HK1234049B - Imidazo isoindole derivative, preparation method therefor and medical use thereof - Google Patents

Description

Imidazolidinone derivatives, preparation methods thereof and their use in medicine

Technical Field

The present invention belongs to the field of medicine and relates to imidazoisoindole derivatives, their preparation methods and their applications in medical research. The present invention discloses that the derivatives are used as IDO inhibitors for treating diseases with pathological characteristics of the IDO-mediated tryptophan metabolic pathway, including cancer, Alzheimer's disease, autoimmune diseases, depression, anxiety, cataracts, psychological disorders and AIDS.

Background Art

Cancer is one of the major diseases that seriously endangers human life, with over half of cases occurring in developing countries. The incidence of malignant tumors in my country is generally on the rise, increasing at an average annual rate of 3% to 5%. It is estimated that by 2020, 4 million people in my country will develop cancer and 3 million will die from it. The main causes are aging, urbanization, industrialization, and changes in lifestyle. In China's hospital drug market, sales of anti-cancer drugs have steadily increased in recent years, reaching 66.42 billion yuan in 2012, a year-on-year increase of 13.07%. By 2017, the market size of anti-cancer drugs is expected to reach 105.57 billion yuan, a year-on-year increase of 7.57%.

Due to the unrestricted growth, infiltration, and metastasis of malignant tumors, the three conventional treatments currently used in clinical practice (surgery, radiotherapy, and chemotherapy) are unable to completely remove or eradicate tumor cells, leading to frequent tumor metastasis or recurrence. Tumor biotherapy is a new approach to cancer prevention and treatment that utilizes modern biotechnology and related products. Due to its safety, effectiveness, and low adverse reaction profile, it has become the fourth mode of cancer treatment after surgery, radiotherapy, and chemotherapy. It achieves anti-tumor effects by mobilizing the host's natural defense mechanisms (such as inhibiting IDO-mediated tumor immune escape) or administering naturally occurring, highly targeted substances.

Indoleamine-pyrrole-2,3-dioxygenase (IDO) is a heme-containing monomeric protein composed of 403 amino acid residues, including two folded α-helical domains. The larger domain contains the catalytic pocket, where substrates interact with IDO through hydrophobic interactions. IDO catalyzes the conversion of tryptophan to formylkynurenine. It is widely distributed in tissues other than the liver in humans and other mammals (rabbits and mice). It is the only rate-limiting enzyme outside the liver that catalyzes the catabolism of tryptophan, an essential amino acid for cell activation and proliferation and an essential component of proteins. IDO is closely associated with various cytokines, including interferons (IFNs), interleukins (ILs), and tumor necrosis factor, which can activate IDO under certain conditions. There is a regulatory point in the T cell cycle that is highly sensitive to tryptophan levels. On the one hand, IDO depletes local tryptophan, causing T cells to arrest in the mid-G1 phase, thereby inhibiting T cell proliferation. On the other hand, kynurenine, the main product of IDO-catalyzed tryptophan metabolism, induces changes in intracellular oxidants and antioxidants through the mediation of oxygen free radicals, inducing T cell apoptosis. This is an inherent immunosuppressive mechanism in the body. Numerous studies have demonstrated that IDO is highly expressed in leukemia cells, inhibiting local T cell proliferation, suppressing T cell-mediated immune responses, and blocking T cell activation signaling, thereby mediating tumor cell evasion from the immune system. It has been found that most human tumors constitutively express IDO. Therefore, IDO is a potential target for cancer immunotherapy.

Disclosed patent applications for inhibitors that selectively inhibit IDO include WO2012142237, WO2004094409, WO2006122150, WO2007075598, WO2010005958, and WO2014066834.

IDO inhibitors hold great promise as pharmaceuticals, but currently no promising marketable IDO inhibitors have been identified. To achieve better tumor treatment outcomes and better meet market demand, we seek to develop a new generation of highly effective, low-toxic, and selective IDO inhibitors. This invention provides a novel selective IDO inhibitor structure and demonstrates that compounds with this novel structure exhibit excellent efficacy and effects, particularly superior pharmacokinetic and absorption activity.

Summary of the Invention

The object of the present invention is to provide a compound represented by general formula (I) or its tautomers, mesomers, racemates, enantiomers, diastereomers, or mixtures thereof, or pharmaceutically acceptable salts thereof, wherein the compound represented by general formula (I) has the following structure:

in:

M is an inorganic acid or an organic acid, preferably trifluoroacetic acid;

A is selected from cycloalkyl, heterocyclyl, aryl and heteroaryl, wherein the cycloalkyl, heterocyclyl, aryl and heteroaryl are optionally further substituted with one or more substituents selected from alkyl, halogen, amino, nitro, hydroxy, cyano, alkoxy, hydroxyalkyl, cycloalkyl, heterocyclyl, aryl and heteroaryl;

R ¹ is selected from a hydrogen atom, an alkyl group, a cycloalkyl group, a heterocyclic group, an aryl group, a heteroaryl group, -OR ⁴ , -C(O)R ⁴ and -C(O)OR ⁴ , -S(O) _m R ⁴ , -NR ⁵ R ⁶ , -C(O)NR ⁵ R ⁶ , -C(O)NHR ⁵ , -NR ⁵ C(O)R ⁶ and -NR ⁵ S(O) _m R ⁶ ;

^R2 are the same or different and are each independently selected from hydrogen, alkyl, haloalkyl, alkoxy, haloalkoxy, halogen, amino, nitro, hydroxy, cyano, cycloalkyl, heterocyclic, aryl and heteroaryl;

^R3 are the same or different and are each independently selected from hydrogen, alkyl, haloalkyl, alkoxy, haloalkoxy, halogen, amino, nitro, hydroxy, cyano, cycloalkyl, heterocyclic, aryl, heteroaryl, -OR4, -C(O) ^R4 , -C(O) ^OR4 , ^-S (O) _mR4 , ^-NR5R6 , -C(O) ^NR5R6 , ^-NR5C (O) ^R6 and ^-NR5S (O _)mR6 ^{; wherein} the alkyl, haloalkyl ^{, cycloalkyl} , heterocyclic, aryl and heteroaryl are optionally further selected from alkyl, haloalkyl, halogen, amino, nitro, cyano, hydroxy, alkoxy, haloalkoxy, hydroxyalkyl ^, cycloalkyl, heterocyclic, aryl, heteroaryl, -Ra, ^-OR7 , -C(O) ^R7 , -C(O) ^OR7 , -S(O) _m R ⁷ , -NR ⁷ R ⁸ , -C(O)NR ⁷ R ⁸ , -NR ⁷ C(O)R ⁸ and -NR ⁷ S(O) _m R ⁸ ;

^Ra is selected from the group consisting of alkyl, haloalkyl, cycloalkyl, heterocyclyl, aryl and heteroaryl, wherein the alkyl, haloalkyl, cycloalkyl, heterocyclyl, aryl and heteroaryl are optionally further substituted with one or more substituents selected from the group consisting of alkyl, halogen, amino, nitro, hydroxy, alkoxy, hydroxyalkyl, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, ^-OR4 , -C(O) ^R4 , -C(O) ^OR4 , -S ⁽ O ⁾ _mR4 , ^-NR5R6 , -C(O) ^NR5R6 , ^-NR5C (O) ^R6 and ^-NR5S (O ⁾ _mR6 ^;

^R4 is selected from the group consisting of hydrogen, alkyl, haloalkyl, hydroxy, amino, alkoxy, haloalkoxy, cycloalkyl, heterocyclic, aryl and heteroaryl; wherein the alkyl, haloalkyl, cycloalkyl, heterocyclic, aryl and heteroaryl are optionally further substituted by one or more substituents selected from the group consisting of alkyl, halogen, amino, nitro, cyano, hydroxy, hydroxyalkyl, alkoxy, cycloalkyl, heterocyclic, aryl, heteroaryl, -R ^a , -OR ⁷ , -C(O)R ⁷ , -C(O)OR ⁷ , -S(O) _m R ⁷ , -NR ⁷ R ⁸ , -C(O)NR ⁷ R ⁸ , -NR ⁷ C(O)R ⁸ and -NR ⁷ S(O) _m R ⁸ ;

R ⁵ and R ⁶ are the same or different and are each independently selected from a hydrogen atom, an alkyl group, a hydroxyl group, an amino group, a cycloalkyl group, a heterocyclic group, an aryl group and a heteroaryl group, wherein the alkyl group, the cycloalkyl group, the heterocyclic group, the aryl group and the heteroaryl group are optionally further substituted by one or ^more substituents selected from an alkyl group, a halogen group, a hydroxyl group, an amino group, a nitro group, a cyano group, an alkoxy group, a hydroxyalkyl group, a cycloalkyl group, a heterocyclic group, an aryl group and a heteroaryl group, -R ^a , -OR ⁷ , -C(O) _R ⁷ , -C(O)OR ⁷ , -S(O) m ^{R 7} , -NR ⁷ R 8 ^, -C(O)NR 7 R ^{8 , -NR 7 C(O)R 8 and} -NR ⁷ S(O) _m R ⁸ ;

R ⁷ and R ⁸ are the same or different and are each independently selected from a hydrogen atom, an alkyl group, a hydroxyl group, an amino group, a cycloalkyl group, a heterocyclic group, an aryl group, and a heteroaryl group, wherein the alkyl group, the cycloalkyl group, the heterocyclic group, the aryl group, and the heteroaryl group are optionally further substituted with one or more substituents selected from an alkyl group, a halogen group, a hydroxyl group, an amino group, a nitro group, a cyano group, an alkoxy group, a hydroxyalkyl group, a cycloalkyl group, a heterocyclic group, an aryl group, and a heteroaryl group;

p is an integer of 0, 1, 2, 3 or 4;

y is an integer of 0, 1, 2 or 3;

m is an integer of 0, 1 or 2; and

n is an integer of 0, 1, 2, 3, 4 or 5.

In a preferred embodiment of the present invention, a compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein y is 0, 1 or 3, especially 0.

In a preferred embodiment of the present invention, a compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein A is selected from heterocyclic groups and cycloalkyl groups, wherein the cycloalkyl groups and heterocyclic groups are optionally further substituted with one or more substituents selected from alkyl groups, halogen, amino groups, nitro groups, hydroxyl groups, cyano groups, alkoxy groups, hydroxyalkyl groups, cycloalkyl groups, heterocyclic groups, aryl groups and heteroaryl groups.

In a preferred embodiment of the present invention, a compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, wherein n is an integer of 0, 1 and 2.

In a preferred embodiment of the present invention, a compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound represented by general formula (II) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof,

in:

X is CH or N;

R ¹ to R ³ , M, p, n and y are as defined in the general formula (I);

a is an integer of 0, 1, 2 or 3; and

b is an integer of 0, 1, 2 or 3.

In a preferred embodiment of the present invention, a compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound represented by general formula (II-A) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof,

in:

R ¹ to R ³ , M, p, n and y are as defined in the general formula (I).

In a preferred embodiment of the present invention, a compound represented by general formula (II-A) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound represented by general formula (II-B) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof,

in:

R ² , R ⁵ , R ⁶ , M, p and y are as defined in the general formula (I).

In a preferred embodiment of the present invention, a compound represented by general formula (II) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound represented by general formula (III) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof,

in:

R ¹ to R ³ , M, p, n and y are as defined in the general formula (I).

In a preferred embodiment of the present invention, a compound represented by general formula (III) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound represented by general formula (IV) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof,

in:

R ² , R ³ , M, p, n and y are as defined in the general formula (I).

In a preferred embodiment of the present invention, a compound represented by general formula (III) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound represented by general formula (IV-1) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof,

in:

R ² , ^Ra , M, p and y are as defined in the general formula (I).

In a preferred embodiment of the present invention, a compound represented by general formula (III) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound represented by general formula (IV-2) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof,

in:

R ² , R ⁴ , M, p and y are as defined in the general formula (I).

In a preferred embodiment of the present invention, a compound represented by general formula (IV) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, is a compound represented by general formula (IV-A) and general formula (IV-B) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof,

in:

R ² , R ³ , M, p, n and y are as defined in the general formula (I).

In a preferred embodiment of the present invention, a compound represented by general formula (V) or its tautomer, mesomer, racemate, enantiomer, diastereomer or a mixture thereof is an intermediate for preparing a compound represented by general formula (II) or its tautomer, mesomer, racemate, enantiomer, diastereomer or a mixture thereof,

in:

Q is an inorganic acid or an organic acid, preferably trifluoroacetic acid;

X is CH or N;

R ² , p, a and b are as defined in formula (II); and

x is an integer of 0, 1, 2, or 3.

In a preferred embodiment of the present invention, a method for preparing a compound of formula (II-1) or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, the method comprising:

The compound of general formula (V) undergoes a coupling reaction with a halide of R ¹ in the presence of a catalyst under alkaline conditions, and the obtained product is optionally further reacted with a boronic acid or boronic ester of R ³ to obtain a compound of general formula (II-1);

in:

X is N;

R ¹ to R ³ , p, n, a and b are as defined in formula (II); and

Q and x are defined as described in formula (V).

In a preferred embodiment of the present invention, a method for preparing a salt of a compound of formula (II) or a tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof comprises:

The compound of general formula (II-1) is salified under acidic conditions to obtain a compound of general formula (II);

in:

X is CH or N;

R ¹ to R ³ , M, p, y, n, a and b are as defined in the general formula (II).

Another aspect of the present invention relates to a pharmaceutical composition comprising a therapeutically effective dose of a compound of the formula or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents, or excipients. The present invention also relates to a method for preparing the above composition, comprising mixing the compound of the formula or its tautomer, mesomer, racemate, enantiomer, diastereomer, or mixture thereof, or a pharmaceutically acceptable salt thereof, with a pharmaceutically acceptable carrier, diluent, or excipient.

The present invention further relates to the use of a compound represented by general formula (I) or its tautomers, mesomers, racemates, enantiomers, diastereomers or mixtures thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the same, in the preparation of a medicament for preventing and/or treating diseases having pathological characteristics of the IDO-mediated tryptophan metabolic pathway. IDO inhibitors can be used to inhibit cardiac disorders and treat other diseases with pathological features of the IDO-mediated tryptophan metabolic pathway, including viral infections such as AIDS, cellular infections such as Lyme disease and streptococcal infections, neurodegenerative disorders (e.g., Alzheimer's disease, Huntington's disease, and Huntington's disease), autoimmune diseases, depression, anxiety, cataracts, psychological disorders, AIDS, cancer (including T-cell leukemia and colon cancer), eye disease states (e.g., cataracts and age-related yellowing), and autoimmune diseases, wherein the cancer can be selected from breast cancer, cervical cancer, colon cancer, lung cancer, stomach cancer, rectal cancer, pancreatic cancer, brain cancer, skin cancer, oral cancer, prostate cancer, bone cancer, kidney cancer, ovarian cancer, bladder cancer, liver cancer, fallopian tube tumors, ovarian tumors, peritoneal tumors, stage IV melanoma, glioma, glioblastoma, hepatocellular carcinoma, papillary nephroma, head and neck tumors, leukemia, lymphoma, myeloma, and non-small cell lung cancer.

The present invention also relates to a compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the same, for use in preventing and/or treating diseases having pathological characteristics of the IDO-mediated tryptophan metabolic pathway. These diseases include viral infections such as AIDS, cellular infections such as Lyme disease and streptococcal infections, neurodegenerative disorders (e.g., Alzheimer's disease, Huntington's disease, and Huntington's disease), autoimmune diseases, depression, anxiety, cataracts, psychological disorders, AIDS, cancer (including T-cell leukemia and colon cancer), eye disease states (e.g., cataracts and age-related yellowing), and autoimmune diseases, wherein the cancer can be selected from breast cancer, cervical cancer, colon cancer, lung cancer, stomach cancer, rectal cancer, pancreatic cancer, brain cancer, skin cancer, oral cancer, prostate cancer, bone cancer, kidney cancer, ovarian cancer, bladder cancer, liver cancer, fallopian tube tumors, ovarian tumors, peritoneal tumors, stage IV melanoma, glioma, glioblastoma, hepatocellular carcinoma, papillary nephroma, head and neck tumors, leukemia, lymphoma, myeloma, and non-small cell lung cancer.

The present invention also relates to a method for treating and/or preventing a disease having pathological characteristics of an IDO-mediated tryptophan metabolic pathway, comprising administering to a patient a therapeutically effective dose of a compound represented by general formula (I) or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the same. These diseases include viral infections such as AIDS, cellular infections such as Lyme disease and streptococcal infections, neurodegenerative disorders (e.g., Alzheimer's disease, Huntington's disease, and Huntington's disease), autoimmune diseases, depression, anxiety, cataracts, psychological disorders, AIDS, cancer (including T-cell leukemia and colon cancer), eye disease states (e.g., cataracts and age-related yellowing), and autoimmune diseases, wherein the cancer can be selected from breast cancer, cervical cancer, colon cancer, lung cancer, stomach cancer, rectal cancer, pancreatic cancer, brain cancer, skin cancer, oral cancer, prostate cancer, bone cancer, kidney cancer, ovarian cancer, bladder cancer, liver cancer, fallopian tube tumors, ovarian tumors, peritoneal tumors, stage IV melanoma, glioma, glioblastoma, hepatocellular carcinoma, papillary nephroma, head and neck tumors, leukemia, lymphoma, myeloma, and non-small cell lung cancer.

Another aspect of the present invention relates to a method for treating cancer, comprising administering to a patient a therapeutically effective dose of a compound of formula (I) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereoisomer, or mixture thereof, or a pharmaceutically acceptable salt thereof. The method shows outstanding efficacy and fewer side effects, wherein the cancer can be selected from breast cancer, cervical cancer, colon cancer, lung cancer, stomach cancer, rectal cancer, pancreatic cancer, brain cancer, skin cancer, oral cancer, prostate cancer, bone cancer, kidney cancer, ovarian cancer, bladder cancer, liver cancer, fallopian tube tumor, ovarian tumor, peritoneal tumor, stage IV melanoma, glioma, glioblastoma, hepatocellular carcinoma, papillary nephroma, head and neck tumor, leukemia, lymphoma, myeloma, and non-small cell lung cancer, preferably fallopian tube tumor, peritoneal tumor, stage IV melanoma, myeloma, and breast cancer, more preferably breast cancer.

Pharmaceutical compositions containing the active ingredient may be in a form suitable for oral administration, such as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs. Oral compositions may be prepared according to any method known in the art for preparing pharmaceutical compositions, and such compositions may contain one or more ingredients selected from the group consisting of sweeteners, flavoring agents, colorants, and preservatives to provide a pleasing and palatable pharmaceutical formulation. Tablets contain the active ingredient and, in admixture, nontoxic, pharmaceutically acceptable excipients suitable for tablet preparation. These excipients may be inert excipients, granulating agents, disintegrants, and lubricants. These tablets may be uncoated or coated using known techniques that mask the taste of the drug or delay disintegration and absorption in the gastrointestinal tract, thereby providing a sustained-release effect over a longer period of time.

Oral preparations may also be provided in soft gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, or with a water-soluble carrier or an oily vehicle or olive oil.

Aqueous suspensions contain the active substance in admixture with excipients suitable for the preparation of aqueous suspensions. Such excipients are suspending agents, dispersing agents, or wetting agents. Aqueous suspensions may also contain one or more preservatives, such as ethylparaben or n-propylparaben, one or more coloring agents, one or more flavoring agents, and one or more sweetening agents.

Oil suspensions can be prepared by suspending the active ingredient in a vegetable oil or mineral oil. The oil suspension may contain a thickener. The sweeteners and flavorings mentioned above may be added to provide a palatable preparation. These compositions may be preserved by the addition of antioxidants.

Dispersible powders and granules suitable for preparing aqueous suspensions can be prepared by adding water to provide the active ingredient and a dispersant or wetting agent, a suspending agent, or one or more preservatives for mixing. Suitable dispersants or wetting agents and suspending agents are exemplified above. Other excipients such as sweeteners, flavorings, and coloring agents may also be added.

The pharmaceutical compositions of the present invention may also be in the form of oil-in-water emulsions. The oil phase may be a vegetable oil or mineral oil, or a mixture thereof. Suitable emulsifiers may be naturally occurring phospholipids or partial ester emulsions. They may also contain sweeteners, flavorings, preservatives, and antioxidants. The pharmaceutical compositions may be in the form of sterile injectable aqueous solutions. Acceptable solvents or solvents that may be used include water, Ringer's solution, and isotonic sodium chloride solution. Sterile injectable formulations may be sterile injectable oil-in-water microemulsions in which the active ingredient is dissolved in the oil phase. The injection or microemulsion may be administered into the patient's bloodstream via local, bulk injection.

The pharmaceutical composition can be in the form of a sterile injectable aqueous or oily suspension for intramuscular and subcutaneous administration. The suspension can be prepared using suitable dispersants or wetting agents and suspending agents as described above according to known techniques. Sterile injectable formulations can also be sterile injectable solutions or suspensions prepared in a parenterally acceptable nontoxic diluent or solvent. In addition, sterile fixed oils can be conveniently used as solvents or suspending media.

