Pyridothiophene-2-carboxylic acid derivative, preparation method therefor and use thereof

Abstract

The present invention relates to a pyridothiophene-2-carboxylic acid derivative, a preparation method therefor and a use thereof, and in particular to a compound as shown in formula (I) or a pharmaceutical salt thereof, which has a good inhibition / degradation effect on BCKDK. Substituents in the formula are as defined in the description.

Description

PYRIDOTHIOPHENE-2-CARBOXYLIC ACID DERIVATIVE, PREPARATION METHOD THEREFOR AND USE THEREOF TECHNICAL FIELDThe present disclosure relates to the field of medicinal chemistry, and in particular to a pyridothiophene-2-carboxylic acid derivative, a preparation method therefor, and use thereof. BACKGROUNDBranched-chain keto acid dehydrogenase kinase (BCKDK) is associated with various human diseases. In addition to regulating the catabolism of branched-chain amino acids (BCAAs), BCKDK can also enhance MEK / ERK signaling associated with malignant proliferation of cancer.BCAAs (leucine, isoleucine, and valine) are essential amino acids in the human body, accounting for about 40% of the essential amino acids in healthy subjects. They must be obtained through a balanced diet and play an important role in nutrient sensing and cell signaling.Branched-chain amino acids are toxic in excess, but are necessary for protein synthesis and cell signaling processes. Branched-chain amino acids are converted by the branched-chain aminotransferase (BCAT) into the α-keto acid form: α-ketoisocaproic acid (KIC / ketoleucine), 2-keto-3-methylpentanoic acid (KMV / ketoisoleucine), and α-ketoisopentanoic acid (KIV / ketovaline). Branched-chain keto acids (BCKAs) are then oxidized and decarboxylated by a branched-chain keto acid dehydrogenase (BCKDH) complex. This complex consists of multiple copies of BCKDH Ela / p tetramer, BCKDH E2, and BCKDH E3 subunits. This complex is regulated by the inhibitory phosphorylation mediated by a BCKDH kinase (BCKDK), and the same phosphorylation site is dephosphorylated by a phosphatase PPM1K. Inhibiting complex phosphorylation promotes BCKDH activity, thereby promoting irreversible catabolism of BCKAs (Lynch CJ, Adams SH: Branched-chain amino acids in metabolic signalling and insulin resistance. Nat Rev Endocrinol 2014, 10:723-36.). The absence of BCKDK in mice confirms this regulation, as BCKDK-deficient mice exhibit enhanced BCKDH activity in various tissues (Joshi MA, Jeoung NH, Obayashi M, Hattab EM, Brocken EG, Liechty EA, Kubek MJ, Vattem KM, Wek RC, Harris RA: Impaired growth and neurological abnormalities in branched-chain alpha-keto acid dehydrogenase kinase-deficient mice. Biochem J 2006, 400:153-62.).Insufficient catabolism of branched-chain amino acids (BCAAs) is associated with insulin resistance, diabetes, congenital heart defect, and heart failure. Circulating branched-chain amino acids and their breakdown products, branched-chain α-keto acids (BCKAs), are also indicators of the onset of diabetes. The α-keto acid dehydrogenase (BCKDH) complex is a rate-limiting step in the breakdown and clearance of branched-chain amino acids; while BCKDK phosphorylates E1α subunits of BCKDH, thereby inhibiting its function.In recent years, although there have been some early studies related to BCKDK, such as WO2020056155A, WO2020261144A, WO2020261205A, WO2022175959A, and WO2023100061A, there remains a need to develop novel and effective BCKDK inhibitors for the treatment of diseases or disorders associated with elevated BCAA concentrations. SUMMARYThe present disclosure provides a compound represented by formula (I) or a pharmaceutically acceptable salt thereof,wherein R1, R2, and R3 are each independently selected from the group consisting of hydrogen, fluorine, chlorine, and methyl;R4 is selected from the group consisting of fluorine and chlorine;R5 is selected from the group consisting of C1-4 alkyl, C1-4 haloalkyl, 3- to 4-membered cycloalkyl, and -L1-R9, wherein L1 is selected from the group consisting of sulfur and oxygen, and R9 is selected from the group consisting of C1-4 alkyl, C1-4 haloalkyl, deuterated C1-4 alkyl, and 3- to 4-membered cycloalkyl; X1 is selected from the group consisting of nitrogen and CR6, wherein R6 is selected from the group consisting of hydrogen, fluorine, and chlorine;R7 and R8 are each independently selected from the group consisting of hydrogen, fluorine, and chlorine.In some embodiments, in the compound represented by formula (I) or the pharmaceutically acceptable salt thereof provided in the present disclosure, X1 is nitrogen.In some embodiments, the compound represented by formula (I) or the pharmaceutically acceptable salt thereof provided in the present disclosure is a compound represented by formula (II) or a pharmaceutically acceptable salt thereof,wherein R1, R2, R3, R4, R5, R6, R7, and R8 are each as defined in formula (I).In some embodiments, in the compound represented by formula (I) or (II) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of C1-4 alkyl and C1-4 haloalkyl.In an optional embodiment, in the compound represented by formula (I) or (II) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of methyl, ethyl, and isopropyl.In some embodiments, in the compound represented by formula (I) or (II) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of methyl and ethyl.In some embodiments, in the compound represented by formula (I) or (II) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of 3- to 4-membered cycloalkyl.In an optional embodiment, in the compound represented by formula (I) or (II) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is cyclopropyl.In some embodiments, the compound represented by formula (I) or (II) or the pharmaceutically acceptable salt thereof provided in the present disclosure is a compound represented by formula (II-1) or a pharmaceutically acceptable salt thereof,wherein R1, R2, R3, R4, R6, R7, R8, and R9 are each as defined in formula (I).In some embodiments, in the compound represented by formula (I), (II), or (II-1) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R9 is selected from the group consisting of C1-4 alkyl, C1-4 haloalkyl, and deuterated C1-4 alkyl.In an optional embodiment, in the compound represented by formula (I), (II), or (II-1) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl.In an optional embodiment, in the compound represented by formula (I), (II), or (II-1) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R9 is selected from the group consisting of methyl, difluoromethyl, trifluoromethyl, and deuterated methyl.In some embodiments, the compound represented by formula (I) or (II) or the pharmaceutically acceptable salt thereof provided in the present disclosure is a compound represented by formula (II-2) or a pharmaceutically acceptable salt thereof,wherein R1, R2, R3, R4, R6, R7, R8, and R9 are each as defined in formula (I).In some embodiments, in the compound represented by formula (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R9 is selected from the group consisting of C1-4 alkyl, C1-4 haloalkyl, and deuterated C1-4 alkyl.In an optional embodiment, in the compound represented by formula (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl.In an optional embodiment, in the compound represented by formula (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R9 is methyl.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, at least one of R1, R2, and R3 is selected from the group consisting of fluorine and chlorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R1 and R3 are each independently selected from the group consisting of hydrogen, fluorine, and chlorine.In an optional embodiment, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, one of R1 and R3 is hydrogen, and the other is selected from the group consisting of hydrogen, fluorine, and chlorine.In an optional embodiment, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R1 and R3 are each independently hydrogen.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R2 is fluorine or chlorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R2 is fluorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R2 is chlorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R1 and R3 are each independently hydrogen; R2 is fluorine or chlorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R4 is selected from the group consisting of fluorine and chlorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R4 is fluorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R1 and R3 are each independently hydrogen, R2 is fluorine or chlorine, and R4 is fluorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, at least one of R6, R7, and R8 is selected from the group consisting of fluorine and chlorine.In an optional embodiment, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, at least two of R6, R7, and R8 are selected from the group consisting of fluorine and chlorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R6 is selected from the group consisting of fluorine and chlorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, at least one of R6, R7, and R8 is chlorine.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R7 is selected from the group consisting of fluorine and chlorine, and R8 is hydrogen.In an optional embodiment, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R7 is fluorine, and R8 is hydrogen.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R8 is selected from the group consisting of fluorine and chlorine, and R7 is hydrogen.In some embodiments, in the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R1 and R3 are each independently hydrogen; R2 is fluorine or chlorine; R4 is fluorine; R6 is selected from the group consisting of fluorine and chlorine; R7 is selected from the group consisting of fluorine and chlorine, and R8 is hydrogen; or R8 is selected from the group consisting of fluorine and chlorine, and R7 is hydrogen; and at least one of R6, R7, and R8 is chlorine.In some embodiments, the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure is a compound represented by formula (II-A) or a pharmaceutically acceptable salt thereof,wherein R2, R5, R6, and R7 are each as defined in formulas (I), (II), (II-1), and (II-2).In an optional embodiment, in the compound represented by formula (II-A) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R2 is chlorine.In an optional embodiment, in the compound represented by formula (II-A) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R2 is fluorine.In some embodiments, the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure is a compound represented by formula (II-B) or a pharmaceutically acceptable salt thereof,wherein R2, R5, R6, and R7 are each as defined in formulas (I), (II), (II-1), and (II-2).In an optional embodiment, in the compound represented by formula (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R2 is chlorine.In an optional embodiment, in the compound represented by formula (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R2 is fluorine.In some embodiments, in the compound represented by formula (II-A) or (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of methyl and ethyl.In some embodiments, in the compound represented by formula (II-A) or (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is oxygen, and R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl.In some embodiments, in the compound represented by formula (II-A) or (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is oxygen, and R9 is selected from the group consisting of methyl and deuterated methyl.In some embodiments, in the compound represented by formula (II-A) or (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is oxygen, and R9 is selected from the group consisting of difluoromethyl and trifluoromethyl.In some embodiments, in the compound represented by formula (II-A) or (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is sulfur, and R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl.In some embodiments, in the compound represented by formula (II-A) or (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is sulfur, and R9 is methyl or deuterated methyl.In some embodiments, in the compound represented by formula (II-A) or (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R6 is fluorine or chlorine, and R7 is hydrogen.In an optional embodiment, in the compound represented by formula (II-A) or (II-B) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R6 is fluorine, and R7 is hydrogen.In