in a microwave reactor for 20 minutes. The mixture was cooled to room temperature, and water (50 mL) was added. The mixture was extracted with ethyl acetate (50 mL x 2). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by silica gel column chromatography (petroleum ether / ethyl acetate = 8 / 1-2 / 1) to give 2-(6-(4,4-difluoropiperidin-1-yl)-5-methoxypyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (1E) (yellow solid, 540 mg, 85% yield).

[0202] LCMS m / z (ESI) = 608.3 [M+H].

[0203] Step 5: N-(4-(5-(6-(4,4-difluoropiperidin-1-yl)-5-methoxypyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-)6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 1)

[0204] Under nitrogen, 2-(6-(4,4-difluoropiperidin-1-yl)-5-methoxypyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (1E) (200 mg, 0.33 mmol), 2-hydroxyethane-1-sulfonamide (62 mg, 0.49 mmol), CuI (31 mg, 0.165 mmol), N,N-dimethylethylenediamine (15 mg, 0.165 mmol), and K2CO3 (136 mg, 0.987 mmol) were added to acetonitrile (3 mL). The reaction mixture was stirred at 50°C for 3 hours. After cooling to room temperature, water (100 mL) was added, and the mixture was extracted with ethyl acetate (30 mL x 2). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by preparative thin layer chromatography (DCM / MeOH=20 / 1) to give N-(4-(5-(6-(4,4-difluoropiperidin-1-yl)-5-methoxypyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-)6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 1) (white solid, 15 mg, 7.5% yield).

[0205] 1 H NMR (400MHz, DMSO-d6) δ9.64(s,1H),7.83(d,J=7.6Hz,1H),7.82(d,J=8.5H z,1H),7.49(d,J=7.6Hz,1H),7.04(s,1H),6.93(d,J=8.5Hz,1H),4.82(s,1 H),3.93(s,3H),3.77(t,J=6.4Hz,2H),3.62-3.55(m,4H),3.36(t,J=6.4Hz ,2H),2.97-2.86(m,4H),2.20-2.03(m,4H),1.60-1.44(m,4H),0.29(s,4H);

[0206] LCMS m / z (ESI) = 605.4 [M+H].

[0207] Example 2: N-(4-(5-(6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 2)

[0208] Step 1: Ethyl 6-(4,4-difluoropiperidin-1-yl)picolinate (2B)

[0209] Under nitrogen, ethyl 6-chloropicolinate (1 g, 5.39 mmol), 4,4-difluoropiperidine hydrochloride (1.27 g, 8.08 mmol), and DIPEA (2.1 g, 16.17 mmol) were added to NMP (15 mL), and the reaction mixture was stirred at 150°C for 2 hours. After cooling to room temperature, water (100 mL) was added, and the mixture was extracted with ethyl acetate (30 mL x 3). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, and filtered. The organic phases were collected and concentrated by rotary evaporation to afford ethyl 6-(4,4-difluoropiperidin-1-yl)picolinate (2B) as a white solid, 400 mg, 27% yield.

[0210] LC-MS m / z(ESI)=271.5[M+H].

[0211] Step 2: 6-(4,4-difluoropiperidin-1-yl)pyridine hydrazide (2C)

[0212] Ethyl 6-(4,4-difluoropiperidin-1-yl)picolinate (2B) (400 mg, 1.48 mmol) and hydrazine hydrate (370 mg, 7.4 mmol) were added to MeOH (5 mL), and the reaction mixture was stirred at 85°C for 2 hours. After cooling to room temperature, a solid precipitated and was filtered. The filter cake was washed with a small amount of methanol and water and dried to afford 6-(4,4-difluoropiperidin-1-yl)picolinohydrazide (2C) (white solid, 300 mg, 79% yield).

[0213] LC-MS m / z(ESI)=257.4[M+H].

[0214] Step 3: 6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinylhydrazide (2D)

[0215] 6-(4,4-difluoropiperidin-1-yl)picolinohydrazide (2C) (300 mg, 1.09 mmol), 4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoic acid (Intermediate 1), and NMI (268 mg, 3.27 mmol) were added to acetonitrile (3 mL). TCFH (367 mg, 1.31 mmol) was slowly added under ice-water cooling. The reaction mixture was stirred at room temperature for 2 hours. The mixture was filtered, and the filter cake was washed with a small amount of acetonitrile and dried to give 6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinohydrazide (2D) (white solid, 450 mg, 67% yield).

[0216] LC-MS m / z(ESI)=296.8[M+H].

[0217] Step 4: 2-(6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (2E)

[0218] Under nitrogen, 6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinohydrazide (2D) (450 mg, 0.76 mmol), T3P (1.9 g, 3.04 mmol), and triethylamine (230 mg, 2.28 mmol) were added to 1,4-dioxane (5 mL). The mixture was heated in a microwave oven at 150°C and stirred for 0.5 hours. After cooling to room temperature, water (20 mL) was added and the mixture was extracted with ethyl acetate (10 mL x 3). The combined organic phases were washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 410 / 1) to give 2-(6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (2E) (white solid, 300 mg, 69% yield).

[0219] LC-MS m / z(ESI)=578.7[M+H].

[0220] Step 5: N-(4-(5-(6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 2)

[0221] Under nitrogen, 2-(6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (2E) (300 mg, 0.52 mmol), 2-hydroxyethane-1-sulfonamide (98 mg, 0.78 mmol), N,N-dimethylethylenediamine (23 mg, 0.26 mmol), CuI (50 mg, 0.26 mmol), and K2CO3 (216 mg, 1.56 mmol) were added to acetonitrile (5 mL). The reaction mixture was stirred at 50°C for 2 hours. After cooling to room temperature, water (20 mL) was added, and the mixture was extracted with ethyl acetate (10 mL x 3). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by reverse phase column chromatography (methanol / water = 4:1) to give N-(4-(5-(6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 2) (white solid, 110 mg, 37% yield).