The compounds of this invention may be administered in the form of suppositories for rectal administration. These pharmaceutical compositions can be prepared by mixing the drug with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid in the rectum and will therefore melt in the rectum to release the drug.

As is well known to those skilled in the art, the dosage of a drug depends on a variety of factors, including but not limited to the following: the activity of the specific compound used, the patient's age, the patient's weight, the patient's health condition, the patient's behavior, the patient's diet, the administration time, the administration method, the excretion rate, the combination of drugs, etc.; in addition, the optimal treatment method such as the treatment mode, the daily dosage of the compound of formula (I) or the type of pharmaceutically acceptable salt can be verified according to traditional treatment plans.

Detailed Description of the Invention

Unless otherwise stated, the terms used in the specification and claims have the following meanings.

The term "alkyl" refers to a saturated aliphatic hydrocarbon group, which is a straight or branched chain group containing 1 to 20 carbon atoms, preferably an alkyl group containing 1 to 12 carbon atoms, more preferably an alkyl group containing 1 to 6 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 2, 3-Dimethylpentyl, 2,4-dimethylpentyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 2-ethylpentyl, 3-ethylpentyl, n-octyl, 2,3-dimethylhexyl, 2,4-dimethylhexyl, 2,5-dimethylhexyl, 2,2-dimethylhexyl, 3,3-dimethylhexyl, 4,4-dimethylhexyl, 2-ethylhexyl, 3-ethylhexyl, 4-ethylhexyl, 2-methyl-2-ethylpentyl, 2-methyl-3-ethylpentyl, n-nonyl, 2-methyl-2-ethylhexyl, 2-methyl-3-ethylhexyl, 2,2-diethylpentyl, n-decyl, 3,3-diethylhexyl, 2,2-diethylhexyl, and various branched chain isomers thereof. More preferred are lower alkyl groups containing 1 to 6 carbon atoms, non-limiting examples of which include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, and the like. The alkyl group may be substituted or unsubstituted. When substituted, the substituent may be substituted at any available point of attachment. The substituent is preferably one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl or carboxylate.

The term "alkylene" refers to an alkyl group in which one hydrogen atom is further substituted, for example: "methylene" refers to _-CH2- , "ethylene" refers to -( _CH2 ) _2- , "propylene" refers to -( _CH2 ) _3- , "butylene" refers to -( _CH2 ) _4- , etc. The term "alkenyl" refers to an alkyl group as defined above consisting of at least two carbon atoms and at least one carbon-carbon double bond, for example, ethenyl, 1-propenyl, 2-propenyl, 1-, 2- or 3-butenyl, etc. The alkenyl group may be substituted or unsubstituted, and when substituted, the substituent is preferably one or more of the following groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio.

The term "cycloalkyl" refers to a saturated or partially unsaturated monocyclic or polycyclic hydrocarbon substituent, wherein the cycloalkyl ring contains 3 to 20 carbon atoms, preferably 3 to 12 carbon atoms, more preferably 3 to 8 carbon atoms, and most preferably 3 to 6 carbon atoms. Non-limiting examples of monocyclic cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl, and the like; polycyclic cycloalkyls include spirocyclic, fused, and bridged cycloalkyls.

The term "spiroalkyl" refers to a polycyclic group having a carbon atom (called a spiro atom) shared between 5 to 20 monocyclic rings, which may contain one or more double bonds, but no ring has a completely conjugated π electron system. Preferably, it is 6 to 14 yuan, more preferably 7 to 10 yuan. According to the number of spiro atoms shared between the rings, the spiroalkyl group is divided into a single spiroalkyl group, a double spiroalkyl group or a multi-spiroalkyl group, preferably a single spiroalkyl group and a double spiroalkyl group. More preferably, it is a 4 yuan/4 yuan, 4 yuan/5 yuan, 4 yuan/6 yuan, 5 yuan/5 yuan or 5 yuan/6 yuan single spiroalkyl group. Non-limiting examples of spiroalkyl groups include:

The term "fused cycloalkyl" refers to a 5 to 20-membered, all-carbon polycyclic group in which each ring in the system shares a pair of adjacent carbon atoms with the other rings in the system, wherein one or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron system. Preferably, it is 6 to 14 members, more preferably 7 to 10 members. Depending on the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic fused cycloalkyl groups, preferably bicyclic or tricyclic, more preferably 5-membered/5-membered or 5-membered/6-membered bicyclic alkyl groups. Non-limiting examples of fused cycloalkyl groups include:

The term "bridged cycloalkyl" refers to a 5-20 membered, all-carbon polycyclic group in which any two rings share two carbon atoms that are not directly connected, which may contain one or more double bonds, but no ring has a completely conjugated π electron system. Preferably, it is 6-14 members, more preferably 7-10 members. Depending on the number of constituent rings, it can be classified as a bicyclic, tricyclic, tetracyclic or polycyclic bridged cycloalkyl group, preferably a bicyclic, tricyclic or tetracyclic group, more preferably a bicyclic or tricyclic group. Non-limiting examples of bridged cycloalkyl groups include:

The cycloalkyl ring may be fused to an aryl, heteroaryl or heterocycloalkyl ring, wherein the ring attached to the parent structure is a cycloalkyl, non-limiting examples of which include indanyl, tetrahydronaphthyl, benzocycloheptanyl, etc. The cycloalkyl group 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, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl or carboxylate.

The term "heterocyclyl" refers to a saturated or partially unsaturated monocyclic or polycyclic hydrocarbon substituent containing 3 to 20 ring atoms, one or more of which is a heteroatom selected from nitrogen, oxygen, or S(O) _m (wherein m is an integer from 0 to 2), but excluding the ring moieties of -OO-, -OS-, or -SS-, and the remaining ring atoms are carbon. Preferably, it contains 3 to 12 ring atoms, of which 1 to 4 are heteroatoms; more preferably, it contains 3 to 6 ring atoms. Non-limiting examples of monocyclic heterocyclyls include pyrrolidinyl, imidazolidinyl, tetrahydrofuranyl, tetrahydrothienyl, dihydroimidazolyl, dihydrofuranyl, dihydropyrazolyl, dihydropyrrolyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, homopiperazinyl, and the like, preferably piperidinyl and pyrrolidinyl. Polycyclic heterocyclyls include spirocyclic, fused, and bridged heterocyclyls.

The term "spiroheterocyclic radical" refers to a polycyclic heterocyclic group having one atom (called spiral atom) shared between 5 to 20 rings, wherein one or more ring atoms are heteroatoms selected from nitrogen, oxygen or S(O) _m (wherein m is an integer from 0 to 2), and the remaining ring atoms are carbon. It may contain one or more double bonds, but no ring has a completely conjugated π electron system. It is preferably 6 to 14 members, more preferably 7 to 10 members. Spiroheterocyclic radicals are divided into single spiral heterocyclic radicals, double spiral heterocyclic radicals or multiple spiral heterocyclic radicals according to the number of shared spiral atoms between rings, preferably single spiral heterocyclic radicals and double spiral heterocyclic radicals. It is more preferably 4/4 members, 4/5 members, 4/6 members, 5/5 members or 5/6 members single spiral heterocyclic radicals. Non-limiting examples of spiroheterocyclic radicals include:

The term "fused heterocyclic group" refers to a polycyclic heterocyclic group of 5 to 20 members, each ring in the system sharing a pair of atoms adjacent to the other rings in the system, one or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron system, wherein one or more ring atoms are heteroatoms selected from nitrogen, oxygen or S(O) _m (wherein m is an integer from 0 to 2), and the remaining ring atoms are carbon. Preferably, it is 6 to 14 members, more preferably 7 to 10 members. According to the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic fused heterocyclic groups, preferably bicyclic or tricyclic, more preferably 5-membered/5-membered or 5-membered/6-membered bicyclic fused heterocyclic groups. Non-limiting examples of fused heterocyclic groups include:

The term "bridged heterocyclic group" refers to a polycyclic heterocyclic group of 5 to 14 members, wherein any two rings share two atoms that are not directly connected, which may contain one or more double bonds, but no ring has a completely conjugated π electron system, wherein one or more ring atoms are heteroatoms selected from nitrogen, oxygen or S(O) _m (wherein m is an integer from 0 to 2), and the remaining ring atoms are carbon. Preferably, it is 6 to 14 members, more preferably 7 to 10 members. According to the number of constituent rings, it can be divided into bicyclic, tricyclic, tetracyclic or polycyclic bridged heterocyclic groups, preferably bicyclic, tricyclic or tetracyclic, more preferably bicyclic or tricyclic. Non-limiting examples of bridged heterocyclic groups include:

The heterocyclyl ring may be fused to an aryl, heteroaryl or cycloalkyl ring, wherein the ring attached to the parent structure is a heterocyclyl, non-limiting examples of which include:

wait.

The heterocyclyl group may be optionally substituted or unsubstituted. When substituted, the substituents are preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl or carboxylate.

The term "aryl" refers to a 6- to 14-membered all-carbon monocyclic or fused polycyclic (i.e., rings sharing adjacent pairs of carbon atoms) group having a conjugated π electron system, preferably 6- to 10-membered, such as phenyl and naphthyl. More preferably, phenyl. The aryl ring may be fused to a heteroaryl, heterocyclyl, or cycloalkyl ring, wherein the ring attached to the parent structure is the aryl ring, non-limiting examples of which include:

Preferred

The aryl group may be substituted or unsubstituted. When substituted, the substituents are preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, carboxyl or carboxylate.

The term "heteroaryl" refers to a heteroaromatic system containing 1 to 4 heteroatoms and 5 to 14 ring atoms, wherein the heteroatoms are selected from oxygen, sulfur and nitrogen. The heteroaryl group is preferably 5 to 10-membered, more preferably 5-membered or 6-membered, such as imidazolyl, furyl, thienyl, thiazolyl, pyrazolyl, oxazolyl, pyrrolyl, tetrazolyl, pyridinyl, pyrimidinyl, thiadiazole, pyrazinyl, etc., preferably imidazolyl, pyrazolyl or pyrimidinyl, thiazolyl; more preferably pyrazolyl. The heteroaryl ring can be fused to an aryl, heterocyclyl or cycloalkyl ring, wherein the ring connected to the parent structure is a heteroaryl ring, non-limiting examples of which include:

Preferred

The heteroaryl group may be optionally substituted or unsubstituted, and when substituted, the substituents are preferably one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, carboxyl or carboxylate.

The term "alkoxy" refers to-O-(alkyl) and-O-(unsubstituted cycloalkyl), wherein the definition of alkyl is as described above. The limiting examples of alkoxy include: methoxy, ethoxy, propoxy, butoxy, cyclopropyloxy, cyclobutyloxy, cyclopentyloxy, cyclohexyloxy. Alkoxy can be optionally substituted or unsubstituted, and when substituted, substituents are preferably one or more following groups, which are independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, sulfydryl, hydroxyl, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkyloxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio, carboxyl or carboxylate.

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

The term "haloalkoxy" refers to an alkoxy group substituted with one or more halogens, wherein alkoxy is as defined above.

The term "hydroxyalkyl" refers to an alkyl group substituted with a hydroxy group, wherein alkyl is as defined above.

The term "hydroxy" refers to an -OH group.

The term "halogen" refers to fluorine, chlorine, bromine or iodine.

The term "amino" refers to _-NH2 .

The term "cyano" refers to -CN.

The term "nitro" refers to _-NO2 .

The term "oxo" refers to =0.

The term "carboxy" refers to -C(O)OH.

The term "carboxylate" refers to -C(O)O(alkyl) or -C(O)O(cycloalkyl), wherein alkyl is as defined above.

The term "acyl halide" refers to a compound containing the group -C(O)-halogen.

Different expressions such as “X is selected from A, B, or C”, “X is selected from A, B and C”, “X is A, B or C”, and “X is A, B and C” all express the same meaning, that is, X can be any one or more of A, B, and C.

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

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

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

"Pharmaceutically acceptable salts" refer to salts of the compounds of the present invention that are safe and effective when used in mammals and have the desired biological activity.

Synthesis method of the compound of the present invention

In order to achieve the purpose of the present invention, the present invention adopts the following technical solutions:

The method for preparing the compound represented by the general formula (II) and the general formula (II-1) of the present invention or its tautomer, mesomer, racemate, enantiomer, diastereomer or mixture thereof, or its pharmaceutically acceptable salt comprises the following steps:

Option 1

Under low temperature alkaline conditions (the reagent providing alkaline conditions is preferably lithium diisopropylamide), the compound of general formula (II-a) reacts with a bromobenzene compound to obtain a compound of general formula (II-b); the compound of general formula (II-b) reacts with a methanesulfonyl chloride compound under alkaline conditions (the reagent providing alkaline conditions is preferably sodium hydride or triethylamine) to obtain a compound of general formula (II-c); the obtained compound of general formula (II-c) reacts with imidazole under alkaline conditions to obtain a compound of general formula (II-d); the obtained compound of general formula (II-d) is subjected to intramolecular coupling in the presence of heating, a base and a phosphine palladium catalyst (the catalyst is preferably triphenylphosphine and palladium acetate) to obtain a compound of general formula (II-e); the obtained compound of general formula (II-e) is deprotected under acidic conditions to obtain a compound of general formula (V) or its salt; the compound of general formula (V) is further reacted with R in the presence of alkaline conditions (the reagent providing alkaline conditions is preferably sodium tert-butoxide) and a phosphine palladium catalyst. The halide of ^{R 3} undergoes coupling reaction, and the obtained product is optionally further coupled with the boronic acid or boric acid ester of R ³ to obtain a compound of general formula (II-a); the compound of general formula (II-a) is salified under acidic conditions to obtain a compound of general formula (II-b).

Reagents providing alkaline conditions include organic bases and inorganic bases, wherein the organic bases include but are not limited to triethylamine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, potassium acetate, sodium tert-butoxide or potassium tert-butoxide, and the inorganic bases include but are not limited to sodium hydride, potassium phosphate, sodium carbonate, potassium carbonate or cesium carbonate.

The phosphine palladium catalysts involved include but are not limited to 2-dicyclohexylphosphine-2,4,6-triisopropylbiphenyl, (±)-2,2'-bis-(diphenylphosphine)-1,1'-binaphthyl, tris(dibenzylideneacetone)dipalladium, palladium acetate, [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium, triphenylphosphine, and tetrakis(triphenylphosphine)palladium.

in:

R ¹ to R ³ , M, p, n, a, b and y are as defined in the general formula (II); Q and x are as defined in the general formula (V).

Option 2

The compound of the general formula (II-f) reacts with p-toluenesulfonyl hydrazide to obtain a compound of the general formula (II-g); the obtained compound of the general formula (II-g) reacts with a bromobenzaldehyde compound under alkaline conditions to obtain a compound of the general formula (II-h); the obtained compound of the general formula (II-h) is reduced in the presence of a reducing agent (the reducing agent is preferably sodium borohydride) to obtain a compound of the general formula (II-b); the compound of the general formula (II-b) reacts with a sulfonyl chloride compound under alkaline conditions (the reagent providing alkaline conditions is preferably sodium hydride or triethylamine) to obtain a compound of the general formula (II-c ) compound; the obtained compound of formula (II-c) is reacted with imidazole under alkaline conditions to obtain a compound of formula (II-d); the obtained compound of formula (II-d) is subjected to intramolecular coupling in the presence of heating, alkali and a phosphine palladium catalyst (the catalyst is preferably triphenylphosphine and palladium acetate) to obtain a compound of formula (II-e); the obtained compound of formula (II-e) is deprotected under acidic conditions to obtain a compound of formula (V) or a salt thereof; the compound of formula (V) is further subjected to coupling reaction with a halide of R ³ in the presence of alkaline conditions (the reagent providing alkaline conditions is preferably sodium tert-butoxide) and a phosphine palladium catalyst, and the obtained product is optionally further coupled with a boronic acid or boric ester of R ³ to obtain a compound of formula (II-a); the compound of formula (II-a) is salified under acidic conditions to obtain a compound of formula (II-b).

Reagents providing alkaline conditions include organic bases and inorganic bases, wherein the organic bases include but are not limited to triethylamine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, potassium acetate, sodium tert-butoxide or potassium tert-butoxide, and the inorganic bases include but are not limited to sodium hydride, potassium phosphate, sodium carbonate, potassium carbonate or cesium carbonate.

The phosphine palladium catalysts involved include but are not limited to 2-dicyclohexylphosphine-2,4,6-triisopropylbiphenyl, (±)-2,2'-bis-(diphenylphosphine)-1,1'-binaphthyl, tris(dibenzylideneacetone)dipalladium, palladium acetate, [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium, triphenylphosphine, and tetrakis(triphenylphosphine)palladium.

The reducing agent includes but is not limited to Fe powder, Zn powder, H ₂ , sodium borohydride, sodium triacetoxyborohydride, sodium cyanoborohydride or lithium aluminum hydride.

in:

R ¹ to R ³ , M, p, n, a, b and y are as defined in the general formula (II); Q and x are as defined in the general formula (V).

Option 3

There are several specific synthesis methods for synthesizing the compound of formula (II-1) from the compound of formula (V), depending on the different compounds:

Method 1: Under the protection of inert gas, high temperature, alkalinity, and phosphine palladium catalyst conditions, the compound of general formula (V) and the halide of R ¹ are directly heated or coupled in a microwave reactor, and the obtained product is optionally further coupled with the boronic acid or boric ester of R ¹ to obtain the compound of general formula (II-1); the halide of R ³ in this reaction is preferably an aromatic halogenated compound; the alkaline reagent is preferably sodium tert-butoxide.

Method 2: Under high temperature and alkaline conditions, the compound of general formula (V) directly undergoes a coupling reaction with the halide of ^R1 , and the obtained product is optionally further coupled with the boronic acid or boric ester of R3 to obtain the compound of general formula (II-1); the halide of ^R1 in this reaction is preferably a heteroaryl halogenated compound; the alkaline reagent is preferably triethylamine.

Method 3: At room temperature and alkaline conditions, the compound of general formula (V) directly reacts with a halide of R ³ to obtain a compound of general formula (II-1); the halide in this reaction is preferably an iodide and a highly active acyl halide compound; the alkaline reagent under these conditions is preferably potassium carbonate.

The phosphine palladium catalysts involved include but are not limited to 2-dicyclohexylphosphine-2,4,6-triisopropylbiphenyl, (±)-2,2'-bis-(diphenylphosphine)-1,1'-binaphthyl, tris(dibenzylideneacetone)dipalladium, palladium acetate, [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium, triphenylphosphine, and tetrakis(triphenylphosphine)palladium.

The reagents providing alkaline conditions include organic bases and inorganic bases. The organic bases include but are not limited to sodium tert-butoxide, triethylamine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, sodium tert-butoxide, potassium acetate or potassium tert-butoxide, preferably sodium tert-butoxide; the inorganic bases include but are not limited to sodium hydride, potassium phosphate, sodium carbonate, potassium carbonate or cesium carbonate.

in:

X is N;

R ¹ to R ³ , p, n, a, and b are as defined in the general formula (II); Q and x are as defined in the general formula (V).

DETAILED DESCRIPTION

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

Example

The structures of the compounds were determined by nuclear magnetic resonance (NMR) or mass spectrometry (MS). NMR measurements were performed using a Bruker AVANCE-400 NMR spectrometer. The solvents used were deuterated dimethyl sulfoxide (DMSO-d ₆ ), deuterated chloroform (CDCl ₃ ), and deuterated methanol (CD ₃ OD). The internal standard was tetramethylsilane (TMS). Chemical shifts are reported in units of 10 ^-6 (ppm).

MS was measured using a FINNIGAN LCQAd (ESI) mass spectrometer (manufacturer: Thermo, model: Finnigan LCQadvantage MAX).

HPLC analysis 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).

The average kinase inhibition rate and _IC50 value were determined using a NovoStar microplate reader (BMG, Germany).

The thin layer chromatography silica gel plate uses Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plate. The silica gel plate used in thin layer chromatography (TLC) adopts a specification of 0.15mm-0.2mm. The specification used for thin layer chromatography separation and purification of products is 0.4mm-0.5mm silica gel plate.

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

The known starting materials of the present invention can be synthesized by methods known in the art, or can be purchased from ABCR GmbH & Co. KG, Acros Organnics, Aldrich Chemical Company, AccelaChemBio Inc, Darui Chemicals, and other companies.

Unless otherwise specified in the examples, all reactions were carried out under an argon or nitrogen atmosphere.

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

Hydrogen atmosphere means that the reaction bottle is connected to a hydrogen balloon with a capacity of about 1L.

The pressurized hydrogenation reaction uses a Parr 3916EKX hydrogenator and a Qinglan QL-500 hydrogen generator or an HC2-SS hydrogenator.

The hydrogenation reaction is usually carried out by evacuating the chamber and filling it with hydrogen, and the operation is repeated three times.

A CEM Discover-S 908860 microwave reactor was used for the microwave reaction.

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

Unless otherwise specified in the examples, the reaction temperature is room temperature.

Room temperature is the most suitable reaction temperature, and the temperature range is 20℃～30℃.

