Deuterium-modified thienopyridone compounds

Abstract

The invention relates to a deuterium-modified thienopyridone compound, a preparation method thereof, a pharmaceutical composition thereof and a medical use thereof. The deuterium-modified thienopyridone compound is shown in Formula I, and it inhibits the activity of HPK1 kinase and can achieve an anti-tumor effect. #imgabs0#

Description

Technical Field

[0001] The present application belongs to the field of pharmaceutical chemistry and relates to a deuterium-modified thienopyridone compound, and specifically relates to a compound shown in Formula I or a pharmaceutically acceptable salt thereof, a preparation method thereof, a pharmaceutical composition thereof and its medical use. The deuterium-modified thienopyridone compound of the present application shows inhibition of HPK1 kinase activity and can achieve an anti-tumor effect. Background Art

[0002] Hematopoietic progenitor kinase 1 (HPK1), also known as mitogen-activated protein kinase 1 (MAP4K1), is a mammalian Ste20-related serine / threonine protein kinase, a microtubule-associated protein, and a member of the mitogen-activated protein kinase (MAP4K) family. The family also includes five subtypes: GCK / MAP4K2, GLK / MAP4K3, HGK / MAP4K4, KHS1 / MAP4K5, and MINK1 / MAP4K6. Unlike the other five MAP4K subtypes that are widely expressed in tissue cells, HPK1 is only expressed in hematopoietic tissue cells. It can participate in regulating the signal transduction of the hematopoietic system, including lymphocytes, by mediating multiple cell signaling pathways (including MAPK signals, antigen receptor signals, and cytokine signals).

[0003] Studies have found that HPK1 mainly acts through the c-Jun N-terminal kinase (JNK) and extracellular signal-regulated protein kinases (ERK) signaling pathways to inhibit immune cell responses. In T cells, after T cell receptor (TCR) protein activation, HPK1 interacts with a large number of TCRs and is phosphorylated by tyrosine kinases Lck and Zap70. Activated HPK1 further phosphorylates T cell receptor adaptor protein SLP-76, establishing a docking site for negative regulator 14-3-3, ultimately destroying the stability of the TCR signaling complex (lato-gads-SLP76) and hindering the transmission of downstream mitogen-activated protein (MAP) kinase signals, negatively regulating TCR signaling, and then inhibiting T cell proliferation. In B cells, there is also a similar negative feedback mechanism. The B cell receptor (BCR) signal is phosphorylated and activated by HPK1, which in turn blocks downstream signal transmission and inhibits B cell proliferation. In addition, HPK1 also has a negative feedback regulatory effect on NK cells (Natural killer) and dendritic cells (DC).

[0004] Due to its important role in immunity, HPK1 inhibitors play an important role in malignant solid tumors or blood cancers (such as acute myeloid leukemia, breast cancer, lung cancer, etc.), autoimmune diseases (such as systemic lupus erythematosus, psoriatic arthritis) and inflammatory responses.

[0005] Currently, there is no drug on the market targeting HPK1, and there is still a need to develop compounds with selective inhibitory activity, or better pharmacodynamics, or better pharmacokinetics in this field. Summary of the invention

[0006] The present application provides a compound represented by formula I or a pharmaceutically acceptable salt thereof,

[0007]

[0008] in,

[0009] R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R9 , R 10 , R 11 , R 12 , R 13 , R 14 , R 15 and R 16 Each is independently selected from hydrogen (H) or deuterium (D);

[0010] The condition is R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 , R 14 , R 15 and R 16 At least one of is selected from deuterium.

[0011] In some embodiments, the R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 , R 14 , R 15 and R 16 At least two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen or sixteen of the are selected from deuterium.

[0012] In some embodiments, the R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 , R 14 , R 15and R 16 Any one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen or sixteen of the are selected from deuterium.