[0222] 1 H NMR (400MHz, DMSO-d6) δ7.86(d,J=8.0Hz,1H),7.80(t,J=8.0Hz,1H),7.55(d,J=7.9Hz,1H),7.21(d,J=8.0Hz,1H),7.06(s,1H),6.96(d,J=7.9 Hz,1H),3.90-3.81(m,4H),3.77(t,J=6.6Hz,2H),3.37(t,J=6.6Hz,2H) ,2.97-2.88(m,4H),2.13-1.98(m,4H),1.58-1.44(m,4H),0.31(s,4H).

[0223] LC-MS m / z(ESI)=575.9[M+H].

[0224] Example 3: N-(4-(6-(4,4-difluoropiperidin-1-yl)-5-fluoropyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 3)

[0225] Step 1: 6-(4,4-difluoropiperidin-1-yl)-5-fluoropicolinic acid methyl ester (3B)

[0226] Under nitrogen, methyl 6-chloro-5-fluoropicolinate (1 g, 4.27 mmol), 4,4-difluoropiperidine hydrochloride (0.8 g, 5.12 mmol), Pd2(dba)3 (0.20 g, 0.21 mmol), XantPhos (0.25 g, 0.43 mmol), and Cs2CO3 (4.18 g, 12.81 mmol) were added to 1,4-dioxane (10 mL). The reaction mixture was stirred at 100°C for 4 hours. The mixture was allowed to return to room temperature. Water (100 mL) was added, and the mixture was extracted with ethyl acetate (30 mL x 3). The combined organic phases were washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by column chromatography (petroleum ether / ethyl acetate = 50 / 1) to give methyl 6-(4,4-difluoropiperidin-1-yl)-5-fluoropicolinate (3B) (yellow solid, 640 mg, 55% yield).

[0227] LC-MS m / z(ESI)=275.0[M+H].

[0228] Step 2: 6-(4,4-difluoropiperidin-1-yl)-5-fluoropicolinohydrazide (3C)

[0229] Methyl 6-(4,4-difluoropiperidin-1-yl)-5-fluoropicolinate (3B) (640 mg, 2.33 mmol) and hydrazine hydrate (583 mg, 11.65 mmol) were added to MeOH (8 mL). The reaction mixture was stirred at 85°C for 2 hours. After cooling to room temperature, a solid precipitated and was filtered. The filter cake was washed with a small amount of methanol and water and dried to yield 6-(4,4-difluoropiperidin-1-yl)-5-fluoropicolinohydrazide (3C) (white solid, 630 mg, 99% yield).

[0230] LC-MS m / z(ESI)=275.0[M+H].

[0231] Step 3: 6-(4,4-difluoropiperidin-1-yl)-5-fluoro-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinylhydrazide (3D)

[0232] 6-(4,4-difluoropiperidin-1-yl)-5-fluoropicolinohydrazide (3C) (630 mg, 2.30 mmol), 4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoic acid (Intermediate 1) (905 mg, 2.53 mmol), and NMI (566 mg, 6.90 mmol) were added to acetonitrile (20 mL). TCFH (774 mg, 2.76 mmol) was slowly added under ice-water cooling. The reaction mixture was stirred at room temperature for 2 hours. Filtered, and the filter cake was washed with a small amount of acetonitrile to obtain 6-(4,4-difluoropiperidin-1-yl)-5-fluoro-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinohydrazide (3D) (white solid, 1 g, 71% yield).

[0233] LC-MS m / z(ESI)=614.0[M+H].

[0234] Step 4: 2-(6-(4,4-difluoropiperidin-1-yl)-5-fluoropyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (3E)

[0235] 6-(4,4-difluoropiperidin-1-yl)-5-fluoro-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinohydrazide (3D) (1 g, 1.63 mmol), T3P (4.15 g, 6.52 mmol), and triethylamine (0.49 g, 4.89 mmol) were added to 1,4-dioxane (30 mL). The mixture was heated in a microwave oven at 150°C for 0.5 h. After cooling to room temperature, water (100 mL) was added and the mixture was extracted with ethyl acetate (20 mL x 3). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 4 / 1) to give 2-(6-(4,4-difluoropiperidin-1-yl)-5-fluoropyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (3E) (yellow solid, 0.5 g, 52% yield).

[0236] LC-MS m / z(ESI)=596.4[M+H].

[0237] Step 5: N-(4-(6-(4,4-difluoropiperidin-1-yl)-5-fluoropyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 3)

[0238] Under nitrogen, 2-(6-(4,4-difluoropiperidin-1-yl)-5-fluoropyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (3E) (500 mg, 0.84 mmol), 2-hydroxyethane-1-sulfonamide (160 mg, 1.26 mmol), N,N-dimethylethylenediamine (37 mg, 0.42 mmol), CuI (80 mg, 0.42 mmol), and K2CO3 (349 mg, 2.52 mmol) were added to acetonitrile (10 mL). The reaction mixture was stirred at 50°C for 2 hours. After cooling to room temperature, water (100 mL) was added, and the mixture was extracted with ethyl acetate (20 mL x 3). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated by rotary evaporation. The residue was separated and purified by reverse phase chromatography (methanol / water = 3:1) to give N-(4-(6-(4,4-difluoropiperidin-1-yl)-5-fluoropyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 3) (white solid, 270 mg, 54% yield).

