Compound capable of stabilizing mutant phenylalanine hydroxylase, and pharmaceutical composition and use thereof

Abstract

The present invention provides a compound (I) capable of stabilizing mutant phenylalanine hydroxylase, a pharmaceutical composition comprising same, and a preparation method therefor and a use thereof. The compound can effectively restore the activity of a mutant phenylalanine hydroxylase protein, is used for treating disorders and / or diseases caused by phenylalanine hydroxylase mutations, such as phenylketonuria, and is used in preparation of drugs for such disorders or diseases.

Description

Compounds capable of stabilizing phenylalanine hydroxylase mutations, pharmaceutical compositions, and uses thereof

[0001] The present invention claims the right to the patent application number 202410353484.1 filed on March 26, 2024 with the State Intellectual Property Office of China, entitled “Compounds, pharmaceutical compositions and uses thereof capable of stabilizing phenylalanine hydroxylase mutations”; the patent application number 202410451080.6 filed on April 15, 2024 with the State Intellectual Property Office of China, entitled “Compounds, pharmaceutical compositions and uses thereof capable of stabilizing phenylalanine hydroxylase mutations”; the patent application number 202410524217.6 filed on April 28, 2024 with the State Intellectual Property Office of China, entitled “Compounds, pharmaceutical compositions and uses thereof capable of stabilizing phenylalanine hydroxylase mutations”. the patent application number 202410692809.9 filed on May 30, 2024 to the State Intellectual Property Office of China, entitled “Compounds capable of stabilizing phenylalanine hydroxylase mutations, pharmaceutical compositions and uses thereof”; the patent application number 202410788286.8 filed on June 18, 2024 to the State Intellectual Property Office of China, entitled “Compounds capable of stabilizing phenylalanine hydroxylase mutations, pharmaceutical compositions and uses thereof”; the patent application number 20241103 filed on July 30, 2024 to the State Intellectual Property Office of China, entitled “Compounds capable of stabilizing phenylalanine hydroxylase mutations, pharmaceutical compositions and uses thereof” 4386.8, entitled “Compounds, pharmaceutical compositions and uses thereof capable of stabilizing phenylalanine hydroxylase mutations”; Patent application number 202411287832.6, filed with the State Intellectual Property Office of China on September 13, 2024, entitled “Compounds, pharmaceutical compositions and uses thereof capable of stabilizing phenylalanine hydroxylase mutations”; Patent application number 202411433007.2, filed with the State Intellectual Property Office of China on October 14, 2024, entitled “Compounds, pharmaceutical compositions and uses thereof capable of stabilizing phenylalanine hydroxylase mutations”; Patent application number 202411433007.2, filed with the State Intellectual Property Office of China on November 14, 2024 Priority is claimed to the prior application, patent application number 202411628970.6, entitled “Compounds, pharmaceutical compositions and uses thereof capable of stabilizing phenylalanine hydroxylase mutations” filed with the State Intellectual Property Office of China on January 16, 2025, patent application number 202510072400.1, entitled “Compounds, pharmaceutical compositions and uses thereof capable of stabilizing phenylalanine hydroxylase mutations” filed with the State Intellectual Property Office of China on March 20, 2025, patent application number 202510336009.8, entitled “Compounds, pharmaceutical compositions and uses thereof capable of stabilizing phenylalanine hydroxylase mutations” filed with the State Intellectual Property Office of China on March 20, 2025. The entire contents of the above-mentioned prior applications are incorporated herein by reference. Technical Field

[0002] The present invention belongs to the field of pharmaceutical compounds, and in particular relates to a small molecule compound capable of stabilizing phenylalanine hydroxylase mutations, a pharmaceutical composition and uses thereof. Background Art

[0003] Phenylketonuria (PKU) is an inherited metabolic disorder caused by a defect or absence of the enzyme phenylalanine hydroxylase (PAH), preventing patients from properly metabolizing phenylalanine. The global prevalence of PKU is approximately one in 10,000 individuals.

[0004] Phenylalanine hydroxylase (PAH) is an enzyme present in humans and other organisms that catalyzes the conversion of phenylalanine to tyrosine. Normal phenylalanine metabolism requires both PAH and BH4. PAH is the rate-limiting enzyme in the conversion of phenylalanine to tyrosine; BH4 serves as a coenzyme for PAH. Approximately 80-90% of daily phenylalanine intake must be metabolized through this pathway. PAH mutations or BH4 deficiency can lead to accumulation of phenylalanine in the blood. In patients with phenylketonuria, phenylalanine accumulates in the blood. If left untreated, it can lead to intellectual disability and other serious health problems. Children with PKU generally have no special symptoms at birth. Untreated children begin to gradually develop the disease 3-4 months after birth. Their hair turns from black to yellow, their skin becomes pale, and their urine and sweat have a rat urine odor. Adult patients may experience lower limb spasticity and cerebellar ataxia, tremors, encephalopathy and abnormal vision. Severe cases may lead to death.

[0005] Currently, the treatment for PKU mainly consists of food therapy and drug therapy. Food therapy strictly limits the intake of natural amino acids and amino acid mixtures containing phenylalanine, and consumes low-protein foods. Since protein and / or phenylalanine deficiency can also have adverse effects, including growth restriction, anorexia, hair loss, lethargy, and eczematous outbreaks, food therapy is very challenging and difficult for patients to adhere to. Drug treatments mainly include tetrahydrobiopterin and polyethylene glycol phenylalanine ammonia lyase. Each drug has its own limitations. For example, tetrahydrobiopterin is only effective for a small number of patients (usually patients with mild PKU or BH4 deficiency); polyethylene glycol phenylalanine ammonia lyase requires daily subcutaneous injection, and long-term injection is prone to adverse immune reactions, and the treatment cost is expensive.

[0006] Mutated phenylalanine hydroxylase (PHH) proteins become unstable and susceptible to degradation. The compounds of the present invention can stabilize mutated PHH, restore its activity, mitigate its degradation, and metabolize phenylalanine to tyrosine, demonstrating their potential for treating PKU caused by PHH mutations. Summary of the Invention

[0007] To improve the above technical problems, the present invention provides a compound represented by formula (I), its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or prodrug compound:

[0008] in,

[0009] The fused ring group is formed by the fusion of ring A and ring B, and ring A is selected from a benzene ring, a 5-6 membered heteroaromatic ring, a saturated or partially unsaturated C 3-12 Carbocyclic ring or 3-14 membered heterocyclic ring; Ring B is selected from 5 membered heteroaromatic ring, saturated or partially unsaturated C 3-12 carbocyclic ring or 3-14 membered heterocyclic ring; and when ring A is a 5-membered heteroaromatic ring, ring B is a 5-membered heteroaromatic ring;

[0010] Each R a The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: OH, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, halo C 1-12 Alkyl, C 1-12 Alkoxy, C 1-12 Alkylthio, C 3-12 Cycloalkyl, 3-14 membered heterocyclic group, C 6-14 Aryl, 5-14 membered heteroaryl, -N(R 11 )(R 12 ) or -S(O)2R 13 ; or, two adjacent R a Together with the atoms to which they are attached, they form an unsubstituted or optionally substituted R a1 Substituted C 3-6 Cycloalkyl ring, 3-6 membered heterocyclic ring, C 6-10 aromatic ring or 5-10 membered heteroaromatic ring; each R a1 The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R a2 Substituted with the following groups: OH, NH2, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group, C 6-10 Aryl, 5-10 membered heteroaryl, -N(R 14 )(R 15 ) or -S(O)2R 16; Each R a2 the same or different, independently selected from OH, -N(R 17 )(R 18 ), CN, halogen, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl or 3-6 membered heterocyclic group; R 11 、R 12 、R 13 、R 14 、R 15 、R 16 、R 17 、R 18 The same or different, independently selected from H, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 3-12 Cycloalkyl;

[0011] Each R c the same or different, independently selected from deuterium or C 1-12 alkyl;

[0012] X1 is selected from O or S;

[0013] Y1 is absent or selected from unsubstituted or optionally substituted with one, two or more R d Substituted with the following groups: C 6-14 Arylene, 5-14 membered heteroarylene, C 6-14 Arylene-C 1-12 Alkylene, 5-14 membered heteroaryl-C 1-12 Alkylene, C 6-14 Arylene-O-, 5-14 membered heteroarylene-O-, C 6-14 Arylene-NH-, 5-14 membered heteroarylene-NH-, C 6-14 Arylene-CO- or 5-14 membered heteroarylene-CO-; each R d The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R d1 Substituted with the following groups: OH, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 1-12 Alkoxy, C 1-12 Alkylthio, C 3-12 Cycloalkyl, 3-14 membered heterocyclic group, C 6-14 Aryl, 5-14 membered heteroaryl, -N(R 21 )(R 22 ), or -S(O)2R 23 ; Each R d1the same or different, independently selected from OH, -N(R 24 )(R 25 ), CN, halogen, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group; R 21 、R 22 、R 23 、R 24 、R 25 The same or different, independently selected from H, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 3-12 Cycloalkyl;

[0014] Ring C is selected from C 6-14 Aromatic ring or 5-14 membered heteroaromatic ring;

[0015] Each R b The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R b1 Substituted with the following groups: OH, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, halo C 1-12 Alkyl, C 1-12 Alkoxy, C 1-12 Alkylthio, C 3-12 Cycloalkyl, C 3-12 Carbocyclic ring, 3-14 membered heterocyclic ring, C 6-14 Aryl, 5-14 membered heteroaryl, -N(R 31 )(R 32 ), hydroxyl C 1-12 Alkyl, C 1-12 Alkyl-N(R 31 )(R 32 )、-S(O)2R 33 、 Each R b1 The same or different, independently selected from oxo (=O), OH, -N(R 34 )(R 35 ), CN, halogen, C 1- 6 alkyl, C 1-6 Alkoxy, halogenated C 1-6 Alkyl, hydroxyl C 1-12 Alkyl, C 1-12 Alkyl-N(R 34 )(R 35 ), C 3-6 Cycloalkyl, 3-6 membered heterocyclic group; R 31 、R32 、R 33 、R 34 、R 35 、R 36 、R 37 、R 38 、R 39 The same or different, independently selected from H, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 3-12 Cycloalkyl;

[0016] m1 is selected from 0, 1, 2 or 3;

[0017] m2 is selected from 0, 1, 2, 3, 4, 5 or 6;

[0018] m3 is selected from 0, 1, 2, 3, 4, 5, 6, 7 or 8;

[0019] n is selected from 0, 1 or 2.

[0020] According to some embodiments, m3 is 0.

[0021] According to some embodiments, m3 is 1.

[0022] According to some embodiments, R c Selected from methyl.

[0023] According to some embodiments, n is 1.

[0024] According to some embodiments, Y1 is absent or selected from unsubstituted or optionally substituted with one, two or more R d substituted 5-membered heteroarylene, 5-membered heteroarylene-CH2-, 5-membered heteroarylene-O-, 5-membered heteroarylene-NH-.

[0025] According to some embodiments, Y1 is absent or selected from unsubstituted or optionally substituted with one, two or more R d substituted 5-membered heteroarylene, 5-membered heteroarylene-CH2-, 5-membered heteroarylene-O-.

[0026] According to some embodiments, Y1 is absent or selected from unsubstituted or optionally substituted with one, two or more R d Substituted 5-membered heteroarylene.

[0027] According to some embodiments, Y1 is absent.

[0028] According to some embodiments, Y1 is selected from unsubstituted or optionally substituted with one R d Substituted groups: The “*” side is connected to the carbonyl group, and the “#” side is connected to the ring C.

[0029] According to some embodiments, Y1 is selected from unsubstituted or optionally substituted with one R d Substituted groups: The “*” side is connected to the carbonyl group, and the “#” side is connected to the ring C.

[0030] According to some embodiments, Y1 is selected from unsubstituted or optionally substituted with one R d Substituted groups: The “*” side is connected to the carbonyl group, and the “#” side is connected to the ring C.

[0031] According to some embodiments, each R d the same or different, independently selected from CN, halogen, OH, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy or C 3-6 Cycloalkyl.

[0032] According to some embodiments, each R d are identical or different and are independently selected from methyl, difluoromethyl, trifluoromethyl, Cl, Br or cyclopropyl.

[0033] According to some embodiments, Y1 is absent or selected from The “*” side is connected to the carbonyl group, and the “#” side is connected to the ring C.

[0034] According to some embodiments, Y1 is absent or selected from The “*” side is connected to the carbonyl group, and the “#” side is connected to the ring C.

[0035] According to some embodiments, Y1 is absent or selected from The “*” side is connected to the carbonyl group, and the “#” side is connected to the ring C.

[0036] According to some embodiments, ring C is selected from phenyl or 5-10 membered heteroaryl.

[0037] According to some embodiments, ring C is selected from a benzene ring, a pyrrole ring, a thiophene ring, a furan ring, a pyrazole ring, an imidazole ring, a thiazole ring, an oxazole ring, an isoxazole ring, a dioxazole ring, a pyridine ring, a pyridone ring, a pyrimidine ring, and a pyrazolopyridine ring.

[0038] According to some embodiments, ring C is selected from a benzene ring, a pyrrole ring, a thiophene ring, a furan ring, a pyrazole ring, an imidazole ring, a thiazole ring, an oxazole ring, a dioxazole ring, a pyridine ring, a pyridone ring, a pyrimidine ring, and a pyrazolopyridine ring.

[0039] According to some embodiments, ring C is selected from a benzene ring, a pyrazole ring, an imidazole ring, a thiazole ring, an oxazole ring, a dioxazole ring, a pyridine ring, a pyridone ring, a pyrimidine ring, and a pyrazolopyridine ring.

[0040] According to some embodiments, ring C is selected from a benzene ring, a pyrazole ring, a pyridine ring, a pyridone ring, a pyrimidine ring, and a pyrazolopyridine ring.

[0041] According to some embodiments, ring C is selected from a benzene ring, a pyrazole ring, a pyridine ring, a pyrimidine ring, and a pyrazolopyridine ring.

[0042] According to some embodiments, ring C is selected from

[0043] According to some embodiments, ring C is selected from

[0044] According to some embodiments, ring C is selected from

[0045] According to some embodiments, ring C is selected from

[0046] According to some embodiments, ring C is selected from

[0047] According to some embodiments, ring C is selected from

[0048] According to some embodiments, ring C is selected from

[0049] According to some embodiments, ring C is selected from

[0050] According to some embodiments, each R b The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R b1 Substituted with the following groups: OH, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 1-12 Alkoxy, C 1-12 Alkylthio, C3-12 Cycloalkyl, 3-14 membered heterocyclic group, C 6-14 Aryl, 5-14 membered heteroaryl, -N(R 31 )(R 32 )、-S(O)2R 33 、 Each R b1 the same or different, independently selected from OH, -N(R 34 )(R 35 ), CN, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, halogenated C1 -6 Alkyl, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group; R 31 、R 32 、R 33 、R 34 、R 35 、R 36 、R 37 、R 38 、R 39 The same or different, independently selected from H, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 3-12 Cycloalkyl.

[0051] According to some embodiments, each R b The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R b1 Substituted with the following groups: OH, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, C 3-6 Carbocyclic ring, 3-6 membered heterocyclic ring, -N(R 31 )(R 32 ), hydroxyl C 1-6 Alkyl, C 1-6 Alkyl-N(R 31 )(R 32 )、-S(O)2H、 Each R b1 the same or different, independently selected from OH, -N(R 34 )(R 35 ), CN, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, halogenated C 1-6 Alkyl or C 3-6 Cycloalkyl; R 31 、R 32 、R 34、R 35 、R 36 、R 37 、R 38 、R 39 The same or different, independently selected from H or C 1-6 alkyl.

[0052] According to some embodiments, each R b The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R b1 Substituted with the following groups: OH, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group, -N(R 31 )(R 32 ) or -S(O)2H; each R b1 the same or different, independently selected from OH, -N(R 34 )(R 35 ), CN, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, halogenated C 1-6 Alkyl or C 3-6 Cycloalkyl; R 31 、R 32 、R 34 、R 35 The same or different, independently selected from H or C 1-6 alkyl.

[0053] According to some embodiments, each R b the same or different, independently selected from CN, F, Cl, Br, CH3, CHF2, CF3, CH2CH3, CH2CF3, -C≡CCH3, CHOCH3, CH2CN, CH2NHCH3, CH(CH3)2, C(CH3)3, CH2CH2OH, C(CH3)2OH, NH2, NHCH3, N(CH3)2, CH2CH2N(CH3)2, OH, OCH3, OCF3, cyclopropyl, cyclobutyl, cyclopentyl, -S(O)2-cyclopropyl,

[0054] According to some embodiments, each R bthe same or different, independently selected from CN, F, Cl, Br, CH3, CHF2, CF3, CH2CH3, CH2CF3, -C≡CCH3, CHOCH3, CH2CN, CH2NHCH3, CH(CH3)2, C(CH3)3, CH2CH2OH, C(CH3)2OH, NH2, NHCH3, N(CH3)2, CH2CH2N(CH3)2, OH, OCH3, OCF3, cyclopropyl, cyclobutyl, cyclopentyl, -S(O)2-cyclopropyl,

[0055] According to some embodiments, each R b the same or different, independently selected from CN, F, Cl, Br, CH3, CHF2, CF3, CH2CH3, CH2CF3, -C≡CCH3, CHOCH3, CH2CN, CH2NHCH3, CH(CH3)2, C(CH3)3, CH2CH2OH, C(CH3)2OH, NH2, NHCH3, N(CH3)2, CH2CH2N(CH3)2, OH, OCH3, OCF3, cyclopropyl, cyclobutyl, cyclopentyl, -S(O)2-cyclopropyl,

[0056] According to some embodiments, each R b the same or different, independently selected from CN, F, Cl, Br, CH3, CHF2, CF3, CH2CH3, CH2CF3, -C≡CCH3, CH2OCH3, CH2CN, CH2NHCH3, CH(CH3)2, CH2CH2OH, C(CH3)2OH, NH2, NHCH3, N(CH3)2, CH2CH2N(CH3)2, OH, OCH3, OCF3, cyclopropyl, cyclobutyl, -S(O)2-cyclopropyl,

[0057] According to some embodiments, each R b the same or different, independently selected from CN, F, Cl, Br, CH3, CHF2, CF3, CH2CH3, CH2CF3, -C≡CCH3, CH2OCH3, CH2CN, CH2NHCH3, CH(CH3)2, CH2CH2OH, C(CH3)2OH, NH2, NHCH3, N(CH3)2, CH2CH2N(CH3)2, OH, OCH3, OCF3, cyclopropyl, cyclobutyl, -S(O)2-cyclopropyl,

[0058] According to some embodiments, each R b the same or different, independently selected from CN, F, Cl, Br, CH3, CHF2, CF3, CH2CH3, CH2CF3, CH2OCH3, CH2CN, CH2NHCH3, CH(CH3)2, C(CH3)2OH, NH2, NHCH3, N(CH3)2, OH, OCH3, OCF3, cyclopropyl, cyclobutyl, -S(O)2-cyclopropyl,

[0059] According to some embodiments, each R b the same or different, independently selected from CN, F, Cl, CH3, CHF2, CF3, CH2CH3, CH2CF3, CH2OCH3, CH2CN, CH2NHCH3, CH(CH3)2, C(CH3)2OH, NH2, NHCH3, OH, OCH3, OCF3, cyclobutyl, -S(O)2-cyclopropyl,

[0060] According to some embodiments, each R b the same or different, independently selected from CN, F, Cl, CH3, CHF2, CF3, CH2CH3, CH2CF3, CH2OCH3, CH2CN, CH2NHCH3, CH(CH3)2, C(CH3)2OH, NH2, NHCH3, OH, OCH3, OCF3, cyclobutyl, -S(O)2-cyclopropyl or

[0061] According to some embodiments, m2 is selected from 0, 1 or 2.

[0062] According to some embodiments, Selected from

[0063] According to some embodiments, Selected from

[0064] According to some embodiments, Selected from

[0065] According to some embodiments, Selected from

[0066] According to some embodiments, Selected from

[0067] According to some embodiments, Selected from

[0068] According to some embodiments, Selected from

[0069] According to some embodiments, Selected from

[0070] According to some embodiments, Selected from

[0071] According to some embodiments, ring A is selected from a benzene ring or a 5-6 membered heteroaromatic ring.

[0072] According to some embodiments, Ring A is selected from a 5-6 membered heteroaromatic ring.

[0073] According to some embodiments, ring A is selected from a benzene ring, a pyrazole ring, a triazole ring, a thiazole ring, an oxazole ring, a pyridine ring, a pyrimidine ring, a pyridone ring, a pyridazine ring, and a pyrazine ring.

[0074] According to some embodiments, ring A is selected from a benzene ring, a pyrazole ring, a triazole ring, a thiazole ring, an oxazole ring, and a pyridine ring.

[0075] According to some embodiments, ring A is selected from a pyrazole ring, a triazole ring, a thiazole ring, an oxazole ring, and a pyridine ring.

[0076] According to some embodiments, Ring A is selected from in It indicates that it is fused with ring B at this position.

[0077] According to some embodiments, Ring A is selected from in It indicates that it is fused with ring B at this position.

[0078] According to some embodiments, Ring A is selected from in It indicates that it is fused with ring B at this position.

[0079] According to some embodiments, Ring A is selected from in It indicates that it is fused with ring B at this position.

[0080] According to some embodiments, ring B is selected from a 5-membered heteroaromatic ring and a 5-8-membered heteroaromatic ring; and when ring A is a 5-membered heteroaromatic ring, ring B is a 5-membered heteroaromatic ring.

[0081] According to some embodiments, Ring B is a 5-membered heteroaryl ring.

[0082] According to some embodiments, ring B is selected from a thiazole ring, an isothiazole ring, an oxazole ring, a pyrazole ring, an imidazole ring, a triazole ring, a thiophene ring, a pyrrole ring, a furan ring, a dihydrofuran ring, a tetrahydrofuran ring, and an isoxazol-3-one ring.

[0083] According to some embodiments, ring B is selected from a thiazole ring, an oxazole ring, an imidazole ring, a thiophene ring, a pyrrole ring, and a furan ring.

[0084] According to some embodiments, ring B is selected from a thiazole ring, an oxazole ring, an imidazole ring, a thiophene ring, and a pyrrole ring.

[0085] According to some embodiments, Ring B is selected from in It indicates that it is fused with ring A at this position.

[0086] According to some embodiments, Ring B is selected from in It indicates that it is fused with ring A at this position.

[0087] According to some embodiments, Ring B is selected from in It indicates that it is fused with ring A at this position.

[0088] According to some embodiments, Ring B is selected from in It indicates that it is fused with ring A at this position.

[0089] According to some embodiments, Selected from

[0090] According to some embodiments, Selected from

[0091] According to some embodiments, Selected from

[0092] According to some embodiments, Selected from

[0093] According to some embodiments, Selected from

[0094] According to some embodiments, each R a The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: OH, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, halo C 1-6 Alkyl, C 1-6 Alkoxy, C 1-6 Alkylthio, C 3-6 Cycloalkyl, 3-8 membered heterocyclic group, phenyl, 5-6 membered heteroaryl, -N(R 11 )(R 12 ); or, two adjacent R a Together with the atoms to which they are attached, they form C 3-6 Cycloalkyl, 3-8 membered heterocyclic group, phenyl, 5-6 membered heteroaryl; each R a1 the same or different, independently selected from CN, halogen, OH, NH2, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group, C 6-10 Aryl, 5-10 membered heteroaryl, -N(R 14 )(R 15 );R 11 、R 12 、R 14 、R 15 The same or different, independently selected from H, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 3-6 Cycloalkyl.

[0095] According to some embodiments, each R a The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: OH, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, halo C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-8 membered heterocyclic group, phenyl, 5-6 membered heteroaryl, -N(R 11 )(R 12 ); or, two adjacent R a Together with the atoms to which they are attached, they form C 3-6 Cycloalkyl, 3-8 membered heterocyclic group, phenyl, 5-6 membered heteroaryl; each R a1 the same or different, independently selected from CN, halogen, OH, NH2, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group, C 6-10 Aryl, 5-10 membered heteroaryl, -N(R 14 )(R 15 );R 11 、R 12 、R 14 、R 15 The same or different, independently selected from H, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 3-6 Cycloalkyl.

[0096] According to some embodiments, each R a The same or different, independently selected from CN, F, Cl, Br, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: OH, C 1-4 Alkyl, C 2-3 Alkenyl, C 2-3 Alkynyl, halo C 1-4 Alkyl, C 1-4 Alkoxy, C 1-4 Alkylthio, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group, phenyl, 5-6 membered heteroaryl, -N(R 11 )(R 12 ); or two adjacent R a Together with the atoms to which they are attached, they form a benzene ring; each R a1are the same or different and are independently selected from CN, F, Cl, Br, OH, NH2, methyl, ethyl, isopropyl, trifluoromethyl, difluoromethyl; R 11 、R 12 are the same or different and are independently selected from H or methyl.

[0097] According to some embodiments, each R a The same or different, independently selected from CN, F, Cl, Br, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: OH, C 1-4 Alkyl, C 2-3 Alkenyl, C 2-3 Alkynyl, halo C 1-4 Alkyl, C 1-4 Alkoxy, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group, phenyl, 5-6 membered heteroaryl, -N(R 11 )(R 12 ); or two adjacent R a Together with the atoms to which they are attached, they form a benzene ring; each R a1 are the same or different and are independently selected from CN, F, Cl, Br, OH, NH2, methyl, ethyl, isopropyl, trifluoromethyl, difluoromethyl; R 11 、R 12 are the same or different and are independently selected from H or methyl.

[0098] According to some embodiments, each R a are the same or different and are independently selected from methyl, ethyl, isopropyl, tert-butyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, vinyl, propenyl, 1-fluorovinyl, 2-fluorovinyl, ethynyl, CN, F, Cl, Br, CD3, CH2OH, C(CH3)2OH, NH2, NHCH3, N(CH3)2, OCH3, OCH2CH3, OCH(CH3)2, OCF3, OCHF2, SCH3, CF3, CHF2, CF2CH3, CH2CF3, CH2CN, Tetrahydropyranyl (such as ), morpholinyl (such as ), phenyl (such as ), pyridyl (such as ), 3-fluoropyridyl (such as ), 3-chloropyridyl (such as ), 3-methylpyridyl (such as ); or two adjacent R a Together with the atoms to which they are attached, they form a benzene ring.

[0099] According to some embodiments, each R a are the same or different and are independently selected from methyl, ethyl, isopropyl, tert-butyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, vinyl, propenyl, 1-fluorovinyl, 2-fluorovinyl, ethynyl, CN, F, Cl, Br, C(CH3)2OH, NH2, NHCH3, N(CH3)2, OCH3, OCH2CH3, OCH(CH3)2, OCF3, OCHF2, SCH3, CF3, CHF2, CF2CH3, CH2CF3, CH2CN, Tetrahydropyranyl (such as ), morpholinyl (such as ), phenyl (such as ), pyridyl (such as ), 3-fluoropyridyl (such as ), 3-chloropyridyl (such as ), 3-methylpyridyl (such as ); or two adjacent R a Together with the atoms to which they are attached, they form a benzene ring.

[0100] According to some embodiments, each R a The same or different, independently selected from methyl, ethyl, isopropyl, tert-butyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, vinyl, propenyl, 1-fluorovinyl, 2-fluorovinyl, ethynyl, CN, F, Cl, C (CH 3 ) 2 OH, NH 2 , NHCH 3 , N (CH 3 ) 2 , OCH 3 , OCH 2 CH 3 , OCH (CH 3 ) 2 , OCF 3 , OCHF 2 , CF 3 , CHF 2 , tetrahydropyranyl (such as ), morpholinyl (such as ), phenyl (such as ), pyridyl (such as ), 3-fluoropyridyl (such as ), 3-chloropyridyl (such as ), 3-methylpyridyl (such as ); or two adjacent R a Together with the atoms to which they are attached, they form a benzene ring.

[0101] According to some embodiments, each R aThe same or different, independently selected from methyl, ethyl, isopropyl, tert-butyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, vinyl, propenyl, 1-fluorovinyl, ethynyl, CN, F, Cl, C (CH 3 ) 2 OH, NH 2 , NHCH 3 , N (CH 3 ) 2 , OCH 3 , OCH 2 CH 3 , OCH (CH 3 ) 2 , OCF 3 , OCHF 2 , CF 3 , CHF 2 , tetrahydropyranyl (such as ), morpholinyl (such as ), phenyl (such as ), pyridyl (such as ), 3-fluoropyridyl (such as ), 3-chloropyridyl (such as ), 3-methylpyridyl (such as ); or two adjacent R a Together with the atoms to which they are attached, they form a benzene ring.

[0102] According to some embodiments, m1 is selected from 0, 1 or 2.

[0103] According to some embodiments, m1 is selected from 0 or 1.

[0104] According to some embodiments, Selected from

[0105] According to some embodiments, Selected from

[0106] According to some embodiments, Selected from

[0107] According to some embodiments, Selected from

[0108] According to some embodiments, Selected from

[0109] According to some embodiments, Selected from

[0110] According to some embodiments, Selected from

[0111] According to some embodiments, the compound represented by formula (I) has the structure shown below:

[0112] Among them, ring A, ring B, ring C, Y1, R a 、R b 、R c , m1, m2, m3, n have the definitions as described in this article.

[0113] According to some embodiments, the compound represented by formula (I) has the structure shown below:

[0114] Among them, ring A, ring B, ring C, Y1, R a 、R b , m1, m2 have the definitions as described in this article.

[0115] According to some embodiments, the compound represented by formula (I) has the structure shown below:

[0116] Among them, R a , Rb, m1, m2 have the definitions as described herein.

[0117] According to some embodiments, the compound represented by formula (I) has the structure shown below:

[0118] Among them, R a Selected from halogen (such as F, Cl, Br), C 1-4 Alkyl (such as methyl, ethyl), halogenated C 1-4 Alkyl (e.g., trifluoromethyl, difluoromethyl);

[0119] R b , m2 have the definitions as described herein.

[0120] According to some embodiments, the compound represented by formula (I) is selected from the following structures:

[0121] The present invention also provides a method for preparing the compound represented by formula (I), comprising the following steps A:

[0122] Wherein, R' is H or an alkali metal ion (such as K + 、Li + );

[0123] Ring A, Ring B, Ring C, Y1, R a 、R b 、R c , m1, m2, m3, and n have the definitions described herein.

[0124] The present invention also provides a pharmaceutical composition comprising a therapeutically effective amount of at least one of the compound represented by formula (I), its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or prodrug compound thereof.

[0125] According to some embodiments, the pharmaceutical composition further comprises one or more pharmaceutically acceptable excipients.

[0126] According to some embodiments, the pharmaceutical composition may further contain one or more additional therapeutic agents.

[0127] The present invention also provides a method for treating or preventing a disease or condition caused by a phenylalanine hydroxylase mutation, comprising administering to a patient a preventive or therapeutically effective amount of at least one of the compounds represented by formula (I), its racemates, stereoisomers, tautomers, solvates, polymorphs, pharmaceutically acceptable salts, or prodrug compounds thereof.

[0128] The present invention also provides a method for treating or preventing diseases or conditions caused by phenylalanine hydroxylase mutation, comprising administering to a patient a preventive or therapeutically effective amount of the above-mentioned pharmaceutical composition.

[0129] According to some embodiments, the phenylalanine hydroxylase mutation contains at least one R408W, R261Q, R243Q, Y414C, L48S, A403V, ​​I65T, R241C, L348V, R408Q or V388M mutation.

[0130] According to some embodiments, the phenylalanine hydroxylase mutation contains at least one R408W, Y414C, 165T, F39L, R408Q, L348V, R261Q, A300S or L48S mutation.

[0131] According to some embodiments, the phenylalanine hydroxylase mutation contains at least one R408W mutation.

[0132] According to some embodiments, the phenylalanine hydroxylase mutation contains two R408W mutations.

[0133] According to some embodiments, the disease or disorder caused by the phenylalanine hydroxylase mutation is phenylketonuria.

[0134] According to some embodiments, the patient comprises a mammal, preferably a human.

[0135] The present invention also provides a compound represented by formula (I), its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or at least one of its prodrug compounds, or a pharmaceutical composition thereof for treating or preventing diseases or conditions caused by phenylalanine hydroxylase mutations.

[0136] According to some embodiments, the phenylalanine hydroxylase mutation contains at least one R408W, R261Q, R243Q, Y414C, L48S, A403V, ​​I65T, R241C, L348V, R408Q or V388M mutation.

[0137] According to some embodiments, the phenylalanine hydroxylase mutation contains at least one R408W, Y414C, 165T, F39L, R408Q, L348V, R261Q, A300S or L48S mutation.

[0138] According to some embodiments, the phenylalanine hydroxylase mutation contains at least one R408W mutation.

[0139] According to some embodiments, the phenylalanine hydroxylase mutation contains two R408W mutations.

[0140] According to some embodiments, the disease or disorder caused by the phenylalanine hydroxylase mutation is phenylketonuria.

[0141] The present invention also provides the use of at least one of the compound represented by formula (I), its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or prodrug compound thereof in the preparation of a drug.

[0142] According to some embodiments, the use may be use in preparing a medicament for treating or preventing a disease or disorder caused by a phenylalanine hydroxylase mutation.

[0143] According to some embodiments, the phenylalanine hydroxylase mutation contains at least one R408W, R261Q, R243Q, Y414C, L48S, A403V, ​​I65T, R241C, L348V, R408Q or V388M mutation.

[0144] According to some embodiments, the phenylalanine hydroxylase mutation contains at least one R408W, Y414C, 165T, F39L, R408Q, L348V, R261Q, A300S or L48S mutation.

[0145] According to some embodiments, the phenylalanine hydroxylase mutation contains at least one R408W mutation.

[0146] According to some embodiments, the phenylalanine hydroxylase mutation contains two R408W mutations.

[0147] According to some embodiments, the disease or disorder caused by the phenylalanine hydroxylase mutation is phenylketonuria. Beneficial effects

[0148] The compounds provided by the present invention can effectively restore the activity of phenylalanine hydroxylase mutant proteins and can be used to treat or prevent conditions and diseases associated with phenylalanine hydroxylase mutations. Through structural optimization, the series of compounds obtained in the present invention not only enhance their in vitro activity but also significantly improve their pharmacokinetic properties and / or increase their safety.

[0149] Definitions and Explanations of Terms

[0150] Unless otherwise indicated, the definitions of groups and terms in this specification and claims, including definitions used as examples, exemplary definitions, preferred definitions, definitions in tables, and definitions of specific compounds in the Examples, may be arbitrarily combined and coupled with one another. The group definitions and compound structures resulting from such combinations and couplings should be understood to be within the scope of this specification and / or claims.

[0151] The term "optional" (or "optionally", "optionally") in the general formula definitions of this application means the situation of being substituted by zero, one or more substituents, for example, "optionally substituted by one, two or more R" means that it may not be substituted by R (unsubstituted) or may be optionally substituted by one, two or more R.

[0152] "More" means three or more, for example, 3, 4, 5, 6, 7, 8, 9 or 10.

[0153] Unless otherwise indicated, numerical ranges recited in this specification and claims are equivalent to reciting at least each specific integer value therein. For example, the numerical range "1-12" is equivalent to reciting each integer value in the numerical range "1-12", namely, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and 12.

[0154] The term "C 1-12 "Alkyl" is understood to mean straight-chain and branched alkyl groups having 1 to 12 carbon atoms, "C 1-8 "Alkyl" means straight chain and branched chain alkyl groups having 1, 2, 3, 4, 5, 6, 7, or 8 carbon atoms, "C 1-6 The term "alkyl" refers to straight-chain and branched alkyl groups having 1, 2, 3, 4, 5 or 6 carbon atoms. The alkyl group is, for example, methyl, ethyl, propyl, butyl, pentyl, hexyl, isopropyl, isobutyl, sec-butyl, tert-butyl, isopentyl, 2-methylbutyl, 1-methylbutyl, 1-ethylpropyl, 1,2-dimethylpropyl, neopentyl, 1,1-dimethylpropyl, 4-methylpentyl, 3-methylpentyl, 2-methylpentyl, 1-methylpentyl, 2-ethylbutyl, 1-ethylbutyl, 3,3-dimethylbutyl, 2,2-dimethylbutyl, 1,1-dimethylbutyl, 2,3-dimethylbutyl, 1,3-dimethylbutyl or 1,2-dimethylbutyl, or the like or isomers thereof.

[0155] The term "C 2-12 "Alkenyl" is understood to mean a linear or branched monovalent hydrocarbon radical having 1 to 12 carbon atoms, containing one or more double bonds and having 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms. For example, a radical having 2, 3, 4, 5, 6, 7 or 8 carbon atoms (i.e., C 2-8 alkenyl), for example, having 2, 3, 4, 5, or 6 carbon atoms (i.e., C 2-6 alkenyl), having 2 or 3 carbon atoms (i.e., C 2-3It is understood that when the alkenyl group contains more than one double bond, the double bonds may be separated from one another or conjugated. The alkenyl group is, for example, vinyl, allyl, (E)-2-methylvinyl, (Z)-2-methylvinyl, (E)-but-2-enyl, (Z)-but-2-enyl, (E)-but-1-enyl, (Z)-but-1-enyl, pent-4-enyl, (E)-pent-3-enyl, (Z)-pent-3-enyl, (E)-pent-2-enyl, (Z)-pent-2-enyl, (E)- Pent-1-enyl, (Z)-pent-1-enyl, hex-5-enyl, (E)-hex-4-enyl, (Z)-hex-4-enyl, (E)-hex-3-enyl, (Z)-hex-3-enyl, (E)-hex-2-enyl, (Z)-hex-2-enyl, (E)-hex-1-enyl, (Z)-hex-1-enyl, isopropenyl, 2-methylprop-2-enyl, 1-methylprop-2-enyl , 2-methylprop-1-enyl, (E)-1-methylprop-1-enyl, (Z)-1-methylprop-1-enyl, 3-methylbut-3-enyl, 2-methylbut-3-enyl, 1-methylbut-3-enyl, 3-methylbut-2-enyl, (E)-2-methylbut-2-enyl, (Z)-2-methylbut-2-enyl, (E)-1-methylbut-2-enyl, (Z)-1-methyl But-2-enyl, (E)-3-methylbut-1-enyl, (Z)-3-methylbut-1-enyl, (E)-2-methylbut-1-enyl, (Z)-2-methylbut-1-enyl, (E)-1-methylbut-1-enyl, (Z)-1-methylbut-1-enyl, 1,1-dimethylprop-2-enyl, 1-ethylprop-1-enyl, 1-propylvinyl, 1-isopropylvinyl.

[0156] The term "C 2-12 “Alkynyl” is understood to mean a straight or branched monovalent hydrocarbon radical having 1 to 12 carbon atoms, containing one or more triple bonds and having 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms, for example, having 2, 3, 4, 5, 6, 7 or 8 carbon atoms (i.e., “C 2-8 alkynyl”), having 2, 3, 4, 5, or 6 carbon atoms (i.e., “C 2-6 Alkynyl”), having 2 or 3 carbon atoms (“C 2-3The alkynyl group is, for example, ethynyl, prop-1-ynyl, prop-2-ynyl, but-1-ynyl, but-2-ynyl, but-3-ynyl, pent-1-ynyl, pent-2-ynyl, pent-3-ynyl, pent-4-ynyl, hex-1-ynyl, hex-2-ynyl, hex-3-ynyl, hex-4-ynyl, hex-5-ynyl, 1-methylprop-2-ynyl, 2-methylbut-3-ynyl, 1-methylbut-3-ynyl, 1-methylbut-2-ynyl, 3-methylbut-1-ynyl, 1-ethylprop-2-ynyl, 3-methylpent-4-ynyl, 2-methylpent-4-ynyl, 1-methylpent-4-ynyl, In some embodiments, the alkynyl group is ethynyl, prop-1-ynyl or prop-2-ynyl.