[0013] In some embodiments, the R 1 , R 2 and R 3 Selected from deuterium, R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0014] In some embodiments, the R 4 , R 5 , R 10 and R 11 Selected from deuterium, R 1 , R 2 , R 3 , R 6 , R 7 , R 8 , R 9 , R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0015] In some embodiments, the R 6 , R 7 , R 8 and R 9 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 10 , R 11 , R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0016] In some embodiments, the R 1 , R2 , R 3 , R 4 , R 5 , R 10 and R 11 Selected from deuterium, R 6 , R 7 , R 8 , R 9 , R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0017] In some embodiments, the R 1 , R 2 , R 3 , R 6 , R 7 , R 8 and R 9 Selected from deuterium, R 4 , R 5 , R 10 , R 11 , R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0018] In some embodiments, the R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 and R 11 Selected from deuterium, R 1 , R 2 , R 3 , R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0019] In some embodiments, the R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 and R11 Selected from deuterium, R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0020] In some embodiments, the R 12 and R 14 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 13 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0021] In some embodiments, the R 13 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0022] In some embodiments, the R 12 , R 13 and R 14 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0023] In some embodiments, the R 1 , R 2 , R 3 , R 12 , R 13 and R 14 Selected from deuterium, R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 15 and R 16 are each independently selected from hydrogen or deuterium.

[0024] In some embodiments, the R 15 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 , R 14 and R 16 are each independently selected from hydrogen or deuterium.

[0025] In some embodiments, the R 16 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 , R 14 and R 15 are each independently selected from hydrogen or deuterium.

[0026] In some embodiments, the R 15 and R 16 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 and R 14 are each independently selected from hydrogen or deuterium.

[0027] In some embodiments, the R 1 , R 2 , R 3 , R 15 and R 16 Selected from deuterium, R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 13 and R 14 are each independently selected from hydrogen or deuterium.

[0028] In some embodiments, the compound represented by Formula I or a pharmaceutically acceptable salt thereof is selected from the following compounds or a pharmaceutically acceptable salt thereof:

[0029] Compound <![CDATA[R 1 ]]> <![CDATA[R 2 ]]> <![CDATA[R 3 ]]> <![CDATA[R 4 ]]> <![CDATA[R 5 ]]> <![CDATA[R 6 ]]> <![CDATA[R 7 ]]> <![CDATA[R 8 ]]> <![CDATA[R 9 ]]> <![CDATA[R 10 ]]> <![CDATA[R 11 ]]> <![CDATA[R 12 ]]> <![CDATA[R 13 ]]> <![CDATA[R 14 ]]> <![CDATA[R 15 ]]> <![CDATA[R 16 ]]> I-1 D D D H H H H H H H H H H H H H I-2 H H H D D D D D D D D H H H H H I-3 H H H D D H H H H D D H H H H H I-4 H H H H H D D D D H H H H H H H I-5 D D D D D D D D D D D H H H H H I-6 D D D D D H H H H D D H H H H H I-7 H H H H H H H H H H H D D D H H I-8 H H H H H H H H H H H H H H D D I-9 D D D H H H H H H H H D D D H H I-10 D D D H H H H H H H H H H H D D I-11 H H H H H H H H H H H D H D H D

[0030] In some embodiments, the compound represented by Formula I or a pharmaceutically acceptable salt thereof is selected from the following compounds or a pharmaceutically acceptable salt thereof:

[0031]

[0032]

[0033] On the other hand, the present application provides a pharmaceutical composition, which includes a compound of the present application or a pharmaceutically acceptable salt thereof, and optionally further includes a pharmaceutically acceptable excipient. In one embodiment, the pharmaceutical composition also includes a second active agent, which is another anticancer agent (e.g., a small molecule anticancer agent or an antibody anticancer agent).

[0034] On the other hand, the present application provides the use of the compound of the present application or its pharmaceutically acceptable salt, or the above-mentioned pharmaceutical composition in the preparation of a drug for treating a disease that benefits from inhibiting HPK1 kinase activity. In one embodiment, the drug further comprises a second active agent for treating the disease, and the second active agent is another anticancer agent (such as a small molecule anticancer agent or an antibody anticancer agent).

[0035] Another aspect of the present application provides use of the compound of the present application or a pharmaceutically acceptable salt thereof, or the above-mentioned pharmaceutical composition in treating diseases that benefit from inhibiting HPK1 kinase activity.

[0036] Another aspect of the present application provides the compound of the present application or a pharmaceutically acceptable salt thereof, or the above-mentioned pharmaceutical composition for use in treating a disease that benefits from the inhibition of HPK1 kinase activity.

[0037] On the other hand, the present application provides a method for treating diseases that benefit from inhibiting HPK1 kinase activity, comprising administering a therapeutically effective amount of the compound of the present application or a pharmaceutically acceptable salt thereof, or the above-mentioned pharmaceutical composition to an individual in need of such treatment (the individual is selected from mammals, preferably humans).