[0239] 1 H NMR (400MHz, DMSO-d6) δ9.82(s,1H),7.90-7.73(m,3H),7.05(s,1H),6.94(d,J=8.5Hz,1H),4.72(s,1H),3.77(t,J=6.5 Hz,2H),3.73-3.65(m,4H),3.37(t,J=6.5Hz,2H),2.97-2.88(m,4H),2.22-2.08(m,4H),1.59-1.44(m,4H),0.30(s,4H).

[0240] LC-MS m / z(ESI)=593.1[M+H].

[0241] Example 4: N-(5-(5-bromo-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-1,3Z,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 4)

[0242] Step 1: 5-Bromo-6-(4,4-difluoropiperidin-1-yl)picolinic acid methyl ester (4B)

[0243] Methyl 5-bromo-6-chloropicolinate (1 g, 3.99 mmol), 4,4-difluoropiperidine hydrochloride (0.94 g, 5.99 mmol), and DIPEA (1.54 g, 11.97 mmol) were added to NMP (10 mL). The reaction mixture was stirred in a microwave at 150°C for 1 hour. After cooling to room temperature, water (50 mL) was added, and the mixture was extracted with ethyl acetate (20 mL x 3). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated by rotary evaporation to afford methyl 5-bromo-6-(4,4-difluoropiperidin-1-yl)picolinate (4B) as a yellow solid, 1 g, 75% yield.

[0244] LC-MS m / z(ESI)=335.2,337.2[M+H].

[0245] Step 2: 5-Bromo-6-(4,4-difluoropiperidin-1-yl)pyridine hydrazide (4C)

[0246] Methyl 5-bromo-6-(4,4-difluoropiperidin-1-yl)picolinate (4B) (500 mg, 1.96 mmol) and hydrazine hydrate (261 mg, 4.44 mmol) were added to MeOH (5 mL). The reaction mixture was stirred at room temperature for 1 hour. Solid precipitated and was filtered. The filter cake was washed with a small amount of methanol and water and dried to yield 5,5-bromo-6-(4,4-difluoropiperidin-1-yl)picolinohydrazide (4C) (yellow solid, 259 mg, 69% yield).

[0247] LC-MS m / z(ESI)=335.2,337.2[M+H].

[0248] Step 3: 5-Bromo-6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinylhydrazide (4D)

[0249] 5-Bromo-6-(4,4-difluoropiperidin-1-yl)picolinohydrazide (Compound 4C) (259 mg, 0.77 mmol), 4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoic acid (Intermediate 1) (305 mg, 0.85 mmol), and NMI (190 mg, 2.31 mmol) were added to acetonitrile (3 mL). TCFH (260 mg, 0.92 mmol) was slowly added under ice-water cooling. The reaction mixture was stirred at room temperature for 2 hours. Filtered, the filter cake was washed with a small amount of acetonitrile to obtain 5-bromo-6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinohydrazide (4D) (yellow solid, 418 mg, 80% yield).

[0250] LC-MS m / z(ESI)=674.3,676.3[M+H].

[0251] Step 4: 2-(5-bromo-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (4E)

[0252] 5-Bromo-6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinohydrazide (4D) (400 mg, 0.59 mmol), T3P (1.5 g, 2.36 mmol), and triethylamine (180 mg, 1.77 mmol) were added to 1,4-dioxane (5 mL). The reaction mixture was stirred in a microwave at 150°C for 1 hour. After cooling to room temperature, water (100 mL) was added, and the mixture was extracted with ethyl acetate (10 mL x 3). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated by rotary evaporation. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 4 / 1) to give 2-(5-bromo-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (4E) (white solid, 250 mg, 64% yield).

[0253] LC-MS m / z(ESI)=656.7,658.0[M+H].

[0254] Step 5: N-(5-(5-bromo-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 4)

[0255] Under nitrogen, 2-(5-bromo-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (4E) (100 mg, 0.15 mmol), 2-hydroxyethane-1-sulfonamide (28 mg, 0.22 mmol), N,N-dimethylethylenediamine (7 mg, 0.08 mmol), CuI (15 mg, 0.08 mmol), and K2CO3 (62 mg, 0.45 mmol) were added to acetonitrile (1 mL). The reaction mixture was stirred at 50°C for 2 hours. After cooling to room temperature, water (50 mL) was added, and the mixture was extracted with ethyl acetate (10 mL x 3). The combined organic phases were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated by rotary evaporation. The residue was purified by preparative thin layer chromatography (petroleum ether / ethyl acetate = 1 / 2) to give N-(5-(5-bromo-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 4) (off-white solid, 10 mg, 10% yield).

[0256] 1 H NMR (400MHz, DMSO-d6) δ10.17(s,1H),8.26(d,J=8.0Hz,1H),7.89(d,J=8.5Hz,1H),7.79(d,J=8.0Hz,1H),7.05(s,1H),6.95(d,J=8.5Hz,1H),4. 87(s,1H),3.77(t,J=6.5Hz,2H),3.54-3.46(m,4H),3.37(t,J=6.5Hz,2H ),2.97-2.88(m,4H),2.26-2.11(m,4H),1.59-1.46(m,4H),0.30(s,4H);

[0257] LC-MS m / z(ESI)=653.1,655.1[M+H].

