WO2025130971A1 - Fused ring compound, pharmaceutical composition comprising same, and use thereof - Google Patents

Abstract

The present invention relates to a compound of formula (I), a pharmaceutical composition comprising same, and a use of the compound in the prevention or treatment of diseases.

Description

Condensed ring compound, pharmaceutical composition containing the same and use thereof Field of the Invention

The present invention relates to a condensed ring compound, a pharmaceutical composition containing the condensed ring compound, and use thereof for preventing or treating diseases.

Background of the Invention

Microsatellite instability (MSI) is a type of genomic damage caused by deficient mismatch repair (dMMR). Despite advances in the treatment of microsatellite instability-high (MSI-H) cancers, tumor evolution and drug resistance remain the main causes of treatment failure and death in cancer patients. For example, approximately half of patients with dMMR colorectal cancer treated with PD-1 and PD-L1 checkpoint inhibitors experience primary drug resistance. See, e.g., Overman MJ et al., J Clin Oncol. 2018 Mar 10;36(8):773-779. There remains an unmet clinical need for new treatment options for patients who are refractory to currently available therapies.

Werner helicase (WRN) is considered a synthetic lethal target for MSI-H cancers (see, e.g., Chan EM et al., Nature. 2019 Apr; 568(7753): 551-556). WRN is a member of the RecQ family of DNA helicases and plays an important role in maintaining genome stability, DNA repair, replication, transcription, and telomere maintenance. Studies on WRN-dependent mechanisms have shown that dinucleotide TA repeats are massively amplified in MSI cells. These amplified TA repeats form secondary DNA structures that require WRN helicases to unwind (see, e.g., van Wietmarschen N et al., Nature. 2020 Oct; 586(7828): 292-298). In the absence of WRN (or when WRN helicases are inhibited), amplified TA repeats in MSI cells are cleaved by nucleases, ultimately leading to chromosome breaks. Therefore, inhibition of WRN helicase is an effective strategy for the treatment of cancers characterized by microsatellite instability-high (MSI-H) or mismatch repair deficiency (dMMR), including colorectal, gastric, or endometrial cancer.

SUMMARY OF THE INVENTION

The present application provides compounds used as WRN inhibitors, which can be used to prevent or treat cancers characterized by microsatellite high instability (MSI-H) or mismatch repair deficiency (dMMR). In addition, the compounds of the present invention also have excellent properties such as good physicochemical properties (such as solubility, physical and/or chemical stability), good pharmacokinetic properties (such as improved bioavailability, good metabolic stability, suitable half-life and duration of action), good safety (lower toxicity (such as reduced cardiotoxicity) and/or less side effects), less prone to drug resistance.

One aspect of the present invention provides a compound or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, wherein the compound has a structure of formula (I):

in:

Selected from

W is CR ¹ or N;

^R1 at each occurrence is independently selected from H, halogen, -OH, _-NH2 , -CN, _-NO2 , _-SF5 , _C1-6 alkyl, deuterated _C1-6 alkyl, halogenated _C1-6 alkyl, _C2-6 alkenyl, _C2-6 alkynyl, _C3-6 cycloalkyl, 3-10 membered heterocyclyl, _C6-10 aryl, 5-14 membered heteroaryl, _C6-12 aralkyl, -C(=O) ^Ra , -OC(=O) ^Ra , -C(=O) ^ORa , ^-ORa , ^-SRa , -S(=O) ^Ra , -S(=O) _2Ra , ^-S (=O ^{)2NRaRb , -NRaRb} , -C ₍ =O) ^NRaRb , -NRa ^{- C} (=O) ^{Rb , -NRa} -C(═O)OR ^b , -NR ^a -S(═O) ₂ -R ^b , -NR ^a -C(═O)-NR ^a R ^b , -P(═O)R ^a R ^b , -C _1-6 alkylene-R ^a , -C _1-6 alkylene-OR ^a , -C _1-6 alkylene-NR ^a R ^b , -OC _1-6 alkylene-NR ^a R ^b , (—C _3-6 cycloalkylene)-CN and (—C _3-6 cycloalkylene)-C _1-6 haloalkyl;

Alternatively, two R ^1s in the ortho position together with the groups to which they are attached optionally together constitute a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring;

^R2 is

R ³ , R ²¹ , R ²² , R ²³ and R ²⁴ are each independently selected at each occurrence from H, halogen, -OH, -NH ₂ , -CN, -NO ₂ , -SF ₅ , C _1-6 alkyl, deuterated C _1-6 alkyl, halogenated C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, C _6-12 aralkyl, -C(═O)R ^{a , -OC(═O)R a ,} -C(═O)OR ^a , -OR ^a , -SR ^a , -S(═O)R ^a , -S(═O) ₂ R ^a , -S(═O) ₂ NR ^a R ^b , -S(═O)(═NR ^a )R ^b , -NR ^a R ^b , —C(═O)NR ^a R ^b , —NR ^a -C(═O)R ^b , —NR ^a -C(═O)R b , —NR ^a -C(═O)OR ^b , —NR ^a -S(═O) ₂ -R ^b , —NR ^{a -C} (═O)-NR ^a R ^b , —P(═O)R a R b , —C _1-6 alkylene-R ^a , —C _1-6 alkylene-OR ^a , —C _1-6 alkylene-NR ^a R ^b , —OC _1-6 alkylene-NR ^a R ^b , (—C _3-6 cycloalkylene)-CN and (—C _3-6 cycloalkylene)-C _1-6 haloalkyl;

Alternatively, R ³ and R ²¹ or R ²² together with the group to which they are attached optionally together form a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring;

^R4 is

R ⁴¹ is selected from a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring, and a 5-14 membered heteroaromatic ring;

Ring X and Ring Z are each independently selected from a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring, and a 5-14 membered heteroaromatic ring;

Ring Y is absent or is selected from a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring and a 5-14 membered heteroaromatic ring; when Ring Y is absent, R ²⁴ is also absent;

^L2 is selected from -O-, -C(=O)-, -NRC(=O)-, -S-, -S(=O)-, -S(=O) _2- , _C1-6 alkylene and -O-( _C1-6 alkylene)-;

R, ^Ra and ^Rb are each independently selected at each occurrence from H, _C1-6 alkyl, _C3-10 cycloalkyl, 3-10 membered heterocyclyl, _C6-10 aryl, 5-14 membered heteroaryl and _C6-12 aralkyl;

The above alkylene, alkyl, alkenyl, alkynyl, cycloalkylene, cycloalkyl, hydrocarbon ring, heterocyclyl, heterocycle, aryl, aromatic ring, heteroaryl, heteroaromatic ring and aralkyl are each optionally substituted at each occurrence by one or more substituents independently selected from the following: deuterium atom, halogen, -OH, =O, -NH2, -CN, _-NO2 , = _CH2 , _C1-6 alkyl, deuterated _{C1-6 alkyl} , _C2-6 alkenyl, _C2-6 alkynyl, _C3-6 cycloalkyl, 3-10 membered heterocyclyl, _C6-10 aryl, 5-14 membered heteroaryl, _C6-12 aralkyl, -C(=O) ^Rc , -OC(=O) ^Rc , -C(=O) ^ORc , ^-ORc , ^-SRc , -S(=O) ^Rc , -S(= ^O ) _2Rc -S(＝O) _2NRcRd , ^-NRcRd , ^-C (＝O) ^NRcRd , ^-NRc- C( ^＝ O) ^Rd , -NRc ^- C(＝O) ^ORd , -NRc ^- S(＝O) ₂ - ^Rd , -NRc ^{- C} (＝O)-NRcRd, -C1-6alkylene ^{- ORc} , _{-C1-6alkylene} - ^NRcRd and _{-OC1-6alkylene} - ^NRcRd ; when the same ^ring atom or adjacent ring atoms of a cycloalkylene, ^cycloalkyl , hydrocarbon ^ring , _heterocyclyl , heterocycle, ^aryl , aromatic ring, heteroaryl and heteroaromatic ring are substituted by two substituents, the two substituents together with the group to which they are attached optionally constitute a _C3-6 hydrocarbon ring, a 3-10 membered heterocycle, a C1-6alkylene ring or a heteroaryl ring. _6-10 membered aromatic ring or 5-14 membered heteroaromatic ring; each of the alkylene, alkyl, alkenyl, =CH ₂ , alkynyl, cycloalkyl, hydrocarbon ring, heterocyclyl, heterocycle, aryl, aromatic ring, heteroaryl, heteroaromatic ring and aralkyl is further optionally substituted by one or more substituents independently selected from the following: halogen, -OH, =O, -C(=O)O-tert-butyl, -NH ₂ , -CN, -NO ₂ , C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, C _6-12 aralkyl, -C _1-6 alkylene-C _3-6 cycloalkyl, -OC _1-6 alkyl and -C _1-6 alkylene-OC _1-6 alkyl;

R ^c and R ^d are each independently selected at each occurrence from H, C _1-6 alkyl, C _3-10 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, and C _6-12 aralkyl, said alkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and aralkyl being further optionally substituted with one or more substituents independently selected from the group consisting of halogen, -OH, =O, -C(=O)O-tert-butyl, -NH ₂ , -CN, -NO ₂ , C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, C _6-12 aralkyl _{, and -C 1-6 alkylene} -OC _1-6 alkyl; and

p, q and t are each independently an integer selected from 1, 2 or 3.

Another aspect of the present invention provides a pharmaceutical composition comprising a prophylactically or therapeutically effective amount of a compound of the present invention or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof and one or more pharmaceutically acceptable carriers.

Another aspect of the present invention provides use of a compound of the present invention or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, or a pharmaceutical composition of the present invention in the preparation of a medicament for use as a WRN inhibitor.

Another aspect of the present invention provides a compound of the present invention or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, or a pharmaceutical composition of the present invention, for use as a WRN inhibitor.

Another aspect of the present invention provides a method for preventing or treating cancer (preferably a cancer characterized by microsatellite high instability (MSI-H) or mismatch repair deficiency (dMMR)), which comprises administering to an individual in need thereof an effective amount of a compound of the present invention or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, or a pharmaceutical composition of the present invention.

DETAILED DESCRIPTION OF THE INVENTION

definition

Unless otherwise defined below, the meanings of all technical terms and scientific terms used herein are intended to be the same as those generally understood by those skilled in the art. Reference to the technology used herein is intended to refer to the technology generally understood in the art, including those changes in technology or replacement of equivalent technology that are obvious to those skilled in the art. Although it is believed that the following terms are well understood by those skilled in the art, the following definitions are still set forth to better explain the present invention.

The terms "comprises," "comprising," "having," "containing," or "involving" and other variations thereof herein are inclusive or open-ended and do not exclude additional unrecited elements or method steps.

As used herein, the term "alkylene" refers to a saturated divalent hydrocarbon group, preferably a saturated divalent hydrocarbon group having 1, 2, 3, 4, 5 or 6 carbon atoms, such as methylene, ethylene, propylene or butylene.

As used herein, the term "alkyl" is defined as a straight or branched saturated aliphatic hydrocarbon. In some embodiments, the alkyl has 1 to 12, for example 1 to 6 carbon atoms. For example, as used herein, the term "C _1-6 alkyl" refers to a linear or branched group of 1 to 6 carbon atoms (e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl or n-hexyl), which is optionally substituted by 1 or more (such as 1 to 3) suitable substituents such as halogen (in this case, the group is referred to as "haloalkyl") (e.g., CF ₃ , C ₂ F ₅ , CHF ₂ , CH ₂ F, CH ₂ CF ₃ , CH ₂ Cl or -CH ₂ CH ₂ CF _3, etc.). The term "C _1-4 alkyl" refers to a linear or branched aliphatic hydrocarbon chain of 1 to 4 carbon atoms (ie, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl or tert-butyl).

As used herein, the term "alkenyl" means a linear or branched monovalent hydrocarbon group containing one or more double bonds and having 2 to 6 carbon atoms (" _C2-6 alkenyl"). The alkenyl group is, for example, -CH= _CH2 , _-CH2CH = _CH2 , -C( _CH3 )= _CH2 , _-CH2- CH=CH- _CH3 , 2-pentenyl, 3-pentenyl, 4-pentenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 5-hexenyl, 2-methyl-2-propenyl and 4-methyl-3-pentenyl. When the compound of the present invention contains an alkenyl group, the compound may exist in the pure E (entgegen) form, the pure Z (zusammen) form or any mixture thereof. The term "alkenylene" is a corresponding divalent group, including, for example, "C _2-6 alkenylene", "C _2-4 alkenylene", etc., and specific examples include, but are not limited to: -CH=CH-, -CH ₂ CH=CH-, -C(CH ₃ )=CH-, butenylene, pentenylene, hexenylene, etc.

As used herein, the term "alkynyl" refers to a monovalent hydrocarbon group containing one or more triple bonds, preferably having 2, 3, 4, 5 or 6 carbon atoms, such as ethynyl, 2-propynyl, 2-butynyl, 1,3-butadiynyl, etc. The alkynyl group is optionally substituted with one or more (such as 1 to 3) identical or different substituents. The term "alkynylene" is a corresponding divalent group, including, for example, "C _2-8 alkynylene", "C _2-6 alkynylene", "C _2-4 alkynylene", etc. Examples thereof include, but are not limited to The alkynylene group is optionally substituted with one or more (such as 1 to 3) identical or different substituents.

As used herein, the term "paracyclic ring" or "fused ring" refers to a ring system formed by two or more cyclic structures sharing two adjacent atoms with each other.

As used herein, the term "spirocycle" refers to a ring system formed by two or more cyclic structures that share one ring atom with each other.

As used herein, the term "bridged ring" refers to a ring system formed by two or more cyclic structures sharing two atoms that are not directly connected to each other.

As used herein, the terms "cycloalkylene", "cycloalkyl" and "hydrocarbon ring" refer to saturated (i.e., "cycloalkylene" and "cycloalkyl") or partially unsaturated (i.e., having one or more double bonds and/or triple bonds within the ring) monocyclic or polycyclic hydrocarbon rings (including spiro, fused or bridged ring systems) having, for example, 3-10 (suitably 3-8, more suitably 3-6) ring carbon atoms, including but not limited to (cyclo)propyl (ring), (cyclo)butyl (ring), (cyclo)pentyl (ring), (cyclo)hexyl (ring), (cyclo)heptyl (ring), (cyclo)octyl (ring), (cyclo)nonyl (ring), (cyclo)hexenyl (ring) and the like.