[0038] In some embodiments, the disease that benefits from inhibition of HPK1 kinase activity is selected from tumors or cancers, such as blood cancers or solid tumors, such as acute myeloid leukemia, breast cancer or lung cancer.

[0039] In one embodiment, in the above-mentioned disease treatment methods, the compound of the present application or its pharmaceutically acceptable salt or the above-mentioned pharmaceutical composition is administered in combination with other anticancer agents (such as small molecule anticancer agents or antibody anticancer agents) or other anticancer therapies (such as radiotherapy and / or chemotherapy).

[0040] The compounds of the present application have good HPK1 in vitro kinase inhibitory activity, cell p-SLP76 phosphorylation inhibitory activity, and good in vitro liver microsome stability. DETAILED DESCRIPTION

[0041] definition

[0042] Unless otherwise specified, the following terms used in this application have the following meanings. A particular term should not be considered to be uncertain or unclear in the absence of a special definition, but should be understood according to the common meaning in the art. When a trade name appears in this article, it is intended to refer to the corresponding commodity or its active ingredient.

[0043] The word "comprise" or "comprises" and its English variations such as comprises or comprising should be construed in an open and non-exclusive sense, ie, "including but not limited to".

[0044] The term "independently" means that when any variable (e.g., R) appears more than once in the composition or structure of a compound, its definition in each case is independent. For example, if a group is substituted by 2 Rs, each R has an independent option. Or, for example, if multiple groups are substituted by a certain R, each group substituted by R is independent of each other.

[0045] The term "pharmaceutically acceptable" refers to those compounds, materials, compositions and / or dosage forms which, within the scope of sound medical judgment, are suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0046] As the pharmaceutically acceptable salt, for example, metal salts, ammonium salts, salts with organic bases, salts with inorganic acids, salts with organic acids, salts with basic or acidic amino acids and the like can be mentioned.

[0047] The term "pharmaceutical composition" refers to a mixture of one or more compounds of the present application or their salts and pharmaceutically acceptable excipients. The purpose of a pharmaceutical composition is to facilitate administration of the compounds of the present application to an organism.

[0048] The term "pharmaceutically acceptable excipients" refers to those excipients that have no significant irritation to the organism and do not impair the biological activity and performance of the active compound. Suitable excipients are well known to those skilled in the art, such as carbohydrates, waxes, water-soluble and / or water-swellable polymers, hydrophilic or hydrophobic materials, gelatin, oils, solvents, water, etc.

[0049] The pharmaceutical composition of the present application can be prepared by combining the compound of the present application with suitable pharmaceutically acceptable excipients, for example, it can be formulated into solid, semi-solid, liquid or gaseous preparations, such as tablets, pills, capsules, powders, granules, ointments, emulsions, suspensions, suppositories, injections, inhalants, gels, microspheres and aerosols, etc.

[0050] Typical routes of administration of the compounds of the present application or their pharmaceutically acceptable salts or their pharmaceutical compositions include, but are not limited to, oral, rectal, topical, inhalation, parenteral, sublingual, vaginal, intranasal, intraocular, intraperitoneal, intramuscular, subcutaneous, and intravenous administration.

[0051] The term "treat" generally refers to obtaining a desired pharmacological and / or physiological effect. The effect may be therapeutic, partially or completely stabilizing or curing a disease and / or side effects resulting from a disease. As used herein, "treat" encompasses any treatment of a patient's disease that: (a) inhibits the symptoms of a disease, i.e., prevents its development; or (b) alleviates the symptoms of a disease, i.e., causes regression of the disease or symptoms.

[0052] The term "effective amount" means the amount of the compound of the present application that (i) treats a given disease, condition or disorder, (ii) alleviates, improves or eliminates one or more symptoms of a specific disease, condition or disorder, or (iii) delays the onset of one or more symptoms of a specific disease, condition or disorder described herein. The amount of active substance (e.g., antibody or compound of the present application) constituting a "therapeutically effective amount" may vary according to factors such as the disease state, age, sex and weight of the individual, and the ability of the therapeutic agent or therapeutic agent combination to induce a desired response in the individual. The effective amount may also be routinely determined by those skilled in the art based on their own knowledge and the present disclosure.

[0053] In this application, the term "individual" includes humans and animals, for example, mammals (such as primates, cattle, horses, pigs, dogs, cats, mice, rats, rabbits, goats, sheep and birds, etc.). In some embodiments, the subject or patient is a mouse. In some embodiments, the subject or patient is a human.