[0157] The term "C 3-12 "Cycloalkyl" is understood to mean a saturated monovalent monocyclic, bicyclic (such as condensed, bridged, spiro) hydrocarbon ring or tricyclic alkane having 3 to 12 carbon atoms, preferably "C 3-10 Cycloalkyl", more preferably "C 3-8 Cycloalkyl". The term "C 3-12 "Cycloalkyl" is understood to mean a saturated monovalent monocyclic, bicyclic (eg bridged, spiro) hydrocarbon ring or tricyclic alkane having 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms. 3-12 The cycloalkyl group may be a monocyclic hydrocarbon group such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl or cyclodecyl, or a bicyclic hydrocarbon group such as borneol, indolyl, hexahydroindolyl, tetrahydronaphthyl, decahydronaphthyl, bicyclo[2.1.1]hexyl, bicyclo[2.2.1]heptyl, bicyclo[2.2.1]heptenyl, 6,6-dimethylbicyclo[3.1.1]heptyl, 2,6,6-trimethylbicyclo[3.1.1]heptyl, bicyclo[2.2.2]octyl, 2,7-diazaspiro[3,5]nonyl, 2,6-diazaspiro[3,4]octyl, or a tricyclic hydrocarbon group such as adamantyl.

[0158] The term "C 6-14 "Aryl" should be understood to mean preferably a monovalent aromatic or partially aromatic monocyclic, bicyclic (such as fused, bridged, spiro) or tricyclic hydrocarbon ring having 6 to 14 carbon atoms, which can be a single aromatic ring or a polyaromatic ring fused together, preferably "C6-10 Aryl". The term "C 6-14 "Aryl" is understood to mean preferably a monovalent aromatic or partially aromatic monocyclic, bicyclic or tricyclic hydrocarbon ring ("C 6-14 or a ring having 9 carbon atoms ("C9 aryl"), for example indanyl or indenyl, or a ring having 10 carbon atoms ("C 10 aryl) such as tetrahydronaphthyl, dihydronaphthyl or naphthyl, or a ring having 13 carbon atoms ("C 13 aryl), such as fluorenyl, or a ring having 14 carbon atoms ("C 14 aryl”), such as anthracenyl. When the C 6-20 When the aryl group is substituted, it may be monosubstituted or polysubstituted. Furthermore, there is no limitation on the position of substitution, and for example, substitution may be at the ortho, para or meta position.

[0159] The term "5-14 membered heteroaryl" is understood to include monovalent monocyclic, bicyclic (e.g. fused, bridged, spiro) or tricyclic aromatic ring systems having 5 to 14 ring atoms and containing 1 to 5 heteroatoms independently selected from N, O and S, for example "5-10 membered heteroaryl". The term "5-14 membered heteroaryl" is understood to include monovalent monocyclic, bicyclic or tricyclic aromatic ring systems having 5, 6, 7, 8, 9, 10, 11, 12, 13 or 14 ring atoms, in particular 5 or 6 or 9 or 10 carbon atoms, and containing 1 to 5, preferably 1 to 3, heteroatoms each independently selected from N, O and S and, in each case, may additionally be benzofused. "Heteroaryl" also refers to a radical in which a heteroaromatic ring is fused to one or more aryl, alicyclic or heterocyclyl rings, wherein the radical or point of attachment is on the heteroaromatic ring. Non-limiting examples include 1-, 2-, 3-, 5-, 6-, 7-, or 8-indolizinyl, 1-, 3-, 4-, 5-, 6-, or 7-isoindolyl, 2-, 3-, 4-, 5-, 6-, or 7-indolyl, 2-, 3-, 4-, 5-, 6-, or 7-indazolyl, 2-, 4-, 5-, 6-, 7-, or 8-purinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8-, or 9-quinolizinyl, 2-, 3-, 4-, 5-, 6-, 7-, or 8-quinolinyl, 1-, 3-, 4- 1-, 2-, 3-, 4-, 5-, 6-, 7-, or 8-isoquinolyl, 1-, 4-, 5-, 6-, 7-, or 8-phthalazinyl, 2-, 3-, 4-, 5-, or 6-naphthyridinyl, 2-, 3-, 5-, 6-, 7-, or 8-quinazolinyl, 3-, 4-, 5-, 6-, 7-, or 8-cinnolinyl, 2-, 4-, 6-, or 7-pteridinyl, 1-, 2-, 3-, 4-, 5-, 6-, 7-, or 8-4aHcarbazolyl, 1-, 2-, 3-, 4-, 5-, 6- , 7- or 8-carbazolyl, 1-, 3-, 4-, 5-, 6-, 7-, 8- or 9-carbolinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8-, 9- or 10-phenanthridinyl, 1-, 2-, 3-, 4-, 5-, 6-, 7-, 8- or 9-acridinyl, 1-, 2-, 4-, 5-, 6-, 7-, 8- or 9-pyridinyl, 2-, 3-, 4-, 5-, 6-, 8-, 9- or 10-phenanthrolinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8- or 9- phenazinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8-, 9- or 10-phenothiazinyl, 1-, 2-, 3-, 4-, 6-, 7-, 8-, 9- or 10-phenazinyl, 2-, 3-, 4-, 5-, 6- or 1-, 3-, 4-, 5-, 6-, 7-, 8-, 9- or 10-benzoisoquinolinyl, 2-, 3-, 4- or thieno[2,3-b]furanyl, 2-, 3-, 5-, 6-, 7-, 8-, 9-, 10- or 11-7H-pyrazino[2,3-c]carbazolyl, 2-, 3-, 5-, 6- or 7-2H-furo[3,2-b]-pyranyl, 2-, 3-, 4-, 5-, 7- or 8-5H-pyrido[2,3-d]-o-oxazinyl, 1-, 3- or 5-1H-pyrazolo[4,3-d]oxazolyl, 2-, 4- or 54H-imidazo[4,5-d]thiazolyl, 3-, 5- or 8-pyrazino[2,3-d]pyridazinyl, 2-, 3-, 5- or 6-imidazo[2,1-b]thiazolyl, 1-, 3-, 6-, 7-, 8- or 9-furo[3,4-c]cinnolinyl, 1-, 2-, 3-, 4-, 5-, 6-, 8-, 9-, 10 or 11-4H-pyrido[2,3

[0015] In some embodiments, the present invention further comprises carbazolyl, 2-, 3-, 6-, or 7-imidazo[1,2-b][1,2,4]triazinyl, 7-benzo[b]thienyl, 2-, 4-, 5-, 6-, or 7-benzoxazolyl, 2-, 4-, 5-, 6-, or 7-benzimidazolyl, 2-, 4-, 4-, 5-, 6-, or 7-benzothiazolyl, 1-, 2-, 4-, 5-, 6-, 7-, 8-, or 9-benzoxapinyl, 2-, 4-, 5-, 6-, 7-, or 8-benzoxazinyl, 1-, 2-, 3-, 5-, 6-, 7-, 8-, 9-, 10-, or 11-4H-pyrrolo[1,2-b][2]benzazepinyl. Typical fused heteroaryl groups include, but are not limited to, 2-, 3-, 4-, 5-, 6-, 7-, or 8-quinolyl, 1-, 3-, 4-, 5-, 6-, 7-, or 8-isoquinolyl, 2-, 3-, 4-, 5-, 6-, or 7-indolyl, 2-, 3-, 4-, 5-, 6-, or 7-benzo[b]thienyl, 2-, 4-, 5-, 6-, or 7-benzoxazolyl, 2-, 4-, 5-, 6-, or 7-benzimidazolyl, and 2-, 4-, 5-, 6-, or 7-benzothiazolyl. When the 5- to 14-membered heteroaryl group is linked to other groups to form a compound of the present invention, the carbon atoms on the 5- to 14-membered heteroaryl ring may be linked to the other groups, or heteroatoms on the 5- to 14-membered heteroaryl ring may be linked to the other groups. When the 5- to 14-membered heteroaryl group is substituted, it may be monosubstituted or polysubstituted. Furthermore, there is no limitation on the substitution site, for example, a hydrogen atom connected to a carbon atom on a heteroaryl ring may be substituted, or a hydrogen atom connected to a heteroatom on a heteroaryl ring may be substituted.

[0160] The term "carbocycle" refers to a saturated or unsaturated non-aromatic monocyclic or polycyclic (such as bicyclic) hydrocarbon ring (for example, a monocyclic ring such as a cyclopropane ring, a cyclobutane ring, a cyclopentane ring, a cyclohexane ring, a cycloheptane ring, a cyclooctane ring, a cyclononane ring, or a bicyclic ring, including a spirocyclic, fused or bridged system (such as a bicyclo[11.1]pentane ring, a bicyclo[2.2.1]heptane ring, a bicyclo[3.2.1]octane ring or a bicyclo[5.2.0]nonane ring, a tertiary naphthalene ring, etc.), which may be optionally substituted with one or more (such as one, two or three) suitable substituents. The term "3-6 membered carbocycle" refers to a carbocycle containing 3, 4, 5 or 6 ring-forming carbon atoms.

[0161] Unless otherwise defined, the term "3-14 membered heterocyclyl" refers to a saturated or unsaturated non-aromatic ring or ring system, for example, a 4-, 5-, 6- or 7-membered monocyclic ring, a 7-, 8-, 9-, 10-, 11- or 12-membered bicyclic ring (such as a fused ring, a bridged ring, a spirocyclic ring) or a 10-, 11-, 12-, 13- or 14-membered tricyclic ring system, and contains at least one, for example 1, 2, 3, 4, 5 or more heteroatoms selected from O, S and N, wherein N and S may also be optionally oxidized to various oxidation states to form nitrogen oxides, -S(O)- or -S(O)2- states. For example, the "3-14 membered heterocyclyl" may be a 3-14 membered N-containing heterocyclyl (containing at least one N). Preferably, the heterocyclyl may be selected from a "3-10 membered heterocyclyl". The term "3-10 membered heterocyclyl" means a saturated or unsaturated non-aromatic ring or ring system, and contains at least one heteroatom selected from O, S and N. The heterocyclyl can be connected to the rest of the molecule through any one of the carbon atoms or the nitrogen atom (if present). The heterocyclyl can include fused or bridged rings and spirocyclic rings. In particular, the heterocyclyl can include, but is not limited to, 4-membered rings such as azetidinyl, oxetane; 5-membered rings such as tetrahydrofuranyl, dioxolyl, pyrrolidinyl, imidazolidinyl, pyrazolidinyl, pyrrolinyl; or 6-membered rings such as tetrahydropyranyl, piperidinyl, morpholinyl, dithianyl, thiomorpholinyl, piperazinyl or trithianyl; or 7-membered rings such as diazepanyl. Optionally, the heterocyclyl can be benzo-fused. The heterocyclic group may be bicyclic, such as, but not limited to, a 5,5-membered ring, such as a hexahydrocyclopenta[c]pyrrol-2(1H)-yl ring, or a 5,6-membered bicyclic ring, such as a hexahydropyrrolo[1,2-a]pyrazin-2(1H)-yl ring. The heterocyclic group may be partially unsaturated, i.e., it may contain one or more double bonds, such as, but not limited to, dihydrofuranyl, dihydropyranyl, 2,5-dihydro-1H-pyrrolyl, 4H-[1,3,4]thiadiazinyl, 1,2,3,5-tetrahydrooxazolyl, or 4H-[1,4]thiazinyl, or it may be benzo-fused, such as, but not limited to, dihydroisoquinolinyl. When the 3-14-membered heterocyclic group is linked to other groups to form the compounds of the present invention, the linking may be to a carbon atom of the 3-14-membered heterocyclic group or to a heterocyclic atom on the 3-14-membered heterocyclic group ring. For example, when the 3-14 membered heterocyclic group is selected from piperazinyl, the nitrogen atom on the piperazinyl group may be connected to the other group. Or when the 3-14 membered heterocyclic group is selected from piperidinyl, the nitrogen atom on the piperidinyl ring and the carbon atom at the para position thereof may be connected to the other group.

[0162] The term "halogen" refers to fluorine, chlorine, bromine and iodine.

[0163] The term "nitrogen oxide" refers to a compound formed by oxidation of a nitrogen atom in a tertiary amine or nitrogen-containing (aromatic) heterocyclic compound structure.

[0164] The term "spirocyclic" refers to a ring system in which two rings share one ring atom.

[0165] The term "fused ring" refers to a ring system in which two rings share two ring atoms.

[0166] The term "bridged ring" refers to a ring system in which two rings share three or more ring atoms.

[0167] Unless otherwise specified, a heterocyclyl, heterocyclylene, heteroaryl, or heteroarylene group includes all possible isomeric forms thereof, such as positional isomers thereof. Thus, for some illustrative, non-limiting examples, 1-, 2-, 3-, 4-, 5-, 6-, 7-, 8-, 9-, 10-, 11-, 12-, etc. (if present) substituted at one, two, or more positions thereof or bonded to other groups include pyridin-2-yl, pyridin-2-ylene, pyridin-3-yl, pyridin-3-ylene, pyridin-4-ylene, and pyridin-4-ylene; thienyl or thienylene group includes thien-2-yl, thien-2-ylene, thien-3-ylene, and thien-3-ylene; pyrazol-1-yl, pyrazol-3-yl, pyrazol-4-yl, and pyrazol-5-yl.

[0168] The compounds of the present disclosure may 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 a structural isomer that exists in equilibrium and is easily converted from one isomeric form to another isomeric form. It includes all possible tautomers, i.e., existing in the form of a single isomer or in the form of a mixture of any proportions of the tautomers. Non-limiting examples include: keto-enol, imine-enamine, lactam-lactim, etc.

[0169] "Halo" means substituted with one or more halogens.

[0170] The term "haloalkyl" refers to an alkyl group substituted with one or more halogens, wherein alkyl is as defined above.

[0171] The term "oxo" refers to a substituent in which a carbon atom, a nitrogen atom, or a sulfur atom is oxidized to form an oxy group (=O).

[0172] The term "alkylamino" refers to -NH-(alkyl) or -N-(alkyl)2, wherein alkyl is as defined above. Non-limiting examples of alkylamino include methylamino, ethylamino, propylamino, isopropylamino, butylamino, dimethylamino, methylethylamino, diethylamino, dipropylamino, methylpropylamino, diisopropylamino, dibutylamino, and the like.

[0173] "Hydroxyalkyl" refers to an alkyl group substituted with one or more hydroxy groups, wherein alkyl is as defined above. Non-limiting examples of hydroxyalkyl groups include hydroxymethyl, hydroxyethyl, hydroxypropyl, hydroxymethylpropyl, dihydroxypropyl, and the like.

[0174] The term "alkyloxy" refers to -O-(alkyl), wherein alkyl is as defined above. Non-limiting examples of alkoxy include: methoxy, ethoxy, propoxy, butoxy. Alkoxy groups may be optionally substituted or unsubstituted, and when substituted, the substituents are preferably one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkyloxy, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, or heterocycloalkyloxy.

[0175] The terms "alkyleneoxy" and "oxyalkylene" refer to -alkylene-O- or -O-alkylene-, where alkylene represents a linear or branched saturated divalent hydrocarbon radical. The definition of "alkyl" with respect to the number of carbon atoms in "alkylene" applies as defined above. It will be understood by those skilled in the art that an alkyleneoxy or oxyalkylene group can be attached to the remainder of the molecule in which it is contained in any orientation, i.e., the two terms are used interchangeably.

[0176] Wavy lines intersecting chemical bonds Used to indicate the connection position between a group and other atoms in the molecular structure. Indicates connection with the 3-position of pyridyl. When the group connection position is not fixed, taking pyridyl as an example, The method is shown in FIG. 1 , which indicates that the pyridyl group can be connected to any position that can be connected. It means that it can be connected to any connectable position on the heteroaryl ring, for example, it can be connected to any of the four carbon atoms on the pyridine ring on the right side of the heteroaryl group, or it can be connected to a carbon atom on the pyrazole ring on the left side. Unless otherwise specified, similar expressions in this application are interpreted the same as above.

[0177] In the chemical structure of the compound of the present invention, the bond Indicates that the configuration is not specified. Indicates the absolute configuration, that is, if there are stereoisomers in the chemical structure, the bond Can be or include both Two configurations.

[0178] In the present invention, the compounds referred to also include isotopically labeled compounds, which are the same as those shown in Formula I, but in which one or more atoms are replaced by atoms 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 invention include isotopes of H, C, N, O, S, F, and Cl, such as 2 H. 3 H. 13 C. 11 C. 14 C. 15 N. 18 O. 17 O. 32 P. 35 S. 18 F and 36 Cl. Compounds of the invention, prodrugs thereof, or pharmaceutically acceptable salts of said compounds or prodrugs containing the aforementioned isotopes and / or other isotopes of other atoms are within the scope of the invention. Certain isotopically labeled compounds of the invention, for example, those incorporating radioactive isotopes (such as 3 H and 14 C) compounds can be used in drug and / or substrate tissue distribution assays. 3 H) and carbon 14 (i.e. 14 C) isotopes are particularly preferred due to their ease of preparation and detectability. 2 H or D) substitution may provide certain therapeutic advantages (e.g., increased in vivo half-life or reduced dosage requirements) derived from greater metabolic stability and may therefore be preferred in certain circumstances. The presence of hydrogen in a substituent of the present invention without the separate listing of the term deuterium or tritium does not exclude deuterium or tritium, but rather may also include deuterium or tritium.

[0179] It will be appreciated by those skilled in the art that the compounds of formula (I) may exist in the form of various pharmaceutically acceptable salts. If these compounds have a basic center, they may form acid addition salts; if these compounds have an acidic center, they may form base addition salts; if these compounds contain both an acidic center (e.g., a carboxyl group) and a basic center (e.g., an amino group), they may also form internal salts.

[0180] The compounds of the present invention may exist in the form of solvates (e.g., hydrates), wherein the compounds of the present invention contain a polar solvent as a structural element of the crystal lattice of the compound, in particular water, methanol or ethanol. The amount of polar solvent, in particular water, may be present in a stoichiometric or non-stoichiometric ratio.

[0181] Depending on their molecular structure, the compounds of the present invention may be chiral and therefore may exist in various enantiomeric forms. Thus, these compounds may exist in racemic or optically active forms. The compounds of the present invention encompass isomers or mixtures thereof, racemates, in which each chiral carbon is in the R or S configuration. The compounds of the present invention or their intermediates can be separated into enantiomeric compounds by chemical or physical methods well known to those skilled in the art, or used in this form for synthesis. In the case of racemic amines, diastereomers are prepared from the mixture by reaction with an optically active resolving agent. Examples of suitable resolving agents are optically active acids, such as R and S forms of tartaric acid, diacetyltartaric acid, dibenzoyltartaric acid, mandelic acid, malic acid, lactic acid, appropriate N-protected amino acids (e.g., N-benzoylproline or N-phenylsulfonylproline) or various optically active camphorsulfonic acids. Chromatographic enantiomer resolution can also be advantageously performed with the aid of optically active resolving agents (e.g., dinitrobenzoylphenylglycine, cellulose triacetate or other carbohydrate derivatives or chirally derivatized methacrylate polymers immobilized on silica gel). Suitable eluents for this purpose are aqueous or alcoholic solvent mixtures, e.g., hexane / isopropanol / acetonitrile.

[0182] The corresponding stable isomers can be separated according to known methods, for example by extraction, filtration or column chromatography.

[0183] The term "patient" refers to any animal including mammals, preferably mice, rats, other rodents, rabbits, dogs, cats, pigs, cows, sheep, horses or primates, and most preferably humans.

[0184] The term "therapeutically effective amount" refers to that amount of an active compound or drug that will elicit the biological or medical response that a researcher, veterinarian, physician, or other clinician is seeking in a tissue, system, animal, individual, or human, and includes one or more of the following: (1) prevents disease, e.g., prevents a disease, disorder, or condition in an individual who is susceptible to the disease, disorder, or condition but who is not yet experiencing or developing the pathology or symptoms of the disease. (2) inhibits disease, e.g., inhibits the disease, disorder, or condition (i.e., prevents further development of the pathology and / or symptoms) in an individual who is experiencing or developing the pathology or symptoms of the disease, disorder, or condition. (3) alleviates disease, e.g., alleviates the disease, disorder, or condition (i.e., reverses the pathology and / or symptoms) in an individual who is experiencing or developing the pathology or symptoms of the disease, disorder, or condition. DETAILED DESCRIPTION

[0185] The technical solutions of the present invention will be described in further detail below with reference to specific embodiments. It should be understood that the following embodiments are merely illustrative and explanations of the present invention and should not be construed as limiting the scope of protection of the present invention. All technologies implemented based on the above content of the present invention are encompassed within the scope of protection that the present invention is intended to protect.

[0186] Unless otherwise specified, the raw materials and reagents used in the following examples are commercially available or can be prepared by known methods.

[0187] The structures of the compounds of the present invention are determined by nuclear magnetic resonance (NMR) and / or liquid chromatography-mass spectrometry (LC-MS). NMR chemical shifts (δ) are given in parts per million (ppm). NMR measurements are performed using a Bruker AVANCE-400 NMR spectrometer, using deuterated dimethyl sulfoxide (DMSO-d6), deuterated methanol (CD3OD), or deuterated chloroform (CDCl3) as the solvent, and tetramethylsilane (TMS) as the internal standard.

[0188] Liquid chromatography-mass spectrometry (LC-MS) was performed using an Agilent 1200 Infinity Series mass spectrometer. HPLC was performed using an Agilent 1200DAD high-pressure liquid chromatograph (Sunfire C18 150×4.6 mm column) and a Waters 2695-2996 high-pressure liquid chromatograph (Gimini C18 150×4.6 mm column).

[0189] Thin-layer chromatography silica gel plates use Yantai Huanghai HSGF254 or Qingdao Ocean Chemical GF254 silica gel plates. The specifications used for TLC are 0.15mm-0.20mm, and the specifications used for thin-layer chromatography separation and purification products are 0.4mm-0.5mm. Column chromatography generally uses Yantai Huanghai 200-300 mesh silica gel as the carrier.

[0190] Unless otherwise stated, the raw materials and reagents used in the following examples are commercially available or can be prepared by known methods. Unless otherwise stated, all reactions of the present invention were carried out under a dry nitrogen or argon atmosphere with continuous magnetic stirring, with dry solvents, and reaction temperatures are reported in degrees Celsius.

[0191] Preparation of intermediate INT 1

[0192] Note: "*" indicates that the carbon atom at the position is chiral carbon and the configuration is one of R or S, for example express One of them.

[0193] Step 1: Synthesis of compound INT 1-1

[0194] To a solution of 3-methyl-6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[3,2-c]pyridine (compound 103-5, 8.6 g, 31.81 mmol) in dichloromethane (100 mL) were added di-tert-butyl dicarbonate (22 mL, 95.43 mmol) and triethylamine (8.82 mL, 63.62 mmol). The mixture was reacted at room temperature for 2 hours. After the reaction was completed, water (500 mL) was added to the reaction solution, and ethyl acetate (100 mL×3) was added for extraction. The organic phase was washed with saturated brine (100 mL×3), dried over anhydrous sodium sulfate, filtered and concentrated to obtain a crude product, which was then purified by normal phase purification (petroleum ether:ethyl acetate=5:1) to obtain compound INT 1-1 (13.3 g, 88.85%).

[0195] LCMS: (ESI, m / z): 471.4 [M+H] + .

[0196] Separation by SFC (system: Waters SFC 150; column name: Column size: 250*50mm 10μm; mobile phase A: supercritical CO2, mobile phase B: ethanol (+0.1% 7.0mol / L amine methanol solution); wavelength: 214nm; flow rate: 140mL / min; temperature: RT; injection volume: 1.5mL; cycle time: 8.26min; single injection time: 15min; solvent: ethanol: distillation grade; supercritical CO2: food grade) were separated to obtain INT 1-1A (5.5g, 41.35%, front peak, elution time: 10.68min) and INT 1-1B (back peak, elution time: 12.82min).

[0197] Step 2: Synthesis of compound INT 1

[0198] Compound INT 1-1A (5.5 g, 11.69 mmol) was dissolved in a hydrogen chloride 1,4-dioxane (60 mL) solution and reacted at room temperature for 12 hours. After the reaction was completed, the reaction solution was concentrated to obtain compound INT 1 (4.1 g, 114.34%).

[0199] LCMS: (ESI, m / z): 271.0 [M+H] + .

[0200] Preparation of intermediate INT 2

[0201] Note: "*" indicates that the carbon atom at the position is chiral carbon and the configuration is one of R or S, for example express One of them.

[0202] Step 1: Synthesis of compound INT 2-1

[0203] To a solution of 3-chloro-6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[3,2-c]pyridine (compound 105-3, 6.4 g, 22.01 mmol, 1 eq) in dichloromethane (60 mL) were added di-tert-butyl dicarbonate (14.41 g, 66.03 mmol, 3 eq) and triethylamine (6.68 g, 66.03 mmol, 3 eq). The reaction was stirred at room temperature for 16 hours. After the reaction was completed, the mixture was concentrated to give a crude product, which was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 1:1) to give compound INT 2-1 (8 g, 74.02%).

[0204] LCMS: (ESI, m / z): 491.1 [M+H] + .

[0205] Separation by SFC (system: Waters SFC 150; column name: Column size: 250*40mm 10μm; mobile phase A: supercritical CO2, mobile phase B: ethanol (+0.1% 7.0mol / L amine methanol solution); A:B=45:55; wavelength: 214nm; flow rate: 145mL / min; temperature: RT; injection volume: 5mL; cycle time: 9.11min; solvent: ethanol: distillation grade, supercritical CO2: food grade) to obtain INT 2-1A (2.8g, 40%, front peak, elution time: 6.37min) and INT 2-1B (2.8g, 40%, back peak, elution time: 11.15min).

[0206] Step 2: Synthesis of compound INT 2

[0207] To INT 2-1A (2.8 g, 5.70 mmol) was added dioxane hydrochloride solution (50 mL, 4 M), and the reaction was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was concentrated to obtain compound INT 2 (2.8 g, 100%).

[0208] LCMS: (ESI, m / z): 291.0 [M+H] + .

[0209] Preparation of intermediate INT 3

[0210] Note: "*" indicates that the carbon atom at the position is chiral carbon and the configuration is one of R or S, for example express One of them.

[0211] Based on the similar steps to synthesize compound INT 2, compound INT 3 was obtained.

[0212] Example 1

[0213] Step 1: Synthesis of Compound 001-2

[0214] Under nitrogen protection, to a dioxane solution (100 mL) of 2-bromo-1,3-thiazole (compound 001-1, 10 g, 60.97 mmol, 1 eq) was added prop-2-yn-1-yl acetate (8.37 g, 85.36 mmol, 1.4 eq), cuprous iodide (0.58 g, 3.05 mmol, 0.05 eq), triethylamine (18.51 g, 182.91 mmol, 3 eq), bis(triphenylphosphine) dichloride. Palladium (II) (2.14 g, 3.05 mmol, 0.05 eq) was added and the system was stirred at 50°C for 16 hours. After the reaction was completed, the reaction solution was filtered through a pad of celite, the filtrate was collected and added with water (300 mL), extracted with ethyl acetate (200 mL × 3), dried over anhydrous sodium sulfate, filtered, and dried to give a crude product. The crude product was purified by silica gel column chromatography (ethyl acetate / petroleum ether (0-30%)) to give compound 001-2 (5.9 g, 53.4%).

[0215] LCMS: (ESI, m / z): 182.1 [M+H] + .

[0216] Step 2: Synthesis of Compound 001-3

[0217] To a solution of 3-(1,3-thiazol-2-yl)prop-2-yn-1-yl acetate (compound 1-2, 3 g, 16.56 mmol, 1 eq) in tetrahydrofuran (30 mL) and water (15 mL) was slowly added lithium hydroxide hydrate (0.73 g, 17.39 mmol, 1.05 eq) at 0°C. The mixture was stirred at room temperature for 1 hour. After the reaction was complete, water (80 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (2×80 mL). The organic phase was collected, washed with water (100 mL) and brine (100 mL), dried over anhydrous sodium sulfate, filtered, and dried to give a crude product 001-3 (2 g). The crude product was used directly in the next step.

[0218] 1H NMR (400MHz, CDCl3) δ7.83 (d, J = 3.2Hz, 1H), 7.36 (d, J = 3.2Hz, 1H), 4.55 (s, 2H).

[0219] Step 3: Synthesis of Compound 001-4

[0220] To a solution of water (7.39 mL, 410.18 mmol, 17.3 eq) and trifluoroacetic acid (43.54 mL, 569.04 mmol, 24 eq) was added (E)-(N-[(2,4,6-trimethylbenzenesulfonyl)oxy]ethylcarboximidate) (7.10 g, 24.90 mmol, 1.05 eq) at 0°C. After the addition was complete, the system was stirred at 0°C for 2 hours. After the reaction was complete, water (80 mL) was added at 0°C. A solid precipitated, which was filtered and rinsed twice with water (20 mL). The upper solid was dissolved in dichloromethane (40 mL), dried over anhydrous sodium sulfate, and the filtrate was collected by filtration. 3-(1,3-thiazol-2-yl)prop-2-yn-1-ol (Compound 1-3, 3.3 g, 23.71 mmol, 1 eq) was slowly added to the filtrate at 0°C and stirred at room temperature for 16 hours. After the reaction was complete, methyl tert-butyl ether (120 mL) was slowly added at 0°C to precipitate a solid. The solid was filtered and rinsed twice with methyl tert-butyl ether (60 mL). The supernatant solid was dried to obtain Compound 001-4 (6.3 g). The crude product was used directly in the next step.

[0221] LCMS: (ESI, m / z): 155.1[M] + .

[0222] Step 4: Synthesis of Compound 001-5

[0223] At room temperature, compound 001-4 (6.3 g, 17.77 mmol, 1 eq) was dissolved in N,N-dimethylformamide (70 mL) solution, and silver carbonate (0.2 g, 0.71 mmol, 0.04 eq) was added. The reaction solution was stirred at 30°C for 16 hours. After the reaction was completed, water (200 mL) was added to the reaction mixture, and the mixture was extracted with ethyl acetate (100 mL×3). The organic phases were combined, backwashed with saturated sodium chloride solution (200 mL×2), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (ethyl acetate / petroleum ether (0-100%)) to give compound 001-5 (1.4 g, 51.08%).

[0224] LCMS: (ESI, m / z): 155.2 [M+H] + .

[0225] Step 5: Synthesis of Compound 001-6

[0226] At room temperature, (pyrazolo[3,2-b][1,3]thiazol-6-yl)methanol (compound 001-5, 1.4 g, 9.08 mmol, 1 eq) was dissolved in acetonitrile (20 mL), and a solution of 2-iodoacylbenzoic acid (5.09 g, 18.16 mmol, 2 eq) was slowly added. The mixture was stirred at 80°C for 1 hour. After the reaction was completed, the reaction solution was filtered, and the filtrate was added with water (80 mL). The mixture was extracted with ethyl acetate (2×60 mL), and the organic phase was washed with sodium bicarbonate (3×80 mL) and brine (80 mL). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and spin-dried to give the crude product, which was purified by silica gel column chromatography (ethyl acetate / petroleum ether = 0-50%) to give compound 001-6 (700 mg, 50.66%).

[0227] 1 H NMR (400MHz, CDCl3) δ10.05 (s, 1H), 7.85 (dd, J = 4.4, 0.8Hz, 1H), 7.12 (d, J = 4.2Hz, 1H), 6.96 (s, 1H).

[0228] Step 6: Synthesis of Compound 001-7

[0229] Under nitrogen protection, sodium hydroxide (552 mg, 13.8 mmol, 3 eq) and histamine dihydrochloride (1016.07 mg, 5.52 mmol, 1.2 eq) were added to a solution of pyrazolo[3,2-b][1,3]thiazole-6-carbaldehyde (compound 001-6, 700 mg, 4.60 mmol, 1 eq) in ethanol (12 mL) and water (6 mL) at 25°C, and the mixture was stirred at 85°C for 16 hours. After the reaction was completed, the reaction mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography (methanol / dichloromethane (0-30%)) to give compound 001-7 (850 mg, 75.64%).

[0230] LCMS: (ESI, m / z): 246.1 [M+H] + .

[0231] Step 6: Synthesis of Compound 001

[0232] Under nitrogen protection, phosphorus oxychloride (0.22 mL, 2.46 mmol, 3 eq) was added dropwise to a solution of 4-(pyrazolo[3.2.b][1.3]thiazol-6-yl)-4,5,6,7-tetrahydro-1H-imidazo[4.5-c]pyridine (compound 001-7, 200 mg, 0.82 mmol, 1 eq) and 5-(1-methyl-1H-pyrazol-4-yl)-1,3,4-oxadiazole-2-carboxylic acid (compound 001-8, 196.88 mg, 0.98 mmol, 1.2 eq) in pyridine (5 mL) at 0°C. The reaction was stirred at room temperature for 12 hours. After the reaction was completed, the reaction solution was dried and the residue was purified by dryness-based silica gel column chromatography (methanol / dichloromethane (0-10%)) to give compound 001 (100 mg, 29%). (5-(1-methyl-1H-pyrazol-4-yl)-1,3,4-oxadiazol-2-yl)(4-(pyrazolo[5,1-b]thiazol-6-yl)-1,4,6,7-tetrahydro-5H-imidazo[4,5-c]pyridin-5-yl)methanone (Compound 001, 100 mg) was separated by SFC (System: Waters SFC 150; Column: Chromatographic column specifications: 250*40mm 10μm; mobile phase A: supercritical CO2, mobile phase B: methanol (+0.1% 7.0mol / L amine methanol solution); A:B=65:35; wavelength: 214nm; flow rate: 140mL / min; column temperature: RT; back pressure: 100bar; injection volume: 4.0mL; cycle time: 14.7min; solvent: MeOH: distillation grade, supercritical CO2: food grade) to obtain compounds 001-1A (front peak, retention time: 2.522min, 41.59mg, 12.10%) and 001-1B (back peak, retention time: 4.549min, 15.34mg, 4.46%).

[0233] Compound 001-1A

[0234] LCMS: (ESI, m / z): 422.1 [M+H]+.

[0235] 1 H NMR (400MHz, DMSO-d6) δ12.02(s,1H),8.59(s,1H),8.19–8.10(m,1H),8.08(d,J=3.0Hz,1H),7.59(d,J=7.7Hz,1H),7.35– 7.26(m,1H),6.60–6.40(m,1H),4.85–4.70(m,1H),3.95(s,3H),3.78–3.60(m,1H),3.31–3.23(m,1H),3.03–2.66(m,2H).

[0236] Compound 001-1B

[0237] 1 H NMR (400MHz, DMSO-d6) δ12.01(s,1H),8.59(s,1H),8.19–8.10(m,1H),8.08(d,J=3.0Hz,1H),7.59(d,J=7.7Hz,1H),7.35– 7.26(m,1H),6.60–6.40(m,1H),4.86–4.70(m,1H),3.95(s,3H),3.78–3.55(m,1H),3.32–3.23(m,1H),3.07–2.66(m,2H).

[0238] Example 2

[0239] Step 1: Synthesis of Compound 094-1

[0240] Under nitrogen protection, methyl 2-(dimethoxyphosphoryl)-2-acetamidoacetate (26.28 g, 109.88 mmol, 1.1 eq) was dissolved in dichloromethane (500 mL), and DBU (16.73 g, 109.88 mmol, 1.1 eq) was added. The mixture was added to a solution of thiophene-2,3-dicarbaldehyde (14 g, 99.89 mmol, 1 eq) in tetrahydrofuran (1000 mL) at 0°C. The reaction solution was stirred at 0°C for 1 hour, then warmed to room temperature and reacted for 16 hours. After the reaction was completed, the reaction mixture was concentrated under reduced pressure and the residue was purified by silica gel column chromatography (ethyl acetate / petroleum ether (0-50%)) to give compounds 094-1 (8 g) and 093-1 (10 g).

[0241] Compound 094-1

[0242] LCMS: (ESI, m / z): 194.0 [M+H] + .

[0243] 1 H NMR (400MHz, CDCl3) δ9.22 (s, 1H), 8.73 (s, 1H), 7.74 (d, J = 4.0Hz, 1H), 7.55 (d, J = 8.0, 0.5Hz, 1H), 4.05 (s, 3H).

[0244] Compound 093-1

[0245] LCMS: (ESI, m / z): 194.0 [M+H] + .

[0246] 1 H NMR (400MHz, CDCl3) δ9.26 (s, 1H), 8.61 (s, 1H), 7.84 (d, J = 8.0Hz, 1H), 7.52 (d, J = 4.0Hz, 1H), 4.05 (s, 3H).

[0247] Step 2: Synthesis of Compound 094-2

[0248] To a solution of methyl thieno[3,2-c]pyridine-6-carboxylate (compound 094-1, 1 g, 5.18 mmol, 1 eq) in toluene (20 mL) was slowly added dropwise diisobutylaluminum hydride (6.91 mL, 10.36 mmol, 2 eq) under nitrogen at -65°C. The mixture was stirred at -65°C for 2 hours. After completion of the reaction, methanol (6.91 mL) was added to the reaction mixture at low temperature. The mixture was warmed to room temperature and quenched with saturated aqueous sodium chloride (100 mL). The mixture was stirred at room temperature for 10 minutes, filtered through celite, and the filtrate was extracted with ethyl acetate (3 × 100 mL). The organic phases were combined, backwashed with saturated brine (2 × 100 mL), and dried over anhydrous sodium sulfate. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure to afford compound 094-2 (790 mg). The crude product was used directly in the next step.

[0249] 1 H NMR (400MHz, CDCl3) δ10.26 (s, 1H), 9.28–9.23 (m, 1H), 8.57–8.53 (m, 1H), 7.80 (d, J = 4.0Hz, 1H), 7.62–7.54 (m, 1H).

[0250] Step 3: Synthesis of Compound 094-3

[0251] Under nitrogen protection, sodium hydroxide (294 mg, 7.35 mmol, 3 eq) and 2-(1H-imidazol-5-yl)ethane-1-amine dihydrochloride (541.11 mg, 2.94 mmol, 1.2 eq) were added to a solution of thieno[3,2-c]pyridine-6-carbaldehyde (compound 094-2, 400 mg, 2.45 mmol, 1 eq) in ethanol (10 mL) and water (10 mL) at room temperature. The mixture was stirred at 80°C for 16 hours. After the reaction was completed, the reaction mixture was concentrated and the crude product was purified by silica gel column chromatography (methanol / dichloromethane (0-20%)) to give compound 094-3 (570 mg, 90%).

[0252] LCMS: (ESI, m / z): 257.1 [M+H] + .

[0253] Step 4: Synthesis of Compound 094

[0254] Under nitrogen protection, phosphorus oxychloride (0.27 mL, 0.60 mmol, 3 eq) was added dropwise to a solution of 4-(thieno[3,2-c]pyridin-6-yl)-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine (compound 094-3, 250 mg, 0.98 mmol, 1 eq) and 5-(pyridin-2-yl)-1,3,4-oxadiazole-2-carboxylic acid lithium salt (compound 094-5, 203.82 mg, 1.03 mmol, 1.05 eq) in pyridine (10 mL) at 0°C. The mixture was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was dried and the residue was purified by silica gel column chromatography (methanol / dichloromethane (0-10%)) to give compound 094 (100 mg, 23.88%).

[0255] LCMS: (ESI, m / z): 430.1 [M+H] + .

[0256] 1 H NMR(400MHz,DMSO-d6)δ14.44(s,1H),9.20–9.11(m,1H),9.05–8.98(m,1H),8.87–8.8 0(m,1H),8.40–8.33(m,1H),8.30–8.23(m,1H),8.16–8.08(m,1H),7.97(t,J=5.5Hz,1 H),7.74–7.69(m,1H),7.68–7.63(m,1H),7.42(s,0.3H),6.95(s,0.7H),5.12–5.04(m ,0.7H),4.84–4.77(m,0.3H),3.84–3.71(m,1H),3.22–3.07(m,1H),3.01–2.88(m,1H).

[0257] Compound 094 was separated by SFC (System: Waters SFC 80; Column: Chromatographic column specifications: 250*30mm 10μm; mobile phase A: supercritical CO2, mobile phase B: methanol (+0.1% 7.0mol / L amine methanol solution): acetonitrile = 50:50; A:B = 45:55; wavelength: 214nm; flow rate: 80mL / min; column temperature: RT; back pressure: 100bar; injection volume: 4.0mL; cycle time: 14.7min; solvent: MeOH: distillation grade, acetonitrile: distillation grade, supercritical CO2: food grade), to obtain compounds 094-A (front peak, retention time: 2.599min, 45.2mg, 10.5%) and 094-B (back peak, retention time: 5.048min, 49.73mg, 11.8%).