As used herein, the term "cycloalkyl" refers to a saturated monocyclic or polycyclic (such as bicyclic) hydrocarbon ring (e.g., a monocyclic ring such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, or a bicyclic ring, including a spirocyclic, fused or bridged system (such as bicyclo[1.1.1]pentyl, bicyclo[2.2.1]heptyl, bicyclo[3.2.1]octyl or bicyclo[5.2.0]nonyl, decalinyl, etc.), which is optionally substituted with 1 or more (such as 1 to 3) suitable substituents. The cycloalkyl group has 3 to 15 carbon atoms. For example, the term " _{C3-6cycloalkyl} " refers to a saturated monocyclic or polycyclic (such as bicyclic) hydrocarbon ring of 3 to 6 ring carbon atoms (e.g., cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl), which is optionally substituted with 1 or more (such as 1 to 3) suitable substituents, for example, methyl-substituted cyclopropyl.

As used herein, the term "heterocyclyl" (or "heterocycle") refers to a saturated or partially unsaturated monocyclic or bicyclic group having 2, 3, 4, 5, 6, 7, 8, 9 or 10 carbon atoms and one or more (e.g., one, two, three or four) heteroatoms selected from O, S, N and P in the ring, and the "heterocyclyl" (or "heterocycle") may contain -C(=O)- as a ring member. The heterocyclyl may be attached to the rest of the molecule via the carbon atoms and/or heteroatoms (if present). In particular, a 3-10 membered heterocyclyl group is a group having 3-10 carbon atoms and heteroatoms in the ring, such as, but not limited to, an oxirane, an aziridinyl, an azetidinyl, an oxetanyl, a tetrahydrofuranyl, a dioxolinyl, a pyrrolidinyl, a pyrrolidonyl, an imidazolidinyl, a pyrazolidinyl, a pyrrolinyl, a tetrahydropyranyl, a piperidinyl, a morpholinyl, a dithianyl, a thiomorpholinyl, a piperazinyl or a trithianyl.

As used herein, the term "heterocyclyl" (or "heterocycle") encompasses fused ring structures, where the point of attachment to other groups can be on either ring of the fused ring structure. Therefore, the heterocyclic group of the present invention also includes, but is not limited to, heterocyclic and heterocyclic groups, heterocyclic and cycloalkyl groups, monoheterocyclic and monoheterocyclic groups, monoheterocyclic and monocycloalkyl groups, aryl and heterocyclic groups, heteroaryl and heterocyclic groups, such as 3-7 membered (mono) heterocyclic groups and 3-7 membered (mono) heterocyclic groups, 3-7 membered (mono) heterocyclic groups and (mono) cycloalkyl groups, 3-7 membered (mono) heterocyclic groups and C _4-6 (mono) cycloalkyl groups, C _6-10 aryl and 3-7 membered heterocyclic groups, 5-6 membered heteroaryl and 3-7 membered heterocyclic groups, examples of which include, but are not limited to, pyrrolidinyl and cyclopropyl, cyclopentyl and aziridine, pyrrolidinyl and cyclobutyl, pyrrolidinyl and pyrrolidinyl, pyrrolidinyl and piperidinyl, pyrrolidinyl and piperazinyl, piperidinyl and morpholinyl,

As used herein, the term "heterocyclyl" (or "heterocycle") encompasses bridged heterocyclyls (bridged heterocycle) and spiro heterocyclyls (spiro heterocycle).

As used herein, the term "bridged heterocycle" refers to a cyclic structure containing one or more (e.g., 1, 2, 3 or 4) heteroatoms (e.g., oxygen atoms, nitrogen atoms and/or sulfur atoms) formed by two rings sharing two ring atoms that are not directly connected, including but not limited to 7-10 membered bridged heterocycles, 8-10 membered bridged heterocycles, 7-10 membered nitrogen-containing bridged heterocycles, 7-10 membered oxygen-containing bridged heterocycles, 7-10 membered sulfur-containing bridged heterocycles, etc., for example The "nitrogen-containing bridged heterocycle", "oxygen-containing bridged heterocycle" and "sulfur-containing bridged heterocycle" optionally further contain one or more other heteroatoms selected from oxygen, nitrogen and sulfur.

As used herein, the term "spiroheterocycle" refers to a cyclic structure containing one or more (e.g., 1, 2, 3, or 4) heteroatoms (e.g., oxygen atoms, nitrogen atoms, sulfur atoms) formed by two or more rings sharing a ring atom, including but not limited to 5-10 membered spiroheterocycle, 6-10 membered spiroheterocycle, 6-10 membered nitrogen-containing spiroheterocycle, 6-10 membered oxygen-containing spiroheterocycle, 6-10 membered sulfur-containing spiroheterocycle, etc., for example The "nitrogen-containing spiro heterocycle", "oxygen-containing spiro heterocycle" and "sulfur-containing spiro heterocycle" optionally further contain one or more other heteroatoms selected from oxygen, nitrogen and sulfur. The term "6-10 membered nitrogen-containing spiro heterocyclic group" refers to a spiro heterocyclic group containing a total of 6-10 ring atoms and at least one of the ring atoms being a nitrogen atom.

As used herein, the terms "(ylidene)aryl" and "aromatic ring" refer to an all-carbon monocyclic or fused-ring polycyclic aromatic group having a conjugated π electron system. For example, as used herein, the terms "C _6-10 (ylidene)aryl" and "C _6-10 aromatic ring" mean an aromatic group containing 6 to 10 carbon atoms, such as (ylidene)phenyl (benzene ring) or (ylidene)naphthyl (naphthalene ring). The (ylidene)aryl group and the aromatic ring are optionally substituted with 1 or more (such as 1 to 3) suitable substituents (e.g., halogen, -OH, -CN, -NO ₂ , C _1-6 alkyl, etc.).

The term "aralkyl" refers to an alkyl substituted with an aryl group, wherein the aryl group and the alkyl group are as defined herein. Typically, the aryl group may have 6-14 carbon atoms, and the alkyl group may have 1-6 carbon atoms. Exemplary aralkyl groups include, but are not limited to, benzyl, phenylethyl, phenylpropyl, phenylbutyl.

As used herein, the terms "heteroaryl(ene)" and "heteroaromatic ring" refer to a monocyclic, bicyclic or tricyclic aromatic ring system having 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14 ring atoms, in particular 1 or 2 or 3 or 4 or 5 or 6 or 9 or 10 carbon atoms, and which contains at least one heteroatom which may be identical or different (the heteroatom being, for example, oxygen, nitrogen or sulfur) and, in each case additionally may be benzo-fused. In particular, "(ene)heteroaryl" or "heteroaryl ring" is selected from (ene)thienyl (ring), (ene)furanyl (ring), (ene)pyrrolyl (ring), (ene)oxazolyl (ring), (ene)thiazolyl (ring), (ene)imidazolyl (ring), (ene)pyrazolyl (ring), (ene)isoxazolyl (ring), (ene)isothiazolyl (ring), (ene)oxadiazolyl (ring), (ene)triazolyl (ring), (ene)thiadiazolyl (ring), etc., and their benzo derivatives; or (ene)pyridinyl (ring), (ene)pyridazinyl (ring), (ene)pyrimidinyl (ring), (ene)pyrazinyl (ring), (ene)triazinyl (ring), etc., and their benzo derivatives.

As used herein, the term "halo" or "halogen" group is defined to include F, Cl, Br, or I.

As used herein, the term "alkylthio" refers to an alkyl group as defined above attached to the parent molecular moiety through a sulfur atom. Representative examples of C _1-6 alkylthio include, but are not limited to, methylthio, ethylthio, tert-butylthio and hexylthio.

As used herein, the term "nitrogen-containing heterocycle" refers to a saturated or partially unsaturated monocyclic or bicyclic group having 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 or 13 carbon atoms and at least one nitrogen atom in the ring, which may also optionally contain one or more (e.g., one, two, three or four) ring members selected from N, O, S, S=O and S(=O) ₂ ; the nitrogen-containing heterocycle is connected to the rest of the molecule via any ring member. The nitrogen-containing heterocycle is preferably a saturated nitrogen-containing monocyclic ring. In particular, the 3- to 14-membered nitrogen-containing heterocycle is a group having 3-14 carbon atoms and heteroatoms (at least one of which is a nitrogen atom) in the ring, including but not limited to a three-membered nitrogen-containing heterocycle (such as aziridine), a four-membered nitrogen-containing heterocycle (such as azetidinyl), a five-membered nitrogen-containing heterocycle (such as pyrrolyl, pyrrolidinyl (pyrrolidine ring), pyrrolinyl, pyrrolidonyl, imidazolyl, imidazolidinyl, imidazolinyl, pyrazolyl, pyrazolinyl), a six-membered nitrogen-containing heterocycle (such as piperidinyl (piperidine ring), morpholinyl, thiomorpholinyl, piperazinyl), a seven-membered nitrogen-containing heterocycle, and the like.

The term "substituted" means that one or more (e.g., one, two, three, or four) hydrogens on the designated atom are replaced by a selection from the indicated group, provided that the normal valence of the designated atom in the present context is not exceeded and the substitution forms a stable compound. Combinations of substituents and/or variables are permitted only if such combinations form stable compounds.

If a substituent is described as being "optionally substituted," the substituent may be (1) unsubstituted or (2) substituted. If a carbon of a substituent is described as being optionally substituted with one or more of the listed substituents, then one or more hydrogens on the carbon (to the extent of any hydrogens present) may be replaced, individually and/or together, with independently selected optional substituents. If a nitrogen of a substituent is described as being optionally substituted with one or more of the listed substituents, then one or more hydrogens on the nitrogen (to the extent of any hydrogens present) may each be replaced with an independently selected optional substituent.

If substituents are described as being "independently selected" from a group, each substituent is selected independently of the other. Thus, each substituent may be the same as or different from another (other) substituent.

As used herein, the term "one or more" means 1 or more than 1, such as 2, 3, 4, 5 or 10, where reasonable.

Unless otherwise indicated, as used herein, the point of attachment of a substituent may be from any suitable position of the substituent.

When a bond to a substituent is shown to pass through a bond connecting two atoms in a ring, then such substituent may be bonded to any ring atom in the substitutable ring.

The present invention also includes all pharmaceutically acceptable isotopically labeled compounds which are identical to the compounds of the present invention except that one or more atoms are replaced by an atom having the same atomic number but an atomic mass or mass number different from the atomic mass or mass number prevalent in nature. Examples of isotopes suitable for inclusion in the compounds of the present invention include, but are not limited to, isotopes of hydrogen (e.g., deuterium (D, ² H), tritium (T, ³ H)); isotopes of carbon (e.g., ¹¹ C, ¹³ C, and ¹⁴ C); isotopes of chlorine (e.g., ³⁶ Cl); isotopes of fluorine (e.g., ¹⁸ F); isotopes of iodine (e.g., ¹²³ I and ¹²⁵ I); isotopes of nitrogen (e.g., ¹³ N and ¹⁵ N); isotopes of oxygen (e.g., ¹⁵ O, ¹⁷ O, and ¹⁸ O); isotopes of phosphorus (e.g., ³² P); and isotopes of sulfur (e.g., ³⁵ S). Certain isotopically labeled compounds of the invention (e.g., those incorporating radioisotopes) are useful in drug and/or substrate tissue distribution studies (e.g., assays). The radioisotopes tritium (i.e., ³ H) and carbon-14 (i.e., ¹⁴ C) are particularly useful for this purpose because they are easily incorporated and easily detected. Substitution with positron emitting isotopes (e.g., ¹¹ C, ¹⁸ F, ¹⁵ O, and ¹³ N) can be used to examine substrate receptor occupancy in positron emission tomography (PET) studies. Isotopically labeled compounds of the invention can be prepared by methods similar to those described in the accompanying routes and/or in the examples and preparations by using appropriate isotopically labeled reagents in place of the non-labeled reagents previously employed. Pharmaceutically acceptable solvates of the invention include those in which the crystallization solvent may be isotopically substituted, for example, D ₂ O, acetone-d ₆ or DMSO-d ₆ .

The term "stereoisomer" means an isomer formed due to at least one asymmetric center. In compounds with one or more (e.g., one, two, three, or four) asymmetric centers, it can produce racemic mixtures, single enantiomers, diastereomeric mixtures, and individual diastereomers. Specific individual molecules can also exist as geometric isomers (cis/trans). Similarly, the compounds of the present invention can exist as mixtures (commonly referred to as tautomers) of two or more structurally different forms in rapid equilibrium. Representative examples of tautomers include keto-enol tautomers, phenol-ketone tautomers, nitroso-oxime tautomers, imine-enamine tautomers, etc. It is to be understood that the scope of the present application covers all such isomers or mixtures thereof in any proportion (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%).

In this article, solid lines can be used Solid wedge Virtual wedge Carbon-carbon bonds of the compounds of the invention are depicted. The use of solid lines to depict bonds to asymmetric carbon atoms is intended to indicate that all possible stereoisomers at that carbon atom are included (e.g., specific enantiomers, racemic mixtures, etc.). The use of solid or dashed wedges to depict bonds to asymmetric carbon atoms is intended to indicate that the stereoisomer shown is present. When present in a racemic mixture, the solid and dashed wedges are used to define relative stereochemistry, not absolute stereochemistry. Unless otherwise indicated, the compounds of the invention are intended to exist in the form of stereoisomers, which include cis and trans isomers, optical isomers (e.g., R and S enantiomers), diastereomers, geometric isomers, rotational isomers, conformational isomers, atropisomers, and mixtures thereof. The compounds of the invention may exhibit more than one type of isomerism and consist of mixtures thereof (e.g., racemic mixtures and diastereomeric pairs).

Atropisomers are compounds that can be separated into their rotationally restricted isomers.