[0054] The therapeutic dose of the compounds of the present application may be determined, for example, based on the specific use of the treatment, the method of administering the compound, the health and condition of the patient, and the judgment of the prescribing physician. The ratio or concentration of the compounds of the present application in the pharmaceutical composition may not be fixed, depending on a variety of factors, including dosage, chemical properties (e.g., hydrophobicity), and route of administration. For example, the compounds of the present application may be provided by a physiologically buffered aqueous solution containing about 0.1 to 10% w / v of the compound for parenteral administration. Some typical dosage ranges are about 1 μg / kg to about 1 g / kg body weight / day. In certain embodiments, the dosage range is about 0.01 mg / kg to about 100 mg / kg body weight / day. The dosage is likely to depend on such variables as the type and degree of development of the disease or condition, the general health status of the specific patient, the relative biological efficacy of the selected compound, the excipient formulation, and its route of administration. The effective dose can be obtained by extrapolation of a dose-response curve derived from an in vitro or animal model test system.

[0055] The compounds of the present application can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthesis methods, and equivalent substitution methods well known to those skilled in the art. Preferred embodiments include but are not limited to the examples of the present application.

[0056] The chemical reactions of the specific embodiments of the present application are carried out in a suitable solvent, which must be suitable for the chemical changes of the present application and the reagents and materials required. In order to obtain the compounds of the present application, it is sometimes necessary for those skilled in the art to modify or select the synthesis steps or reaction processes based on the existing embodiments.

[0057] Example

[0058] For the sake of clarity, the present invention is further described with examples, but the examples do not limit the scope of the present application. All reagents used in the present application are commercially available and can be used without further purification.

[0059] In addition to the following examples, other compounds disclosed in the present application can be prepared by the preparation methods of the reference examples or preparation methods known in the art. Those skilled in the art can detect the mass spectrum (MS) or nuclear magnetic resonance (NMR) of the compounds by the methods of the present application or preparation methods known in the art.

[0060] Example 1: Preparation of 4-amino-5-(5-(4-(methyl-d3)piperazin-1-yl)-1H-benzo[d]imidazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one (Compound I-1)

[0061]

[0062] Step 1: Preparation of tert-butyl 4-(methyl-d3)piperazine-1-carboxylate (Compound 1A)

[0063] In a 250 mL reaction bottle, tert-butyl piperazine-1-carboxylate (5 g) and potassium carbonate (7.42 g) were dissolved in tetrahydrofuran (100 mL), stirred at 25°C under nitrogen protection, and then iodomethane-d3 (4.28 g) was added dropwise at -30°C, and stirred at 25°C under nitrogen protection. After the reaction was complete, the mixture was filtered and concentrated under reduced pressure to obtain compound 1A (5.5 g).

[0064] MS: m / z = 204.2 [M+H] + .

[0065] Step 2: Preparation of 1-(methyl-d3)piperazine (Compound 1B)

[0066] Compound 1A (5.5 g), trifluoroacetic acid (12 g), and dichloromethane (30 mL) were added to a 100 mL reaction bottle in sequence, and stirred at 25°C for reaction. After the reaction was complete, the reaction solution was adjusted to alkaline, extracted with dichloromethane (50 mL*3), and the organic phases were combined, dried, filtered, and concentrated under reduced pressure to obtain compound 1B (2.8 g).

[0067] MS: m / z = 104.2 [M+H] + .

[0068] Step 3: Preparation of 5-(4-(methyl-d3)piperazine-1-yl)-2-nitroaniline (Compound 1C)

[0069] Compound 1B (2.8 g), 5-chloro-2-nitroaniline (2.7 g), potassium carbonate (8.6 g), and N-methylpyrrolidone (20 mL) were added to a 100 mL reaction bottle in sequence, and stirred at 160° C. After the reaction was complete, the mixture was concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol=9 / 1) to obtain compound 1C (1.12 g).

[0070] MS: m / z = 240.2 [M+H] + .

[0071] Step 4: Preparation of 4-(4-(methyl-d3)piperazin-1-yl)benzene-1,2-diamine (Compound 1D)

[0072] 1C (1.12 g), ethanol (20 mL), water (10 mL), zinc powder (1.6 g), and ammonium chloride (0.131 g) were added to a 100 mL reaction bottle in sequence. The mixture was stirred at 80°C for reaction. After the reaction was complete, the mixture was filtered and the filtrate was concentrated under reduced pressure to obtain compound 1D (1 g).