some embodiments, the compound represented by formula (I), (II), (II-1), or (II-2) or the pharmaceutically acceptable salt thereof provided in the present disclosure is a compound represented by formula (II-C) or a pharmaceutically acceptable salt thereof,, wherein R2, R5, R6, and R7 are each as defined in formulas (I), (II), (II-1), and (II-2).In an optional embodiment, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R2 is chlorine.In an optional embodiment, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R2 is fluorine.In some embodiments, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of methyl and ethyl.In some embodiments, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is oxygen, and R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl.In some embodiments, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is oxygen, and R9 is selected from the group consisting of methyl and deuterated methyl.In some embodiments, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is oxygen, and R9 is selected from the group consisting of difluoromethyl and trifluoromethyl.In some embodiments, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is sulfur, and R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl.In some embodiments, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R5 is selected from the group consisting of -L1-R9, wherein L1 is sulfur, and R9 is methyl or deuterated methyl.In some embodiments, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, at least one of R6 and R7 is selected from the group consisting of fluorine and chlorine.In an optional embodiment, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R6 is selected from the group consisting of fluorine and chlorine.In an optional embodiment, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R6 is selected from the group consisting of fluorine, and R7 is selected from the group consisting of fluorine and chlorine.In an optional embodiment, in the compound represented by formula (II-C) or the pharmaceutically acceptable salt thereof provided in the present disclosure, R6 is selected from the group consisting of chlorine, and R7 is selected from the group consisting of fluorine and chlorine.The present disclosure further provides a compound or a pharmaceutically acceptable salt thereof selected from the group consisting of the following:, , , , .The present disclosure further provides an isotopically substituted form of the aforementioned compound or the pharmaceutically acceptable salt thereof; optionally, the isotopically substituted form is a deuterated form.The present disclosure provides a pharmaceutical composition comprising the compound described above, and one or more pharmaceutically acceptable excipients.In some embodiments, a unit dose of the pharmaceutical composition is 0.001 mg-1000 mg.In certain embodiments, the pharmaceutical composition comprises 0.01%-99.99% of the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof based on the total weight of the composition. In certain embodiments, the pharmaceutical composition comprises 0.1%-99.9% of the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof. In certain embodiments, the pharmaceutical composition comprises 0.5%-99.5% of the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof. In certain embodiments, the pharmaceutical composition comprises 1%-99% of the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof. In certain embodiments, the pharmaceutical composition comprises 2%-98% of the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof.In certain embodiments, the pharmaceutical composition comprises 0.01%-99.99% of the pharmaceutically acceptable excipient based on the total weight of the composition. In certain embodiments, the pharmaceutical composition comprises 0.1%-99.9% of the pharmaceutically acceptable excipient. In certain embodiments, the pharmaceutical composition comprises 0.5%-99.5% of the pharmaceutically acceptable excipient. In certain embodiments, the pharmaceutical composition comprises 1%-99% of the pharmaceutically acceptable excipient. In certain embodiments, the pharmaceutical composition comprises 2%-98% of the pharmaceutically acceptable excipient.The present disclosure further provides use of the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof or the aforementioned pharmaceutical composition as a medicament.The present disclosure further provides use of the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof or the aforementioned pharmaceutical composition in the manufacture of a medicament for preventing and / or treating a BCKDK-related disease.The present disclosure further provides use of the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof or the aforementioned pharmaceutical composition in the manufacture of a medicament for preventing and / or treating a disease, wherein the disease includes, but is not limited to, a glucose metabolism disorder or an abnormal blood glucose disease, a heart disease, or a kidney disease.The present disclosure further provides a method for preventing and / or treating a BCKDK-related disease, comprising administering to a patient the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof or the aforementioned pharmaceutical composition.The present disclosure further provides a method for preventing and / or treating a disease, comprising administering to a patient the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof or the aforementioned pharmaceutical composition, wherein the disease includes, but is not limited to, a glucose metabolism disorder or an abnormal blood glucose disease, a heart disease, or a kidney disease.In an optional embodiment, in the method provided in the present disclosure, the aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof or the aforementioned pharmaceutical composition is administered to a patient in a therapeutically effective amount.In some embodiments, the BCKDK-related disease includes, but is not limited to, a glucose metabolism disorder or an abnormal blood glucose disease, a heart disease, or a kidney disease.In some embodiments, the glucose metabolism disorder or the abnormal blood glucose disease is diabetes.In some embodiments, the heart disease is heart failure.The aforementioned compound or the pharmaceutically acceptable salt thereof or the isotopically substituted form thereof or the aforementioned pharmaceutical composition provided in the present disclosure exhibits a good inhibition / degradation effect on BCKDK.In another aspect, the present disclosure provides a preparation method for a compound represented by formula (I) or a pharmaceutically acceptable salt thereof, comprising a step of removing a protecting group PG from a compound represented by formula (I-a) or a pharmaceutically acceptable salt thereof under an alkaline condition, and further comprising a step of adjusting the pH to acidity,wherein PG is a carboxyl protecting group; R1, R2, R3, R4, R5, X1, R7, and R8 are each as defined in formula (I).In an optional embodiment, in the preparation method for the compound represented by formula (I) or the pharmaceutically acceptable salt thereof provided in the present disclosure, PG is C1-6 alkyl.In another aspect, the present disclosure provides a compound represented by formula (I-a) or a pharmaceutically acceptable salt thereof,wherein PG is a carboxyl protecting group; R1, R2, R3, R4, R5, X1, R7, and R8 are each as defined in formula (I).In an optional embodiment, in the compound represented by formula (I-a) or the pharmaceutically acceptable salt thereof, PG is C1-6 alkyl.Definitions of TermsWhere the present disclosure does not define a particular configuration, the compounds of the present disclosure may exist in particular geometric or stereoisomeric forms. The present disclosure contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers, (D)-isomer, (L)-isomer, atropisomers (i.e., stereoisomers with hindered rotation), and racemic mixtures and other mixtures thereof, such as enantiomerically or diastereomerically enriched mixtures, all of which fall within the scope of the present disclosure. Additional asymmetric carbon atoms may be present in substituents such as an alkyl group. All such isomers and mixtures thereof are included within the scope of the present disclosure.In addition, the compounds and intermediates of the present disclosure may also exist in different tautomeric forms, and all such forms are included within the scope of the present disclosure. The term “tautomer” or “tautomeric form” refers to structural isomers of different energies that can interconvert via a low-energy barrier.The compounds of the present disclosure may be asymmetric, for example, having one or more stereoisomers. Unless otherwise specified, all stereoisomers are included, for example, enantiomers and diastereomers. The compounds of the present disclosure containing asymmetric carbon atoms may be separated in an optically active pure form or in a racemic form. The optically active pure form may be isolated from a racemic mixture or synthesized using chiral starting materials or chiral reagents.Optically active (R)- and (S)-isomers, and D- and L-isomers may be prepared by chiral synthesis, chiral reagents, or other conventional techniques. If one enantiomer of a certain compound of the present disclosure is desired, it may be prepared by asymmetric synthesis or derivatization with a chiral auxiliary, wherein the resulting mixture of diastereomers is separated, and the auxiliary group is cleaved to provide the pure desired enantiomer. Alternatively, when the molecule contains a basic functional group (e.g., amino) or an acidic functional group (e.g., carboxyl), salts of diastereomers are formed with an appropriate optically active acid or base, diastereomeric resolution is then performed by conventional methods well-known in the art, and pure enantiomers are then recovered. In addition, the separation of enantiomers and diastereomers is generally accomplished by chromatography using a chiral stationary phase, optionally in combination with chemical derivatization (e.g., formation of a carbamate from an amine).In the chemical structures of the compounds of the present disclosure, the bond “” indicates an unspecified configuration; that is, if chiral isomers exist in the chemical structures, the bond “” may be “” or “”, or includes both the configurations “” and “” simultaneously. In the chemical structures of the compounds of the present disclosure, the bond “” does not specify a configuration; that is, the configuration of the bond “” may be an E configuration or a Z configuration, or includes both the E configuration and the Z configuration simultaneously.The present disclosure also includes some isotopically labeled compounds of the present disclosure that are identical to those recited herein but have one or more atoms replaced with an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into the compounds of the present disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine, and chlorine, such as 2H, 3H, 11C, 13C, 14C, 13N, 15N, 15O, 17O, 18O, 31P, 32P, 35S, 18F, 123I, 125I, and 36Cl.Unless otherwise specified, when a position is specifically designated as deuterium (D), the position shall be understood to be deuterium having an abundance that is at least 1000 times greater than the natural abundance of deuterium (which is 0.015%) (i.e., at least 10% deuterium incorporation). The compounds in examples comprise deuterium having an abundance that is greater than at least 1000 times the natural abundance, at least 2000 times the natural abundance, at least 3000 times the natural abundance, at least 4000 times the natural abundance, at least 5000 times the natural abundance, at least 6000 times the natural abundance, or higher times the natural abundance. The present disclosure also includes various deuterated forms of the compounds. Each available hydrogen atom linked to a carbon atom may be independently replaced with a deuterium atom. Those skilled in the art can synthesize the deuterated forms of the compounds by referring to the relevant literature. Commercially available deuterated starting materials can be used in preparing the deuterated forms of the compounds, or they can be synthesized using conventional techniques with deuterated reagents, including but not limited to deuterated borane, borane-d3 in tetrahydrofuran, deuterated lithium aluminum hydride, deuterated iodoethane, deuterated iodomethane, and the like.The term “alkyl” refers to a saturated aliphatic hydrocarbon group, preferably an alkyl group containing 1 to 6 carbon atoms (e.g., 1, 2, 3, 4, 5, or 6 carbon atoms). Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, and 2,3-dimethylbutyl.The term “cycloalkyl” or “carbocycle” refers to a saturated or partially unsaturated, monocyclic or polycyclic hydrocarbon substituent, and the cycloalkyl ring contains 3 to 20 carbon atoms, preferably 3 to 6 carbon atoms. Non-limiting examples of monocyclic cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, etc. Polycyclic cycloalkyl groups include spiro-ring, fused-ring, and bridged-ring cycloalkyl groups.The term “alkoxy” refers to -O-(alkyl) and -O-(unsubstituted cycloalkyl), wherein the alkyl and cycloalkyl are as defined above. Non-limiting examples of alkoxy groups include methoxy, ethoxy, propoxy, butoxy, cyclopropyloxy, cyclobutoxy, cyclopentyloxy, and cyclohexyloxy.The term “hydroxy” refers to -OH.The term “halogen” refers to fluorine, chlorine, bromine, or iodine.The term “haloalkyl” refers to an alkyl group substituted with halogen, wherein the alkyl group is as defined above.The term “cyano” refers to -CN.The term “amino” refers to -NH2.“Optionally” or “optional” means that the event or circumstance subsequently described may, but does not necessarily, occur; this description includes an instance where the event or circumstance occurs or does not occur. For example, “C1-6 alkyl optionally substituted with halogen or cyano” means that the halogen or cyano may, but does not necessarily, exist; this description includes an instance where the alkyl is substituted with halogen or cyano and an instance where the alkyl is not substituted with halogen or cyano.The term “substituted” means that one or more, preferably up to 5, and more preferably 1 to 3, hydrogen atoms in the group are independently substituted with a corresponding number of substituents. It goes without saying that a substituent is only in its possible chemical position, and those skilled in the art can determine (experimentally or theoretically) possible or impossible substitution without undue effort.“Substituted with one or more...” refers to substitution with a single substituent or multiple substituents. In the case of substitution with multiple substituents, there may be a plurality of identical substituents, or one or a group of a plurality of different substituents.In the present disclosure, the terms “comprise” and “include” can be replaced with “consist of”.The term “composition” refers to a mixture of a drug containing one or more of the compounds described herein or physiologically and pharmaceutically acceptable salts or precursors thereof, and other chemical components, as well as other components such as physiologically and pharmaceutically acceptable carriers and excipients. The composition is intended to promote administration to an organism and facilitate the absorption of the active ingredient, thereby exerting biological activity.The term “pharmaceutically acceptable excipient” includes, but is not limited to, any auxiliary, carrier, excipient, glidant, sweetener, diluent, preservative, dye / colorant, flavoring agent, surfactant, wetting agent, dispersant, suspending agent, stabilizer, isotonic agent, solvent, or emulsifier that has been approved by the U.S. Food and Drug Administration as acceptable for use in humans or livestock animals.Unless otherwise specified, the “compound” of the present disclosure can be present in the form of a salt or mixed salt or in a non-salt form (e.g., a free acid or free base). When present in the form of a salt or mixed salt, it may be a pharmaceutically acceptable salt.The term “pharmaceutically acceptable salt” includes pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts.As used herein, the term “inhibit” is used interchangeably with “decrease”, “silence”, “down-regulate”, “repress”, and other similar terms, and includes any level of inhibition. Inhibition can be assessed in terms of a decrease in the absolute or relative level of one or more of these variables relative to a control level. The control level may be any type of control level used in the art, such as a pre-dose baseline level or a level determined from a similar untreated or control (e.g., buffer-only control or inert agent control) treated subject, cell, or sample.“Effective amount”, “effective dose”, “effective therapeutic amount”, or “therapeutically effective amount” refers to the amount of a drug, a compound, or a pharmaceutical composition necessary to obtain any one or more beneficial or desired therapeutic results. For prophylactic use, the beneficial or desired results include elimination or reduction of risk, reduction of severity, or delay of the onset of a disorder, including the biochemistry, histology, and / or behavioral symptoms of the disorder, complications thereof, and intermediate pathological phenotypes that appear during the progression of the disorder.As used herein, “object”, “patient”, “subject”, and “individual” are used interchangeably and include human or non-human animals, e.g., mammals, e.g., humans or monkeys. DETAILED DESCRIPTIONThe present disclosure is further described below with reference to examples; however, these examples are not intended to limit the scope of the present disclosure. Experimental methods without specific conditions indicated in the examples of the present disclosure were generally conducted under conventional conditions, or conditions recommended by the manufacturers of the starting materials or commercial products. Where the specific source of a reagent is not indicated, the reagent may be obtained from any supplier of molecular biology reagents at a quality / purity level for molecular biology applications.Unless otherwise specified, all reagents used in the following examples are commercially available.Analysis conditions used for all mass spectrometry data in the following examples: column model: ACQUITY UPLC BEH C18 1.7 μM, 2.1 × 50 mm Column; mobile phase A: 0.05% ammonia water + 0.01% formic acid in water; mobile phase B: 0.01% formic acid in acetonitrile; the gradient of mobile phase B in 0-1.7 min was 5%-95%; the gradient of mobile phase B was maintained at 95% in 1.7-2.5 min.In the following examples, the instrument used for all proton nuclear magnetic resonance spectroscopic data was a Bruker 400 MHz nuclear magnetic resonance spectrometer; the instrument used for all fluorine nuclear magnetic resonance spectroscopic data was a Bruker 376 MHz nuclear magnetic resonance spectrometer. Example 16-Chloro-3-(2,4,5-trifluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (1)Step 1: synthesis of methyl 3-amino-6-chlorothieno[3,2-b]pyridine-2-carboxylate 1cAt 0 °C, methyl 2-mercaptoacetate 1b (2.4 mL, 27.24 mmol) was slowly added dropwise to a solution of 5-chloro-3-nitropyridin-2-carbonitrile 1a (5.00 g, 27.24 mmol) in N,N-dimethylformamide (60 mL), and then a solution of potassium hydroxide (3.06 g, 54.50 mmol) in water (10.0 mL) was added dropwise. The reaction mixture was stirred at 0-5 °C for 1 h. Ice water (60 mL) was added, followed by extraction with ethyl acetate (2 × 100 mL). The organic phases were combined, washed with saturated brine (4 × 80 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product of 1c (6.5 g, yield: 98%).MS(ESI) m / z = 243.2 [M+H]+.Step 2: synthesis of methyl 3-bromo-6-chlorothieno[3,2-b]pyridine-2-carboxylate 1dAt 25 °C, in a nitrogen atmosphere, tert-butyl nitrite (3.92 g, 1.3 mL, 10.7 mmol) was added dropwise to a three-necked flask containing copper (II) bromide (2.05 g, 9.13 mmol) and acetonitrile (15.0 mL). A suspension of compound 1c (2.0 g, 8.3 mmol) in acetonitrile (5.0 mL) was then added at 20 °C. The reaction mixture was stirred at 25 °C for 2 h, then slowly poured into a hydrochloric acid solution (2 N, 20 mL), and extracted with ethyl acetate (2 × 70 mL). The organic phases were combined, washed with saturated brine (2 × 100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 1d (1.0 g, yield: 39%).MS(ESI) m / z = 306.1 [M+H]+.Step 3: synthesis of methyl 6-chloro-3-(2,4,5-trifluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylate 1fAt room temperature, compound 1d (75 mg, 0.25 mmol) and compound 1e (50 mg, 0.25 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.4 mL), and 1,1'-di-tert-butylphosphinoferrocene palladium(II) dichloride (15.9 mg, 0.024 mmol) and potassium carbonate (101 mg, 0.73 mmol) were added. The reaction mixture was stirred at 100 °C for 1 h in a nitrogen atmosphere. After the mixture was cooled to room temperature, ethyl acetate (60 mL) was added. The mixture was washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 1f (60 mg, yield: 61%).MS(ESI) m / z = 388.2 [M+H]+.Step 4: synthesis of 6-chloro-3-(2,4,5-trifluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid 1At room temperature, compound 1f (60 mg, 0.16 mmol) was dissolved in a mixed solvent of tetrahydrofuran (1 mL), methanol (0.5 mL), and water (0.5 mL), and lithium hydroxide (37 mg, 1.6 mmol) was added. The reaction mixture was stirred at room temperature for 1 h, and water (30 mL) was added. The pH was adjusted to 4-5 with hydrochloric acid (1 N), followed by extraction with ethyl acetate (2 × 10 mL). The organic phases were combined, washed with saturated brine (2 × 10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was separated and purified by preparative high performance liquid chromatography (column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 40%-95%, 18 min; flow rate: 30 mL / min; peak time: 11.06 min) to give the title compound 1 (15 mg, yield: 25%).MS(ESI) m / z = 374.2 [M+H]+.1H-NMR (400 MHz, DMSO-d6) δ 8.87 (s, 1H), 8.78 (s, 1H),7.44-7.37 (m, 1H), 4.02 (s, 3H).19F-NMR (376 MHz, DMSO-d6) δ -132.28(dd, 1F), -142.60 (dd, 1F), -151.65 (dd, 1F). Example 26-Chloro-3-(6-chloro-2,4-difluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (2); 6-Chloro-3-(6-chloro-2,4-difluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (2-P1); 6-Chloro-3-(6-chloro-2,4-difluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (2-P2) Step 1: synthesis of methyl 6-chloro-3-(6-chloro-2,4-difluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylate 2bAt room temperature, compound 1d (60 mg, 0.20 mmol) and compound 2a (89 mg, 0.30 mmol) were dissolved in toluene (3 mL) and water (0.3 mL), and tris(dibenzylideneacetone)dipalladium(0) (18 mg, 0.02 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (16 mg, 0.04 mmol), and potassium phosphate (125 mg, 0.59 mmol) were added. The reaction mixture was stirred at 70 °C for 2 h in a nitrogen atmosphere. After the mixture was cooled to room temperature, ethyl acetate (40 mL) was added. The mixture was washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 2b (50 mg, yield: 61%).MS(ESI) m / z = 404.2 [M+H]+.Step 2: synthesis of 6-chloro-3-(6-chloro-2,4-difluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid 2At room temperature, compound 2b (50 mg, 0.12 mmol) was dissolved in a mixed solvent of tetrahydrofuran (2 mL), methanol (1 mL), and water (1 mL), and lithium hydroxide (30 mg, 1.2 mmol) was added. The reaction mixture was stirred at room temperature for 1 h. Water (30 mL) was added. The pH was adjusted to 4-5 with hydrochloric acid (1 N), followed by extraction with ethyl acetate (2 × 10 mL). The organic phases were combined, washed with saturated brine (2 × 10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was separated and purified by preparative high performance liquid chromatography (column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 40%-95%, 18 min; flow rate: 30 mL / min; peak time: 11.06 min) to give the title compound 2 (18 mg, yield: 38%).MS(ESI) m / z = 390.1 [M+H]+.1H-NMR (400 MHz, DMSO-d6) δ 8.88 (s, 1H), 8.76 (s, 1H),7.63-7.60 (m, 1H), 3.96 (s, 3H).19F-NMR (376 MHz, DMSO-d6) δ -124.92 (d, 1F), -126.84(d, 1F).Step 3: 6-chloro-3-(6-chloro-2,4-difluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (2-P1) and 6-chloro-3-(6-chloro-2,4-difluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (2-P2)The purified product 2 of step 2 was further resolved by SFC. Resolution method: column: ChiralPak AD, 250 × 30 mm I.D., 5 μm; mobile phase: A: CO2; B: isopropanol [0.1% NH3 (7 M in MeOH)]; gradient: B 30%; flow rate: 100 mL / min; back pressure: 100 bar; column temperature: 35 °C; wavelength: 214 nm; cycle time: about 2 min. Analysis method: column: ChiralPak AD, 100 × 4.6 mm I.D., 3 μm; mobile phase: A: CO2; B: isopropanol (0.1% DEA); gradient: B 5%-45%, 4 