[0258] Example 5: N-(4-(5-(6-(4,4-difluoropiperidin-1-yl)-5-(2-hydroxyethoxy)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 5)

[0259] Step 1: 5-(2-(Benzyloxy)ethoxy)-6-bromopicolinic acid methyl ester (5B)

[0260] Under nitrogen, methyl 6-bromo-5-hydroxypicolinate (5A) (1 g, 4.31 mmol) was dissolved in DMF (15 mL), and ((2-bromoethoxy)methyl)benzene (1 g, 4.65 mmol) and K2CO3 (1.5 g, 10.85 mmol) were added. The reaction mixture was heated to 80°C and stirred for 4 hours. After cooling to room temperature, water (30 mL) was added and the mixture was extracted with ethyl acetate (30 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 2 / 1) to obtain methyl 5-(2-(benzyloxy)ethoxy)-6-bromopicolinate (5B) (yellow solid, 1.49 g, 94% yield). LC-MS m / z (ESI) = 266.1 [M+H].

[0261] Step 2: Methyl 5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)picolinate (5C)

[0262] Under nitrogen, methyl 5-(2-(benzyloxy)ethoxy)-6-bromopicolinate (5B) (700 mg, 1.91 mmol) and 4,4-difluoropiperidine hydrochloride (326 mg, 2.1 mmol) were dissolved in 1,4-dioxane (15 mL). Cs2CO3 (1.87 g, 5.73 mmol), Pd2(dba)3 (87 mg, 0.095 mmol), and XantPhos (111 mg, 0.191 mmol) were added. The reaction mixture was heated to 100°C and stirred for 5 hours. After cooling to room temperature, water (30 mL) was added and the mixture was extracted with ethyl acetate (30 mL x 2). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography (petroleum ether / ethyl acetate = 4 / 1) to give methyl 5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)picolinate (5C) (yellow solid, 330 mg, 42% yield).

[0263] LC-MS m / z(ESI)=407.2[M+H].

[0264] Step 3: 5-(2-(Benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)pyridine hydrazide (5D)

[0265] Methyl 5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)picolinate (Compound 5C) (330 mg, 0.812 mmol) was dissolved in MeOH (10 mL) and slowly added dropwise with ice-water cooling. Hydrazine hydrate (163 mg, 3.25 mmol) was added. The reaction mixture was stirred at room temperature for 2 hours. The mixture was cooled with ice-water, filtered, and the filter cake was washed with a small amount of methanol and water, and dried to obtain 5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)picolinohydrazide (5D) (white solid, 260 mg, 78% yield).

[0266] LC-MS m / z(ESI)=407.4[M+H].

[0267] Step 4: 5-(2-(Benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinohydrazide (5E)

[0268] 5-(2-(Benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)picolinohydrazide (5D) (260 mg, 0.64 mmol), 4-iodo-2-(6-azaspiro[2.5]octyl-6-yl)benzoic acid (Intermediate 1) (274 mg, 0.77 mmol) and NMI (158 mg, 1.92 mmol) were dissolved in CH3CN (10 mL) and TCFH (216 mg, 0.77 mmol) was slowly added under ice-water cooling. The mixture was stirred at room temperature overnight, cooled in an ice-water bath, filtered, and the filter cake was washed with a small amount of acetonitrile and dried to give 5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinohydrazide (5E) (yellow solid, 360 mg, 75% yield).

[0269] LC-MS m / z(ESI)=746.6[M+H].

[0270] Step 5: 2-(5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (5F)

[0271] To a microwave reaction vial, add 5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)-N'-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)benzoyl)picolinohydrazide (5E) (360 mg, 0.483 mmol), T3P (922 mg, 1.45 mmol), triethylamine (147 mg, 1.45 mmol), and 1,4-dioxane (4 mL). Heat to 150°C in a microwave reactor and react for 15 minutes. Cool to room temperature, add water (50 mL), and extract with ethyl peroxide (20 mL x 2). The organic phase is washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by column chromatography (petroleum ether / ethyl acetate = 8 / 1-2 / 1) to give 2-(5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (5F) (yellow solid, 256 mg, 72% yield).

[0272] LC-MS m / z(ESI)=728.8[M+H].

[0273] Step 6: N-(4-(5-(5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (5G)

[0274] Under nitrogen, 2-(5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-5-(4-iodo-2-(6-azaspiro[2.5]octan-6-yl)phenyl)-1,3,4-oxadiazole (5F) (256 mg, 0.352 mmol), 2-hydroxyethane-1-sulfonamide (66 mg, 0.528 mmol), CuI (34 mg, 0.176 mmol), N,N'-dimethylethylenediamine (16 mg, 0.176 mmol), and K2CO3 (146 mg, 1.056 mmol) were added to CH3CN (4 mL). The reaction mixture was heated to 60°C and stirred for 2 hours. After cooling to room temperature, water (50 mL) was added, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phase was washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, and concentrated. The residue was separated and purified by reverse phase column chromatography (methanol / water = 4:1) to obtain N-(4-(5-(5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (5G) (white solid, 180 mg, 70% yield).

[0275] LC-MS m / z(ESI)=725.2[M+H].

[0276] Step 7: N-(4-(5-(6-(4,4-difluoropiperidin-1-yl)-5-(2-hydroxyethoxy)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 5)

[0277] Dissolve N-(4-(5-(5-(2-(benzyloxy)ethoxy)-6-(4,4-difluoropiperidin-1-yl)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (5G) (180 mg, 0.25 mmol) in MeOH (8 mL), and add palladium / carbon (90 mg, 0.85 mmol). Stir at room temperature under a hydrogen atmosphere for 72 hours. Filter, and concentrate the filtrate. The residue was separated and purified by reverse phase column (methanol / water = 4:1) to give N-(4-(5-(6-(4,4-difluoropiperidin-1-yl)-5-(2-hydroxyethoxy)pyridin-2-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Compound 5) (white solid, 10 mg, 6% yield).