[0073] MS: m / z = 210.3 [M+H] + .

[0074] Step 5: Preparation of ethyl 2-(5-(4-(methyl-d3)piperazine-1-yl)-1H-benzo[d]imidazol-2-yl)acetate (Compound 1E)

[0075] To a 100 mL reaction bottle, ethyl 3-ethoxy-3-iminopropionate hydrochloride (1.4 g), compound 1D (1 g), and dimethylformamide (20 mL) were added in sequence, and the mixture was stirred at 80° C. under nitrogen protection. After the reaction was complete, the mixture was concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol=4 / 1) to obtain compound 1E (1.416 g).

[0076] MS: m / z = 306.1 [M+H] + .

[0077] Step 6: Preparation of 4-amino-5-(5-(4-(methyl-d3)piperazin-1-yl)-1H-benzo[d]imidazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one (Compound I-1)

[0078] Compound 1E (1.416 g), 2-aminothiophene-3-carbonitrile (0.576 g), tetrahydrofuran (20 mL), and lithium diisopropylamide (4.9 g) were added to a 100 mL reaction bottle in sequence. The mixture was stirred at 60°C under nitrogen protection. After the reaction was complete, the mixture was quenched with saturated ammonium chloride, extracted with ethyl acetate (30 mL*3), and the organic phases were combined, dried, filtered, and concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol=9 / 1) to obtain compound I-1 (0.1 g).

[0079] MS: m / z = 384.6 [M+H] + .

[0080] 1 H NMR(500MHz,DMSO-d6)δ12.62(s,1H),12.10(s,1H),10.67(d,1H),8.00(s, 1H),7.45-7.60(m,2H),7.19(s,2H),6.89(s,1H),3.13(s,4H),2.60(s,4H).

[0081] Example 2: Preparation of 4-amino-5-(5-(4-methylpiperazin-1-yl-2,2,3,3,5,5,6,6-d8)-1H-benzo[d]imidazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one (Compound I-2)

[0082]

[0083] Step 1: Preparation of 5-(4-methylpiperazine-1-yl-2,2,3,3,5,5,6,6-d8)-2-nitroaniline (Compound 2A)

[0084] In a 100 mL reaction bottle, compound 1-methylpiperazine-2,2,3,3,5,5,6,6-d8 (0.847 g), 5-chloro-2-nitroaniline (1.356 g), potassium carbonate (4.330 g), N-methylpyrrolidone (20 mL) were added in sequence, stirred at 160°C for reaction, and concentrated under reduced pressure after the reaction was complete. The residue was separated by column chromatography (dichloromethane / methanol = 9 / 1) to obtain compound 2A (0.619 g).

[0085] MS: m / z = 245.1 [M+H] + .

[0086] Step 2: Preparation of 4-(4-methylpiperazine-1-yl-2,2,3,3,5,5,6,6-d8)benzene-1,2-diamine (Compound 2B)

[0087] Compound 2A (0.618 g), ethanol (30 mL), water (5 mL), zinc powder (0.828 g) and ammonium chloride (1.354 g) were added to a 100 mL reaction bottle in sequence. The mixture was stirred at 80°C for reaction. After the reaction was complete, the mixture was filtered and the filtrate was concentrated under reduced pressure to obtain compound 2B (0.543 g).

[0088] MS: m / z = 215.2 [M+H] + .

[0089] Step 3: Preparation of ethyl 2-(5-(4-methylpiperazin-1-yl-2,2,3,3,5,5,6,6-d8)-1H-benzo[d]imidazol-2-yl)acetate (Compound 2C)

[0090] To a 100 mL reaction bottle, ethyl 3-ethoxy-3-iminopropionate hydrochloride (0.826 g), compound 2B (0.543 g) and dimethylformamide (10 mL) were added in sequence, and the mixture was stirred at 80° C. under nitrogen protection. After the reaction was complete, the mixture was concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol=4 / 1) to obtain compound 2C (0.314 g).

[0091] MS: m / z = 311.5 [M+H] + .