min; flow rate: 3.0 mL / min; back pressure: 2000 psi; column temperature: 40 °C; wavelength: 214 nm; cycle time: about 2 min.2-P1, retention time t = 2.985 min, 1.7 mg;2-P2, retention time t = 2.807 min, 2.1 mg. Example 36-Chloro-3-(2,4,5-trifluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (3)Step 1: synthesis of methyl 6-chloro-3-(2,4,5-trifluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylate 3bAt room temperature, compound 1d (100 mg, 0.33 mmol) and compound 3a (133 mg, 0.49 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.40 mL), and 1,1'-di-tert-butylphosphinoferrocene palladium(II) dichloride (21.3 mg, 0.03 mmol) and potassium carbonate (135 mg, 0.09 mmol) were added. The reaction mixture was stirred at 70 °C for 1 h in a nitrogen atmosphere. After the mixture was cooled to room temperature, ethyl acetate (40 mL) was added. The mixture was washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 3b (80 mg, yield: 65%).MS(ESI) m / z = 372.2 [M+H]+.Step 2: synthesis of 6-chloro-3-(2,4,5-trifluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylic acid 3At room temperature, compound 3b (80 mg, 0.22 mmol) was dissolved in a mixed solvent of tetrahydrofuran (2 mL), methanol (1 mL), and water (1 mL), and lithium hydroxide (52 mg, 2.2 mmol) was added. The reaction mixture was stirred at room temperature for 1 h. Water (30 mL) was added. The pH was adjusted to 4-5 with hydrochloric acid (1 N), followed by extraction with ethyl acetate (2 × 10 mL). The organic phases were combined, washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was separated and purified by preparative high performance liquid chromatography (column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 35%-85%, 18 min; flow rate: 30 mL / min; peak time: 13.50 min) to give the title compound 3 (18 mg, yield: 22%).MS(ESI) m / z = 358.2 [M+H]+.1H-NMR (400 MHz, DMSO-d6) δ 8.86 (s, 1H), 8.77 (s, 1H), 7.54-7.48 (m, 1H),2.26 (s, 3H). 19F-NMR (376 MHz, DMSO-d6) δ -118.78(dd, 1F), -137.93(dd, 1F), -144.56 (dd, 1F). Example 46-Fluoro-3-[2,4,5-trifluoro-3-(methylthio)phenyl]thieno[3,2-b]pyridine-2-carboxylic acid (4)Step 1: synthesis of methyl 3-amino-6-fluorothieno[3,2-b]pyridine-2-carboxylate 4bAt 0 °C, methyl mercaptoacetate (1.1 mL, 12.57 mmol) was added dropwise to a solution of compound 4a (2 g, 11.97 mmol) in N,N-dimethylformamide (20 mL), and then an aqueous solution of potassium hydroxide (5 M, 4.8 mL, 23.94 mmol) was slowly added dropwise. The mixture was left to react at 0 °C for 30 min. The reaction mixture was poured into ice water (60 mL) and extracted with ethyl acetate (3 × 50 mL). The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure, and the resulting crude product was purified by column chromatography (petroleum ether / ethyl acetate) to give compound 4b (2 g, yield: 73%).MS(ESI) m / z = 227.2 [M+H]+.Step 2: synthesis of methyl 6-fluoro-3-iodothieno[3,2-b]pyridine-2-carboxylate 4cAt room temperature, compound 4b (1 g, 4.42 mmol) and diiodomethane (0.54 mL, 6.63 mmol) were dissolved in acetonitrile (20 mL). The mixture was heated to 70 °C, and then isoamyl nitrite (0.89 mL, 6.63 mmol) was slowly added dropwise to the system, with the temperature maintained at 70-80 °C. After the addition, the mixture was left to react at 70 °C for 2 h. The reaction mixture was directly concentrated under reduced pressure to dryness, and the residue was purified by column chromatography (petroleum ether / ethyl acetate) to give a crude product of 4c (350 mg, yield: 23%).MS(ESI) m / z = 338.2 [M+H]+.Step 3: synthesis of methyl 6-fluoro-3-(2,4,5-trifluoro-3-hydroxyphenyl)thieno[3,2-b]pyridine-2-carboxylate 4eAt room temperature, compound 4c (300 mg, 0.89 mmol) and compound 4d (366 mg, 1.34 mmol) were dissolved in 1,4-dioxane (2 mL) and water (0.40 mL), and 1,1'-di-tert-butylphosphinoferrocene palladium(II) dichloride (58 mg, 0.09 mmol) and potassium carbonate (369 mg, 2.67 mmol) were added. The reaction mixture was stirred at 100 °C for 1 h in a nitrogen atmosphere. After the mixture was cooled to room temperature, ethyl acetate (20 mL) was added. The mixture was washed with saturated brine (2 × 10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 4e (100 mg, yield: 31%).MS(ESI) m / z = 358.2 [M+H]+.Step 4: synthesis of 2,5,6-trifluoro-3-[6-fluoro-2-(methoxycarbonyl)thieno[3,2-b]pyridin-3-yl]phenyl trifluoromethanesulfonate 4fAt room temperature, triethylamine (85 mg, 0.84 mmol) and trifluoromethanesulfonic anhydride (118 mg, 0.42 mmol) were added to a solution of compound 4e (100 mg, 0.28 mmol) in dichloromethane (4 mL). After the addition, the mixture was left to react at room temperature for 1 h. The reaction mixture was concentrated under reduced pressure to remove dichloromethane, and ethyl acetate (20 mL) was added to the residue. The mixture was washed with saturated brine (2 × 10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 4f (98 mg, yield: 71%).MS(ESI) m / z = 490.2 [M+H]+.Step 5: synthesis of methyl 6-fluoro-3-[2,4,5-trifluoro-3-(methylthio)phenyl]thieno[3,2-b]pyridine-2-carboxylate 4gAt room temperature, compound 4f (80 mg, 0.16 mmol), sodium thiomethoxide (57 mg, 0.82 mmol), palladium acetate (7.3 mg, 0.033 mmol), 2,2'-bis(diphenylphosphino)-1,1'-binaphthyl (20.4 mg, 0.033 mmol), and toluene (2 mL) were added to a round-bottom flask. In a nitrogen atmosphere, the mixture was heated to 120 °C and left to react overnight. After the reaction mixture was cooled to room temperature, ethyl acetate (20 mL) was added. The mixture was washed with saturated brine (2 × 10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 4g (50 mg, yield: 79%).MS(ESI) m / z = 388.2 [M+H]+.Step 6: synthesis of 6-fluoro-3-[2,4,5-trifluoro-3-(methylthio)phenyl]thieno[3,2-b]pyridine-2-carboxylic acid 4At room temperature, compound 4g (50 mg, 0.13 mmol) was dissolved in a mixed solvent of tetrahydrofuran (2 mL), methanol (1 mL), and water (1 mL), and lithium hydroxide (31 mg, 1.3 mmol) was added. The reaction mixture was stirred at room temperature for 1 h. Water (30 mL) was added. The pH was adjusted to 4-5 with hydrochloric acid (1 N, 4 mL), followed by extraction with ethyl acetate (2 × 20 mL). The organic phases were combined, washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was separated and purified by preparative high performance liquid chromatography (column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 40%-95%, 18 min; flow rate: 30 mL / min; peak time: 10.99 min) to give the title compound 4 (18 mg, yield: 37%).MS(ESI) m / z = 374.2 [M+H]+.1H-NMR (400 MHz, DMSO-d6) δ 8.80 (s, 1H), 8.65 (d, 1H), 2.52 (s, 3H).19F-NMR (376 MHz, DMSO-d6) δ -110.02 (d, 1F), -127.38(s, 1F), -129.29 (d, 1F), -142.40 (dd, 1F). Example 56-Fluoro-3-(2,4,5-trifluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (5)6-Fluoro-3-(2,4,5-trifluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (5) was prepared by using the synthesis method in Example 1 and replacing compound 1d with compound 4b.Preparative high performance liquid chromatography separation and purification conditions for compound 5: column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 40%-90%, 18 min; flow rate: 30 mL / min; peak time: 10.5 min.MS(ESI) m / z = 358.30 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.80 (d,1H), 8.64 (dd,1H), 7.44-7.38 (m,1H), 4.01(s,3H).19F-NMR(376 MHz, DMSO-d6) δ -127.42 (s, 1F), -132.32 (dd, 1F), -142.65 (dd, 1F), -151.72 (dd, 1F). Example 63-(3-Ethyl-2,4,5-trifluorophenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (6)Step 1: synthesis of methyl 6-fluoro-3-(2,4,5-trifluoro-3-vinylphenyl)thieno[3,2-b]pyridine-2-carboxylate 6gAt room temperature, compound 4f (160 mg, 0.32 mmol), potassium vinyltrifluoroborate (127 mg, 0.95 mmol), tris(dibenzylideneacetone)dipalladium(0) (28.9 mg, 0.032 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (13 mg, 0.032 mmol), and potassium phosphate (168 mg, 0.79 mmol) were dissolved in toluene (5 mL). The mixture was left to react at 100 °C for 1 h in a nitrogen atmosphere. The reaction mixture was poured into water (50 mL) and extracted with ethyl acetate (3 × 30 mL). The organic phases were combined, washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by column chromatography (petroleum ether / ethyl acetate) to give 6g (112 mg, yield: 95%).MS(ESI) m / z = 368.2 [M+H]+.Step 2: synthesis of methyl 3-(3-ethyl-2,4,5-trifluorophenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylate 6hAt room temperature, compound 6g (100 mg, 0.26 mmol) and 10% palladium on carbon (29 mg, 0.27 mmol) were dissolved in methanol (15 mL). The mixture was stirred for 2 h in a hydrogen atmosphere. The reaction mixture was directly filtered, and the filtrate was concentrated under reduced pressure to give a crude product of 6h (95 mg, 98%).MS(ESI) m / z = 370.2 [M+H]+.Step 3: synthesis of 3-(3-ethyl-2,4,5-trifluorophenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid 6At room temperature, compound 6h (100 mg, 0.27 mmol) was dissolved in a mixed solvent of tetrahydrofuran (6 mL), methanol (3 mL), and water (3 mL), and lithium hydroxide (114 mg, 2.71 mmol) was added. The reaction mixture was stirred at room temperature for 1 h. Water (30 mL) was added. The pH was adjusted to 4-5 with hydrochloric acid (1 N, 4 mL), followed by extraction with ethyl acetate (3 × 30 mL). The organic phases were combined, washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was separated and purified by preparative high performance liquid chromatography (column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 40%-95%, 18 min; flow rate: 30 mL / min; peak time: 11.52 min) to give the title compound 6 (26.8 mg, yield: 27%).MS(ESI) m / z = 356.2 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.79 (s, 1H), 8.63 (d, 1H), 7.52 (dd, 1H), 2.73 (q, 2H), 1.19(q, 3H).19F-NMR(376 MHz, DMSO-d6) δ -120.81 (dd, 1F), -127.60 (s, 1F), -140.39 (dd, 1F) -144.30 (dd, 1F). Example 76-Chloro-3-(3-ethyl-2,4,5-trifluorophenyl)thieno[3,2-b]pyridine-2-carboxylic acid (7)Step 1: synthesis of methyl 6-chloro-3-(2,4,5-trifluoro-3-hydroxyphenyl)thieno[3,2-b]pyridine-2-carboxylate 7eAt room temperature, compound 1d (500 mg, 1.64 mmol) and compound 4d (676 mg, 2.47 mmol) were dissolved in 1,4-dioxane (5 mL) and water (0.80 mL), and 1,1'-di-tert-butylphosphinoferrocene palladium(II) dichloride (103 mg, 0.16 mmol) and potassium carbonate (680 mg, 4.92 mmol) were added. The reaction mixture was stirred at 100 °C for 1 h in a nitrogen atmosphere. After the mixture was cooled to room temperature, ethyl acetate (40 mL) was added. The mixture was washed with saturated brine (2 × 15 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 7e (171 mg, yield: 28%).MS(ESI) m / z = 374.1 [M+H]+.Step 2: synthesis of 2,5,6-trifluoro-3-[6-chloro-2-(methoxycarbonyl)thieno[3,2-b]pyridin-3-yl]phenyl trifluoromethanesulfonate 7fAt room temperature, triethylamine (255 mg, 1.38 mmol) and trifluoromethanesulfonic anhydride (177 mg, 0.69 mmol) were added to a solution of compound 7e (171 mg, 0.46 mmol) in dichloromethane (6 mL). After the addition, the mixture was left to react at room temperature for 1 h. The reaction mixture was concentrated under reduced pressure to remove dichloromethane, and ethyl acetate (40.0 mL) was added to the residue. The mixture was washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 7f (200 mg, yield: 86%).MS(ESI) m / z = 506.1 [M+H]+.6-Chloro-3-(3-ethyl-2,4,5-trifluorophenyl)thieno[3,2-b]pyridine-2-carboxylic acid (7) was synthesized from compound 7f in three steps by using the synthesis method in Example 6.Preparative high performance liquid chromatography separation and purification conditions for compound 7: column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 45%-95%, 18 min; flow rate: 30 mL / min; peak time: 12.37 min.MS(ESI) m / z = 372.1 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.86 (s, 1H), 8.78 (s, 1H), 7.52 (q, 1H), 2.73 (q, 2H), 1.19 (t, 3H). 