The reaction progress in the examples was monitored by thin layer chromatography (TLC). The developing solvent systems used in the reactions were: A: dichloromethane and methanol system, B: n-hexane and ethyl acetate system, C: petroleum ether and ethyl acetate system, and D: acetone. The volume ratio of the solvents was adjusted according to the polarity of the compounds.

The eluent system for column chromatography and the developing solvent system for thin-layer chromatography used for purifying compounds include: A: dichloromethane and methanol system, B: n-hexane and ethyl acetate system, C: n-hexane, ethyl acetate and dichloromethane system, D: petroleum ether and ethyl acetate system, and E: ethyl acetate. The volume ratio of the solvent is adjusted according to the polarity of the compound, and can also be adjusted by adding a small amount of triethylamine and acidic or alkaline reagents.

Example 1

6-Fluoro-5-(1-phenylpiperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

tert-Butyl 4-((2-bromo-6-fluorophenyl)(hydroxy)methyl)piperidine-1-carboxylate 1c

Lithium diisopropylamide (32.5 mL, 65.0 mmol) was added to 50 mL of tetrahydrofuran. A pre-prepared solution of 1-bromo-3-fluorobenzene 1a (8.75 g, 50.0 mmol) in tetrahydrofuran (25 mL) was added dropwise at -78°C, and the mixture was stirred at -78°C for 1 hour. A pre-prepared solution of tert-butyl 4-formylpiperidine-1-carboxylate 1b (8.75 g, 50.0 mmol) in tetrahydrofuran (25 mL) was then added dropwise at -78°C, and the mixture was stirred at -78°C for 1 hour. After completion of the reaction, 25 mL of methanol was added dropwise at -78°C to quench the reaction. The reaction solution was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography using eluent System B to afford compound 1c (16.3 g, yellow syrupy solid) in an 84.0% yield.

MS m/z (LC-MS): 332.0 [M-56]

Step 2

Tert-Butyl 4-((2-bromo-6-fluorophenyl)(p-toluenesulfonyloxy)methyl)piperidine-1-carboxylate 1d

Compound 1c (15 g, 38.63 mmol) was dissolved in 350 mL of tetrahydrofuran, and sodium hydride (3.09 g, 77.26 mmol) was added portionwise. The mixture was stirred until gas evolution ceased. 250 mL of a pre-prepared solution of p-toluenesulfonyl chloride (8.10 g, 42.49 mmol) in tetrahydrofuran was added dropwise. The mixture was stirred at room temperature for 30 minutes, refluxed for 4 hours, and stirred at 70°C for 48 hours. After the reaction was completed, the mixture was cooled to 0°C and quenched by the dropwise addition of 50 mL of water. 50 mL of saturated sodium chloride solution was added, and the layers were separated. The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System B to afford compound 1d (6.6 g, light yellow viscous solid) in a yield of 31.80%.

MS m/z(LC-MS): 314.0/316.0[M-56-TsO]

Step 3

tert-Butyl 4-((2-bromo-6-fluorophenyl)(1H-imidazol-1-yl)methyl)piperidine-1-carboxylate 1e

Imidazole (12.5 g, 184.3 mmol) was dissolved in 50 mL of N,N-dimethylformamide. Sodium hydride (7.40 g, 184.3 mmol) was added portionwise and stirred at room temperature for 1 hour. A 20 mL solution of preformed compound 1d (10.0 g, 18.43 mmol) in N,N-dimethylformamide was then added dropwise, and the mixture was stirred at 100°C for 12 hours. After the reaction, 300 mL of ethyl acetate was added, and the mixture was washed with saturated sodium chloride solution (150 mL x 3). The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A to afford compound 1e (1.90 g, brown viscous solid) in a yield of 23.5%.

MS m/z(LC-MS): 438.1/440.1[M+1]

Step 4

tert-Butyl 4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidine-1-carboxylate 1f

Compound 1e (1.90 g, 4.33 mmol), N,N-dicyclohexylmethylamine (1.35 g, 6.93 mmol), and triphenylphosphine (908 mg, 3.46 mmol) were added to a 10 mL N,N-dimethylformamide solution. Palladium acetate (390 mg, 1.74 mmol) was added under an argon atmosphere, and the mixture was stirred at 100°C for 4.5 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by silica gel column chromatography with eluent System B to obtain compound 1f (1.30 g, yellow viscous solid) in an 83.8% yield.

MS m/z (LC-MS): 358.1[M+1]

Step 5

6-Fluoro-5-(piperidin-4-yl)-5H-imidazo[5,1-a]isoindole ditrifluoroacetate 1g

Compound 1f (1.30 g, 3.64 mmol) was dissolved in 5 mL of dichloromethane, and 5 mL of trifluoroacetic acid was added dropwise. The mixture was stirred at room temperature for 1 hour. After the reaction, the reaction solution was concentrated under reduced pressure to obtain crude compound 1g (1.77 g, brown viscous solid). The product was directly used in the next reaction without purification.

MS m/z (LC-MS): 258.3[M+1]

Step 6

6-Fluoro-5-(1-phenylpiperidin-4-yl)-5H-imidazo[5,1-a]isoindole 1h

The crude product Compound 1g (230 mg, 0.50 mmol), bromobenzene (314 mg, 2.00 mmol), (±)-2,2′-bis-(diphenylphosphino)-1,1′-binaphthyl (39 mg, 0.0625 mmol), and sodium tert-butoxide (192 mg, 2.0 mmol) were dissolved in 5 mL of 1,4-dioxane. Tris(dibenzylideneacetone)dipalladium (46 mg, 0.05 mmol) was added under an argon atmosphere, and the mixture was stirred at 100°C for 4 hours. After completion of the reaction, 20 mL of ethyl acetate was added, and the mixture was washed with water (10 mL x 3). The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to obtain Compound 1h (6.3 mg, brown viscous solid) in a yield of 3.6%.

MS m/z(ESI):334.3[M+1]

Step 7

6-Fluoro-5-(1-phenylpiperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 1

Compound 1h (6.3 mg, 0.018 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.01 mL of trifluoroacetic acid was added. The mixture was stirred at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 1 (8.3 mg, light brown solid) with a yield of 100%.

MS m/z(ESI):334.3[M+1]

¹ H NMR (400MHz, CD ₃ OD) δ9.35 (s, 1H), 7.93 (s, 1H), 7.74 (d, 1H), 7.69-7.64 (m, 1H), 7.60-7.50 (m, 5H), 7.38-7.34 (m, 1H), 6.15 (d, 1H), 3.79-3.71(m, 1H), 3.69-3.61(m, 2H), 3.61-3.51(m, 1H), 3.02-2.92(m, 1H), 2.14-2.04(m, 2H), 1.76-1.59(m, 2H).

Example 2

7-Fluoro-5-(1-phenylpiperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

tert-Butyl 4-(2-p-toluenesulfonylhydrazono)piperidine-1-carboxylate 2c

Toluenesulfonylhydrazide 2a (9.31 g, 50.0 mmol) was added to 100 mL of methanol, and N-tert-butyloxycarbonylpiperidone 2b (9.96 g, 50.0 mmol, prepared by the known method "ACS Medicinal Chemistry Letters, 2014, 5(5), 550-555") was added. The reaction system was reacted at room temperature for 2 hours. After the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was dried to obtain crude compound 2c (18.37 g, white solid). The product was directly used in the next reaction without purification.

MS m/z (LC-MS): 368.0 [M+1]

Step 2

tert-Butyl 4-(2-bromo-5-fluorobenzoyl)piperidine-1-carboxylate 2e

Crude compound 2c (5.52 g, 15.0 mmol), 2-bromo-5-fluorobenzaldehyde 2d (3.05 g, 15.0 mmol), and cesium carbonate (7.33 g, 22.5 mmol) were placed in a sealed tube. Under an argon atmosphere, 60 mL of 1,4-dioxane was added, and the reaction system was stirred at 110°C for 10 hours. After the reaction, 20 mL of ethyl acetate was added, and the mixture was washed with water (100 mL × 1) and then with saturated sodium chloride solution (100 mL × 2). The mixture was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain crude compound 2e (6.17 g, a yellow viscous solid). The product was directly used in the next reaction without purification.

MS m/z(LC-MS): 330.0/332.0[M-56]

Step 3

tert-Butyl 4-((2-bromo-5-fluorophenyl)(hydroxy)methyl)piperidine-1-carboxylate 2f

Crude compound 2e (6.17 g, 16.0 mmol) was dissolved in 100 mL of methanol. The reaction system was cooled to 0°C, and sodium borohydride (1.21 g, 32.0 mmol) was slowly added at 0°C. The reaction system was stirred at 0°C for 1 hour. After the reaction, 200 mL of ethyl acetate was added, and the mixture was washed with saturated sodium chloride solution (100 mL × 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain the crude title product, compound 2f (6.21 g, yellow syrup). The product was directly used in the next reaction without purification.

MS m/z(LC-MS): 332.0/334.0[M-56]

Step 4

tert-Butyl 4-((2-bromo-5-fluorophenyl)((methylsulfonyl)oxy)methyl)piperidine-1-carboxylate 2g

Crude compound 2f (6.21 g, 16.0 mmol) was dissolved in 100 mL of dichloromethane and cooled to 0°C. Triethylamine (3.24 g, 32.0 mmol) was added, followed by the dropwise addition of methylsulfonyl chloride (2.75 g, 24.0 mmol). The reaction system was stirred at 0°C for 1 hour. After completion of the reaction, the reaction solution was washed with water (50 mL × 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain crude compound 2g (7.46 g, brown viscous solid). The product was directly used in the next reaction without purification.

Step 5

tert-Butyl 4-((2-bromo-5-fluorophenyl)(1H-imidazol-1-yl)methyl)piperidine-1-carboxylate 2h

The crude compound 2g (7.46 g, 16.0 mmol), 1-H-imidazole (10.89 g, 160.0 mmol), and N,N-diisopropylethylamine (20.68 g, 160.0 mmol) were added to a sealed tube, followed by 50 mL of acetonitrile, and the reaction system was stirred at 120°C for 12 hours. After completion of the reaction, the reaction was cooled to room temperature, and 200 mL of ethyl acetate was added. The mixture was washed with water (100 mL × 3), then with saturated sodium chloride solution (100 mL × 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with eluent System A to obtain compound 2h (4.68 g, brown-yellow foamy solid) in a yield of 66.7%.

MS m/z(LC-MS): 438.1/440.1[M+1]

Step 6

tert-Butyl 4-(7-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidine-1-carboxylate 2i

Compound 2h (4.68 g, 10.68 mmol), N,N-dicyclohexylmethylamine (3.34 g, 17.08 mmol), and triphenylphosphine (2.24 g, 8.54 mmol) were added to a 50 mL N,N-dimethylformamide solution. Palladium acetate (960 mg, 4.28 mmol) was added under an argon atmosphere, and the reaction system was allowed to react at 100°C for 17 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by silica gel column chromatography with eluent System A to obtain compound 2i (3.81 g, yellow solid) in a 99.0% yield.

MS m/z (LC-MS): 358.1[M+1]

Step 7

7-Fluoro-5-(piperidin-4-yl)-5H-imidazo[5,1-a]isoindole ditrifluoroacetate 2j

Compound 2i (3.81 g, 10.68 mmol) was dissolved in 10 mL of dichloromethane, and 10 mL of trifluoroacetic acid was added dropwise. The mixture was reacted at room temperature for 2 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain crude compound 2j (5.18 g, brown viscous solid), which was directly used in the next reaction without purification.

MS m/z (LC-MS): 258.3[M+1]

Step 8

7-Fluoro-5-(1-phenylpiperidin-4-yl)-5H-imidazo[5,1-a]isoindole 2k

Crude compound 2j (115 mg, 0.25 mmol), bromobenzene (47 mg, 0.30 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (6 mg, 0.025 mmol), sodium tert-butoxide (96 mg, 1.0 mmol), and palladium acetate (6.0 mg, 0.025 mmol) were added to a mixed solvent of toluene and tert-butanol (V:V = 5:1) (1.8 mL). The reaction system was microwaved at 120°C for 1 hour. After completion of the reaction, 50 mL of dichloromethane was added, the mixture was washed with saturated sodium sulfite solution (25 mL x 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to afford compound 2k (12.6 mg, brown syrup) in a yield of 15.2%.

MS m/z(ESI):334.3[M+1]

Step 9

7-Fluoro-5-(1-phenylpiperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 2

Compound 2k (12.6 mg, 0.038 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.01 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 2 (16.9 mg, light brown solid) with a yield of 100%.

MS m/z(ESI):334.3[M+1]

Example 3

6-Fluoro-5-(1-(3-fluorophenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

6-Fluoro-5-(1-(3-fluorophenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 3a

The crude compound 1g (230 mg, 0.5 mmol), 1-bromo-3-fluorobenzene (105 mg, 0.6 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (24 mg, 0.05 mmol), and sodium tert-butoxide (192 mg, 2.0 mmol) were added to a 3 mL mixture of toluene and tert-butanol (V:V = 5:1). Under an argon atmosphere, palladium acetate (12.0 mg, 0.05 mmol) was added, and the reaction system was reacted at 120°C for 12 hours. After completion of the reaction, 20 mL of ethyl acetate was added, the mixture was washed with water (10 mL x 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to obtain compound 3a (17.3 mg, black syrup) in a 10% yield.

MS m/z(ESI):352.3[M+1]

Step 2

6-Fluoro-5-(1-(3-fluorophenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 3

Compound 3a (17.3 mg, 0.05 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.01 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 3 (23 mg, light brown solid) with a yield of 100%.

MS m/z(ESI):352.3[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.51(s, 1H), 8.06(s, 1H), 7.76(d, 1H), 7.67-7.62(m, 1H), 7.47-7.32(m, 1H), 7.23-7.10(m, 1H), 6.76-6.62(m, 2H), 6.57-6.45(m, 1H), 6.12(d, 1H), 3 .85-3.73(m, 1H), 3.73-3.63(m, 1H), 2.79-2.67(m, 1H), 2.67-2.50(m, 2H), 1.83-1.68(m, 1H), 1.61-1.45(m, 1H), 1.35-1.23(m, 1H), 1.10-0.94(m, 1H).

Example 4

6-Fluoro-5-(1-(4-methoxyphenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

6-Fluoro-5-(1-(4-methoxyphenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 4a

The crude product, compound 1g (115 mg, 0.25 mmol), 4-iodoanisole (70 mg, 0.3 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (12 mg, 0.025 mmol), and sodium tert-butoxide (96 mg, 1.0 mmol) were added to 1.8 mL of a mixed solvent of toluene and tert-butanol (V:V = 5:1). Under an argon atmosphere, palladium acetate (6.0 mg, 0.025 mmol) was added, and the reaction system was microwaved at 160°C for 30 minutes. After completion of the reaction, the mixture was washed with saturated sodium sulfite solution (25 mL × 3) and extracted with dichloromethane (20 mL × 3). The organic phases were combined, washed with saturated sodium sulfite solution (25 mL × 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to obtain the title product, compound 4a (29 mg, brown syrup), in a yield of 33.6%.

MS m/z(ESI):364.3[M+1]

Step 2

6-Fluoro-5-(1-(4-methoxyphenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 4

Compound 4a (29 mg, 0.084 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 4 (40 mg, light brown solid) with a yield of 100%.

MS m/z(ESI):364.3[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.41(s, 1H), 8.02(s, 1H), 7.75(d, 1H), 7.68-7.63(m, 1H), 7.44-7.39(m , 1H), 7.21-7.10(m, 2H), 6.97-6.87(m, 2H), 6.14(d, 1H), 3.71(s, 3H), 3.5 9-3.50(m, 1H), 3.49-3.40(m, 1H), 3.18-2.82(m, 2H), 2.69-2.55(m, 2H), 1 .95-1.83(m, 1H), 1.83-1.66(m, 1H), 1.42-1.30(m, 1H), 1.26-1.10(m, 1H).

Example 5

4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)benzonitrile trifluoroacetate

first step

4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)benzonitrile 5a

The crude product 1g (230 mg, 0.5 mmol), p-iodobenzonitrile (137 mg, 0.6 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (24 mg, 0.05 mmol), sodium tert-butoxide (192 mg, 2.0 mmol), and palladium acetate (12.0 mg, 0.05 mmol) were added to a mixed solvent of toluene and tert-butanol (V:V = 5:1) (3 mL). The reaction system was microwave-treated at 120°C under an argon atmosphere for 1 hour. After completion of the reaction, 20 mL of dichloromethane was added, and the product was washed with saturated sodium sulfite solution (10 mL x 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to obtain compound 5a (67 mg, brown syrup) in a yield of 29.7%.

MS m/z(ESI):359.1[M+1]

Step 2

4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)benzonitrile trifluoroacetate 5

Compound 5a (67 mg, 0.188 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.1 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 5 (70 mg, light brown solid) with a yield of 100%.

MS m/z(ESI):359.1[M+1]

Example 6

4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)benzamide trifluoroacetate

first step

4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)benzamide 6a

Compound 5 (60 mg, 0.127 mmol), zinc powder (340 mg, 5.2 mmol), 3 mL of acetic acid, and 0.2 mL of concentrated hydrochloric acid were added to a reaction flask and reacted at 125°C for 19 hours. After completion of the reaction, the zinc powder was filtered off, the filtrate was concentrated under reduced pressure, and the resulting residue was purified by HPLC to obtain compound 6a (4.5 mg, brown syrup) in a yield of 9.4%.

MS m/z(ESI):377.4[M+1]

Step 2

4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)benzamide trifluoroacetate 6

Compound 6a (4.5 mg, 0.012 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.1 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 6 (5.8 mg, light brown solid) with a yield of 100%.

MS m/z(ESI):377.4[M+1]

Example 7

6-Fluoro-5-(1-(4-(methylsulfonyl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

6-Fluoro-5-(1-(3-(methylsulfonyl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 7b

Crude compound 1g (150 mg, 0.33 mmol), 1-bromo-3-(methylsulfonyl)benzene 7a (93 mg, 0.40 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (16 mg, 0.033 mmol), sodium tert-butoxide (158 mg, 1.65 mmol), and palladium acetate (20.0 mg, 0.083 mmol) were added to 3 mL of a mixed solvent of toluene and tert-butanol (V:V = 5:1). The reaction system was microwave-treated at 160°C for 30 minutes. After completion of the reaction, the product was filtered using celite, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to afford compound 7b (14.5 mg, light orange solid) in a yield of 13.0%.

MS m/z(ESI):412.0[M+1]

Step 2

6-Fluoro-5-(1-(3-(methylsulfonyl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 7

Compound 7b (14.5 mg, 0.035 mmol) was dissolved in 5 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 12 hours. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 7 (18 mg, light orange solid) with a yield of 100%.

MS m/z(ESI):412.0[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ9.05(br.s, 1H), 7.60-7.56(m, 3H), 7.45-7.38(m, 3H), 7.26-7.22(t, 1H), 7.14-7.12(d, 1H), 5.7 5(s, 1H), 3.06(s, 3H), 2.91-2.75(m, 2H), 1.94-1.74(m, 2H), 1.49-1.46(m, 1H), 1.37-1.19(m, 4H).

Example 8

6-Fluoro-5-(1-(3-(1-methyl-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole

The crude compound 1g (229 mg, 0.5 mmol) and 4-(3-bromophenyl)-1-methyl-1H-pyrazole 8a (236 mg, 1 mmol, prepared using the method disclosed in patent application "WO2013043946") were dissolved in 10 mL of toluene. (±)-2,2′-bis-(diphenylphosphino)-1,1′-binaphthyl (311 mg, 0.05 mmol), sodium tert-butoxide (192 mg, 2 mmol), and tris(dibenzylideneacetone)dipalladium (45.78 mg, 0.05 mmol) were added, and the reaction system was microwaved at 120°C for 1 hour. After completion of the reaction, the filtrate was concentrated under reduced pressure, and the resulting residue was purified by high-performance liquid chromatography to obtain compound 8 (30 mg, white solid) in a yield of 11.3%.

MS m/z(ESI):414.2[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ8.18-8.16(m, 2H), 7.99-7.93(m, 1H), 7.90-7.85(m, 2H), 7.74-7.72(m, 1H), 7.69-7.65(m, 4H), 7.36-7.32(m, 1H), 6.66(s, 1H), 3.96 (s, 3H), 3.36 (m, 1H), 3.15 (m, 1H), 2.78 (m, 1H), 2.66 (m, 1H), 2.40 (m, 1H), 2.03 (m, 1H), 1.78 (m, 1H), 1.28 (m, 1H), 1.01 (m, 1H).

Example 9

6-Fluoro-5-(1-(4-(1-methyl-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole

first step

5-(1-(4-bromophenyl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole 9a

1 g (1.45 g, 3 mmol) of the crude compound was dissolved in 30 mL of toluene, and p-dibromobenzene (1.41 g, 6 mmol) was added. (±)-2,2′-bis-(diphenylphosphino)-1,1′-binaphthyl (187 mg, 0.3 mmol), sodium tert-butoxide (1.15 mg, 12 mmol), and tris(dibenzylideneacetone)dipalladium (275 mg, 0.3 mmol) were also added. The reaction system was allowed to react at 80°C under an argon atmosphere for 12 hours. After completion of the reaction, the filtrate was concentrated under reduced pressure, and the resulting residue was purified by silica gel column chromatography using eluent System A to obtain compound 9a (670 mg, yellow solid) in a 50% yield.