[0258] Compound 094-A

[0259] LCMS: (ESI, m / z): 430.1 [M+H] + .

[0260] 1 H NMR(400MHz,MeOD-d4)δ9.02(d,J=27.8Hz,1H),8.77(t,J=5.4Hz,1H),8.33–8.27(m, 1H),8.18(s,0.6H),8.11–8.04(m,1H),7.90(s,0.4H),7.75(d,J=5.4Hz,1H),7.72–7. 63(m,2H),7.53(dd,J=20.6,5.4Hz,1H),7.41(s,0.4H),6.88(s,0.6H),5.05–4.90(m, 1H),4.02–3.92(m,0.6H),3.28–3.13(m,1H),3.08–2.94(m,0.4H),2.92–2.80(m,1H).

[0261] Compound 094-B

[0262] LCMS: (ESI, m / z): 430.1 [M+H] + .

[0263] 1H NMR(400MHz,MeOD-d4)δ9.01(d,J=27.9Hz,1H),8.77(t,J=5.8Hz,1H),8.30(dd,J=7.9, 3.5Hz,1H),8.18(s,0.6H),8.12–8.03(m,1H),7.90(s,0.4H),7.75(d,J=5.3Hz,1H),7.7 2–7.63(m,2H),7.53(dd,J=20.6,5.5Hz,1H),7.41(s,0.4H),6.88(s,0.6H),5.06–4.90( m,1H),4.02–3.90(m,0.6H),3.29–3.14(m,1H),3.08–2.95(m,0.4H),2.92–2.81(m,1H).

[0264] Using conditions similar to those in the above examples, the compounds listed in Table 1 were prepared. The structural characterization data of these compounds are listed in Table 1.

[0265] Table 1

[0266] Example 3

[0267] Step 1: Synthesis of Compound 093-2

[0268] To a solution of methyl thieno[2,3-c]pyridine-5-carboxylate (compound 093-1, 1 g, 5.18 mmol, 1 eq) in toluene (20 mL) was slowly added dropwise diisobutylaluminum hydride (6.91 mL, 10.36 mmol, 2 eq) under nitrogen at -65°C. The mixture was stirred at -65°C for 2 hours. After completion of the reaction, methanol (6.91 mL) was added to the reaction mixture at low temperature. The mixture was warmed to room temperature and quenched with saturated aqueous sodium chloride (100 mL). The mixture was stirred at room temperature for 10 minutes, filtered through celite, and the filtrate was extracted with ethyl acetate (3 × 100 mL). The organic phases were combined, backwashed with saturated brine (2 × 100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to afford compound 093-2 (790 mg). The crude product was used directly in the next step.

[0269] 1 H NMR (400MHz, CDCl3) δ10.24(s,1H),9.32–9.26(m,1H),8.41(s,1H),7.88–7.81(m,1H),7.59–7.53(m,1H).

[0270] Step 2: Synthesis of Compound 093-3

[0271] Under nitrogen protection, sodium hydroxide (294 mg, 7.35 mmol, 3 eq) and 2-(1H-imidazol-5-yl)ethane-1-amine dihydrochloride (541.11 mg, 2.94 mmol, 1.2 eq) were added to a solution of thieno[2,3-c]pyridine-5-carboxaldehyde (compound 093-2, 400 mg, 2.45 mmol, 1 eq) in ethanol (10 mL) and water (10 mL) at room temperature. The mixture was stirred at 80°C for 16 hours. After the reaction was completed, the reaction mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography (methanol / dichloromethane (0-20%)) to give compound 093-3 (690 mg, 88%).

[0272] LCMS: (ESI, m / z): 257.1 [M+H] + .

[0273] Step 3: Synthesis of Compound 093

[0274] Under nitrogen protection, phosphorus oxychloride (0.27 mL, 2.94 mmol, 3 eq) was added dropwise to a solution of 4-(thieno[2,3-c]pyridin-5-yl)-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine (compound 093-3, 250 mg, 0.98 mmol, 1 eq) and 5-(pyridin-2-yl)-1,3,4-oxadiazole-2-carboxylic acid lithium salt (compound 094-5, 203.82 mg, 1.03 mmol, 1.05 eq) in pyridine (10 mL) at 0°C. The mixture was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was dried and the residue was purified by silica gel column chromatography (methanol / dichloromethane (0-10%)) to give compound 093 (100 mg, 23.88%).

[0275] LCMS: (ESI, m / z): 430.1 [M+H] + .

[0276] 1H NMR (400MHz, DMSO-d6) δ14.40 (s, 1H), 9.28 (d, J = 9.6Hz, 1H), 9.03–8.95 (m, 1H), 8. 83(d,J=4.0Hz,1H),8.31–8.20(m,2H),8.15–8.07(m,2H),7.71(dd,J=7.4,5.0Hz, 1H),7.63(dd,J=14.0,5.4Hz,1H),7.40(s,0.3H),6.96(s,0.7H),5.10–4.95(m,0. 7H),4.87–4.76(m,0.3H),3.89–3.57(m,1H),3.23–3.09(m,1H),3.02–2.88(m,1H).

[0277] Compound 093 was separated by SFC (System: Waters SFC 80; Column: Chromatographic column specifications: 250*30mm 10μm; mobile phase A: supercritical CO2, mobile phase B: methanol (+0.1% 7.0mol / L amine methanol solution): acetonitrile = 50:50; A:B = 45:55; wavelength: 214nm; flow rate: 80mL / min; column temperature: RT; back pressure: 100bar; injection volume: 4.0mL; cycle time: 9.4min; solvent: MeOH: distillation grade, acetonitrile: distillation grade, supercritical CO2: food grade) to obtain compounds 093-A (front peak, retention time: 2.478min, 44.24mg, 10.6%) and 093-B (back peak, retention time: 4.779min, 44.83mg, 10.7%).

[0278] Compound 093-A

[0279] LCMS: (ESI, m / z): 430.1 [M+H] + .

[0280] 1H NMR(400MHz,MeOD-d4)δ9.07(d,J=28.9Hz,1H),8.77(t,J=4.9Hz,1H),8.33–8.26(m,1H),8.11 –8.05(m,1H),8.04(s,0.6H),7.97(dd,J=22.8,5.4Hz,1H),7.75(d,J=2.6Hz,0.8H),7.68(s,0. 6H),7.67–7.62(m,1H),7.52(d,J=5.4Hz,0.6H),7.41(d,J=4.9Hz,0.8H),6.89(s,0.6H),5.03 –4.88(m,1H),4.02–3.92(m,0.6H),3.28–3.15(m,1H),3.06–2.95(m,0.4H),2.91–2.81(m,1H).

[0281] Compound 093-B

[0282] LCMS: (ESI, m / z): 430.1 [M+H] + .

[0283] 1 H NMR(400MHz,MeOD-d4)δ9.07(d,J=28.9Hz,1H),8.77(t,J=4.8Hz,1H),8.29(dd,J=7.9,3.0Hz ,1H),8.11–8.02(m,1.6H),7.97(dd,J=22.8,5.4Hz,1H),7.77–7.73(m,0.8H),7.68(s,0.6H) ,7.67–7.63(m,1H),7.51(d,J=5.1Hz,0.6H),7.41(d,J=4.9Hz,0.8H),6.89(s,0.6H),5.04–4 .90(m,1H),4.03–3.91(m,0.6H),3.28–3.14(m,1H),3.08–2.96(m,0.4H),2.91–2.81(m,1H).

[0284] Example 4

[0285] Step 1: Synthesis of Compound 035-2

[0286] Under nitrogen protection, prop-2-yn-1-ol (3.68 g, 65.72 mmol, 1.3 eq), copper iodide (0.96 g, 5.05 mmol, 0.1 eq), and bis(triphenylphosphine)palladium(II) chloride (1.77 g, 2.53 mmol, 0.05 eq) were added to a solution of 2-bromo-4-methyl-1,3-thiazole (compound 035-1, 9 g, 50.55 mmol, 1 eq) in triethylamine (90 mL) at room temperature. The mixture was stirred at 50° C. for 16 hours. After the reaction was completed, the reaction solution was dried and the residue was purified by dryness-based silica gel column chromatography (ethyl acetate / petroleum ether (0-50%)) to give compound 035-2 (0.33 g, 38.35%).

[0287] LCMS: (ESI, m / z): 154.1 [M+H] + .

[0288] Step 2: Synthesis of compound 035-3

[0289] To a solution of water (8.45 mL, 468.66 mmol, 17.3 eq) and trifluoroacetic acid (49.75 mL, 650.16 mmol, 24 eq) was added (E)-(N-[(2,4,6-trimethylbenzenesulfonyl)oxy]ethylcarboximidate) (8.12 g, 28.44 mmol, 1.05 eq) at 0°C, and stirring was continued at 0°C for 2 hours. After the reaction was completed, water (120 mL) was added at 0°C, and a solid precipitated. The solid was filtered and rinsed twice with water (100 mL). The upper solid was dissolved in DCM (40 mL), dried over anhydrous sodium sulfate, and the filtrate was collected by filtration. 3-(4-Methyl-1,3-thiazol-2-yl)prop-2-yn-1-ol (Compound 035-2, 4.15 g, 27.09 mmol, 1 eq) was slowly added to the filtrate at 0°C and stirred at room temperature for 16 hours. After the reaction was complete, methyl tert-butyl ether (500 mL) was slowly added at 0°C to precipitate the product. The product was filtered and rinsed twice with methyl tert-butyl ether (300 mL). The supernatant solid was dried to obtain Compound 035-3 (4.8 g). The crude product was used directly in the next step.

[0290] LCMS: (ESI, m / z): 169.5[M] + .

[0291] Step 3: Synthesis of Compound 035-4

[0292] At room temperature, compound 035-3 (4.8 g, 28.37 mmol, 1 eq) was dissolved in methanol (50 mL) solution, and sodium methoxide (0.21 mL, 1.12 mmol, 2 eq) solution was slowly added and stirred at room temperature for 1 hour. After the reaction was completed, water (200 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (200 mL×2). The organic phases were combined, backwashed with saturated sodium chloride solution (200 mL×2), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (ethyl acetate / petroleum ether (0-50%)) to give compound 035-4 (0.1 g).

[0293] LCMS: (ESI, m / z): 169.1 [M+H] + .

[0294] Step 4: Synthesis of compound 035-5

[0295] At room temperature, (3-methylpyrazolo[3,2-b][1,3]thiazol-6-yl)methanol (compound 035-4, 80 mg, 0.48 mmol, 1 eq) was dissolved in acetonitrile (8 mL) solution, and 2-iodineoxybenzoic acid (268.82 mg, 0.96 mmol, 2 eq) was slowly added. After the addition was complete, the system was stirred at 80 ° C for 1 hour. After the reaction was completed, water (60 mL) was added to the reaction solution and extracted with ethyl acetate (2×60 mL). The organic phase was collected and washed with sodium bicarbonate (3×60 mL) and brine (80 mL). The collected organic phase was dried over anhydrous sodium sulfate, filtered, and dried to give the crude product, which was purified by silica gel column chromatography (ethyl acetate / petroleum ether = 0-30%) to give compound 035-5 (80 mg, 81%).

[0296] LCMS: (ESI, m / z): 167.4 [M+H] + .

[0297] Step 5: Synthesis of Compound 035-6

[0298] Under nitrogen protection, sodium hydroxide (43.2 mg, 1.08 mmol, 3 eq) and 2-(1H-imidazol-4-yl)ethane-1-amine dihydrochloride (79.52 mg, 0.43 mmol, 1.2 eq) were added to a solution of 3-methylpyrazolo[3,2-b][1,3]thiazole-6-carbaldehyde (compound 035-5, 60 mg, 0.36 mmol, 1 eq) in ethanol (3 mL) and water (3 mL) at room temperature. The mixture was stirred at 80°C for 16 hours. After the reaction was completed, the reaction solution was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography (methanol / dichloromethane (0-20%)) to give compound 035-6 (850 mg, 75.64%).

[0299] LCMS: (ESI, m / z): 260.1 [M+H] + .

[0300] Step 5: Synthesis of Compound 035

[0301] To a solution of 4-(pyrazolo[3,2-b][1,3]thiazol-6-yl)-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine (compound 035-6, 50 mg, 0.19 mmol, 1 eq) and 5-(1-methyl-1H-pyrazol-4-yl)-1,3,4-oxadiazole-2-carboxylate lithium (compound 094-5, 201.09 mg, 0.20 mmol, 1.05 eq) in pyridine (10 mL) was added dropwise phosphorus oxychloride (0.052 mL, 0.57 mmol, 3 eq) under nitrogen at 0°C. The reaction was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was dried and the residue was purified by dry-phase silica gel column chromatography (methanol / dichloromethane (0-10%)) and then purified by preparative HPLC (Waters 2767 / Qda, column: Xbridge C18 19*250mm, 10μm; mobile phase A: 0.03% NH3H2O / H2O, B: acetonitrile; flow rate: 20mL / min; elution gradient: 24% to 24%; retention time: 6.10-8.50min) to obtain compound 002 (14.71mg, 17.52%).

[0302] LCMS: (ESI, m / z): 436.2 [M+H] + .

[0303] Example 5

[0304] Step 1: Synthesis of compound 123-2

[0305] Under nitrogen protection, sodium hydroxide (245 mg, 6.12 mmol, 3 eq) and 2-(1H-imidazol-5-yl)ethane-1-amine dihydrochloride (446 mg, 2.45 mmol, 1.2 eq) were added to a solution of furano[2,3-c]pyridine-5-carbaldehyde (compound 123-1, 300 mg, 2.04 mmol, 1 eq) in ethanol (7 mL) and water (7 mL) at room temperature. The mixture was stirred at 80°C for 16 hours. After the reaction was completed, the reaction mixture was concentrated and the pH was adjusted to 9 with dilute hydrochloric acid. The crude product was purified by C18 silica gel column chromatography (acetonitrile / water (0-35%)) to give compound 123-2 (320 mg, 61%).

[0306] LCMS: (ESI, m / z): 241.2 [M+H]+ .

[0307] Step 2: Synthesis of compound 123

[0308] To a solution of 5-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)furo[2,3-c]pyridine (compound 123-2, 100 mg, 0.42 mmol, 1 eq) and 5-(pyridin-2-yl)-1,3,4-oxadiazole-2-carbonyl chloride (compound 123-3, 96.83 mg, 0.46 mmol, 1.1 eq) in N,N-dimethylformamide (1.5 mL) was added dropwise triethylamine (0.12 mL, 0.84 mmol, 2 eq) at 0°C under nitrogen protection. The mixture was stirred at room temperature for 1 hour. After the reaction, the crude product was prepared by HPLC under the following conditions (column: XBridge C18 19*250mm, 10μm; mobile phase A: 0.03% ammonia water, mobile phase B: acetonitrile; flow rate: 20 ml / min; gradient: 25% to 35%; retention time: 7.8-9.1 minutes) to obtain compound 123 (23.1 mg, 13.43%).

[0309] LCMS: (ESI, m / z): 414.1 [M+H] + .

[0310] 1 H NMR(400MHz,DMSO-d6)δ12.16(s,1H),8.91(s,0.6H),8.86–8.80(m,1.4H),8.28–8.21(m,2H), 8.12–8.07(m,1H),7.88(s,0.5H),7.71–7.67(m,2H),7.62(s,0.5H),7.12(d,J=1.6Hz,0.6H),7 .11(s,0.4H),7.07(d,J=1.6Hz,0.4H),6.67(s,0.6H),4.86–4.73(m,1H),3.93–3.80(m,0.6H) ,3.24–3.16(m,0.4H),3.04–2.96(m,0.6H),2.94–2.83(m,0.4H),2.77(dd,J=15.7,3.7Hz,1H).

[0311] Using conditions similar to those in the above examples, the compounds listed in Table 2 were prepared. The structural characterization data of these compounds are listed in Table 2.

[0312] Table 2

[0313] Example 6

[0314] Step 1: Synthesis of compound 105-1

[0315] At room temperature, methyl thieno[3,2-c]pyridine-6-carboxylate (compound 094-1, 300 mg, 1.55 mmol, 1 eq) was dissolved in N,N-dimethylformamide (5 mL), and N-chlorosuccinimide (620.91 mg, 4.65 mmol, 3 eq) was added. The mixture was stirred at room temperature overnight. After the reaction was completed, water (30 mL) was added to the reaction solution to dilute it, and the mixture was extracted with ethyl acetate (20 mL × 3). The organic phases were combined, washed with saturated brine (50 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 15:1) to give compound 105-1 (330 mg, 93%).

[0316] LCMS: (ESI, m / z): 228.0 [M+H] + .

[0317] Step 2: Synthesis of compound 105-2

[0318] Under nitrogen protection, diisobutylaluminum hydride (2.46 mL, 3.69 mmol, 3 eq) was slowly added dropwise to a solution of methyl 3-chlorothieno[3,2-c]pyridine-6-carboxylate (compound 105-1, 280 mg, 1.23 mmol, 1 eq) in tetrahydrofuran (3 mL) at -78°C, and the mixture was stirred at -78°C for 2 hours. After the reaction was completed, the reaction mixture was added to a stirred saturated aqueous solution of potassium tartrate at -20°C and stirred at room temperature for 2 hours. The mixture was extracted with ethyl acetate (3×20 mL), the organic phases were combined, washed with saturated brine (2×10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate (0-20%)) to give compound 105-2 (150 mg, 62%).

[0319] LCMS: (ESI, m / z): 198.0 [M+H] + .

[0320] Step 3: Synthesis of Compound 105-3

[0321] Under nitrogen protection, 2-(1H-imidazol-5-yl)ethane-1-amine dihydrochloride (145.77 mg, 0.79 mmol, 1.2 eq) and sodium hydroxide (79.2 mg, 1.98 mmol, 3.0 eq) were added to a mixed solvent of EtOH (1.5 mL) and water (1.5 mL) at 45 ° C. The mixture was stirred at 80 ° C. for 16 hours. After the reaction was completed, 1 M aqueous hydrochloric acid was added to the reaction mixture to adjust the pH to 7. The ethanol was spin-dried to give a crude product. The crude product was purified by reverse column chromatography (0.1% NH3·H2O / acetonitrile = 29 / 71) to give compound 105-3 (30 mg, 15.69%).

[0322] LCMS: (ESI, m / z): 291.1 [M+H] + .

[0323] Step 4: Synthesis of Compound 105

[0324] To a solution of 3-chloro-6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[3,2-c]pyridine (compound 105-3, 30 mg, 0.10 mmol, 1 eq) and 5-(pyridin-2-yl)-1,3,4-oxadiazole-2-carboxylic acid potassium salt (compound 105-4, 34.38 mg, 0.15 mmol, 1.5 eq) in pyridine (1 mL) was added dropwise phosphorus oxychloride (46.0 mg, 0.3 mmol, 3 eq) at 0°C under nitrogen atmosphere. The mixture was stirred at room temperature for 2 hours. After the reaction, the reaction solution was dried and the crude product was prepared by HPLC under the following conditions (column: XBridge C18 19*250mm, 10μm; mobile phase A: 0.03% ammonium bicarbonate, mobile phase B: acetonitrile; flow rate: 20 ml / min; gradient: 33% to 33%; retention time: 8.1–9.3 min / 16 min) to obtain compound 105 (3.33 mg, 6.96%).

[0325] LCMS: (ESI, m / z): 464.2 [M+H] + .

[0326] The above method was repeated to enrich compound 105 (85.67 mg), which was separated by SFC (System: Waters SFC 150; Column: Chromatographic column specifications: 250*25mm 10μm; mobile phase A: supercritical CO2, mobile phase B: methanol (+0.2% 7.0mol / L amine methanol solution); A:B=75:25; wavelength: 214nm; flow rate: 140mL / min; column temperature: RT; back pressure: 100bar; injection volume: 2.5mL; cycle time: 6.0min) to obtain compounds 105-A (front peak, retention time: 4.81min, 16.95mg) and 105-B (back peak, retention time: 7.05min, 20.49mg).

[0327] Compound 105-A

[0328] LCMS: (ESI, m / z): 464.1 [M+H] + .

[0329] 1 H NMR(400MHz,DMSO-d6)δ12.13(s,1H),8.99(d,J=36.6Hz,1H),8.84–8.78(m ,1H),8.32(s,0.6H),8.24(t,J=8.4Hz,1H),8.14(s,0.4H),8.12–8.06(m,1H ),8.01(d,J=11.5Hz,1H),7.72–7.55(m,2H),6.91(d,J=186.4Hz,1H),4.91 –4.77(m,1H),4.05–3.85(m,0.6H),3.26–3.16(m,0.4H),3.09–2.73(m,2H).

[0330] Compound 105-B

[0331] LCMS: (ESI, m / z): 464.1 [M+H] + .

[0332] 1 H NMR(400MHz,DMSO-d6)δ12.14(s,1H),8.99(d,J=36.6Hz,1H),8.85–8.78(m ,1H),8.32(s,0.6H),8.24(t,J=8.4Hz,1H),8.14(s,0.4H),8.12–8.06(m,1H ),8.01(d,J=11.5Hz,1H),7.72–7.55(m,2H),6.91(d,J=186.7Hz,1H),4.93 –4.76(m,1H),4.00–3.83(m,0.6H),3.26–3.18(m,0.4H),3.09–2.72(m,2H).

[0333] Using conditions similar to those in the above examples, the compounds listed in Table 3 were prepared. The structural characterization data of these compounds are listed in Table 3.

[0334] Table 3

[0335] Example 7

[0336] Step 1: Synthesis of compound 103-1

[0337] Thieno[3,2-c]pyridine-6-carboxylic acid methyl ester (compound 094-1, 2.0 g, 10.35 mmol, 1 eq) was dissolved in chloroform (20 mL) at room temperature, and sodium bicarbonate (0.87 g, 10.35 mmol, 1.0 eq), potassium dihydrogen phosphate (2.70 g, 15.52 mmol, 1.5 eq) and magnesium sulfate (1.62 g, 13.46 mmol, 1.3 eq) were added in sequence. Bromine (2.4 8g, 15.52mmol, 1.5eq) was slowly added to the reaction solution, and the mixture was stirred at 50°C overnight. After the reaction was completed, water (50mL) was added to the reaction solution to dilute it, and the mixture was extracted with dichloromethane (50mL×3). The organic phases were combined, washed with saturated brine (50mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated. The resulting residue was purified by silica gel column chromatography (petroleum ether\ethyl acetate=5:1) to give compound 103-1 (1.2g, 43%).

[0338] LCMS: (ESI, m / z): 273.8 [M+2+H] + .

[0339] Step 2: Synthesis of compound 103-2

[0340] At room temperature, methyl 3-bromothieno[3,2-c]pyridine-6-carboxylate (compound 103-1, 1.1 g, 4.04 mmol, 1.0 eq) was dissolved in tetrahydrofuran (10 mL), and lithium borohydride (0.18 g, 8.08 mmol, 2.0 eq) and methanol (0.5 mL) were added. The mixture was stirred at room temperature for 1 hour. After the reaction was completed, water (10 ml) was added to the reaction solution to quench the mixture, and the mixture was extracted with ethyl acetate (20 mL × 3). The combined organic phases were washed with saturated brine (20 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The resulting residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate (0-50%)) to give compound 103-2 (650 mg, 66%).

[0341] LCMS: (ESI, m / z): 245.9 [M+2+H] + .

[0342] Step 3: Synthesis of Compound 103-3

[0343] To a solution of (3-bromothieno[3,2-c]pyridin-6-yl)methanol (compound 103-2, 650 mg, 2.66 mmol, 1 eq) in dioxane (2 mL) and water (0.5 mL) was added trimethylcyclotriboroxane (667.82 mg, 5.32 mmol, 2.0 eq), dichloro[1,1-bis(di-tert-butylphosphino)ferrocene]palladium(II) (173.37 mg, 0.27 mmol, 0.1 eq) and phosphorus at 45 ° C under nitrogen protection. The mixture was stirred at 100 ° C overnight. After the reaction was completed, water (20 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with ethyl acetate (20 mL × 3). The organic phases were combined, washed with saturated brine (10 mL × 2), dried over anhydrous sodium sulfate, filtered and concentrated to give a crude product, which was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 5: 1) to give compound 103-3 (150 mg, 31.43%).

[0344] LCMS: (ESI, m / z): 180.0 [M+H] + .

[0345] Step 4: Synthesis of compound 103-4

[0346] At room temperature, (3-methylthieno[3,2-c]pyridin-6-yl)methanol (compound 103-3, 130 mg, 0.73 mmol, 1.0 eq) was dissolved in dichloromethane (8 mL), and Dess-Martin periodinane (619.24 mg, 1.46 mmol, 2.0 eq) was slowly added. The reaction was stirred at room temperature for 30 minutes. After the reaction was completed, saturated aqueous sodium bicarbonate solution (20 mL) was added to the reaction solution for dilution, and dichloromethane (20 mL×3) was added for extraction. The organic phases were combined, washed with saturated brine (20 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 8:1) to give compound 103-4 (120 mg, 93%).

[0347] LCMS: (ESI, m / z): 178.0 [M+H] + .

[0348] Step 4: Synthesis of compound 103-5

[0349] 3-Methylthieno[3,2-c]pyridine-6-carbaldehyde (compound 103-4, 120 mg, 0.68 mmol, 1.0 eq) was dissolved in ethanol (1 mL) and water (1 mL), and 2-(1H-imidazol-5-yl)ethane-1-amine dihydrochloride (150.18 mg, 0.82 mmol, 1.2 eq) and sodium hydroxide (81.6 mg, 2.04 mmol, 2.5 eq) were added. The mixture was stirred at 80°C for 16 hours. After the reaction was completed, 1M aqueous hydrochloric acid solution was added to the reaction solution to adjust the pH to 7. The ethanol was spin-dried to obtain a crude product. The crude product was purified by reverse column purification (0.1% NH3·H2O / acetonitrile = 20%) to obtain compound 103-5 (30 mg, 16.39%).

[0350] LCMS: (ESI, m / z): 271.1 [M+H] + .

[0351] Step 5: Synthesis of Compound 103

[0352] Under nitrogen protection, phosphorus oxychloride (50.60 mg, 0.33 mmol, 3 eq) was added dropwise to a solution of 3-methyl-6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[3,2-c]pyridine (compound 103-5, 30 mg, 0.11 mmol, 1 eq) and potassium (5-(pyridin-2-yl)-1,3,4-oxadiazole-2-carboxylate) (compound 105-4, 37.82 mg, 0.17 mmol, 1.5 eq) in pyridine (1 mL) at 0°C. The mixture was stirred at room temperature for 2 hours. After the reaction, the reaction solution was dried and the crude product was purified by HPLC (column: XBridge C18 19*250mm, 10μm; mobile phase A: 0.1% formic acid, mobile phase B: acetonitrile; flow rate: 20 ml / min; gradient: 18% to 28%; retention time: 7.6–8.9 min / 16 min) to obtain compound 103 (7.37 mg, 14.98%).

[0353] LCMS: (ESI, m / z): 444.1 [M+H] + .

[0354] The above method was repeated to enrich compound 103 (150 mg) and separate by SFC (System: Waters SFC 150; Column: Chromatographic column specifications: 250*30mm 10μm; mobile phase A: supercritical CO2, mobile phase B: methanol (+0.1% 7.0mol / L amine methanol solution); A:B=60:40; wavelength: 214nm; flow rate: 120mL / min; column temperature: RT; back pressure: 100bar; injection volume: 8.0mL; cycle time: 5.0min) to obtain compound 103-A (front peak, retention time: 11.17min, 44.97mg) and compound 103-B (back peak, retention time: 17.72min, 45.03mg).

[0355] Compound 103-A

[0356] LCMS: (ESI, m / z): 444.2 [M+H] + .

[0357] 1 H NMR (400MHz, DMSO-d6) δ12.12(d,J=37.5Hz,1H),8.96(d,J=34.8Hz,1H),8.83(t,J= 5.4Hz,1H),8.28–8.21(m,1H),8.16(s,0.6H),8.13–8.06(m,1H),7.95(s,0.4H),7.7 4–7.57(m,2H),7.48(d,J=13.4Hz,1H),6.86(d,J=190.6Hz,1H),4.90–4.71(m,1H),4 .03–3.78(m,0.6H),3.23–3.11(m,0.4H),3.08–2.74(m,2H),2.45(d,J=18.6Hz,3H).

[0358] Compound 103-B

[0359] LCMS: (ESI, m / z): 444.2 [M+H] + .

[0360] 1H NMR (400MHz, DMSO-d6) δ12.14(s,1H),8.97(d,J=35.8Hz,1H),8.83(t,J=5.6Hz,1H), 8.25(dd,J=7.7,4.2Hz,1H),8.16(s,0.6H),8.10(t,J=7.8Hz,1H),7.95(s,0.4H),7. 72–7.58(m,2H),7.49(d,J=13.2Hz,1H),6.90(d,J=186.3Hz,1H),4.89–4.73(m,1H), 4.00–3.83(m,0.6H),3.23–3.11(m,0.4H),3.07–2.72(m,2H),2.45(d,J=17.5Hz,3H).

[0361] Using conditions similar to those in the above examples, the compounds listed in Table 4 were prepared. The structural characterization data of these compounds are listed in Table 4.

[0362] Table 4

[0363] Example 8

[0364] Step 1: Synthesis of compound 132-2

[0365] To a solution of thieno[2,3-b]pyridine (compound 132-1, 2.1 g, 15.53 mmol, 1 eq) in dichloromethane (20 mL) was added 3-chloroperoxybenzoic acid (3.22 g, 18.64 mmol, 1.2 eq) at 0°C. The mixture was stirred at room temperature for 40 hours. After the reaction was complete, the mixture was filtered and dried to give compound 132-2 (2.9 g, crude product), which was used directly in the next reaction.

[0366] LCMS: (ESI, m / z): 152.1 [M+H] + .

[0367] Step 2: Synthesis of compound 132-3

[0368] Under nitrogen protection, tetrabutylammonium bromide (4.49 g, 13.93 mmol, 1.17 eq) was added to a solution of thieno[2,3-b]pyridine 7-oxide (compound 132-2, 1.8 g, 11.91 mmol, 1 eq) in N,N-dimethylformamide (10.6 mL) and ethylene glycol dimethyl ether (62 mL) at once at 0°C, and methanesulfonic anhydride (4.02 g, 23.11 mmol, 1.94 eq) was slowly added in batches. The mixture was stirred at 0°C for 2 hours, then heated to room temperature and stirred for 16 hours. After the reaction was completed, the reaction solution was spin-dried and the residue was dissolved in dichloromethane (150 mL) and water (150 mL). The mixture was washed with 2N ethanol and 4% ethanol. The pH was adjusted to 7 with NaOH, extracted with dichloromethane (3×150 mL), and the organic layer was washed with saturated brine (3×100 mL), dried over anhydrous sodium sulfate, filtered and concentrated. The resulting residue was purified by silica gel column chromatography (ethyl acetate / petroleum ether (0-30%)) to give compound 132-3 (2.2 g, 86.32%).

[0369] LCMS: (ESI, m / z): 213.9 [M+H] + .

[0370] Step 3: Synthesis of compound 132-4

[0371] 6-Bromothieno[2,3-b]pyridine (compound 132-3, 2 g, 9.34 mmol, 1 eq), triethylamine (4.73 g, 46.7 mmol, 5 eq) and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (0.68 g, 0.93 mmol, 0.1 eq) were dissolved in N,N-dimethylformamide (8 mL) and methanol (32 mL), and CO (15 Psi) was introduced. The mixture should be stirred at 70°C for 12 hours. After the reaction is completed, the reaction solution is spin-dried, quenched with water (200 mL), extracted with ethyl acetate (3×300 mL), and the organic phases are combined, backwashed with saturated brine (3×200 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate is concentrated under reduced pressure. The resulting residue is purified by dryness method by silica gel column chromatography (ethyl acetate / petroleum ether (0-20%)) to give compound 132-4 (900 mg, 49.86%).

[0372] LCMS: (ESI, m / z): 194.0 [M+H] + .

[0373] Step 4: Synthesis of compound 132-5

[0374] To a solution of methyl thieno[2,3-b]pyridine-6-carboxylate (compound 132-4, 900 mg, 4.66 mmol, 1 eq) in toluene (20 mL) was slowly added dropwise diisobutylaluminum hydride (6.21 mL, 9.32 mmol, 2 eq) under nitrogen at -65°C. The mixture was stirred at -65°C for 2 hours. After completion of the reaction, methanol (6.21 mL) was added to the reaction mixture at low temperature. The mixture was warmed to room temperature and quenched with saturated aqueous sodium chloride (200 mL). The mixture was stirred at room temperature for 10 minutes, filtered through celite, and the filtrate was extracted with ethyl acetate (100 mL × 3). The organic phases were combined, backwashed with saturated brine (200 mL × 2), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to afford compound 132-5 (0.5 g). The crude product was used directly in the next step.

[0375] Step 5: Synthesis of compound 132-6

[0376] Under nitrogen protection, sodium hydroxide (367.2 mg, 9.18 mmol, 3 eq) and 2-(1H-imidazol-5-yl)ethane-1-amine dihydrochloride (675.83 mg, 3.67 mmol, 1.2 eq) were added to a solution of thieno[2,3-b]pyridine-6-carbaldehyde (compound 132-5, 500 mg, 3.06 mmol, 1 eq) in ethanol (10 mL) and water (10 mL) at room temperature. The mixture was stirred at 85°C for 12 hours. After the reaction was completed, the reaction mixture was concentrated under reduced pressure and the crude product was purified by silica gel column chromatography (methanol / dichloromethane (0-20%)) to give compound 132-5 (720 mg, 91.68%).

[0377] LCMS: (ESI, m / z): 257.1 [M+H] + .

[0378] Step 6: Synthesis of Compound 132

[0379] Under nitrogen protection, phosphorus oxychloride (0.32 mL, 3.51 mmol, 3 eq) was added dropwise to a solution of 6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[2,3-b]pyridine (compound 132-6, 0.3 g, 1.17 mmol, 1 eq) and 5-(pyridin-2-yl)-1,3,4-oxadiazole-2-carboxylate lithium (compound 94-5, 0.24 g, 1.23 mmol, 1.05 eq) in pyridine (10 mL) at 0°C. The mixture was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was dried and the residue was purified by dryness-based silica gel column chromatography (methanol / dichloromethane (0-10%)) and then slurried with methanol to give compound 132 (120 mg, 23.88%).

[0380] LCMS: (ESI, m / z): 430.2 [M+H] + .

[0381] 1 H NMR(400MHz,DMSO-d6)δ14.38(s,1H),9.02–8.88(m,1H),8.83(d,J=4.8Hz,1H),8.47–8.37 (m,1H),8.26(dd,J=16.4,7.8Hz,1H),8.16–8.07(m,1H),7.94(dd,J=9.0,6.0Hz,1H),7.82( d,J=8.4Hz,0.3H),7.73–7.68(m,1.7H),7.54–7.46(m,1H),7.38(s,0.3H),7.00(s,0.7H), 5.04(dd,J=14.3,5.2Hz,0.7H),4.84–4.76(m,0.3H),3.74–3.58(m,1H),3.22–2.87(m,2H).

[0382] The experiment was repeated using the above method to enrich compound 132 (200 mg) and separate it by SFC (System: Waters SFC 150; Column: Chromatographic column specifications: 250*25mm 10μm; mobile phase: A supercritical CO2, mobile phase, B: methanol (+0.1% 7.0mol / L amine methanol solution); A:B=50:50; wavelength: 214nm; flow rate: 120mL / min; column temperature: RT; back pressure: 100bar; injection volume: 2.0mL; cycle time: 7.7min) to obtain compound 132-A (front peak, retention time: 2.862min, 54.16mg) and compound 132-B (back peak, retention time: 4.853min, 68.85mg).

[0383] Compound 132-A

[0384] LCMS: (ESI, m / z): 430.2 [M+H] + .

[0385] 1H NMR(400MHz,MeOD-d4)δ8.79–8.74(m,1H),8.29(d,J=7.9Hz,1H),8.20(dd,J=35.7,8 .3Hz,1H),8.10–8.04(m,1H),7.74–7.61(m,3.6H),7.49(d,J=8.3Hz,0.4H),7.36(d,J =6.0Hz,0.6H),7.34(s,0.4H),7.31(d,J=6.0Hz,0.4H),6.87(s,0.6H),5.08–4.92(m, 1H),4.07–3.93(m,0.6H),3.41–3.34(m,0.4H),3.25–2.96(m,1H),2.92–2.81(m,1H).

[0386] Compound 132-B

[0387] LCMS: (ESI, m / z): 430.1 [M+H] + .

[0388] 1 H NMR(400MHz,MeOD-d4)δ8.77(d,J=4.7Hz,1H),8.34–8.18(m,2H),8.17–8.03(m,2H),7.74–7.55(m,3H),7.44(s,0.4H),7.36(dd,J=16.2,6.0Hz, 1H),6.98(s,0.6H),5.15–4.93(m,1H),3.96–3.79(m,0.6H),3.43–3.36 (m,0.4H),3.27–3.18(m,0.6H),3.10–2.98(m,0.4H),2.97–2.84(m,1H).

[0389] Using conditions similar to those in the above examples, the compounds listed in Table 5 were prepared. The structural characterization data of these compounds are listed in Table 5.

[0390] Table 5

[0391] Example 9

[0392] Step 1: Synthesis of compound 162-2

[0393] To a solution of methyl 1-methyl-1H-pyrazole-3-carboxylate (compound 162-1, 5 g, 35.68 mmol) in ethanol (50 mL) was added hydrazine hydrate (10.72 g, 214.08 mmol), and the mixture was stirred at room temperature for 3 hours. After the reaction was completed, the reaction solution was filtered, washed with ethanol, and concentrated to give compound 162-2 (4 g, 76%).

[0394] LCMS: (ESI, m / z): 141.1 [M+H] + .

[0395] Step 2: Synthesis of compound 162-3

[0396] At 0°C, methyl 2-chloro-2-oxoacetate (2.10 g, 17.12 mmol) and triethylamine (4.33 g, 42.81 mmol) were added dropwise to a solution of 1-methyl-1H-pyrazole-3-carboxylic acid hydrazide (compound 162-2, 2 g, 14.27 mmol) in dichloromethane (20 mL). The reaction solution was stirred at room temperature for 1 hour and concentrated to obtain compound 162-3 (3.1 g, 96%).

[0397] LCMS: (ESI, m / z): 227.0 [M+H] + .

[0398] Step 3: Synthesis of compound 162-4

[0399] To a solution of methyl 2-[(1-methyl-1H-pyrazol-3-yl)formylhydrazide]-2-oxoacetate (compound 162-3, 2 g, 8.84 mmol) in dichloromethane (20 mL) at 0°C were added p-toluenesulfonyl chloride (1.69 g, 8.84 mmol) and triethylamine (2.68 g, 26.52 mmol). The reaction solution was stirred at room temperature for 1 hour. After the reaction was completed, it was diluted with water (30 mL) and extracted with ethyl acetate (100 mL×3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 0%-20%) to give compound 162-4 (1.5 g, 73%).

[0400] LCMS: (ESI, m / z): 209.0 [M+H] + .

[0401] 1 H NMR (400MHz, CDCl3) δ7.52 (d, J = 2.4Hz, 1H), 7.01 (d, J = 2.4Hz, 1H), 4.08 (s, 3H), 4.06 (s, 3H).

[0402] Step 4: Synthesis of compound 162-5

[0403] To a mixed solution of methyl 5-(1-methyl-1H-pyrazol-3-yl)-1,3,4-oxadiazole-2-carboxylate (compound 162-4, 300 mg, 1.44 mmol) in methyl tert-butyl ether (5 mL) and dichloromethane (5 mL) was added trimethyl(potassium oxy)silane (184.7 mg, 1.44 mmol). The mixture was stirred at room temperature for 2 hours. After the reaction was completed, 1N hydrochloric acid was added to adjust the pH to 7, the mixture was diluted with water (30 mL), and extracted with ethyl acetate (100 mL×3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated to give compound 162-5 (190 mg, 51%).