19F-NMR(376 MHz, DMSO-d6) δ -120.80 (dd, 1F), -140.28 (dd, 1F), -144.24 (dd, 1F). Example 83-{6-chloro-3-[(difluoromethyl)oxy]-2,4-difluorophenyl}-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (8)Step 1: synthesis of tert-butyl (4-chloro-2,6-difluorophenoxy)dimethylsilane 8bTriethylamine (9.2 g, 91.2 mmol) was added to a solution of compound 8a (10 g, 60.8 mmol) in dichloromethane (200 mL) in an ice bath. After 10 min of stirring, tert-butyldimethylsilyl trifluoromethanesulfonate (24.1 g, 91.2 mmol) was added dropwise. After the addition, the mixture was stirred at room temperature for 1 h. The reaction mixture was washed with saturated brine (3 × 80 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by column chromatography (petroleum ether / ethyl acetate) to give 8b (15.1 g, yield: 89%) in the form of an oil.Step 2: synthesis of (3-((tert-butyldimethylsilyl)oxy)-6-chloro-2,4-difluorophenyl)boronic acid 8cAt -78 °C, in a nitrogen atmosphere, n-butyllithium (2.5 M, 23.6 mL, 59.1 mmol) was slowly added dropwise to a solution of compound 8b (15 g, 53.8 mmol) in anhydrous tetrahydrofuran (200 mL). After the addition, the mixture was stirred at -78 °C for 1 h, and then a solution of trimethyl borate (7.3 g, 70 mmol) in anhydrous tetrahydrofuran (20 mL) was slowly added dropwise. After the addition, the mixture was stirred at -78 °C for another 1 h. The reaction mixture was slowly added to a saturated ammonium chloride solution (200 mL) stirred in an ice bath, and after 5 min, ethyl acetate (100 mL) was added. After the mixture was mixed well, the organic phase was isolated, washed with saturated brine (3 × 80 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by column chromatography (petroleum ether / ethyl acetate) to give 8c as an oil (11.2 g, yield: 64%).1H-NMR (400 MHz, DMSO-d6) δ 8.71 (s, 2H), 7.28 (d, 1H), 0.98 (s, 9H), 0.18 (s, 6H).Step 3: synthesis of methyl 3-(3-((tert-butyldimethylsilyl)oxy)-6-chloro-2,4-difluorophenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylate 8dAt room temperature, compound 8c (334 mg, 1.03 mmol) and compound 5d (300 mg, 1.03 mmol) were dissolved in toluene (5 mL) and water (1 mL), and tris(dibenzylideneacetone)dipalladium(0) (94.6 mg, 0.10 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (84.9 mg, 0.20 mmol), and potassium phosphate (548 mg, 2.6 mmol) were added. The reaction mixture was stirred at 100 °C for 1 h in a nitrogen atmosphere. After the mixture was cooled to room temperature, ethyl acetate (40 mL) was added. The mixture was washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was purified by column chromatography (petroleum ether / ethyl acetate) to give the title compound 8d (197 mg, yield: 39%).MS(ESI) m / z = 488.4 [M+H]+.Step 4: synthesis of methyl 3-(6-chloro-2,4-difluoro-3-hydroxyphenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylate 8eAt room temperature, compound 8d (180 mg, 0.37 mmol) and tetrabutylammonium fluoride (1 M, 0.6 mL, 0.6 mmol) were dissolved in tetrahydrofuran (10 mL). The mixture was stirred at room temperature for 2 h. The reaction mixture was poured into water (30 mL) and extracted with ethyl acetate (3 × 20 mL). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product of 8e (201 mg, crude product yield: 145%).MS(ESI) m / z = 374.2 [M+H]+.Step 5: synthesis of methyl 3-{6-chloro-3-[(difluoromethyl)oxy]-2,4-difluorophenyl}-6-fluorothieno[3,2-b]pyridine-2-carboxylate 8fAt room temperature, compound 8e (180 mg, 0.48 mmol), sodium difluorochloroacetate (147 mg, 0.96 mmol), and potassium carbonate (80 mg, 0.58 mmol) were dissolved in N,N-dimethylformamide (5 mL). The mixture was left to react at 100 °C for 2 h in a nitrogen atmosphere. The reaction mixture was poured into water (30 mL) and extracted with ethyl acetate (3 × 20 mL). The organic phases were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was then purified by column chromatography (petroleum ether / ethyl acetate) to give compound 8f (100 mg, 49%).MS(ESI) m / z = 424.2 [M+H]+.Step 6: synthesis of 3-{6-chloro-3-[(difluoromethyl)oxy]-2,4-difluorophenyl}-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid 8At room temperature, compound 8f (100 mg, 0.236 mmol) was dissolved in a mixed solvent of tetrahydrofuran (4 mL), methanol (2 mL), and water (2 mL), and lithium hydroxide (100 mg, 2.4 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. Water (30 mL) was added. The pH was adjusted to 4-5 with hydrochloric acid (1 N), followed by extraction with ethyl acetate (2 × 30 mL). The organic phases were combined, washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was separated and purified by preparative high performance liquid chromatography (column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 35%-90%, 18 min; flow rate: 30 mL / min; peak time: 11.37 min) to give the title compound 8 (27.8 mg, yield: 28%).MS(ESI) m / z = 410.1 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.78 (s,1H), 8.67 (d,1H), 7.84 (d, 1H), 7.34 (t,1H).19F-NMR(376 MHz, DMSO-d6) δ -82.76 (t, 2F), -121.98 (dd, 1F), -124.08 (s, 1F), -126.95 (s, 1F). Example 93-{3-[(Difluoromethyl)oxy]-2,4,5-trifluorophenyl}-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (9)3-{3-[(Difluoromethyl)oxy]-2,4,5-trifluorophenyl}-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (9) was prepared by using the synthesis method in Example 8 and replacing compound 8e with compound 4e.Preparative high performance liquid chromatography separation and purification conditions for compound 9: column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 25%-70%, 18 min; flow rate: 30 mL / min; peak time: 11.72 min.MS(ESI) m / z = 392.1 [M-H]-.1H-NMR(400 MHz, DMSO-d6) δ 8.81 (s,1H), 8.65 (d, 1H), 7.76 (dd, 1H), 7.36 (t, 1H).19F-NMR(376 MHz, DMSO-d6) δ -82.48 (s, 2F), -127.30 (3, 1F), -129.43~-129.50 (m, 1F), -141.61 (dd, 1F), -147.72 (d, 1F). Example 106-Chloro-3-{3-[(difluoromethyl)oxy]-2,4,5-trifluorophenyl}thieno[3,2-b]pyridine-2-carboxylic acid (10)6-Chloro-3-{3-[(difluoromethyl)oxy]-2,4,5-trifluorophenyl}thieno[3,2-b]pyridine-2-carboxylic acid (10) was prepared by using the synthesis method in Example 8 and replacing compound 8e with compound 7e.Preparative high performance liquid chromatography separation and purification conditions for compound 10: column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 45%-90%, 18 min; flow rate: 30 mL / min; peak time: 11.28 min.MS(ESI) m / z = 392.1 [M-H]-.1H-NMR(400 MHz, DMSO-d6) δ 8.81 (s,1H), 8.65 (d,1H), 7.76 (dd, 1H), 7.36 (t, 1H).19F-NMR(376 MHz, DMSO-d6) δ -82.48 (s, 2F), -127.30 (3, 1F), -129.43~-129.50 (m, 1F), -141.61 (dd, 1F), -147.72 (d, 1F). Example 116-Chloro-3-{6-chloro-3-[(difluoromethyl)oxy]-2,4-difluorophenyl}thieno[3,2-b]pyridine-2-carboxylic acid (11)6-Chloro-3-{6-chloro-3-[(difluoromethyl)oxy]-2,4-difluorophenyl}thieno[3,2-b]pyridine-2-carboxylic acid (11) was prepared by using the synthesis method in Example 8 and replacing compound 5d with compound 1d.Preparative high performance liquid chromatography separation and purification conditions for compound 11: column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 50%-95%, 18 min; flow rate: 30 mL / min; peak time: 10.18 min.MS(ESI) m / z = 426.1 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.89 (s,1H), 8.77 (s,1H), 7.83(d,1H), 7.33(t,1H).19F-NMR(376 MHz, DMSO-d6) δ -82.75(t, 2F), -121.93(d, 1F), -124.02(s, 1F). Example 126-Chloro-3-(4-chloro-2-fluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (12)6-Chloro-3-(4-chloro-2-fluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (12) was prepared by using the synthesis method in Example 8, replacing compound 5d with compound 1d, and replacing compound 8c with compound 12a.Preparative high performance liquid chromatography separation and purification conditions for compound 12: column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 50%-90%, 18 min; flow rate: 30 mL / min; peak time: 10.65 min.MS(ESI) m / z = 372.2 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.86 (s, 1H), 8.76 (s, 1H), 7.43 (d, 1H), 7.24 (t, 1H), 3.90 (s, 3H).19F-NMR(376 MHz, DMSO-d6) δ -127.61 (s, 1F). Example 136-Chloro-3-(4-chloro-2-fluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (13)6-Chloro-3-(4-chloro-2-fluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (13) was synthesized by using the synthesis method in Example 8.Preparative high performance liquid chromatography separation and purification conditions for compound 13: column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 50%-95%, 18 min; flow rate: 30 mL / min; peak time: 11.22 min.MS(ESI) m / z = 356.1 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.85(s,1H), 8.74(s,1H), 7.40(d, 1H), 7.33(t,1H), 2.31(s,3H).19F-NMR(376 MHz, DMSO-d6) δ -112.58(s, 1F). Example 146-Chloro-3-{6-chloro-3-[(difluoromethyl)oxy]-2,4-difluorophenyl}thieno[3,2-b]pyridine-2-carboxylic acid (11-P1); 6-Chloro-3-{6-chloro-3-[(difluoromethyl)oxy]-2,4-difluorophenyl}thieno[3,2-b]pyridine-2-carboxylic acid (11-P2)Compound 11 (60 mg) was resolved by SFC. Resolution method: column: ChiralPak IC, 250 × 30 mm I.D., 5 μm; mobile phase: A: CO2; B: isopropanol [0.1% NH3 (7 M in MeOH)]; gradient: B 20%; flow rate: 100 mL / min; back pressure: 100 bar; column temperature: 35 °C; wavelength: 214 nm; cycle time: about 2.5 min.11-P1, retention time t = 2.862 min, 24 mg;11-P2, retention time t = 2.263 min, 18 mg. Example 156-Chloro-3-(4-chloro-2,5-difluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (15)Step 1: synthesis of 2,5-difluoro-3-methylaniline 15bAt room temperature, compound 15a (1.00 g, 5.78 mmol), iron powder (0.97 g, 17.37 mmol), and ammonium chloride (1.54 g, 28.88 mmol) were added to ethanol (10 mL) and water (3 mL). The mixture was stirred at room temperature overnight in a nitrogen atmosphere. The reaction mixture was filtered. The filter residue was washed with ethyl acetate (3 × 30 mL), and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (petroleum ether / ethyl acetate) to give compound 15b (700 mg, yield: 84%).MS(ESI) m / z = 144.1 [M+H]+.Step 2: synthesis of 4-chloro-2,5-difluoro-3-methylaniline 15cAt room temperature, compound 15b (650 mg, 4.54 mmol) and N-chlorosuccinimide (606 mg, 4.54 mmol) were dissolved in dichloromethane (10 mL). The mixture was stirred at 50 °C overnight in a nitrogen atmosphere. The reaction mixture was concentrated under reduced pressure, and the residue was purified by column chromatography (petroleum ether / ethyl acetate) to give compound 15c (300 mg, yield: 37%).Step 3: synthesis of 1-bromo-4-chloro-2,5-difluoro-3-methylbenzene 15dAt room temperature, tert-butyl nitrite (0.25 mL, 2.11 mmol) and copper bromide (408 mg, 1.83 mmol) were dissolved in acetonitrile (10 mL), and then compound 15c (250 mg, 1.41 mmol) was added in portions to the above system. The mixture was stirred at room temperature for 1 h. The reaction mixture was poured into water (30 mL), extracted with ethyl acetate (3 × 30 mL), washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by column chromatography (petroleum ether / ethyl acetate) to give compound 15d (300 mg, yield: 88%).Step 4: synthesis of methyl 6-chloro-3-(4-chloro-2,5-difluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylate 15eAt room temperature, compound 15d (200 mg, 0.83 mmol), bis(pinacolato)diboron (210 mg, 0.83 mmol), 1,1'-bis(diphenylphosphino)ferrocene palladium(II) dichloride (61.44 mg, 0.08 mmol), and potassium acetate (203 mg, 2.07 mmol) were dissolved in 1,4-dioxane (5 mL). The mixture was stirred at 80 °C overnight in a nitrogen atmosphere. Then, compound 1d (100 mg, 0.33 mmol), potassium carbonate (239 mg, 1.73 mmol), 1,1'-di-tert-butylphosphinoferrocene palladium(II) dichloride (45 mg, 0.07 mmol), and water (1 mL) were added to the above system. The mixture was stirred at 80 °C for 1 h in a nitrogen atmosphere. The reaction mixture was poured into water (30 mL), extracted with ethyl acetate (3 × 30 mL), washed with saturated brine, dried over anhydrous sodium sulfate, and filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by column chromatography (petroleum ether / ethyl acetate) to give compound 15e (120 mg, yield: 44%).Step 5: synthesis of 6-chloro-3-(4-chloro-2,5-difluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylic acid 15At room temperature, compound 15e (120 mg, 0.31 mmol) and lithium hydroxide (130 mg, 3.09 mmol) were dissolved in a mixed solvent of tetrahydrofuran (5 mL), methanol (2 mL), and water (2 mL). The mixture was stirred at room temperature for 1 h. Water (30 mL) was added. The pH was adjusted to 4-5 with hydrochloric acid (1 N, 4 mL), followed by extraction with ethyl acetate (2 × 20 mL). The organic phases were combined, washed with saturated brine (2 × 20 mL), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure to give a crude product, which was separated and purified by preparative high performance liquid chromatography (column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 45%-95%, 15 min; flow rate: 30 mL / min; peak time: 9.85 min) to give the title compound 15 (54 mg, yield: 46%). MS(ESI) m / z = 374.1 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.86 (s,1H), 8.76 (s,1H), 7.50 (t,1H), 2.35(s,3H).19F-NMR(376 MHz, DMSO-d6) δ -117.41 (d, 1F), -120.46 (d, 1F). Example 166-Chloro-3-(5-chloro-2,4-difluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (16)6-Chloro-3-(5-chloro-2,4-difluoro-3-methoxyphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (16) was prepared by using the synthesis method in Example 1 and replacing compound 1e with compound 16a.Preparative high performance liquid chromatography separation and purification conditions for compound 16: column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 40%-95%, 18 min; flow rate: 30 mL / min; peak time: 12.26 min.MS(ESI) m / z = 390.1 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.87 (s, 1H), 8.79 (s, 1H), 7.50 (t, 1H), 3.99 (s, 3H).19F-NMR(376 MHz, DMSO-d6) δ -127.96 (s, 1F), -128.99 (s, 1F). Example 176-Chloro-3-(5-chloro-2,4-difluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylic acid (17)Step 1: synthesis of 6-chloro-2,4-difluoro-3-methylaniline 17bAt room temperature, compound 17a (2 g, 13.97 mmol) and N-chlorosuccinimide (1.96 mg, 14.67 mmol) were dissolved in dichloromethane (20 mL). The mixture was stirred at 50 °C overnight and concentrated to remove the solvent. Ethyl acetate (50 mL) was added to the residue. The mixture was then washed with water (1 × 10 mL) and saturated brine (2 × 10 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give a crude product, which was then purified by column chromatography (petroleum ether / ethyl acetate) to give 17b (700 mg, yield: 28%).Step 2: synthesis of 2-bromo-1-chloro-3,5-difluoro-4-methylbenzene 17cAt room temperature, copper bromide (973 mg, 4.34 mmol) and acetonitrile (15 mL) were added to a three-necked flask, and tert-butyl nitrite (528 mg, 5.12 mmol) was added dropwise in a nitrogen atmosphere. At 20 °C, a suspension of compound 17b (700 mg, 3.94 mmol) in acetonitrile (4 mL) was added dropwise. After the addition, the mixture was stirred at room temperature for 2 h. Diluted hydrochloric acid (1 N, 10 mL) was added, followed by extraction with ethyl acetate (2 × 20 mL). The mixture was washed with water (1 × 10 mL) and saturated brine (2 × 10 mL) in sequence, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was then purified by column chromatography (petroleum ether / ethyl acetate) to give compound 17c (450 mg, yield: 47%).Step 3: synthesis of 2-(6-chloro-2,4-difluoro-3-methylphenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane 17dIn a nitrogen atmosphere, at the temperature of a dry ice-acetonitrile system, a solution of isopropylmagnesium chloride in tetrahydrofuran (2 M, 2.8 mL) was slowly added dropwise to a solution of compound 17c (450 mg, 1.86 mmol) in tetrahydrofuran (5 mL). After 3 h of stirring, 4,4,5,5-tetramethyl-2-(propan-2-yloxy)-1,3,2-dioxaborolane (0.76 mL, 3.73 mmol) was slowly added dropwise to the reaction mixture. After the addition, the mixture was slowly heated to 20 °C and stirred for 2 h. Diluted hydrochloric acid (1 N, 10 mL) was added to the reaction mixture, followed by extraction with ethyl acetate (2 × 20 mL). The mixture was washed with water (1 × 10 mL) and saturated brine (2 × 10 mL) in sequence, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was then purified by column chromatography (petroleum ether / ethyl acetate) to give compound 17d (140 mg, yield: 26%).Step 4: synthesis of methyl 6-chloro-3-(6-chloro-2,4-difluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylate 17eAt room temperature, compound 17d (100 mg, 0.32 mmol) and compound 1d (140 mg, 0.48 mmol) were dissolved in toluene / water (2 mL / 0.5 mL), and then tris(dibenzylideneacetone)dipalladium(0) (30 mg, 0.032 mmol), 2-dicyclohexylphosphino-2',6'-dimethoxybiphenyl (29 mg, 0.064 mmol), and potassium phosphate (216 mg, 0.96 mmol) were added. The mixture was stirred at 100 °C for 4 h in a nitrogen atmosphere. Ethyl acetate (60 mL) was added to the reaction system. The mixture was washed with water (1 × 10 mL) and saturated brine (2 × 10 mL) in sequence, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was then purified by column chromatography (petroleum ether / ethyl acetate) to give compound 17e (90 mg, yield: 72%).MS(ESI) m / z = 388.2 [M+H]+.Step 5: synthesis of 6-chloro-3-(5-chloro-2,4-difluoro-3-methylphenyl)thieno[3,2-b]pyridine-2-carboxylic acid 17At room temperature, compound 17e (90 mg, 0.23 mmol) was dissolved in a mixed solvent of tetrahydrofuran (2 mL), methanol (1 mL), and water (1 mL), and lithium hydroxide (56 mg, 2.3 mmol) was added. The mixture was stirred for 1 h. The pH was adjusted to 4-5 with diluted hydrochloric acid (1 N). Ethyl acetate (40 mL) was added. The mixture was washed with water (1 × 10 mL) and saturated brine (2 × 10 mL) in sequence, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product of 17, which was then separated and purified by preparative high performance liquid chromatography (column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; flow rate: 30 mL / min, 18 min; gradient: 40%-95%; retention time: 11.99 min) to give the title compound 17 (37 mg, yield: 43%).MS(ESI) m / z = 374.1 [M+H]+.1H-NMR(400 MHz, DMSO-d6) δ 8.82 (s, 1H), 8.74 (s, 1H), 7.48 (s, 1H), 2.19 (s, 3H).19F-NMR(376 MHz, DMSO-d6) δ -111.12(s, 1F), δ =-112.74(s, 1F). Example 186-Chloro-3-{6-chloro-2,4-difluoro-3-[(trideuteriomethyl)oxy]phenyl}thieno[3,2-b]pyridine-2-carboxylic acid (18); 6-Chloro-3-{6-chloro-2,4-difluoro-3-[(trideuteriomethyl)oxy]phenyl}thieno[3,2-b]pyridine-2-carboxylic acid (18-P1); 6-Chloro-3-{6-chloro-2,4-difluoro-3-[(trideuteriomethyl)oxy]phenyl}thieno[3,2-b]pyridine-2-carboxylic acid (18-P2)Step 1: synthesis of methyl 6-chloro-3-(6-chloro-2,4-difluoro-3-hydroxyphenyl)thieno[3,2-b]pyridine-2-carboxylate 18aCompound 2b (5 g, 12.4 mmol) was dissolved in dichloromethane (10 mL) in an ice bath, and a solution of boron tribromide in dichloromethane (1 M, 50 mL) was slowly added dropwise. After the addition, the mixture was stirred at room temperature for 2 h. Methanol was slowly added to the system in an ice bath. After the addition, the mixture was stirred for 10 min and concentrated under reduced pressure to give a crude product of 18a (4 g, yield: 82%), which was directly used in the next step.MS(ESI) m / z = 390.3 [M+H]+.Step 2: synthesis of methyl 6-chloro-3-(6-chloro-2,4-difluoro-3-(methoxy-d3)phenyl)thieno[3,2-b]pyridine-2-carboxylate 18bAt room temperature, the crude product of 18a (4 g, 10.3 mmol), deuterated iodomethane (0.71 mL, 11.3 mmol), and potassium carbonate (2.1 g, 15.4 mmol) were dissolved in N,N-dimethylformamide (5 mL). The mixture was stirred at 50 °C overnight. Water (30 mL) was added, followed by extraction with ethyl acetate (3 × 10 mL). The organic phases were combined, washed with saturated brine (2 × 10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product of 18b (3.5 g, yield: 83%), which was directly used in the next step.MS(ESI) m / z = 407.3 [M+H]+.Step 3: synthesis of 6-chloro-3-(6-chloro-2,4-difluoro-3-(methoxy-d3)phenyl)thieno[3,2-b]pyridine-2-carboxylic acid 18At room temperature, compound 18b (3.5 g, 8.6 mmol) was dissolved in a mixed solvent of tetrahydrofuran (6 mL), methanol (3 mL), and water (3 mL), and lithium hydroxide (2.1 g, 87 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. Water (30 mL) was added. The pH was adjusted to 4-5 with hydrochloric acid (1 N), followed by extraction with ethyl acetate (2 × 30 mL). The organic phases were combined, washed with saturated brine (2 × 10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a crude product, which was separated and purified by preparative high performance liquid chromatography (column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 35%-85%, 15 min; flow rate: 30 mL / min; peak time: 9.82 min) to give the title compound 18 (1.8 g, yield: 52%).MS (ESI) m / z = 393.3 [M+H]+.1H-NMR (400 MHz, DMSO-d6) δ 8.87 (d, 1H), 8.76 (d, 1H),7.67 (d, 1H).19F-NMR (376 MHz, DMSO-d6) δ -124.93 (s, 1F), -126.95 (s, 1F).Step 4: 6-chloro-3-{6-chloro-2,4-difluoro-3-[(trideuteriomethyl)oxy]phenyl}thieno[3,2-b]pyridine-2-carboxylic acid (18-P1); 6-chloro-3-{6-chloro-2,4-difluoro-3-[(trideuteriomethyl)oxy]phenyl}thieno[3,2-b]pyridine-2-carboxylic acid (18-P2)Compound 18 (92 mg) was resolved by SFC. Resolution method: column: ChiralPak AD, 250 × 30 mm I.D., 5 μm; mobile phase: A: CO2; B: isopropanol [0.1% NH3 (7 M in MeOH)]; gradient: B 20%; flow rate: 100 mL / min; back pressure: 100 bar; column temperature: 35 °C; wavelength: 214 nm; cycle time: about 5 min.18-P1, retention time t = 2.819 min, 32 mg;18-P2, retention time t = 2.626 min, 34 mg. Example 193-(6-Chloro-2,4-difluoro-3-methoxyphenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (19); 3-(6-Chloro-2,4-difluoro-3-methoxyphenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (19-P1); 3-(6-Chloro-2,4-difluoro-3-methoxyphenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (19-P2)Step 1: synthesis of 3-(6-chloro-2,4-difluoro-3-methoxyphenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (19)3-(6-Chloro-2,4-difluoro-3-methoxyphenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (19) was prepared by using the synthesis method in Example 2 and replacing compound 1d with compound 5d.Preparative high performance liquid chromatography separation and purification conditions for compound 19: column: Waters Xbridge, 30 × 250 mm; mobile phase A: 0.1% formic acid in water; mobile phase B: acetonitrile; gradient: 35%-95%, 18 min; flow rate: 30 mL / min; peak time: 10.89 min.MS (ESI) m / z = 374.2 [M+H]+.1H-NMR (400 MHz, DMSO-d6) δ 8.77 (d, 1H), 8.65 (dd, 1H), 7.61 (dd, 1H), 3.95 (s, 3H).19F-NMR (376 MHz, DMSO-d6) δ -124.96 (d, 1F), -126.96 (d, 1F), -127.19 (s, 1F).Step 2: 3-(6-chloro-2,4-difluoro-3-methoxyphenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (19-P1); 3-(6-chloro-2,4-difluoro-3-methoxyphenyl)-6-fluorothieno[3,2-b]pyridine-2-carboxylic acid (19-P2)The product 19 (520 mg) was resolved by SFC. Resolution method: column: ChiralPak IC, 250 × 30 mm I.D., 5 μm; mobile phase: A: CO2; B: methanol [0.2% TFA]; gradient: B 20%; flow rate: 100 mL / min; back pressure: 100 bar; column temperature: 35 °C; wavelength: 210 nm; cycle time: about 2 min.19-P1, retention time t = 2.409 min, 240 mg;19-P2, retention time t = 1.993 min, 234 mg. Biological EvaluationsThe following test examples further describe and illustrate the present disclosure, but these examples are not intended to limit the scope of the present disclosure.Test Example 1: Assay on Inhibitory Effect of Compounds on BCKDK Protein in HEK293 Cells1. Experimental materialsName of reagentManufacturerCat. No.DMEM (4.5 g / L) complete culture mediumGibco11995040Phospho-BCKDH-E1α (Ser293) (E2V6B) rabbit mAbCST40368SCellTag 520S tainLicor926-41094Intercept® (TBS) blocking bufferLicor927-60001IRDye® 800CW goat anti-rabbit IgG secondary antibodyLicor926-32211Odyssey imager M Licor / HEK293 cellsATCCCRL-15732. Test methodHEK293 cells were seeded in a 384-well black clear-bottom plate at 10,000 cells / well and incubated overnight. The cells were treated with different concentrations of compounds (100 μM maximum concentration, 3-fold dilution, 10 concentrations) at 37 °C with 5% carbon dioxide for 4 h. Pre-cooled 8% paraformaldehyde was added to the plate at 50 μL / well, and the cells were fixed for 1 h in the dark. The fixative solution was pipetted and discarded. A permeabilization solution was added at 50 μL / well, and the mixture was left to stand for 5 min each time. The process was repeated 6 times. A blocking buffer was then added at 50 μL / well, and the mixture was incubated on a shaker at room temperature for one hour. The primary antibody, Phospho-BCKDH-E1α (Ser293) (1000×), was added, followed by incubation at 4 °C for 24 h. After the permeabilization step was repeated again, the secondary antibodies, Cell Tag520 (500×) & IRDye 800CW (1000×), were added, followed by incubation at room temperature for 1 h in the dark. The permeabilization step was repeated once more, and the permeabilization solution was completely pipetted away at the last time. The signals were detected using the Odyssey imager M imaging system, data were processed using XL fit, and IC50 was calculated.3. Experimental resultsThe inhibition of the BCKDK protein in HEK293 cells by the compounds of the present disclosure was determined by the above assay. The IC50 values obtained are shown in Table 1.Table 1. IC50 (μM) of compounds of the present disclosure against HEK293AExample No.IC50 (μM) 15.662-P10.9634.7242.6159.6662.8371.3684.3695.92103.5711-P10.56154.1318-P10.8919-P11.56PF-073289481.69PF-07328948 