MS m/z (LC-MS): 412.2[M+1]

Step 2

6-Fluoro-5-(1-(4-(1-methyl-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 9

Crude compound 9a (165 mg, 0.4 mmol) was dissolved in 5 mL of ethylene glycol dimethyl ether. 1-Methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (166 mg, 0.8 mmol), sodium carbonate (127 mg, 1.2 mmol), and 0.5 mL of water were added. After stirring, [1,1′-bis(diphenylphosphino)ferrocene]palladium dichloride (29 mg, 0.04 mmol) was added. The mixture was microwaved at 120°C under an argon atmosphere for 40 minutes. After completion of the reaction, 50 mL of ethyl acetate and 20 mL of water were added, the layers were separated, the aqueous phase was extracted with ethyl acetate (30 mL), and the organic phases were combined. The organic phase was washed with saturated sodium chloride solution (40 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by HPLC to give compound 9 (10 mg, white solid) in a yield of 6.06%.

MS m/z (LC-MS): 414.4 [M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ7.99(s, 1H), 7.95(s, 1H), 7.71(s, 1H), 7.51-7.46(m, 2H), 7.35(d, 2H), 7.23 (s, 1H), 7.16-7.13 (m, 1H), 6.87 (d, 2H), 5.70 (s, 1H), 3.83 (s, 3H), 3.76-3.73 (m , 1H), 3.63-3.60(m, 1H), 2.70-2.64(m, 1H), 2.37-2.34(m, 1H), 1.79-1.76(m, 1 H), 1.70-1.65(m, 1H), 1.34-1.30(m, 1H), 1.20-1.17(m, 1H), 0.91-0.87(m, 1H).

Example 10

6-Fluoro-5-(1-(4-(piperazin-1-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

tert-Butyl 4-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)piperazine-1-carboxylate 10a

Compound 9a (82 mg, 2 mmol) and 1-tert-butyloxycarbonylpiperazine (760 mg, 4 mmol) were dissolved in 15 mL of toluene. (±)-2,2′-bis-(diphenylphosphino)-1,1′-binaphthyl (125 mg, 0.2 mmol), sodium tert-butoxide (576 mg, 6 mmol), and tris(dibenzylideneacetone)dipalladium (183 mg, 0.2 mmol) were added. The reaction system was microwaved at 120°C for 75 minutes. After completion of the reaction, the product was filtered through celite, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A to obtain compound 10a (517 mg, yellow solid) in a 50% yield.

MS m/z (LC-MS): 518.2[M+1]

Step 2

6-Fluoro-5-(1-(4-(piperazin-1-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 10b

Compound 10a (517 mg, 1 mmol) was dissolved in 16 mL of dichloromethane, and 4 mL of trifluoroacetic acid was added. The mixture was allowed to react overnight at room temperature. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by HPLC to obtain compound 10b (15.4 mg, light yellow solid) in a 3.7% yield.

MS m/z (LC-MS): 418.2 [M-1]

Step 3

6-Fluoro-5-(1-(4-(piperazin-1-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 10

Compound 10b (14.5 mg, 0.035 mmol) was dissolved in 5 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 12 hours. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 10 (20 mg, light yellow solid) in a yield of 100%.

MS m/z(ESI):418.2[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ7.77(s, 1H), 7.34-7.38(m, 3H), 7.22(s, 1H), 6.98-6.96(m, 1H), 6.85-6.84(m, 3H), 5.38(s, 1H), 3.65(m, 1H) , 3.62 (m, 1H), 3.34 (m, 1H), 3.09 (m, 1H), 2.70 (m, 1H), 2.56 (m, 2H), 2.38 (m, 2H), 1.86 (m, 3H), 1.31-1.16 (m, 5H).

Example 11

4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-N-(tetrahydro-2H-pyran-4-yl)benzamide trifluoroacetate

first step

4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)benzoic acid 11a

Compound 5a (630 mg, 1.76 mmol) was added to 15 mL of 6N hydrochloric acid, and the reaction system was reacted at 100°C for 12 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was dried to obtain crude compound 11a (664 mg, light brown solid), which was directly used in the next reaction without purification.

MS m/z (LC-MS): 378.1[M+1]

Step 2

4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-N-(tetrahydro-2H-pyran-4-yl)benzamide 11b

Crude compound 11a (75 mg, 0.2 mmol), 4-aminotetrahydropyran (40 mg, 0.4 mmol), and triethylamine (0.14 mL, 1.0 mmol) were added to 1.0 mL of N,N-dimethylformamide, followed by 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (152 mg, 0.4 mmol). The reaction system was incubated at 50°C for 12 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by high-performance liquid chromatography to obtain compound 11b (48 mg, light brown solid) in a yield of 52.1%.

MS m/z (LC-MS): 461.4 [M+1]

Step 3

4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-N-(tetrahydro-2H-pyran-4-yl)benzamide trifluoroacetate 11

Compound 11b (48 mg, 0.104 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.1 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 11 (60 mg, light brown solid) with a yield of 100%.

MS m/z(ESI):461.4[M+1]

¹ H NMR (400 MHz, DMSO-d ₆ )δ9.46 (s, 1H), 8.04 (s, 1H), 7.98 (d, 1H), 7.76-7.63 (m, 4H), 7.44-7.39 (m, 1H), 6.89 (d, 2H), 6.11 (d, 1H), 3.98-3.80 (m, 5H), 3.38-3.32 (m , 2H), 2.83-2.73(m, 1H), 2.73-2.63(m, 1H), 2.62-2.54(m, 1H), 1.79- 1.68(m, 3H), 1.59-1.48(m, 3H), 1.34-1.25(m, 1H), 1.05-0.95(m, 1H).

Example 12

N-(3-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)acetamide trifluoroacetate

first step

6-Fluoro-5-(1-(3-nitrophenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 12a

The crude product, Compound 1g (230 mg, 0.5 mmol), 1-iodo-3-nitrobenzene (149 mg, 0.6 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (24 mg, 0.05 mmol), sodium tert-butoxide (192 mg, 2.0 mmol), and palladium acetate (12.0 mg, 0.05 mmol) were added to a mixture of toluene and tert-butanol (V:V = 5:1) in a microwave oven at 160°C for 30 minutes. After completion of the reaction, 20 mL of saturated sodium sulfite solution was added, and the mixture was extracted with dichloromethane (20 mL x 3). The organic phases were combined, washed with saturated sodium sulfite solution (20 mL x 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A to afford Compound 12a (50 mg, brown viscous material) in a yield of 26.5%.

MS m/z (LC-MS): 379.1[M+1]

Step 2

3-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)aniline 12b

Compound 12a (50 mg, 0.132 mmol) and 10 mg of 10% palladium on carbon were added to 2 mL of a mixture of methanol and tetrahydrofuran (V:V = 1:1). The reaction system was purged with hydrogen three times and allowed to react at room temperature for 3 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain crude compound 12b (46 mg, a reddish-brown viscous solid), which was directly used in the next reaction without purification.

MS m/z (LC-MS): 349.0 [M+1]

Step 3

N-(3-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)acetamide 12c

Crude compound 12b (46 mg, 0.132 mmol), acetic acid (16 mg, 0.264 mmol), 1-hydroxybenzotriazole (36 mg, 0.264 mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (51 mg, 0.264 mmol), and N,N-diisopropylethylamine (85 mg, 0.66 mmol) were added to 1 mL of N,N-dimethylformamide and the reaction system was allowed to react at room temperature for 2 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by HPLC to obtain compound 12c (8.4 mg, brown syrup) in a 16.3% yield.

MS m/z (LC-MS): 391.4 [M+1]

Step 4

N-(3-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)acetamide trifluoroacetate 12

Compound 12c (8.4 mg, 0.021 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.05 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 12 (10.8 mg, light brown solid) with a yield of 100%.

MS m/z(ESI):391.4[M+1]

Example 13

2-(3-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-N-methylacetamide trifluoroacetate

first step

Methyl 2-(3-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)acetate 13b

To the crude product compound 1g (1.21g, 2.5mmol), methyl 2-(3-bromophenyl)acetate 13a (1.15g, 5.0mmol, prepared by the known method "Journal of Medicinal Chemistry, 2008, 51(3), 392-395"), cuprous iodide (95mg, 0.5mmol), L-proline (115mg, 1mmol), and potassium carbonate (1.38g, 10.0mmol) were added 10mL of dimethyl sulfoxide, and the reaction system was reacted at 90°C under an argon atmosphere for 24 hours. After the reaction, 100mL of ethyl acetate was added, and the mixture was washed with water (250mL×3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent system A to obtain the title product, compound 13b (410mg, brown syrup), in a yield of 40.5%.

MS m/z (LC-MS): 406.0 [M+1]

Step 2

2-(3-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)acetic acid 13c

Compound 13b (410 mg, 1.0 mmol) was dissolved in 4 mL of tetrahydrofuran, and 4 mL of 1 M sodium hydroxide solution was added. The mixture was allowed to react at room temperature for 12 hours. After completion of the reaction, 40 mL of water was added, and the mixture was extracted with ethyl acetate (25 mL × 3). The aqueous phase was adjusted to pH 6 by dropwise addition of acetic acid, and then extracted with dichloromethane (20 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain crude compound 13c (190 mg, brown solid). The product was directly used in the next reaction without purification.

MS m/z (LC-MS): 392.0 [M+1]

Step 3

2-(3-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-N-methylacetamide 13d

Crude compound 13c (190 mg, 0.485 mmol), monomethylamine hydrochloride (65 mg, 0.97 mmol), 2-(7-azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (369 mg, 0.097 mmol), and triethylamine (245 mg, 2.425 mmol) were added to 2 mL of N,N-dimethylformamide, and the reaction system was reacted at 50°C for 12 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by HPLC to obtain compound 13d (39 mg, brown solid) in a yield of 19.9%.

MS m/z (LC-MS): 405.0 [M+1]

Step 4

2-(3-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-N-methylacetamide trifluoroacetate 13

Compound 13d (39 mg, 0.096 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.05 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 13 (50 mg, light brown solid) in a yield of 100%.

MS m/z(ESI):405.0[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.52(s, 1H), 8.07(s, 1H), 7.97-7.89(m, 1H), 7.76(d, 1H), 7.70-7.64(m, 1H), 7. 46-7.41(m, 1H), 7.22-7.14(m, 1H), 7.04-6.73(m, 3H), 6.14(d, 1H), 3.73-3.63(m, 1H), 3.63-3.53(m, 1H), 3.32(s, 2H), 3.00-2.70(m, 2H), 2.63-2.53(m, 1H), 2.55(d , 3H), 1.91-1.81(m, 1H), 1.73-1.58(m, 1H), 1.38-1.28(m, 1H), 1.10-1.00(m, 1H).

Example 14

6-Fluoro-5-(1-(1-methyl-1H-indol-4-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

6-Fluoro-5-(1-(1-methyl-1H-indol-4-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 14a

The crude compound 1g (367 mg, 0.76 mmol) and 4-bromo-1-methyl-1H-indole (150 mg, 0.714 mmol) were dissolved in 15 mL of a mixed solvent of toluene and tert-butanol (V:V = 5:1). 2-Dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (34 mg, 0.071 mmol), sodium tert-butoxide (274 mg, 2.86 mmol), and palladium acetate (16.0 mg, 0.071 mmol) were added. The reaction system was reacted at 120°C under an argon atmosphere for 12 hours. After completion of the reaction, the reaction solution was filtered, the filtrate was concentrated under reduced pressure, and the resulting residue was purified by high-performance liquid chromatography to obtain compound 14a (23 mg, yellow solid) in an 8.4% yield.

MS m/z (LC-MS): 387.4 [M-1]

Step 2

6-Fluoro-5-(1-(1-methyl-1H-indol-4-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 14

Compound 14a (23 mg, 0.06 mmol) was dissolved in 2 mL of dichloromethane, and 0.1 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 14 (30 mg, yellow solid) with a yield of 100%.

MS m/z (LC-MS): 387.4 [M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.54 (s, 1H), 8.09 (s, 1H), 7.78 (d, 1H), 7.65-7.71 (m, 1H), 7.45 (t, 1H), 7.02-7.33 (m, 3H), 6.75 (br.s, 1H), 6.41 (br.s, 1H) ), 6.12(s, 1H), 3.75(s, 3H), 3.55-3.74(m, 2H), 2.52-2.80(m, 2H), 1.70-2.00(m, 3H), 1.30-1.40(m, 1H), 1.20-1.30(m, 1H).

Example 15

6-Fluoro-5-(1-(pyridin-2-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

6-Fluoro-5-(1-(pyridin-2-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 15a

Crude compound 1g (115 mg, 0.25 mmol), 2-bromopyridine (47 mg, 0.30 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (12.0 mg, 0.025 mmol), and sodium tert-butoxide (96 mg, 1.0 mmol) were added to a 1.8 mL mixture of toluene and tert-butanol (V:V = 5:1) and stirred until homogeneous. Palladium acetate (6.0 mg, 0.025 mmol) was added, and the reaction system was microwaved at 160°C for 30 minutes. After completion of the reaction, 20 mL of water was added, and the mixture was extracted with dichloromethane (20 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to afford compound 15a (7.1 mg, brown syrup) in a yield of 6.8%.

MS m/z (LC-MS): 335.0 [M+1]

Step 2

6-Fluoro-5-(1-(pyridin-2-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 15

Compound 15a (7.1 mg, 0.21 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.1 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 15 (9.5 mg, light brown solid) in a yield of 100%.

MS m/z (LC-MS): 335.0 [M+1]

Example 16

6-Fluoro-5-(1-(pyrimidin-2-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

6-Fluoro-5-(1-(pyrimidin-2-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 16a

The crude compound 1g (230 mg, 0.5 mmol), 2-chloropyrimidine (57 mg, 0.50 mmol), and triethylamine (202 mg, 2.0 mmol) were added to a sealed tube, followed by 5 mL of ethanol. The reaction system was incubated at 100°C for 12 hours. After completion of the reaction, the reaction mixture was concentrated under reduced pressure, and the resulting residue was purified by HPLC to obtain compound 16a (127 mg, light brown syrup) in a yield of 75.5%.

MS m/z (LC-MS): 336.0 [M+1]

Step 2

6-Fluoro-5-(1-(pyrimidin-2-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 16

16a (127 mg, 0.38 mmol) was dissolved in 2 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 16 (170 mg, light brown solid) in a yield of 100%.

MS m/z(ESI):336.0[M+1]

¹ H NMR (400 MHz, DMSO-d ₆ )δ9.48(s, 1H), 8.31(d, 2H), 8.04(s, 1H), 7.75(d, 1H), 7.67-7.61(m, 1H), 7.42-7.37(m, 1H), 6.58-6.56(m, 1H), 6.10(d, 1H), 4.85-4.75(m , 1H), 4.71-4.61(m, 1H), 2.93-2.81(m, 1H), 2.81-2.62(m, 3H), 1.85- 1.74(m, 1H), 1.46-1.36(m, 1H), 1.32-1.22(m, 1H), 0.87-0.77(m, 1H).

Example 17

4-(2-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)pyrimidin-4-yl)morpholine trifluoroacetate

first step

6-Fluoro-5-(piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 17a

Compound 1f (17.1 g, 47.9 mmol) was dissolved in 100 mL of dichloromethane, and 50 mL of trifluoroacetic acid was added dropwise. The mixture was allowed to react at room temperature for 12 hours. After the reaction, most of the dichloromethane and trifluoroacetic acid were removed by concentration under reduced pressure. 100 mL of dichloromethane was added, and saturated sodium bicarbonate solution was added dropwise until no bubbles were released. The pH was adjusted to 7, and the layers were separated. The aqueous phase was extracted with dichloromethane (200 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain crude compound 17a (12.3 g, brown syrup). The product was directly used in the next reaction without purification.

MS m/z (LC-MS): 258.3[M+1]

Step 2

4-(2-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)pyrimidin-4-yl)morpholine 17c

The crude compound 17a (260 mg, 1.005 mmol) was dissolved in 2.5 mL of ethanol, and 4-(2-chloropyrimidin-4-yl)morpholine 17b (100 mg, 0.503 mmol, prepared using the known method "Chemistry & Biology Interface, 2012, 2(5), 347-361") and triethylamine (203 mg, 2.012 mmol) were added. The mixture was stirred at 100°C for 48 hours. After the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by high-performance liquid chromatography to obtain compound 17c (59 mg, yellow solid) in a 30% yield.

MS m/z (LC-MS): 421.3 [M+1]

Step 3

4-(2-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)pyrimidin-4-yl)morpholine trifluoroacetate 17

Compound 17c (59 mg, 0.14 mmol) was dissolved in 3 mL of dichloromethane, and 0.1 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 30 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 17 (75 mg, yellow gum) with a yield of 100%.

MS m/z(ESI):421.3[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.36 (s, 1H), 8.03 (s, 1H), 7.86 (d, 1H), 7.75 (d, 1H), 7.63-7.68 (m, 1H), 7.41 (t, 1H), 6.52 (d, 1H), 6.10 (d, 1H), 4.30-4.55 (m, 2H), 3.6 0-3.85(m, 8H), 3.09(t, 1H), 2.99(t, 1H), 2.70-2.80(m, 1H), 1.82-1.88(m, 1H), 1.36-1.55(m, 1H), 1.30-1.36(m, 1H), 0.88-1.05(m, 1H).

Example 18

6-Fluoro-5-(1-(5-(1-(tetrahydro-2H-pyran-4-yl)-1H-pyrazol-4-yl)pyrimidin-2-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole

first step

5-(1-(5-bromopyrimidin-2-yl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole 18a

In a sealed tube, crude compound 1g (1.0g, 2.2mol), 5-bromo-2-chloropyrimidine (468mg, 2.42mmol), and triethylamine (1.1g, 11mmol) were added to 20mL of ethanol and reacted at 90°C for 12 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure and the resulting residue was purified by silica gel column chromatography with eluent System A to obtain compound 18a (730mg, orange-yellow solid) in a yield of 80.2%.

MS m/z (LC-MS): 414.2[M+2]

Step 2

6-Fluoro-5-(1-(5-(1-(tetrahydro-2H-pyran-4-yl)-1H-pyrazol-4-yl)pyrimidin-2-yl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 18

Compound 18a (124 mg, 0.3 mmol) was dissolved in 5 mL of ethylene glycol dimethyl ether and 0.5 mL of water, and 1-(tetrahydro-2H-pyran-4-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-1H-pyrazole 18b (125 mg, 0.45 mmol, prepared by the known method "Bioorganic & Medicinal Chemistry, 2013, 21(21), 6804-6820"), tetrakis(triphenylphosphine)palladium (69 mg, 0.06 mmol), and sodium carbonate (63.6 mg, 0.6 mmol) were added. The mixture was reacted at 80°C under an argon atmosphere for 12 hours. After the reaction, 30 mL of water was added, and the mixture was extracted with dichloromethane (40 mL×3). The organic phases were combined, washed with saturated sodium chloride solution (40 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by high performance liquid chromatography to obtain compound 18 (30 mg, white solid) with a yield of 20.7%.

MS m/z (LC-MS): 486.5[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ8.56(s, 2H), 8.18(s, 1H), 7.94(s, 1H), 7.82(s, 1H), 7.48-7.46(m, 2H), 7.20(s, 1H), 7.16 -7.11(m, 1H), 5.68(s, 1H), 4.81-4.78(m, 1H), 4.64-4.61(m, 1H), 4.42-4.36(m, 1H), 3.98-3 .95(m, 2H), 3.50-3.44(m, 2H), 2.92-2.86(m, 1H), 2.78-2.72(m, 1H), 2.01-1.89(m, 4H), 1.8 1-1.78(m, 1H), 1.53-1.45(m, 1H), 1.26-1.24(m, 1H), 1.19-1.17(m, 1H), 0.73-0.63(m, 1H).

Example 19

6-Fluoro-5-(1-methylpiperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

6-Fluoro-5-(1-methylpiperidin-4-yl)-5H-imidazo[5,1-a]isoindole 19a

The crude compound 1g (121 mg, 0.25 mmol) was dissolved in 2 mL of N,N-dimethylformamide, potassium carbonate (173 mg, 1.25 mmol) was added, and the mixture was stirred until uniform. Methyl iodide (21 mg, 0.15 mmol) was added, and the mixture was allowed to react at room temperature for 48 hours. After the reaction, 20 mL of ethyl acetate was added, and the mixture was washed with water (10 mL × 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to obtain compound 19a (16.0 mg, light brown viscous substance) in a yield of 38.9%.

MS m/z (LC-MS): 272.0 [M+1]

Step 2

6-Fluoro-5-(1-methylpiperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 19

The crude compound 19a (16.0 mg, 0.59 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.05 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 19 (22.6 mg, light brown solid) in a yield of 100%.