[0278] 1 H NMR (400MHz, DMSO-d6) δ7.86(d,J=8.5Hz,1H),7.80(d,J=8.2Hz,1H),7.51(d,J=8.2Hz,1H),7.07(d,J=1.8Hz,1H),6.97(dd,J=8.5, 1.8Hz,1H),4.97(s,1H),4.18(t,J=4.8Hz,2H),3.86-3.76(m,4H),3.70-3.61(m,4H),3.38(t,J=6.4Hz,2H),2.99-2.90(m,4H),2.22 -2.04(m,4H),1.61-1.49(m,4H),0.32(s,4H).

[0279] LCMS m / z (ESI) = 635.4 [M+H].

[0280] Table 1: Examples 6 to 9 were prepared according to similar procedures as Examples 1 to 5:

[0281] Table 2: NMR of the compounds in Table 1:

[0282] Example 10: N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)-pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)-2-(dihydrophosphinoyloxyethyl)-1-sulfonamide

[0283] Step 1: N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)-pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)-2-(dihydrophosphinoyloxyethyl)-1-sulfonamide (Compound 10)

[0284] Under nitrogen protection, N-(3-(6-azaspiro[2.5]octan-6-yl)-4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 2) (120 mg, 0.21 mmol) was added to acetonitrile (2 mL). The reaction solution was cooled to 0 ° C, pyrophosphoryl chloride (264 mg, 1.05 mmol) was slowly added dropwise, and stirred at room temperature for 2 hours. The reaction solution was poured into ice water (10 mL) and stirred at room temperature overnight. Extracted with DCM / MeOH (10 / 1) (10 mL x 3). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by reverse phase column (methanol / water = 4:1) to give N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)-pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)-2-(dihydrophosphinoyloxyethyl)-1-sulfonamide (Compound 10) (yellow solid, 60 mg, 44% yield).

[0285] 1 H NMR (400MHz, DMSO-d6) δ10.36 (s, 1H), 8.66 (dd, J=5.1, 2.8Hz, 1H), 7.91 (dd, J= 8.5,2.8Hz,1H),7.42(dd,J=5.1,2.8Hz,1H),7.06(d,J=2.8Hz,1H),6.97(dt,J =8.5,2.8Hz,1H),4.18-4.12(m,2H),4.05-3.98(m,4H),3.58(t,J=8.0Hz,2H), 2.96-2.90(m,J=11.5Hz,4H),2.12-2.0(m,4H),1.56-1.46(m,4H),0.30(s,4H)

[0286] LCMS m / z (ESI) = 656.4 [M+H].

[0287] Table 3: Examples 11 to 16 were prepared according to a similar procedure to Example 10:

[0288] Table 4: NMR of compounds in Table 3:

[0289] Example 17: 2-(N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)sulfamoyl)ethyl-L-valine ester hydrochloride (Compound 17)

[0290] Step 1: 2-(N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)sulfamoyl)ethyl(tert-butyloxycarbonyl)-L-valine ester (17A)

[0291] N-(3-(6-azaspiro[2.5]octan-6-yl)-4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 2) (200 mg, 0.35 mmol), N-Boc-L-valine (76 mg, 0.35 mmol), EDCI (74 mg, 0.39 mmol), and DMAP (4 mg, 0.035 mmol) were added to acetonitrile (3 mL) and stirred at room temperature overnight. Water (20 mL) was added, and the mixture was extracted with DCM / MeOH = 10 / 1 (10 mL x 3). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by reverse phase column (methanol / water = 4:1) to give 2-(N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)sulfamoyl)ethyl(tert-butoxycarbonyl)-L-valine ester (17A) (yellow solid, 140 mg, 52% yield).

[0292] LCMS m / z (ESI) = 775.2 [M+H].

[0293] Step 2: 2-(N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)sulfamoyl)ethyl-L-valine ester hydrochloride (Compound 17)

[0294] 2-(N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)sulfamoyl)ethyl(tert-butoxycarbonyl)-L-valine ester (17A) (100 mg, 0.13 mmol) was added to ethyl acetate (3 mL) and a solution of hydrogen chloride in ethyl acetate (1 mL, 4 M). The reaction was stirred at room temperature for 3 hours. The reaction solution was concentrated, added with water and freeze-dried to give 2-(N-(4-(5-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1,3,4-oxadiazol-2-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)sulfamoyl)ethyl-L-valine ester hydrochloride (Compound 17) (yellow solid, 81 mg, 93% yield).

[0295] 1 H NMR (400MHz, DMSO-d6) δ10.56(s,1H),8.70(d,J=4.9Hz,1H),8.47(s,3H),7.95(d,J=8.5Hz ,1H),7.45(d,J=4.9Hz,1H),7.12(d,J=2.1Hz,1H),7.02(dd,J=8.5,2.1Hz,1H),4.60-4.46( m,2H),4.08-4.01(m,4H),3.88-3.83(m,1H),3.72-3.70(m,2H),2.98-2.94(m,4H),2.17-2 .03(m,5H),1.60-1.48(m,4H),0.95(d,J=7.0,Hz,3H),0.93(d,J=7.0,Hz,3H),0.34(s,4H);

[0296] LCMS m / z (ESI) = 710.2 [M+H].

[0297] Example 18: 2-(N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)sulfamoyl)ethyl-L-valine ester hydrochloride (Compound 18)

[0298] Step 1: 2-(N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)sulfamoyl)ethyl(tert-butyloxycarbonyl)-L-valine (18A)

[0299] N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyridin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)-2-hydroxyethane-1-sulfonamide (Intermediate 4) (200 mg, 0.35 mmol), N-Boc-L-valine (76 mg, 0.35 mmol), EDCI (74 mg, 0.39 mmol), and DMAP (4 mg, 0.035 mmol) were added to acetonitrile (2 mL). The reaction mixture was stirred at room temperature overnight. Water (100 mL) was added, and the mixture was extracted with DCM / MeOH = 10 / 1 (20 mL x 3). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by reverse phase column (methanol / water = 4:1) to give 2-(N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)sulfamoyl)ethyl(tert-butoxycarbonyl)-L-valine (18A) (yellow oil, 80 mg, 30% yield).