[0404] LCMS: (ESI, m / z): 159.0 [M+H] + .

[0405] Step 5: Synthesis of Compound 162

[0406] 4-(Thieno[3,2-c]pyridin-6-yl)-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine (Compound 94-3, 150 mg, 0.59 mmol) and 5-(1-methyl-1H-pyrazol-3-yl)-1,3,4-oxadiazole-2-carboxylate potassium (Compound 162-5, 137 mg, 0.59 mmol) were dissolved in pyridine (5 mL), and trichlorophosphine oxide (271 mg, 1.77 mmol) was added dropwise thereto at 0°C. The mixture was stirred at room temperature for 1 hour. After the reaction was completed, the reaction solution was concentrated and the residue was purified by high performance liquid chromatography under the following conditions: Chromatographic column specifications: Waters 2767 / QDA column: Atlatis T3Prep OBD 19*250mm*10μm; mobile phase A: water (0.05% trifluoroacetic acid), mobile phase B: acetonitrile; flow rate: 20mL / min; elution gradient: from 14% B to 19% B in 16min; detection wavelength: 254nm / 220nm; retention time (min) 6.2-9.0, to obtain compound 162 (16.85mg, 6.53%).

[0407] LCMS: (ESI, m / z): 433.1 [M+H] + .

[0408] 1H NMR (400MHz, DMSO-d6) δ9.15(d,J=14.6Hz,1H),9.00(s,1H),8.33(d,J=5.0Hz,1H),8. 01(d,J=2.3Hz,1H),7.96(t,J=5.8Hz,1H),7.69–7.62(m,1H),7.45(s,0.3H),6.97(dd ,J=11.8,2.3Hz,1H),6.93(s,1H),5.10(dd,J=14.1,5.1Hz,0.7H),4.79(dd,J=13.0,4 .9Hz,1H),4.00(s,3H),3.76–3.74(m,0.7H),3.33–3.28(m,0.3H),3.13–2.90(m,2H).

[0409] Using conditions similar to those in the above examples, the compounds listed in Table 6 were prepared. The structural characterization data of these compounds are listed in Table 6.

[0410] Table 6 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0411] Example 10

[0412] Step 1: Synthesis of compound 163-2

[0413] At room temperature, sodium 2-chloro-2,2-difluoroacetate (7.07 g, 46.38 mmol, 1.2 eq) and cesium carbonate (23.25 g, 71.36 mmol, 2 eq) were added to a solution of ethyl 1H-pyrazole-3-carboxylate (compound 163-1, 5 g, 35.68 mmol, 1 eq) in N,N-diethylacetamide (50 mL). The mixture was stirred at 100 ° C for 16 hours. After the reaction was completed, water (50 mL) was added to the reaction solution, and the mixture was diluted with ethyl acetate (50 mL × 3). The organic phases were combined, washed with saturated brine (30 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 3 / 1) to give compound 163-2 (2.2 g, 32.43%).

[0414] LCMS: (ESI, m / z): 191.0 [M+H] + .

[0415] Based on the similar procedures to synthesize compound 162, compound 163 (23.97 mg, 3.92%) was obtained.

[0416] LCMS: (ESI, m / z): 469.1 [M+H] + .

[0417] 1 H NMR(400MHz, DMSO-d6)δ9.06(d,J=36.7Hz,1H),8.58(t,J=2.6Hz,1H),8.38( s,1H),8.22–8.14(m,1H),8.04–7.98(m,1H),7.96–7.82(m,1H),7.68(s,0.4H ),7.63–7.54(m,1.6H),7.28–7.21(m,1H),7.12(s,0.4H),6.65(s,0.6H),4.9 2–4.72(m,1H),3.94–3.90(m,0.6H),3.23–3.18(m,0.4H),3.01–2.73(m,2H).

[0418] Using conditions similar to those in the above examples, the compounds listed in Table 7 were prepared. The structural characterization data of these compounds are listed in Table 7.

[0419] Table 7 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0420] Example 11

[0421] Step 1: Synthesis of compound 193-2

[0422] To a solution of 3-bromo-6-(((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 193-1, 300 mg, 0.75 mmol) in 1,4-dioxane (3 mL) and water (0.3 mL) were added 4,4,5,5-tetramethyl-2-vinyl-1,3,2-dioxaborolane (347 mg, 0.25 mmol), [1,1'-bis(di-tert-butylphosphino)ferrocene]palladium dichloride (49 mg, 0.075 mmol) and potassium phosphate (478 mg, 2.25 mmol). The mixture was reacted at 100°C under nitrogen for 2 h. After the reaction was completed, the reaction solution was concentrated to obtain a crude product, which was purified by normal phase purification (petroleum ether:ethyl acetate = 3:1) to obtain compound 193-2 (160 mg, 61.44%).

[0423] LCMS: (ESI, m / z): 348.3 [M+H] + .

[0424] Step 2: Synthesis of compound 193-3

[0425] To a solution of 6-(((triisopropylsilyl)oxy)methyl)-3-vinylthieno[3,2-c]pyridine (compound 193-2, 160 mg, 0.46 mmol) in methanol (3 mL) was added palladium carbon (30 mg), and the mixture was reacted at room temperature under hydrogen protection for 1 h. After the reaction was completed, the reaction liquid was filtered and the filtrate was concentrated to obtain a crude product. The crude product was purified by normal phase purification (petroleum ether: ethyl acetate = 2:1) to obtain compound 193-3 (120 mg, 74.57%).

[0426] LCMS: (ESI, m / z): 350.5 [M+H] + .

[0427] Step 3: Synthesis of Compound 193-4

[0428] To a solution of 3-ethyl-6-(((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 193-3, 120 mg, 0.34 mmol) in tetrahydrofuran (2 mL) was added tetrabutylammonium fluoride (190 mg, 0.51 mmol) and the mixture was reacted at room temperature for 12 h. After the reaction was completed, the reaction solution was concentrated to obtain a crude product, which was purified by normal phase purification (petroleum ether: ethyl acetate = 1:1) to obtain compound 193-4 (40 mg, 60.30%).

[0429] LCMS: (ESI, m / z): 194.0 [M+H] + .

[0430] Based on the similar procedures to synthesize compound 103, compound 193 (4 mg, 9.95%) was obtained.

[0431] LCMS: (ESI, m / z): 458.1 [M+H] + .

[0432] 1 H NMR (400MHz, DMSO-d6) δ12.13(d,J=36.2Hz,1H),9.01(d,J=36.4Hz,1H),8.82(t,J=5.0H z,1H),8.29–8.21(m,1H),8.17(s,0.6H),8.13–8.06(m,1H),7.96(s,0.4H),7.72–7.66(m ,1.4H),7.59(s,0.6H),7.50(d,J=12.5Hz,1H),7.10(s,0.4H),6.63(s,0.6H),4.93–4.72 (m,1H),4.01–3.82(m,0.5H),3.24–3.17(m,0.5H),2.96–2.73(m,4H),1.33–1.25(m,3H).

[0433] Separation was performed by SFC (System: Waters SFC 150; Column: Chromatographic column specifications: 250*25mm10μm; mobile phase: A supercritical CO2, mobile phase, B: methanol (+0.1% 7.0mol / L amine methanol solution); wavelength: 254nm; flow rate: 40mL / min; column temperature: RT; back pressure: 100bar; injection volume: 3.0mL; cycle time: 9.3min) to obtain compound 193-A (front peak, retention time: 9.5-12.5min, 24mg) and compound 193-B (back peak, retention time: 13.6-17.0min, 21mg).

[0434] Compound 193-A

[0435] LCMS: (ESI, m / z): 458.2 [M+H] + .

[0436] 1H NMR (400MHz, DMSO-d6) δ12.09 (s, 1H), 9.02 (dd, J = 33.7, 20.0Hz, 1H), 8.82 (t ,J=5.2Hz,1H),8.30–8.21(m,1H),8.19–7.91(m,2H),7.73–7.56(m,2H),7.4 9(d,J=12.3Hz,1H),6.94(dd,J=193.0,62.7Hz,1H),4.90–4.71(m,1H),4.02 –3.82(m,0.6H),3.25–3.15(m,0.4H),3.09–2.69(m,4H),1.34–1.21(m,3H).

[0437] Compound 193-B

[0438] LCMS: (ESI, m / z): 458.2 [M+H] + .

[0439] 1 H NMR(400MHz,DMSO-d6)δ12.13(d,J=34.7Hz,1H),9.02(dd,J=33.9,20.3Hz,1H) ,8.82(t,J=5.2Hz,1H),8.29–8.21(m,1H),8.19–7.92(m,2H),7.73–7.57(m,2H ),7.50(t,J=11.4Hz,1H),6.94(dd,J=192.9,62.4Hz,1H),4.94–4.69(m,1H),4 .06–3.78(m,0.6H),3.26–3.15(m,0.4H),3.10–2.69(m,4H),1.37–1.19(m,3H).

[0440] Using conditions similar to those in the above examples, the compounds listed in Table 8 were prepared. The structural characterization data of these compounds are listed in Table 8.

[0441] Table 8 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0442] Example 12

[0443] Step 1: Synthesis of Compound 107-1

[0444] Under nitrogen protection, 1.3 M isopropylmagnesium chloride-lithium chloride (2.88 mL, 3.75 mmol, 3 eq) was added to a solution of 3-bromo-6-(((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 193-1, 500 mg, 1.25 mmol, 1 eq) in tetrahydrofuran (10 mL) at 0°C. The mixture was stirred at 0°C for 1 hour, and then N,N-dimethylformamide (0.29 mL, 3.75 mmol, 3 eq) was added. The mixture was returned to room temperature and stirred for 1 hour. After the reaction was completed, the reaction solution was concentrated to obtain a crude product, which was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 5:1) to obtain compound 107-1 (320 mg, 73.32%).

[0445] LCMS: (ESI, m / z): 350.2 [M+H] + .

[0446] Step 2: Synthesis of Compound 107-2

[0447] To a solution of 6-(((triisopropylsilyloxy)methyl)thieno[3,2-c]pyridine-3-carbaldehyde (compound 107-1, 300 mg, 0.86 mmol, 1 eq) in dichloromethane (5 mL) was added diethylaminosulfur trifluoride (0.45 mL, 3.44 mmol, 4 eq) at room temperature, and the mixture was stirred at 50°C for 16 hours. After the reaction, the reaction solution was added dropwise to 20 ml of saturated sodium bicarbonate solution, extracted with dichloromethane (20 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product, which was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 3:1) to give compound 107-2 (230 mg, 72.13%).

[0448] LCMS: (ESI, m / z): 372.1 [M+H] + .

[0449] Based on the similar procedures to synthesize compound 193, compound 107 (17.68 mg, 16.14%) was obtained.

[0450] LCMS: (ESI, m / z): 480.2 [M+H] + .

[0451] 1H NMR (400MHz, DMSO-d6) δ12.30–11.94(m,1H),9.10(d,J=39.2Hz,1H),8.87–8.78(m,1 H),8.38–8.29(m,1.6H),8.25(t,J=7.8Hz,1H),8.14(s,0.4H),8.12–8.06(m,1H),7.7 5–7.65(m,1.4H),7.60(s,0.6H),7.41(td,J=54.6,11.8Hz,1H),7.12(s,0.4H),6.66 (s,0.6H),4.96–4.74(m,1H),3.95(s,0.5H),3.26–3.17(m,0.5H),3.07–2.74(m,2H).

[0452] Separation was performed by SFC (System: Waters SFC 150; Column: Chromatographic column specifications: 250*30mm10μm; mobile phase A: supercritical CO2, mobile phase B: methanol (+0.1% 7.0mol / L amine methanol solution); A:B: 55:45; wavelength: 214nm; flow rate: 140mL / min; column temperature: RT; back pressure: 100bar; injection volume: 8.0mL; cycle time: 15.0min), to obtain compound 107-A (front peak, retention time: 2.252min, 3.6mg) and compound 107-B (back peak, retention time: 3.621min, 4.6mg).

[0453] Compound 107-A

[0454] LCMS: (ESI, m / z): 480.2 [M+H] + .

[0455] 1 H NMR(400MHz, DMSO-d6)δ12.14(d,J=34.1Hz,1H),9.21–8.99(m,1H),8.87–8.78(m,1H),8.38–8.06(m,4H),7.75–7.58(m,2H), 7.58–7.23(m,1H),6.89(d,J=184.8Hz,1H),4.96–4.74(m,1H),4.07–3.92(s,0.5H),3.26–3.17(m,0.5H),3.09–2.74(m,2H).

[0456] Compound 107-B

[0457] LCMS: (ESI, m / z): 480.2 [M+H]+ .

[0458] 1 H NMR (400MHz, DMSO-d6) δ12.14(d,J=32.8Hz,1H),9.10(d,J=39.2Hz,1H),8.87–8.78(m,1H),8.38–8.06(m,4H),7.75–7.58(m ,2H),7.57–7.23(m,1H),6.89(d,J=184.8Hz,1H),4.96–4.74(m,1H),3.95(s,0.5H),3.25–3.17(m,0.5H),3.07–2.74(m,2H).

[0459] Using conditions similar to those in the above examples, the compounds listed in Table 9 were prepared. The structural characterization data of these compounds are listed in Table 9.

[0460] Table 9

[0461] Example 13

[0462] Step 1: Synthesis of compound 108-1

[0463] Under nitrogen protection, isopropylmagnesium chloride and lithium chloride (5.78 mL, 1.3 M, 3 eq) were added dropwise to a solution of 3-bromo-6-(((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 193-1, 1 g, 2.51 mmol, 1 eq) in tetrahydrofuran (10 mL) at 0°C. The mixture was stirred at 0°C for 1 hour, and trimethyl borate (783 mg, 7.53 mmol, 3 eq) was added to the system. The mixture was stirred at room temperature for 1 hour. After the reaction was completed, the reaction system was directly injected into C18 silica gel column chromatography for purification (acetonitrile / water (0-35%)) to obtain compound 108-1 (500 mg, 55%).

[0464] LCMS: (ESI, m / z): 366.1 [M+H] + .

[0465] Step 2: Synthesis of compound 108-2

[0466] (6-(((Triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridin-3-yl)boronic acid (Compound 108-1, 500 mg, 1.37 mmol) was dissolved in ethylene glycol dimethyl ether (5 mL), and the atmosphere was replaced with nitrogen. Cuprous iodide (260.92 mg, 1.37 mmol), 1,10-phenanthroline (271.56 mg, 1.37 mmol) and Togni reagent (542.52 mg, The reaction solution was stirred at 35 ° C for 16 hours. After the reaction was completed, water (50 mL) was added to the reaction solution to dilute it, and it was extracted with ethyl acetate (50 mL × 3). The organic phases were combined, washed with saturated brine (50 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 3 / 1) to give compound 108-2 (310 mg, 58.15%).

[0467] LCMS: (ESI, m / z): 390.2 [M+H] + .

[0468] Based on the similar steps to synthesize compound 193, compound 108 (12 mg, 8.69%) was obtained

[0469] LCMS: (ESI, m / z): 498.1 [M+H] + .

[0470] 1H NMR (400MHz, DMSO-d6) δ12.15(d,J=30.2Hz,1H),9.08(d,J=37.5Hz,1H),8.82(d, J=4.4Hz,1H),8.69(d,J=11.8Hz,1H),8.44(s,0.6H),8.30–8.22(m,1.4H),8.15–8 .03(m,1H),7.70(t,J=5.8Hz,1.4H),7.61(s,0.6H),7.13(s,0.4H),6.65(s,0.6H) ,4.95–4.79(m,1H),4.00–3.95(m,0.5H),3.05–3.00(m,0.5H),2.97–2.69(m,2H).

[0471] Separation was performed by SFC (System: Waters SFC 150; Column: Chromatographic column specifications: 250*30mm10μm; mobile phase A: supercritical CO2, mobile phase B: methanol (+0.1% 7.0mol / L amine methanol solution); A:B: 60:40; wavelength: 214nm; flow rate: 120mL / min; column temperature: RT; back pressure: 100bar; injection volume: 5.0mL; cycle time: 15.0min), to obtain compound 108-A (front peak, retention time: 1.583min, 4.3mg) and compound 108-B (back peak, retention time: 2.697min, 3.7mg).

[0472] Compound 108-A

[0473] LCMS: (ESI, m / z): 498.1 [M+H] + .

[0474] 1 H NMR (400MHz, DMSO-d6) δ12.15(s,1H),9.08(d,J=37.5Hz,1H),8.82(d,J=4.4Hz,1H),8.69(d,J=11.8Hz,1H),8.49–8.22(m,2H) ,8.15–8.03(m,1H),7.76–7.57(m,2H),6.89(d,J=192.8Hz,1H),4.95–4.79(m,1H),4.00–3.95(m,0.5H),3.17–2.69(m,2.5H).

[0475] Compound 108-B

[0476] LCMS: (ESI, m / z): 498.1 [M+H] + .

[0477] 1 H NMR (400MHz, DMSO-d6) δ12.22(s,1H),9.08(d,J=37.5Hz,1H),8.82(d,J=4.4Hz,1H),8.69(d,J=11.8Hz,1H),8.47–8.22(m,2H) ,8.15–8.03(m,1H),7.76–7.57(m,2H),6.89(d,J=192.8Hz,1H),4.95–4.77(m,1H),4.00–3.95(m,0.5H),3.17–2.69(m,2.5H).

[0478] Using conditions similar to those in the above examples, the compounds listed in Table 10 were prepared. The structural characterization data of these compounds are listed in Table 10.

[0479] Table 10

[0480] Example 14

[0481] Step 1: Synthesis of compound 243-2

[0482] To a solution of ethyl 5-bromo-1,3,4-oxadiazole-2-carboxylate (compound 243-1, 200 mg, 0.90 mmol, 1 eq) in dichloromethane (3 mL) were added 4-fluoropiperidine hydrochloride (37.7 mg, 0.27 mmol, 1 eq) and N,N-diisopropylethylamine (104.68 mg, 0.81 mmol, 3 eq) at room temperature and stirred at room temperature for 1 hour. After the reaction was completed, water (30 mL) was added to the reaction mixture, and the mixture was extracted with ethyl acetate (3×30 mL) and washed with saturated brine (2×30 mL). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate (0-30%)) to give compound 243-2 (200 mg, 90.8%).

[0483] LCMS: (ESI, m / z): 244.0 [M+H] + .

[0484] Step 2: Synthesis of compound 243-3

[0485] Under nitrogen protection, a solution of ethyl 5-(4-fluoropiperidin-1-yl)-1,3,4-oxadiazole-2-carboxylate (compound 243-2, 200 mg, 0.82 mmol, 1 eq) in dichloromethane (2 mL) was added to a solution of potassium trimethylsilanolate (105.2 mg, 0.82 mmol, 1 eq) in methyl tert-butyl ether (2 mL) at 0°C. The mixture was stirred at 0°C for 2 hours. After the reaction was completed, the reaction mixture was concentrated and the crude product was slurried with methyl tert-butyl ether: dichloromethane = 5:1 (30 mL) to give compound 243-3 (150 mg, 72.0%).

[0486] LCMS: (ESI, m / z): 216.0 [M-K+2H] + .

[0487] Step 3: Synthesis of Compound 243

[0488] To a solution of potassium 5-(4-fluoropiperidin-1-yl)-1,3,4-oxadiazole-2-carboxylate (compound 243-3, 100 mg, 0.39 mmol, 1 eq) and 4-(thieno[3,2-c]pyridin-6-yl)-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine (compound 94-3, 100 mg, 0.39 mmol, 1 eq) in pyridine (3 mL) was added dropwise phosphorus oxychloride (179.4 mg, 1.17 mmol, 3 eq) at 0°C under nitrogen atmosphere. The mixture was stirred at room temperature for 1 hour. After the reaction was complete, the reaction solution was dried and the crude product was prepared by HPLC under the following conditions: (XBridge C18 The purified product was purified by HPLC (19*250mm, 10μm; mobile phase A: 0.03% ammonia water, mobile phase B: acetonitrile; flow rate: 20 ml / min; gradient: 29% to 29%; retention time: 8.0–9.0 min / 16 min) to obtain compound 243 (12.5 mg, 6.9%).

[0489] LCMS: (ESI, m / z): 454.2 [M+H] + .

[0490] Using conditions similar to those in the above examples, the compounds listed in Table 11 were prepared. The structural characterization data of these compounds are listed in Table 11.

[0491] Table 11 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0492] Example 15

[0493] Step 1: Synthesis of compound 244-2

[0494] At room temperature, 3-bromo-2-fluoroaniline (compound 244-1, 5 g, 26.31 mmol, 1 eq) was added to water (100 mL), concentrated hydrochloric acid (200 mL) was added, the temperature was raised to 90 ° C to dissolve, the temperature was lowered to 0 ° C, sodium nitrite (2.00 g, 28.94 mmol, 1.1 eq) aqueous solution (40 mL) was added dropwise, and stirred at 0 ° C for 1 hour. Ethoxy (potassium sulfide) methylthioketone (5.06 g, 31.57 mmol, 1.2 eq) aqueous solution (100 mL) was added dropwise, and stirred at room temperature for 2 hours. After the reaction was completed, the aqueous phase was washed with water. The mixture was extracted three times with dichloromethane (200 mL), the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated. The crude product was dissolved in ethanol (100 mL), potassium hydroxide (2.95 g, 52.62 mmol, 2 eq) was added, stirred at room temperature for 1 h, concentrated, added with water (200 mL), extracted with dichloromethane (200 mL), the aqueous phase was adjusted to pH 3 with 2M HCl, and the aqueous phase was extracted with ethyl acetate (200 mL×2). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated to give compound 244-2 (2.4 g, 44.5%), which was directly used for the next reaction.

[0495] LCMS: (ESI, m / z): 206.6 [MH].

[0496] Step 2: Synthesis of compound 244-3

[0497] At room temperature, 3-bromo-2-fluorobenzene-1-thiol (compound 244-2, 2.4 g, 11.59 mmol, 1 eq) was dissolved in N,N-dimethylformamide (30 mL), and 2-bromo-1,1-dimethoxyethane (1.96 g, 11.59 mmol, 1 eq), potassium carbonate (3.20 g, 23.18 mmol, 2 eq) and tetrabutylammonium iodide (0.43 g, 1.16 mmol, 0.1 eq) were added. The mixture was stirred at room temperature overnight. After the reaction was completed, the reaction solution was quenched with 100 mL of water, extracted with ethyl acetate (50 mL), dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography (PE / EA: 0-10%) to give compound 244-3 (1.8 g, 52.61%).

[0498] Step 3: Synthesis of Compound 244-4

[0499] At room temperature, 1-bromo-3-[(2,2-dimethoxyethyl)thio]-2-fluorobenzene (compound 244-3, 2.3 g, 7.79 mmol, 1 eq) was dissolved in toluene (30 mL), and polyphosphoric acid (3.5 g) was added. The reaction temperature was raised to 120°C and stirred for 16 h. After the reaction was completed, the reaction solution was filtered, the filtrate was concentrated, and the crude product was purified by silica gel column chromatography (100% PE) to give compound 244-4 (650 mg, 36.10%).

[0500] GCMS:(ESI,m / z):230[M].

[0501] Step 4: Synthesis of compound 244-5

[0502] At room temperature, 6-bromo-7-fluoro-1-benzothiophene (compound 244-4, 450 mg, 1.95 mmol, 1 eq) was dissolved in ultra-dry tetrahydrofuran (7 mL), cooled to -78 ° C. Under nitrogen protection, n-butyl lithium (0.86 mL, 2.15 mmol, 1.1 eq) was added, and the reaction solution was stirred at -78 ° C for half an hour. Ultra-dry N,N-dimethylformamide (213.79 mg, 2.92 mmol) was added and stirred for 1 hour. The reaction solution was quenched with saturated aqueous ammonium chloride (30 mL) and extracted with ethyl acetate (20 mL × 2). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (100% PE) to give compound 244-5 (170 mg, 48.44%).

[0503] GCMS:(ESI,m / z):180[M].

[0504] Based on the similar procedures to synthesize compound 103, compound 244 (20.72 mg, 10.57%) was obtained.

[0505] LCMS: (ESI, m / z): 447.2 [M+H] + .

[0506] 1 H NMR (400MHz, DMSO-d6) δ12.30–12.10(m,1H),8.86–8.79(m,1H),8.32–8.23(m,1H),8.19–8.06(m,1H),7.75–7.66(m,3H),7.48 –7.37(m,2H),7.31–7.14(m,1.4H),6.74(s,0.6H),4.95–4.70(m,1H),3.57–3.39(m,1H),3.11–2.99(m,1H),2.89–2.73(m,1H).

[0507] Example 16

[0508] Step 1: Synthesis of compound 185-2

[0509] To a solution of nicotinate ethyl ester (compound 185-1, 1.5 g, 9.92 mmol) in ethanol (10 mL) was added hydrazine hydrate (1.99 g, 39.68 mmol) and reacted at room temperature for 2 h. After the reaction was completed, the reaction solution was concentrated to obtain compound 185-2 (1.3 g, 95.53%).

[0510] LCMS: (ESI, m / z): 138.0 [M+H] + .

[0511] Step 2: Synthesis of compound 185-3

[0512] Nicotinic acid hydrazide (compound 185-2, 1.3 g, 9.48 mmol) was dissolved in dichloromethane (20 mL), and triethylamine (3.94 mL, 28.44 mmol) and monomethyl oxalyl chloride (1.16 g, 9.48 mmol) were added at 0°C. The mixture was reacted at room temperature for 2 h. After the reaction was completed, the reaction solution was concentrated to obtain compound 185-3 (700 mg, 33.09%).

[0513] LCMS: (ESI, m / z): 224.1 [M+H] + .

[0514] Step 3: Synthesis of compound 185-4

[0515] To a solution of methyl 2-(2-nicotinoylhydrazide)-2-oxoacetate (compound 185-3, 700 mg, 3.14 mmol) in dichloromethane (10 mL) were added triethylamine (1.31 mL, 9.42 mmol) and p-toluenesulfonyl chloride (599 mg, 3.14 mmol), and the mixture was reacted at room temperature for 2 h. After the reaction was completed, the reaction solution was concentrated to obtain a crude product, which was purified by normal phase purification (petroleum ether: ethyl acetate = 1:1) to obtain compound 185-4 (540 mg, purity 50%, 41.96%).

[0516] LCMS: (ESI, m / z): 206.0 [M+H] + .

[0517] Step 4: Synthesis of compound 185-5

[0518] To a solution of methyl 5-(pyridin-3-yl)-1,3,4-oxadiazole-2-carboxylate (compound 185-4, 70 mg, 0.34 mmol) in dichloromethane (1 mL) and methyl tert-butyl ether (1 mL) was added potassium trimethylsilanol (44 mg, 0.34 mmol) and reacted at room temperature for 2 h. After the reaction was completed, the reaction solution was concentrated and methyl tert-butyl ether (2 mL) was added. The mixture was stirred for 15 min and filtered. The filter cake was concentrated in vacuo to give compound 185-5 (75 mg, 95.90%).

[0519] LCMS: (ESI, m / z): 192.0 [M+H] + .

[0520] Step 4: Synthesis of Compound 185

[0521] To a solution of potassium 5-(pyridin-3-yl)-1,3,4-oxadiazole-2-carboxylate (compound 185-5, 75 mg, 0.33 mmol) in pyridine (1 mL) was added 6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[3,2-c]pyridine (compound 094-3, 85 mg, 0.33 mmol), and phosphorus oxychloride (152 mg, 0.99 mmol) was added at 0°C. The mixture was reacted at room temperature for 12 h. After the reaction was completed, the reaction solution was concentrated and purified by reverse phase chromatography (acetonitrile: water = 1:1) to obtain a crude product. The crude product was then subjected to reverse phase preparative chromatography (conditions as follows: chromatography column specifications: prep-HPLC (Waters 2767 / QDA), column: SunFire C18, 19*250mm, 10μm; mobile phase A: 0.1% formic acid / water, mobile phase B: acetonitrile; flow rate: 20mL / min; elution gradient: 52% to 52%; retention time: 8.4-9.5min) to obtain compound 185 (11.7mg, 8.33%).

[0522] LCMS: (ESI, m / z): 430.1 [M+H] + .

[0523] Example 17

[0524] Step 1: Synthesis of compound 183-2

[0525] At room temperature, (dimethoxymethyl)dimethylamine (5.03 g, 42.20 mmol) was slowly added to methyl 3-cyclopropyl-3-oxopropanoate (compound 183-1, 5.0 g, 35.17 mmol), and the reaction solution was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was concentrated to obtain crude compound 183-2 (6.94 g), which was directly used in the next reaction.

[0526] 1 H NMR (400MHz, CDCl3) δ7.66–7.59(m,1H),3.77(s,3H),3.20–2.68(m,6H),2.44–2.36(m,1H),1.07–1.00(m,2H),0.88–0.78(m,2H).

[0527] Step 2: Synthesis of compound 183-3

[0528] At room temperature, methylhydrazine sulfate (5.07 g, 35.14 mmol) was dissolved in ethanol (300 mL), and cesium carbonate (22.92 g, 70.34 mmol) was added. The reaction solution was stirred at room temperature for 10 minutes, and then (2E)-2-[(E)-cyclopropylcarbonyl]-3-(dimethylamino)prop-2-enoic acid methyl ester (compound 183-2, 6.58 g, 33.38 mmol) was added. The reaction solution was stirred at 90°C for 16 hours. After the reaction was completed, it was filtered through diatomaceous earth and washed with ethyl acetate (100 mL). The filtrate was concentrated to obtain a crude product, which was purified by normal phase column chromatography (PE:EA=100:0-3) to obtain compound 183-3 (94 g).

[0529] 1 H NMR (400MHz, CDCl3) δ7.74(s,1H),3.81(s,3H),3.79(s,3H),2.53–2.45(m,1H),0.97–0.90(m,4H).

[0530] Based on the similar procedures to synthesize compound 162, compound 183 (85 mg, 16.3%) was obtained.

[0531] LCMS: (ESI, m / z): 473.2 [M+H] + .

[0532] 1 H NMR (400MHz, DMSO-d6) δ12.43(s,1H),9.06(d,J=30.0Hz,1H),8.17(d,J=31.7Hz,1H) ,8.01–7.84(m,2H),7.70(d,J=30.3Hz,1H),7.59(dd,J=12.8,5.5Hz,1H),6.89(d,J= 193.7Hz,1H),4.83(ddd,J=17.9,13.3,5.1Hz,1H),3.96–3.89(m,3.5H),3.21–3.14( m,0.5H),3.05–2.75(m,2H),2.02–1.93(m,1H),1.12–1.05(m,2H),0.88–0.79(m,2H).

[0533] Using conditions similar to those in the above examples, the compounds listed in Table 12 were prepared. The structural characterization data of these compounds are listed in Table 12.

[0534] Table 12 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0535] Example 18

[0536] Step 1: Synthesis of compound 192-2

[0537] Under nitrogen protection, to a solution of 2-cyanobenzoic acid (compound 192-1, 2 g, 13.59 mmol, 1 eq) in dichloromethane (20 mL) was added (isocyanimido)triphenylphosphane (4.1 g, 13.59 mmol, 1 eq) at room temperature, and the mixture was stirred at room temperature overnight. After the reaction was completed, the reaction mixture was concentrated and the crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate (0-60%)) to give compound 192-2 (300 mg, 12.9%).

[0538] LCMS: (ESI, m / z): 172.1 [M+H] + .

[0539] Step 1: Synthesis of compound 192-3

[0540] Under carbon dioxide gas protection, potassium tert-butoxide (589.1 mg, 5.25 mmol, 3 eq) and 18-crown-6 (1387.6 mg, 5.25 mmol, 3 eq) were added to a solution of 2-(1,3,4-oxadiazol-2-yl)benzonitrile (compound 192-2, 300 mg, 1.75 mmol, 1 eq) in N,N-dimethylformamide (5 mL) at room temperature. The mixture was stirred at room temperature for 2 hours, and iodomethane (745.18 mg, 5.25 mmol, 3 eq) was added dropwise. The mixture was then stirred at room temperature for 16 hours. After the reaction was complete, the mixture was quenched with water (30 mL), extracted with ethyl acetate (30 mL×3), washed with saturated brine (30 mL×2), dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate (0-40%)) to give compound 192-3 (250 mg, 62.2%).

[0541] LCMS: (ESI, m / z): 230.0 [M+H] + .

[0542] Based on the similar procedure to synthesize compound 162, compound 192 (85 mg, 16.3%) was obtained.

[0543] LCMS: (ESI, m / z): 454.2 [M+H] + .

[0544] 1H NMR (400MHz, DMSO-d6) δ12.25–11.95(m,1H),9.08(d,J=26.6Hz,1H),8.23(d,J=7.1Hz,1H),8.17(d,J=7.5Hz,1H),8.08(d,J=44.3Hz,1H),7.99–7. 86(m,3H),7.69–7.57(m,2H),6.83(d,J=150.8Hz,1H),4.81(dd,J=12.9, 4.9Hz,1H),4.03–3.93(m,0.5H),3.27–3.19(m,0.5H),3.08–2.76(m,2H).

[0545] Using conditions similar to those in the above examples, the compounds listed in Table 13 were prepared. The structural characterization data of these compounds are listed in Table 13.

[0546] Table 13 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0547] Example 19

[0548] Step 1: Synthesis of compound 165-2

[0549] 1,4-Dimethyl-1H-pyrazole-3-carboxylic acid (compound 165-1, 1 g, 7.14 mmol) was dissolved in dichloromethane (10 mL), oxalyl chloride (1.09 g, 8.57 mmol) was added at 0°C, and N,N-dimethylformamide (0.052 g, 0.71 mmol) was added dropwise. The reaction solution was stirred at room temperature for 1 hour. After the reaction was completed, it was concentrated to obtain compound 165-2 (1.1 g, 97%).

[0550] Step 2: Synthesis of compound 165-3

[0551] 1,4-Dimethyl-1H-pyrazole-3-carbonyl chloride (compound 165-2, 1.1 g, 6.94 mmol) was dissolved in tetrahydrofuran (20 mL), and ethyl (hydrazinecarbonyl)formate (1.01 g, 7.63 mmol) and N,N-diisopropylethylamine (2.69 g, 20.82 mmol) were added. The reaction solution was stirred at 60°C for 1 hour. After the reaction was completed, the reaction solution was concentrated, and methyl tert-butyl ether (50 mL) and ethyl acetate (5 mL) were added. The mixture was vigorously stirred for 30 minutes and filtered to obtain compound 165-3 (1.7 g, 96.40%).

[0552] LCMS: (ESI, m / z): 255.1 [M+H] + .

[0553] Based on the similar procedures to synthesize compound 162, compound 165 (10 mg, 1.9%) was obtained.

[0554] LCMS: (ESI, m / z): 477.2 [M+H] + .

[0555] 1 H NMR(400MHz, DMSO-d6)δ12.11(d,J=37.2Hz,1H),9.20–8.95(m,1H),8.25–7. 96(m,1H),7.88(dd,J=10.7,5.4Hz,1H),7.78(s,1H),7.72–7.53(m,2H),6.93 (dd,J=197.6,58.7Hz,1H),4.95–4.72(m,1H),4.02–3.96(m,0.5H),3.92(d,J =6.2Hz,3H),3.19–3.11(m,0.5H),3.06–2.74(m,2H),2.30(d,J=11.0Hz,3H).

[0556] Using conditions similar to those in the above examples, the compounds listed in Table 14 were prepared. The structural characterization data of these compounds are listed in Table 14.

[0557] Table 14 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0558] Example 20

[0559] Step 1: Synthesis of compound 168-2

[0560] 1-Ethyl-1H-pyrazole-3-carboxylic acid (compound 168-1, 2 g, 14.27 mmol, 1 eq) was dissolved in DMF (15 mL), and (tert-butoxy)formic hydrazide (2.26 g, 17.12 mmol, 1.2 eq), HATU (6.51 g, 17.12 mmol, 1.2 eq) and DIEA (7.09 mL, 42.81 mmol, 3 eq) were added. The reaction solution was stirred at room temperature for 16 hours. After the reaction was completed, water (50 mL) was added, and the mixture was washed with ethyl acetate (50 mL×3). It was washed with saturated sodium chloride aqueous solution (50 mL), dried over anhydrous sodium sulfate, filtered, and dried to give the crude product, which was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 2:1) to give compound 168-2 (3.5 g, 96.44%).

[0561] LCMS: (ESI, m / z): 199.1 [M-56+H] + .

[0562] Step 2: Synthesis of compound 168-3

[0563] Tert-butyl 2-(1-ethyl-1H-pyrazole-3-carbonyl)hydrazine-1-carboxylate (compound 168-2, 3.5 g, 13.76 mmol, 1 eq) was dissolved in a 1,4-dioxane solution of hydrochloric acid (35 mL), and the reaction solution was stirred at room temperature for 1 hour. After the reaction was completed, the reaction solution was concentrated to obtain compound 168-3 (2 g, 94.25%).

[0564] LCMS: (ESI, m / z): 155.1 [M+H] + .

[0565] Based on the similar procedures to synthesize compound 162, compound 168 (16.8 mg, 3.09%) was obtained.

[0566] LCMS: (ESI, m / z): 447.2 [M+H] + .

[0567] 1H NMR (400MHz, DMSO-d6) δ12.16 (s, 1H), 9.06 (d, J = 36.8Hz, 1H), 8.19 (d, J = 14.4Hz, 1H),8.05(t,J=2.2Hz,1H),7.88(dd,J=10.7,5.5Hz,1H),7.72–7.52(m,2H),7.15 (s,0.4H),6.99–6.94(m,1H),6.64(s,0.6H),4.97–4.69(m,1H),4.39–4.22(m,2H ),4.00–3.86(m,0.5H),3.19–3.15(m,0.5),3.05–2.72(m,2H),1.48–1.37(m,3H).

[0568] Using conditions similar to those in the above examples, the compounds listed in Table 15 were prepared. The structural characterization data of these compounds are listed in Table 15.

[0569] Table 15

[0570] Example 21

[0571] Based on the similar procedures to synthesize compound 183, compound 182 (104.58 mg, 16.52%) was obtained.

[0572] LCMS: (ESI, m / z): 461.3 [M+H] + .

[0573] Example 22

[0574] Step 1: Synthesis of compound 218-2

[0575] To a solution of 2-chlorothieno[2,3-d]pyrimidine (compound 218-1, 1.5 g, 8.79 mmol, 1.0 eq) in methanol (20 mL) were added [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (640 mg, 0.88 mmol, 0.1 eq) and triethylamine (1.78 g, 17.58 mmol, 2.0 eq). The reaction was stirred at 120°C under a carbon monoxide atmosphere for 16 hours. After completion of the reaction, the mixture was diluted with water (50 mL) and extracted with ethyl acetate (500 mL × 3). The organic phases were combined, washed with saturated brine (30 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to give the crude product. The crude product was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 10:1) to give compound 218-1 (1.4 g, 82.00%).

[0576] LCMS: (ESI, m / z): 195.1 [M+H] + .

[0577] Based on the similar procedures to synthesize compound 105, compound 218 (10 mg, 1.9%) was obtained.

[0578] LCMS: (ESI, m / z): 431.2 [M+H] + .

[0579] 1 H NMR (400MHz, DMSO-d6) δ12.29–11.97(m,1H),9.33(dd,J=29.0,20.3Hz,1H),8.80(dt,J=17.9,5.7Hz,1H),8.31–8.03(m,2H),8.02–7.92(m,1H),7.7 3–7.65(m,1H),7.64–7.50(m,2H),6.88(dd,J=148.3,110.7Hz,1H),5.04– 4.87(m,1H),4.47–4.31(m,0.5H),4.04–3.77(m,0.5H),3.10–2.63(m,2H).