MS m/z (LC-MS): 272.0 [M+1]

¹ H NMR (400MHz, CD ₃ OD) δ9.31 (s, 1H), 7.91 (s, 1H), 7.73 (d, 1H), 7.67-7.62 (m, 1H), 7.36-7.31 (m, 1H), 6.08 (d, 1H), 3.61-3.52 (m, 1H), 3.52-3.43 (m, 1 H), 3.12-2.98(m, 2H), 2.88-2.76(m, 1H), 2.83(s, 3H), 2.01-1.92(m, 1H), 1.87-1.73(m, 1H), 1.73-1.62(m, 1H), 1.54-1.39(m, 1H).

Example 20

1-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)ethanone trifluoroacetate

first step

1-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)ethanone 20a

Dissolve the crude compound 1g (31 mg, 0.084 mmol) in 2 mL of dichloromethane, add triethylamine (34 mg, 0.336 mmol), stir thoroughly, and then add acetyl chloride (13 mg, 0.168 mmol). Allow to react at room temperature for 48 hours. After completion of the reaction, concentrate the reaction mixture under reduced pressure, and purify the resulting residue by HPLC to obtain compound 20a (7.2 mg, brown solid) in a 40% yield.

MS m/z (LC-MS): 300.0 [M+1]

Step 2

1-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)ethanone trifluoroacetate

The crude compound 20a (7.2 mg, 0.024 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.01 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 20 (10 mg, light brown solid) in a yield of 100%.

MS m/z (LC-MS): 300.0 [M+1]

Example 21

4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)-N-phenylpiperidine-1-carboxamide trifluoroacetate

first step

4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)-N-phenylpiperidine-1-carboxamide 21a

Bis(trichloromethyl)carbonate (297 mg, 1.0 mmol) was dissolved in 2 mL of dichloromethane, and a pre-prepared dichloromethane solution of 2 mL of aniline (93 mg, 1.0 mmol) was added dropwise, followed by a pre-prepared dichloromethane solution of 1 mL of triethylamine (0.28 mL). The mixture was allowed to react at room temperature for 10 minutes. After concentration under reduced pressure, 5 mL of tetrahydrofuran and 0.28 mL of triethylamine were added, followed by compound 1g (243 mg, 0.5 mmol), and the mixture was allowed to react at room temperature for 12 hours. After completion of the reaction, 20 mL of ethyl acetate and 20 mL of water were added, the liquids were separated, the organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high performance liquid chromatography to afford compound 21a (92 mg, light yellow solid) in a yield of 48.9%.

MS m/z (LC-MS): 377.0 [M-1]

Step 2

4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)-N-phenylpiperidine-1-carboxamide trifluoroacetate 21

The crude compound 21a (92 mg, 0.244 mmol) was dissolved in 2 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 21 (120 mg, light brown solid) in a yield of 100%.

MS m/z (LC-MS): 377.0 [M-1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.47(s, 1H), 8.45(s, 1H), 8.04(s, 1H), 7.75(d, 1H), 7.68-7.63(m, 1H), 7.43-7.37(m, 3H), 7.21-7.17(m, 2H), 6.91-6.88(m, 1H), 6.08(d, 1H), 4.2 8-4.18(m, 1H), 4.15-4.05(m, 1H), 2.83-2.68(m, 2H), 2.64-2.54(m, 1H), 1 .76-1.67(m, 1H), 1.42-1.32(m, 1H), 1.28-1.19(m, 1H), 0.91-0.81(m, 1H).

Example 22

N-(4-Benzonitrile)-4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidine-1-carboxamide trifluoroacetate

first step

N-(4-Benzonitrile)-4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidine-1-carboxamide 22a

Bis(trichloromethyl)carbonate (297 mg, 1.0 mmol) was dissolved in 2 mL of dichloromethane, and a pre-prepared dichloromethane solution of 2 mL of p-aminobenzonitrile (118 mg, 1.0 mmol) was added dropwise. A pre-prepared dichloromethane solution of 1 mL of triethylamine (0.28 mL) was also added dropwise. The mixture was allowed to react at room temperature for 1 hour. After concentration under reduced pressure, 5 mL of tetrahydrofuran, 0.28 mL of triethylamine, and compound 1g (243 mg, 0.5 mmol) were added, and the mixture was allowed to react at room temperature for 12 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by high-performance liquid chromatography to obtain compound 22a (101 mg, light brown solid) in a yield of 50.5%.

MS m/z (LC-MS): 402.0 [M+1]

Step 2

N-(4-Benzonitrile)-4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidine-1-carboxamide trifluoroacetate 22

Compound 22a (101 mg, 0.25 mmol) was dissolved in 2 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 22 (130 mg, light brown solid) in a yield of 100%.

MS m/z (LC-MS): 402.0 [M+1]

¹ H NMR (400 MHz, DMSO-d ₆ )δ9.47 (s, 1H), 8.99 (s, 1H), 8.06 (s, 1H), 7.76 (d, 1H), 7.68-7.60 (m, 5H), 7.44-7.39 (m, 1H), 6.09 (d, 1H), 4.30-4.20 (m, 1H), 4.15-4.05 (m , 1H), 2.91-2.81(m, 1H), 2.81-2.71(m, 1H), 2.68-2.58(m, 1H), 1.79- 1.68(m, 1H), 1.44-1.34(m, 1H), 1.31-1.21(m, 1H), 0.93-0.83(m, 1H).

Example 23

2-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-5-phenylthiazole trifluoroacetate

first step

5-Bromo-2-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)thiazole 23a

The crude compound 1g (2.43 g, 5.0 mmol), 2,5-dibromothiazole (1.82 g, 7.5 mmol), and triethylamine (2.02 g, 20.0 mmol) were added to 15 mL of dimethyl sulfoxide, and the reaction system was microwave-activated at 120°C for 1.5 hours. After completion of the reaction, 250 mL of ethyl acetate was added, and the mixture was washed with water (150 mL × 3) and saturated sodium chloride solution (150 mL). The mixture was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A to obtain compound 23a (710 mg, brown solid) in a yield of 33.9%.

MS m/z (LC-MS): 420.0 [M+1]

Step 2

2-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-5-phenylthiazole 23b

Compound 23a (105 mg, 0.25 mmol), phenylboronic acid (46 mg, 0.375 mmol), tetrakis(triphenylphosphine)palladium (58 mg, 0.05 mmol), and potassium phosphate trihydrate (133 mg, 0.5 mmol) were added to 1.4 mL of a mixed solvent of N,N-dimethylformamide and water (V:V = 6:1). The reaction system was microwaved at 120°C for 35 minutes. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by high-performance liquid chromatography to obtain compound 23b (24 mg, brown syrup) in a yield of 22.7%.

MS m/z (LC-MS): 417.0 [M+1]

Step 3

2-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-5-phenylthiazole trifluoroacetate 23

Compound 23b (24 mg, 0.057 mmol) was dissolved in 0.5 mL of dichloromethane, and 0.05 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 23 (30 mg, light brown solid) with a yield of 100%.

MS m/z (LC-MS): 417[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.42(s, 1H), 8.02(s, 1H), 7.74(d, 1H), 7.68-7.62(m, 1H), 7.57(s, 1H), 7.44 -7.38(m, 3H), 7.35-7.31(m, 2H), 7.18-7.22(m, 1H), 6.11(d, 1H), 4.06-3.99(m , 1H), 3.92-3.84(m, 1H), 3.17-3.07(m, 1H), 3.04-2.94(m, 1H), 2.70-2.60(m, 1 H), 1.91-1.81(m, 1H), 1.67-1.52(m, 1H), 1.34-1.25(m, 1H), 1.04-0.93(m, 1H).

Example 24

5-(1-(1H-benzo[d]imidazol-2-yl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole

The crude compound 1g (257g, 1mmol) and 2-chlorobenzimidazole (153mg, 1mmol) were dissolved in 10mL of N-methylpyrrolidone, and N,N-diisopropylethylamine (390mg, 3mmol) was added. The mixture was reacted at 90°C for 12 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by silica gel column chromatography with eluent System A to obtain compound 24 (20mg, white solid) in a 41% yield.

MS m/z (LC-MS): 374.2 [M-1]

¹ H NMR (400MHz, CDCl ₃ ) δ7.74 (s, 1H), 7.38-7.17 (m, 5H), 7.17 (s, 1H), 6.98 (s, 2H), 5.32 (s, 1H), 4.38-4.21 (m, 2H), 3.04-2.93 (m, 2H), 2.48 (m 1H), 1.82-1.71(m, 2H), 1.29-1.23(m, 1H), 0.92(m, 1H).

Example 25

6-Fluoro-5-(1-phenylpyrrolidin-3-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

Tert-Butyl 3-((2-bromo-6-fluorophenyl)(hydroxy)methyl)pyrrolidine-1-carboxylate 25b

Lithium diisopropylamide (13 mL, 26 mmol) was added to 20 mL of tetrahydrofuran and cooled to -78°C under argon. A pre-prepared tetrahydrofuran solution of 1-bromo-3-fluorobenzene 1a (3.5 g, 20.0 mmol) (10 mL) was added dropwise at -78°C, and the mixture was allowed to react at -78°C for 1 hour. Furthermore, a pre-prepared tetrahydrofuran solution of tert-butyl 3-formylpyrrolidine-1-carboxylate 25a (3.985 g, 20.0 mmol, prepared by the known method "Jpn. Tokyo Koh, 2009, 3 Jun, 4272338") (10 mL) was added dropwise at -78°C, and the mixture was allowed to react at -78°C for 1 hour. After the reaction, 10 mL of methanol was added dropwise at -78°C to quench the reaction. The reaction solution was concentrated under reduced pressure and the residue was purified by silica gel column chromatography using eluent System B to obtain compound 25b (5.15 g, light yellow oil) in a yield of 68.8%.

MS m/z (LC-MS): 320.0 [M-56]

Step 2

Tert-butyl 3-((2-bromo-6-fluorophenyl)((methylsulfonyl)oxy)methyl)pyrrolidine-1-carboxylate 25c

Compound 25b (5.1 g, 13.6 mmol) was dissolved in 50 mL of dichloromethane, and triethylamine (3.8 mL, 27.7 mmol) was added. Methanesulfonyl chloride (1.639 g, 14.3 mmol) was added dropwise under an ice bath, and the mixture was allowed to react at room temperature for 1 hour. After the reaction, 50 mL of dichloromethane was added, and the mixture was washed with water (60 mL) and saturated sodium chloride solution (60 mL). The mixture was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain crude compound 25c (5.9 g, yellow viscous solid). The product was directly used in the next reaction without purification.

MS m/z (LC-MS): 3000 [M-56-95]

Step 3

Tert-butyl 3-((2-bromo-6-fluorophenyl)(1H-imidazol-1-yl)methyl)pyrrolidine-1-carboxylate 25d

The crude product, compound 25c (2.4 g, 5.3 mmol), was dissolved in 10 mL of acetonitrile, and imidazole (3.6 g, 53 mmol) and N,N-diisopropylethylamine (6.85 g, 53 mmol) were added. The mixture was microwaved at 120°C for 1 hour and 40 minutes. After completion of the reaction, 100 mL of ethyl acetate was added, and the mixture was washed with water (60 mL × 2) and saturated sodium chloride solution (60 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain compound 25d (2.7 g, brown viscous solid). The product was directly used in the next reaction without purification.

MS m/z (LC-MS): 424.3[M+1]

Step 4

tert-Butyl 3-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)pyrrolidine-1-carboxylate 25e

The crude product, compound 25d (2.4 g, 5.66 mmol), was dissolved in 10 mL of N,N-dimethylformamide solution. N,N-dicyclohexylmethylamine (1.77 g, 9.05 mmol), triphenylphosphine (594 mg, 2.264 mmol), and palladium acetate (254 mg, 1.132 mmol) were added and microwave-treated at 120°C for 1 hour. After completion of the reaction, the N,N-dimethylformamide solution was removed by concentration under reduced pressure. 100 mL of ethyl acetate was added, and the mixture was washed with water (40 mL x 2) and saturated sodium chloride solution (60 mL). The mixture was dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated under reduced pressure and the resulting residue was purified by silica gel column chromatography using eluent System B to afford compound 25e (1.62 g, light brown oil) in a yield of 78.3%.

MS m/z (LC-MS): 344.2[M+1]

Step 5

6-Fluoro-5-(pyrrolidin-3-yl)-5H-imidazo[5,1-a]isoindole ditrifluoroacetate 25f

Compound 25e (1.62 g, 4.71 mmol) was dissolved in 20 mL of dichloromethane, and 2.69 mL of trifluoroacetic acid was added dropwise. The mixture was reacted at room temperature for 4 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain crude compound 25f (2.76 g, brown oil), which was directly used in the next reaction without purification.

MS m/z (LC-MS): 243.9 [M+1]

Step 6

6-Fluoro-5-(1-phenylpyrrolidin-3-yl)-5H-imidazo[5,1-a]isoindole 25g

Crude compound 25f (292 mg, 0.50 mmol) was dissolved in 1.5 mL of a mixed solvent of toluene and ethanol (V:V = 5:1). Bromobenzene (94 mg, 0.6 mmol), 2-dicyclohexylphosphino-2',4',6'-triisopropylbinaphthyl (24 mg, 0.05 mmol), sodium tert-butoxide (240 mg, 2.50 mmol), and palladium acetate (11 mg, 0.05 mmol) were added. The mixture was microwaved at 120°C under an argon atmosphere for 50 minutes. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by high-performance liquid chromatography to afford compound 25g (23 mg, brown oil) in a yield of 14.3%.

MS m/z(ESI):320.3[M+1]

Step 7

6-Fluoro-5-(1-phenylpyrrolidin-3-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 25

Compound 25g (23 mg, 0.072 mmol) was dissolved in 5 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 60 minutes. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 25 (31 mg, brown oily solid) with a yield of 100%.

MS m/z(ESI):320.3[M+1]

Example 26

6-Fluoro-5-(1-(3-fluorophenyl)piperidin-3-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

tert-Butyl 3-((2-bromo-6-fluorophenyl)(hydroxy)methyl)piperidine-1-carboxylate 26b

15 mL of tetrahydrofuran was added to a reaction flask, cooled to -78°C under an argon atmosphere, and lithium diisopropylamide (9.3 mL, 18.6 mmol) was added. 1-Bromo-3-fluorobenzene 1a (2.5 g, 14.3 mmol) was added dropwise, and the mixture was allowed to react at -78°C for 1 hour. A pre-prepared tetrahydrofuran solution of tert-butyl 3-formylpiperidine-1-carboxylate 26a (3.0 g, 14.3 mmol) was then added dropwise, and the mixture was allowed to react at -78°C for 1 hour. After completion of the reaction, 15 mL of methanol was added dropwise at -78°C to quench the reaction. The reaction solution was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography using eluent System B to afford compound 26b (3.9 g, slightly yellow solid) in a 34% yield.

MS m/z (LC-MS): 334.0 [M-55]

Step 2

Tert-butyl 3-((2-bromo-6-fluorophenyl)((methylsulfonyl)oxy)methyl)piperidine-1-carboxylate 26c

Compound 26b (3.9 g, 10 mmol) was dissolved in 40 mL of dichloromethane, and triethylamine (2.02 g, 20 mmol) was added dropwise. Methanesulfonyl chloride (1.2 g, 10.05 mmol) was then added dropwise. The mixture was allowed to react at room temperature for 12 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure and the resulting residue was purified by silica gel column chromatography using eluent System B to afford compound 26c (1.9 g, light yellow oil) in a yield of 40.7%.

MS m/z (LC-MS): 314.0 [M-152]

Step 3

Tert-butyl 3-((2-bromo-6-fluorophenyl)(1H-imidazol-1-yl)methyl)piperidine-1-carboxylate 26d

Compound 26c (2.0 g, 4.28 mmol) was dissolved in 5 mL of acetonitrile, and imidazole (2.9 g, 42.8 mmol) and N,N-diisopropylethylamine (5.5 g, 42.8 mmol) were added. The mixture was microwaved at 120°C for 1 hour. After completion of the reaction, the reaction solution was concentrated under reduced pressure, and the resulting residue was purified by silica gel column chromatography using eluent System A to obtain the title product, compound 26d (0.85 g, light brown oil) in a 47% yield.

MS m/z (LC-MS): 440.0 [M+2]

Step 4

tert-Butyl 3-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidine-1-carboxylate 26e

Compound 26d (1.0 g, 2.28 mmol) was dissolved in 5 mL of N,N-dimethylformamide solution, and N,N-dicyclohexylmethylamine (712 mg, 3.65 mmol), triphenylphosphine (239 mg, 0.91 mmol), and palladium acetate (100 mg, 0.45 mmol) were added. The mixture was microwaved at 120°C for 1 hour. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain crude compound 26e (1.21 g, orange oil), which was directly used in the next reaction without purification.

MS m/z (LC-MS): 358.2[M+1]

Step 5

6-Fluoro-5-(piperidin-3-yl)-5H-imidazo[5,1-a]isoindole ditrifluoroacetate 26f

Crude compound 26e (1.7 g, 4.76 mmol) was dissolved in 20 mL of dichloromethane, and 1 mL of trifluoroacetic acid was added dropwise. The mixture was reacted at room temperature for 48 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure to obtain crude compound 26f (2.5 g, brown oil), which was directly used in the next reaction without purification.

MS m/z (LC-MS): 258.0 [M+1]

Step 6

6-Fluoro-5-(1-(3-fluorophenyl)piperidin-3-yl)-5H-imidazo[5,1-a]isoindole 26g

Crude compound 26f (200 mg, 0.44 mmol), 1-bromo-3-fluorobenzene (93 mg, 0.53 mmol), 2-dicyclohexylphosphino-2′,4′,6′-triisopropylbiphenyl (21 mg, 0.044 mmol), sodium tert-butoxide (211 mg, 2.2 mmol), and palladium acetate (20 mg, 0.089 mmol) were dissolved in 5 mL of a mixed solvent of toluene and ethanol (V:V = 5:1) and microwave-treated at 120°C for 1 hour. After completion of the reaction, the mixture was filtered through celite, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to afford compound 26g (3.7 mg, brown oil) in a 3% yield.

MS m/z(ESI):352.0[M+1]

Step 7

6-Fluoro-5-(1-(3-fluorophenyl)piperidin-3-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 26

Compound 26g (3.7 mg, 0.01 mmol) was dissolved in 5 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was reacted at room temperature for 12 hours. After the reaction, the reaction solution was concentrated under reduced pressure to obtain compound 26 (4.8 mg, brown oil) with a yield of 100%.

MS m/z(ESI):352.0[M+1]

Examples 27, 28

(R)-6-Fluoro-5-(1-(4-(1-methyl-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 27

(S)-6-Fluoro-5-(1-(4-(1-methyl-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 28

Compound 9 (1.9 g, 4.59 mmol) was chirally prepared (separation conditions: chiral column CHIRALPAKIF, mobile phase: dichloromethane: methanol = 70:30, flow rate: 30 mL/min), and the corresponding components were collected and concentrated under reduced pressure to obtain compound 27 (700 mg, 1.69 mmol) in a yield of 73.7%; compound 28 (640 mg, 1.54 mmol) in a yield of 67.4%.

27:

MS m/z(ESI):414.4[M+1]

Chiral HPLC analysis: retention time 2.466 minutes, ee value >99.0%. (Chromatographic column: CHIRALPAK ID; mobile phase: DCM/MeOH/TEA = 80/20/0.1 (V/V/V))

¹ H NMR (400 MHz, DMSO-d ₆ )δ7.98(s, 1H), 7.94(s, 1H), 7.70(s, 1H), 7.44-7.51(m, 2H), 7.34(d, 2H ), 7.22(s, 1H), 7.11-7.17(m, 1H), 6.86(d, 2H), 5.69(s, 1H), 3.82(s, 3H ), 3.73(d, 1H), 3.60(d, 1H), 2.63-2.69(m, 1H), 2.33-2.36(m, 1H), 1.75 -1.78(m, 1H), 1.64-1.69(m, 1H), 1.16-1.33(m, 2H), 0.87-0.90(m, 1H).

28:

MS m/z(ESI):414.4[M+1]

Chiral HPLC analysis: retention time 4.122 minutes, ee value >99.0%. (Chromatographic column: CHIRALPAK ID; mobile phase: DCM/MeOH/TEA = 80/20/0.1 (V/V/V))

¹ H NMR (400 MHz, DMSO-d ₆ )δ7.98(s, 1H), 7.94(s, 1H), 7.70(s, 1H), 7.44-7.51(m, 2H), 7.34(d, 2H ), 7.22(s, 1H), 7.11-7.17(m, 1H), 6.86(d, 2H), 5.69(s, 1H), 3.82(s, 3H ), 3.73(d, 1H), 3.60(d, 1H), 2.63-2.69(m, 1H), 2.33-2.36(m, 1H), 1.75 -1.78(m, 1H), 1.64-1.69(m, 1H), 1.16-1.33(m, 2H), 0.87-0.90(m, 1H).