[0300] LCMS m / z (ESI) = 774.2 [M+H].

[0301] Step 2: 2-(N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)sulfamoyl)ethyl-L-valine ester hydrochloride (Compound 18)

[0302] 2-(N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octan-6-yl)phenyl)sulfamoyl)ethyl(tert-butoxycarbonyl)-L-valine (18A) (80 mg, 0.10 mmol) was dissolved in ethyl acetate (3 mL) and a solution of hydrogen chloride in ethyl acetate (1 mL, 4 M). The reaction was stirred at room temperature for 2 hours. The reaction solution was directly concentrated, added with water and freeze-dried to obtain 2-(N-(4-(1-(2-(4,4-difluoropiperidin-1-yl)pyrimidin-4-yl)-1H-1,2,3-triazol-4-yl)-3-(6-azaspiro[2.5]octyl-6-yl)phenyl)sulfamoyl)ethyl-L-valine ester hydrochloride (Compound 18) (yellow solid, 70 mg, 95% yield).

[0303] 1H NMR (400MHz, DMSO-d6) δ10.28(s,1H),9.48(s,1H),8.66(d,J=5.3Hz,1H),8.53(d ,J=5.5Hz,3H),8.10(d,J=8.5Hz,1H),7.36(d,J=5.3Hz,1H),7.23(d,J=2.1Hz,1H) ,7.11(dd,J=8.5,2.1Hz,1H),4.56-4.43(m,2H),3.85-3.78(m,1H),3.61(t,J=6.1 Hz,2H),2.15-2.01(m,5H),1.68-1.41(m,4H),0.91(d,J=6.9Hz,6H),0.37(s,4H);

[0304] LCMS m / z (ESI) = 711.2 [M+H].

[0305] Biological Example 1 In vitro inhibition test of KIF18A enzyme activity by the compounds of the present invention

[0306] The inhibition of kinesin motility was measured using the ADP-Glo ​​luminescence assay (Promega). The test compound was serially diluted 3-fold in DMSO over a 10-point concentration range using a 384-well plate (Corning). Human KIF18A (1-467) protein (4 nM) was incubated with a 3-fold serial dilution of the test compound or DMSO at room temperature in a reaction buffer (15 mM Tris, pH 7.5, 10 mM MgCl2, 0.01% Pluronic F-68, 2% DMSO, 1 μM paclitaxel, 30 μg / mL polymerized microtubules (purchased from Cytoskeleton Inc)) for less than 30 minutes. 30 μM adenosine triphosphate (ATP) was then added to initiate the enzymatic reaction, and the reaction mixture was incubated at room temperature for 15 minutes, after which the ADP-Glo ​​reagent was added according to the kit instructions. The luminescence intensity, which was proportional to the presence of ADP, was measured using a microplate reader. Inhibition rate (%) = 100-100 × (sample signal - negative control signal) ÷ ​​(positive control signal - negative control signal). The positive control (enzyme + substrate) and the negative control (substrate only) had the same concentration of DMSO. Curve fitting and IC were determined using GraphPad Prism 7 with a 3-parameter nonlinear regression equation (variable slope). 50 Value, where IC 50 The concentration of the compound at which KIF18A activity is inhibited by 50%. The results are shown in Table 5 below.

[0307] Table 5 Inhibitory activity of the compounds of the present invention on KIF18A

[0308] Biological Example 2 In vitro antiproliferative activity of the compounds of the present invention on OVCAR3 cells

[0309] 90 μL of a suspension of OVCAR-3 tumor cells (2000 cells per well) was seeded into a 96-well plate. The plate was then placed in a CO2 incubator for 24 hours to allow the cells to fully adhere. The next day, the compound was dissolved in DMSO and a 2 mM stock solution was prepared. The compound was serially diluted 3-fold with DMSO to 10 different concentrations. 5 μL of this serially diluted compound solution was transferred to a 96-well plate containing 95 μL of culture medium and mixed thoroughly to obtain a 10× compound stock solution. 10 μL of this 10× compound solution was then transferred to a 96-well plate seeded with 90 μL of cells to obtain a final 1× concentration. The final DMSO concentration was 0.5%. The plate was gently shaken to thoroughly mix the drug, and the cell culture plate was placed back in a CO2 incubator for another 4 days. On the 4th day, CellTiter-Glo reagent (25 μL) was added to each test well, mixed on a shaker for 2 minutes, and then incubated at room temperature for 25 minutes. Luminescence intensity was measured using an ID5 multi-function microplate reader. Inhibition rate (%) = 100-100 × (sample signal - negative control signal) ÷ ​​(positive control signal - negative control signal). The positive control (cells + CTG) and negative control (medium + CTG) had the same concentration of DMSO. Curve fitting and IC were determined using GraphPad Prism 7 with a 3-parameter nonlinear regression equation (variable slope). 50 The results are shown in Table 6 below.