It should also be understood that certain compounds of the present invention may exist in free form for treatment, or, where appropriate, in the form of pharmaceutically acceptable derivatives thereof. In the present invention, pharmaceutically acceptable derivatives include, but are not limited to, pharmaceutically acceptable salts, esters, solvates, metabolites or prodrugs, which, after being administered to a patient in need thereof, can directly or indirectly provide a compound of the present invention or a metabolite or residue thereof. Therefore, when referring to "compounds of the present invention" herein, it is also intended to cover the above-mentioned various derivative forms of the compound.

Pharmaceutically acceptable salts of the compounds of the present invention include acid addition salts and base addition salts thereof.

For a review of suitable salts, see Stahl and Wermuth, "Handbook of Pharmaceutical Salts: Properties, Selection, and Use" (Wiley-VCH, 2002). Methods for preparing pharmaceutically acceptable salts of the compounds of the invention are known to those skilled in the art.

As used herein, the term "ester" means an ester derived from the compounds of the general formulae herein, including physiologically hydrolyzable esters (which can be hydrolyzed under physiological conditions to release the compounds of the present invention in free acid or alcohol form). The compounds of the present invention themselves may also be esters.

The compounds of the present invention may exist in the form of solvates (preferably hydrates), wherein the compounds of the present invention contain polar solvents as structural elements of the crystal lattice of the compounds, in particular water, methanol or ethanol. The amount of polar solvents, in particular water, may exist in a stoichiometric or non-stoichiometric ratio.

Also included within the scope of the present invention are metabolites of the compounds of the present invention, i.e., substances formed in vivo upon administration of the compounds of the present invention. Such products may be produced, for example, by oxidation, reduction, hydrolysis, amidation, deamidation, esterification, defatting, enzymatic hydrolysis, etc. of the administered compound. Thus, the present invention includes metabolites of the compounds of the present invention, including compounds prepared by contacting the compounds of the present invention with a mammal for a period of time sufficient to produce a metabolic product thereof.

The present invention further includes within its scope prodrugs of the compounds of the present invention, which are certain derivatives of the compounds of the present invention that may themselves have little or no pharmacological activity and can be converted into compounds of the present invention having the desired activity when administered into or onto the body, for example, by hydrolytic cleavage. Typically such prodrugs will be functional group derivatives of the compounds that are easily converted into the desired therapeutically active compounds in vivo. Additional information on the use of prodrugs can be found in "Pro-drugs as Novel Delivery Systems," Vol. 14, ACS Symposium Series (T. Higuchi and V. Stella) and "Bioreversible Carriers in Drug Design," Pergamon Press, 1987 (E. B. Roche, ed., American Pharmaceutical Association). Prodrugs of the present invention can be prepared, for example, by replacing appropriate functional groups present in the compounds of the present invention with certain moieties known to those skilled in the art as "pro-moieties" (e.g. as described in "Design of Prodrugs", H. Bundgaard (Elsevier, 1985)).

The present invention also encompasses compounds of the present invention containing protecting groups. In any process for preparing the compounds of the present invention, it may be necessary and/or desirable to protect sensitive or reactive groups on any of the molecules involved, thereby forming a chemically protected form of the compounds of the present invention. This can be achieved by conventional protecting groups, for example, those described in Protective Groups in Organic Chemistry, ed. J.F.W. McOmie, Plenum Press, 1973; and T.W. Greene & P.G.M. Wuts, Protective Groups in Organic Synthesis, John Wiley & Sons, 1991, which references are incorporated herein by reference. The protecting groups may be removed at an appropriate subsequent stage using methods known in the art.

As used herein, the term "about" means within ±10% of the stated numerical value, preferably within ±5%, and more preferably within ±2%.

Compound

In some embodiments, the present disclosure provides a compound or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound, or prodrug thereof, wherein the compound has the structure of Formula (I):

in:

Selected from

W is CR ¹ or N;

^R1 at each occurrence is independently selected from H, halogen, -OH, _-NH2 , -CN, _-NO2 , _-SF5 , _C1-6 alkyl, deuterated _C1-6 alkyl, halogenated _C1-6 alkyl, _C2-6 alkenyl, _C2-6 alkynyl, _C3-6 cycloalkyl, 3-10 membered heterocyclyl, _C6-10 aryl, 5-14 membered heteroaryl, _C6-12 aralkyl, -C(=O) ^Ra , -OC(=O) ^Ra , -C(=O) ^ORa , ^-ORa , ^-SRa , -S(=O) ^Ra , -S(=O) _2Ra , ^-S (=O ^{)2NRaRb , -NRaRb} , -C ₍ =O) ^NRaRb , -NRa ^{- C} (=O) ^{Rb , -NRa} -C(═O)OR ^b , -NR ^a -S(═O) ₂ -R ^b , -NR ^a -C(═O)-NR ^a R ^b , -P(═O)R ^a R ^b , -C _1-6 alkylene-R ^a , -C _1-6 alkylene-OR ^a , -C _1-6 alkylene-NR ^a R ^b , -OC _1-6 alkylene-NR ^a R ^b , (—C _3-6 cycloalkylene)-CN and (—C _3-6 cycloalkylene)-C _1-6 haloalkyl;

Alternatively, two R ^1s in the ortho position together with the groups to which they are attached optionally together form a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring;

^R2 is

R ³ , R ²¹ , R ²² , R ²³ and R ²⁴ are each independently selected at each occurrence from H, halogen, -OH, -NH ₂ , -CN, -NO ₂ , -SF ₅ , C _1-6 alkyl, deuterated C _1-6 alkyl, halogenated C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, C _6-12 aralkyl, -C(═O)R ^{a , -OC(═O)R a ,} -C(═O)OR ^a , -OR ^a , -SR ^a , -S(═O)R ^a , -S(═O) ₂ R ^a , -S(═O) ₂ NR ^a R ^b , -S(═O)(═NR ^a )R ^b , -NR ^a R ^b , —C(═O)NR ^a R ^b , —NR ^a -C(═O)R ^b , —NR ^a -C(═O)R b , —NR ^a -C(═O)OR ^b , —NR ^a -S(═O) ₂ -R ^b , —NR ^{a -C} (═O)-NR ^a R ^b , —P(═O)R a R b , —C _1-6 alkylene-R ^a , —C _1-6 alkylene-OR ^a , —C _1-6 alkylene-NR ^a R ^b , —OC _1-6 alkylene-NR ^a R ^b , (—C _3-6 cycloalkylene)-CN and (—C _3-6 cycloalkylene)-C _1-6 haloalkyl;

Alternatively, R ³ and R ²¹ or R ²² together with the group to which they are attached optionally together form a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring;

^R4 is

R ⁴¹ is selected from a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring, and a 5-14 membered heteroaromatic ring;

Ring X and Ring Z are each independently selected from a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring, and a 5-14 membered heteroaromatic ring;

Ring Y is absent or is selected from a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring and a 5-14 membered heteroaromatic ring; when Ring Y is absent, R ²⁴ is also absent;

^L2 is selected from -O-, -C(=O)-, -NRC(=O)-, -S-, -S(=O)-, -S(=O) _2- , _C1-6 alkylene and -O-( _C1-6 alkylene)-;

R, ^Ra and ^Rb are each independently selected at each occurrence from H, _C1-6 alkyl, _C3-10 cycloalkyl, 3-10 membered heterocyclyl, _C6-10 aryl, 5-14 membered heteroaryl and _C6-12 aralkyl;

The above alkylene, alkyl, alkenyl, alkynyl, cycloalkylene, cycloalkyl, hydrocarbon ring, heterocyclyl, heterocycle, aryl, aromatic ring, heteroaryl, heteroaromatic ring and aralkyl are each optionally substituted at each occurrence by one or more substituents independently selected from the following: deuterium atom, halogen, -OH, =O, -NH2, -CN, _-NO2 , = _CH2 , _C1-6 alkyl, deuterated _{C1-6 alkyl} , _C2-6 alkenyl, _C2-6 alkynyl, _C3-6 cycloalkyl, 3-10 membered heterocyclyl, _C6-10 aryl, 5-14 membered heteroaryl, _C6-12 aralkyl, -C(=O) ^Rc , -OC(=O) ^Rc , -C(=O) ^ORc , ^-ORc , ^-SRc , -S(=O) ^Rc , -S(= ^O ) _2Rc -S(＝O) _2NRcRd , ^-NRcRd , ^-C (＝O) ^NRcRd , ^-NRc- C( ^＝ O) ^Rd , -NRc ^- C(＝O) ^ORd , -NRc ^- S(＝O) ₂ - ^Rd , -NRc ^{- C} (＝O)-NRcRd, -C1-6alkylene ^{- ORc} , _{-C1-6alkylene} - ^NRcRd and _{-OC1-6alkylene} - ^NRcRd ; when the same ^ring atom or adjacent ring atoms of a cycloalkylene, ^cycloalkyl , hydrocarbon ^ring , _heterocyclyl , heterocycle, ^aryl , aromatic ring, heteroaryl and heteroaromatic ring are substituted by two substituents, the two substituents together with the group to which they are attached optionally constitute a _C3-6 hydrocarbon ring, a 3-10 membered heterocycle, a C1-6alkylene ring or a heteroaryl ring. _6-10 membered aromatic ring or 5-14 membered heteroaromatic ring; each of the alkylene, alkyl, alkenyl, =CH ₂ , alkynyl, cycloalkyl, hydrocarbon ring, heterocyclyl, heterocycle, aryl, aromatic ring, heteroaryl, heteroaromatic ring and aralkyl is further optionally substituted by one or more substituents independently selected from the following: halogen, -OH, =O, -C(=O)O-tert-butyl, -NH ₂ , -CN, -NO ₂ , C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, C _6-12 aralkyl, -C _1-6 alkylene-C _3-6 cycloalkyl, -OC _1-6 alkyl and -C _1-6 alkylene-OC _1-6 alkyl;

R ^c and R ^d are each independently selected at each occurrence from H, C _1-6 alkyl, C _3-10 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, and C _6-12 aralkyl, said alkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and aralkyl being further optionally substituted with one or more substituents independently selected from the group consisting of halogen, -OH, =O, -C(=O)O-tert-butyl, -NH ₂ , -CN, -NO ₂ , C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, C _6-12 aralkyl _{, and -C 1-6 alkylene} -OC _1-6 alkyl; and

p, q and t are each independently an integer selected from 1, 2 or 3.

In some embodiments, R ¹ is independently selected from H, halogen, C _1-6 alkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, -OR ^a and -NR ^a R ^b at each occurrence, preferably, R ¹ is independently selected from H, halogen, C _1-6 alkyl, C _3-6 cycloalkyl and -OR ^a at each occurrence; wherein the alkyl, cycloalkyl, heterocyclyl, aryl and heteroaryl are each optionally substituted with one or more substituents independently selected from the following: halogen, C _1-6 alkyl, C _2-6 alkenyl, =CH ₂ , C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl and 5-14 membered heteroaryl; the alkyl, alkenyl, =CH ₂ , cycloalkyl, heterocyclyl, aryl and heteroaryl are each further optionally substituted by one or more substituents independently selected from the group consisting of halogen, C _1-6 alkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl and -C _1-6 alkylene-C _3-6 cycloalkyl.

In a preferred embodiment, R ¹ is H, methyl, halogen, methoxy,

In some embodiments, two R ^1s in the ortho position together with the group to which they are attached optionally constitute a C _3-6 hydrocarbon ring, a 3-10 membered heterocycle, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring, wherein each of the hydrocarbon ring, heterocycle, aromatic ring and heteroaromatic ring is optionally substituted by one or more substituents independently selected from the following: halogen, C _1-6 alkyl and halogenated C _1-6 alkyl.

In a preferred embodiment, the two R ¹ in the ortho position together with the group to which they are attached optionally constitute A benzene ring or a pyridine ring, each of which is optionally substituted by one or more substituents independently selected from the group consisting of halogen, C _1-6 alkyl and halogenated C _1-6 alkyl.

In some embodiments, ring X is a benzene ring, a 5-6 membered heterocyclic ring, or a 5-6 membered heteroaromatic ring (preferably a thiophene ring, a thiazole ring, or a pyridine ring), and ring Y is absent.

In some embodiments, ring X is a benzene ring or a 5-6 membered heteroaromatic ring, and ring Y is a C _3-6 hydrocarbon ring, a 5-6 membered heterocyclic ring, or a 5-6 membered heteroaromatic ring.

In a preferred embodiment, for More preferably

In some embodiments, R ²¹ , R ²² , R ²³ and R ²⁴ are each independently selected at each occurrence from H, halogen, -SF ₅ , C _1-6 alkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, -O-(C _1-6 alkyl), -S(＝O) ₂ -(C _1-6 alkyl), -S(＝O) ₂ -(C _3-6 cycloalkyl), -P(＝O)(C _1-6 alkyl) ₂ , (-C _3-6 cycloalkylene)-CN and (-C _3-6 cycloalkylene)-C _1-6 alkyl, said alkyl, cycloalkylene, cycloalkyl and heterocyclyl are each optionally substituted with one or more substituents independently selected from: halogen, C _1-6 alkyl and halo-substituted C _1-6 alkyl.

In a preferred embodiment, R ²¹ , R ²² , R ²³ and R ²⁴ are each independently selected from H, halogen, -SF ₅ , C _1-6 alkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl and -O-(C _1-6 alkyl), said alkyl, cycloalkyl and heterocyclyl being each optionally substituted with one or more substituents independently selected from halogen, _{C 1-6 alkyl} and halo-substituted C _1-6 alkyl.

In some embodiments, Selected from:

In some embodiments, Selected from:

In some embodiments, R ³ is H, C _1-6 alkyl, -OR ^a or -SR ^a ; preferably, R ³ is H, ethyl, -O-CH ₃ or -S-CH ₃ .

In some embodiments, ^R3 and ^R21 or ^R22 together with the group to which they are attached optionally together constitute a _C3-6 hydrocarbon ring or a 3-10 membered heterocycle, and the hydrocarbon ring and the heterocycle are optionally substituted with one or more substituents selected from a deuterium atom, a halogen, -OH, a _C1-6 alkyl group, a halogenated _C1-6 alkyl group and a _-OC1-6 alkyl group; when the same ring atom or adjacent ring atoms of the hydrocarbon ring and the heterocycle are substituted with two substituents, the two substituents together with the group to which they are attached optionally together constitute a _C3-6 hydrocarbon ring or a 3-10 membered heterocycle.