Example 29

(2S)-3-(4-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)propane-1,2-diol

first step

(S)-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-4-(4,4,5,5-tetramethyl-1-1,3,2-dioxaborolan-2-yl)-1H-pyrazole 29c

4-(4,4,5,5-Tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole 29a (1 g, 5.15 mmol) was dissolved in 6 mL of N,N-dimethylformamide. 250 mg of sodium hydride (60%) was added at room temperature and the mixture was stirred for 20 minutes. (R)-4-(chloromethyl)-2,2-dimethyl-1,3-dioxolane 29b (1.164 g, 7.73 mmol) was then added. The mixture was heated to 100°C and stirred for 12 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and the resulting residue was purified by silica gel column chromatography using eluent System A to afford the title compound 29c (790 mg, yellow oil) in a 50% yield.

Step 2

5-(1-(4-(1-(((S)-2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole 29d

Compounds 9a (250 mg, 0.606 mmol) and 29c (560 mg, 1.82 mmol) were added to 7 mL of n-butanol. Potassium phosphate (386 mg, 1.82 mmol), tris(dibenzylideneacetone)dipalladium (41.7 mg, 0.0455 mmol), and 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl (87 mg, 0.182 mmol) were also added. The mixture was heated to 100°C and reacted for 12 hours. The reaction solution was cooled to room temperature and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A, followed by thin-layer chromatography using developing solvent System A to obtain the title compound 29d (120 mg, yellow solid) in a 38.6% yield.

Step 3

(2S)-3-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)propane-1,2-diol 29

Dissolve 29d (120 mg, 0.234 mmol) in 4 mL of methanol, add 2 mL of 2N hydrochloric acid, and stir at room temperature for 3 hours. The reaction solution is concentrated under reduced pressure, and the resulting residue is purified by HPLC to obtain the title compound 29 (95 mg, orange solid) in a yield of 79%.

MS m/z(ESI):474.5[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.59 (s, 1H), 8.17 (s, 1H), 8.12 (s, 1H), 7.92 (s, 1H), 7.65-7.83 (m, 6H), 7.46 (t, 1H), 6.24 (s, 1H), 4.23 (dd, 1H), 3.95-4.0 5(m, 1H), 3.80-3.86(m, 1H), 3.50-3.75(m, 2H), 3.25-3.50(m, 4H), 2.78-2.95(m, 1H), 2.00-2.30(m, 2H), 1.50-1.70(m, 2H).

Example 30

(2R)-3-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)propane-1,2-diol trifluoroacetate

first step

(R)-1-((2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-4-(4,4,5,5-tetramethyl-1-1,3,2-dioxaborolan-2-yl)-1H-pyrazole 30b

29a (500 mg, 2.58 mmol) was dissolved in 3 mL of N,N-dimethylformamide. 113 mg of sodium hydroxide (60%) was added at room temperature and the reaction was stirred at room temperature for 20 minutes. (S)-4-(chloromethyl)-2,2-dimethyl-1,3-dioxolane 30a (505 mg, 3.35 mmol) was added and the temperature was raised to 100°C with stirring for 12 hours. 0.5 mL of methanol was added to the reaction solution to quench the reaction. The N,N-dimethylformamide was removed by concentration under reduced pressure. 50 mL of ethyl acetate and 5 mL of water were added to the resulting residue. The organic phase was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain the crude title compound 30b (400 mg, yellow oil). The product was directly used in the next step without purification.

Step 2

5-(1-(4-(1-(((R)-2,2-dimethyl-1,3-dioxolan-4-yl)methyl)-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole 30c

9a (100 mg, 0.242 mmol) and crude product 30b (223 mg, 0.726 mmol) were dissolved in 5 mL of n-butanol. Potassium phosphate (154 mg, 0.726 mmol), tris(dibenzylideneacetone)dipalladium (22 mg, 0.0242 mmol), and 2-dicyclohexylphosphino-2′,4′,6′-triisopropylbiphenyl (46 mg, 0.0968 mmol) were added. The mixture was heated to 100°C and stirred for 12 hours. The reaction solution was cooled to room temperature to obtain the crude title compound 30c (150 mg, brown oil), which was directly used in the next step without purification.

Step 3

(2R)-3-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)propane-1,2-diol trifluoroacetate 30

30c (150 mg, 0.292 mmol) was dissolved in 5 mL of methanol, and 5 mL of 2N hydrochloric acid was added. The mixture was stirred at room temperature for 12 hours. The reaction solution was concentrated under reduced pressure and the resulting residue was purified by HPLC to afford the title compound 30 (8 mg, white solid) in a 5.8% yield.

MS m/z(ESI):474.4[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.51(s, 1H), 8.07(s, 1H), 8.01(s, 1H), 7.75-7.80(m, 2H), 7.65-7.70(m, 1H) ,7.40-7.55(m,3H),7.00-7.10(m,2H),6.15(s,1H),4.20(dd,1H),3.94-4.02(m , 1H), 3.60-3.85(m, 3H), 3.25-3.40(m, 2H), 2.70-3.02(m, 2H), 2.60-2.70(m, 1 H), 1.82-1.90(m, 1H), 1.65-1.80(m, 1H), 1.30-1.40(m, 1H), 1.15-1.20(m, 1H).

Example 31

6-Fluoro-5-(1-(4-(1-(1-(methylsulfonyl)piperidin-4-yl)-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

tert-Butyl 4-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)piperidine-1-carboxylate 31b

9a (250 mg, 0.606 mmol) and tert-butyl 4-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-1H-pyrazol-1-yl)piperidine-1-carboxylate 31a (343 mg, 0.91 mmol, prepared by the known method "Bioorganic & Medicinal Chemistry, 2013, 21(21), 6804-6820") were dissolved in 5 mL of n-butanol, and potassium phosphate (260 mg, 1.212 mmol), tris(dibenzylideneacetone)dipalladium (17 mg, 0.018 mmol) and 2-dicyclohexylphosphine-2′,4′,6′-triisopropylbiphenyl (35 mg, 0.073 mmol) were added, and the temperature was raised to 100°C and stirred for 12 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and the resulting residue was purified by silica gel column chromatography using eluent System A to give the title compound 31b (160 mg, yellow solid) in a yield of 45.3%.

Step 2

6-Fluoro-5-(1-(4-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 31c

31b (160 mg, 0.275 mmol) was dissolved in 3 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was stirred at room temperature for 2 hours. The reaction solution was concentrated under reduced pressure to obtain the crude title compound 31c (250 mg, yellow oil), which was directly used in the next step without purification.

Step 3

6-Fluoro-5-(1-(4-(1-(1-(methylsulfonyl)piperidin-4-yl)-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 31

Crude product 31c (125 mg, 0.138 mmol) was dissolved in 3 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid and methylsulfonyl chloride (47 mg, 0.412 mmol) were added. The mixture was stirred at room temperature for 2 hours. 10 mL of dichloromethane and 2 mL of water were added to the reaction solution, and the layers were separated. The organic phase was concentrated under reduced pressure and the resulting residue was purified by HPLC to afford the title compound 31 (12 mg, white solid) in a yield of 12.9%.

MS m/z(ESI):561.5[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.47(s, 1H), 8.16(s, 1H), 8.05(s, 1H), 7.74-7.80(m, 2H), 7.63-7.70(m, 1H) ,7.38-7.50(m,3H),6.96(d,2H),6.13(s,1H),4.25-4.34(m,1H),3.60-3.80(m , 4H), 2.90-3.00(m, 5H), 2.50-2.70(m, 3H), 2.10-2.20(m, 2H), 1.92-2.05(m, 2 H), 1.81-1.85(m, 1H), 1.61-.165(m, 1H), 1.30-1.34(m, 1H), 1.00-1.15(m, 1H).

Example 32

6-Fluoro-5-(1-(4-(1-((R)-tetrahydrofuran-3-yl)-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

9a (103 mg, 0.25 mmol) and (R)-1-(tetrahydrofuran-3-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-1H-pyrazole 32a (99 mg, 0.375 mmol, prepared by the method disclosed in patent application "WO201493647") were dissolved in 3 mL of n-butanol, and potassium phosphate (106 mg, 0.5 mmol), tris(dibenzylideneacetone)dipalladium (17 mg, 0.018 mmol) and 2-dicyclohexylphosphine-2′,4′,6′-triisopropylbiphenyl (36 mg, 0.075 mmol) were added, and the temperature was raised to 100 ° C and the reaction was carried out for 12 hours. The reaction solution was cooled to room temperature, filtered through a pad of celite, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by HPLC to give the title compound 32 (55 mg, white solid) in a yield of 37.9%.

MS m/z (ESI): 470.5 [M+1];

¹ H NMR (400MHz, DMSO-d ₆ )δ9.49 (s, 1H), 8.10 (s, 1H), 8.06 (s, 1H), 7.8 (s, 1H), 7.76-7.78 (m, 1H), 7.41-7.45 (m, 3H), 6.97 (d, 2H), 6.14(s, 1H), 4.98-5.00(m, 1H), 3.97-4.02(m, 2H), 3.89-3.92(m, 1H), 3.80-3.86(m, 1H), 3.73-3 .76(m, 1H), 3.63-3.66(m, 1H), 2.75-2.85(m, 1H), 2.62-2.75(m, 1H), 2.53-2.59(m, 1H), 2.29-2.41(m , 2H), 1.82-1.85(m, 1H), 1.62-1.64(m, 1H), 1.31-1.34(m, 1H), 1.07-1.13(m, 1H), 0.85-0.87(m, 1H).

Example 33

(1S,4s)-4-((R)-6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)-N-(3-methoxyphenyl)cyclohexylcarboxamide

first step

(1S,4s)-4-((R)-(2-bromo-6-fluorophenyl)(hydroxy)methyl)cyclohexanecarboxylic acid methyl ester 33b

26.8 mL of lithium diisopropylamide was dissolved in 50 mL of tetrahydrofuran and cooled to -78°C. A pre-prepared solution of 1a (7.2 g, 41.18 mmol) in tetrahydrofuran (20 mL) was added dropwise and stirred for 1 hour. Then, a pre-prepared solution of (1r,4r)-methyl 4-formylcyclohexanecarboxylate 33a (7 g, 41.18 mmol, prepared using the method disclosed in patent application WO2013050334) in tetrahydrofuran (50 mL) was added dropwise and stirred for 2 hours. The reaction mixture was quenched by the addition of 10 mL of methanol, warmed to room temperature, and 100 mL of ethyl acetate was added. The mixture was washed with water (100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System B to afford the title compound 33b (6.8 g, brown oil) in a 47.9% yield.

Step 2

(1S,4s)-4-((R)-(2-bromo-6-fluorophenyl)((methylsulfonyl)oxy)methyl)cyclohexanecarboxylic acid methyl ester 33c

33b (6.8 g, 19.7 mmol) was dissolved in 100 mL of dichloromethane, and triethylamine (3.99 g, 39.4 mmol) was added, followed by dropwise addition of methylsulfonyl chloride (2.48 g, 21.67 mmol). The mixture was stirred at room temperature for 12 hours. 50 mL of dichloromethane was added to the reaction solution, which was washed with saturated sodium chloride solution (100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to afford the crude title compound 33c (8.33 g, brown oil). The product was carried on to the next step without further purification.

Step 3

(1S,4s)-4-((R)-(2-bromo-6-fluorophenyl)(1H-imidazol-1-yl)methyl)cyclohexanecarboxylate 33d

Crude product 33c (1.4 g, 3.3 mmol) was dissolved in 5 mL of acetonitrile, and 1H-imidazole (2.25 g, 33 mmol) and N,N-diisopropylethylamine (4.26 g, 33 mmol) were added. The mixture was microwaved at 130°C for 45 minutes. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and purified by silica gel column chromatography with eluent System B to afford the title compound 33d (2.8 g, brown oil) in a 35.95% yield.

Step 4

(1S,4s)-4-((R)-6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)cyclohexanecarboxylic acid methyl ester 33e

33d (2.8 g, 7.08 mmol) was dissolved in 12 mL of N,N-dimethylformamide, and N,N-dicyclohexylmethylamine (2.2 g, 11.328 mmol), triphenylphosphine (743 mg, 2.8 mmol), and palladium acetate (318 mg, 1.4 mmol) were added. The mixture was microwaved at 120°C for 1 hour. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and purified by silica gel column chromatography with eluent System B to afford the title compound 33e (628 mg, brown solid) in a 28.5% yield.

Step 5

(1S,4s)-4-((R)-6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)cyclohexanecarboxylic acid 33f

33e (628 mg, 2 mmol) was dissolved in 20 mL of methanol, 5 mL of water was added, and then sodium hydroxide (400 mg, 10 mmol) was added, and the reaction was stirred at room temperature for 12 hours. The reaction solution was concentrated under reduced pressure to remove the methanol, 30 mL of water was added to the resulting residue, and the mixture was extracted with ethyl acetate (50 mL). 6 M hydrochloric acid was added dropwise to the aqueous phase until the pH was 5-6, and the mixture was extracted with a mixed solvent of dichloromethane and methanol (V/V = 5:1) (60 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain the title compound 33f (600 mg, brown oil), which was directly used for the next step without purification.

Step 6

(1S,4s)-4-((R)-6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)-N-(3-methoxyphenyl)cyclohexylcarboxamide 33

The crude product 33f (60 mg, 0.2 mmol) was dissolved in 2 mL of N,N-dimethylformamide, and 3-methoxyaniline (25 mg, 0.2 mmol), 1-hydroxybenzotriazole (32 mg, 0.24 mmol), 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (46 mg, 0.24 mmol), and N,N-diisopropylethylamine (129 mg, 1 mmol) were added. The reaction was stirred at room temperature for 48 hours. The reaction solution was purified by HPLC to obtain the title compound 33 (13 mg, white solid) in a yield of 16.25%.

MS m/z(ESI):406.4[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ9.72(s, 1H), 7.97(s, 1H), 7.44-7.49(m, 2H), 7.26-7.3(m, 1H), 7.21(s, 1H), 7.11-7.18(m, 2H), 7.05-7.10(m, 1H), 6.56-6.59( m, 1H), 5.60 (s, 1H), 3.69 (s, 3H), 2.10-2.36 (m, 3H), 1.69-1.92 (m, 3H), 1.44-1.58 (m, 1H), 1.28-1.43 (m, 2H), 0.55-0.68 (m, 1H).

Example 34

6-Fluoro-5-(1-(2-fluoro-4-(1-methyl-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole

first step

6-Fluoro-5-(1-(2-fluoro-4-nitrophenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 34a

1 g (1.7 g, 3.51 mmol) of the crude product was dissolved in 20 mL of dimethyl sulfoxide, and 1,2-difluoro-4-nitrobenzene (0.557 g, 3.51 mmol) and triethylamine (1.42 g, 14.04 mmol) were added. The mixture was stirred at room temperature for 2.5 hours. 150 mL of ethyl acetate was added to the reaction solution, which was washed with water (100 mL x 2). The organic phase was concentrated under reduced pressure and the residue was purified by silica gel column chromatography with eluent System A to obtain the title compound 34a (970 mg, yellow solid) in a 53.8% yield.

Step 2

3-Fluoro-4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)aniline 34b

34a (970 mg, 2.45 mmol) was dissolved in 20 mL of methanol, and 200 mg of 10% palladium on carbon was added. The reaction system was replaced with hydrogen three times and stirred at room temperature for 12 hours. The reaction solution was concentrated under reduced pressure to obtain the crude title compound 34b (920 mg, yellow solid), which was directly used in the next step without purification.

Step 3

5-(1-(4-Bromo-2-fluorophenyl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole 34c

Crude product 34b (620 mg, 1.692 mmol) was dissolved in 40% hydrobromic acid solution and cooled to 0-5°C. 1 mL of pre-prepared sodium nitrite solution (128 mg, 1.86 mmol) was added dropwise. After the addition was complete, the mixture was reacted at 0°C for 40 minutes. The reaction solution was poured into 0°C hydrobromic acid solution and heated to 60°C for 2 hours. 2N sodium hydroxide solution was added dropwise to the reaction solution until the pH reached 8-9. The mixture was extracted with ethyl acetate (100 mL x 2). The combined organic phases were washed sequentially with water (100 mL) and saturated sodium chloride solution (100 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A to afford the title compound 34c (450 mg, white solid) in a 61.8% yield.

Step 4

6-Fluoro-5-(1-(2-fluoro-4-(1-methyl-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 34

34c (200 mg, 0.465 mmol) was dissolved in 11 mL of a mixed solvent of ethylene glycol dimethyl ether and water (V:V=10:1). 1-Methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole (145 mg, 0.697 mmol), tetrakistriphenylphosphine palladium (54 mg, 0.0465 mmol), and sodium carbonate (99 mg, 0.93 mmol) were added. The mixture was heated to 80°C and stirred for 48 hours. The reaction solution was cooled to room temperature, concentrated under reduced pressure, and the resulting residue was purified by high performance liquid chromatography to give the title compound 34 (160 mg, brown solid) in a 35% yield.

MS m/z(ESI):432.4[M+1]

¹ HNMR (400MHz, DMSO-d ₆ )δ8.06(s, 2H), 7.80(s, 1H), 7.45-7.55(m, 2H), 7.22-7.40(m, 3H), 7.16-7.21(m, 1H), 6.97(t, 1H), 5.71(s, 1H), 3.83(s, 3H), 3.20-3.45(m, 2H), 2.68(t, 1H), 2.56(t, 1H), 2.35(t, 1H), 1.65-1.85(m, 2H), 1.14-1.25(m, 1H), 0.85-1.00(m, 1H).

Example 35

6-Fluoro-5-(1-(3-fluoro-4-(1-methyl-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

4-(4-Bromo-2-fluorophenyl)-1-methyl-1H-pyrazole 35b

4-Fluoro-2-bromo-1-iodobenzene 35a (301 mg, 1 mmol) was dissolved in 3 mL of dimethyl sulfoxide, and 1-methyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-1H-pyrazole (139 mg, 0.67 mmol), [1,1′-bis(diphenylphosphino)ferrocene]palladium dichloride (49 mg, 0.067 mmol), potassium acetate (66 mg, 0.67 mmol) and cesium carbonate (650 mg, 2.01 mmol) were added. The temperature was raised to 80°C and the reaction was stirred for 1 hour. The reaction solution was cooled to room temperature, 350 mL of water was added, and the mixture was extracted with ethyl acetate (60 mL × 3). The organic phases were combined, washed with water (120 mL × 2), washed with saturated sodium chloride solution (60 mL), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give the crude title compound 35b (170 mg, tan solid). The product was directly used for the next step without purification.

Step 2

6-Fluoro-5-(1-(3-fluoro-4-(1-methyl-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate 35

The crude product 1 g (200 mg, 0.41 mmol) was dissolved in 10 mL of toluene, and crude product 35b (170 mg, 0.67 mmol), tris(dibenzylideneacetone)dipalladium (61 mg, 0.067 mmol), (±)-2,2′-bis-(diphenylphosphino)-1,1′-binaphthyl (442 mg, 0.067 mmol), and sodium tert-butoxide (257 mg, 2.68 mmol) were added. The mixture was stirred at 80°C for 12 hours. The reaction solution was cooled to room temperature, 30 mL of water was added, and the mixture was extracted with dichloromethane (30 mL x 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A to afford the title compound 35 (5 mg, white solid) in a 2.8% yield.

MS m/z(ESI):432.3[M+1]

Example 36

4-(5-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-1,3,4-thiadiazol-2-yl)pyridin-2-amine trifluoroacetate

first step

2-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-1,3,4-thiadiazole 36a

1 g (4.85 g, 10.0 mmol) of the crude product, 2-bromo-1,3,4-thiadiazole (1.65 g, 10.0 mmol), and triethylamine (10.1 g, 100.0 mmol) were added to 20 mL of dimethyl sulfoxide and the mixture was heated to 120°C in a sealed tube and reacted for 12 hours. The reaction solution was cooled to room temperature, 200 mL of dichloromethane was added, and the mixture was washed with water (200 mL x 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography with eluent System A to obtain the title compound 36a (1.09 g, light brown solid) in a 31.9% yield.

Step 2

2-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-5-(2-fluoropyridin-4-yl)-1,3,4-thiadiazole 36b

36a (800 mg, 2.34 mmol), 4-bromo-2-fluoropyridine (824 mg, 4.68 mmol), palladium acetate (76 mg, 0.334 mmol), tri(tert-butyl)phosphine (1.35 g, 0.668 mmol), and cesium carbonate (1.52 g, 4.68 mmol) were added to 10 mL of N,N-dimethylformamide and stirred at 150°C for 4 hours. The reaction solution was cooled to room temperature, and 150 mL of dichloromethane was added. The product was washed with water (100 mL x 3), dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A to afford the title compound 36b (260 mg, brown solid) in a 25.5% yield.

Step 3

4-(5-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-1,3,4-thiadiazol-2-yl)-N-(4-methoxyphenyl)pyridin-2-amine 36c

36b (260 mg, 0.60 mmol) was dissolved in 3 mL of dimethyl sulfoxide, and (4-methoxyphenyl)methanamine (823 mg, 6.0 mmol) was added. The mixture was heated to 130°C and stirred for 4 hours. The reaction solution was cooled to room temperature, and 100 mL of dichloromethane was added. The mixture was washed with water (100 mL x 3), dried over anhydrous sodium sulfate, and filtered. The filtrate was concentrated under reduced pressure and the resulting residue was purified by silica gel column chromatography with eluent System A to give the title compound 36c (130 mg, light brown solid) in a yield of 39.1%.