[0580] Example 23

[0581] Step 1: Synthesis of compound 190-2

[0582] To a solution of methyl 6-aminopyridine-2-carboxylate (compound 190-1, 3 g, 19.72 mmol) and sodium bicarbonate (1.1 g, 13.2 mmol) in acetonitrile (18 mL) and water (12 mL) was added benzyl chloroformate (3.06 mL, 21.69 mmol) at 0°C and stirred at room temperature for 16 hours. After the reaction was complete, the mixture was diluted with water and filtered to obtain compound 190-2 (3.5 g, 62.01%).

[0583] LCMS: (ESI, m / z): 287.1 [M+H] + .

[0584] Based on the similar steps to synthesize compound 183, compound 190-7 (120 mg, 19.6%) was obtained

[0585] Step 7: Synthesis of Compound 190

[0586] To a solution of benzyl N-(6-(5-(4-(thieno[3,2-c]pyridin-6-yl)-1,4,6,7-tetrahydro-5H-imidazo[4,5-c]pyridine-5-carbonyl)-1,3,4-oxadiazol-2-yl)pyridin-2-yl)carbamate (compound 190-7, 80 mg, 0.14 mmol) in acetic acid (1 mL) was added hydrogen bromide (0.5 mL) and the reaction was completed in 2 hours. The mixture was concentrated to give a crude product, which was then subjected to reverse phase preparative chromatography (conditions as follows: chromatography column specifications: prep-HPLC (Waters 2767 / QDA), column: SunFire C18, 19×250 mm, 10 μm; mobile phase A: 0.1% formic acid / water, mobile phase B: acetonitrile; flow rate: 20 mL / min; elution gradient: 60% to 70%; retention time: 5.5 -9.5 min) to afford compound 190 (4.52 mg, 7.36%).

[0587] LCMS: (ESI, m / z): 445.1 [M+H] + .

[0588] 1 H NMR (400MHz, DMSO-d6) δ12.18(s,1H),9.06(d,J=39.1Hz,1H),8.09(d,J=92.2Hz,1H),7.88(dd,J=12.3,5.4Hz,1H),7.71–7.54(m,3H),7.35(t,J=6.9 Hz,1H),7.16(s,0.4H),6.71–6.60(m,1.6H),6.52(d,J=15.5Hz,2H),4.90– 4.73(m,1H),3.94–3.86(m,0.5H),3.16–3.12(m,0.5H),3.04–2.73(m,2H).

[0589] Using conditions similar to those in the above examples, the compounds listed in Table 16 were prepared. The structural characterization data of these compounds are listed in Table 16.

[0590] Table 16

[0591] Example 24

[0592] Step 1: Synthesis of compound 133-2

[0593] To a solution of thieno[3,2-c]pyridin-6-ylmethanol (compound 133-1, 1 g, 6.05 mmol) in dichloromethane (12 mL) was added triisopropylsilyl chloride (1.75 g, 9.07 mmol) at room temperature and stirred at room temperature overnight. After the reaction was completed, the reaction mixture was concentrated and the residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate (0-20%)) to give compound 133-2 (1.9 g, 97.6%).

[0594] LCMS: (ESI, m / z): 322.2 [M+H] + .

[0595] Step 2: Synthesis of compound 133-3

[0596] Under nitrogen protection, m-chloroperbenzoic acid (805.0 mg, 4.67 mmol, 1.5 eq) was added portionwise to a solution of 6-(((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 133-2, 1000 mg, 3.11 mmol, 1 eq) in dichloromethane (12 mL) at room temperature. The mixture was stirred at room temperature for 2 hours. After the reaction was completed, the reaction mixture was quenched with saturated aqueous sodium sulfite solution (50 mL), extracted with ethyl acetate (50 mL×3), washed with saturated brine (50 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate (0-100%)) to give compound 133-3 (800 mg, 76.2%).

[0597] LCMS: (ESI, m / z): 338.2 [M+H] + .

[0598] Step 3: Synthesis of compound 133-4

[0599] Under nitrogen protection, phosphorus oxybromide (409.4 mg, 1.43 mmol, 1.2 eq) and DMF (43.5 mg, 0.59 mmol, 0.5 eq) were added to a solution of 6-(((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine 5-oxide (compound 133-3, 400 mg, 1.19 mmol, 1 eq) in dichloromethane (6 mL) at 0°C and stirred at room temperature for 12 hours. After the reaction, the reaction mixture was adjusted to pH 7-8 with saturated aqueous sodium bicarbonate solution, extracted with dichloromethane (50 mL×3), washed with saturated brine (50 mL×2), dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate (0-50%)) to give compound 133-4 (250 mg, 52.6%).

[0600] LCMS: (ESI, m / z): 402.2 [M+2+H] + .

[0601] Step 4: Synthesis of compound 133-5

[0602] To a solution of 4-bromo-6-(((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 133-4, 250 mg, 0.62 mmol, 1 eq) in 1,4-dioxane (3 mL) and water (0.1 mL) was added trimethylboroxane (155.6 mg, 1.24 mmol, 2 eq), [1,1'-bis(di-tert-butylphosphino)ferrocene]palladium(II) dichloride (40.4 mg, 0.062 mmol, 1 eq) at room temperature under nitrogen atmosphere. mmol, 0.1eq) and potassium phosphate (263.21 mg, 1.24 mmol, 2eq), stirred at 100 ° C for 4 hours. After the reaction, water (20 mL) was added to the reaction mixture to quench it, extracted with ethyl acetate (20 mL × 3), washed with saturated brine (20 mL × 2), dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate (0-50%)) to give compound 133-5 (145 mg, 69.2%).

[0603] LCMS: (ESI, m / z): 336.3 [M+H] + .

[0604] Based on the similar procedures to synthesize compound 193, compound 133 (120 mg, 19.6%) was obtained.

[0605] LCMS: (ESI, m / z): 444.2 [M+H] + .

[0606] 1 H NMR(400MHz,DMSO-d6)δ12.30–11.91(m,1H),8.83(dd,J=12.7,4.4Hz,1H),8.3 1–8.22(m,1H),8.15–8.06(m,1H),7.99(s,0.5H),7.85(dd,J=11.7,5.5Hz,1H), 7.72–7.55(m,3.5H),6.78(d,J=147.2Hz,1H),4.93–4.67(m,1H),4.16–3.92(m ,0.4H),3.16–3.05(m,0.6H),3.03–2.78(m,2H),2.75(s,1.5H),2.56(s,1.5H).

[0607] Example 25

[0608] Step 1: Synthesis of compound 255-2

[0609] Under nitrogen protection, 1H-imidazole-5-carboxaldehyde (compound 255-1, 2 g, 20.81 mmol) was dissolved in nitroethane (15 mL, 208.1 mmol), and cyclohexylamine (12 mL, 104.05 mmol) was added at 0°C. The reaction solution was stirred at 0°C for 20 minutes, and glacial acetic acid (36 mL, 624.3 mmol) was added. The reaction solution was stirred at 70°C for 2 hours. After the reaction was completed, the reaction solution was concentrated, diluted with saturated sodium bicarbonate (100 mL), and extracted with ethyl acetate (100 mL×3). The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1 / 1) to give compound 255-2 (1.8 g, 56.47%).

[0610] LCMS: (ESI, m / z): 154.0 [M+H] + .

[0611] Step 2: Synthesis of compound 255-3

[0612] (Z)-5-(2-nitroprop-1-en-1-yl)-1H-imidazole (compound 255-2, 800 mg, 5.22 mmol) was dissolved in tetrahydrofuran (10 mL), and lithium aluminum tetrahydride (594.30 mg, 15.66 mmol) was added. The reaction solution was stirred at 70°C for 16 hours. After the reaction was completed, it was cooled to room temperature. Saturated anhydrous sodium sulfate solution (2 mL) and ethyl acetate (50 mL) were added dropwise to the reaction system. A large amount of anhydrous sodium sulfate was added, and the mixture was vigorously stirred for 1 hour. After filtration and concentration, compound 255-3 (400 mg, 61.17%) was obtained.

[0613] LCMS: (ESI, m / z): 126.2 [M+H] + .

[0614] Step 3: Synthesis of compound 255-4

[0615] Compound 1-(1H-imidazol-5-yl)propan-2-amine (compound 255-3, 300 mg, 2.40 mmol) was dissolved in a mixed solvent of ethanol (3 mL) and water (3 mL), and thieno[3,2-c]pyridine-6-carboxylic acid (compound 094-3, 470.02 mg, 2.88 mmol) and sodium hydroxide (288 mg, 7.20 mmol) were added. The reaction solution was stirred at 80°C for 1 hour. After cooling, 1M hydrochloric acid solution was added to adjust the pH to 6-7. The reaction solution was purified by C18 reverse phase (0.05% ammonia water) to obtain compounds 255-4-A (40 mg, 6.18%) and 255-4-B (10 mg, 1.54%).

[0616] 255-4-A

[0617] LCMS: (ESI, m / z): 271.1 [M+H] + .

[0618] 255-4-B

[0619] LCMS: (ESI, m / z): 271.1 [M+H] + .

[0620] Step 4: Synthesis of compound 255

[0621] Compound 255-4-A (35 mg, 0.13 mmol, 1 eq) and potassium 5-(pyridin-2-yl)-1,3,4-oxadiazole-2-carboxylate (compound 105-4, 35.76 mg, 0.16 mmol, 1.2 eq) were dissolved in pyridine (1 mL). Under nitrogen protection, phosphorus oxychloride (0.036 mL, 0.39 mmol, 3 eq) was added dropwise at 0°C. The mixture was stirred and reacted at 0°C for 1 hour. After the reaction was completed, the crude product was purified by high performance liquid chromatography (column: XBridge C18 19*250 mm, 10 μm; mobile phase A: 10 mmol aqueous ammonium bicarbonate solution, mobile phase B: acetonitrile; flow rate: 20 mL / min; gradient: 45% to 47%; retention time: 7.50-9.50 min / 16 min) to obtain compound 255-A (5.37 mg, 9.35%).

[0622] LCMS: (ESI, m / z): 444.1 [M+H] + .

[0623] Compound 255-B was synthesized from compound 255-4-B using the same method.

[0624] LCMS: (ESI, m / z): 444.2 [M+H] + .

[0625] Using conditions similar to those in the above examples, the compounds listed in Table 17 were prepared. The structural characterization data of these compounds are listed in Table 17.

[0626] Table 17

[0627] Example 26

[0628] Step 1: Synthesis of compound 222-1

[0629] At room temperature, N-chlorosuccinimide (1866.75 mg, 13.98 mmol, 3 eq) was added to a solution of methyl thieno[2,3-b]pyridine-6-carboxylate (compound 132-4, 900 mg, 4.66 mmol, 1 eq) in N,N-dimethylformamide (10 mL) and the reaction was allowed to proceed for 5 hours. After completion of the reaction, the reaction solution was diluted with 100 ml of water and extracted twice with an equal amount of ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by dry-phase silica gel column chromatography (petroleum ether / ethyl acetate = 9:1) to give compound 222-1 (850 mg, 80.15%).

[0630] LCMS: (ESI, m / z): 228.0 [M+H] + .

[0631] Step 2: Synthesis of compound 222-2

[0632] 3-Chlorothieno[2,3-b]pyridine-6-carboxylic acid methyl ester (compound 222-1, 750 mg, 3.29 mmol, 1 eq) was dissolved in anhydrous toluene (30 mL) and protected with nitrogen. 1 mol / L diisobutylaluminum hydride toluene solution (9.87 mL, 9.87 mmol, 3 eq) was added at -78 ° C. The mixture was reacted for 2 hours at -78 ° C. After the reaction, 10 ml of methanol was added to the reaction solution at -78 ° C., and then 80 ml of saturated brine was added to quench the reaction. Then 100 ml of ethyl acetate was added, filtered, and the filtrate was separated. The organic phase was dried over anhydrous sodium sulfate, filtered, and dried to give a crude compound 222-2 (650 mg), which was directly used in the next reaction.

[0633] LCMS: (ESI, m / z): 198.0 [M+H] + .

[0634] Step 3: Synthesis of compound 222-3

[0635] 3-Chlorothieno[2,3-b]pyridine-6-carbaldehyde (compound 222-2, 200 mg, 1.01 mmol, 1 eq) and histamine dihydrochloride (223.09 mg, 1.21 mmol, 1.2 eq) were dissolved in ethanol (4 mL), and a solution of sodium hydroxide (121.20 mg, 3.03 mmol, 3 eq) in water (4 mL) was added. The mixture was reacted at 80° C. for 16 hours. After the reaction was completed, the reaction solution was dried and the crude product was purified by silica gel column chromatography (dichloromethane / methanol = 4:1) to give compound 222-3 (300 mg).

[0636] LCMS: (ESI, m / z): 291.1 [M+H] + .

[0637] Step 4: Synthesis of compound 222

[0638] 3-Chloro-6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[2,3-b]pyridine (compound 222-3, 300 mg, 1.03 mmol, 1 eq) and 5-(pyridin-2-yl)-1,3,4-oxadiazole-2-carboxylate lithium ester (compound 094-5, 213.14 mg, 1.08 mmol, 1.05 eq) were dissolved in pyridine (10 mL), phosphorus oxychloride (473.79 mg, 3.09 mmol, 3 eq) was added at 0°C, and the mixture was reacted at 25°C for 2 hours. After the reaction, the reaction solution was dried and purified by silica gel column chromatography (dichloromethane / methanol = 4:1). The mixture was then slurried with methanol and filtered. The filter cake was compound 222 (10 mg), which was separated by SFC (System: Waters SFC 150; chromatographic column: Chromatographic column specifications: 250×30mm 10μm; mobile phase A: supercritical CO2, mobile phase B: methanol (+0.1% 7.0mol / L amine methanol solution); A:B=75:25; wavelength: 214nm; flow rate: 140mL / min; column temperature: RT; back pressure: 100bar; injection volume: 1.0mL; cycle time: 4.3min; solvent: methanol: distillation grade, supercritical CO2: food grade) to obtain compounds 222-A (front peak, retention time: 3.424min, 22.25mg) and 222-B (back peak, retention time: 4.566min, 28.71mg).

[0639] Compound 222-A

[0640] LCMS: (ESI, m / z): 464.1 [M+H] + .

[0641] 1H NMR(400MHz,MeOD-d4)δ8.77(d,J=4.8Hz,1H),8.29(t,J=6.7Hz,1H),8.20( dd,J=30.4,8.4Hz,1H),8.11–8.03(m,1H),7.88–7.61(m,4H),7.12(d,J=193 .2Hz,1H),5.02(ddd,J=27.0,13.5,5.1Hz,1H),4.04–3.92(m,0.6H),3.43–3 .34(m,0.4H),3.23–3.15(m,0.6H),3.07–2.95(m,0.4H),2.92–2.80(m,1H).

[0642] Compound 222-B

[0643] LCMS: (ESI, m / z): 464.1 [M+H] + .

[0644] 1 H NMR(400MHz,DMSO-d6)δ12.15(s,1H),8.86–8.78(m,1H),8.32–8.17(m,2H),8.13–7.98(m,2H),7.95–7.76(m,1H),7.73 –7.60(m,2H),6.88(d,J=161.2Hz,1H),4.97–4.85(m,1H),3.98–3.79(m,0.6H),3.39–3.34(m,0.4H),3.12–2.74(m,2H).

[0645] Example 27

[0646] Step 1: Synthesis of compound 260-2

[0647] 2-Methylthiazole-4-carboxylic acid (compound 260-1, 2 g, 13.97 mmol, 1 eq) was dissolved in thionyl chloride (30 mL) and refluxed at 60° C. overnight. After the reaction was completed, the reaction solution was directly spin-dried to obtain compound 260-2 (2.2 g, crude product).

[0648] Step 2: Synthesis of compound 260-3

[0649] To a solution of 2-methylthiazole-4-carbonyl chloride (compound 260-2, 2.2 g, 13.61 mmol, 1 eq) in dichloromethane (25 mL) were added ethyl 2-hydrazino-2-oxoacetate (2.16 g, 16.33 mmol, 1.2 eq) and triethylamine (5.66 mL, 40.83 mmol, 3 eq) at 0°C. The mixture was stirred at 0°C for 1 hour. After the reaction was complete, the reaction solution was used directly in the next step without further treatment.

[0650] LCMS: (ESI, m / z): 258.0 [M+H] + .

[0651] Step 3: Synthesis of compound 260-4

[0652] To a dichloromethane solution (25 mL) of ethyl 2-(2-(2-methylthiazole-4-carbonyl)hydrazine)-2-oxoacetate (compound 260-3, 3.5 g, 13.60 mmol, 1 eq) was added triethylamine (5.66 mL, 40.8 mmol, 3 eq) at 0°C. After stirring, p-toluenesulfonyl chloride (3.11 g, 16.32 mmol, 1.2 eq) was added at 0°C. The mixture was stirred at 0°C for 1 hour and returned to room temperature overnight. After the reaction was completed, water (100 mL) was added to the reaction solution to quench the mixture. The mixture was extracted with ethyl acetate (100 mL×3). The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1 / 4) to give compound 260-4 (830 mg, 25.5%).

[0653] LCMS: (ESI, m / z): 240.0 [M+H] + .

[0654] Step 4: Synthesis of compound 260-5

[0655] To a solution of ethyl 5-(2-methylthiazol-4-yl)-1,3,4-oxadiazole-2-carboxylate (compound 260-4, 830 mg, 3.47 mmol, 1 eq) in dichloromethane (8 mL) and methyl tert-butyl ether (8 mL) was added potassium trimethylsilanol (445.17 mg, 3.47 mmol, 1 eq), and the mixture was stirred at 0°C for 1 hour. After the reaction, the reaction solution was spin-dried, slurried with methyl tert-butyl ether (10 mL), and filtered to give compound 260-5 (800 mg, 92.5%).

[0656] LCMS: (ESI, m / z): 212.0 [M-K+2H] + .

[0657] Step 5: Synthesis of compound 260-6

[0658] To a solution of potassium 5-(2-methyl-1,3-thiazol-4-yl)-1,3,4-oxadiazole-2-carboxylate (compound 260-5, 350 mg, 1.4 mmol, 1 eq) in dichloromethane (5 mL) was added dropwise DMF (10.23 mg, 0.14 mmol, 0.1 eq) at 0°C. After stirring for 1 minute, oxalyl chloride (0.24 mL, 2.8 mmol, 2 eq) was added dropwise. The mixture was stirred at 0°C for 30 minutes. After completion of the reaction, the reaction solution was spin-dried to give a crude compound 260-6 (320 mg, 99.25%), which was used directly in the next reaction.

[0659] LCMS: (ESI, m / z): 225.9 [M+H] + .

[0660] Step 5: Synthesis of Compound 260

[0661] To a solution of 5-(2-methyl-1,3-thiazol-4-yl)-1,3,4-oxadiazole-2-carbonyl chloride (compound 260-6, 320 mg, 1.39 mmol, 1 eq) in dichloromethane (3 mL) was added 6-(4,5,6,7-tetrahydro-1H-imidazo(4,5-c)pyridin-4-yl)-3-methylthieno[3,2-c]pyridine (compound 103-5, 225.47 mg, 0.83 mmol, 0.6 eq) at 0°C, and triethylamine (0.58 mL, 4.17 mmol, 3 eq) was added dropwise. The mixture was stirred at room temperature for 2 hours. After the reaction, the crude product was purified by silica gel column chromatography (dichloromethane / methanol = 10 / 1) to give a crude product, which was slurried with acetonitrile to give compound 260 (90 mg, 13.93%). SFC (column size: 250×25 mm 10 μm; mobile phase: methanol (+0.1% 7.0 mol / L amine methanol solution); wavelength: 214 nm; flow rate: 40 mL / min; temperature: RT; column pressure: 100 bar; injection volume: 8 mL; cycle time: 14.7 min; solvent: methanol: distillation grade, supercritical CO2: food grade) gave compounds 260-A (front peak, Rf=6.0-8.5 min, 33.14 mg, 5.13%) and 260-B (back peak, Rf=10.25-14.25 min, 36.61 mg, 5.67%).

[0662] Compound 260-A

[0663] LCMS: (ESI, m / z): 464.2 [M+H] + .

[0664] 1 H NMR (400MHz, DMSO-d6) δ12.27–11.97(m,1H),9.07–8.87(m,1H),8.55(d,J= 4.9Hz,1H),8.18–7.84(m,1H),7.74–7.55(m,1H),7.54–7.42(m,1H),6.95(d d,J=206.5,61.0Hz,1H),5.05–4.64(m,1H),4.07–3.73(m,0.6H),3.25–3.12 (m,0.4H),3.12–2.79(m,2H),2.78(d,J=4.5Hz,3H),2.45(d,J=15.6Hz,3H).

[0665] Compound 260-B

[0666] LCMS: (ESI, m / z): 464.2 [M+H] + .

[0667] 1 H NMR (400MHz, DMSO-d6) δ12.27–11.97(m,1H),8.97(d,J=32.3Hz,1H),8.55(d,J=4.9 Hz,1H),8.03(d,J=93.3Hz,1H),7.66(d,J=39.1Hz,1H),7.54–7.42(m,1H),6.91(d, J=200.8Hz,1H),4.82(ddd,J=17.6,13.3,5.1Hz,1H),3.97–3.86(m,0.6H),3.19–3. 09(m,0.4H),3.07–2.79(m,2H),2.78(d,J=4.4Hz,3H),2.45(dd,J=14.9,0.8Hz,3H).

[0668] Using conditions similar to those in the above examples, the compounds listed in Table 18 were prepared. The structural characterization data of these compounds are listed in Table 18.

[0669] Table 18 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0670] Example 28

[0671] Step 1: Synthesis of compound 258-1

[0672] To a solution of compound 5-(pyridin-3-yl)-1,3,4-oxadiazole-2-carboxylic acid potassium (compound 185-5, 260 mg, 1.13 mmol) in dichloromethane (5 mL) was added N,N-dimethylformamide (0.0087 mL, 0.11 mmol), and oxalyl chloride (0.19 mL, 2.26 mmol) was added at 0°C. The mixture was reacted at room temperature for 1 h. After the reaction was completed, the reaction solution was concentrated to give compound 258-1 (230 mg, 96.75%).

[0673] Step 2: Synthesis of Compound 258

[0674] To a solution of 5-(pyridin-3-yl)-1,3,4-oxadiazole-2-carbonyl chloride (compound 258-1, 230 mg, 1.10 mmol) in dichloromethane (5 mL) was added 3-methyl-6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[3,2-c]pyridine (compound 103-5, 178 mg, 0.66 mmol), and triethylamine (0.46 mL, 3.30 mmol) was added at 0°C. The mixture was reacted at 0°C for 1 h. After completion of the reaction, the reaction solution was concentrated to obtain a crude product, which was purified by normal phase purification (dichloromethane:methanol=8:1) to obtain compound 258 (100 mg, 20.55%). Compounds 258-A (front peak, Rf = 9.5 min, 7 mg, 7.00%) and 258-B (back peak, Rf = 15.5 min, 5 mg, 5.00%) were obtained by SFC (column size: 250×25 mm 10 μm; mobile phase: methanol (+0.1% 7.0 mol / L amine methanol solution); wavelength: 214 nm; flow rate: 40 mL / min; temperature: RT; injection volume: 2.0 mL; cycle time: 111 min; solvent: methanol: distillation grade, supercritical CO2: food grade).

[0675] Compound 258-A

[0676] LCMS: (ESI, m / z): 444.2 [M+H] + .

[0677] Compound 258-B

[0678] LCMS: (ESI, m / z): 444.2 [M+H] + .

[0679] Example 29

[0680] Step 1: Synthesis of compound 259-2

[0681] To a solution of potassium 5-(4-methylthiazol-2-yl)-1,3,4-oxadiazole-2-carboxylate (compound 259-1, 190 mg, 0.76 mmol) in dichloromethane (3 mL) was added N,N-dimethylformamide (0.0058 mL, 0.076 mmol), and oxalyl chloride (0.13 mL, 1.52 mmol) was added at 0°C. The mixture was reacted at room temperature for 1 h. After the reaction was completed, the reaction solution was concentrated to give compound 259-2 (160 mg, 91.42%).

[0682] Step 2: Synthesis of Compound 259

[0683] To a solution of 5-(4-methylthiazol-2-yl)-1,3,4-oxadiazole-2-carbonyl chloride (compound 259-2, 160 mg, 0.70 mmol) in dichloromethane (3 mL) was added 3-methyl-6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[3,2-c]pyridine (134 mg, 0.42 mmol), and triethylamine (0.29 mL, 2.10 mmol) was added at 0°C. The mixture was reacted at 0°C for 1 h. After the reaction was completed, the reaction solution was concentrated to obtain a crude product. The crude product was purified by normal phase chromatography (dichloromethane:methanol=8:1) to obtain the crude compound, which was then subjected to reverse phase preparative chromatography (conditions as follows: chromatographic column specifications: prep-HPLC (Waters 2767 / QDA), column: SunFire C18, 19×250 mm, 10 μm; mobile phase A: 0.1% formic acid / water, mobile phase B: acetonitrile; flow rate: 20 mL / min; elution gradient: 57% to 57%; retention time: 9.4-10.3 min) to give compound 259 (5 mg, 1.55%).

[0684] LCMS: (ESI, m / z): 464.1 [M+H] + .

[0685] Example 30

[0686] Step 1: Synthesis of compound 267-2

[0687] 2-Bromo-1-cyclopropyl-1-one (2.49 g, 15.25 mmol, 1 eq) was added to a solution of ethyl 2-amino-2-thioacetate (2 g, 15.25 mmol, 1 eq) in ethanol (20 mL), and the mixture was stirred at 70°C for 1 hour. After the reaction was completed, the reaction solution was spin-dried and the crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 0% to 20%) to give compound 267-2 (1.7 g, 50.86%).

[0688] LCMS: (ESI, m / z): 198.0 [M+H] + .

[0689] Based on the similar procedures to synthesize compound 183, compound 267-A (29.93 mg, 15.51%) was obtained.

[0690] 1 H NMR (400MHz, DMSO-d6) δ12.12(d,J=34.2Hz,1H),8.96(d,J=30.0Hz,1H),8.05(d,J=78 .1Hz,1H),7.80(s,1H),7.64(d,J=40.6Hz,1H),7.49(d,J=11.3Hz,1H),6.83(d,J=181 .3Hz,1H),4.97–4.69(m,1H),4.04–3.75(m,0.6H),3.19–3.13(m,0.4H),3.07–2.71(m ,2H),2.45(d,J=14.2Hz,3H),2.28–2.18(m,1H),1.04–0.98(m,2H),0.94–0.88(m,2H).

[0691] LCMS: (ESI, m / z): 490.1 [M+H] + .

[0692] Example 31

[0693] Step 1: Synthesis of compound 245-2

[0694] At room temperature, 3-bromo-2-fluoroaniline (compound 245-1, 10 g, 52.62 mmol, 1 eq) was added to water (200 mL), concentrated hydrochloric acid (400 mL) was added, the temperature was raised to 90 ° C to dissolve, the temperature was lowered to 0 ° C, and an aqueous solution (80 mL) of sodium nitrite (4.00 g, 57.88 mmol, 1.1 eq) was added dropwise. The mixture was stirred at 0 ° C for 1 hour, and ethoxy (potassium sulfonyl) methylthioketone (10.12 g, 63.1 4mmol, 1.2eq) in an aqueous solution (200 mL), stirred at room temperature for 2 hours. After the reaction was completed, the aqueous phase was extracted three times with dichloromethane (400 mL), the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated. The crude product was dissolved in ethanol (200 mL), potassium hydroxide (5.9 g, 105.24 mmol, 2eq) was added, stirred at room temperature for 1 hour, concentrated, water (400 mL) was added, and extracted once with dichloromethane (400 mL). The aqueous phase was adjusted to pH 3 with 2M HCl, and the aqueous phase was extracted twice with ethyl acetate (400 mL), dried over anhydrous sodium sulfate, and concentrated to give compound 245-2 (3.7 g, 34%), which was directly used for the next step reaction.

[0695] LCMS: (ESI, m / z): 204.7 [M+H] + .

[0696] Step 2: Synthesis of compound 245-3

[0697] At room temperature, 3-bromo-2-fluorobenzene-1-thiol (compound 245-2, 3.7 g, 11.59 mmol, 1 eq) was dissolved in N,N-dimethylformamide (37 mL), and 1-bromo-2,2-dimethoxypropane (2.01 g, 11.59 mmol, 1 eq), potassium carbonate (2.7 g, 23.18 mmol, 2 eq) and tetrabutylammonium iodide (0.44 g, 1.16 mmol, 0.1 eq) were added. The mixture was stirred at 150 ° C for 2 hours. After the reaction was completed, the reaction solution was quenched with 100 mL of water, extracted twice with ethyl acetate (50 mL), dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography (PE / EA: 0-10%) to obtain compound 245-3 (1.2 g, 21%).

[0698] Step 3: Synthesis of Compound 245-4

[0699] At room temperature, 1-bromo-3-[(2,2-dimethoxypropyl)thio]-2-fluorobenzene (compound 245-3, 1.2 g, 3.89 mmol, 1 eq) was dissolved in toluene (20 mL), and polyphosphoric acid (1.5 g) was added. The reaction temperature was raised to 120°C and stirred for 16 h. After the reaction was completed, the reaction solution was filtered and concentrated. The crude product was purified by silica gel column chromatography (100% PE) to give compound 245-4 (800 mg, 84%).

[0700] 1 H NMR (400MHz, CDCl3) δ7.68–7.47(m,1H),7.36–7.24(m,1H),7.27(s,1H),2.43–2.40(m,3H).

[0701] Step 4: Synthesis of compound 245-5

[0702] At room temperature, 2-methyl-6-bromo-7-fluoro-1-benzothiophene (compound 245-4, 800 mg, 3.28 mmol, 1 eq) was dissolved in ultra-dry tetrahydrofuran (10 mL), cooled to 0°C, and under nitrogen, isopropylmagnesium chloride lithium chloride complex (7.56 mL, 9.84 mmol, 3 eq) was added. The reaction solution was stirred at 0°C for 1 hour, ultra-dry N,N-dimethylformamide (718 mg, 9.84 mmol, 3 eq) was added, and the mixture was stirred at room temperature for 1 hour. After the reaction was completed, saturated aqueous ammonium chloride solution (50 mL) was added dropwise to the reaction solution to quench the reaction, and the mixture was extracted twice with ethyl acetate (50 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (100% PE) to give compound 245-5 (300 mg, 47%).

[0703] 1 H NMR (400MHz, CDCl3) δ10.49 (s, 1H), 7.86 (dd, J = 8.1, 6.4Hz, 1H), 7.57 (d, J = 8.3Hz, 1H), 7.38 (s, 1H), 2.47 (s, 3H).

[0704] Based on the similar procedures to synthesize compound 193, compound 245 (80 mg, 20%) was obtained.

[0705] LCMS: (ESI, m / z): 461.1 [M+H] + .

[0706] Using conditions similar to those in the above examples, the compounds listed in Table 19 were prepared. The structural characterization data of these compounds are listed in Table 19.

[0707] Table 19

[0708] Example 32

[0709] Step 1: Synthesis of compound 302-2

[0710] At room temperature, 1H-pyrazole-3-carboxylic acid ethyl ester (compound 302-1, 2.8 g, 20.13 mmol), bromocyclopentane (3.0 g, 20.13 mmol), K2CO 3( 5.5 g, 40.26 mmol) was dissolved in DMF (50 mL) and reacted at 80 ° C overnight. After the reaction, the reaction mixture was added to water (100 mL), extracted with ethyl acetate (100 mL × 3), washed with saturated brine (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 0-50%) to give compound 302-2 (2.6 g, 62.0%).

[0711] LCMS: (ESI, m / z): 209.0 [M+H] + .

[0712] Based on the similar procedures to synthesize compound 183, compound 302-A (62.9 mg, 38.1%) was obtained.

[0713] LCMS: (ESI, m / z): 501.1 [M+H] + .

[0714] 1 H NMR(400MHz, DMSO-d6)δ12.28–11.91(m,1H),8.97(d,J=32.0Hz,1H),8.15(s,0.6H),8.10–8.05(m, 1H),7.92(s,0.4H),7.64(d,J=43.4Hz,1H),7.49(d,J=11.9Hz,1H),7.17(s,0.4H),6.96(d,J=2.4H z,1H),6.64(s,0.6H),4.91–4.73(m,2H),3.97–3.85(m,0.5H),3.18–3.09(m,0.5H),3.05–2.70(m, 2H),2.49–2.42(m,3H),2.21–2.08(m,2H),2.04–1.92(m,2H),1.87–1.77(m,2H),1.73–1.60(m,2H).

[0715] Using conditions similar to those in the above examples, the compounds listed in Table 20 were prepared. The structural characterization data of these compounds are listed in Table 20.

[0716] Table 20 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0717] Example 33

[0718] Step 1: Synthesis of compound 323-2

[0719] At 0°C, methyl 2-methyl-1,3-oxazole-4-carboxylate (5 g, 35.4 mmol, 1 eq) was dissolved in ethanol (60 mL), and hydrazine hydrate (7.09 g, 141.7 mmol, 4 eq) was slowly added. After the addition was completed, the mixture was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was directly dried to obtain compound 323-2 (5 g, crude product), which was directly used in the next reaction.

[0720] LCMS: (ESI, m / z): 142.2 [M+H] + .

[0721] Step 2: Synthesis of compound 323-3

[0722] 2-Methyl-1,3-oxazole-4-carboxylic acid hydrazide (compound 323-2, 5 g, 35.43 mmol, 1 eq) was dissolved in dichloromethane (60 mL). Triethylamine (7.2 g, 70.9 mmol, 2 eq) was slowly added at 0°C, followed by ethyl oxalyl chloride (6.8 g, 49.6 mmol, 1.4 eq). After the addition was complete, the mixture was stirred at room temperature for 2 hours. After the reaction was complete, the reaction solution was directly used in the next reaction.

[0723] LCMS: (ESI, m / z): 242.1 [M+H] + .

[0724] Step 3: Synthesis of compound 323-4

[0725] At 0°C, dichloromethane (20 mL) was added to the reaction solution of the previous step, followed by triethylamine (10.1 g, 99.5 mmol, 3 eq) and then p-toluenesulfonyl chloride (19.0 g, 99.5 mmol, 3 eq). After the addition was complete, the mixture was stirred at room temperature for 16 hours. After the reaction was complete, the reaction solution was concentrated under vacuum, the residue was dissolved in 50 ml of ethyl acetate, and the insoluble matter was removed by filtration. The filtrate was diluted with 100 ml of water and extracted three times with 150 ml of ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 5:1) to give compound 323-4 (7 g, 94.5%).

[0726] LCMS: (ESI, m / z): 224.1 [M+H] + .

[0727] Step 4: Synthesis of compound 323-5

[0728] To a solution of ethyl 5-(2-methyl-1,3-oxazol-4-yl)-1,3,4-oxadiazole-2-carboxylate (compound 323-4, 2 g, 8.9 mmol, 1 eq) in tetrahydrofuran (20 mL) and water (5 mL) was added lithium hydroxide monohydrate (4.3.6 mg, 9.7 mmol, 1.1 eq) at room temperature, and the reaction was stirred at room temperature for 2 h. After the reaction was completed, the reaction solution was spin-dried, and the resulting residue was dissolved in a small amount of water and purified by a C18 reverse phase column (pure water / acetonitrile = 0-0%) to give compound 323-5 (1 g, 55.5%).

[0729] LCMS: (ESI, m / z): 196.1 [M-Li+2H] + .

[0730] Step 5: Synthesis of compound 323

[0731] Compound INT 1 (0.07 g, 0.26 mmol, 1 eq) and lithium 5-(2-methyloxazol-4-yl)-1,3,4-oxadiazole-2-carboxylate (compound 323-5, 0.052 g, 0.26 mmol, 1 eq) were dissolved in pyridine (4 mL), and phosphorus oxychloride (0.12 g, 0.78 mmol, 3 eq) was added. The mixture was reacted at room temperature for 2 hours. After the reaction, the reaction solution was dried and the crude product was purified by silica gel column chromatography (dichloromethane:methanol=0-20%) and then prepared by high performance liquid chromatography. Preparation conditions (Waters 2767 / QDA, column: Sunfire C18 19*250mm, 10μm; mobile phase A: 0.1% formic acid / water, B: acetonitrile; flow rate: 20 ml / min; gradient: 21-25%; retention time: 8.8-10.8 minutes (16 minutes)), to give compound 323 (21 mg, 18%).

[0732] LCMS: (ESI, m / z): 448.1 [M+H] + .

[0733] 1H NMR(400MHz,MeOD-d4)δ8.92(d,J=19.8Hz,1H),8.67(d,J=5.4Hz,1H),7.97(d ,J=112.1Hz,1H),7.74(d,J=21.5Hz,1H),7.38(d,J=22.2Hz,1H),7.09(d,J=1 80.4Hz,1H),5.08-4.96(m,1H),3.94–3.89(m,0.6H),3.26–3.14(m,1H),3.05 –2.93(m,0.4H),2.90-2.80(m,1H),2.57(d,J=5.2Hz,3H),2.52–2.48(m,3H).

[0734] Example 34 Note: "*" indicates that the carbon atom at the position is chiral carbon and the configuration is one of R or S, for example express One of them.

[0735] Step 1: Synthesis of compound 313-2

[0736] To a solution of 4,6-dichloronicotinic acid (10 g, 52.08 mmol) in dichloromethane (100 mL) was added N,O-dimethylhydroxylamine hydrochloride (6.6 g, 67.7 mmol), and N,N-carbonyldiimidazole (16.89 g, 104.16 mmol) and triethylamine (14.44 mL, 104.16 mmol) were added at 0°C. The mixture was reacted at room temperature for 4 h. After the reaction, saturated aqueous ammonium chloride solution (1000 mL) was added to the reaction solution, and dichloromethane (100 mL×3) was added for extraction. The organic phase was washed with saturated brine (100 mL×3), dried over anhydrous sodium sulfate, filtered and concentrated to obtain a crude product, which was then purified by normal phase purification (petroleum ether:ethyl acetate = 5:1) to obtain compound 313-2 (11.9 g, 97.20%).

[0737] LCMS: (ESI, m / z): 234.9 [M+H] + .

[0738] Step 2: Synthesis of compound 313-3

[0739] To a solution of 4,6-dichloro-N-methoxy-N-methylnicotinamide (compound 313-2, 11.7 g, 49.77 mmol) in tetrahydrofuran (100 mL) was added deuterated methylmagnesium iodide (150 mL, 124.43 mmol) at 0°C, and the reaction was carried out at 0°C for 4 h. After the reaction, saturated aqueous ammonium chloride solution (1000 mL) was added to the reaction solution, and the mixture was extracted with dichloromethane (100 mL×3). The organic phase was washed with saturated brine (100 mL×3), dried over anhydrous sodium sulfate, filtered and concentrated to obtain a crude product, which was then purified by normal phase purification (petroleum ether: ethyl acetate = 3:1) to obtain compound 313-3 (6.5 g, 67.65%).

[0740] LCMS: (ESI, m / z): 192.9 [M+H] + .

[0741] Step 3: Synthesis of compound 313-4

[0742] To a solution of 1-(4,6-dichloropyridin-3-yl)ethan-1-one-2,2,2-d3 (compound 313-3, 6.5 g, 33.85 mmol) in 1,2-dichloroethane (70 mL) were added methyl thioglycolate (3.77 g, 35.55 mmol) and 1,8-diazacyclo[5,4,0]undecene-7 (12.6 mL, 84.63 mmol) at 0°C, and the mixture was reacted at room temperature for 3 h. After the reaction was completed, the mixture was concentrated, water (100 mL) was added to the system, and dichloromethane (50 mL×3) was added for extraction. The organic phase was washed with saturated brine (50 mL×3), dried over anhydrous sodium sulfate, and concentrated to obtain a crude product. Acetonitrile (50 mL) was added to the crude product, and the mixture was filtered to obtain compound 313-4 (6.5 g, 78.69%).

[0743] LCMS: (ESI, m / z): 244.9 [M+H] + .