In a preferred embodiment, R ³ and R ²¹ or R ²² together with the group to which they are attached optionally together constitute a cyclopentene ring, a cyclohexene ring, a pyrrolidine ring, a piperidine ring or a morpholine ring, and the cyclopentene ring, the cyclohexene ring, the pyrrolidine ring, the piperidine ring and the morpholine ring are optionally substituted with one or more substituents selected from deuterium atoms, halogen, -OH, C _1-6 alkyl, halogenated C _1-6 alkyl and -OC _1-6 alkyl; when the same ring atom of the cyclopentene ring, the cyclohexene ring, the pyrrolidine ring, the piperidine ring and the morpholine ring is substituted with two substituents, the two substituents together with the group to which they are attached optionally together constitute a C _3-6 hydrocarbon ring.

In some embodiments, ring Z is a 3-10 membered heterocycle or a benzene ring; preferably a 5-10 membered heterocycle; more preferably a 5-6 membered heterocycle; and

The heterocyclic ring and the benzene ring are each optionally substituted at each occurrence with one or more substituents independently selected from the group consisting of halogen, C _1-6 alkyl and halo-substituted C _1-6 alkyl.

In a preferred embodiment, ring Z is

In some embodiments, L ² is -C(=O)- or -NRC(=O)-, wherein R is H or C _1-6 alkyl;

In a preferred embodiment, ^L2 is -C(=O)-.

In some embodiments, R ⁴¹ is selected from a 3-10 membered heterocycle, a C _6-10 aromatic ring, and a 5-14 membered heteroaromatic ring, wherein the heterocycle, aromatic ring, and heteroaromatic ring are each optionally substituted by one or more substituents independently selected from the following: halogen, -OH, C _1-6 alkyl, -OC _1-6 alkyl, and -SC _1-6 alkyl, preferably, the heterocycle, aromatic ring, and heteroaromatic ring are at least substituted by -OH or -OC _1-6 alkyl.

In some embodiments, -L ² -R ⁴¹ is

In some embodiments, R ⁴¹ is a 5-6 membered heteroaromatic ring, preferably a 6 membered heteroaromatic ring, more preferably a pyridine ring or a pyrimidine ring, which is substituted by at least one -OH.

In a preferred embodiment, -L ² -R ⁴¹ is

In some embodiments, the present disclosure provides a compound or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound, or prodrug thereof, wherein the compound has a structure of the following formula:

in:

Ring C and Ring D are each independently a C _3-6 hydrocarbon ring, a 3-10 membered heterocycle, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring, and the hydrocarbon ring, heterocycle, aromatic ring and heteroaromatic ring are each optionally substituted by one or more substituents independently selected from the following: a deuterium atom, a halogen, -OH, a C _1-6 alkyl group, a halogenated C _1-6 alkyl group and a -OC _1-6 alkyl group; when the same ring atom or adjacent ring atoms of the hydrocarbon ring, heterocycle, aromatic ring and heteroaromatic ring are substituted by two substituents, the two substituents together with the group to which they are attached optionally constitute a C _3-6 hydrocarbon ring, a 3-10 membered heterocycle, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring;

Preferably, ring C is a benzene ring or a pyridine ring, each of which is optionally substituted by one or more substituents independently selected from the group consisting of halogen, C _1-6 alkyl and halogenated C _1-6 alkyl; and/or

Ring D is a cyclopentene ring, a cyclohexene ring, a pyrrolidine ring, a piperidine ring or a morpholine ring, each of which is optionally substituted by one or more substituents independently selected from the following: a deuterium atom, a halogen, -OH, a C _1-6 alkyl group, a halogenated C _1-6 alkyl group and a -OC _1-6 alkyl group; when the same ring atom or adjacent ring atoms of ring D are substituted by two substituents, the two substituents together with the groups to which they are attached optionally constitute a C _3-6 hydrocarbon ring or a 3-10 membered heterocyclic ring;

The remaining groups are as defined above.

The present disclosure covers technical solutions/compounds obtained by any combination of various embodiments.

In a preferred embodiment, the present disclosure provides a compound or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, wherein the compound is selected from:

Pharmaceutical compositions and methods of treatment

In some embodiments, the present invention provides a pharmaceutical composition comprising a preventive or therapeutically effective amount of a compound of the present invention or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof and one or more pharmaceutically acceptable carriers. The pharmaceutical composition is preferably a solid preparation, a semisolid preparation, a liquid preparation or a gaseous preparation. In some embodiments, the pharmaceutical composition may also include one or more other therapeutic agents.

In some embodiments, the present invention provides the use of a compound of the present invention or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, or a pharmaceutical composition of the present invention in the preparation of a medicament for use as a WRN inhibitor.

In some embodiments, the present invention provides a compound of the present invention or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, or a pharmaceutical composition of the present invention, for use as a WRN inhibitor.

In some embodiments, the present invention provides a method for preventing or treating cancer, preferably a cancer characterized by microsatellite high instability (MSI-H) or mismatch repair deficiency (dMMR), comprising administering to an individual in need thereof an effective amount of a compound of the present invention or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, or a pharmaceutical composition of the present invention.

In some embodiments, the cancer includes colorectal cancer, gastric cancer, endometrial cancer, uterine cancer, adrenocortical cancer, cervical cancer, esophageal cancer, breast cancer, kidney cancer, prostate cancer, and ovarian cancer.

In the present invention, "pharmaceutically acceptable carrier" refers to a diluent, adjuvant, excipient or vehicle that is administered together with a therapeutic agent and is suitable for contact with the tissues of humans and/or other animals without excessive toxicity, irritation, allergic reaction or other problems or complications corresponding to a reasonable benefit/risk ratio within the scope of reasonable medical judgment.

As used herein, unless otherwise indicated, the terms "treat," ...

As used herein, "individual" includes humans or non-human animals. Exemplary human individuals include human individuals (referred to as patients) suffering from diseases (e.g., diseases described herein) or normal individuals. "Non-human animals" in the present invention include all vertebrates, such as non-mammals (e.g., birds, amphibians, reptiles) and mammals, such as non-human primates, livestock and/or domesticated animals (e.g., sheep, dogs, cats, cows, pigs, etc.).

In another embodiment, the pharmaceutical compositions of the present invention may further comprise one or more additional therapeutic or prophylactic agents.

Example

The present invention is further described below in conjunction with examples, but these examples are not intended to limit the scope of the present invention.

The abbreviations in this invention have the following meanings:

General synthetic route:

Route 1

Route 2

Wherein, in Scheme 1 and Scheme 2, PG is a protecting group, and the remaining groups are as defined herein.

Example 1: Synthesis of Compound B1

1) Synthesis of intermediate B1-3:

Compound B1-1 (9.0 g, 38.03 mmol), compound B1-2 (21.72 g, 68.45 mmol) and TEA (11.52 g, 114.09 mmol) were added to 1,4-dioxane (100 mL), replaced with nitrogen three times, and then heated to 120 ° C. The reaction was stirred overnight under a nitrogen atmosphere. TLC monitoring showed that the raw material reaction was complete, the reaction solution was concentrated under reduced pressure, and the residue was purified by column chromatography (PE/EA=3/1) to obtain a yellow solid compound B1-3 (15.0 g, yield 76%). LCMS (ESI) m/z=518.2[M+H] ⁺ .

2) Synthesis of intermediate B1-4:

Compound B1-3 (12 g, 23.2 mmol) was added to a mixed solution of ethanol and tetrahydrofuran (2:1) (450 mL), cooled to 0°C, NaOH (1.85 g, 46.4 mmol) was dissolved in 10 mL of water, slowly added dropwise to the reaction solution, stirred overnight at room temperature, TLC monitoring showed that the raw material reaction was complete, HCl (1 M) was added dropwise to the reaction solution to make the pH value reach 5, washed with water, extracted with ethyl acetate, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, the residue was added with ether and then filtered to obtain a yellow solid compound B1-4 (9.0 g, yield 79%). LCMS (ESI) m/z = 490.3 [M + H] ⁺ .

3) Synthesis of intermediate B1-6:

Compound B1-4 (9.6 g, 19.6 mmol), compound B1-5 (3.78 g, 39.26 mmol), TEA (7.92 g, 78.4 mmol) and HATU (8.94 g, 23.52 mmol) were added to DMF (100 mL), replaced with nitrogen three times, and stirred at room temperature under nitrogen atmosphere overnight. TLC monitoring showed that the raw material was completely reacted, washed with water, extracted with ethyl acetate, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (PE/EA=4/1) to obtain yellow solid compound B1-6 (4.5 g, yield 60%). LCMS (ESI) m/z=383.2[M+H] ⁺ .

4) Synthesis of intermediate B1-7:

Compound B1-6 (5.0 g, 13.09 mmol) was added to tetrahydrofuran solution (100 mL), replaced with nitrogen three times, cooled to 0°C, methyl Grignard reagent (1M, 52.4 mL, 52.4 mmol) was slowly added dropwise, and stirred at 0°C for 1 hour. LC-MS showed that the raw material reaction was complete, and ammonium chloride aqueous solution was slowly added dropwise to quench, extracted with ethyl acetate, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (PE/EA=6/1) to obtain yellow solid compound B1-7 (3.6 g, yield 81%). LCMS (ESI) m/z=338.1[M+H] ⁺ .

5) Synthesis of intermediate B1-8:

Compound B1-7 (3.6 g, 10.7 mmol) was added to dichloromethane (10 mL), and then trifluoroacetic acid (10 mL) was added, and then the temperature was raised to 40°C, and the reaction was stirred for 2 hours. LC-MS showed that the raw material reaction was complete, and a saturated aqueous solution of sodium bicarbonate was slowly added to the reaction solution until the pH value was equal to 7, and ethyl acetate was extracted. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (PE/EA=4/1) to obtain a yellow solid compound B1-8 (1.7 g, yield 85%). LCMS (ESI) m/z=188.3[M+H] ⁺ .

6) Synthesis of intermediate B1-9:

Compound B1-8 (1.0 g, 5.3 mmol) was added to dichloromethane (20 mL), cooled to 0°C, propionyl chloride (0.99 g, 10.6 mmol) and pyridine (2.09 g, 26.5 mmol) were slowly added dropwise, and the reaction solution was stirred at 0°C for 3 hours. LC-MS showed that the raw material reaction was complete, and water was added to quench, and ethyl acetate was extracted. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (PE/EA=3/1) to obtain yellow solid compound B1-9 (0.98 g, yield 75%). LCMS (ESI) m/z=244.1[M+H] ⁺ .

7) Synthesis of intermediate B1-10:

Compound B1-9 (950 mg, 3.9 mmol) was added to tert-butyl alcohol (50 mL), potassium tert-butylate (2190 mg, 19.54 mmol) was slowly added, nitrogen was replaced three times, the reaction solution was heated to 95°C, and stirred for 2 hours under nitrogen atmosphere. LC-MS showed that the raw materials were completely reacted. The reaction solution was cooled to room temperature, HCl (1 M) was added dropwise to the reaction solution to make the pH value reach 7, and ethyl acetate was extracted. The organic phase was then washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (DCM/MeOH=50/1) to obtain yellow solid compound B1-10 (450 mg, yield 51%). LCMS (ESI) m/z=226.3[M+H] ⁺ .

8) Synthesis of intermediate B1-12:

Compound B1-11 (3.0 g, 15.34 mmol) was dissolved in dichloromethane (20 mL), triethylamine (3.10 g, 15.34 mmol) and bromoacetyl bromide (3.0 g, 15.34 mmol) were added at 0°C, and the mixture was stirred at room temperature for 5 hours. LC-MS showed that the raw material reaction was complete, and aqueous solution was slowly added dropwise to quench, and ethyl acetate was extracted. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (PE/EA=50/1) to obtain an off-white solid compound B1-12 (4.0 g, yield 83%). LCMS (ESI) m/z=315.9[M+H] ⁺ .

9) Synthesis of intermediate B1-13:

Compound B1-12 (950.mg, 3.02mmol) and compound B1-10 (678.6mg, 3.02mmol) were added to acetonitrile (10mL), DIEA (1168.7mg, 9.06mmol) was slowly added dropwise, nitrogen was replaced three times, the reaction solution was heated to 80°C and stirred overnight, LC-MS showed that the raw materials were completely reacted, the reaction solution was concentrated under reduced pressure, and the residue was purified by column chromatography (PE/EA=6/4) to obtain white solid compound B1-13 (850mg, yield 61.27%). LCMS (ESI) m/z=461.0[M+H] ⁺ .

10) Synthesis of intermediate B1-14:

Compound B1-13 (600 mg, 1.30 mmol) was added to dichloromethane (20 mL), NBS (232.17 mg, 1.3 mmol) was added at 0°C, and the reaction was stirred at room temperature for 1 hour. LC-MS showed that the raw material reaction was complete, and aqueous solution was slowly added dropwise to quench, extracted with ethyl acetate, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (PE/EA=2/1) to obtain an off-white solid compound B1-14 (550 mg, yield 78%). LCMS (ESI) m/z=538.9[M+H] ⁺ .

11) Synthesis of intermediate B1-15:

Compound B1-14 (180 mg, 0.33 mmol), piperazine (567.6 mg, 6.6 mmol) and silver tetrafluoroborate (130 mg, 0.66 mmol) were added to dimethyl sulfoxide (4 mL), and the reaction solution was heated to 120°C for reaction. LC-MS showed that the raw material reaction was complete, and aqueous solution was added dropwise to quench, extracted with ethyl acetate, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (DCM/MeOH=10/1) to obtain an off-white solid compound B1-15 (25 mg, yield 14%). LCMS (ESI) m/z=545.1[M+H] ⁺ .