Step 4

4-(5-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)-1,3,4-thiadiazol-2-yl)pyridin-2-amine trifluoroacetate 36

36c (130 mg, 0.235 mmol) was dissolved in 5 mL of trifluoroacetic acid and stirred at 60°C for 4 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and purified by HPLC to afford the title compound 36 (30 mg, light brown solid) in a 23.4% yield.

MS m/z (LC-MS): 434.3[M+1]

Example 37

6-Fluoro-5-(1-(5-((1-(methylsulfonyl)piperidin-3-yl)oxy)pyrimidin-2-yl)piperidin-4-yl-5H-imidazo[5,1-a]isoindole trifluoroacetate

first step

5-(1-(5-(Benzyloxy)pyrimidin-2-yl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole 37a

The crude product 17a (1.5 g, 5.83 mmol) and 5-(benzyloxy)-2-chloropyrimidine 37a (1.29 g, 5.83 mmol) were dissolved in 10 mL of N,N-dimethylacetamide, and N,N-diisopropylethylamine (3.76 g, 29.1 mmol) was added. The mixture was microwaved at 150°C for 1 hour. The reaction mixture was cooled to room temperature and filtered. 100 mL of ethyl acetate and 50 mL of water were added to the filtrate. The layers were separated, and the organic phase was washed with water and saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A to afford the title compound 37a (600 mg, brown oil) in a 40% yield.

Step 2

2-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)pyrimidin-5-ol 37c

37a (600 mg, 1.36 mmol) was dissolved in 15 mL of methanol, and 120 mg of palladium on carbon (10%) was added. The atmosphere was replaced with hydrogen three times and the reaction was stirred for 3 hours. The reaction mixture was filtered to remove the palladium on carbon, and the filtrate was concentrated under reduced pressure to obtain the crude title compound 37c (455 mg, yellow solid). This product was directly used in the next reaction without purification.

Step 3

Tert-butyl 3-((2-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)pyrimidin-5-yl)oxy)piperidine-1-carboxylate 37d

Crude product 37c (200 mg, 0.57 mmol), tert-butyl 3-hydroxypiperidine-1-carboxylate (114 mg, 0.57 mmol), diisopropyl azodicarboxylate (162 mg, 0.856 mmol), and triphenylphosphine (224 mg, 0.856 mmol) were dissolved in 5 mL of tetrahydrofuran and stirred at room temperature for 12 hours. The reaction solution was concentrated under reduced pressure and the residue was purified by silica gel column chromatography using eluent System A to afford the title compound 37d (100 mg, light brown solid) in a 33.1% yield.

Step 4

6-Fluoro-5-(1-(5-((1-(methylsulfonyl)piperidin-3-yl)oxy)pyrimidin-2-yl)piperidin-4-yl-5H-imidazo[5,1-a]isoindole trifluoroacetate 37

Dissolve 37d (38 mg, 0.087 mmol) in 5 mL of dichloromethane, add 0.25 mL of trifluoroacetic acid and 11 drops of methanesulfonyl chloride, and stir at room temperature for 12 hours. The reaction solution is concentrated under reduced pressure, and the resulting residue is purified by HPLC to afford the title compound 37 (14 mg, white solid) in a 20% yield.

MS m/z(ESI):513.3[M+1]

Example 38

6-Fluoro-5-(1-(4-((tetrahydrofuran-3-yl)oxy)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole

first step

4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenol 38b

1 g (5.82 g, 12 mmol) and 1,4-cyclohexanedione 38a (1.61 g, 14.4 mmol) were dissolved in 40 mL of ethanol. Triethylamine (2.424 g, 24 mmol) and 200 mg of palladium on carbon (10%) were added, and the mixture was heated to 85°C and stirred for 12 hours. The reaction solution was cooled to room temperature, water was added, and extraction was performed with dichloromethane (50 mL x 3). The organic phases were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to obtain the crude title product 38b (4.19 g, brown solid), which was directly used in the next reaction without purification.

Step 2

6-Fluoro-5-(1-(4-((tetrahydrofuran-3-yl)oxy)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 38

The crude product 38b (349 mg, 1 mmol) was dissolved in 5 mL of ethanol, and tetrahydrofuran-3-methanesulfonic acid 38c (333 mg, 2 mmol, prepared using the method disclosed in patent application "WO2014049133") and potassium carbonate (420 mg, 3 mmol) were added. The reaction was microwaved at 125°C for 1 hour. The reaction solution was cooled to room temperature, filtered, and the filtrate was concentrated under reduced pressure. Water was added to the resulting residue, and the mixture was extracted with dichloromethane (20 mL × 3). The organic phases were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by high-performance liquid chromatography to obtain the title compound 38 (20 mg, light yellow solid) in a yield of 4.8%.

MS m/z(ESI):420.5[M+1]

Example 39

6-Fluoro-5-(1-(4-((1-(methylsulfonyl)piperidin-4-yl)oxy)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole

first step

4-(4-iodophenoxy)-1-(methylsulfonyl)piperidine 39b

4-(4-iodophenoxy)piperidine 39a (600 mg, 2 mmol, prepared using the method disclosed in patent application WO2004089373) was dissolved in 15 mL of dichloromethane. Triethylamine (404 mg, 4 mmol) was added, followed by dropwise addition of methanesulfonyl chloride (273.6 mg, 2.4 mmol). The mixture was stirred at room temperature for 1 hour. Water was added to the reaction solution, which was extracted with ethyl acetate (20 mL x 3). The organic phases were combined, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to afford the crude title compound 39b (100 mg, light yellow solid). This product was directly used in the next step without purification.

Step 2

6-Fluoro-5-(1-(4-((1-(methylsulfonyl)piperidin-4-yl)oxy)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 39

Crude product 39b (114 mg, 0.3 mmol) and 1g (122 mg, 0.25 mmol) were dissolved in 5 mL of toluene. Tris(dibenzylideneacetone)dipalladium (22.9 mg, 0.025 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (12 mg, 0.025 mmol), and sodium tert-butoxide (36 mg, 0.375 mmol) were added, and the mixture was microwaved at 156° C. for 40 minutes. The reaction solution was cooled to room temperature, and the resulting residue was purified by high performance liquid chromatography to obtain the title compound 39 (10 mg, light yellow solid) in a yield of 7.8%.

MS m/z(ESI):511.6[M+1]

Examples 40, 41

(R)-2-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)ethanol 40

(S)-2-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)ethanol 41

first step

6-Fluoro-5-(1-(4-(1-(2-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 40b

4-(4-Bromophenyl)-1-(2-((tetrahydro-2H-pyran-2-yl)oxy)ethyl)-1H-pyrazole 40a (14.8 g, 42 mmol) and 6-fluoro-5-(piperidin-4-yl)-5H-imidazo[5,1-a]isoindole 17a (13.9 g, 42 mmol) were added to 300 mL of N,N-dimethylformamide. Tri-tert-butylphosphine tetrafluoroborate (1.863 g, 64.5 mmol) and potassium phosphate (35 g, 168 mmol) were then added, and the atmosphere was replaced with argon three times. Tris(dibenzylideneacetone)dipalladium (2.92 g, 3.19 mmol) was added, and the atmosphere was replaced with argon once. The reaction mixture was heated to 110°C and stirred for 2 hours. After the reaction, the reaction solution was filtered and the filtrate was concentrated under reduced pressure to remove N,N-dimethylformamide. The resulting residue was purified by silica gel column chromatography using eluent System A to obtain compound 40b (6.38 g, gray oil) in a yield of 29%.

MS m/z (LC-MS): 528.3[M+1]

Step 2

2-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)ethanol 40c

Compound 40b (9 g, 17.1 mmol) was dissolved in 100 mL of methanol, and 5.7 mL of concentrated hydrochloric acid (12 M) was added. The reaction mixture was heated to 45°C and stirred for 1 hour. After completion of the reaction, the reaction mixture was cooled to room temperature and adjusted to pH 8 by adding saturated sodium carbonate. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography using eluent System A to obtain compound 40c (5.2 g, yellow solid) in a 65% yield.

MS m/z (LC-MS): 444.4[M+1]

Step 3

(R)-2-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)ethanol 40

(S)-2-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)ethanol 41

Compound 40c (1.4 g, 3.16 mmol) was subjected to chiral preparation (separation conditions: chiral preparative column Superchiral S-AS (Chiralway), 2 cm ID*25 cm Length, 5 um; mobile phase: CO2/MeOH/DEA=60/ ₄₀ /0.05 (v/v), flow rate: 50 mL/min), and the corresponding fractions were collected and concentrated under reduced pressure to give compound 40 (630 mg, yellow solid) and compound 41 (652 mg, yellow solid).

40:

MS m/z (ESI): 444.5 [M+1];

Chiral HPLC analysis: retention time 3.064 minutes, chiral purity: 97.79% (chromatographic column: Superchiral S-AS (Chiralway), 0.46 cm ID*25 cm Length, 5 μm; mobile phase: CO2/MeOH/DEA=60/40/0.05 (v/v)

¹ H NMR (400 MHz, DMSO-d ₆ )δ9.97(s, 2H), 7.73(s, 1H), 7.44-7.51(m, 2H), 7.36(d, 2H), 7.22(s, 1H) ,7.10-7.19(m,1H),6.87(d,2H),5.70(d,1H),4.91(t,1H),4.12(t,2H), 3.70-3.79(m, 3H), 3.62(d, 1H), 2.62-2.73(m, 1H), 2.52-2.58(m, 1H), 2. 31-2.43(m, 1H), 1.55-1.83(m, 2H), 1.13-1.23(m, 1H), 0.82-0.96(m, 1H).

41:

MS m/z (ESI): 444.5 [M+1];

Chiral HPLC analysis: Retention time 4.280 minutes, chiral purity: 99.52%. (Chromatographic column: Superchiral S-AS (Chiralway), 0.46 cm ID x 25 cm Length, 5 μm; Mobile phase: CO2/MeOH/DEA = 60/40/0.05 (v/v))

¹ H NMR (400 MHz, DMSO-d ₆ )δ9.97(s, 2H), 7.73(s, 1H), 7.44-7.51(m, 2H), 7.36(d, 2H), 7.22(s, 1H) ,7.10-7.19(m,1H),6.87(d,2H),5.70(d,1H),4.91(t,1H),4.12(t,2H), 3.70-3.79(m, 3H), 3.62(d, 1H), 2.62-2.73(m, 1H), 2.52-2.58(m, 1H), 2. 31-2.43(m, 1H), 1.55-1.83(m, 2H), 1.13-1.23(m, 1H), 0.82-0.96(m, 1H).

Example 42

6-Fluoro-5-(1-(4-(1-(2-methoxyethyl)-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

9a (100 mg, 0.242 mmol) and 1-(2-methoxyethyl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrazole 42a (91 mg, 0.363 mmol, prepared using the method disclosed in patent application "WO2014015088") were dissolved in 5 mL of n-butanol. Tris(dibenzylideneacetone)dipalladium (13 mg, 0.0142 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (28 mg, 0.0581 mmol), and potassium phosphate (154 mg, 0.726 mmol) were added. The mixture was heated to 100°C and stirred for 2 hours. The reaction solution was cooled to room temperature, and the resulting residue was purified by high-performance liquid chromatography to obtain the title compound 42 (8 mg, white solid) in a 5.8% yield.

MS m/z(ESI):458.4[M+1]

Example 43

N-(2-(4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)ethyl)acetamide trifluoroacetate

first step

tert-Butyl (2-(4-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)ethyl)carbamate 43b

9a (100 mg, 0.243 mmol) was dissolved in 3 mL of n-butanol, and tert-butyl (2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-1H-pyrazol-1-yl)ethyl)carbamate 43a (123 mg, 0.364 mmol, prepared by the method disclosed in patent application "CN103087050"), tris(dibenzylideneacetone)dipalladium (11 mg, 0.0122 mmol), 2-dicyclohexylphosphine-2′,4′,6′-triisopropylbiphenyl (23 mg, 0.0486 mmol) and potassium phosphate (103 mg, 0.486 mmol) were added. The temperature was raised to 100°C and the reaction was stirred for 2 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and the resulting residue was purified by silica gel column chromatography using eluent System A to afford the title compound 43b (50 mg, yellow solid) in a 38% yield.

Step 2

2-(4-(4-(6-Fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)ethanamine 43c

43b (50 mg, 0.092 mmol) was dissolved in 3 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid was added. The mixture was stirred at room temperature for 2 hours. The reaction solution was concentrated under reduced pressure to obtain the crude title compound 43c (60 mg, brown oil), which was directly used in the next step without purification.

Step 3

N-(2-(4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)phenyl)-1H-pyrazol-1-yl)ethyl)acetamide trifluoroacetate 43

Crude product 43c (60 mg, 0.092 mmol) was dissolved in dichloromethane, and acetyl chloride (14 mg, 0.184 mmol) and triethylamine (28 mg, 0.276 mmol) were added. The mixture was stirred at room temperature for 3 hours. The reaction solution was concentrated under reduced pressure and the resulting residue was purified by HPLC to afford the title compound 43 (7 mg, off-white solid) in a 13% yield.

MS m/z (ESI): 485.5 [M+1];

¹ H NMR (400 MHz, DMSO-d ₆ )δ9.51 (s, 1H), 8.00-8.20 (m, 3H), 7.75-7.95 (m, 2H), 7.64-7.73 (m, 1H), 7.40-7.60 (m, 3H), 7.00-7.18 (m, 2H), 6.16 (s, 1H), 4.14 (t, 2H), 3 .72(d,1H),3.62(d,1H),3.41-3.46(m,2H),2.70-3.00(m,2H),2.55- 2.70(m, 1H), 1.60-1.90(m, 5H), 1.30-1.40(m, 1H), 1.10-1.20(m, 1H).

Example 44

6-Fluoro-5-(1-(4-(1-((R)-tetrahydrofuran-3-yl)-1H-pyrazol-4-yl)phenyl)piperidin-4-yl)-5H-imidazo[5,1-a]isoindole trifluoroacetate

9a (103 mg, 0.25 mmol) was dissolved in 3 mL of n-butanol, and (S)-1-(tetrahydrofuran-3-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolane-2-yl)-1H-pyrazole 44a (99 mg, 0.375 mmol, prepared by the method disclosed in patent application "US20080167287"), tris(dibenzylideneacetone)dipalladium (17 mg, 0.01875 mmol), 2-dicyclohexylphosphine-2′,4′,6′-triisopropylbiphenyl (36 mg, 0.075 mmol) and potassium phosphate (106 mg, 0.5 mmol) were added, and the temperature was raised to 100 ° C. and the reaction was stirred for 2 hours. The reaction mixture was cooled to room temperature and filtered through Celite to remove insoluble matter. The filtrate was concentrated under reduced pressure and the resulting residue was purified by HPLC to give the title compound 44 (30 mg, white solid) in a yield of 40.3%.

MS m/z (ESI): 470.5 [M+1];

¹ H NMR (400MHz, DMSO-d ₆ )δ9.49 (s, 1H), 8.10 (s, 1H), 8.06 (s, 1H), 7.8 (s, 1H), 7.76-7.78 (m, 1H), 7.41-7.45 (m, 3H), 6.97 (d, 2H), 6.14(s, 1H), 4.98-5.00(m, 1H), 3.97-4.02(m, 2H), 3.89-3.92(m, 1H), 3.80-3.86(m, 1H), 3.73-3 .76(m, 1H), 3.63-3.66(m, 1H), 2.75-2.85(m, 1H), 2.62-2.75(m, 1H), 2.53-2.59(m, 1H), 2.29-2.41(m , 2H), 1.82-1.85(m, 1H), 1.62-1.64(m, 1H), 1.31-1.34(m, 1H), 1.07-1.13(m, 1H), 0.85-0.87(m, 1H).

Example 45

2-Fluoro-4-(4-(6-fluoro-5H-imidazo[5,1-a]isoindol-5-yl)piperidin-1-yl)benzonitrile

17a (500 mg, 1.94 mmol), 2,4-difluorobenzonitrile 45a (270 mg, 1.94 mmol), and triethylamine (216 mg, 2.14 mmol) were dissolved in 5 mL of dimethyl sulfoxide and stirred at 80°C for 12 hours. The reaction solution was cooled to room temperature, and the resulting residue was purified by HPLC to obtain the title compound 45 (190 mg, light brown solid) in a 26% yield.

MS m/z(ESI):377.1[M+1]

¹ H NMR (400MHz, DMSO-d ₆ )δ8.1 (s, 1H), 7.74 (dd, 1H), 7.46-7.49 (m, 2H), 7.24 (s, 1H), 7.17-7.12 (m, 1H), 6.96 (dd, 1H), 6.88 (td, 1H), 5.72 (s, 1H), 3.49-3.5 9(m, 2H), 2.88(t, 1H), 2.73(t, 1H), 2.37-3.43(m, 1H), 1.76-1.84(m, 1H), 1.61-1.71(m, 1H), 1.20-1.29(m, 1H), 0.91-1.01(m, 1H).

Example 46

5-(1-(3,4-difluorophenyl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole

first step

6-Fluoro-5-(piperidin-4-yl)-5H-imidazo[5,1-a]isoindole hydrochloride 46a

1f (17.7 g, 49.6 mmol) was dissolved in 180 mL of a mixture of dichloromethane and 1,4-dioxane (V/V = 5:1). After cooling in an ice bath, 41.2 mL of concentrated hydrochloric acid was added dropwise. The mixture was stirred at room temperature for 2 hours. The reaction solution was concentrated under reduced pressure to obtain the crude title compound 46a (16.37 g, white solid), which was directly used in the next step without purification.

Step 2

5-(1-(3,4-difluorophenyl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole 46

Crude product 46a (165 mg, 0.5 mmol) and 4-bromo-1,2-difluorobenzene 46b (116 mg, 0.6 mmol) were dissolved in 6 mL of a mixed solvent of toluene and tert-butanol (V/V = 5:1). Palladium acetate (11.22 mg, 0.05 mmol), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (24 mg, 0.05 mmol), and sodium tert-butoxide (200 mg, 2 mmol) were added, and the mixture was microwaved at 160°C for 0.5 hour. The reaction solution was cooled to room temperature, concentrated under reduced pressure, and the resulting residue was purified by high performance liquid chromatography to obtain the title compound 46 (10 mg, white solid) in a 5.4% yield.

MS m/z(ESI):370.4[M+1]

¹ H NMR (400MHz, CDCl ₃ )δ7.72(s, 1H), 7.34(dd, 1H), 7.22-7.26(m, 2H), 6.96(s, 2H), 6.54-6.65(m, 2H), 5.57(dd, 1H), 3.49-3.59(m, 2H), 2.61 (t, 1H), 2.58 (t, 1H), 2.54 (m, 1H), 1.76-1.84 (m, 1H), 1.61-1.71 (m, 1H), 1.20-1.29 (m, 1H), 0.91-1.01 (m, 1H).

Example 47

5-(1-(5-chloropyridin-2-yl)piperidin-4-yl)-6-fluoro-5H-imidazo[5,1-a]isoindole

Crude product 46a (500 mg, 1.52 mmol) and 2,5-dichloropyridine 47a (292 mg, 1.97 mmol) were dissolved in 10 mL of dimethyl sulfoxide. N,N-diisopropylethylamine (980 mg, 7.6 mmol) was added and the mixture was microwaved at 140°C for 3.5 hours. The reaction mixture was cooled to room temperature and the resulting residue was purified by HPLC to afford the title compound 47 (105 mg, orange-yellow solid) in an 18.7% yield.

MS m/z(ESI):369.8[M+1]

¹ H NMR (400 MHz, DMSO-d ₆ ): δ8.05-8.04(d, 1H), 7.97(s, 1H), 7.54-7.51(dd, 1H), 7.48-7.46(m, 2H), 7.23(s, 1H), 7.17-7.12(m, 1H), 6.81-6.79(d, 1H), 6.70-6.69(d ,1H),4.39-4.36(d,1H),4.22-4.19(d,1H),2.85-2.79(m,1H),2.72- 2.66(m, 1H), 1.54-1.45(m, 1H), 1.30-1.24(m, 2H), 1.97-1.14(m, 2H).

Biological evaluation

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

Test Example 1: Determination of the inhibitory activity of the compounds of the present invention on human IDO1 protease

The in vitro protease activity of human IDO1 was tested by the following method.

This method is used to determine the inhibitory effect of the compounds of the present invention on the activity of human IDO1 protease.

1. Experimental Materials and Instruments

1. Microplate reader (Synergy HT, BIOTEK)

2. Tryptophan (T0254-5G, Sigma-Aldrich)

3. Catalase from bovine liver (C1345-1G, Sigma-Aldrich)

4. Methylene blue (M9140-25G, Sigma-Aldrich)

5. Sodium L-ascorbate (A7631-25G, Sigma-Aldrich)

6. 4-(Dimethylamino)benzaldehyde (D2004-25G, Sigma-Aldrich)

7. Trichloroacetic acid (T9159-100G, Sigma-Aldrich)

8. Human IDO1 gene (SC126221, Origene)

2. Experimental steps

Homemade IDO1 protease

The human IDO1 gene was transferred into the Pet30a plasmid through gene cloning technology and then transferred into competent Escherichia coli rosseta; the culture was amplified in liquid LB (Luria-Bertani) medium [prepared according to the "Molecular Cloning Laboratory Manual" (by J. Sambrook and D.W. Russell)], and the cells were collected, ultrasonically disrupted, and passed through a column to obtain purified IDO1 protease by elution.