[0744] Step 4: Synthesis of compound 313-5

[0745] To a solution of methyl 6-chloro-3-(methyl-d3)thieno[3,2-c]pyridine-2-carboxylate (compound 313-4, 6.5 g, 26.64 mmol) in methanol (50 mL) and water (10 mL) was added lithium hydroxide (1.28 g, 53.28 mmol), and the reaction was carried out at room temperature for 5 h. After the reaction was completed, the mixture was concentrated, and water (100 mL) was added to the system. The pH was adjusted to 3 with 6N hydrochloric acid, and the mixture was filtered. The filter cake was concentrated in vacuo to obtain compound 313-5 (6.3 g).

[0746] LCMS: (ESI, m / z): 230.9 [M+H]+ .

[0747] Step 5: Synthesis of compound 313-6

[0748] 6-Chloro-3-(methyl-d3)thieno[3,2-c]pyridine-2-carboxylic acid (compound 313-5, 6.3 g, 27.31 mmol, 1 eq) was dissolved in DMSO (60 mL), and silver carbonate (1.51 g, 5.46 mmol, 0.2 eq) and acetic acid (0.082 g, 1.37 mmol, 0.05 eq) were added. The mixture was heated to 120°C and reacted for 24 hours. After the reaction, the reaction solution was diluted with water (100 mL) and extracted with ethyl acetate (100 mL × 3). The organic phases were combined, washed with saturated brine (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 1 / 1) to obtain compound 313-6 (4.6 g, 90.23%).

[0749] LCMS: (ESI, m / z): 186.9 [M+H] + .

[0750] Step 6: Synthesis of compound 313-7

[0751] To a solution of 6-chloro-3-(methyl-d3)thieno[3,2-c]pyridine (compound 313-6, 4.6 g, 24.94 mmol, 1 eq) in methanol (50 mL) were added 1,1'-bis(diphenylphosphino)ferrocenepalladium(II) dichloride (1.8 g, 2.46 mmol, 2 eq) and triethylamine (7.48 g, 73.92 mmol, 3 eq). The mixture was stirred at 140°C in a carbon monoxide atmosphere for 16 hours. After the reaction was completed, the reaction solution was spin-dried to give a crude product, which was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 4:1) to give compound 313-7 (4.56 g, 88.01%).

[0752] LCMS: (ESI, m / z): 211.1 [M+H] + .

[0753] Step 7: Synthesis of compound 313-8

[0754] To a solution of methyl 3-(methyl-d3)thieno[3,2-c]pyridine-6-carboxylate (compound 313-7, 4.56 g, 21.69 mmol) in tetrahydrofuran (50 mL) was added diisobutylaluminum hydride (18.8 mL, 28.20 mmol) at -78°C and the mixture was reacted at -78°C for 2 h. After the reaction, saturated aqueous ammonium chloride solution (1000 mL) was added to the reaction solution, and dichloromethane (100 mL×3) was added for extraction. The organic phase was washed with saturated brine (100 mL×3), dried over anhydrous sodium sulfate, filtered and concentrated to obtain a crude product. The crude product was purified by normal phase purification (petroleum ether: ethyl acetate = 3:1) to obtain compound 313-8 (3.5 g, 89.54%).

[0755] LCMS: (ESI, m / z): 181.2 [M+H] + .

[0756] Step 8: Synthesis of compound 313-9

[0757] To a solution of 3-(methyl-d3)thieno[3,2-c]pyridine-6-carbaldehyde (compound 313-8, 3.5 g, 19.42 mmol) in ethanol (15 mL) and water (15 mL) were added histamine dihydrochloride (4.29 g, 23.30 mmol) and sodium hydroxide (2.33 g, 58.26 mmol), and the mixture was reacted at 80°C for 3 h. After the reaction, the reaction solution was concentrated, water (30 mL) was added, and the mixture was stirred for 30 min and filtered. The filter cake was dried in vacuo to obtain compound 313-9 (4.4 g, 82.89%).

[0758] LCMS: (ESI, m / z): 274.1 [M+H] + .

[0759] Step 9: Synthesis of compound 313-10

[0760] To a dichloromethane solution (40 mL) of 3-(methyl-d3)-6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[3,2-c]pyridine (compound 313-9, 4.2 g, 15.36 mmol, 1 eq) were added triethylamine (4.26 mL, 30.72 mmol, 2 eq) and di-tert-butyl dicarbonate (10.06 g, 46.08 mmol, 3 eq). The mixture was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was dried and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to obtain compound 313-10 (6.2 g, 85.21%).

[0761] LCMS: (ESI, m / z): 474.3 [M+H] + .

[0762] Purified by SFC separation (system: Waters SFC 150; column name: Column size: 250*25mm; mobile phase A: supercritical CO2, mobile phase B: ethanol (+0.1% 7.0mol / L ammonia in methanol); A:B=50:50; wavelength: 214nm; flow rate: 140ml / min; column temperature: room temperature; back pressure: 100bar; injection: 1.0mL; circulation time: 15.5min; solvent: ethanol: redistilled grade, supercritical CO2: food grade) to obtain 313-10-A (front peak, 2.7g, SFC, Rf=1.430min, 43.55%) and 313-10-B (back peak, 2.9g, SFC, Rf=2.762min, 46.77%).

[0763] Step 10: Synthesis of compound 313-11-A

[0764] At room temperature, compound 313-10-A (2.7 g, 5.7 mmol, 1 eq) was dissolved in a solution of hydrochloric acid in 1,4-dioxane (30 mL), and the mixture was stirred at room temperature for 2 hours. After the reaction, the reaction solution was dried to obtain compound 313-11-A (2.5 g, crude product).

[0765] LCMS: (ESI, m / z): 273.6 [M+H] + .

[0766] Step 11: Synthesis of Compound 313-A

[0767] Compound 313-11-A (250 mg, 0.91 mmol, 1 eq) and potassium 5-(pyridin-2-yl)-1,3,4-oxadiazole-2-carboxylate (312.90 mg, 1.36 mmol, 1.5 eq) were dissolved in a mixed solution of acetonitrile (5 mL) and pyridine (0.5 mL). Phosphorus oxychloride (0.25 mL, 2.73 mmol, 3 eq) was added dropwise at -20°C, and the mixture was stirred at -20°C for 1 hour. After the reaction was completed, the reaction solution was spin-dried and purified by reverse column chromatography (ammonia:methanol = 50%) to give compound 313-A (125.3 mg, 30.69%).

[0768] LCMS: (ESI, m / z): 447.2 [M+H] + .

[0769] 1H NMR (400MHz, DMSO-d6) δ12.13(s,1H),8.97(d,J=35.7Hz,1H),8.82(t,J=5.5Hz ,1H),8.25(dd,J=7.9,3.8Hz,1H),8.16(s,0.6H),8.09(td,J=7.8,1.4Hz,1H), 7.95(s,0.4H),7.73–7.56(m,2H),7.48(d,J=13.0Hz,1H),6.90(d,J=187.4Hz, 1H),4.89(m,1H),4.02–3.87(m,0.5H),3.26–3.13(m,0.5H),3.08–2.70(m,2H).

[0770] Using conditions similar to those in the above examples, the compounds listed in Table 21 were prepared. The structural characterization data of these compounds are listed in Table 21.

[0771] Table 21

[0772] Example 35

[0773] Step 1: Synthesis of compound 310-1

[0774] 5-Bromo-1,3,4-oxadiazole-2-carboxylic acid ethyl ester (compound 243-1, 6 g, 27.15 mmol, 1 eq) and (tert-butoxy)carbohydrazide (3.59 g, 27.15 mmol, 0.1 eq) were dissolved in tetrahydrofuran (60 mL) and reacted at room temperature for 2 hours. After the reaction, the reaction solution was dried to obtain a crude product, which was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 5 / 1) to obtain compound 310-1 (4 g, 54.12%).

[0775] LCMS: (ESI, m / z): 216.9 [M+H] + .

[0776] Step 2: Synthesis of compound 310-2

[0777] To a solution of ethyl 5-(2-(tert-butoxycarbonyl)hydrazine)-1,3,4-oxadiazole-2-carboxylate (compound 310-1, 4 g, 14.69 mmol, 1 eq) in dichloromethane (40 mL) were added di-tert-butyl dicarbonate (7.69 g, 35.26 mmol, 2.4 eq), triethylamine (12.22 mL, 88.14 mmol, 6 eq), and 4-dimethylaminopyridine (0.18 g, 1.47 mmol, 0.1 eq) at room temperature. The mixture was reacted at room temperature for 4 hours. After the reaction was completed, the reaction solution was dried and the crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 5 / 1) to give compound 310-2 (2.5 g, 36.01%).

[0778] LCMS: (ESI, m / z): 473.2 [M+H] + .

[0779] Step 3: Synthesis of compound 310-3

[0780] At 0°C, tri-tert-butyl 2-(5-(ethoxycarbonyl)-1,3,4-oxadiazol-2-yl)hydrazine-1,1,2-tricarboxylate (compound 310-2, 2.5 g, 5.29 mmol, 1 eq) and potassium trimethylsilanol (0.68 g, 5.29 mmol, 1 eq) were dissolved in a mixed solvent of dichloromethane (15 mL) and methyl tert-butyl ether (15 mL). The mixture was stirred at 0°C for 1 hour. After the reaction was completed, the reaction solution was dried to give compound 310-3 (2.47 g, 96.74%).

[0781] LCMS: (ESI, m / z): 445.3 [M-K+2H] + .

[0782] Step 4: Synthesis of compound 310-4

[0783] Potassium 5-(1,2,2-tri(tert-butoxycarbonyl)hydrazine)-1,3,4-oxadiazole-2-carboxylate (compound 310-3, 2.47 g, 5.55 mmol, 1.5 eq) and INT 1 (1 g, 3.70 mmol, 1 eq) were dissolved in a mixed solvent of acetonitrile (20 mL) and pyridine (2 mL). Phosphorus oxychloride (1.01 mL, 11.10 mmol, 3 eq) was added at -20°C and stirred at -20°C for 1 hour. After the reaction was completed, the reaction solution was quenched with saturated aqueous sodium bicarbonate solution (10 mL), the reaction solution was spin-dried, and purified by C18 (ammonia-acetonitrile system 0-30%) to give compound 310-4 (1.8 g, 69.84%).

[0784] LCMS: (ESI, m / z): 697.3 [M+H] + .

[0785] Step 5: Synthesis of compound 310-5

[0786] Tri-tert-butyl 2-(5-(4-(3-methylthieno[3,2-c]pyridin-6-yl)-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine-5-carbonyl)-1,3,4-oxadiazol-2-yl)hydrazine-1,1,2-tricarboxylate (compound 310-4, 1.8 g, 2.58 mmol, 1 eq) was dissolved in a 1,4-dioxane solution of hydrochloric acid (20 mL), stirred at room temperature for 1 hour, and after completion of the reaction, the reaction solution was spin-dried to give compound 310-5 (0.9 g, 87.88%).

[0787] LCMS: (ESI, m / z): 397.1 [M+H] + .

[0788] Step 6: Synthesis of compound 310

[0789] (5-Hydrazino-1,3,4-oxadiazol-2-yl)(4-(3-methylthieno[3,2-c]pyridin-6-yl)-1,4,6,7-tetrahydro-5H-imidazo[4,5-c]pyridin-5-yl)methanone (compound 310-5, 0.8 g, 2.02 mmol, 1 eq) was dissolved in ethanol (10 mL), and 4,4-dimethoxy-2-butanone (0.32 g, 2.42 mmol, 1.2 eq) was added. The mixture was reacted at 80°C for 2 h. After the reaction was completed, the reaction solution was dried and purified by HPLC (Waters 2767 / QDA) column: Sunfire C18 19*250mm*10μm; flow rate: 20 ml / min; mobile phase A: 0.1% formic acid aqueous solution, B: ACN; gradient: 20-20%; retention time: 7.6-10.0 minutes, a total of 16 minutes), to obtain compounds 310-A ​​(front peak, 65.79 mg, 7.3%) and 310-B (back peak, 12.13 mg, 1.34%).

[0790] 310-A

[0791] LCMS: (ESI, m / z): 447.1 [M+H] + .

[0792] 1H NMR (400MHz, DMSO-d6) δ12.37(s,1H),8.98(d,J=20.8Hz,1H),8.46(d,J=2.4Hz,1H),8 .06(d,J=64.4Hz,1H),7.68(d,J=29.4Hz,1H),7.49(d,J=9.4Hz,1H),6.91(d,J=207.1 Hz,1H),6.61–6.56(m,1H),4.85(ddd,J=69.8,13.4,5.2Hz,1H),3.96–3.84(m,0.6H), 3.21–3.18(m,0.4H),3.03–2.73(m,2H),2.46(d,J=11.1Hz,3H),2.33(d,J=5.6Hz,3H).

[0793] 310-B

[0794] LCMS: (ESI, m / z): 447.1 [M+H] + .

[0795] Example 36

[0796] Step 1: Synthesis of compound 315-2

[0797] To a solution of 5-chlorothiazol-2-amine (compound 315-1) hydrochloride (2 g, 11.69 mmol, 1 eq.) in acetonitrile (30 mL) was added copper bromide (2.8 g, 12.86 mmol, 1.1 eq.) at room temperature. The temperature was lowered to 0°C, and tert-butyl nitrite (1.4 g, 14.03 mmol, 1.2 eq.) was added. The mixture was warmed to room temperature and stirred for 2 hours. After the reaction was completed, water (100 mL) was added to the reaction solution, and ethyl acetate (100 mL × 3) was added for extraction. The organic phases were combined, washed with saturated brine (50 mL × 3), dried over anhydrous sodium sulfate, filtered and concentrated to obtain a crude product, which was then purified by normal phase purification (petroleum ether: ethyl acetate = 100:1) to obtain compound 315-2 (1.5 g, 64.63%).

[0798] LCMS: (ESI, m / z): 199.8 [M+2+H] + .

[0799] Step 2: Synthesis of compound 315-3

[0800] To a solution of 2-bromo-5-chlorothiazole (compound 315-2, 1.5 g, 7.56 mmol, 1 eq.) in methanol (20 mL) were added 1,1'-bis(diphenylphosphino)ferrocenepalladium(II) dichloride (0.55 g, 0.76 mmol, 0.1 eq.) and triethylamine (2.29 g, 22.68 mmol, 3 eq.), and the mixture was reacted at 80°C under carbon monoxide protection for 5 h. After the reaction, water (100 mL) was added to the reaction solution, and ethyl acetate (80 mL×3) was added for extraction. The organic phases were combined, washed with saturated brine (80 mL×2), dried over anhydrous sodium sulfate, filtered and concentrated to obtain a crude product, which was then purified by normal phase purification (petroleum ether:ethyl acetate = 50:1) to obtain compound 315-3 (400 mg, 29.8%).

[0801] LCMS: (ESI, m / z): 177.9 [M+H] + .

[0802] Based on the similar procedure to synthesize compound 162, compound 315 (8 mg) was obtained.

[0803] LCMS: (ESI, m / z): 484.0 [M+H] + .

[0804] 1 H NMR (400MHz, DMSO-d6) δ12.16(s,1H),8.97(d,J=29.3Hz,1H),8.32(s,1H),8.06(d,J=74.9Hz,1H),7.65(d,J=33.5Hz,1H),7.49(d,J=9.6 Hz,1H),6.85(d,J=161.1Hz,1H),4.90–4.72(m,1H),3.98–3.87(m,0.6H),3.26–3.19(m,0.4H),3.04–2.73(m,2H),2.46(d,J=12.2Hz,3H).

[0805] Using conditions similar to those in the above examples, the compounds listed in Table 22 were prepared. The structural characterization data of these compounds are listed in Table 22.

[0806] Table 22 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0807] Example 37

[0808] Step 1: Synthesis of compound 31-2

[0809] To a solution of 2-bromothiazole-5-carboxaldehyde (compound 316-1, 5 g, 26.04 mmol) in dichloromethane (50 mL) was added diethylaminosulfur trifluoride (6.9 mL, 52.08 mmol) and the reaction was carried out at room temperature for 3 h. After the reaction was completed, saturated aqueous ammonium chloride solution (1000 mL) was added to the reaction solution, and dichloromethane (50 mL×3) was added for extraction. The organic phases were combined, washed with saturated brine (50 mL×3), dried over anhydrous sodium sulfate, filtered and concentrated to obtain a crude product, which was then purified by normal phase purification (petroleum ether: ethyl acetate = 80:1) to obtain compound 315-2 (3.54 g, 63.52%).

[0810] Based on the similar procedure to synthesize compound 315, compound 316 (3.8 mg) was obtained.

[0811] LCMS: (ESI, m / z): 500.1 [M+H] + .

[0812] 1 H NMR (400MHz, DMSO-d6) δ12.12(d,J=33.7Hz,1H),8.97(d,J=28.8Hz,1H),8.56(s,1H),8.07(d,J=71.9Hz,1H),7.73–7 .43(m,3H),6.81(d,J=158.9Hz,1H),4.89–4.71(m,1H),4.02–3.91(m,0.6H),3.03–2.70(m,2.4H),2.48–2.42(m,3H).

[0813] Example 38

[0814] Step 1: Synthesis of compound 322-2

[0815] To a solution of ethyl 5-methyl-1,3-oxazole-2-carboxylate (compound 322-1, 1 g, 6.45 mmol, 1 eq) in ethanol (10 mL) was added hydrazine hydrate (1.29 g, 25.8 mmol, 4 eq) and the mixture was reacted at 25°C for 12 hours. After the reaction, the reaction solution was spin-dried and mixed with the sample and purified by a forward column using a dichloromethane-methanol system as the mobile phase (dichloromethane: methanol = 9:1) to obtain compound 322-2 (800 mg, 87.95%).

[0816] LCMS: (ESI, m / z): 142.4 [M+H] + .

[0817] Step 2: Synthesis of compound 322-3

[0818] To a solution of 5-methyl-1,3-oxazole-2-carbohydrazide (compound 322-2, 0.8 g, 5.67 mmol, 1 eq) and triethylamine (1.15 g, 11.34 mmol, 2 eq) in dichloromethane (12 mL) was added ethyl oxalyl chloride (0.93 g, 6.8 mmol, 1.2 eq) at 0°C and the mixture was reacted at 25°C for 2 hours. After the reaction was complete, the reaction solution was directly used in the next reaction.

[0819] LCMS: (ESI, m / z): 242.1 [M+H] + .

[0820] Step 3: Synthesis of compound 322-4

[0821] The amount of raw materials was calculated according to the theoretical yield of the previous step, i.e., compound 322-3 (1.3 g, 5.39 mmol, 1 eq). Triethylamine (1.64 g, 1617 mmol, 3 eq) and p-toluenesulfonyl chloride (3.08 g, 16.17 mmol, 3 eq) were added to the reaction solution at 0°C and reacted at 25°C for 3 hours. After the reaction, the reaction solution was diluted with 80 ml of dichloromethane and washed twice with 50 ml of saturated brine. The organic phase was dried over anhydrous sodium sulfate, spin-dried, and purified by a forward column (petroleum ether: ethyl acetate = 2:3) to obtain 322-4 (0.8 g, 66.5%).

[0822] LCMS: (ESI, m / z): 224.1 [M+H] + .

[0823] Step 4: Synthesis of compound 322-5

[0824] Ethyl 5-(5-methyl-1,3-oxazol-2-yl)-1,3,4-oxadiazole-2-carboxylate (compound 322-4, 300 mg, 1.34 mmol, 1 eq) was dissolved in tetrahydrofuran (10 mL), and a solution of lithium hydroxide monohydrate (84.34 mg, 2.01 mmol, 1.5 eq) in water (2 mL) was added. The mixture was reacted at 25°C for 2 hours. After the reaction, the reaction solution was dried and passed through a reverse column (acetonitrile and pure water system as the mobile phase, pure water produced the product peak), and then lyophilized to obtain compound 322-5 (220 mg, 81.4%).

[0825] LCMS: (ESI, m / z): 196.1 [M-Li+2H] + .

[0826] Step 5: Synthesis of compound 322

[0827] INT 1 (70 mg, 0.26 mmol, 1 eq) and 5-(5-methyl-1,3-oxazol-2-yl)-1,3,4-oxadiazole-2-carboxylic acid lithium salt (57.51 mg, 0.29 mmol, 1.1 eq) were dissolved in pyridine (6 mL), and phosphorus oxychloride (119.60 mg, 0.78 mmol, 3 eq) was added at 0°C. The mixture was reacted at 25°C for 3 hours. After the reaction, the reaction solution was dried and passed through a reverse phase column (ammonia methanol as the mobile phase, 80% methanol as the product peak). After drying, the product was purified by high performance liquid chromatography (Waters 2767 / QDA) column: Sunfire C18 19*250mm*10μm; flow rate: 20 ml / min; mobile phase A: 0.1% formic acid aqueous solution, B: ACN; gradient: 20-25%; retention time: 8.3-9.3 minutes, a total of 16 minutes) to give compound 322 (20 mg, 17.3%).

[0828] LCMS: (ESI, m / z): 448.3 [M+H] + .

[0829] 1 H NMR (400MHz, MeOD-d4) δ8.92(d,J=20.3Hz,1H),7.96(d,J=116.5Hz,1H),7.96(d,J= 116.5Hz,1H),7.37(s,0.4H),7.36–7.30(m,1H),7.20(dd,J=6.8,1.1Hz,1H),6.86( s,0.6H),5.03–4.88(m,1H),4.00–3.87(m,0.6H),3.26–3.10(m,1H),3.05–2.94(m, 0.4H), 2.90–2.79(m,1H),2.51(dd,J=3.5,1.1Hz,3H),2.49(dd,J=4.8,1.0Hz,3H).

[0830] Using conditions similar to those in the above examples, the compounds listed in Table 23 were prepared. The structural characterization data of these compounds are listed in Table 23.

[0831] Table 23 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0832] Example 39

[0833] Step 1: Synthesis of compound 319-2

[0834] 3-Bromo-1H-pyrrole-2-carboxylic acid methyl ester (compound 319-1, 6 g, 29.41 mmol) was dissolved in tetrahydrofuran (40 ml), sodium hydride (1.41 g, 58.82 mmol) was added, and iodomethane (6.26 g, 44.12 mmol) was added at room temperature. The mixture was reacted at room temperature for 2 hours. After the reaction, the reaction solution was poured into water to quench the reaction, extracted with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (PE:EA = 30%) to obtain compound 319-2 (5 g, 77.97%).

[0835] LCMS: (ESI, m / z): 220.0 [M+2+H] + .

[0836] Step 2: Synthesis of compound 319-3

[0837] 3-Bromo-1-methyl-1H-pyrrole-2-carboxylic acid methyl ester (compound 319-2, 6 g, 27.52 mmol) and sodium hydroxide (1.10 g, 27.52 mmol) were dissolved in tetrahydrofuran (1 mL) and water (0.2 mL) and reacted at room temperature for 2 hours. After the reaction, the pH was adjusted to 5 and the mixture was filtered to obtain compound 319-3 (4 g, 71.25%).

[0838] Step 3: Synthesis of compound 319-4

[0839] 3-Bromo-1-methyl-1H-pyrrole-2-carboxylic acid (compound 319-3, 2 g, 9.80 mmol), ammonium chloride (0.79 g, 14.70 mmol), 2-(7-aza-1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (5.59 g, 14.70 mmol), and ethyldiisopropylamine (2.53 g, 19.60 mmol) were dissolved in dimethylformamide (20 mL) and reacted at room temperature for 2 hours. After the reaction, the reaction solution was poured into water, extracted with ethyl acetate, dried over anhydrous sodium sulfate, and spin-dried. The crude product was purified by silica gel column chromatography (PE:EA=50%) to obtain compound 319-4 (1.5 g, 75.36%).

[0840] LCMS: (ESI, m / z): 203.0 [M+2+H] + .

[0841] Step 4: Synthesis of compound 319-5

[0842] 3-Bromo-1-methyl-1H-pyrrole-2-carboxamide (compound 319-4, 2 g, 9.85 mmol) was dissolved in phosphorus oxychloride (10 ml) and reacted at 80°C for 2 hours. After the reaction, the system was spin-dried, the pH was adjusted to 9 with sodium bicarbonate, extracted with ethyl acetate, dried over anhydrous sodium sulfate, and spin-dried to obtain compound 319-5 (1.5 g, 82.30%).

[0843] Step 5: Synthesis of compound 319-6

[0844] 3-Bromo-1-methyl-1H-pyrrole-2-carbonitrile (compound 319-5, 1 g, 5.40 mmol), bis(pinacol)diboron (2.74 g, 10.8 mmol), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (0.40 g, 0.54 mmol), and potassium acetate (2.65 g, 27 mmol) were dissolved in 1,4-dioxane (20 mL) and reacted at 100°C for 12 hours. After the reaction, the compound was poured into water (10 mL) and extracted with ethyl acetate (20 mL×2). The organic phases were combined and dried over anhydrous sodium sulfate. The crude product was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 5%) to obtain compound 319-6 (0.8 g, 63.78%).

[0845] Step 6: Synthesis of compound 319-7

[0846] 1-Methyl-3-(tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrrole-2-carbonitrile (compound 319-6, 0.2 g, 0.86 mmol), ethyl 5-bromo-1,3,4-oxadiazole-2-carboxylate (compound 243-1, 0.19 g, 0.86 mmol), and [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (0.063 g, 0.086 mmol) were dissolved in 1,4-dioxane (1 mL) and water (0.2 mL), and the mixture was reacted at 100°C for 2 hours. After the reaction, the system was poured into water (10 mL), extracted with ethyl acetate (20 mL), dried over anhydrous sodium sulfate, and spin-dried. The crude product was purified by silica gel column chromatography (PE:EA = 30%) to obtain compound 319-7 (0.15 g, 70.70%).

[0847] Based on the similar procedure to synthesize compound 322, compound 319 (3 ​​mg) was obtained.

[0848] LCMS: (ESI, m / z): 471.2 [M+H] + .

[0849] Example 40

[0850] Step 1: Synthesis of compound 334-2

[0851] To a solution of ethyl 4-formyl-1H-pyrazole-3-carboxylate (compound 334-1, 5 g, 29.74 mmol, 1 eq) in tetrahydrofuran (50 mL) was added sodium hydride (1.43 g, 59.48 mmol, 2 eq) at 0°C. The mixture was reacted at 0°C for 20 minutes, followed by addition of iodomethane (6.33 g, 44.61 mmol, 1.5 eq). The mixture was reacted at 25°C for 2 hours. After completion of the reaction, the reaction solution was poured into water to quench the reaction, extracted with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (PE:EA=5:1) to give 334-2 (4.5 g, 83.07%).

[0852] LCMS: (ESI, m / z): 183.1 [M+H] + .

[0853] Step 2: Synthesis of compound 334-3

[0854] To a solution of ethyl 4-formyl-1-methyl-1H-pyrazole-3-carboxylate (compound 334-2, 4 g, 21.96 mmol, 1 eq) in dichloromethane (40 mL) was added diethylaminosulfur trifluoride (17.7 g, 109.8 mmol, 5 eq) at 0°C and the mixture was reacted at 25°C for 12 hours. After completion of the reaction, the reaction solution was slowly added to ice water to quench the reaction, extracted with dichloromethane, and the organic phases were combined, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography (PE:EA=5:1) to give compound 334-3 (2.1 g, 46.85%).

[0855] Based on the similar procedure to synthesize compound 322, compound 334 (3 mg) was obtained.

[0856] LCMS: (ESI, m / z): 497.2 [M+H] + .

[0857] Example 41

[0858] Step 1: Synthesis of compound 347-2

[0859] To a solution of tert-butyl (2-(methoxy(methyl)amino)-2-oxoethyl)carbamate (compound 347-1, 10 g, 45.82 mmol, 1 eq) in THF (100 mL) was added dropwise cyclopropylmagnesium bromide (114.55 ml, 114.55 mmol, 1 M, 2.5 eq) at -78°C under a nitrogen atmosphere. The reaction was stirred at room temperature for 4 hours. After the reaction, aqueous ammonium chloride solution (50 mL) was slowly poured into the reaction solution and diluted and stirred for 10 minutes. Ethyl acetate (100 mL × 2) was added for extraction. The combined organic phases were washed with saturated brine (50 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 4:1) to give compound 347-2 (3.5 g, 38.34%).

[0860] 1 H NMR (400MHz, CDCl3) δ4.20 (d, J = 4.6Hz, 2H), 1.96–1.83 (m, 1H), 1.45 (s, 9H), 1.13–1.08 (m, 2H), 0.99–0.94 (m, 2H).

[0861] Step 2: Synthesis of compound 347-3

[0862] At room temperature, a solution of hydrochloric acid in dioxane (30 mL) was added to tert-butyl (2-cyclopropyl-2-oxoethyl)carbamate (compound 347-2, 3.4 g, 17.06 mmol, 1 eq), and the reaction was stirred for 16 hours. After the reaction was completed, the mixture was concentrated to give compound 347-3 (2.3 g, 99.41%).

[0863] LCMS: (ESI, m / z): 100.3 [M+H] + .

[0864] Step 3: Synthesis of compound 347-4

[0865] To a solution of 2-amino-1-cyclopropylethane-1-one hydrochloride (compound 347-3, 1.65 g, 16.64 mmol, 1 eq) and triethylamine (5.05 g, 49.92 mmol, 3 eq) in toluene (20 mL) was added ethyl oxalyl chloride (2.73 g, 19.97 mmol, 1.2 eq) at 0°C, and the reaction was stirred at room temperature for 1 hour. After the reaction was completed, water (50 mL) was added to the reaction solution for dilution, and the mixture was extracted with ethyl acetate (50 mL × 2). The organic phases were combined, washed with saturated brine (30 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to give compound 347-4 (2.8 g, 84.45%).

[0866] LCMS: (ESI, m / z): 200.2 [M+H] + .

[0867] Step 4: Synthesis of compound 347-5

[0868] Phosphorus oxychloride (21.56 g, 140.6 mmol, 10 eq) was added to a solution of ethyl 2-((2-cyclopropyl-2-oxoethyl)amino)-2-oxoacetate (compound 347-4, 2.8 g, 14.06 mmol, 1 eq) at room temperature, and the reaction was stirred at 120°C for 16 hours. After completion of the reaction, the mixture was cooled to room temperature, and methanol (20 mL) was added to the reaction system under ice bath and stirred for 20 minutes. The mixture was concentrated, diluted with water (50 mL), and extracted with ethyl acetate (50 mL × 2). The organic phases were combined, washed with saturated brine (50 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to give the crude product, which was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 1:1) to give compound 347-5 (950 mg, 37.30%).

[0869] LCMS: (ESI, m / z): 182.1 [M+H] + .

[0870] Based on the similar procedures to synthesize compound 183, compound 347 (25 mg, 11.4%) was obtained.

[0871] LCMS: (ESI, m / z): 474.2 [M+H] + .

[0872] Using conditions similar to those in the above examples, the compounds listed in Table 24 were prepared. The structural characterization data of these compounds are listed in Table 24.

[0873] Table 24 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0874] Example 42

[0875] Step 1: Synthesis of compound 338-2

[0876] Ethyl 2-methyl-1,3-oxazole-4-carboxylate (compound 338-1, 10 g, 64.45 mmol) and N-bromosuccinimide (17.21 g, 96.68 mmol) were dissolved in acetonitrile (100 mL) and reacted at 80°C for 2 hours. After the reaction was completed, the reaction solution was poured into water (20 mL), extracted with ethyl acetate (30 mL × 2), dried over anhydrous sodium sulfate, and the solvent was dried by spin drying. The product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 0-12%) to obtain compound 338-2 (4 g, 26.52%).

[0877] LCMS: (ESI, m / z): 236.2 [M+2+H] + .

[0878] Step 2: Synthesis of compound 338-3

[0879] To a solution of ethyl 5-bromo-2-methyl-1,3-oxazole-4-carboxylate (compound 338-2, 2.4 g, 10.25 mmol, 1 eq) in tetrahydrofuran (24 mL) and water (6 mL) were added cyclopropylboronic acid (1.76 g, 20.5 mmol, 2 eq), tricyclohexylphosphine (0.57 g, 2.05 mmol, 0.1 eq), tris(dibenzylideneacetone) diphosphine (compound 338-2, 0.94 g, 1.03 mmol, 0.2 eq), and tripotassium phosphate (6.53 g, 30.75 mmol, 3 eq). The reaction system was reacted at 80°C for 16 hours. After the reaction, the reaction solution was extracted with ethyl acetate (30 mL × 2), dried over anhydrous sodium sulfate, and dried by spin drying. The product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 9:1) to obtain compound 338-3 (1.45 g, 72.43%).

[0880] Based on the similar procedures to synthesize compound 183, compound 338 (12 mg, 9.5%) was obtained.

[0881] LCMS: (ESI, m / z): 488.4 [M+H] + .

[0882] Using conditions similar to those in the above examples, the compounds listed in Table 25 were prepared. The structural characterization data of these compounds are listed in Table 25.

[0883] Table 25 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0884] Example 43

[0885] Step 1: Synthesis of compound 335-2

[0886] To a solution of ethyl 2-chlorooxazole-4-carboxylate (compound 335-1, 3.5 g, 19.9 mmol, 1 eq) in toluene (50 mL) were added cyclopropylboronic acid (5.3 g, 29.9 mmol, 1.5 eq), palladium acetate (420 mg, 2.0 mmol, 0.1 eq), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex (1.4 g, 4.0 mmol, 0.2 eq) and cesium carbonate (19.3 g, 59.8 mmol, 3 eq), and the mixture was reacted at 100°C for 16 hours. After the reaction, the reaction solution was dried and purified by silica gel column chromatography (petroleum ether:ethyl acetate = 9:1) to obtain compound 335-2 (3.3 g, 91.6%).

[0887] LCMS: (ESI, m / z): 182.1 [M+H] + .

[0888] Based on the similar procedures to synthesize compound 183, compound 335 (9.8 mg, 8.0%) was obtained.

[0889] LCMS: (ESI, m / z): 474.2 [M+H] + .

[0890] Example 44

[0891] Step 1: Synthesis of compound 355-1

[0892] To a solution of methyl 2-methyloxazole-4-carboxylate (compound 323-1, 10 g, 64.45 mmol, 1 eq) in DMF (120 mL) was added N-bromosuccinimide (17.2 g, 96.68 mmol, 1.5 eq) and stirred at 80°C for 12 hours. After the reaction was completed, the reaction was quenched with water (100 mL), extracted with ethyl acetate (100 mL × 3), washed with saturated brine (100 mL × 2), and the organic phases were combined and dried over anhydrous sodium sulfate. The residue was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 5:1) to give compound 355-1 (5.5 g, 36.4%).

[0893] LCMS: (ESI, m / z): 222.0 [M+H] + .

[0894] Step 1: Synthesis of compound 355-2

[0895] To a solution of methyl 5-bromo-2-methyloxazole-4-carboxylate (compound 355-1, 3.5 g, 15.91 mmol, 1 eq) in DMF (40 mL) were added cuprous iodide (6.06 g, 31.82 mmol, 2.0 eq) and methyl 2,2-difluoro-2-(fluorosulfonyl)acetate (6.11 g, 31.82 mmol, 2.0 eq). The reaction solution was stirred at 80°C for 12 hours. After completion of the reaction, the mixture was quenched with water (100 mL), extracted with ethyl acetate (100 mL × 3), washed with saturated brine (100 mL × 2), and the organic phases were combined and dried over anhydrous sodium sulfate. The residue was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 8:1) to give compound 355-2 (0.75 g, 22.5%).

[0896] LCMS: (ESI, m / z): 210.1 [M+H] + .

[0897] Based on the similar procedures to synthesize compound 323, compound 355 (32 mg, 14.3%) was obtained.

[0898] LCMS: (ESI, m / z): 516.5 [M+H] + .

[0899] 1 H NMR (400MHz, DMSO-d6) δ12.19(s,1H),8.96(d,J=41.6Hz,1H),8.08(d,J=84.8Hz,1H),7.66(d,J=39.0Hz,1H),7.49(dd,J=8.7,1.1Hz,1H),6.84(d,J=1 63.5Hz,1H),4.86–4.73(m,1H),3.94(t,J=11.3Hz,0.6H),3.22–3.12(m,0. 4H),3.06–2.73(m,2H),2.65(d,J=2.9Hz,3H),2.46(dd,J=12.7,1.0Hz,3H).

[0900] Using conditions similar to those in the above examples, the compounds listed in Table 26 were prepared. The structural characterization data of these compounds are listed in Table 26.

[0901] Table 26 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[0902] Example 45

[0903] Step 1: Synthesis of compound 248-2

[0904] To a solution of DMP (19.85 g, 46.8 mmol) and NaHCO3 (6.55 g, 78 mmol) in THF (100 mL) at 0°C was added 2-(thiophen-3-yl)ethane-1-ol (compound 248-1, 4 g, 31.20 mmol) dropwise. The mixture was stirred at room temperature for 2 h. After the reaction was completed, water (100 mL) was added to the reaction solution for dilution, and the mixture was extracted with ethyl acetate (30 mL × 3). The combined organic phases were washed with saturated brine (50 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 5:1) to give compound 248-2 (1.4 g, 35.56%).

[0905] Step 2: Synthesis of compound 248-3

[0906] At room temperature, 2-(thiophen-3-yl)acetaldehyde (compound 248-2, 1500 mg, 11.89 mmol), methyl 3-oxobutanoate (1380.67 mg, 11.89 mmol) and Sc(OTf)3 (585.18 mg, 1.19 mmol) were added to acetonitrile (20 mL) and reacted at 60°C for 4 hours. After the reaction, water (100 mL) was added to the reaction solution for dilution, and the mixture was extracted with ethyl acetate (30 mL×3). The combined organic phases were washed with saturated brine (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 2:1) to give compound 248-3 (400 mg, 16.31%).

[0907] Step 3: Synthesis of Compound 248-4

[0908] To a solution of methyl 7-methyl-1-benzothiophene-6-carboxylate (compound 248-3, 400 mg, 1.94 mmol) in THF (5 mL) was added DIBAL-H (1.55 mL, 2.33 mmol) at -78°C, and the reaction was stirred for 16 hours. After the reaction was completed, the reaction solution was slowly poured into a saturated aqueous solution of potassium sodium tartrate (30 mL) and diluted and stirred for 2 hours. Ethyl acetate (20 mL × 3) was added and extracted. The combined organic phases were washed with saturated brine (20 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to obtain compound 248-4 (250 mg, 73.15%).

[0909] Based on the similar procedures to synthesize compound 133, compound 248 (10.34 mg, 7.87%) was obtained.

[0910] 1H NMR (400MHz, DMSO-d6) δ12.13(s,1H),8.81(d,J=4.2Hz,1H),8.25(d,J=7.9Hz,1H),8.09(td,J=7.8,1.7Hz,1H),7.80(d,J=5.4Hz,1H),7.74–7. 59(m,3H),7.47(d,J=5.5Hz,1H),6.99(d,J=7.6Hz,2H),4.72(dd,J=14. 3,5.2Hz,1H),3.63–3.55(m,1H),3.07–2.96(m,1H),2.91–2.69(m,4H).

[0911] LCMS: (ESI, m / z): 443.1 [M+H] + .

[0912] Using conditions similar to those in the above examples, the compounds listed in Table 27 were prepared. The structural characterization data of these compounds are listed in Table 27.

[0913] Table 27

[0914] Example 46

[0915] Step 1: Synthesis of compound 231-1

[0916] To a solution of tert-butyl 5-aminopentanoate hydrochloride (209.71 mg, 1 mmol, 1 eq) in THF (2 mL) were added ethyl 5-bromo-1,3,4-oxadiazole-2-carboxylate (compound 243-1, 220 mg, 1.0 mmol, 1 eq) and DIEA (155.09 mg, 1.2 mmol, 1.2 eq). The reaction was stirred at room temperature for 2 hours. After the reaction was completed, the mixture was concentrated to obtain a crude product, which was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to obtain compound 231-1 (220 mg, 70.53%).

[0917] LCMS: (ESI, m / z): 258.0 [M+H] + .