12) Synthesis of Product B1:

Compound B1-16 (8.87 mg, 0.059 mmol), HOBT (5.94 mg, 0.044 mmol) and EDCI (11.52 mg, 0.06 mmol) were added to DMF (4 mL), replaced with nitrogen three times, and stirred for 2 hours under a nitrogen atmosphere. Compound B1-15 (23 mg, 0.04 mmol) and DIEA (15.48 mg, 0.12 mmol) were added to the reaction solution, and then stirring was continued at room temperature for another 2 hours. LC-MS showed that the raw material reaction was complete, and aqueous solution was added dropwise to quench, extracted with dichloromethane, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by preparative high performance liquid chromatography to obtain compound B1 (5.4 mg, yield 19%). LCMS (ESI) m/z＝680.9[M+H] ^+.1 H NMR (400MHz, CD ₃ OD): δ8.53(s,1H),8.32(d,J＝7.6Hz,1H),8.11-8.07(m,2H),7.99-7.95(m,1H),7.90-7.86(m,1H),7.81(s,1H),7.58(dd,J ₁ ＝1.2Hz,J ₂ ＝8.8Hz,1H),5.70(s,2H),4.74-4.70(m,1H),4.07-3.91(m,3H),3.52-3.40(m,3H),3.22 -3.13(m,1H),2.99-2.96(m,1H),2.84-2.81(m,1H),2.53(s,3H),1.43(t,J＝7.6Hz,3H).

Example 2: Synthesis of Compound B6

1) Synthesis of intermediate B6-2:

Compound B6-1 (9 g, 38.31 mmol) was dissolved in methanol (100 mL) solution, cooled to -30 °C and stirred for ten minutes, then 2,3-butanedione (3.3 g, 38.31 mmol) and sodium acetate (10.18 g, 122.59 mmol) in aqueous solution (100 mL) were added, and after the addition was complete, 40 mL (3.6 mol/L) of sodium hydroxide solution was added, and then the reaction was reacted at 0 °C for 30 min, and then transferred to room temperature for overnight reaction. LCMS showed that the reaction was complete, and the mixture was extracted with water (200 mL) and ethyl acetate (200 mL*3), and the organic phases were combined and then washed with saturated brine (20 mL*3). The organic phase was dried over anhydrous sodium sulfate and concentrated to obtain crude product B6-2 (3.85 g, yield: 81.6%). LCMS (ESI) m/z = 124.1 [M+H] ⁺ .

2) Synthesis of intermediate B6-3:

Compound B6-2 (3.8 g, 24.99 mmol) was dissolved in acetonitrile (40 mL), cooled to 0°C, and NBS (4.45 g, 24.99 mmol) was added thereto. The reaction solution was stirred at 25°C for 0.5 hours. LCMS showed that the reaction was complete, and the mixture was extracted with water (100 mL) and ethyl acetate (100 mL*3). The organic phases were combined and then washed with saturated brine (20 mL*3). The organic phase was dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain B6-3 (3.9 g, yield: 77.23%). LCMS (ESI) m/z=202.0 [M+H] ⁺ .

3) Synthesis of intermediate B6-4:

Compound B6-3 (3.9 g, 19.3 mmol) and cuprous cyanide (4.13 g, 46.13 mmol) were dissolved in DMF (40 mL), and then reacted at 120 ° C for 2 h. LCMS showed that the reaction was complete, and extracted with water (100 mL) and ethyl acetate (100 mL*3), and the organic phases were combined and then washed with saturated brine (20 mL*3). The organic phase was dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain B6-4 (0.74 g, yield: 25.85%). LCMS (ESI) m/z = 149.2 [M+H] ⁺ .

4) Synthesis of intermediate B6-5:

Compound B6-4 (0.57 g, 3.85 mmol) was dissolved in tetrahydrofuran (6 mL), cooled to 0°C, and then methylmagnesium bromide (3 M, 13 mL, 38.5 mmol) was added, followed by reaction at 50°C overnight. The reaction was completed after LCMS monitoring, and extracted with water (100 mL) and ethyl acetate (100 mL*3), and the organic phases were combined and then washed with saturated brine (20 mL*3). The organic phase was dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain B6-5 (0.5 g, yield: 78.68%). LCMS (ESI) m/z=166.5[M+H] ⁺ .

5) Synthesis of intermediate B6-6:

Compound B6-5 (0.7 g, 4.24 mmol) was added to dichloromethane (20 mL), cooled to 0°C, propionyl chloride (0.78 g, 8.48 mmol) and pyridine (1.68 g, 21.20 mmol) were slowly added dropwise, and the reaction solution was stirred at 0°C for 3 hours. LC-MS showed that the raw material reaction was complete, and water was added to quench, and ethyl acetate was extracted. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography to obtain yellow solid compound B6-6 (0.6 g, yield 63.99%). LCMS (ESI) m/z = 222.2 [M + H] ⁺ .

6) Synthesis of intermediate B6-7:

Compound B6-6 (0.35 g, 1.58 mmol) was added to 1,4-dioxane (5 mL), potassium tert-butoxide (0.27 g, 2.37 mmol) was slowly added, nitrogen was replaced three times, the reaction solution was heated to 110°C, stirred for 1 hour under nitrogen atmosphere, LC-MS showed that the raw material reaction was complete, the reaction solution was cooled to room temperature, extracted with ethyl acetate, the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the residue was purified by column chromatography to obtain yellow solid compound B6-7 (0.3 g, yield 93.31%). LCMS (ESI) m/z = 204.2 [M + H] ⁺ .

7) Synthesis of intermediate B6-8:

Compound B6-7 (0.28 g, 1.38 mmol) and compound B1-12 (0.48 g, 1.52 mmol) were added to acetonitrile (3 mL), and DIPEA (0.89 g, 6.9 mmol) was slowly added dropwise, and nitrogen was replaced three times. The reaction solution was heated to 45°C and stirred for 4 hours. LC-MS showed that the raw materials reacted completely. The reaction solution was concentrated under reduced pressure, and the residue was purified by column chromatography to obtain a white solid compound B6-8 (0.5 g, yield 82.7%). LCMS (ESI) m/z = 439.2 [M+H] ⁺ .

8) Synthesis of intermediate B6-9:

Compound B6-8 (0.45 g, 1.03 mmol) was added to dichloromethane (10 mL), and NBS (0.32 g, 1.77 mmol) was added at 0°C. The reaction was stirred at room temperature for 1 hour. LC-MS showed that the raw material reaction was complete. The aqueous solution was slowly added dropwise to quench the reaction, and the mixture was extracted with ethyl acetate. The organic phase was then washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography to obtain an off-white solid compound B6-9 (0.2 g, yield 37.67%). LCMS (ESI) m/z = 517.2 [M+H] ⁺ .

9) Synthesis of intermediate B6-11:

Compound B6-9 (0.15 g, 0.29 mmol), compound B6-10 (0.45 g, 2.12 mmol), silver tetrafluoroborate (0.3 g, 1.54 mmol) and DIPEA (0.39 g, 2.9 mmol) were added to dimethyl sulfoxide (4 mL), and the reaction solution was heated to 140°C. LC-MS showed that the raw material reaction was complete, and aqueous solution was added dropwise to quench, extracted with ethyl acetate, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (DCM/MeOH=10/1) to obtain an off-white solid compound B6-11 (20 mg, yield 10.63%). LCMS (ESI) m/z=649.2[M+H] ⁺ .

10) Synthesis of intermediate B6-12:

Compound B6-11 (20 mg, 0.031 mmol) was dissolved in dichloromethane (0.5 mL), and trifluoroacetic acid (0.5 mL) was added dropwise thereto. The reaction solution was stirred at 25° C. for 1.5 hours. LCMS showed that the reaction was complete, and the reaction solution was concentrated to obtain the trifluoroacetate salt of B6-12 (60 mg, crude product) as a yellow oil. LCMS (ESI) m/z=549.2 [M+H] ⁺ .

11) Synthesis of product B6:

Compound B1-16 (7.2 mg, 0.059 mmol), HOAT (6.7 mg, 0.046 mmol) and EDCI (11.52 mg, 0.056 mmol) were added to DMF (0.5 mL), replaced with nitrogen three times, stirred for 1 hour under nitrogen atmosphere, trifluoroacetate of compound B6-12 (60 mg, crude product) and DIPEA (12 mg, 0.093 mmol) were added to the reaction solution, and then stirred for another 0.5 hour at room temperature. LC-MS showed that the raw material reaction was complete, and compound B6 (3.5 mg, two-step yield 16.5%) was obtained by purification by reverse phase C18 column preparative chromatography (0.5% formic acid aqueous solution: acetonitrile = 9:1 to 1:9). LCMS (ESI) m/z = 685.2 [M+H] ⁺ . ¹ H NMR (400MHz, DMSO-d ₆ )δ10.37(s,1H),8.50(s,1H),8.01–7.94(m,2H),7.69(dd,J＝8.8,2.1Hz,1H),5.44(s,2H),4.48(d,J＝13.0Hz,1H),3.88(d,J＝8.9Hz,1H),3.60– 3.11(m,6H),2.60(s,3H),2.59(s,3H),2.41(s,3H),1.99(dt,J＝13.4,7 .0Hz,1H),1.65-1.45(m,2H),1.34–1.27(m,3H),1.06(t,J＝8.2Hz,1H).

Example 3: Synthesis of Compound B18

1) Synthesis of intermediate B18-2:

B18-1 (9.0 g, 44.53 mmol) and anhydrous tetrahydrofuran (225 mL) were added to a 1000 mL three-necked flask in sequence, stirred at -70 to -75 °C under nitrogen protection, and n-butyl lithium n-hexane solution (2.5 mol/L, 48 mL, 120 mmol) was slowly added, stirred at -70 to -75 °C for 20 minutes, slowly heated to -5 to 0 °C, stirred at -5 to 0 °C for 70 minutes, cooled to -70 to -75 °C, and hexachloroethane (31.6 g, 133.6 mmol) was added at -70 to -75 °C, stirred at -70 to -75 °C for 40 minutes, then slowly returned to room temperature and continued to stir for 40 minutes. The reaction was quenched with 10% w/w citric acid aqueous solution, extracted twice with ethyl acetate, and the organic phases were combined, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and column chromatography to obtain B18-2 (3.5 g, 33%).

2) Synthesis of intermediate B18-3:

To a solution of B18-2 (3 g, 12.68 mmol) in anhydrous tert-butyl alcohol (30 mL) was added diphenylphosphoryl azide (4.08 g, 14.84 mmol). After the addition was completed, N,N-diisopropylethylamine (1.92 g, 19.02 mmol) was added to the reaction solution. The reaction solution was heated to 80 ° C and stirred overnight. The reaction solution was concentrated under reduced pressure, and the residue was added with saturated sodium bicarbonate solution, extracted with ethyl acetate (3×50 mL), and the organic phase was washed with brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by column chromatography to obtain a white solid compound B18-3 (2.8 g, 71%).

3) Synthesis of intermediate B18-4:

B18-3 (450 mg, 1.46 mmol) was dissolved in dichloromethane (3 mL) solution, and trifluoroacetic acid (1 mL) was added thereto, and the reaction solution was stirred at 25°C for 2 hours. LCMS showed that the reaction was complete, and the reaction solution was concentrated to obtain a brown oil B18-4 (300 mg, mixture, yield: 98.8%). ESI m/z [M+H] ⁺ = 208.1.

4) Synthesis of intermediate B18-5:

Compound B18-4 (0.3 g, 1.45 mmol) and potassium carbonate (0.6 g, 4.35 mmol) were dissolved in dichloromethane (2 mL) solution, bromoacetyl bromide (0.44 g, 2.17 mmol) was added dropwise under stirring at 0°C, and the mixture was stirred at 25°C for 1 hour. LCMS showed that the reaction was complete, and the reaction solution was poured into water, extracted with dichloromethane (50 mL*2), and concentrated to obtain a yellow solid B18-5 (0.3 g, 63%).

5) Synthesis of product B18:

Referring to the synthesis method of compound B6 in Example 2, compound B18-5 was used instead of B1-12 to synthesize compound B18. LCMS(ESI)m/z＝697.2[M+H] ^+.1 H NMR(400MHz, DMSO-d ₆ )δ10.37(s,1H),8.57(s,1H),7.41(m,2H),5.39(s,2H),4.57–4.41(m,1H),3.91(d,J＝8.6Hz,1H),3.75–3.06(m,6H) ,2.64(s,3H),2.63(s,3H),2.46(s,3H),1.62-1.50(m,1H),1.40-1.28(m,2H),1.28–1.22(m,3H),1.20-1.14(m,1H).

Example 4: Synthesis of Compound B113

1) Synthesis of intermediate B113-9:

Will (0.6 g, 2.38 mmol, prepared according to the synthesis method disclosed in patent application WO2008082484) and methylboronic acid (1.42 g, 23.8 mmol) were dissolved in a mixed solution of 1,4-dioxane (5 mL) and water (0.5 mL), potassium carbonate (0.99 g, 7.14 mmol) was added under stirring, and 1,1-bis(diphenylphosphino)ferrocenepalladium dichloride (II) (0.17 g, 0.24 mmol) was added after three times of gas replacement, and the reaction was stirred at 80°C for 16 hours. LCMS showed that the reaction was complete, and the reaction solution was concentrated and purified by column chromatography (ethyl acetate/petroleum ether) to obtain a white solid B113-9 (0.4 g, yield: 89.8%). m/z[M+H] ⁺ =188.1.

2) Synthesis of intermediate B113-2:

4-Nitro-2H-1,2,3-triazole (1.0 g, 8.76 mmol, 1.0 eq), phenylboronic acid (2.14 g, 17.50 mmol, 2.0 eq.) and copper acetate monohydrate (524 mg, 2.63 mmol, 0.3 eq.) were dissolved in DMSO (15 mL). The reaction solution was replaced with oxygen three times and stirred at 110 ° C overnight. After the reaction was completed, the reaction solution was diluted with water (150 mL), filtered, and the filtrate was extracted with ethyl acetate (3×60 mL). The organic phases were combined and washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was separated and purified by silica gel column chromatography (eluent: ethyl acetate: petroleum ether = 1:10) to obtain B113-2 (210 mg, 12%). LCMS: m/z=191.1[M+H] ⁺ ; ¹ H NMR (400MHz, CDCl ₃ ) δ8.37 (s, 1H), 8.17–8.12 (m, 2H), 7.57–7.48 (m, 3H).