[0310] Table 6 Antiproliferative activity of the preferred compounds of the present invention on OVCAR3 cells

[0311] Biological Example 3 Pharmacokinetic Study in Mice

[0312] Instrument: Liquid chromatography-mass spectrometry (Shimadzu liquid chromatography, API 3000 mass spectrometer, AB). All data were collected and processed by Analyst 1.4.2 software, and calculated and processed by Microsoft Excel. Pharmacokinetic parameters were calculated using WinNonlin 5.2 software and NCA. The main kinetic parameters included T max 、T 1 / 2 、C max , AUC 0-24h Chromatographic column: Venusil XBP Phenyl (4.6◇50 mm, 5 μm); column temperature: 25°C; mobile phase A: water (0.1% formic acid), mobile phase B: methanol, flow rate: 1 mL / min, split ratio: 1:5, isocratic elution, mobile phase B: 85%; injection volume: 5 μL.

[0313] Animals: 30 ICR male mice, weighing 18-22 g, were housed in the laboratory for four days prior to use. They were fed 12 hours before and four hours after dosing and had free access to water during the experiment. Blood samples were collected at designated time points after gavage.

[0314] Solvent: 10% sulfobutylcyclodextrin solution (pH = 4.6) in 50 mM citric acid buffer. Preparation of oral administration solution: Accurately weigh the compound, add it to the solvent, and sonicate at room temperature until the drug is dissolved or a uniform suspension is obtained, to prepare a 1 mg / ml solution.

[0315] Blood was collected from the jugular venous plexus at 0.083, 0.25, 0.5, 1, 2, 4, 6, 8, and 24 hours after oral administration. Fifty μL of plasma sample was added to 250 μL of methanol solution containing the internal standard. The sample was vortexed for 1 minute and centrifuged at 15,000 rpm for 5 minutes. The supernatant was then analyzed by LC-MS / MS.

[0316] The compound was accurately weighed and prepared at various concentrations. Quantitative analysis was performed on a mass spectrometer to establish a standard curve. The plasma concentration of the compound was then measured to determine compound concentrations at different time points. All measured data were collected and processed using relevant software, and pharmacokinetic parameters were fitted using a non-compartmental model. The results are shown in Table 7 below.

[0317] Table 7 Pharmacokinetic parameters of the compounds of the present invention in mice

[0318] Biological Example 4 In vivo efficacy study - mouse OVCAR3 subcutaneous transplant tumor model

[0319] 6x106 OVCAR3 cells were subcutaneously inoculated on the right back of female BALB / c nude mice. When the tumor grew to 150mm3, the animals were randomly divided into groups and gavage-administered. Group 1: vehicle control group; Group 2: compound 11 (3 mg / kg, QD); Group 3: compound 11 (10 mg / kg, QD); Group 4: compound 11 (30 mg / kg, QD); Group 5: compound 11 (100 mg / kg, QD). Tumor volume was measured twice a week and at the end of dosing. The tumor growth inhibition rate of the compound was calculated according to the tumor growth inhibition rate (TGI) = 1-(tumor volume of the dosing group on day 28 - tumor volume of the dosing group on day 0) / (tumor volume of the vehicle control group on day 28 - tumor volume of the dosing group on day 0). The results are shown in Table 8.

[0320] Table 8 Inhibitory effect of the compounds of the present invention on the growth of OVCAR3 subcutaneous transplanted tumors in mice

Claims

1. A compound represented by formula (I) or its stereoisomers, tautomers, pharmaceutically acceptable salts or deuterated derivatives: in: X 1 is C or N; X 3 is C or N; X 2 , X 4 and X 5 are each independently selected from N, O, NH or CH; W 1 N or CR 2 ; R 1 is a 6-membered heterocycloalkyl group, the 6-membered heterocycloalkyl group contains 0, 1 or 2 N atoms and 0 or 1 O atoms, and further, the 6-membered heterocycloalkyl group is substituted by 0, 1, 2 or 3 groups selected from the following: F, Cl, Br, C 1-3 alkyl; R 2 For hydrogen, halogen, CN, C 1-8 Alkyl, C 1-4 Haloalkyl, C 0-4 Alkyl-OR a , C 0-4 Alkyl-NR a R a ; R 3 For hydrogen, C 1-4 Alkyl, C 1-4 Haloalkyl, C 0-4 Alkyl-OR a , C 0-4 Alkyl-NR a R a ; R 4 and R 5 are independently selected from hydrogen or C 1-3 Alkyl; or R 4 , R 5 connected to form a cyclopropyl group; or R 4 , R 5 The carbon atom to which it is connected together forms C=CR j R k ; Among them, R j and R k each independently selected from the group consisting of: hydrogen, halogen; When W 1 When N, R 6 is -OCH2OP(=O)(OH)2, -OP(=O)(OH)2, -OP(=O)(OH)C 1-5 Alkyl, -OC(=O)C 1-5 Alkyl, wherein the C 1-5 The alkyl group is substituted with 0, 1, 2 or 3 substituents selected from the following: OH, NH2, OCH3, or a 5-membered, 6-membered or 7-membered heterocycloalkyl group containing 0, 1 or 2 N atoms and 0 or 1 O atom; When W 1 CR 2 When R 6 is -OH, -OCH2OP(=O)(OH)2, -OP(=O)(OH)2, -OP(=O)(OH)C 1-5 Alkyl, -OC(=O)C 1-5 Alkyl, wherein the C 1-5 The alkyl group is substituted with 0, 1, 2 or 3 substituents selected from the following: OH, NH2, OCH3, or a 5-, 6- or 7-membered group containing 0, 1 or 2 N atoms and 0 or 1 O atom. Heterocycloalkyl; R a is independently H or R b ; and R b In each case independently C 1-6 Alkyl, wherein the C 1-6 The alkyl group is substituted with 0, 1, 2 or 3 substituents selected from the group consisting of halogen, -OH, -OC 1-4 Alkyl, -NH2, -NHC 1-4 Alkyl and -N(C 1-4 Alkyl)C 1-4 alkyl.