[0918] Step 2: Synthesis of compound 231-2

[0919] To a solution of ethyl 5-[(5-(tert-butoxy)-5-oxopentyl)amino]-1,3,4-oxadiazole-2-carboxylate (compound 231-1, 200 mg, 0.64 mmol, 1 eq) in DCM (0.5 mL) and MTBE (0.5 mL) was added potassium trimethylsilanol (98.53 mg, 0.77 mmol, 1.2 eq) at 0°C. The mixture was stirred for 1 hour. After completion of the reaction, the mixture was concentrated to give compound 231-2 (206 mg, 99.80%).

[0920] LCMS: (ESI, m / z): 230.1[MK- t Bu+3H] + .

[0921] Step 3: Synthesis of compound 231-3

[0922] To a solution of potassium 5-((5-(tert-butoxy)-5-oxopentyl)amino)-1,3,4-oxadiazole-2-carboxylate (compound 231-2, 200 mg, 0.64 mmol) in acetonitrile (2 mL) was added 6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)thieno[3,2-c]pyridine (compound 094-3, 115 mg, 0.45 mmol), and phosphorus oxychloride (0.18 mL, 1.92 mmol) was added at -20°C. The mixture was reacted at -20°C for 0.5 hour. After the reaction was completed, the reaction solution was concentrated to obtain a crude product, which was purified by normal phase purification (DCM:MeOH=15:1) to give compound 231-3 (70 mg, 20.95%).

[0923] LCMS: (ESI, m / z): 524.2 [M+H] + .

[0924] Step 4: Synthesis of compound 231-4

[0925] To a solution of tert-butyl 5-((5-(4-(thieno[3,2-c]pyridin-6-yl)-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine-5-carbonyl)-1,3,4-oxadiazol-2-yl)amino)pentanoate (compound 231-3, 27 mg, 0.052 mmol) in dichloromethane (2 mL) was added trifluoroacetic acid (0.2 mL, 2.68 mmol) and the mixture was reacted at room temperature for 2 h. After the reaction was completed, the reaction solution was concentrated to give compound 231-4 (20 mg, 82.96%).

[0926] LCMS: (ESI, m / z): 468.2 [M+H] + .

[0927] Step 5: Synthesis of Compound 231

[0928] To a solution of 5-((5-(4-(thieno[3,2-c]pyridin-6-yl)-4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine-5-carbonyl)-1,3,4-oxadiazol-2-yl)amino)pentanoic acid (compound 231-4, 14 mg, 0.03 mmol) in dichloromethane (2 mL) were added 2-chloro-1-methylpyridine-1-iodide (9 mg, 0.036 mmol) and triethylamine (0.01 mL, 0.06 mmol) and reacted at room temperature for 2 h. After the reaction was complete, saturated aqueous sodium bicarbonate solution (1 mL) was added to the reaction solution to quench the reaction. After concentration, the mixture was subjected to reverse phase preparative chromatography (conditions as follows: chromatographic column specifications: prep-HPLC (Waters 2767 / QDA), Column: SunFire C18, 19*250mm, 10μm; mobile phase A: 0.1% formic acid / water, mobile phase B: acetonitrile; flow rate: 20mL / min; elution gradient: 52% to 52%; retention time: 8.4-9.5min) to obtain compound 231 (4mg, 29.72%).

[0929] LCMS: (ESI, m / z): 450.2 [M+H] + .

[0930] Example 47

[0931] Step 1: Synthesis of compound 358-1

[0932] At 25 ° C, methyl 2-methyl-1,3-oxazole-4-carboxylate (compound 323-1, 5 g, 35.43 mmol, 1 eq) was dissolved in tetrachloromethane (50 mL) solution, and 2,2'-azobis(2-methylpropionitrile) (1.16 g, 7.09 mmol, 0.2 eq) and N-bromosuccinimide (6.31 g, 35.43 mmol, 1 eq) were slowly added. After the addition was completed, the system was stirred at 80 ° C for 16 hours. After the reaction was completed, water (200 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with ethyl acetate (100 mL × 2), washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by normal phase purification (petroleum ether: ethyl acetate = 5:1) to obtain compound 358-1 (2.4 g, 30.79%).

[0933] LCMS: (ESI, m / z): 222.0 [M+2+H] + .

[0934] Step 2: Synthesis of compound 358-2

[0935] At 25 ° C, methyl 2-(bromomethyl)-1,3-oxazole-4-carboxylate (compound 358-1, 2.4 g, 10.91 mmol, 1 eq) was dissolved in N,N-dimethylformamide (30 mL) solution, and methyl 2,2-difluoro-2-(fluorosulfonyl)acetate (4191.84 mg, 21.82 mmol, 2 eq) and copper iodide (415.56 mg, 2.18 mmol, 0.2 eq) were slowly added. After the addition was completed, the system was stirred at 80 ° C for 16 hours. After the reaction was completed, the reaction solution was filtered, water (50 mL) was added, and the mixture was extracted with ethyl acetate (50 mL×2), washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by normal phase purification (petroleum ether: ethyl acetate = 5:1) to obtain compound 358-2 (600 mg, 26.30%).

[0936] LCMS: (ESI, m / z): 210.1 [M+H] + .

[0937] Based on the similar procedures to synthesize compound 323, compound 358 (32 mg, 14.3%) was obtained.

[0938] LCMS: (ESI, m / z): 516.1 [M+H] + .

[0939] Example 48

[0940] Step 1: Synthesis of compound 377-1

[0941] To a mixed solution of 3-bromo-6-((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 193-1, 2 g, 4.99 mmol, 1 eq) in 1,4-dioxane (30 mL) and water (1.5 mL) were added trimethylcyclotriboroxane (3.16 g, 25.06 mmol, 5 eq), dichloro[1,1-bis(di-tert-butylphosphino)ferrocene]palladium(II) (182 mg, 0.25 mm ol, 0.05eq) and potassium phosphate (2.1 g, 9.98 mmol, 2eq) were added under nitrogen protection, and the reaction solution was stirred at 100°C for 2 hours. After the reaction was completed, it was quenched with water (50 mL), extracted with ethyl acetate (50 mL×3), and washed with saturated brine (50 mL×2). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 5:1) to obtain compound 377-1 (1.2 g, 71.6%).

[0942] LCMS: (ESI, m / z): 336.1 [M+H] + .

[0943] Step 2: Synthesis of compound 377-2

[0944] To a solution of 3-methyl-6-((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 377-1, 1.2 g, 3.58 mmol, 1 eq) in tetrahydrofuran (15 mL) was added n-butyllithium (1.72 mL, 4.30 mmol, 1.2 eq) at -78°C, and the mixture was stirred for half an hour. Compound NFSI (1.8 g, 5.73 mmol, 1.6 eq) was then added, and the reaction solution was warmed to room temperature and stirred for 12 hours. After completion of the reaction, the mixture was quenched with water (50 mL), extracted with ethyl acetate (50 mL × 3), washed with saturated brine (50 mL × 2), and the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 5:1) to give compound 377-2 (600 mg, 47.4%).

[0945] LCMS: (ESI, m / z): 354.0 [M+H] + .

[0946] Based on the similar procedures to synthesize compound 193, compound 377 (56 mg, 14.0%) was obtained.

[0947] LCMS: (ESI, m / z): 462.1 [M+H] + .

[0948] 1 H NMR(400MHz,DMSO-d6)δ12.33–11.93(m,1H),8.93–8.80(m,2H),8.25(dd, J=7.9,5.0Hz,1H),8.15(s,0.6H),8.13–8.06(m,1H),7.94(s,0.4H),7.73– 7.57(m,2H),6.85(d,J=190.4Hz,1H),4.88–4.75(m,1H),3.97–3.83(m,0. 5H),3.20–3.14(m,0.5H),3.05–2.71(m,2H),2.30(dd,J=17.2,1.9Hz,3H).

[0949] Example 49

[0950] Step 1: Synthesis of Compound 196-1

[0951] At room temperature, to a solution of 3-bromo-6-((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 193-1, 2 g, 4.99 mmol, 1 eq) in methanol (25 mL) were added compound 1,1'-bis(diphenylphosphino)ferrocenepalladium(II) dichloride (0.37 g, 0.5 mmol, 0.1 eq) and triethylamine (1.5 g, 14.9 mmol, 3.0 eq). The reaction solution was stirred at 120°C for 16 hours. After the reaction was completed, the reaction solution was concentrated, water (100 mL) was added to the residue, extracted with dichloromethane (100 mL×3), washed with saturated brine (100 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 5:1) to give compound 196-1 (1.5 g, 79.1%).

[0952] LCMS: (ESI, m / z): 380.1 [M+H] + .

[0953] Step 2: Synthesis of compound 196-2

[0954] To a solution of 6-((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine-3-carboxylate (compound 196-1, 1.5 g, 3.95 mmol, 1 eq) in tetrahydrofuran (20 mL) was added tetraisopropyl titanate (2.2 g, 7.9 mmol, 2.0 eq) at 0°C, and the mixture was stirred for 10 minutes. Ethylmagnesium bromide (2.1 g, 15.8 mmol, 4.0 eq) was then added dropwise. The reaction mixture was warmed to room temperature and stirred for 8 hours. After completion of the reaction, the mixture was quenched with water (100 mL), extracted with ethyl acetate (100 mL × 3), washed with saturated brine (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 5:1) to give compound 196-2 (450 mg, 30.1%).

[0955] LCMS: (ESI, m / z): 378.2 [M+H] + .

[0956] Step 3: Synthesis of Compound 196-3

[0957] To a solution of 1-(6-((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridin-3-yl)cyclopropan-1-ol (compound 196-2, 430 mg, 1.14 mmol, 1 eq) in dichloromethane (6 mL) was added diethylaminosulfur trifluoride (367.5 mg, 2.28 mmol, 2.0 eq) at room temperature. The reaction solution was stirred at room temperature for 1 hour. After the reaction was completed, the reaction solution was concentrated, water (40 mL) was added to the residue, and the mixture was extracted with ethyl acetate (40 mL×3), washed with saturated brine (40 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 8:1) to give compound 196-3 (320 mg, 74.0%).

[0958] LCMS: (ESI, m / z): 380.2 [M+H] + .

[0959] Based on the similar procedures to synthesize compound 193, compound 196 (11 mg, 11.4%) was obtained.

[0960] LCMS: (ESI, m / z): 462.1 [M+H] + .

[0961] 1 H NMR(400MHz,DMSO-d6)δ12.15(s,1H),9.12(d,J=38.7Hz,1H),8.85–8.78(m,1H ),8.28–8.22(m,1.5H),8.12–8.05(m,1.5H),7.99(dd,J=10.0,2.9Hz,1H),7.73 –7.60(m,2H),6.91(d,J=186.3Hz,1H),4.90–4.77(m,1H),4.00–3.91(m,0.5H) ,3.26–3.21(m,0.5H),3.06–2.76(m,2H),1.56–1.43(m,2H),1.25–1.16(m,2H).

[0962] Using conditions similar to those in the above examples, the compounds listed in Table 28 were prepared. The structural characterization data of these compounds are listed in Table 28.

[0963] Table 28

[0964] Example 50

[0965] Step 1: Synthesis of compound 115-1

[0966] To a solution of 6-(((triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridine (compound 133-2, 1000 mg, 3.11 mmol) in tetrahydrofuran (10 mL) was added n-butyllithium (1.87 mL, 4.67 mmol) at -78°C, and the mixture was stirred at -78°C for 30 minutes. N-fluorobisbenzenesulfonamide (1.47 g, 4.67 mmol) was then added to the system and stirred at room temperature for 1 hour. After the reaction was complete, saturated aqueous ammonium chloride was added, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by normal phase purification (ethyl acetate: petroleum ether = 1:1) to give compound 115-1 (600 mg, 56.82%).

[0967] LCMS: (ESI, m / z): 340.2 [M+H] + .

[0968] Based on the similar procedures to synthesize compound 193, compound 115 (66 mg, 23.8%) was obtained.

[0969] LCMS: (ESI, m / z): 448.2 [M+H] + .

[0970] 1 H NMR (400MHz, DMSO-d6) δ12.15(s,1H),8.96–8.79(m,2H),8.24(t,J=7.9Hz,1H),8.16(d,J=12.5Hz,1H),8.13–8.06(m,1H),7.72–7.57 (m,2H),7.31–7.16(m,1H),6.83(d,J=178.5Hz,1H),4.91–4.74(m,1H),3.98–3.83(m,0.5H),3.24–3.17(m,0.5H),3.04–2.74(m,2H).

[0971] Using conditions similar to those in the above examples, the compounds listed in Table 29 were prepared. The structural characterization data of these compounds are listed in Table 29.

[0972] Table 29

[0973] Example 51

[0974] Step 1: Synthesis of Compound 206-2

[0975] To a solution of 5-fluoropyridin-2-ol (compound 206-1, 10 g, 88.43 mmol, 1 eq) in DMF (100 mL) was added N-iodosuccinimide (29.8 g, 132.65 mmol, 1.5 eq) at -78°C. The reaction solution was stirred at room temperature for 12 hours. After completion of the reaction, water was added to quench the solution (200 mL), and the mixture was extracted with ethyl acetate (200 mL × 3), washed with saturated brine (200 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column (petroleum ether: ethyl acetate = 10:1) to give compound 206-2 (12 g, 56.7%).

[0976] LCMS: (ESI, m / z): 239.9 [M+H] + .

[0977] Step 2: Synthesis of compound 206-3

[0978] At room temperature, bis(triphenylphosphine)palladium(II) chloride (1.32 g, 1.88 mmol, 0.05 eq) and cuprous iodide (0.22 g, 1.13 mmol, 0.03 eq) were added to a solution of 5-fluoro-3-iodopyridin-2-ol (compound 206-2, 9 g, 37.66 mmol, 1 eq) in tetrahydrofuran (100 mL). The atmosphere was replaced with nitrogen, and triethylamine (11.4 g, 112.98 mmol, 3 eq) and ethyl acetate were added. Alkynyltrimethylsilane (6.29 g, 64.02 mmol, 1.7 eq), the reaction solution was stirred at 80 ° C for 16 hours. After the reaction was completed, the reaction solution was concentrated, water (100 mL) was added to the residue, extracted with ethyl acetate (100 mL × 3), washed with saturated brine (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column (petroleum ether: ethyl acetate = 6: 1) to give compound 206-3 (3.0 g, 38.0%).

[0979] LCMS: (ESI, m / z): 210.0 [M+H] + .

[0980] Step 3: Synthesis of Compound 206-4

[0981] To a toluene (30 mL) solution of 5-fluoro-3-((trimethylsilyl)ethynyl)pyridin-2-ol (compound 206-3, 3 g, 14.33 mmol, 1 eq) was added Lawesson's reagent (3.4 g, 8.60 mmol, 0.6 eq) at -78°C under nitrogen protection. The reaction solution was stirred at 120°C for 1 hour. After the reaction was completed, the reaction solution was concentrated and the crude product was purified by silica gel column (petroleum ether: ethyl acetate = 6:1) to obtain compound 206-4 (1.4 g, 43.3%).

[0982] LCMS: (ESI, m / z): 226.0 [M+H] + .

[0983] Step 4: Synthesis of Compound 206-5

[0984] At room temperature, m-chloroperbenzoic acid (2.1 g, 12.42 mmol, 2.0 eq) was added portionwise to a solution of 5-fluoro-2-(trimethylsilyl)thieno[2,3-b]pyridine (compound 206-4, 1.4 g, 6.21 mmol, 1 eq) in dichloromethane (15 mL). The reaction solution was stirred at room temperature for 2 hours. After the reaction, saturated aqueous sodium thiosulfate solution (100 mL) was added to the reaction mixture, extracted with ethyl acetate (100 mL × 3), washed with saturated brine (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column (petroleum ether: ethyl acetate = 1:1) to give compound 206-5 (450 mg, 45.3%).

[0985] LCMS: (ESI, m / z): 242.0 [M+H] + .

[0986] Step 5: Synthesis of Compound 206-6

[0987] At room temperature, 5-fluoro-7λ 4 To a solution of -thieno[2,3-b]pyridine-7-ol (compound 206-5, 650 mg, 2.69 mmol, 1 eq) in 1,2-dichloroethane (8 mL) was added trimethylsilyl cyanide (400.3 mg, 4.04 mmol, 1.5 eq), and the mixture was stirred at room temperature for half an hour. Then, dimethylaminocarbonyl chloride (433.9 mg, 4.04 mmol, 1.5 eq) was added, and the reaction solution was stirred at 80°C overnight. After the reaction was completed, the reaction solution was concentrated, and the crude product was purified by silica gel column (petroleum ether: ethyl acetate = 1:1) to obtain compound 206-6 (300 mg, 44.5%).

[0988] LCMS: (ESI, m / z): 251.0 [M+H] + .

[0989] Step 6: Synthesis of Compound 206-7

[0990] To a solution of 5-fluoro-2-(trimethylsilyl)thieno[2,3-b]pyridine-6-carbonitrile (compound 206-6, 220 mg, 0.88 mmol, 1 eq) in dichloromethane (4 mL) was slowly added diisopropylaluminum hydride (0.88 mL, 1.32 mmol, 1.5 eq) dropwise under nitrogen at -78°C. The reaction solution was stirred at -78°C for 1 hour. After the reaction was completed, the reaction solution was slowly added to an ice-cold saturated aqueous solution of potassium sodium tartrate (5 mL) and stirred until there was no turbidity. The solution was extracted with ethyl acetate (100 mL × 3), washed with saturated brine (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 1:1) to give compound 206-7 (40 mg, 17.9%).

[0991] LCMS: (ESI, m / z): 254.0 [M+H] + .

[0992] Based on the similar procedures to synthesize compound 103, compound 206 (2 mg, 8.9%) was obtained.

[0993] LCMS: (ESI, m / z): 448.0 [M+H] + .

[0994] Using conditions similar to those in the above examples, the compounds listed in Table 30 were prepared. The structural characterization data of these compounds are listed in Table 30.

[0995] Table 30

[0996] Example 52

[0997] Step 1: Synthesis of Compound 201-2

[0998] 1-(6-(((Triisopropylsilyl)oxy)methyl)thieno[3,2-c]pyridin-3-yl)ethan-1-one (compound 201-1, 300 mg, 0.83 mmol, 1 eq) and 1,2-ethanedithiol (0.17 mL, 2.07 mmol, 2.5 eq) were mixed and stirred under nitrogen protection, and boron trifluoride-acetic acid complex (0.13 mL, 0.91 mmol, 1.1 eq) was added and stirred for 30 minutes. After the reaction was completed, the reaction mixture was quenched by adding saturated aqueous sodium bicarbonate solution (10 mL) and extracted with ethyl acetate (3×10 mL). The organic phases were combined, backwashed with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column (PE / EA=3:1) to obtain compound 201-2 (140 mg, 59.86%).

[0999] LCMS: (ESI, m / z): 283.9 [M+H] + .

[1000] Step 2: Synthesis of Compound 201-3

[1001] (3-(2-methyl-1,3-dithiolan-2-yl)thieno[3,2-c]pyridin-6-yl)methanol (compound 201-2, 120 mg, 0.42 mmol, 1 eq) was dissolved in dichloromethane (2 mL), triethylamine (127.50 mg, 1.26 mmol, 3 eq) was added, and acetyl chloride (0.060 mL, 0.84 mmol, 2 eq) was added dropwise. The reaction was carried out at room temperature for 1 hour. After the reaction was completed, the reaction solution was dried and the crude product was directly used in the next step.

[1002] LCMS: (ESI, m / z): 325.9 [M+H] + .

[1003] Step 3: Synthesis of Compound 201-4

[1004] 1,3-Dibromo-5,5-dimethylhydantoin (291.64 mg, 1.02 mmol, 3 eq) was dissolved in dichloromethane (2 mL), and fluoroboric acid-pyridine (0.092 mL, 1.02 mmol, 3 eq) was added under nitrogen protection at -78 ° C. The mixture was stirred for 30 minutes, and (3-(2-methyl-1,3-dithiolan-2-yl)thieno[3,2-c]pyridin-6-yl)acetate (compound 201-3, 110 mg, 0.34 mmol, 1 eq) was added dropwise. The temperature was returned to room temperature and the reaction was allowed to react for 16 hours. After the reaction was completed, the reaction mixture was quenched by adding water (10 mL) and extracted with ethyl acetate (3 x 10 mL). The organic phases were combined, backwashed with saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column (PE / EA=1:5) to obtain compound 201-4 (110 mg).

[1005] LCMS: (ESI, m / z): 272.0 [M+H] + .

[1006] Step 4: Synthesis of Compound 201-5

[1007] Methyl (3-(1,1-difluoroethyl)thieno[3,2-c]pyridin-6-yl)acetate (compound 201-4, 100 mg, 0.37 mmol, 1 eq) was dissolved in methanol (1 mL), and potassium carbonate (102.28 mg, 0.74 mmol, 2 eq) was added. The mixture was stirred at room temperature for 1 hour. After the reaction was completed, the reaction solution was dried and purified by silica gel column chromatography (DCM / MEOH = 5:1) to obtain compound 201-5 (80 mg, 94.67%).

[1008] LCMS: (ESI, m / z): 230.1 [M+H] + .

[1009] Based on the similar procedures to synthesize compound 193, compound 201 (14.84 mg, 16.75%) was obtained.

[1010] LCMS: (ESI, m / z): 494.2 [M+H] + .

[1011] Example 53

[1012] Step 1: Synthesis of compound 314-2

[1013] To a solution of 3-bromo-6-chlorothieno[3,2-c]pyridine (compound 314-1, 560 mg, 2.25 mmol, 1 eq) in THF (10 mL) was added isopropylmagnesium chloride-lithium chloride (5.2 mL, 6.75 mmol, 1.3 M, 3 eq) at 0°C. After stirring for 1 hour, DMF (493.36 mg, 6.75 mmol, 3 eq) was added and the reaction was stirred for 3 hours. After the reaction was completed, the reaction solution was slowly poured into an aqueous ammonium chloride solution (50 mL) and diluted with stirring for 2 hours. Ethyl acetate (50 mL × 3) was added and extracted. The organic phases were combined, washed with saturated brine (30 mL × 3), dried over anhydrous sodium sulfate, filtered and concentrated to give the crude product, which was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 10:1) to give compound 314-2 (445 mg, 99.93%).

[1014] LCMS: (ESI, m / z): 197.9 [M+H] + .

[1015] Step 2: Synthesis of compound 314-3

[1016] To a solution of 6-chlorothieno[3,2-c]pyridine-3-carbaldehyde (compound 314-2, 450 mg, 2.28 mmol, 1 eq) in THF (2 mL) was added sodium borohydride (172.50 mg, 4.56 mmol, 2 eq) at 0°C and stirred at 0°C for 2 hours. After the reaction was completed, the crude product was concentrated and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to give compound 314-3 (344 mg, 75.67%).

[1017] LCMS: (ESI, m / z): 199.9 [M+H] + .

[1018] Steps 3 to 7:

[1019] Based on the similar procedures for the synthesis of compounds 132 and 133, compound 314-8 (120 mg, 46.2%) was obtained.

[1020] LCMS: (ESI, m / z): 544.3 [M+H] + .

[1021] Step 8: Synthesis of compound 314

[1022] (4-(3-(((tetrahydro-2H-pyran-2-yl)oxy)methyl)thieno[3,2-c]pyridin-6-yl)-1,4,6,7-tetrahydro-5H-imidazo[4,5-c]pyridin-5-yl)(5-(pyridin-2-yl)-1,3,4-oxadiazol-2-yl)methanone (compound 314-8, 110 mg, 0.20 mmol, 1 eq) was dissolved in ethanol (2 mL), and PPTS (5.03 mg, 0.020 mmol, 0.1 eq) was added. The reaction solution was stirred at 80° C. for 4 hours. A solid precipitated from the reaction solution, which was filtered and purified by reverse column chromatography (ammonia:methanol = 50%) to give compound 314 (84.41 mg, 90.78%).

[1023] LCMS: (ESI, m / z): 460.2 [M+H] + .

[1024] 1H NMR(400MHz,DMSO-d6)δ12.32–11.87(m,1H),9.21–8.90(m,1H),8.91–8.71(m ,1H),8.25(d,J=7.5Hz,1H),8.18(s,0.5H),8.10(t,J=7.7Hz,1H),7.95(s,0. 5H),7.75–7.55(m,3H),6.93(dd,J=190.9,60.8Hz,1H),5.52–5.22(m,1H),4. 98–4.63(m,3H),4.18–3.72(m,0.5H),3.20–3.11(m,0.5H),3.08–2.66(m,2H).

[1025] Using conditions similar to those in the above examples, the compounds listed in Table 31 were prepared. The structural characterization data of these compounds are listed in Table 31.

[1026] Table 31

[1027] Example 54

[1028] Step 1: Synthesis of compound 116-2

[1029] To a solution of thiophene-2,3-dicarbaldehyde (compound 116-1, 5.35 g, 38.17 mmol, 1 eq) in toluene (57 mL) were added ethane-1,2-diol (11.73 g, 188.98 mmol, 4.95 eq) and p-toluenesulfonic acid hydrate (0.1 g, 0.58 mmol, 0.016 eq) at room temperature, and the mixture was stirred at 110°C for 12 hours. After the reaction was completed, 5% aqueous sodium carbonate solution (100 mL) was added to the reaction mixture, extracted with ethyl acetate (3×100 mL), and washed with saturated brine (2×100 mL). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column (petroleum ether / ethyl acetate (0-20%)) to give compound 116-2 (4.5 g, 52%).

[1030] LCMS: (ESI, m / z): 229.0 [M+H] + .

[1031] Step 2: Synthesis of compound 116-3

[1032] Under nitrogen protection, n-butyl lithium (5.4 mL, 13.5 mmol, 1.1 eq) was added to a solution of 2,2'-(thiophene-2,3-diyl)bis(1,3-dioxolane) (compound 116-2, 2.8 g, 12.27 mmol, 1 eq) in tetrahydrofuran (30 mL) at -78 ° C. The mixture was stirred at -78 ° C for 0.5 h, and N-chlorosuccinimide (2.46 g, 18.41 mmol, 1.5 eq) was added. q), continued stirring at -78 ° C for 30 minutes, then warmed to room temperature and stirred for 2 hours. After the reaction was completed, water (80 mL) was added to the reaction mixture to quench the reaction mixture, extracted with ethyl acetate (3×80 mL), washed with saturated brine (2×80 mL), and the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The resulting residue was purified by silica gel column (petroleum ether / ethyl acetate (0-20%)) to give compound 116-3 (1200 mg, 40.3%).

[1033] LCMS: (ESI, m / z): 262.9 [M+H] + .

[1034] Step 3: Synthesis of compound 116-4

[1035] Under nitrogen protection, to a solution of 2,2'-(thiophene-2,3-diyl)bis(1,3-dioxolane) (compound 116-3, 1.2 g, 4.57 mmol, 1 eq) in tetrahydrofuran (15 mL) was added 5N aqueous hydrochloric acid solution (4 mL) at room temperature, and the mixture was stirred at room temperature overnight. After the reaction was completed, water (50 mL) was added to the reaction mixture, and the mixture was extracted with ethyl acetate (3×50 mL) and washed with saturated brine (2×50 mL). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The resulting residue was purified by silica gel column (petroleum ether / ethyl acetate (0-50%)) to give compound 116-4 (700 mg, 87.7%).

[1036] LCMS: (ESI, m / z): 175.0 [M+H] + .

[1037] Based on the similar procedures to synthesize compound 094, compound 116 (55 mg, 29.42%) was obtained.

[1038] LCMS: (ESI, m / z): 464.1 [M+H] + .

[1039] 1H NMR (400MHz, DMSO-d6) δ12.12(d,J=32.3Hz,1H),8.93(d,J=39.6Hz,1H),8.82(d,J=3.9Hz,1H),8.25(t,J=7.9Hz,1H),8.20(s,0.6H),8.13–8.06(m ,1H),8.01(s,0.4H),7.74–7.54(m,3H),6.82(d,J=183.8Hz,1H),4.96–4 .71(m,1H),4.04–3.82(m,0.5H),3.28–3.14(m,0.5H),3.08–2.72(m,2H).

[1040] Using conditions similar to those in the above examples, the compounds listed in Table 32 were prepared. The structural characterization data of these compounds are listed in Table 32.

[1041] Table 32

[1042] Example 55

[1043] Step 1: Synthesis of Compound 104-2

[1044] To a solution of 6-chlorothieno[3,2-c]pyridine-2-carboxylic acid (7.1 g, 33.23 mmol) in tert-butanol (80 mL) were added di-tert-butyl dicarbonate (14.5 g, 66.46 mmol) and 4,4-dimethylaminopyridine (0.81 g, 6.65 mmol) at room temperature and stirred at room temperature for 16 hours. After the reaction was completed, the reaction mixture was concentrated under reduced pressure and the residue was purified by silica gel column chromatography (ethyl acetate / petroleum ether (10-60%)) to give compound 104-2 (7.1 g, 79.20%).

[1045] LCMS: (ESI, m / z): 270.0 [M+H] + .

[1046] Step 2: Synthesis of Compound 104-3

[1047] To a solution of 2,2,6,6-tetramethylpiperidine (3.28 g, 23.20 mmol) in tetrahydrofuran (40 mL) was added n-butyllithium (9.34 mL, 23.36 mmol) dropwise at -78°C. After stirring for 10 minutes, a solution of tert-butyl 6-chlorothieno[3,2-c]pyridine-2-carboxylate (4.2 g, 15.57 mmol) in tetrahydrofuran (10 mL) was added dropwise. The mixture was stirred at -78°C for 1 hour, then the temperature was raised to room temperature and stirred for 16 hours. After the reaction was completed, the reaction solution was poured into 100 mL of saturated aqueous ammonium chloride solution and extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography (ethyl acetate / petroleum ether (10-60%)) to give compound 104-3 (400 mg, 8.93%).

[1048] LCMS: (ESI, m / z): 288.0 [M+H] + .

[1049] Step 3: Synthesis of Compound 104-4

[1050] To a solution of tert-butyl 6-chloro-3-fluorothieno[3,2-c]pyridine-2-carboxylate (400 mg, 1.39 mmol) in dichloromethane (4 mL) was added trifluoroacetic acid (1 mL) at room temperature and stirred at room temperature for 1 hour. After the reaction was completed, the mixture was concentrated to give compound 104-4 (300 mg, 93.17%).

[1051] LCMS: (ESI, m / z): 231.9 [M+H] + .

[1052] Step 4: Synthesis of compound 104-5

[1053] Under nitrogen protection, silver carbonate (72 mg, 0.26 mmol) and acetic acid (3.9 mg, 0.06 mmol) were added to a solution of 6-chloro-3-fluorothieno[3,2-c]pyridine-2-carboxylic acid (300 mg, 1.30 mmol) in dimethyl sulfoxide (3 mL), and the mixture was stirred at 140°C for 16 hours. After the reaction was completed, the reaction mixture was concentrated under reduced pressure and the residue was purified by silica gel column chromatography (ethyl acetate / petroleum ether (0-50%)) to give compound 104-5 (200 mg, 82.3%).

[1054] LCMS: (ESI, m / z): 188.1 [M+H] + .

[1055] Based on the similar procedures to synthesize compound 094, compound 104 (30 mg, 26.2%) was obtained.

[1056] LCMS: (ESI, m / z): 448.1 [M+H] + .

[1057] Purification by SFC (system: SHIMADZU LC-20AP; column: Column size: 250*30mm*10μm; mobile phase A: supercritical CO2, mobile phase B: ethanol (+0.1% 7.0mol / L ammonia methanol solution); A:B=60:40; wavelength: 214nm; flow rate: 140ml / min; column temperature: room temperature; back pressure: 100bar; injection volume: 8.0mL; cycle time: 8.0min; solvent: ethanol: distillation grade, supercritical CO2: food grade) to obtain compound 104-A (front peak, 12.1mg, SFC, Rf=5.64min, 10.56%) and compound 104-B (back peak, 13.2mg, SFC, Rf=9.98min, 11.52%).

[1058] 104-A

[1059] 1 H NMR(400MHz, DMSO-d6)δ12.13(d,J=29.7Hz,1H),9.00(d,J=35.1Hz,1H),8.85–8.79(m,1H),8.28–8.07(m,3H),7.75–7 .58(m,3H),6.86(d,J=189.3Hz,1H),4.96–4.73(m,1H),4.01–3.88(m,0.5H),3.29–3.17(m,0.5H),3.08–2.74(m,2H).

[1060] LCMS: (ESI, m / z): 448.1 [M+H] + .

[1061] 104-B

[1062] 1 H NMR(400MHz, DMSO-d6)δ12.13(d,J=27.1Hz,1H),9.00(d,J=34.8Hz,1H),8.87–8.78(m,1H),8.28–8.06(m,3H),7.73–7 .57(m,3H),6.87(d,J=186.0Hz,1H),4.91–4.77(m,1H),4.02–3.87(m,0.5H),3.29–3.21(m,0.5H),3.07–2.73(m,2H).

[1063] LCMS: (ESI, m / z): 448.1 [M+H] + .

[1064] Example 56

[1065] Step 1: Synthesis of compound 368-2

[1066] Under nitrogen protection, methyl 3-methylthieno[3,2-c]pyridine-6-carboxylate (compound 368-1, 3 g, 14.48 mmol, 1 eq) was dissolved in DMF (30 mL) and N-chlorosuccinimide (5.80 g, 43.44 mmol, 3 eq) was slowly added. The mixture was stirred at 60°C for 2 hours. After the reaction was completed, the reaction solution was filtered, the filtrate was added to water, and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by spin silica gel column chromatography (ethyl acetate / petroleum ether (0-50%)) to give compound 368-2 (1.1 g, 31.44%).

[1067] LCMS: (ESI, m / z): 242.0 [M+H] + .

[1068] Based on the similar procedures to synthesize compound 094, compound 368 (30 mg, 26.2%) was obtained.

[1069] LCMS: (ESI, m / z): 479.0 [M+H] + .

[1070] Example 57

[1071] Step 1: Synthesis of compound 132

[1072] Under nitrogen protection, zinc powder (2.49 mg, 0.038 mmol), zinc cyanide (22.31 mg, 0.19 mmol), 1,1-di-tert-butyl 4-(3-bromothieno[3,2-c]pyridin-6-yl)-1,4,6,7-tetrahydro-5H-imidazo[4,5-c]pyridine-1,5-dicarboxylate (compound 113-1, 100 mg, 0.19 mmol) in N,N-dimethylformamide (3 mL) were added at 25 °C. '-Bis(diphenylphosphino)ferrocene (21.07 mg, 0.038 mmol) and tris(dibenzylideneacetone)dipalladium (17.40 mg, 0.019 mmol), the mixture was stirred at 120 ° C for 12 hours. After the reaction, the reaction solution was poured into water, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 60%) to obtain compound 113-2 (70 mg, 77.83%).

[1073] LCMS: (ESI, m / z): 382.2 [M+H] + .

[1074] Based on the similar procedures to synthesize compound 310, compound 113 (7 mg, 10.8%) was obtained.

[1075] LCMS: (ESI, m / z): 455.2 [M+H] + .

[1076] Example 58

[1077] Step 1: Synthesis of compound 360-2

[1078] To a solution of potassium 3-ethoxy-3-oxopropanoate (compound 360-1, 16.35 g, 96.08 mmol, 1 eq) in ethyl acetate (160 mL) were added magnesium chloride (27.44 g, 288.24 mmol, 3.0 eq) and triethylamine (48.61 g, 480.4 mmol, 5.0 eq) at 0°C. Under carbon monoxide protection, the mixture was stirred at 120°C for 16 hours. In another reaction flask, to a solution of 1-fluorocyclopropane-1-carboxylic acid (10 g, 96.08 mmol, 1 eq) in tetrahydrofuran (100 mL) was added oxalyl chloride (12.20 g, 96.08 mmol, 1 eq). The reaction was catalyzed by adding 1% 4-[ ...

[1079] LCMS: (ESI, m / z): 175.2 [M+H] + .

[1080] Step 2: Synthesis of compound 360-3

[1081] To a solution of ethyl 3-(1-fluorocyclopropyl)-3-oxopropanoate (compound 360-2, 2.5 g, 14.35 mmol, 3 eq) in glacial acetic acid (10 mL) was added aqueous sodium nitrate solution (0.99 g, 14.35 mmol, 1.0 eq) at 0°C, and the mixture was stirred for 30 minutes, then stirred at room temperature for 0.5 hours. After the reaction, water (100 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (100 mL × 3), washed with saturated brine (100 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated to give crude compound 360-3 (2.8 g, 96.0%), which was directly used in the next reaction.

[1082] LCMS: (ESI, m / z): 204.2 [M+H] + .

[1083] Step 3: Synthesis of compound 360-4

[1084] To a mixed solution of ethyl (E)-3-(1-fluorocyclopropyl)-2-(hydroxyimino)-3-oxopropanoate (compound 360-3, 2.8 g, 7.58 mmol, 1 eq) in glacial acetic acid (30 mL) and acetic anhydride (10 mL) was added palladium carbon (0.31 g, 2.88 mmol, 0.38 eq) at room temperature. The reaction solution was stirred at room temperature for 3 hours. After the reaction was completed, the reaction solution was concentrated and the crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 1:1) to give compound 360-4 (2.5 g, 78.4%).

[1085] LCMS: (ESI, m / z): 232.2 [M+H] + .

[1086] Step 4: Synthesis of compound 360-5

[1087] To a solution of ethyl 2-acetylamino-3-(1-fluorocyclopropyl)-3-oxopropanoate (compound 360-4, 2.3 g, 9.95 mmol, 1 eq) in chloroform (30 mL) was added phosphorus pentoxide (8.47 g, 59.70 mmol, 6.0 eq) at room temperature. The reaction solution was stirred at 600°C for 12 hours. After the reaction was completed, water was added and the mixture was extracted with ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 6:1) to give compound 360-5 (1.3 g, 84.1%).

[1088] LCMS: (ESI, m / z): 214.1 [M+H] + .

[1089] Based on the similar procedures to synthesize compound 193, compound 360 (80 mg, 20%) was obtained.

[1090] LCMS: (ESI, m / z): 506.1 [M+H] + .

[1091] 1H NMR (400MHz, DMSO-d6) δ12.30 (s, 1H), 8.96 (d, J = 35.4Hz, 1H), 8.06 (d, J = 82.1Hz ,1H),7.70(d,J=31.7Hz,1H),7.49(d,J=9.8Hz,1H),6.89(d,J=185.2Hz,1H),5. 04–4.67(m,1H),4.02–3.83(m,0.5H),3.21–3.13(m,0.5H),3.07–2.75(m,2H),2 .60–2.55(m,3H),2.46(d,J=13.4Hz,3H),1.64–1.53(m,2H),1.38–1.28(m,2H).

[1092] Example 59

[1093] Step 1: Synthesis of compound 351-2

[1094] To a solution of 4,6-dichloronicotinonitrile (compound 351-1, 8 g, 46.24 mmol, 1 eq) and methyl thioglycolate (5.15 g, 48.55 mmol, 1.05 eq) in DCE (70 mL) was added DBU (20.70 mL, 138.72 mmol, 3 eq) at 0°C and stirred for 1 hour. After the reaction was complete, water (100 mL) was added to the reaction system, stirred for 10 minutes, and filtered. The filter cake was slurried with ethanol (10 mL) and filtered and dried to give compound 351-2 (8 g, 71.29%).

[1095] LCMS: (ESI, m / z): 243.0 [M+H] + .

[1096] Step 2: Synthesis of compound 351-3

[1097] At room temperature, tert-butyl nitrite (5.49 mL, 46.35 mmol, 1.5 eq) and iodine (8.23 g, 23.45 mmol, 1.05 eq) were added to a solution of methyl 3-amino-6-chlorothieno[3,2-c]pyridine-2-carboxylate (compound 351-2, 7.5 g, 30.90 mmol, 1 eq) in acetonitrile (80 mL). The mixture was stirred at 80°C for 2 hours. After the reaction, water (100 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (100 mL×3). The organic phases were combined, washed with saturated aqueous sodium chloride solution (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated to give a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 3 / 1) to give compound 351-3 (3.5 g, 32.03%).