3) Synthesis of intermediate B113-3:

B113-2 (9.0 g, 47.33 mmol, 1.0 eq), ammonium chloride (7.6 g, 141.98 mmol, 3.0 eq) and iron powder (7.9 g, 141.98 mmol, 3.0 eq) were mixed in ethanol (90 mL) and water (90 mL). The reaction solution was stirred at 70 ° C for 2 h under nitrogen protection. After the reaction was completed, the reaction solution was filtered with diatomaceous earth and the filtrate was extracted with ethyl acetate (200 mL × 3). The organic phases were combined and washed with saturated brine (200 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain a yellow solid B113-3 (8.5 g, 95%). LCMS m/z＝161.1[M+H] ^+.1 H NMR (400MHz, DMSO-d ₆ ) δ7.82–7.78(m,2H),7.48–7.44(m,2H),7.26(s,1H),6.76–6.73(m,1H),5.50(s,2H).

4) Synthesis of intermediate B113-4:

B113-3 (2.0 g, 9.79 mmol, 1.0 eq), methyl 3-oxopentanoate (10.2 g, 78.34 mmol, 8.0 eq) and anhydrous magnesium sulfate (2 g) were dissolved in toluene (10 mL). The reaction solution was stirred at 110 ° C overnight. After the reaction was completed, the reaction solution was diluted with dichloromethane (50 ml) and filtered through diatomaceous earth. The filtrate was concentrated under reduced pressure to obtain a crude product, which was separated and purified by silica gel column chromatography (eluent: ethyl acetate: petroleum ether = 1:2) to obtain B113-4 (2.9 g, 93% yield). LCMS m/z = 273.2 [M + H] ⁺

5) Synthesis of intermediate B113-5:

The solid B113-4 (310 mg, 1.1 mmol, 1.0 eq) was directly poured into diphenyl ether (10 mL) at 235 ° C. The reaction solution was stirred at 235 ° C for 1 h. After the reaction was completed, the reaction solution was cooled to room temperature, diluted with petroleum ether (20 mL) and stirred for 20 minutes. The reaction mixture was filtered and washed with petroleum ether, and the filter cake was dried under reduced pressure to obtain a crude product, which was separated and purified by silica gel column chromatography (eluent: dichloromethane: methanol = 40: 1) to obtain B113-5 (80 mg, 30% yield). LCMS m/z＝241.1[M+H] ^+.1 H NMR (400MHz, DMSO-d ₆ )δ12.40(s,1H),8.14(d,J＝8.0Hz,2H),7.64(t,J＝8.0Hz,2H),7.58–7.49(m,1H),5.93(s,1H),2.63(q,J＝7.6Hz,2H),1.25(t,J＝7.6Hz,3H).

6) Synthesis of intermediate B113-6:

Under nitrogen protection, diisopropylamine (303 mg, 3 mmol, 1.0 eq.) was dissolved in anhydrous tetrahydrofuran (1.3 mL) and cooled to -78 °C. n-Butyl lithium (2.5 M in hexane, 1.2 mL, 3 mmol, 1.0 eq) was slowly added dropwise to the reaction system, and the reaction solution was reacted at -78 °C for 30 minutes.

Under nitrogen protection, B113-5 (29 mg, 0.12 mmol, 1.0 eq) was dissolved in anhydrous tetrahydrofuran (2 mL) and anhydrous DMSO (0.2 mL), and the reaction solution was cooled to 0°C. The above-prepared LDA solution (0.6 mL, 0.6 mmol, 5.0 eq) was slowly added to the reaction solution, and the reaction solution was reacted at 0°C for 30 minutes. (S)-Chloroepoxypropane (22 mg, 0.24 mmol, 2.0 eq) was dissolved in anhydrous tetrahydrofuran (0.4 mL), slowly added dropwise to the reaction solution and stirred at room temperature overnight. After the reaction was completed, the reaction solution was quenched with saturated ammonium chloride (10 mL), extracted with ethyl acetate (3×5 mL), and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was separated and purified by thin layer preparative chromatography (eluent: dichloromethane: methanol = 10:1) to obtain B113-6 (10 mg, 28%) as a light yellow solid. LCMS m/z = 297.1 [M + H] ⁺

7) Synthesis of intermediate B113-7:

Under nitrogen protection, B113-6 (298 mg, 1.0 mmol, 1.0 eq) was dissolved in anhydrous dichloromethane (12 mL), the reaction solution was cooled to 0 ° C, and N-bromosuccinimide (268 mg, 1.5 mmol, 1.5 eq) was slowly added to the reaction solution. The reaction solution was slowly warmed to room temperature and stirred at room temperature overnight. After the reaction was completed, the reaction solution was quenched by adding water (20 mL), extracted with dichloromethane (3×10 mL), and the organic phases were combined and washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain B113-7 (375 mg, 100% crude product), which was a red solid. LCMS m/z＝375.1[M+H] ⁺

8) Synthesis of intermediate B113-8:

B113-7 (375 mg, 1.0 mmol, 1.0 eq), sodium bicarbonate (840 mg, 10 mmol, 10.0 eq), potassium bromide (12 mg, 0.1 mmol, 0.1 eq), TOMAC (21 mg, 0.05 mmol, 0.05 eq) and TEMPO (8 mg, 0.05 mmol, 0.05 eq) were dissolved in dichloromethane (12 mL) and water (9 mL). The reaction solution was cooled to 0 ° C, and a 5% sodium hypochlorite aqueous solution (7.5 g, 5.0 mmol, 5.0 eq) was slowly added dropwise to the reaction solution. The reaction solution was slowly warmed to room temperature and stirred at room temperature for 2.5 hours. After the reaction was completed, the reaction solution was quenched by adding sodium bicarbonate solution and extracted with dichloromethane (3×20 mL). The resulting aqueous phase was adjusted to pH 3 with 3M hydrochloric acid solution and then extracted with ethyl acetate (3×20 mL). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give B113-8 (120 mg, 31%) as a light yellow solid. LCMS m/z = 389.1 [M + H] ⁺

9) Synthesis of intermediate B113-10:

Under nitrogen protection, compound B113-8 (120 mg, 0.31 mmol, 1.0 eq), B113-9 (70 mg, 0.37 mmol, 1.2 eq) and N, N-diisopropylethylamine (160 mg, 1.23 mmol, 4.0 eq) were dissolved in anhydrous dichloromethane (2.5 mL), and then CMPI (119 mg, 0.46 mmol, 1.5 eq) was added, and the reaction solution was stirred at room temperature for 1.5 hours. After the reaction was completed, the reaction solution was quenched by adding water (20 mL), extracted with ethyl acetate (3×20 mL), and the organic phases were combined and washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was separated and purified by silica gel column chromatography (eluent: dichloromethane: methanol = 150: 1) to obtain B113-10 (85 mg, 46% yield), which was a light yellow solid. LCMS m/z＝558.1[M+H] ^+.1 H NMR (400MHz, CDCl ₃ )δ10.09(s,1H),8.12–8.03(m,1H),7.86–7.77(m,1H),7.52–7.41(m,1H),7.38–7.28(m,3H),6.88(d,J＝8.5Hz,1H),5 .43–5.32(m,1H),3.31–3.09(m,1H),2.79–2.63(m,1H),2.41–2.36(m,3H),2.10–1.90(m,1H),1.41(d,J＝7.1Hz,3H).

10) Synthesis of intermediate B113-12:

Under nitrogen protection, B113-10 (53 mg, 0.09 mmol, 1.0 eq), B113-11 (160 mg, 1.42 mmol, 15.0 eq), N,N-diisopropylethylamine (48 mg, 0.47 mmol, 5.0 eq) and silver tetrafluoroborate (28 mg, 0.14 mmol, 1.5 eq) were dissolved in dry DMSO (0.8 mL), and the reaction solution was stirred at 125°C for 3.5 hours. After the reaction was completed, the reaction solution was diluted with methanol (2 mL), filtered, and the filtrate was directly separated and purified by C18 column chromatography (H ₂ O (0.1% formic acid) containing 10-60% ACN) to obtain B113-12 (30 mg, crude product, 50% purity) as a yellow solid. LCMS m/z=590.30 [M+H] ⁺ .

11) Synthesis of product B113:

Under nitrogen protection, B1-16 (9.8 mg, 0.063 mmol) and HOAt (8.8 mg, 0.065 mmol) were dissolved in acetonitrile (2 mL), EDCI (16.1 mg, 0.084 mmol) was added to the reaction solution, and stirred at room temperature for 1 hour. Subsequently, B113-12 (25 mg, crude product, 50% purity) and N, N-diisopropylethylamine (18.9 mg, 0.14 mmol, 3.5 eq) were added to the reaction solution, and the reaction solution continued to stir at room temperature for 0.5 hours. After the reaction was completed, the reaction solution was quenched by adding water (20 mL), extracted with ethyl acetate (3×15 mL), and the organic phases were combined and washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was separated and purified by thin layer preparative chromatography (eluent: dichloromethane: methanol = 10:1) to obtain B113 (1 mg). LCMS m/z＝726.2[M+H] ^+.1 H NMR (400MHz, CD ₃ OD)δ8.56–8.52(m,1H),8.28–8.15(m,2H),7.63–7.57(m,2H),7.55–7.49(m,1H),7 .13–6.96(m,2H),5.50–5.42(m,1H),4.79–4.66(m,1H),4.00–3.88(m,3H),3.78–3. 69(m,2H),3.57–3.48(m,1H),3.18–3.13(m,1H),2.65–2.54(m,1H),2.52–2.50(m,3 H),2.31–2.22(m,4H),1.82–1.67(m,1H),1.61(d,J=7.3Hz,3H),1.55–1.44(m,2H).

Biological testing section

Experimental Example 1 Biological Activity Test

ATPase activity assay of WRN helicase

The commercial ADP-Glo assay kit (Promega, #V9102) was used to detect the ADP content generated by the hydrolysis of ATP by WRN helicase, which can reflect the ATPase activity of WRN helicase.

45 nt oligo DNA single strand FLAP26 (TTTTTTTTTTTTTTTTTTTTTTCCAAGTAAAACGACGGCCAGTGC) was synthesized by Ascent Inc. See, for example, Brosh RM Jr et al., J Biol Chem, 2002 Jun; 277(26):23236-45. Prepare reaction buffer (30mM Tris pH 7.5, 2mM MgCl ₂ , 0.02% BSA, 50mM NaCl, 0.1% pluronic F127), add 5μL 3× test compound (diluted to 0.5% DMSO in reaction buffer, final starting concentration is 10μM, 1:3 dilution, 9 gradients), 5μL 3× WRN recombinant protein and 3× ATP substrate solution (diluted in reaction buffer, final concentrations of WRN and ATP are 10nM and 300μM, respectively) to a 384-well transparent plate, shake and mix, and incubate at 37°C for 3 hours. Next, prepare 3× FLAP26 solution with reaction buffer, take 5μL solution (final concentration of FLAP26 is 0.4nM) and add it to a 384-well transparent plate, shake and mix, start the enzymatic reaction, and incubate at room temperature for 30 minutes.

Take 5 μL of the above mixture and transfer it to a 384-well white plate, add 5 μL of ADP-Glo reagent, shake and mix, and incubate at room temperature in the dark for 40 minutes. Add 10 μL of kinase detection reagent to the above solution, shake and mix, incubate at room temperature in the dark for 30 minutes, and record the chemiluminescence reading. Calculate the inhibition rate of the compound on enzyme activity, and use nonlinear regression (dose response-variable slope) to fit the inhibition rate value and the logarithm of the compound concentration to obtain the IC ₅₀ value of the test compound.

Table 1 WRN ATPase activity data of the compounds in the examples of this application

As can be seen from Table 1, the compounds of the present application have good inhibitory activity on the ATPase activity of the WRN protein.

Tumor cell proliferation inhibitory activity assay

The WRN gene was stably knocked out in DLD1 cells using CRISPR/Cas9 technology to construct the DLD1-WRN-KO cell line, which was used to evaluate the potential off-target effects of the compounds.

After the test compound was treated with the microsatellite unstable SW48 cell line and the control cell DLD1-WRN-KO cell line for 4 days, the ATP level was detected using the CellCounting-Lite kit of Novozymes to evaluate the inhibitory effect of the test compound on the growth of tumor cell lines.

In the present application, SW48 cell line and DLD1-WRN-KO cell line were inoculated in 96-well cell culture plates at an appropriate cell density. After 24 hours, the test compound was used to treat the cells at a maximum concentration of 10 μM, 1:3 dilution ratio of 9 gradients, and a DMSO treatment group was set up. Cultured in a 37°C/5% _CO2 incubator for 4 days. To test the inhibition of tumor cell proliferation by the test compound, the cells were balanced at room temperature for 30 minutes, and then 100 μL of cell proliferation detection reagent CellCounting-Lite (CCL) was added to each well, and incubated in the dark for 10 minutes after oscillation for 5 minutes. The chemiluminescence value was read using a Thermo Varioskan LUX-3020 multifunctional microplate reader to convert it into a proliferation index to calculate the inhibition rate of tumor cell proliferation by the compound, and the inhibition rate value and the logarithm of the compound concentration were fitted using nonlinear regression (dose response-variable slope) to obtain the _IC50 value of the compound.

Table 2 Tumor cell proliferation inhibition activity data of the compounds in the examples of the present application

As can be seen from Table 2, the compounds of the present application have good proliferation inhibition activity on microsatellite unstable SW48 cells, but have no significant proliferation inhibition activity on WRN knockout DLD1 cells, and have good selectivity.

Experimental Example 2 Liver microsome stability test

The first-phase metabolic stability of the test compounds was assessed in liver microsomes of CD-1 mice, Sprague-Dawley rats, beagle dogs, cynomolgus monkeys and humans.

Experimental system:

The animal and human liver microsomes used in this test system were purchased from Xenotech, Corning or other qualified suppliers and stored in a freezer below -60°C before use.

Experimental introduction:

The test sample and the control compound were incubated with animal and human liver microsomes at 37±1°C for a certain period of time, with the longest incubation time being 60 minutes. Samples were taken out at the specified time point and the reaction was terminated with acetonitrile or other organic solvents containing internal standards. After centrifugation, the resulting supernatant was detected by liquid chromatography-tandem mass spectrometry (LC-MS/MS).