was the compound of Example 9 in the patent WO2023100061A1, with the structure being . Test Example 2: Inhibitory Activity of Compounds Against Human CYP450 (CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A)1. Equipment, materials, and reagents1) Human liver microsome (HLM)Human liver microsomes (source: BioIVT) were stored in a freezer at -80 °C. Before use, the human liver microsomes were taken out from the freezer and pre-heated and thawed in a 37 °C water bath, and then placed on ice for later use.2) SubstrateDetails of the preparation of a substrate mixture solution are as follows: The substrate mixture solution was composed of 50% of purified water and 50% of organic solvent, and the prepared substrate mixture solution was stored in a freezer at -20 °C, which was warmed to room temperature and then vortexed for 30 s before use.Substrate mixture solutions of CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3ACYPEnzymeSubstrateMW (g / mol)Stock solutionconcentration (mM)Additionvolume (μL)Totalvolume (μL)Working solutionconcentration (mM)Finalconcentration (μM)1A2Phenacetin179.230 (dissolved in acetonitrile)502506302C9Diclofenac sodium318.110 (dissolved in water)502102C19S-mephenytoin218.235 (dissolved in acetonitrile)507352D6Bufuralol hydrochloride297.85 (dissolved in water)50153AMidazolam325.83 (dissolved in DMSO)500.633) Phosphate buffer (100 mmol / L, pH 7.4)Disodium hydrogen phosphate (analytically pure) and potassium dihydrogen phosphate (analytically pure) were purchased from local suppliers. Preparation of solution A: 7.098 g of disodium hydrogen phosphate was accurately weighed out, and 500 mL of purified water was added. The mixture was subjected to ultrasonic treatment. Preparation of solution B: 3.400 g of potassium dihydrogen phosphate was accurately weighed out, and 250 mL of purified water was added. The mixture was subjected to ultrasonic treatment. The solution B was added to the solution A, and the final pH was 7.4.4) 10 mmol / L NADPH solutionNADPH (MW: 833.4 g / mol) was purchased from MCE and dissolved at 8.334 mg / mL in a phosphate buffer.2. Experimental process1) Preparation of incubation solutionThe incubation system for the mixed solution of the substrate and human liver microsome is as follows:BufferWorking solution concentrationVolumeFinal concentrationLiver microsome20 mg / mL2 μL0.2 mg / mLPhosphate buffer100 mM176 μL100 mMSubstrate-1 μL-2) Compound dilution15 μL of a 10 mmol / L DMSO solution of the test compound was added to a deep-well plate containing sequentially labeled compounds. The dilution steps are as follows:Dilution of test compoundsStock solution / PositionVolume of compound (μL)Volume of DMSO (μL)Working solution concentration (mM)Final concentration (μM)A115 μL of 10 mM stock solution10630B19 μL A118210C19 μL B1210.63D19 μL C1180.21E19 μL D1210.060.3F19 μL E1180.020.13) IncubationEach well of a 96-well deep-well plate contained 179 μL of a mixed solution of the substrate and human liver microsome phosphate buffer, and 1 μL of the test compound or a blank solution. The incubation plate was placed in a 37 °C water bath and pre-incubated for 5 min, and then 20 μL of 10 mM NADPH was added to initiate the reaction. After the addition of NADPH solution, the incubation plate was incubated for another 5 min at 37 °C.4) Quenching of reactionThe reaction was quenched by adding 300 μL of an acetonitrile solution containing 3% formic acid, 200 nM tolbutamide, 200 nM alprazolam, and 200 nM labetalol. The mixture was then centrifuged in a centrifuge at 3220 g for 50 min, and 200 μL of the supernatant was taken for LC / MS / MS analysis.3. Data processingThe peak areas of all samples and the peak area of an internal standard were calculated automatically and imported into Excel software.The percentage of residual activity was calculated according to the following formula:Area ratio = peak areatest compound / peak areainternal standardPercentage of residual activity (%) = area ratiotest compound / area ratioblank × 100The IC50 values (the concentration of the test compound at which 50% inhibition was achieved) were calculated using Excel XLfit 5.3.1.3.Table 2. Inhibitory activity IC50 (μM) of compounds of the present disclosure against CYP450Test compoundCYP1A2CYP2C9CYP2C19CYP2D6CYP3A19-P1>30>30>30>30>30PF-07328948>301.09>30>30>30Reference compound 15.02.3215.76>30>30Reference compound 1 was the compound of Example 16 in the patent WO2023100061A1.According to the results, compound 19-P1 did not inhibit CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A (IC50 > 30 μM), PF-07328948 showed strong inhibition on CYP2C9 (IC50 = 1.09 μM), and reference compound 1 showed relatively strong inhibition on CYP1A2 (IC50 = 5.0 μM), CYP2C9 (IC50 > 2.32 μM), and CYP2C19 (IC50 = 15.76 μM). Test Example 3: Stability Test of Compounds in Hepatocytes1. Experimental steps1) The test compound was dissolved in DMSO to prepare a high-concentration (10 mM) stock solution. Before use, the stock solution was diluted with DMSO to prepare a 100 μM working solution. The final concentration of the test compound was 1 μM.2) A tube of cryopreserved hepatocytes (source of rat, dog, and human hepatocytes: BioIVT; source of monkey hepatocytes: RILD) was taken. It was ensured that the hepatocytes were still in a low-temperature frozen state before thawing. The hepatocytes were quickly placed in a 37 °C water bath, gently shaken until all ice crystals were dispersed, sprayed with 70% ethanol, and then transferred to a biosafety cabinet.3) The contents of small tubes of hepatocytes from different species were poured into a centrifuge tube containing 50 mL of thawing culture medium and centrifuged at 100 g for 10 min. After centrifugation, the thawing culture medium was pipetted off, and a sufficient amount of incubation culture medium was added to obtain a cell suspension with a cell density of about 1.0 × 106 cells / mL.4) Hepatocytes were counted using Cellometer Vision, and the viable cell density was determined. The viability of hepatocytes must be greater than 75%. The hepatocyte suspension was diluted with the incubation culture medium to a viable cell density of 0.5 × 106 cells / mL.5) 198 μL of the viable cell suspension was transferred to a 96-well deep-well plate, and the deep-well plate was placed on a vortexer and pre-heated in an incubator for 10 min. In the experiment, the incubation was performed in duplicate.6) 2 μL of the 100 μM test compound was added to each well to start a reaction, and the deep-well plate was placed back on the vortexer in the incubator.7) The samples were incubated. At 0 min, 15 min, 30 min, 60 min, 90 min, and 120 min, 25 μL of the suspension was taken, and 150 μL of acetonitrile containing an internal standard was added to stop the reaction. After 10 min of vortexing, the mixture was centrifuged at 3220 g at 4 °C for 45 min. 100 μL of the supernatant was transferred to a sample injection plate, and 100 μL of purified water was added. After thorough mixing, the mixture was analyzed by UPLC-MS / MS.2. Data analysisAll data calculations were performed through Microsoft Excel software. The peak areas were determined through an extracted ion chromatogram. Through linear fitting of the natural logarithm of the parent drug elimination percentage versus time, the in vitro clearance rate of the parent drug was determined.The in vitro clearance rate (μL / min / 106 cells) was calculated using the following formula:In vitro CLint = kV / N,where V = the incubation volume per well (0.2 mL);N = the number of cells per well (0.1 × 106 cells).Table 3. Metabolic clearance rates CLint (μL / min / 106 cells) of compounds in hepatocytesTest compoundRatDogMonkeyHuman19-P120.18002.38PF-0732894829.057.0428.6715.03Reference compound 129.2318.7343.9815.35According to the results, compound 19-P1 exhibited significantly greater metabolic stability in dog, monkey, and human hepatocytes than PF-07328948 and reference compound 1. Test Example 4: In Vivo Pharmacokinetic Experiment in Rats and Dogs1. In vivo pharmacokinetic experiment in ratsSD rats (source: Zhejiang Vital River Laboratory Animal Technology Co., Ltd.) were used as test animals. The plasma concentrations of the test compound at different time points after intragastric administration of the test compound to rats were determined using an LC / MS / MS method. The pharmacokinetic behavior of the test compound in rats was studied, and its pharmacokinetic profile was evaluated.Test animals: two healthy male rats aged 6-8 weeks (200-300 g) per group.Compound solution preparation: A certain amount of the compound was weighed out and dissolved in a vehicle (100% normal saline and 1 mol / L hydrochloric acid and sodium hydroxide, allowing the final solution to be adjusted to pH 7) to form a 2 mg / mL colorless, clear solution.Dosing: Rats were fasted overnight and then intragastrically dosed. The test compound was administered at a dose of 20 mg / kg.Procedure: Rats were intragastrically administered with the test compound. At 0.25 h, 0.5 h, 1 h, 2 h, 4 h, 6 h, 8 h, and 24 h after administration, blood samples (about 0.2 mL) were collected via jugular catheterization, placed into test tubes containing EDTA-K2, and centrifuged at around 4 °C at 4000 g per minute for 5 min to separate plasma, and the plasma was stored at -75±15 °C.Determination of the content of the compound to be tested in rat plasma after the test compound was orally administered: 50 μL of the rat plasma at each time point after administration was vortexed with 5 μL of a blank solution and 200 μL of a solution of dexamethasone (internal standard) in acetonitrile for 30 s, and the mixture was centrifuged for 15 min (3900 rpm). 100 μL of the supernatant of the plasma sample was mixed well with 200 μL of ultrapure water, and then 5 μL of the dilution was taken using a syringe for LC / MS / MS analysis.Table 4. Pharmacokinetic parameters of compounds after oral administration to rats (20 mpk)Test compoundT1 / 2 (h)Cmax (ng / mL)AUClast (h*ng / mL)19-P1NA779001297444PF-073289484.1151250223769Note: All the shown values were the mean of two animals.According to the results, compound 19-P1 exhibited significantly greater oral exposure in rats than PF-07328948. 2. In vivo pharmacokinetic experiment in dogsBeagles (source: Beijing Marshall Biotechnology Co., Ltd.) were used as test animals. The plasma concentrations of the test compound at different time points after intragastric administration of the test compound to beagles were determined using an LC / MS / MS method. The pharmacokinetic behavior of the test compound in beagles was studied, and its pharmacokinetic profile was evaluated.Test animals: two healthy male beagles aged 8 months to 3 years (6.0-13.0 kg) per group.Compound solution preparation: A certain amount of the compound was weighed out and dissolved in a vehicle (100% normal saline and 1 mol / L hydrochloric acid and sodium hydroxide, allowing the final solution to be adjusted to pH 7) to form a 0.6 mg / mL colorless, clear solution.Dosing: Beagles were fasted overnight and then intragastrically dosed. The test compound was administered at a dose of 3 mg / kg.Procedure: Beagles were intragastrically administered with the test compound. Before administration and at 0.25 h, 0.5 h, 1 h, 2 h, 4 h, 6 h, 8 h, and 24 h after administration, blood samples (about 0.6 mL) were collected via peripheral venipuncture, placed into test tubes containing EDTA-K2, and centrifuged at around 4 °C at 2000 g per minute for 10 min to separate plasma, and the plasma was stored at -75±15 °C.Determination of the content of the compound to be tested in beagle plasma after the test compound was orally administered: 50 μL of the beagle plasma at each time point after administration was vortexed with 5 μL of a blank solution and 200 μL of a solution of dexamethasone (internal standard) in acetonitrile for 30 s, and the mixture was centrifuged for 15 min (3900 rpm). 100 μL of the supernatant of the plasma sample was mixed well with 200 μL of ultrapure water, and then 5 μL of the dilution was taken using a syringe for LC / MS / MS analysis.Table 5. Pharmacokinetic parameters of compounds after oral administration to dogs (3 mpk)Test compoundT1 / 2 (h)Cmax (ng / mL)AUClast (h*ng / mL)19-P15.7718250102182PF-073289485.421390020741Note: All the shown values were the mean of two animals.According to the results, compound 19-P1 exhibited significantly greater oral exposure in dogs than PF-07328948.