2. The compound according to claim 1 or its stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, W 1 N, R 6 -OP(=O)(OH)2 or 3. The compound according to claim 1 or its stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, W 1 CR 2 , where R 2 -OCH3; R 6 is -OH, -OP(=O)(OH)2 or 4. The compound according to any one of claims 1 to 3, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, wherein R 6 It is -OP(=O)(OH)2.

5. The compound according to any one of claims 1 to 4, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, wherein the group Selected from in, The side with * is the side connected to the benzene ring.

6. The compound according to any one of claims 1 to 5, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, wherein the group Selected from in, The side with * is the side connected to the benzene ring.

7. The compound according to any one of claims 1 to 6, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, wherein the group for 8. The compound according to any one of claims 1 to 7, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, wherein R 1 Selected from 9. The compound according to any one of claims 1 to 8, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, wherein R 1 for 10. The compound according to any one of claims 1 to 9, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, wherein R 3 Selected from hydrogen, 11. The compound according to any one of claims 1 to 10, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, wherein R 3 For hydrogen.

12. The compound according to any one of claims 1 to 11, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, wherein the compound is selected from the group consisting of:

13. The compound according to any one of claims 1 to 12, or a stereoisomer, a tautomer, a pharmaceutically acceptable salt or a deuterated derivative thereof, and a pharmaceutically acceptable carrier, an excipient or a combination thereof.

14. A method for treating a condition treatable with a KIF18A inhibitor, the method comprising administering to a patient in need thereof a therapeutically effective amount of a compound according to any one of claims 1 to 12, or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a composition according to claim 13.

15. The method of claim 14, wherein the condition is a cancer selected from the group consisting of: (a) a solid tumor or a hematogenic tumor selected from the group consisting of bladder cancer, endometrial cancer, squamous cell lung cancer, breast cancer, colon cancer, kidney cancer, liver cancer, lung cancer, small cell lung cancer, esophageal cancer, gallbladder cancer, brain cancer, head and neck cancer, ovarian cancer, pancreatic cancer, gastric cancer, cervical cancer, thyroid cancer, prostate cancer, and skin cancer; (b) a hematopoietic tumor of the lymphoid lineage selected from the group consisting of leukemia, acute lymphocytic leukemia, acute lymphoblastic leukemia, B-cell lymphoma, T-cell lymphoma, and leukemia. (c) a hematopoietic neoplasm of the myeloid lineage selected from the group consisting of acute and chronic myeloid leukemias, myelodysplastic syndromes and promyelocytic leukemias; (d) a tumor of mesenchymal origin selected from the group consisting of fibrosarcoma and rhabdomyosarcoma; (e) a tumor of the central and peripheral nervous system selected from the group consisting of astrocytoma, neuroblastoma, glioma and schwannoma; or (f) a melanoma, a seminoma, a teratoma, an osteosarcoma, a xeroderma pigmentosum, a keratoacanthoma, a follicular thyroid carcinoma or a Kaposi's sarcoma.

16. A method of reducing the size of a solid tumor in a subject, the method comprising administering to a subject in need thereof a therapeutically effective amount of a compound according to any one of claims 1 to 12, or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a composition according to claim 13.

17. A method for treating a cell proliferative disorder in a subject, the method comprising administering to a subject in need thereof a therapeutically effective amount of a compound according to any one of claims 1 to 12, or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a composition according to claim 13.

18. A method of inhibiting KIF18A in a cell, the method comprising treating the cell with a compound according to any one of claims 1 to 12, or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a composition according to claim 13.

19. Use of a compound according to any one of claims 1 to 12 or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a composition according to claim 13, in the preparation of a medicament for a condition treatable with a KIF18A inhibitor.

20. The use of a compound according to claim 19 or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, wherein the disease is a cancer selected from the group consisting of: (a) a solid tumor or a hematological tumor selected from the following cancers: bladder cancer, endometrial cancer, squamous cell lung cancer, breast cancer, colon cancer, kidney cancer, liver cancer, lung cancer, small cell lung cancer, esophageal cancer, gallbladder cancer, brain cancer, head and neck cancer, ovarian cancer, pancreatic cancer, gastric cancer, cervical cancer, thyroid cancer, prostate cancer and skin cancer; (b) a hematopoietic tumor of the lymphoid system selected from the following: leukemia, acute lymphoblastic leukemia, acute lymphoblastic leukemia, (c) a hematopoietic neoplasm of the myeloid lineage selected from the group consisting of acute and chronic myeloid leukemias, myelodysplastic syndromes and promyelocytic leukemias; (d) a tumor of mesenchymal origin selected from the group consisting of fibrosarcoma and rhabdomyosarcoma; (e) a tumor of the central and peripheral nervous system selected from the group consisting of astrocytoma, neuroblastoma, glioma and schwannoma; or (f) a melanoma, a seminoma, a teratoma, an osteosarcoma, a xeroderma pigmentosum, a keratoacanthoma, a follicular thyroid carcinoma or a Kaposi's sarcoma.

21. Use of a compound according to any one of claims 1 to 12, or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a composition according to claim 13, in the preparation of a medicament for reducing the size of a solid tumor in a subject.

22. Use of a compound according to any one of claims 1 to 12, or a stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or a pharmaceutical composition according to claim 13, in the preparation of a medicament for treating a cell proliferative disorder in a subject.

23. Use of the compound according to any one of claims 1 to 12 or its stereoisomer, tautomer, pharmaceutically acceptable salt or deuterated derivative thereof, or the pharmaceutical composition according to claim 13 in the preparation of a medicament for inhibiting KIF18A in a cell.

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