Experimental methods:

1. Preparation of buffer

Dissolve 73.21 g of potassium phosphate trihydrate and 10.78 g of potassium dihydrogen phosphate in 4000 mL of ultrapure water, and adjust the pH value of the solution to 7.40±0.10 using 10% phosphoric acid or 1M potassium hydroxide, with a final concentration of 100 mM.

2. Preparation of working solution

The test sample powder is prepared into a stock solution of a certain concentration using DMSO or other organic solvents, and then further diluted with a suitable organic solvent.

The control compounds testosterone, diclofenac and propafenone were prepared as 10 mM stock solutions in DMSO and then further diluted in appropriate organic solvents.

3. Preparation of Liver Microsome Solution

Dilute each microsome to a 2× working solution using 100 mM potassium phosphate buffer. The final concentration of microsomes in the reaction system is 0.5 mg/mL.

4. Preparation of reduced nicotinamide adenine dinucleotide phosphate (NADPH) regeneration system

Weigh an appropriate amount of nicotinamide adenine dinucleotide phosphate (NADP) and isocitrate (ISO) powder, dissolve in magnesium chloride solution, and shake to mix. Add an appropriate amount of isocitrate dehydrogenase (IDH) and gently mix upside down. The final concentrations in the reaction system are: 1mM NADP, 1mM magnesium chloride, 6mM ISO, and 1unit/mL IDH.

5. Preparation of Stop Solution

The stop solution is prepared with acetonitrile or other organic solvents containing an internal standard (tolbutamide or other suitable compounds). The prepared stop solution is stored in a refrigerator at 2-8°C.

6. Incubation process

Incubations will be done in 96-well plates. Prepare 8 incubation plates, named T0, T5, T15, T30, T45, T60, Blank60, and NCF60. The first 6 plates correspond to reaction time points of 0, 5, 15, 30, 45, and 60 minutes, respectively. No test or control compound is added to the Blank60 plate, and samples are taken after 60 minutes of incubation. In the NCF60 plate, potassium phosphate buffer is used instead of NADPH regeneration system solution for incubation for 60 minutes. All condition samples are three parallels.

Mix the microsomes with the test or control compound, and then place the incubation plates Blank60, T5, T15, T30, T45 and T60, except T0 and NCF60, in a 37°C water bath for pre-incubation for about 10 minutes. Add the stop solution to the incubation plate T0 first, then add the NADPH regeneration system working solution, and add 98μL potassium phosphate buffer to each sample well of the incubation plate NCF60 to start the reaction. After the pre-incubation of the incubation plates Blank60, T5, T15, T30, T45 and T60, add 98μL NADPH regeneration system working solution to each sample well to start the reaction. The reaction temperature is 37±1°C, the final reaction volume is 200μL, and the reaction system includes 0.5mg/mL microsomes, 1.0μM substrate, 1mM NADP, 6mM ISO and 1unit/mL IDH.

At 5, 15, 30, 45 and 60 minutes, cold stop solution containing internal standard was added to the reaction plate to stop the reaction.

All reaction plates after termination were shaken and centrifuged at 4°C, 3220×g for 20 minutes. The supernatant was diluted to a certain ratio and then analyzed by LC-MS/MS.

Sample analysis

Sample analysis was performed using liquid chromatography-tandem mass spectrometry (LC-MS/MS) without standard curves and quality control samples. Semi-quantitative determination was performed using the ratio of the analyte peak area to the internal standard peak area. Retention times of analytes and internal standards, chromatogram acquisition, and chromatogram integration were processed using the software Analyst (Sciex, Framingham, Massachusetts, USA).

The CV of the internal standard peak area in each matrix should be within 20% for each analytical run.

Data analysis

The in vitro elimination rate constant ke of the compound was obtained by converting the ratio of the peak area of the compound to the internal standard into the residual rate in the following formula:

CL _int(mic) = 0.693/T _1/2 /microsomal protein content (microsomal concentration during incubation mg/mL)

CL _int(liver) = CL _int(mic) × amount of microsomal protein in liver (mg/g) × liver weight to body weight ratio

According to the well stir model, the hepatic intrinsic clearance and hepatic clearance can be converted by the following formula.

CL _(liver) = (CL _{int (liver)} *Q _h )/(CL _{int (liver)} +Q _h )

The parameters in the formula are shown in the table below.

Parameters in Data Analysis Formulas

Experimental Example 3 Hepatocyte Metabolic Stability Test

This experiment was used to test the metabolic stability of compounds in hepatocytes.

Prepare 0.5x10 ⁶ /mL hepatocyte suspension with preheated culture medium, then add 198 μL preheated cell suspension to 96-well plate. Add 2 μL of the test compound to each well of the 96-well plate to make a final concentration of 1 μM, and set 2 replicates. For samples with T=0 minutes, mix the compound and cells thoroughly for 1 minute, then immediately take 25 μL of the sample and add it to 125 μL of stop solution (acetonitrile solution containing 200 ng/mL tolbutamide and 200 ng/mL labetalol) in an ice bath and mix. At the same time, place all plates in an incubator at 37°C and 5% CO ₂ , and set the shaker to 600 rpm. The samples were mixed at 15, 30, 60 and 90 minutes of incubation, and 25 μL of the sample was added to 125 μL of stop solution (acetonitrile solution containing 200 ng/mL tolbutamide and 200 ng/mL labetalol) in an ice bath, and then shaken at 500 rpm for 10 minutes. Subsequently, centrifugation was performed at 3220×g for 20 minutes at 4°C. After centrifugation, 80 μL of supernatant was taken from each well and transferred to another 96-well plate containing 240 μL of ultrapure water. The intrinsic clearance (CLint) and half-life (T1/2) were then analyzed and calculated by LC-MS/MS.

Experimental Example 4 Pharmacokinetic Test in Rats

In this experiment, the pharmacokinetic behavior of the test compound in SD rats was investigated after intravenous (IV) and oral (PO) administration.

On the day of administration, the actual body weight of the rats was weighed and the administration volume was calculated. There were 3 rats in each group, and two groups of tests were performed for each compound, one group was administered with a single intravenous injection, and the other group was administered with a single oral gavage. Whole blood samples were collected at the specified time (0.25, 0.5, 1, 2, 4, 8, 24h after administration) by jugular vein blood collection. After blood sample collection, it was immediately transferred to a labeled commercial sample tube containing K2-EDTA (0.85-1.15mg), followed by centrifugation (3200x g, 4°C, 10 minutes) and plasma was collected. The plasma was transferred to a pre-cooled centrifuge tube, snap-frozen in dry ice, and then stored in an ultra-low temperature freezer at -60°C or lower until LC-MS/MS analysis.

Plasma concentrations were determined using LC-MS/MS. The plasma drug concentration data of the compounds were processed using WinNonlin Version 6.3 (Pharsight, Mountain View, CA) pharmacokinetic software with a non-compartmental model. The linear-log trapezoidal method was used to calculate the relevant pharmacokinetic parameters.

Experimental Example 5: Pharmacokinetic test in mice

In this experiment, the pharmacokinetic behavior of the tested compounds was investigated in BALB/c mice after intravenous (IV) and oral (PO) administration.

On the day of administration, the actual body weight of the mice was weighed and the administration volume was calculated. There were 9 mice in each group, and two groups of tests were performed for each compound, one group was administered with a single intravenous injection, and the other group of mice was administered with a single oral gavage. Whole blood samples were collected at the specified time (0.25, 0.5, 1, 2, 4, 8, 24h after administration) by orbital bleeding. After blood sample collection, it was immediately transferred to a labeled commercial sample tube containing K2-EDTA (0.85-1.15mg), followed by centrifugation (3200x g, 4°C, 10 minutes) and plasma was collected. The plasma was transferred to a pre-cooled centrifuge tube, snap-frozen in dry ice, and then stored in an ultra-low temperature freezer at -60°C or lower until LC-MS/MS analysis.

Plasma concentrations were determined using LC-MS/MS. The plasma drug concentration data of the compounds were processed using WinNonlin Version 6.3 (Pharsight, Mountain View, CA) pharmacokinetic software with a non-compartmental model. The linear-log trapezoidal method was used to calculate the relevant pharmacokinetic parameters.

Experimental Example 6 hERG inhibition test

HEK293 cells were cultured in DMEM medium containing 10% fetal bovine serum and 0.8mg/mL G418 at 37℃ and 5% _CO2 . The cells were digested with TrypLE ^TM Express and centrifuged to adjust the cell density to 2×10 ⁶ cells/mL. The cells were then gently mixed on a room temperature balanced shaker for 15-20min and then put on the machine for patch clamp detection. The culture medium of the prepared cells was replaced with extracellular fluid. The intracellular and extracellular fluids were drawn from the liquid pool and added to the intracellular fluid pool, cell and test substance pool of the QPlate chip respectively. The whole-cell patch clamp recorded the voltage stimulation of the whole-cell hERG potassium current, and the experimental data were collected and stored by Qpatch. The compound started at 30μM, diluted 3 times, and 6 concentration points were set. Each drug concentration was set to be administered twice for at least 5 minutes. The current detected in the external solution without the compound for each cell was used as its own control group. At least two cells were used for each concentration and the detection was repeated twice independently. All electrophysiological experiments were performed at room temperature.

Data analysis: First, the current after each drug concentration was normalized with the blank control current. Then calculate the inhibition rate corresponding to each drug concentration For each concentration the mean and standard error were calculated and the half-inhibitory concentration for each compound was calculated: The above equation was used to perform nonlinear fitting of the dose-dependent effect, where Y represents the inhibition rate, C represents the concentration of the test substance, _IC50 represents the half-inhibitory concentration, and HillSlope represents the Hill coefficient. Curve fitting and _IC50 calculation were completed using Graphpad software.

Experimental Example 7 Cytochrome oxidase P450 inhibition test

1) Preparation of buffer:

100mM K-Buffer: Mix 9.5mL of stock solution A into 40.5mL of stock solution B, adjust the total volume to 500mL with ultrapure water, and titrate the buffer to pH 7.4 with KOH or H ₃ PO ₄ .

Raw material A (1M potassium dihydrogen phosphate): 136.5 g potassium dihydrogen phosphate in 1 L water;

Stock B (1M KH2PO4): 174.2 g KH2PO4 in 1 L water.

2) Preparation of test substances

The test substance powder is prepared into a stock solution of a certain concentration using DMSO or other organic solvents, and then further diluted with a suitable organic solvent.

3) In vitro incubation

The in vitro incubation system of liver microsomes for the study of CYP450 enzyme metabolic phenotype is a biochemical reaction carried out under conditions simulating physiological temperature and physiological environment, with the prepared liver microsomes supplemented with redox coenzymes and enzyme-specific selective inhibitors.

4) Detection of parent drug or metabolites

The concentration of parent drug or its metabolites in the incubation solution was determined by LC-MS/MS.

Experimental Example 8 Mouse Tumor Pharmacodynamic Model

In this experimental example, the in vivo efficacy of the tested compounds in a mouse xenograft tumor model was evaluated after intragastric administration (PO).

Human colon adenocarcinoma cell SW48 cells were cultured in monolayer in vitro, and the culture conditions were: DMEM medium containing 10% fetal bovine serum, 1% penicillin and streptomycin, and cultured at 37°C and 5% _CO2 . The cells were digested and passaged twice a week with trypsin. When the cell saturation was 80%-90%, the cells were collected, counted, and inoculated. 0.1mL ( ¹⁰⁷ ) of SW48 cells were subcutaneously inoculated into the right back of each mouse. On the 14th day after cell inoculation, the average tumor volume reached about ^200mm3 , and the mice were randomly divided into groups for drug administration, and the drug was administered by gavage once a day, and the changes in body weight and tumor volume were recorded. After a certain number of days of drug administration, the experiment was terminated. The changes in tumor volume and mouse weight were statistically analyzed.

In addition to those described herein, various modifications of the present invention will be apparent to those skilled in the art based on the foregoing description. Such modifications are also intended to fall within the scope of the appended claims. Each reference cited in this application (including all patents, patent applications, journal articles, books and any other disclosures) is incorporated herein by reference in its entirety.