Compound testing experiments:

Dilute 24 μl of enzyme (IDO1) 100-fold to 2400 μl with 50 mM KPB, resulting in an enzyme solution at a concentration of 2.6 ng/μl. Add 24 μl of enzyme solution to each well of a 96-well reaction plate (AXYGEN, PCR-96-FLT-C) (hereinafter referred to as the reaction plate). Add 24 μl of KPB to a blank well. [Preparation of KPB buffer (50 mM): Use an analytical balance to weigh 6.805 _g of _KH₂PO₄ into a 1000-ml beaker. Use a graduated cylinder to add deionized water to 900 ml. Adjust the pH to 6.5 with 1 M KOH. Pour the solution into a 1-L graduated cylinder and make up to 1 L. Store at 4°C.] Add 1 μl of compound or DMSO to the corresponding reaction wells of the reaction plate. Prepare Solution A: Add 200 μl of 500 mM sodium ascorbate to 1050 μl of KPB and mix on a turbine mixer at maximum speed for 3 seconds. Solution B: Add 100 μl of 10 mM tryptophan to 100 μl of 100,000 units/ml catalase, 5 μl of 10 mM methylene blue, and finally 1050 μl of KPB. Mix thoroughly on a turbo mixer at maximum speed for 3 seconds. Add 1200 μl of Solution A and 1200 μl of Solution B to the reaction plate at maximum speed for 3 seconds. Add 24 μl of this mixture to each well of the reaction plate. Centrifuge the plate at maximum speed for 15 seconds to allow the reaction mixture to pool at the bottom. Mix thoroughly on a shaker for 30 seconds and incubate at 37°C in a constant temperature incubator for 1 hour. Add 10 μl of 30% (w/v) trichloroacetic acid to each well of the plate and incubate at 65°C in a turbo mixer for 15 minutes. Centrifuge the plate at 4700 RPM at room temperature for 5 minutes. Use a dispenser to transfer 40 μl of supernatant from the reaction plate to a corresponding 96-well assay plate (Corning, #3599). Add 40 μl of a 2% (w/v) 4-(dimethylamino)benzaldehyde/glacial acetic acid solution to each well and mix on a shaker at maximum speed for 1 minute. After incubation at room temperature for 2 minutes, read the absorbance at 480 nm on a Synergy HT (BIOTEK).

The inhibitory activity of the compounds of the present invention against human IDO1 protease was determined by the above test, and the measured IC ₅₀ values are shown in Table 1.

Table 1 IC ₅₀ of the compounds of the present invention against human IDO1 protease activity

Conclusion: The compounds of the present invention have a significant inhibitory effect on the activity of human IDO1 protease.

Test Example 2: Determination of the inhibitory activity of the compounds of the present invention on human TDO protease

The in vitro activity of human TDO protease was tested by the following method.

This method is used to determine the inhibitory effect of the compounds of the present invention on the activity of human TDO protease.

1. Experimental Materials and Instruments

1. Microplate reader (Synergy HT, BIOTEK)

2. Tryptophan (T0254-5G, Sigma-Aldrich)

3. Catalase from bovine liver (C1345-1G, Sigma-Aldrich)

4. Methylene blue (M9140-25G, Sigma-Aldrich)

5. Sodium L-ascorbate (A7631-25G, Sigma-Aldrich)

6. 4-(Dimethylamino)benzaldehyde (D2004-25G, Sigma-Aldrich)

7. Trichloroacetic acid (T9159-100G, Sigma-Aldrich)

8. Human TDO (U32989.1, Suzhou Jinweizhi Biotechnology Co., Ltd.)

9. Rosseta (CW0811A, Beijing Kangwei Century Biotechnology Co., Ltd.)

10. Turbine mixer (6776, Corning)

11. Mini Plate Centrifuge (Mini-P25, ABSON life science equipment)

2. Experimental steps

Homemade TDO protease

The constructed plasmid containing the human TDO gene was transformed into competent Escherichia coli Rosseta; the culture was amplified in liquid LB (Luria-Bertani) medium [prepared according to the "Molecular Cloning Laboratory Manual" (written by J. Sambrook and D.W. Russell)], and the cells were collected, ultrasonically disrupted, and eluted through a column to obtain purified TDO1 protease.

Compound testing experiments:

Dilute 24 μl of enzyme (TDO) 100-fold with 50 mM KPB to 2400 μl, for a concentration of 2.6 ng/μl. Add 24 μl of enzyme solution to each well of a 96-well reaction plate (AXYGEN, PCR-96-FLT-C) (hereinafter referred to as the reaction plate). Add 24 μl of KPB buffer to a blank well. [Preparation of KPB buffer (50 mM): Use an analytical balance to weigh 6.805 _g of _KH₂PO₄ into a 1000 ml beaker. Use a graduated cylinder to add deionized water to 900 ml. Adjust the pH to 6.5 with 1 M KOH. Pour the solution into a 1 L graduated cylinder and make up to 1 L. Store at 4°C.] Add 1 μl of compound or DMSO to the corresponding reaction wells of the reaction plate. Prepare Solution A: Add 200 μl of 500 mM sodium L-ascorbate to 1050 μl of KPB. Mix at maximum speed on a turbine mixer for 3 seconds. Solution B: Add 100 μl of 10 mM tryptophan to 100 μl of 100,000 units/ml catalase, 5 μl of 10 mM methylene blue, and finally 1050 μl of KPB. Mix thoroughly on a turbo mixer at maximum speed for 3 seconds. Add 1200 μl of Solution A and 1200 μl of Solution B to the reaction plate at maximum speed for 3 seconds. Add 24 μl of this mixture to each well of the reaction plate. Centrifuge the plate at maximum speed for 15 seconds to allow the reaction mixture to pool at the bottom. Mix thoroughly on a shaker for 30 seconds and incubate in a constant temperature incubator at 37°C for 1 hour. Add 10 μl of 30% (w/v) trichloroacetic acid to each well of the plate and incubate at 65°C for 15 minutes. Centrifuge the plate at 4700 RPM at room temperature for 5 minutes. Use a dispenser to transfer 40 μl of supernatant from the reaction plate to a corresponding 96-well assay plate (Corning, #3599). Add 40 μl of a 2% (w/v) 4-(dimethylamino)benzaldehyde/glacial acetic acid solution to each well and mix on a shaker at maximum speed for 1 minute. After incubation at room temperature for 2 minutes, read the absorbance at 480 nm on a Synergy HT Reader.

The inhibitory activity of the compounds of the present invention against human TDO protease was determined by the above test, and the measured IC ₅₀ values are shown in Table 2.

Table 2 IC ₅₀ of the compounds of the present invention against human TDO protease activity

Conclusion: The compounds of the present invention have a significant inhibitory effect on the activity of human TDO protease.

Test Example 3: Determination of the inhibitory activity of the compounds of the present invention on IDO protease in HeLa cells

The IDO protease activity in HeLa cells was tested by the following method.

This method is used to determine the inhibitory effect of the compounds of the present invention on IDO protease activity in HeLa cells. (Note: HeLa cells express indoleamine 2,3-dioxygenase (IDO) under the induction of interferon gamma (INF-γ))

1. Experimental Materials and Instruments

1. Microplate reader (Synergy HT, BIOTEK)

2. Tryptophan (T0254-5G, Sigma-Aldrich)

3, 4-(Dimethylamino)benzaldehyde (D2004-25G, Sigma-Aldrich)

4. Trichloroacetic acid (T9159-100G, Sigma-Aldrich)

5. HeLa cell line (CCL-2, ATCC)

2. Experimental steps

HeLa cell suspensions were prepared in fresh cell culture medium and plated at 10,000 cells/well in 100 μl of culture medium in a 96-well cell culture plate. The cells were incubated at 37°C in 5% CO2 for 24 hours. The supernatant was removed, and 90 μl of serum-free DMEM high-glucose medium was added to each well. Then, 10 μl of each compound (final concentrations: 10,000, 1,000, 100, 10, 1, and 0.1 nM) prepared in medium containing INF-γ and tryptophan were added to each well. The cells were incubated at 37°C in 5% CO2 for 48 hours. 80 μl of the supernatant from the 96-well cell culture plate was removed and transferred to a 96-well round-bottom plate. 16 μl of 30% (w/v) trichloroacetic acid was added to each well, and the plates were incubated at 65°C in an incubator for 25 minutes. The plates were centrifuged at 4700 RPM for 5 minutes. Use a dispenser to transfer 50 μl of supernatant from the reaction plate to a 96-well flat-bottom transparent plate. Add 50 μl of a 2% (w/v) 4-(dimethylamino)benzaldehyde/glacial acetic acid solution to each well and mix on a shaker for 1 minute. Incubate at room temperature for 2 minutes, and read the absorbance at 480 nm on a Synergy HT Reader.

The inhibitory activity of the compounds of the present invention on IDO protease in HeLa cells was determined by the above test, and the measured IC ₅₀ values are shown in Table 3.

Table 3 IC ₅₀ of the compounds of the present invention against IDO protease activity in HeLa cells

Example No. 3 167.1 4 187.6 5 353.1 8 252.9 9 100.9 11 477.3 12 70.57 15 199.2 16 260.3 18 197.8 twenty one 477 twenty two 129.8 twenty three 438.9 28 85.16 29 292.4 30 333.8 32 225 33 47.23 34 29.34 35 69.06 36 27.76 37 113.8 38 150.7 39 198.3 41 47.7 42 253.8 43 171.1 44 174.7 45 231 46 69.1 47 88.3

Conclusion: The compounds of the present invention have a significant inhibitory effect on IDO protease activity in HeLa cells.

Pharmacokinetic evaluation

Test Example 4: Pharmacokinetics of Compounds 3 and 9 of Examples of the Present Invention

1. Abstract

SD rats were used as test animals, and the drug concentrations in plasma at different time points after oral administration of the compounds of Examples 3 and 9 were determined by LC/MS/MS. The pharmacokinetic behavior of the compounds of the present invention in rats was studied, and their pharmacokinetic characteristics were evaluated.

2. Experimental plan

2.1 Investigational Drugs

Compounds of Examples 3 and 9

2.2 Experimental animals

Eight healthy adult SD rats, half male and half female, were purchased from Shanghai Xipu-Bikai Laboratory Animal Co., Ltd., with animal production license number: SCXK (Shanghai) 2008-0016.

2.3 Drug preparation

Weigh an appropriate amount of sample, add 0.5% CMC-Na, and ultrasonicate to make a 0.5 mg/ml suspension.

2.4 Administration

Eight SD rats, half male and half female, were divided equally into two groups; after fasting overnight, the rats were gavaged and given the drug at a volume of 10 ml/kg.

3. Operation

In the oral administration group, 0.2 ml of blood was collected from the eye socket before administration and 0.5, 1.0, 2.0, 4.0, 6.0, 8.0, 11.0, and 24.0 h after administration. The blood was placed in a heparinized tube and centrifuged at 3500 rpm for 10 min to separate the plasma. The plasma was stored at -20°C and eaten 2 h after administration.

The levels of the test compounds in rat plasma after oral administration of different compounds were determined by LC/MS/MS.

4. Pharmacokinetic parameter results

The pharmacokinetic parameters of the compounds of Examples 3 and 9 of the present invention are as follows:

Conclusion: The compounds of the present invention have good pharmacokinetic absorption and obvious pharmacokinetic absorption effect.

Test Example 5: Pharmacokinetics of Compounds 28 and 41 of Examples of the Present Invention

1. Abstract

C57BL/6 mice were used as test animals. LC/MS/MS was used to determine the plasma drug concentrations at different time points after oral administration of the compounds of Examples 28 and 41 to investigate the pharmacokinetic behavior of the compounds of the present invention in C57BL/6 mice and to evaluate their pharmacokinetic characteristics.

2. Experimental plan

2.1 Investigational Drugs

Compounds of Examples 28 and 41

2.2 Experimental animals

Eighteen female C57BL/6 mice were purchased from Shanghai Xipu-Bikai Laboratory Animal Co., Ltd.

2.3 Drug preparation

Weigh an appropriate amount of sample, add 0.5% CMC-Na, and ultrasonicate to make a 1 mg/ml suspension.

2.4 Administration

Eighteen female C57BL/6 mice were divided into two groups, 9 mice in each group; after fasting overnight, the mice were gavaged and given the drug at a volume of 0.2 ml/kg.

3. Operation

From the oral administration group, 0.2 ml of blood was collected at 0.5, 1.0, 2.0, 4.0, 6.0, 8.0, 11.0, and 24.0 h after administration, placed in heparinized tubes, centrifuged at 3500 rpm for 10 min to separate plasma, and stored at -20°C.

The levels of the test compounds in rat plasma after oral administration of different compounds were determined by LC/MS/MS.

4. Pharmacokinetic parameter results

The pharmacokinetic parameters of the compounds of Examples 28 and 41 of the present invention are as follows:

Conclusion: The compounds of the present invention have good pharmacokinetic absorption and obvious pharmacokinetic absorption effect.

Claims (22)

1. A compound represented by general formula (I): Or in the form of its tautomers, racemates, or mixtures thereof, or its pharmaceutically acceptable salts. in: M is an inorganic acid or an organic acid; A is selected from _C3-8 cycloalkyl or 3- to 6-membered heterocyclic groups; ^R1 is selected from _C1-6 alkyl, _C3-8 cycloalkyl, 3 to 6-membered heterocyclic, _C6-10 aryl, 5 or 6-membered heteroaryl, indolyl, benzimidazolyl and -C(O) ^NHR5 ; ^R2 may be the same or different, and each is independently selected from _C1-6 alkoxy and halogen; ^R3 may be the same or different, and each is independently selected from hydrogen atom, _C1-6 alkyl, _C1-6 haloalkyl, _C1-6 alkoxy, _C1-6 haloalkoxy, halogen, amino, nitro, hydroxyl, cyano, _C6-10 aryl, 5- or 6-membered heteroaryl, ^-OR4 , ^-NR5R6 , -C(O) ^{NR5R6 and -NR5C} (O) ^{R6 ;} wherein the alkyl, haloalkyl, aryl and heteroaryl groups are optionally further substituted by one or more substituents selected from _C1-6 alkyl, _C1-6 haloalkyl, halogen, amino, nitro, cyano, hydroxyl, _C1-6 alkoxy, _C1-6 haloalkoxy, _C1-6 hydroxyalkyl, 3 to ⁶ -membered heterocyclic, ^-Ra , ^-OR7 , ^-S (O) _mR7 and ^-NR7R8 ; ^Ra is a _C1-6 alkyl or a 3- to 6-membered heterocyclic group, wherein the alkyl or heterocyclic group is optionally further substituted by one or more substituents selected from _C1-6 alkyl, halogen, amino, nitro, hydroxy, _C1-6 alkoxy, _C1-6 hydroxyalkyl, ^cyano , 3- to 6-membered heterocyclic group, -S(O) _mR4 and ^-NR5C (O) ^R6 ; ^R4 is selected from hydrogen atoms, _C1-6 alkyl groups or 3 to 6-membered heterocyclic groups; wherein the alkyl and heterocyclic groups are optionally further substituted by one or more substituents selected from -C(O) ^OR7 or -S(O) _mR7 ^{; R5} and ^R6 may be the same or different, and each is independently selected from hydrogen atoms, _C1-6 alkyl groups and 3 to 6-membered heterocyclic groups; ^R7 and ^R8 may be the same or different, and each is independently selected from hydrogen atoms and _C1-6 alkyl groups; p is 0 or 1; y is an integer of 0, 1, 2 or 3; m is 1 or 2; and n is an integer of 0, 1, 2, 3, 4 or 5; The 3- to 6-membered heterocyclic groups mentioned above contain 3 to 6 ring atoms, of which 1 to 4 are heteroatoms selected from nitrogen, oxygen or sulfur; The 5- or 6-membered heteroaryl groups mentioned above are heteroaromatic systems containing 5 or 6 ring atoms, of which 1 to 4 are heteroatoms selected from oxygen, sulfur, and nitrogen. 2. The compound of general formula (I) according to claim 1, wherein M is trifluoroacetic acid. 3. The compound of formula (I) according to claim 1, wherein y is 0, 1 or 3. 4. The compound of general formula (I) according to claim 1, wherein y is 0. 5. The compound of formula (I) according to claim 1, wherein n is 0, 1 or 2. 6. The compound of general formula (I) according to claim 1, wherein it is a compound of general formula (II): Or in the form of its tautomers, racemates, or mixtures thereof, or its pharmaceutically acceptable salts. in: X is CH or N; ^R1 to ^R3 , M, p, n and y are as defined in claim 1; a is an integer of 0, 1, 2, or 3; and b is an integer of 0, 1, or 2. 7. The compound of general formula (I) according to any one of claims 1 to 6, wherein it is a compound of general formula (III): Or in the form of its tautomers, racemates, or mixtures thereof, or its pharmaceutically acceptable salts. in: ^R1 to ^R3 , M, p, n and y are as defined in claim 1. 8. The compound of formula (I) according to claim 1, wherein the compound is a compound of formula (IV): Or in the form of its tautomers, racemates, or mixtures thereof, or its pharmaceutically acceptable salts. in: ^R2 , ^R3 , M, p, n and y are as defined in claim 1. 9. The compound of formula (I) according to any one of claims 1, 6 or 8, wherein it is a compound of formula (IV-1): Or in the form of its tautomers, racemates, or mixtures thereof, or its pharmaceutically acceptable salts. in: ^R2 , Ra, M, p, and y are as defined in claim 1. 10. The compound of formula (I) according to any one of claims 1, 6 or 8, wherein it is a compound of formula (IV-2): Or in the form of its tautomers, racemates, or mixtures thereof, or its pharmaceutically acceptable salts. in: ^R2 , ^R4 , M, p, and y are as defined in claim 1. 11. The compound of formula (I) according to any one of claims 1, 6 or 8, wherein it is a compound of formulas (IV-A) and (IV-B): Or in the form of its tautomers, racemates, or mixtures thereof, or its pharmaceutically acceptable salts. in: ^R2 , ^R3 , M, p, n and y are as defined in claim 1. 12. The compound of formula (I) according to any one of claims 1 to 6, wherein the compound is selected from: 13. The compound of general formula (I) according to any one of claims 1 to 6, wherein the compound is selected from: 14. A compound represented by general formula (V): or its tautomers, racemates, or mixtures thereof in: Q is an inorganic acid or an organic acid; X is CH or N; ^R2 , p, a, and b are as defined in claim 6; and x is an integer of 0, 1, 2, or 3. 15. The compound of general formula (V) according to claim 14, wherein Q is trifluoroacetic acid. 16. A method for preparing a compound of general formula (II-1), the method comprising: The compound of general formula (V) undergoes a coupling reaction with the halide of ^R1 in the presence of a catalyst under basic conditions. The product is optionally further reacted with boric acid or borate ester of ^R3 to give the compound of general formula (II-1). in: X is N; ^R1 to ^R3 , p, n, a, and b are defined as described in claim 6; and Q and x are as defined in claim 14. 17. A method for preparing a compound of general formula (II) according to claim 6, the method comprising: Compounds of general formula (II-1) form salts under acidic conditions to give compounds of general formula (II); in: X is CH or N; ^R1 to ^R3 , M, p, y, n, a, and b are defined as described in claim 6. 18. A pharmaceutical composition comprising a therapeutically effective amount of the compound of general formula (I) according to any one of claims 1 to 6, 12 and 13, or a tautomer, racemate or mixture thereof, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents or excipients. 19. Use of the compound of general formula (I) according to any one of claims 1 to 6, 12 and 13, or a tautomer, racemate or mixture thereof, or a pharmaceutically acceptable salt thereof, or the composition according to claim 18, in the preparation of a medicament for the prevention and/or treatment of diseases having pathological features of the IDO-mediated tryptophan metabolic pathway. 20. The use according to claim 19, wherein the disease having pathological features of the IDO-mediated tryptophan metabolism pathway is selected from cancer, myelodysplastic syndromes, Alzheimer's disease, autoimmune diseases, depression, anxiety disorders, cataracts, mental disorders, and AIDS. 21. The use according to claim 20, wherein the cancer is selected from breast cancer, cervical cancer, colon cancer, lung cancer, gastric cancer, rectal cancer, pancreatic cancer, brain cancer, skin cancer, oral cancer, prostate cancer, bone cancer, kidney cancer, ovarian cancer, bladder cancer, liver cancer, fallopian tube tumors, peritoneal tumors, stage IV melanoma, glioma, papillary renal tumor, head and neck tumors, leukemia, lymphoma, and myeloma. 22. The use according to claim 21, wherein the cancer is selected from non-small cell lung cancer, hepatocellular carcinoma, and glioblastoma.