[1098] LCMS: (ESI, m / z): 353.9 [M+H] + .

[1099] Step 3: Synthesis of compound 351-4

[1100] At room temperature, methyl 6-chloro-3-iodothieno[3,2-c]pyridine-2-carboxylate (compound 351-3, 3.5 g, 9.90 mmol, 1 eq) was dissolved in methanol (40 mL), and a solution of lithium hydroxide (0.71 g, 29.70 mmol, 3 eq) in 10 mL was added. The mixture was stirred at room temperature for 16 hours. After the reaction was completed, the reaction solution was dried and slurried with water (50 mL). The mixture was filtered and dried to give compound 351-4 (2.7 g, 80.33%).

[1101] LCMS: (ESI, m / z): 339.9 [M+H] + .

[1102] Step 4: Synthesis of compound 351-5

[1103] At room temperature, 6-chloro-3-iodothieno[3,2-c]pyridine-2-carboxylic acid (compound 351-5, 2.7 g, 7.95 mmol, 1 eq) was dissolved in dimethyl sulfoxide (40 mL), and silver carbonate (0.44 g, 1.59 mmol, 0.2 eq) and acetic acid (0.024 g, 0.40 mmol, 0.05 eq) were added. The mixture was stirred at 140°C for 24 hours. After the reaction was completed, water (50 mL) was added to the reaction solution to dilute it, and the mixture was extracted with ethyl acetate (50 mL × 3). The organic phases were combined, washed with saturated brine (50 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated to give compound 351-5 (2.3 g, 97.87%).

[1104] LCMS: (ESI, m / z): 295.9 [M+H] + .

[1105] Step 5: Synthesis of compound 351-6

[1106] At room temperature, 6-chloro-3-iodothieno[3,2-c]pyridine (compound 351-5, 2.1 g, 7.11 mmol, 1 eq) was dissolved in NMP (20 mL), and cuprous iodide (1.35 g, 7.11 mmol, 1 eq) and methyl 2,2-difluoro-2-(fluorosulfonyl)acetate (4.10 g, 21.33 mmol, 3 eq) were added. The mixture was stirred at 80°C under nitrogen for 2 hours. After the reaction, water (30 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (30 mL × 3). The organic phase was washed with saturated aqueous sodium chloride solution (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 5 / 1) to obtain compound 351-6 (1.2 g, 71.07%).

[1107] LCMS: (ESI, m / z): 238.0 [M+H] + .

[1108] Step 6: Synthesis of compound 351-7

[1109] 6-Chloro-3-(trifluoromethyl)thieno[3,2-c]pyridine (compound 351-6, 0.9 g, 3.79 mmol, 1 eq) was dissolved in NMP (10 mL), and zinc cyanide (0.89 g, 7.58 mmol, 2 eq) and (1,1'-bis(diphenylphosphino)ferrocene)dichloropalladium(II) (0.55 g, 0.76 mmol, 0.2 eq) were added. The reaction was carried out at 140°C for 16 hours. After the reaction, water (20 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (20 mL×3). The organic phases were combined and washed with saturated aqueous sodium chloride solution (20 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a crude product. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 3 / 1) to obtain compound 351-7 (450 mg, 52.07%).

[1110] LCMS: (ESI, m / z): 229.1 [M+H] + .

[1111] Step 7: Synthesis of compound 351-8

[1112] 3-(Trifluoromethyl)thieno[3,2-c]pyridine-6-carbonitrile (compound 351-7, 0.3 g, 3.79 mmol, 1 eq) was dissolved in 4 M hydrochloric acid in methanol (15 mL) and stirred at 60°C for 16 hours. After the reaction was completed, the reaction solution was concentrated to obtain a crude product, which was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 3 / 1) to obtain compound 351-8 (300 mg, 87.36%).

[1113] LCMS: (ESI, m / z): 262.1 [M+H] + .

[1114] Step 8: Synthesis of compound 351-9

[1115] Methyl 3-(trifluoromethyl)thieno[3,2-c]pyridine-6-carboxylate (compound 351-8, 0.3 g, 1.15 mmol, 1 eq) was dissolved in tetrahydrofuran (4 mL). Under nitrogen protection, diisobutylaluminum hydride (3.07 mL, 4.6 mmol, 4 eq) was added dropwise at -78°C and stirred at -78°C for 1 hour. After the reaction was completed, saturated potassium sodium tartrate solution was added and stirred overnight. The mixture was extracted with ethyl acetate (20 mL×3). The organic phases were combined, washed with saturated sodium chloride aqueous solution (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate = 3 / 2) to obtain compound 351-9 (260 mg, 97.92%).

[1116] LCMS: (ESI, m / z): 232.1 [M+H] + .

[1117] Step 9: Synthesis of compound 351-10

[1118] 3-(Trifluoromethyl)thieno[3,2-c]pyridine-6-carbaldehyde (compound 351-9, 260 mg, 1.12 mmol, 1 eq) was dissolved in a mixed solvent of ethanol (3 mL) and water (3 mL), and histamine dihydrochloride (247.39 mg, 1.34 mmol, 1.2 eq) and sodium hydroxide (134.4 mg, 3.36 mmol, 3 eq) were added. The reaction solution was stirred at 80°C for 1 hour. After the reaction was completed, it was cooled to room temperature and the pH was adjusted to 7-8 with 1M hydrochloric acid. The reaction solution was purified by C18 reverse phase column (0.05% ammonia water) to obtain compound 351-10 (350 mg, 95.96%).

[1119] LCMS: (ESI, m / z): 325.1 [M+H] + .

[1120] Step 10: Synthesis of compound 351

[1121] 6-(4,5,6,7-Tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)-3-(trifluoromethyl)thieno[3,2-c]pyridine (Compound 351-10, 120 mg, 0.37 mmol, 1 eq) and potassium 5-(2-methyloxazol-4-yl)-1,3,4-oxadiazole-2-carboxylate (129.44 mg, 1.55 mmol, 1.5 eq) were dissolved in a mixed solvent of acetonitrile (3 mL) and pyridine (0.5 mL). Phosphorus oxychloride (134.4 mg, 3.36 mmol, 3 eq) was added dropwise at -20°C and stirred at -20°C for 2 hours. After the reaction was completed, saturated aqueous sodium bicarbonate solution (1 mL) was added to the reaction solution to quench the reaction. The product was spin-dried and purified by high performance liquid chromatography under the following conditions (Chromatography column specifications: Waters 2767 / QDA Column: Sunfire C18 21.2×250mm 10μm; mobile phase A: 0.1% FA water, mobile phase B: acetonitrile; flow rate: 20mL / min; elution gradient: 24-27%; detection wavelength: 254nm / 214nm; retention time (min): 7-9min) to obtain compound 351 (71mg, 38.27%).

[1122] SFC (system: Waters SFC 150; column: Column size: 250×30mm 10μm; mobile phase A: supercritical CO2, mobile phase B: ethanol (+0.1% 7.0mol / L amine methanol solution); A:B: 65:35; wavelength: 214nm; flow rate: 140mL / min; column temperature: RT; injection volume: 2.2mL; cycle time: 3.15min; solvent: ethanol: distillation grade, supercritical CO2: food grade) was separated and purified to obtain compound 351-A (front peak, Rf=1.576min, 26.24mg, 14.14%) and compound 351-B (back peak, Rf=2.868min, 27.13mg, 14.62%).

[1123] 351-A

[1124] 1 H NMR (400MHz, DMSO-d6) δ12.31–11.98(m,1H),9.16–8.98(m,2H),8.69(d,J=9.2Hz,1H),8.32(d,J=80.7Hz,1H),7.65(d,J=38.0 Hz,1H),6.91(d,J=202.6Hz,1H),5.00–4.69(m,1H),4.00–3.83(m,0.6H),3.25–3.16(m,0.4H),3.07–2.73(m,2H),2.54(s,3H).

[1125] LCMS: (ESI, m / z): 502.1 [M+H] + .

[1126] 351-B

[1127] 1 H NMR (400MHz, DMSO-d6) δ12.32–11.97(m,1H),9.20–8.98(m,2H),8.69(d,J=9.2Hz,1H),8.32(d,J=80.7Hz,1H),7.65(d,J=38.0 Hz,1H),6.89(d,J=202.6Hz,1H),5.00–4.73(m,1H),4.00–3.83(m,0.6H),3.23–3.16(m,0.4H),3.09–2.71(m,2H),2.54(s,3H).

[1128] LCMS: (ESI, m / z): 502.1 [M+H] + .

[1129] Using conditions similar to those in the above examples, the compounds listed in Table 33 were prepared. The structural characterization data of these compounds are listed in Table 33.

[1130] Table 33

[1131] Example 60

[1132] Step 1: Synthesis of Compound 407-2

[1133] Under nitrogen, 1-fluorocyclopropane-1-carboxylic acid (10 g, 96.08 mmol, 1 eq) and CDI (18.70 g, 115.30 mmol, 1.2 eq) were dissolved in THF (100 mL) and stirred at room temperature for 2 hours. Ethyl 2-isocyanate (11.95 g, 105.69 mmol, 1.1 eq) and LiHMDS (96.08 ml, 96.08 mmol, 1 M, 1 eq) were added at 0°C. The reaction was returned to room temperature and stirred overnight. After the reaction was completed, the reaction solution was slowly poured into saturated aqueous ammonium chloride (200 mL) for dilution and extracted with ethyl acetate (100 mL × 2). The combined organic phases were washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered and concentrated to give the crude product, which was purified by silica gel column chromatography (petroleum ether: ethyl acetate = 2:1) to give compound 407-2 (15 g, 78.38%).

[1134] Step 2: Synthesis of Compound 407-3

[1135] 6N hydrochloric acid (100 mL) was added to ethyl 5-(1-fluorocyclopropyl)oxazole-4-carboxylate (10 g, 50.21 mmol, 1 eq), and the mixture was stirred at 100°C for 2 hours. After the reaction was completed, the reaction solution was concentrated to obtain the hydrochloride salt of compound 407-3 (7.71 g, 100%).

[1136] LCMS: (ESI, m / z): 118.2 [M+H] + .

[1137] Step 3: Synthesis of Compound 407-4

[1138] To a solution of 2-amino-1-(1-fluorocyclopropyl)ethan-1-one hydrochloride (7 g, 45.58 mmol, 1 eq) and TEA (13.84 g, 136.74 mmol, 3 eq) in toluene (100 mL) was added ethyl oxalyl chloride (6.22 g, 45.58 mmol, 1 eq) at 0°C. The reaction was stirred for 1 hour. After completion of the reaction, the reaction solution was concentrated and purified by silica gel column chromatography (petroleum ether:ethyl acetate = 3:1) to give compound 407-4 (2 g, 20.2%).

[1139] LCMS: (ESI, m / z): 218.1 [M+H] + .

[1140] Step 4: Synthesis of Compound 407-5

[1141] To a solution of ethyl 2-((2-(1-fluorocyclopropyl)-2-oxoethyl)amino)-2-oxoacetate (2 g, 9.21 mmol, 1 eq) in toluene (30 mL) was added phosphorus oxychloride (8.41 mL, 92.10 mmol, 10 eq), and the reaction was stirred at 120°C for 16 hours. After the reaction was completed, the reaction solution was concentrated and purified by silica gel column chromatography (petroleum ether:ethyl acetate = 3:1) to give compound 407-5 (1.2 g, 65.43%).

[1142] LCMS: (ESI, m / z): 200.0 [M+H] + .

[1143] Based on the similar procedures to synthesize compound 193, compound 407 (80 mg, 20%) was obtained.

[1144] LCMS: (ESI, m / z): 492.0 [M+H] + .

[1145] 1H NMR (400MHz, DMSO-d6) δ12.14(s,1H),8.98(d,J=30.4Hz,1H),8.06(d,J=78.5Hz,1 H),7.86–7.76(m,1H),7.65(d,J=37.4Hz,1H),7.49(d,J=9.6Hz,1H),6.85(d,J=16 1.1Hz,1H),4.89–4.72(m,1H),4.06–3.84(m,0.5H),3.25–3.15(m,0.5H),3.03–2. 73(m,2H),2.46(d,J=12.8Hz,3H),1.64–1.54(m,2H),1.35–1.27(q,J=8.1Hz,2H).

[1146] Example 61 Note: "*" indicates that the carbon atom at the position is chiral carbon and the configuration is one of R or S, for example express One of them.

[1147] Step 1: Synthesis of compound 399-1

[1148] 6-(4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridin-4-yl)-3-(trifluoromethyl)thieno[3,2-c]pyridine (compound 351-10, 2.15 g, 6.63 mmol, 1 eq) was dissolved in dichloromethane (40 mL) solvent, and NCS (885 mg, 6.63 mmol, 1 eq) and DIEA (2.19 mL, 13.26 mmol, 2 eq) were added. The reaction solution was stirred at 40 ° C for 2 hours, cooled to room temperature, washed with saturated sodium thiosulfate aqueous solution, and the organic phase was concentrated to obtain compound 399-1 (1.8 g, 84.24%).

[1149] LCMS: (ESI, m / z): 323.1 [M+H] + .

[1150] Step 2: Synthesis of compound 399-2

[1151] 6-(6,7-Dihydro-1H-imidazo[4,5-c]pyridin-4-yl)-3-(trifluoromethyl)thieno[3,2-c]pyridine (399-1, 1.8 g, 5.58 mmol, 1 eq) was dissolved in methanol (40 mL), p-toluenesulfonic acid (960 mg, 5.58 mmol, 1 eq) and RuCl[(R,R)-Fsdpen](p-cymene) (200 mg, 0.28 mmol, 0.05 eq) were added, and then the mixture was added to another bottle at 0°C. Formic acid (1.05 mL, 27.9 mmol, 5 eq) and triethylamine (1.55 mL, 11.16 mmol, 2 eq) were added, and the mixed solution of formic acid and triethylamine was added to the mixture of raw materials. The reaction solution was stirred at room temperature for 2 hours. After the reaction was completed, it was concentrated, and dichloromethane and water were added. After extraction, the aqueous phase was retained, and the pH of the aqueous phase was adjusted to 10 with 1N sodium hydroxide aqueous solution, and then extracted three times with dichloromethane. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated to obtain compound 399-2 (1.2 g, 66.25%).

[1152] LCMS: (ESI, m / z): 325.1 [M+H] + .

[1153] Step 3: Synthesis of Compound 399

[1154] (R / S)-6-(4,5,6,7-tetrahydro-1H-imidazo(4,5-c)pyridin-4-yl)-3-(trifluoromethyl)thieno[3,2-c]pyridine (compound 399-2, 80 mg, 0.25 mmol, 1 eq) and potassium 5-(1-methyl-1H-pyrazol-3-yl)-1,3,4-oxadiazole-2-carboxylate (87 mg, 0.38 mmol, 1.5 eq) were dissolved in tetrahydrofuran (2 mL), and DIEA (64.62 mg, 0.50 mmol) was added. , 2eq), stirred at room temperature under nitrogen for 10 minutes, and diphenyl chlorophosphate (100.74 mg, 0.38 mmol, 1.5 eq) was added dropwise, and stirred at room temperature for 2 hours. After the reaction, 1.6 mL of 1 M hydrochloric acid was added to the reaction solution, and the mixture was stirred for 10 minutes. The tetrahydrofuran was spin-dried and the mixture was washed with 1.6 mL of dichloromethane and 0.5 mL of water. The organic phase was washed 4-5 times with 0.8 mL of 15% potassium carbonate aqueous solution, dried over anhydrous sodium sulfate, filtered, concentrated, and purified by Prep-HPLC (column: XBridge C18 19*250 mm, 10 μm; mobile phase A: 10 mmol / L ammonia aqueous solution, mobile phase B: acetonitrile; flow rate: 20 mL / min; gradient: 34% to 38%; retention time: 8.50–9.50 min / 16 min) to give compound 399 (40.02 mg, 32.42%).

[1155] LCMS: (ESI, m / z): 501.0 [M+H] + .

[1156] 1 H NMR (400MHz, DMSO-d6) δ12.23–11.96(m,1H),9.22–8.95(m,1H),8.78–8.59(m,1H),8.49–8.10(m,1H),7.99(s,1H),7.77–7.42(m ,1H),7.39–6.46(m,2H),5.07–4.65(m,1H),3.99(d,J=2.6Hz,3H),3.97–3.74(m,0.5H),3.27–3.16(m,0.5H),3.08–2.68(m,2H).

[1157] Using conditions similar to those in the above examples, the compounds listed in Table 34 were prepared. The structural characterization data of these compounds are listed in Table 34.

[1158] Table 34 Note: “*” indicates that the carbon atom at that position is a chiral carbon and has one of the configurations R or S.

[1159] Biological evaluation

[1160] Test Example 1: Cell-based PAH R408W enzyme activity detection

[1161] 1. Add 90 μL of cells (stable A375 cells with PAH R408W mutation constructed by lentivirus) to a 96-well plate, with a final seeding density of 40,000 cells per well.

[1162] 2. After standing for 1 hour, the compound was added to the above cell wells (initial concentration 20 μM, 2-fold dilution), and the control wells contained an equal amount of DMSO.

[1163] 3. After adding the compound, incubate the plate in a cell culture incubator with 5% CO2 at 37°C for 23 hours. The incubator needs to have enough water in the tray to ensure uniform humidity.

[1164] 4. Prepare a solution of septopterin plus 13C9,15N-phenylalanine by adding 20 μL of 100 mM septopterin stock solution and 800 μL of 100 mM 13C9,15N-phenylalanine stock solution to 10 mL of DMEM medium without puromycin.

[1165] 5. Add 10 μL of this solution directly to each well of the cell plate to a final concentration of 20 μM septerin and 800 μM 13C9,15N-phenylalanine.

[1166] 6. Incubate the cell plate in a 5% CO2, 37°C cell culture incubator for 4 hours.

[1167] 7. After incubation, transfer the culture medium from the plate to a new 96-well plate, seal with aluminum foil and freeze at -80°C until use in analysis (sample plate).

[1168] 8. Transfer 10 μL of each 13C,15N-tyrosine standard to separate wells of a new 96-well plate. Add 200 μL of extraction buffer to each well and seal the plate for analysis. Used for calibration curve preparation (calibration plate).

[1169] 9. Transfer 10 μL of the culture medium from the sample plate wells to a new 96-well plate. Add 200 μL of extraction buffer to each well and seal the plate for analysis.

[1170] 10. The sample plate and the standard curve plate were analyzed by Rapidfire-MS.

[1171] Data Analysis:

[1172] AC 50 The drug concentration that achieves 50% of the maximum biological effect, that is, the drug concentration at which the concentration of tyrosine generated reaches 50% of the maximum concentration of tyrosine that can be generated, is calculated as follows: the tyrosine generation concentration is detected by LCMS, and the tyrosine generation concentration and compound concentration are fitted using GraphPad Prism 9 software to determine the maximum concentration that can generate tyrosine, and the drug concentration at which the tyrosine generation concentration reaches 50% of the maximum concentration of tyrosine generated, that is, AC 50 value and draw a dose-effect curve.

[1173] The representative compounds in this article were tested for their ability to restore the activity of phenylalanine hydroxylase mutant protein through the above experiments. The AC 50 See Table A for values.

[1174] Table A Representative compounds herein for restoring the activity of PAH mutant proteins AC 50 value

[1175] Experimental conclusion: The above representative compounds can effectively restore the activity of phenylalanine hydroxylase mutant protein.

[1176] Test Example 2: Caco-2 cell permeability test

[1177] 1) Cell Culture: Resuscitated Caco-2 cells were cultured in DMEM supplemented with 10% FBS, 1% Penicillin-Streptomycin Liquid (100X), and 1% NEAA in a 37°C incubator containing 5% CO2. The medium was replaced every 2-3 days, and cells were subcultured when confluence reached approximately 80%.

[1178] 2) Preparation of transport buffer: 5 mL of 1 M HEPES was added to 500 mL of HBSS (with or without BSA) to prepare transport buffer (pH 7.4). The final concentration of HEPES (with or without BSA) was 10 mM.

[1179] 3) Before the transport experiment (Days 14-28), remove the culture medium from the cell culture plate, wash the plate twice with prewarmed HBSS buffer (10 mM HEPES, pH 7.4), and add 100 μL of HBSS buffer to each well. After incubation at 37°C for 30 minutes, measure the TEER value using a Millicell-ERS2 cell resistance meter to confirm the integrity and compactness of the cell monolayer.

[1180] 4) Discard the HBSS buffer and transfer the upper plate containing the cell monolayer to the corresponding receiving plate. Add the test compound solution or buffer to the apical side (A side) and basolateral side (B side), respectively. Place the cell plate in a matching 96-well receiving plate and group according to Table B. Add the prepared positive control drug, test compound, or HBSS (with or without BSA) buffer to the A side or B side of the Transwell cell plate filter, respectively. Incubate the cell plate at 37°C.

[1181] Table B Compound grouping scheme LY: Lucifer Yellow TB: Transport buffer (HBSS containing 10 mM HEPES (with or without BSA))

[1182] 1) After 5 minutes of incubation, take 8 μL of sample from each of the A and B dosing sides and add 72 μL of transport buffer (diluted 10-fold) to serve as the initial dosing solution T0.

[1183] 2) After 120 minutes of incubation, samples were collected from both ends of the Transwell cell plate filter membrane. 80 μL of sample solution was taken from the receiving side as the receiving side sample after transport. 8 μL of sample was taken from the dosing side and added to 72 μL of transport buffer as the dosing side sample after transport.

[1184] 3) After collecting bilateral samples, gently tap the plate to discard any remaining solution. Rinse the plate with transport buffer and add 100 μL of acetonitrile containing internal standard working solution to each well to lyse the cells. Mix thoroughly by pipetting up and down, then pipette 80 μL of cell lysate into each well as the post-cell lysis sample.

[1185] 4) After collecting all samples in the transport system (T0 sample, receiving side sample, and donor side sample), add 160 μL of IS / ACN solution to each well, add 80 μL of IS / ACN solution to the collected lysate sample, and then seal.

[1186] 5) Shake for 10 minutes and store the sample plate at -20°C before LC-MS / MS analysis.

[1187] 6) Prepare a standard curve, then pipette 8 μL of the standard curve solution and 72 μL of transport buffer and mix with 160 μL of IS / ACN to serve as the standard curve sample.

[1188] 7) All the above samples were centrifuged at 6000 RPM for 10 min, 100 μL of supernatant was collected, 100 μL of H 2 O was added and mixed thoroughly, and then LC-MS / MS analysis was performed.

[1189] The results of the Caco-2 cell permeability test of the test compounds are shown in Table C.

[1190] Table C Caco-2 cell permeability results of compounds

[1191] Experimental conclusion: The above representative compounds have better cell permeability and lower efflux properties compared with the control compounds.

[1192] Test Example 3 Effect of the Test Compound on hERG Potassium Ion Channel

[1193] 1) Cell Preparation: After subculturing HEK-293-hERG cells to an appropriate state, rinse with PBS (or DPBS), dissociate with Tryple solution, resuspend the cells in culture medium, and store in a centrifuge tube. After centrifugation, discard the supernatant and resuspend the cells in extracellular medium until ready for use. Store at 2-8°C. Prior to patch clamp recording, plate the cells dropwise into a culture dish to ensure that the cells are at a certain density and are individually isolated.

[1194] 2) Electrophysiological testing: hERG currents were recorded using the whole-cell patch clamp technique. Cell suspensions were placed in a small culture dish and placed on an inverted microscope stage. After the cells adhered, extracellular solution was perfused at a recommended flow rate of 1-2 mL / min. Glass microelectrodes were drawn in two steps using a microelectrode puller. After filling with solution, the resistance in water was 2-5 MΩ.

[1195] After establishing whole-cell recording mode, maintain the clamp potential at -80 mV. A depolarizing voltage of +60 mV was applied for 850 ms, followed by a repolarization to -50 mV for 1275 ms to elicit hERG tail currents. This pulse sequence was repeated every 15 seconds throughout the experiment.

[1196] After the current stabilizes, the drug is administered through continuous extracellular perfusion from low to high concentrations. Starting from a low concentration, perfusion is continued until the drug effect stabilizes, and then the next concentration is perfused.

[1197] Experimental results

[1198] The effects of the compounds of the present invention on hERG potassium channels were determined by the above experiments. The results are shown in Table D.

[1199] Table D Effects of compounds on hERG potassium channels

[1200] Experimental conclusion: The inhibitory effects of the above representative compounds on hERG channels are weaker than those of the control compounds, and they have a lower risk of clinical cardiotoxicity and are safer.

[1201] Test Example 4 Pharmacokinetic Study of the Test Compound in Male SD Rats

[1202] Experimental methods

[1203] On the day of administration, the test substance was prepared using a medium of 5% DMSO + 5% Solutol + 90% Saline, and the administration solution was prepared and ready for use.

[1204] The IV dose was 1 mg / kg, and the PO dose was 2 mg / kg. The subjects were fasted overnight before administration.

[1205] Weigh the animals before dosing, calculate the dose based on body weight, and administer the drug once IV and once orally on the day of dosing. Blood (approximately 0.2 mL) was collected from an appropriate vein at 0.083, 0.25, 0.5, 1, 2, 4, 8, and 24 hours after IV dosing, and at 0.083, 0.25, 0.5, 1.0, 2.0, 4.0, 6.0, 8.0, and 24.0 hours after orally dosing. The blood was placed in an EDTA-K2 anticoagulant blood collection tube and centrifuged at 4000 rpm for 6 minutes (2-8°C). Plasma was separated within 1 hour and transferred to labeled EP tubes (the label should at least indicate the study designation, animal number, collection time, and date) and stored at -80°C until testing. Blood collection and centrifugation were performed in an ice bath. Food and water were administered 4 hours after dosing. Test substance concentrations in SD rat plasma were determined using a validated LC-MS / MS method.

[1206] Experimental results

[1207] The pharmacokinetics of the compounds of the present invention in male SD rats were determined by the above experiments. The results are shown in Table E.

[1208] Table E Pharmacokinetic study results of the test compound in male SD rats (IV 1 mpk / PO 2 mpk)

[1209] Experimental conclusion: The above representative compounds have better PK properties in rats than the control compounds.

[1210] Unless otherwise specified, the structures of the reference compounds described in the present invention are as follows:

[1211] The above is an exemplary description of the implementation methods of the technical solution of the present invention. It should be understood that the scope of protection of the present invention is not limited to the above implementation methods. Any modifications, equivalent substitutions, improvements, etc. made by those skilled in the art within the spirit and principles of the present invention shall be included in the scope of protection of the claims of this application.

Claims

1. A compound represented by formula (I), its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt, or prodrug compound: in, The fused ring group is formed by the fusion of ring A and ring B, and ring A is selected from a benzene ring, a 5-6 membered heteroaromatic ring, a saturated or partially unsaturated C 3-12 Carbocyclic ring or 3-14 membered heterocyclic ring; Ring B is selected from 5 membered heteroaromatic ring, saturated or partially unsaturated C 3-12 carbocyclic ring or 3-14 membered heterocyclic ring; and when ring A is a 5-membered heteroaromatic ring, ring B is a 5-membered heteroaromatic ring; Each R a The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: OH, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, halo C 1-12 Alkyl, C 1-12 Alkoxy, C 1-12 Alkylthio, C 3-12 Cycloalkyl, 3-14 membered heterocyclic group, C 6-14 Aryl, 5-14 membered heteroaryl, -N(R 11 )(R 12 ) or -S(O)2R 13 ; or, two adjacent R a Together with the atoms to which they are attached, they form an unsubstituted or optionally substituted R a1 Substituted C 3-6 Cycloalkyl ring, 3-6 membered heterocyclic ring, C 6-10 aromatic ring or 5-10 membered heteroaromatic ring; each R a1 The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R a2 Substituted with the following groups: OH, NH2, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group, C 6-10 Aryl, 5-10 membered heteroaryl, -N(R 14 )(R 15 ) or -S(O)2R 16 ; Each R a2 the same or different, independently selected from OH, -N(R 17 )(R 18 ), CN, halogen, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl or 3-6 membered heterocyclic group; R 11 、R 12 、R 13 、R 14 、R 15 、R 16 、R 17 、R 18 The same or different, independently selected from H, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 3-12 Cycloalkyl; Each R c the same or different, independently selected from deuterium or C 1-12 alkyl; X1 is selected from O or S; Y1 is absent or selected from unsubstituted or optionally substituted with one, two or more R d Substituted with the following groups: C 6-14 Arylene, 5-14 membered heteroarylene, C 6-14 Arylene-C 1-12 Alkylene, 5-14 membered heteroaryl-C 1-12 Alkylene, C 6-14 Arylene-O-, 5-14 membered heteroarylene-O-, C 6-14 Arylene-NH-, 5-14 membered heteroarylene-NH-, C 6-14 Arylene-CO- or 5-14 membered heteroarylene-CO-; each R d The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R d1 Substituted with the following groups: OH, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 1-12 Alkoxy, C 1-12 Alkylthio, C 3-12 Cycloalkyl, 3-14 membered heterocyclic group, C 6-14 Aryl, 5-14 membered heteroaryl, -N(R 21 )(R 22 ), or -S(O)2R 23 ; Each R d1 the same or different, independently selected from OH, -N(R 24 )(R 25 ), CN, halogen, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group; R 21 、R 22 、R 23 、R 24 、R 25 The same or different, independently selected from H, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 3-12 Cycloalkyl; Ring C is selected from C 6-14 Aromatic ring or 5-14 membered heteroaromatic ring; Each R b The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R b1 Substituted with the following groups: OH, C 1-12 Alkyl, C 2-12 Alkenyl, C 2-12 Alkynyl, halo C 1-12 Alkyl, C 1-12 Alkoxy, C 1-12 Alkylthio, C 3-12 Cycloalkyl, C 3-12 Carbocyclic ring, 3-14 membered heterocyclic ring, C 6-14 Aryl, 5-14 membered heteroaryl, -N(R 31 )(R 32 ), hydroxyl C 1-12 Alkyl, C 1-12 Alkyl-N(R 31 )(R 32 )、-S(O)2R 33 、 Each R b1 The same or different, independently selected from oxo (=O), OH, -N(R 34 )(R 35 ), CN, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, halogenated C 1-6 Alkyl, hydroxyl C 1-12 Alkyl, C 1-12 Alkyl-N(R 34 )(R 35 ), C 3-6 Cycloalkyl, 3-6 membered heterocyclic group; R 31 、R 32 、R 33 、R 34 、R 35 、R 36 、R 37 、R 38 、R 39 The same or different, independently selected from H, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 3-12 Cycloalkyl; m1 is selected from 0, 1, 2 or 3; m2 is selected from 0, 1, 2, 3, 4, 5 or 6; m3 is selected from 0, 1, 2, 3, 4, 5, 6, 7 or 8; n is selected from 0, 1 or 2.

2. The compound according to claim 1, its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or prodrug compound, wherein: m3 is 0; Preferably, m3 is 1; Preferably, R c selected from methyl; Preferably, n is 1; Preferably, Y1 is absent or selected from unsubstituted or optionally substituted with one, two or more R d substituted 5-membered heteroarylene, 5-membered heteroarylene-CH2-, 5-membered heteroarylene-O-, 5-membered heteroarylene-NH-; Preferably, Y1 is absent; Preferably, Y1 is selected from unsubstituted or optionally substituted with one R d Substituted groups: The "*" side is connected to the carbonyl group, and the "#" side is connected to the ring C; Preferably, each R d the same or different, independently selected from CN, halogen, OH, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy or C 3-6 Cycloalkyl; Preferably, each R d are the same or different and are independently selected from methyl, difluoromethyl, trifluoromethyl, Cl, Br or cyclopropyl; Preferably, Y1 is absent or selected from The "*" side is connected to the carbonyl group, and the "#" side is connected to the ring C.

3. The compound according to claim 1 or 2, its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or prodrug compound thereof, characterized in that: Ring C is selected from phenyl or 5-10 membered heteroaryl; Preferably, ring C is selected from a benzene ring, a pyrrole ring, a thiophene ring, a furan ring, a pyrazole ring, an imidazole ring, a thiazole ring, an oxazole ring, an isoxazole ring, a dioxazole ring, a pyridine ring, a pyridone ring, a pyrimidine ring, and a pyrazolopyridine ring; Preferably, ring C is selected from Preferably, each R b The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R b1 Substituted with the following groups: OH, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 1-12 Alkoxy, C 1-12 Alkylthio, C 3-12 Cycloalkyl, 3-14 membered heterocyclic group, C 6-14 Aryl, 5-14 membered heteroaryl, -N(R 31 )(R 32 )、-S(O)2R 33 、 Each R b1 the same or different, independently selected from OH, -N(R 34 )(R 35 ), CN, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, halogenated C 1-6 Alkyl, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group; R 31 、R 32 、R 33 、R 34 、R 35 、R 36 、R 37 、R 38 、R 39 The same or different, independently selected from H, C 1-12 Alkyl, halogenated C 1-12 Alkyl, C 3-12 Cycloalkyl; Preferably, each R b the same or different, independently selected from CN, F, Cl, Br, CH3, CHF2, CF3, CH2CH3, CH2CF3, -C≡CCH3, CHOCH3, CH2CN, CH2NHCH3, CH(CH3)2, C(CH3)3, CH2CH2OH, C(CH3)2OH, NH2, NHCH3, N(CH3)2, CH2CH2N(CH3)2, OH, OCH3, OCF3, cyclopropyl, cyclobutyl, cyclopentyl, -S(O)2-cyclopropyl, Preferably, m2 is selected from 0, 1 or 2; Preferably, Selected from 4. The compound according to any one of claims 1 to 3, its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or prodrug compound, characterized in that: Ring A is selected from a benzene ring or a 5-6 membered heteroaromatic ring; Preferably, ring A is selected from a benzene ring, a pyrazole ring, a triazole ring, a thiazole ring, an oxazole ring, a pyridine ring, a pyrimidine ring, a pyridone ring, a pyridazine ring, and a pyrazine ring; Preferably, ring A is selected from in Indicates that it is fused to ring B at this position; Preferably, ring B is selected from a 5-membered heteroaromatic ring and a 5-8-membered heteroaromatic ring; and when ring A is a 5-membered heteroaromatic ring, ring B is a 5-membered heteroaromatic ring; Preferably, ring B is selected from a thiazole ring, an isothiazole ring, an oxazole ring, a pyrazole ring, an imidazole ring, a triazole ring, a thiophene ring, a pyrrole ring, a furan ring, a dihydrofuran ring, a tetrahydrofuran ring, and an isoxazol-3-one ring; Preferably, ring B is selected from in Indicates that it is fused to ring A at this position; Preferably, Selected from Preferably, each R a The same or different, independently selected from CN, halogen, unsubstituted or optionally substituted by one, two or more R a1 Substituted with the following groups: OH, C 1-6 Alkyl, C 2-6 Alkenyl, C 2-6 Alkynyl, halo C 1-6 Alkyl, C 1-6 Alkoxy, C 1-6 Alkylthio, C 3-6 Cycloalkyl, 3-8 membered heterocyclic group, phenyl, 5-6 membered heteroaryl, -N(R 11 )(R 12 ); or, two adjacent R a Together with the atoms to which they are attached, they form C 3-6 Cycloalkyl, 3-8 membered heterocyclic group, phenyl, 5-6 membered heteroaryl; each R a1 the same or different, independently selected from CN, halogen, OH, NH2, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 1-6 Alkoxy, C 3-6 Cycloalkyl, 3-6 membered heterocyclic group, C 6-10 Aryl, 5-10 membered heteroaryl, -N(R 14 )(R 15 );R 11 、R 12 、R 14 、R 15 The same or different, independently selected from H, C 1-6 Alkyl, halogenated C 1-6 Alkyl, C 3-6 Cycloalkyl; Preferably, each R a are the same or different and are independently selected from methyl, ethyl, isopropyl, tert-butyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, vinyl, propenyl, 1-fluorovinyl, 2-fluorovinyl, ethynyl, CN, F, Cl, Br, CD3, CH2OH, C(CH3)2OH, NH2, NHCH3, N(CH3)2, OCH3, OCH2CH3, OCH(CH3)2, OCF3, OCHF2, SCH3, CF3, CHF2, CF2CH3, CH2CF3, CH2CN, Tetrahydropyranyl (such as ), morpholinyl (such as ), phenyl (such as ), pyridyl (such as ), 3-fluoropyridyl (such as ), 3-chloropyridyl (such as ), 3-methylpyridyl (such as ); or two adjacent R a Together with the atoms to which they are attached, they form a benzene ring; Preferably, m1 is selected from 0, 1 or 2; Preferably, Selected from 5. The compound according to any one of claims 1 to 4, its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or prodrug compound, characterized in that: The compound represented by formula (I) has the structure shown below: Among them, ring A, ring B, ring C, Y1, R a 、R b 、R c , m1, m2, m3, n have the definitions in any one of claims 1-4; Preferably, the compound represented by formula (I) has the structure shown below: Among them, ring A, ring B, ring C, R a 、R b , m1, m2 have the definitions in any one of claims 1-4; Preferably, the compound represented by formula (I) has the structure shown below: Among them, R a 、R b , m1, m2 have the definitions in any one of claims 1-4; Preferably, the compound represented by formula (I) has the structure shown below: Among them, R a Selected from halogen (such as F, Cl, Br), C 1-4 Alkyl (such as methyl, ethyl), halogenated C 1-4 Alkyl (e.g., trifluoromethyl, difluoromethyl); R b , m2 have the definitions as described herein.

6. The compound according to any one of claims 1 to 5, its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or prodrug compound, characterized in that: The compound represented by formula (I) is selected from the following structures:

7. A method for preparing the compound of formula (I) according to any one of claims 1 to 6, comprising the following steps A: in, R' is H or an alkali metal ion (such as K + 、Li + ); Ring A, Ring B, Ring C, Y1, R a 、R b 、R c , m1, m2, m3, n have the definitions as described in any one of claims 1-4.

8. A pharmaceutical composition comprising a therapeutically effective amount of at least one of the compound of formula (I) according to any one of claims 1 to 6, its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt, or prodrug compound thereof; Preferably, the pharmaceutical composition further comprises one or more pharmaceutically acceptable excipients; Preferably, the pharmaceutical composition further contains one or more additional therapeutic agents.

9. A method for treating or preventing a disease or condition caused by a phenylalanine hydroxylase mutation, comprising administering to a patient a preventively or therapeutically effective amount of at least one of the compound of formula (I) according to any one of claims 1 to 6, its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt, or prodrug compound thereof, or the pharmaceutical composition according to claim 8; Preferably, the phenylalanine hydroxylase mutation contains at least one R408W, R261Q, R243Q, Y414C, L48S, A403V, ​​I65T, R241C, L348V, R408Q or V388M mutation; Preferably, the phenylalanine hydroxylase mutation contains at least one R408W, Y414C, 165T, F39L, R408Q, L348V, R261Q, A300S or L48S mutation; Preferably, the phenylalanine hydroxylase mutation contains at least one R408W mutation; Preferably, the phenylalanine hydroxylase mutation contains two R408W mutations; Preferably, the disease or condition caused by the phenylalanine hydroxylase mutation is phenylketonuria; Preferably, the patient comprises a mammal, preferably a human.

10. Use of the compound of formula (I) according to any one of claims 1 to 6, its racemate, stereoisomer, tautomer, solvate, polymorph, pharmaceutically acceptable salt or prodrug thereof, or the pharmaceutical composition according to claim 8 in the preparation of a medicament; Preferably, the use is for preparing a drug for treating or preventing a disease or condition caused by a phenylalanine hydroxylase mutation; Preferably, the phenylalanine hydroxylase mutation contains at least one R408W, R261Q, R243Q, Y414C, L48S, A403V, ​​I65T, R241C, L348V, R408Q or V388M mutation; Preferably, the phenylalanine hydroxylase mutation contains at least one R408W, Y414C, 165T, F39L, R408Q, L348V, R261Q, A300S or L48S mutation; Preferably, the phenylalanine hydroxylase mutation contains at least one R408W mutation; Preferably, the phenylalanine hydroxylase mutation contains two R408W mutations; Preferably, the disease or disorder caused by the phenylalanine hydroxylase mutation is phenylketonuria.

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