Claims (14)

A compound or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, wherein the compound has the structure of formula (I):
in: Selected from W is CR ¹ or N; ^R1 at each occurrence is independently selected from H, halogen, -OH, _-NH2 , -CN, _-NO2 , _-SF5 , _C1-6 alkyl, deuterated _C1-6 alkyl, halogenated _C1-6 alkyl, _C2-6 alkenyl, _C2-6 alkynyl, _C3-6 cycloalkyl, 3-10 membered heterocyclyl, _C6-10 aryl, 5-14 membered heteroaryl, _C6-12 aralkyl, -C(=O) ^Ra , -OC(=O) ^Ra , -C(=O) ^ORa , ^-ORa , ^-SRa , -S(=O) ^Ra , -S(=O) _2Ra , ^-S (=O ^{)2NRaRb , -NRaRb} , -C ₍ =O) ^NRaRb , -NRa ^{- C} (=O) ^{Rb , -NRa} -C(═O)OR ^b , -NR ^a -S(═O) ₂ -R ^b , -NR ^a -C(═O)-NR ^a R ^b , -P(═O)R ^a R ^b , -C _1-6 alkylene-R ^a , -C _1-6 alkylene-OR ^a , -C _1-6 alkylene-NR ^a R ^b , -OC _1-6 alkylene-NR ^a R ^b , (—C _3-6 cycloalkylene)-CN and (—C _3-6 cycloalkylene)-C _1-6 haloalkyl; Alternatively, two R ^1s in the ortho position together with the groups to which they are attached optionally together form a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring; ^R2 is R ³ , R ²¹ , R ²² , R ²³ and R ²⁴ are each independently selected at each occurrence from H, halogen, -OH, -NH ₂ , -CN, -NO ₂ , -SF ₅ , C _1-6 alkyl, deuterated C _1-6 alkyl, halogenated C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, C _6-12 aralkyl, -C(═O)R ^{a , -OC(═O)R a ,} -C(═O)OR ^a , -OR ^a , -SR ^a , -S(═O)R ^a , -S(═O) ₂ R ^a , -S(═O) ₂ NR ^a R ^b , -S(═O)(═NR ^a )R ^b , -NR ^a R ^b , —C(═O)NR ^a R ^b , —NR ^a -C(═O)R ^b , —NR ^a -C(═O)R b , —NR ^a -C(═O)OR ^b , —NR ^a -S(═O) ₂ -R ^b , —NR ^{a -C} (═O)-NR ^a R ^b , —P(═O)R a R b , —C _1-6 alkylene-R ^a , —C _1-6 alkylene-OR ^a , —C _1-6 alkylene-NR ^a R ^b , —OC _1-6 alkylene-NR ^a R ^b , (—C _3-6 cycloalkylene)-CN and (—C _3-6 cycloalkylene)-C _1-6 haloalkyl; Alternatively, R ³ and R ²¹ or R ²² together with the group to which they are attached optionally together form a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring; ^R4 is R ⁴¹ is selected from a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring, and a 5-14 membered heteroaromatic ring; Ring X and Ring Z are each independently selected from a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring, and a 5-14 membered heteroaromatic ring; Ring Y is absent or is selected from a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring and a 5-14 membered heteroaromatic ring; when Ring Y is absent, R ²⁴ is also absent; ^L2 is selected from -O-, -C(=O)-, -NRC(=O)-, -S-, -S(=O)-, -S(=O) _2- , _C1-6 alkylene and -O-( _C1-6 alkylene)-; R, ^Ra and ^Rb are each independently selected at each occurrence from H, _C1-6 alkyl, _C3-10 cycloalkyl, 3-10 membered heterocyclyl, _C6-10 aryl, 5-14 membered heteroaryl and _C6-12 aralkyl; The above alkylene, alkyl, alkenyl, alkynyl, cycloalkylene, cycloalkyl, hydrocarbon ring, heterocyclyl, heterocycle, aryl, aromatic ring, heteroaryl, heteroaromatic ring and aralkyl are each optionally substituted at each occurrence by one or more substituents independently selected from the following: deuterium atom, halogen, -OH, =O, -NH2, -CN, _-NO2 , = _CH2 , _C1-6 alkyl, deuterated _{C1-6 alkyl} , _C2-6 alkenyl, _C2-6 alkynyl, _C3-6 cycloalkyl, 3-10 membered heterocyclyl, _C6-10 aryl, 5-14 membered heteroaryl, _C6-12 aralkyl, -C(=O) ^Rc , -OC(=O) ^Rc , -C(=O) ^ORc , ^-ORc , ^-SRc , -S(=O) ^Rc , -S(= ^O ) _2Rc -S(＝O) _2NRcRd , ^-NRcRd , ^-C (＝O) ^NRcRd , ^-NRc- C( ^＝ O) ^Rd , -NRc ^- C(＝O) ^ORd , -NRc ^- S(＝O) ₂ - ^Rd , -NRc ^{- C} (＝O)-NRcRd, -C1-6alkylene ^{- ORc} , _{-C1-6alkylene} - ^NRcRd and _{-OC1-6alkylene} - ^NRcRd ; when the same ^ring atom or adjacent ring atoms of a cycloalkylene, ^cycloalkyl , hydrocarbon ^ring , _heterocyclyl , heterocycle, ^aryl , aromatic ring, heteroaryl and heteroaromatic ring are substituted by two substituents, the two substituents together with the group to which they are attached optionally constitute a _C3-6 hydrocarbon ring, a 3-10 membered heterocycle, a C1-6alkylene ring or a heteroaryl ring. _6-10 membered aromatic ring or 5-14 membered heteroaromatic ring; each of the alkylene, alkyl, alkenyl, =CH ₂ , alkynyl, cycloalkyl, hydrocarbon ring, heterocyclyl, heterocycle, aryl, aromatic ring, heteroaryl, heteroaromatic ring and aralkyl is further optionally substituted by one or more substituents independently selected from the following: halogen, -OH, =O, -C(=O)O-tert-butyl, -NH ₂ , -CN, -NO ₂ , C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, C _6-12 aralkyl, -C _1-6 alkylene-C _3-6 cycloalkyl, -OC _1-6 alkyl and -C _1-6 alkylene-OC _1-6 alkyl; R ^c and R ^d are each independently selected at each occurrence from H, C _1-6 alkyl, C _3-10 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, and C _6-12 aralkyl, said alkyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, and aralkyl being further optionally substituted with one or more substituents independently selected from the group consisting of halogen, -OH, =O, -C(=O)O-tert-butyl, -NH ₂ , -CN, -NO ₂ , C _1-6 alkyl, C _1-6 haloalkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, C _6-12 aralkyl _{, and -C 1-6 alkylene} -OC _1-6 alkyl; and p, q and t are each independently an integer selected from 1, 2 or 3. The compound of claim 1 or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, wherein the compound has the structure of the following formula:

in: Ring C and Ring D are each independently a C _3-6 hydrocarbon ring, a 3-10 membered heterocycle, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring, and the hydrocarbon ring, heterocycle, aromatic ring and heteroaromatic ring are each optionally substituted by one or more substituents independently selected from the following: a deuterium atom, a halogen, -OH, a C _1-6 alkyl group, a halogenated C _1-6 alkyl group and a -OC _1-6 alkyl group; when the same ring atom or adjacent ring atoms of the hydrocarbon ring, heterocycle, aromatic ring and heteroaromatic ring are substituted by two substituents, the two substituents together with the group to which they are attached optionally constitute a C _3-6 hydrocarbon ring, a 3-10 membered heterocycle, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring; Preferably, ring C is a benzene ring or a pyridine ring, each of which is optionally substituted by one or more substituents independently selected from the group consisting of halogen, C _1-6 alkyl and halogenated C _1-6 alkyl; and/or Ring D is a cyclopentene ring, a cyclohexene ring, a pyrrolidine ring, a piperidine ring or a morpholine ring, each of which is optionally substituted by one or more substituents independently selected from the following: a deuterium atom, a halogen, -OH, a C _1-6 alkyl group, a halogenated C _1-6 alkyl group and a -OC _1-6 alkyl group; when the same ring atom or adjacent ring atoms of ring D are substituted by two substituents, the two substituents together with the groups to which they are attached optionally constitute a C _3-6 hydrocarbon ring or a 3-10 membered heterocyclic ring; The remaining groups are as defined in claim 1. The compound of claim 1 or 2, or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, wherein: R ¹ is independently selected from H, halogen, C _1-6 alkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl, 5-14 membered heteroaryl, -OR ^a and -NR ^a R ^b at each occurrence, preferably, R ¹ is independently selected from H, halogen, C _1-6 alkyl, C _3-6 cycloalkyl and -OR ^a at each occurrence; wherein the alkyl, cycloalkyl, heterocyclyl, aryl and heteroaryl are each optionally substituted with one or more substituents independently selected from the following: halogen, C _1-6 alkyl, C _2-6 alkenyl, =CH ₂ , C _3-6 cycloalkyl, 3-10 membered heterocyclyl, C _6-10 aryl and 5-14 membered heteroaryl; the alkyl, alkenyl, =CH ₂ , cycloalkyl, heterocyclyl, aryl and heteroaryl are each further optionally substituted with one or more substituents independently selected from the following: halogen, C 1-6 alkyl, C 3-6 alkenyl, =CH 2 , C 3-6 cycloalkyl, heterocyclyl, aryl and heteroaryl. _{C 1-6} alkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclic group and -C _1-6 alkylene-C _3-6 cycloalkyl; Preferably, R ¹ is H, methyl, halogen, methoxy, Alternatively, two R ^1s in the ortho position together with the groups to which they are attached optionally constitute a C _3-6 hydrocarbon ring, a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring or a 5-14 membered heteroaromatic ring, wherein each of the hydrocarbon ring, heterocyclic ring, aromatic ring and heteroaromatic ring is optionally substituted by one or more substituents independently selected from the following: halogen, C _1-6 alkyl and halogenated C _1-6 alkyl; Preferably, the two R ¹ in the ortho position together with the group to which they are attached optionally constitute A benzene ring or a pyridine ring, each of which is optionally substituted by one or more substituents independently selected from the group consisting of halogen, C _1-6 alkyl and halogenated C _1-6 alkyl. A compound according to any one of claims 1 to 3 or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, wherein Ring X is a benzene ring, a 5-6 membered heterocyclic ring or a 5-6 membered heteroaromatic ring (preferably a thiophene ring, a thiazole ring or a pyridine ring), and Ring Y does not exist; or Ring X is a benzene ring or a 5-6 membered heteroaromatic ring, and Ring Y is a C _3-6 hydrocarbon ring, a 5-6 membered heterocyclic ring or a 5-6 membered heteroaromatic ring; Preferably, for More preferably A compound according to any one of claims 1 to 4, or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, wherein R ²¹ , R ²² , R ²³ and R ²⁴ are each independently selected from H, halogen, -SF ₅ , C _1-6 alkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl, -O-(C _1-6 alkyl), -S(=O) ₂ -(C _1-6 alkyl), -S(=O) ₂ -(C _3-6 cycloalkyl), -P(=O)(C _1-6 alkyl) ₂ , (-C _3-6 cycloalkylene)-CN and (-C _3-6 cycloalkylene)-C C _1-6 alkyl, wherein the alkyl, cycloalkylene, cycloalkyl and heterocyclyl are each optionally substituted by one or more substituents independently selected from the group consisting of halogen, C _1-6 alkyl and halogenated C _1-6 alkyl; Preferably, R ²¹ , R ²² , R ²³ and R ²⁴ are each independently selected from H, halogen, -SF ₅ , C _1-6 alkyl, C _3-6 cycloalkyl, 3-10 membered heterocyclyl and -O-(C _1-6 alkyl), said alkyl, cycloalkyl and heterocyclyl being each optionally substituted with one or more substituents independently selected from halogen, C _1-6 alkyl and halogenated C _1-6 alkyl. A compound according to any one of claims 1 to 5, or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, wherein Selected from: The compound of any one of claims 1 to 6, or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, wherein R ³ is H, C _1-6 alkyl, -OR ^a or -SR ^a ; preferably, R ³ is H, ethyl, -O-CH ₃ or -S-CH ₃ ; Alternatively, R ³ and R ²¹ or R ²² together with the group to which they are attached optionally together form a C _3-6 hydrocarbon ring or a 3-10 membered heterocyclic ring, and the hydrocarbon ring and the heterocyclic ring are optionally substituted by one or more substituents selected from deuterium atoms, halogen, -OH, C _1-6 alkyl, halogenated C _1-6 alkyl and -OC _1-6 alkyl. When the same ring atom or adjacent ring atoms of the hydrocarbon ring and the heterocyclic ring are substituted by two substituents, the two substituents together with the group to which they are attached optionally together form a C _3-6 hydrocarbon ring or a 3-10 membered heterocyclic ring; Preferably, R ³ and R ²¹ or R ²² together with the group to which they are connected optionally together constitute a cyclopentene ring, a cyclohexene ring, a pyrrolidine ring, a piperidine ring or a morpholine ring, and the cyclopentene ring, cyclohexene ring, pyrrolidine ring, piperidine ring and morpholine ring are optionally substituted with one or more substituents selected from deuterium atoms, halogen, -OH, C _1-6 alkyl, halogenated C _1-6 alkyl and -OC _1-6 alkyl; when the same ring atom of the cyclopentene ring, cyclohexene ring, pyrrolidine ring, piperidine ring and morpholine ring is substituted with two substituents, the two substituents together with the group to which they are connected optionally together constitute a C _3-6 hydrocarbon ring. The compound of any one of claims 1 to 7 or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, wherein ring Z is a 3-10 membered heterocycle or a benzene ring; preferably a 5-10 membered heterocycle; more preferably a 5-6 membered heterocycle; and The heterocyclic ring and the benzene ring are each optionally substituted at each occurrence by one or more substituents independently selected from the group consisting of halogen, C _1-6 alkyl and halogenated C _1-6 alkyl; Preferably, ring Z is A compound according to any one of claims 1 to 8, or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, wherein L ² is -C(=O)- or -NRC(=O)-, wherein R is H or C _1-6 alkyl; Preferably, ^L2 is -C(=O)-. A compound according to any one of claims 1 to 9, or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, wherein R ⁴¹ is selected from a 3-10 membered heterocyclic ring, a C _6-10 aromatic ring and a 5-14 membered heteroaromatic ring, wherein the heterocyclic ring, aromatic ring and heteroaromatic ring are each optionally substituted with one or more substituents independently selected from the following: halogen, -OH, C _1-6 alkyl, -OC _1-6 alkyl and -SC _1-6 alkyl, preferably, the heterocyclic ring, aromatic ring and heteroaromatic ring are substituted with at least -OH or -OC _1-6 alkyl; Preferably, -L ² -R ⁴¹ is The compound of any one of claims 1 to 10, or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, wherein R ⁴¹ is a 5-6-membered heteroaromatic ring, preferably a 6-membered heteroaromatic ring, more preferably a pyridine ring or a pyrimidine ring, which is substituted with at least one -OH; Preferably, -L ² -R ⁴¹ is A compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt, ester, stereoisomer, atropisomer, tautomer, polymorph, solvate, metabolite, isotopically labeled compound or prodrug thereof, wherein the compound is selected from:












A pharmaceutical composition comprising a preventively or therapeutically effective amount of a compound according to any one of claims 1 to 12 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, and a pharmaceutically acceptable carrier. Use of a compound according to any one of claims 1 to 12 or a pharmaceutically acceptable salt, ester, stereoisomer, tautomer, polymorph, solvate, metabolite, isotope-labeled compound or prodrug thereof, or a pharmaceutical composition according to claim 13 in the preparation of a medicament for use as a WRN inhibitor, preferably, the medicament is used to prevent or treat cancer (preferably, the cancer is characterized by microsatellite high instability (MSI-H) or mismatch repair deficiency (dMMR)); preferably, the cancer is selected from colorectal cancer, gastric cancer, endometrial cancer, uterine cancer, adrenocortical carcinoma, cervical cancer, esophageal cancer, breast cancer, kidney cancer, prostate cancer and ovarian cancer.