Claims

1. A compound represented by formula (I) or a pharmaceutically acceptable salt thereof,wherein R1, R2, and R3 are each independently selected from the group consisting of hydrogen, fluorine, chlorine, and methyl;R4 is selected from the group consisting of fluorine and chlorine;R5 is selected from the group consisting of C1-4 alkyl, C1-4 haloalkyl, 3- to 4-membered cycloalkyl, and -L1-R9, wherein L1 is selected from the group consisting of sulfur and oxygen, and R9 is selected from the group consisting of C1-4 alkyl, C1-4 haloalkyl, deuterated C1-4 alkyl, and 3- to 4-membered cycloalkyl; X1 is selected from the group consisting of nitrogen and CR6, wherein R6 is selected from the group consisting of hydrogen, fluorine, and chlorine;R7 and R8 are each independently selected from the group consisting of hydrogen, fluorine, and chlorine.

2. The compound represented by formula (I) or the pharmaceutically acceptable salt thereof according to claim 1, wherein X1 is nitrogen.

3. The compound represented by formula (I) or the pharmaceutically acceptable salt thereof according to claim 1, being a compound represented by formula (II) or a pharmaceutically acceptable salt thereof,wherein R1, R2, R3, R4, R5, R6, R7, and R8 are each as defined in claim 1.

4. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, wherein R5 is selected from the group consisting of C1-4 alkyl and C1-4 haloalkyl; preferably, R5 is selected from the group consisting of methyl, ethyl, and isopropyl; most preferably, R5 is selected from the group consisting of methyl and ethyl.

5. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, wherein R5 is selected from the group consisting of 3- to 4-membered cycloalkyl; preferably, R5 is cyclopropyl.

6. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, being a compound represented by formula (II-1) or a pharmaceutically acceptable salt thereof,wherein R1, R2, R3, R4, R6, R7, R8, and R9 are each as defined in claim 1.

7. The compound or the pharmaceutically acceptable salt thereof according to claim 6, wherein R9 is selected from the group consisting of C1-4 alkyl, C1-4 haloalkyl, and deuterated C1-4 alkyl; preferably, R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl; most preferably, R9 is selected from the group consisting of methyl, difluoromethyl, trifluoromethyl, and deuterated methyl.

8. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 3, being a compound represented by formula (II-2) or a pharmaceutically acceptable salt thereof,wherein R1, R2, R3, R4, R6, R7, R8, and R9 are each as defined in claim 1.

9. The compound or the pharmaceutically acceptable salt thereof according to claim 8, wherein R9 is selected from the group consisting of C1-4 alkyl, C1-4 haloalkyl, and deuterated C1-4 alkyl; preferably, R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl; most preferably, R9 is methyl.

10. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 9, wherein at least one of R1, R2, and R3 is selected from the group consisting of fluorine and chlorine.

11. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 10, wherein R1 and R3 are each independently selected from the group consisting of hydrogen, fluorine, and chlorine; preferably, one of R1 and R3 is hydrogen, and the other is selected from the group consisting of hydrogen, fluorine, and chlorine; most preferably, R1 and R3 are each independently hydrogen.

12. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 11, wherein R2 is fluorine or chlorine; preferably, R2 is fluorine; preferably, R2 is chlorine.

13. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 12, wherein R1 and R3 are each independently hydrogen; R2 is fluorine or chlorine.

14. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 13, wherein R4 is selected from the group consisting of fluorine and chlorine; preferably, R4 is fluorine.

15. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 14, wherein R1 and R3 are each independently hydrogen, R2 is fluorine or chlorine, and R4 is fluorine.

16. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 15, wherein at least one of R6, R7, and R8 is selected from the group consisting of fluorine and chlorine; preferably, at least two of R6, R7, and R8 are selected from the group consisting of fluorine and chlorine.

17. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 16, wherein R6 is selected from the group consisting of fluorine and chlorine.

18. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 17, wherein at least one of R6, R7, and R8 is chlorine.

19. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 18, wherein R7 is selected from the group consisting of fluorine and chlorine, and R8 is hydrogen; preferably, R7 is fluorine, and R8 is hydrogen.

20. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 19, wherein R8 is selected from the group consisting of fluorine and chlorine, and R7 is hydrogen.

21. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 20, wherein R1 and R3 are each independently hydrogen; R2 is fluorine or chlorine; R4 is fluorine; R6 is selected from the group consisting of fluorine and chlorine; R7 is selected from the group consisting of fluorine and chlorine, and R8 is hydrogen; or R8 is selected from the group consisting of fluorine and chlorine, and R7 is hydrogen; and at least one of R6, R7, and R8 is chlorine.

22. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 21, being a compound represented by formula (II-A) or formula (II-B) or a pharmaceutically acceptable salt thereof, preferably a compound represented by formula (II-B) or a pharmaceutically acceptable salt thereof,, wherein R2, R5, R6, and R7 are each as defined in claims 1 to 21; preferably, R2 is chlorine; or preferably, R2 is fluorine.

23. The compound or the pharmaceutically acceptable salt thereof according to claim 22, wherein R5 is selected from the group consisting of methyl and ethyl.

24. The compound or the pharmaceutically acceptable salt thereof according to claim 22, wherein R5 is selected from the group consisting of -L1-R9, wherein L1 is oxygen, and R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl; preferably, R9 is selected from the group consisting of methyl and deuterated methyl; or preferably, R9 is selected from the group consisting of difluoromethyl and trifluoromethyl.

25. The compound or the pharmaceutically acceptable salt thereof according to claim 22, wherein R5 is selected from the group consisting of -L1-R9, wherein L1 is sulfur, and R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl; preferably, R5 is selected from the group consisting of -L1-R9, wherein L1 is sulfur, and R9 is methyl or deuterated methyl.

26. The compound or the pharmaceutically acceptable salt thereof according to claim 22, wherein R6 is fluorine or chlorine, and R7 is hydrogen; preferably, R6 is fluorine, and R7 is hydrogen.

27. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 16, 18, 19, and 21, being a compound represented by formula (II-C) or a pharmaceutically acceptable salt thereof,, wherein R2, R5, R6, and R7 are each as defined in claims 1 to 16, 18, 19, and 21; preferably, R2 is chlorine; or preferably, R2 is fluorine.

28. The compound or the pharmaceutically acceptable salt thereof according to claim 27, wherein R5 is selected from the group consisting of methyl and ethyl.

29. The compound or the pharmaceutically acceptable salt thereof according to claim 27, wherein R5 is selected from the group consisting of -L1-R9, wherein L1 is oxygen; R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl; preferably, R9 is selected from the group consisting of methyl and deuterated methyl, or preferably, R9 is selected from the group consisting of difluoromethyl and trifluoromethyl.

30. The compound or the pharmaceutically acceptable salt thereof according to claim 27, wherein R5 is selected from the group consisting of -L1-R9, wherein L1 is sulfur; R9 is selected from the group consisting of methyl, ethyl, difluoromethyl, trifluoromethyl, and deuterated methyl; preferably, R9 is methyl or deuterated methyl.

31. The compound or the pharmaceutically acceptable salt thereof according to claim 27, wherein at least one of R6 and R7 is selected from the group consisting of fluorine and chlorine; preferably, R6 is selected from the group consisting of fluorine and chlorine; most preferably, R6 is selected from the group consisting of fluorine, and R7 is selected from the group consisting of fluorine and chlorine; or R6 is selected from the group consisting of chlorine, and R7 is selected from the group consisting of fluorine and chlorine.

32. The compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 31, being selected from the group consisting of the following compounds or pharmaceutically acceptable salts thereof:, , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , and .

33. An isotopically substituted form of the compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 32, wherein preferably, the isotopically substituted form is a deuterated form.

34. A pharmaceutical composition, comprising the compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 32, or the isotopically substituted form according to claim 33, and one or more pharmaceutically acceptable excipients.

35. Use of the compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 32, or the isotopically substituted form according to claim 33, or the pharmaceutical composition according to claim 34 in the manufacture of a medicament for preventing and / or treating a BCKDK-related disease, wherein preferably, the BCKDK-related disease is selected from the group consisting of a glucose metabolism disorder or an abnormal blood glucose disease, a heart disease, and a kidney disease; more preferably, the glucose metabolism disorder or the abnormal blood glucose disease is diabetes, and the heart disease is heart failure.

36. Use of the compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 32, or the isotopically substituted form according to claim 33, or the pharmaceutical composition according to claim 34 in the manufacture of a medicament for preventing and / or treating a disease, wherein the disease is selected from the group consisting of a glucose metabolism disorder or an abnormal blood glucose disease, a heart disease, and a kidney disease; preferably, the glucose metabolism disorder or the abnormal blood glucose disease is diabetes, and the heart disease is heart failure.

37. A preparation method for the compound or the pharmaceutically acceptable salt thereof according to any one of claims 1 to 32, comprising a step of removing a protecting group PG from a compound represented by formula (I-a) or a pharmaceutically acceptable salt thereof under an alkaline condition, and further comprising a step of adjusting the pH to acidity,wherein PG is a carboxyl protecting group; R1, R2, R3, R4, R5, X1, R7, and R8 are each as defined in claims 1 to 32; preferably, PG is C1-6 alkyl.

38. A compound represented by formula (I-a) or a pharmaceutically acceptable salt thereof,wherein PG is a carboxyl protecting group; R1, R2, R3, R4, R5, X1, R7, and R8 are each as defined in claims 1 to 32; preferably, PG is C1-6 alkyl. 

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