[0092] Step 4: Preparation of 4-amino-5-(5-(4-methylpiperazin-1-yl-2,2,3,3,5,5,6,6-d8)-1H-benzo[d]imidazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one (Compound I-2)

[0093] Compound 2C (0.310 g), 2-aminothiophene-3-carbonitrile (0.200 g), tetrahydrofuran (30 mL), and 2M tetrahydrofuran solution of lithium diisopropylamide (5 mL) were added to a 100 mL reaction bottle in sequence. The mixture was stirred at 60°C under nitrogen protection. After the reaction was complete, the mixture was quenched with saturated ammonium chloride, extracted with ethyl acetate, and the organic phases were combined, dried, filtered, and concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol = 9 / 1) to give compound I-2 (46 mg).

[0094] MS: m / z = 389.5 [M+H] + .

[0095] 1H NMR (500MHz, DMSO-d6) δ12.61(s,1H),12.10(s,1H),10.67(d,J=51.5Hz,1H),7.99(s,1H),7.60( d,J=5.5Hz,1H),7.47(dd,J=8.5,32.0Hz,1H),7.19-7.10(m,2H),6.90-6.87(m,1H),2.29(s,3H).

[0096] Example 3: Preparation of 4-amino-5-(5-(4-methylpiperazin-1-yl-3,3,5,5-d4)-1H-benzo[d]imidazol-2-yl)thiophene[2,3-b]pyridin-6(7H)-one (Compound I-3)

[0097]

[0098] Step 1: Preparation of 1-benzyl-4-nitrosopiperazine-3,3,5,5-d4 (Compound 3A)

[0099] To a 500 mL reaction bottle, heavy water (200 mL), 1-benzyl-4-nitrosopiperazine (30 g) and sodium methoxide (23.69 g) were added in sequence, and the reaction was stirred at 80 ° C under nitrogen protection. Then deuterated ethanol (150 mL) was added, and the reaction was continued to be stirred at 80 ° C under nitrogen protection. After the reaction was complete, the reaction solution was cooled to 0 ° C, filtered, and the filter cake was washed and dried to obtain compound 3A (25 g).

[0100] MS: m / z = 210.5 [M+H] + .

[0101] Step 2: Preparation of 1-benzylpiperazine-3,3,5,5-d4 (Compound 3B)

[0102] In a 1L reaction bottle, compound 3A (20 g), sodium methoxide (23.23 g), deuterated ethanol (150 mL), heavy water (225 mL) and nickel aluminum alloy (60 g) were added in sequence and stirred at 30°C. After the reaction was complete, the mixture was filtered and the filtrate was concentrated under reduced pressure. The residue was extracted with dichloromethane (100 mL*3), and the organic phases were combined, dried, filtered, and concentrated under reduced pressure to obtain compound 3B (13 g).

[0103] MS: m / z = 181.5 [M+H] +.

[0104] 1 H NMR (500MHz, CDCl3) δ7.35-7.20 (m, 5H), 3.49 (d, J = 7.0Hz, 2H), 2.40 (s, 4H).

[0105] Step 3: Preparation of 1-benzyl-4-methylpiperazine-3,3,5,5-d4 (Compound 3C)

[0106] At 0°C, compound 3B (13 g), tetrahydrofuran (50 mL), and sodium hydride (2.88 g) were added to a 100 mL reaction bottle in sequence, and the temperature was slowly raised to 25°C and stirred for 0.5 h. Then, methyl 4-methylbenzenesulfonate (6.71 g) was added to the reaction solution at 0°C, and the mixture was stirred at 25°C. After the reaction was complete, the reaction was quenched and concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol=30 / 1) to obtain compound 3C (4.5 g).

[0107] MS: m / z = 195.5 [M + H] + .

[0108] Step 4: Preparation of 1-methylpiperazine-2,2,6,6-d4 (Compound 3D)

[0109] In a 100 mL reaction bottle, compound 3C (4.5 g), 10% palladium on carbon (4 g) and anhydrous methanol (50 mL) were added in sequence. The mixture was stirred at 25° C. under hydrogen. After the reaction was complete, the mixture was filtered and the filtrate was concentrated under reduced pressure to give compound 3D (1.3 g).

[0110] MS: m / z = 105.4 [M+H] + .

[0111] Step 5: Preparation of 5-(4-methylpiperazine-1-yl-3,3,5,5-d4)-2-nitroaniline (Compound 3E)

[0112] In a 100 mL reaction bottle, compound 3D (1.3 g), 5-chloro-2-nitroaniline (2.15 g), potassium carbonate (6.9 g), and N-methylpyrrolidone (40 mL) were added in sequence. The mixture was stirred at 160°C for reaction. After the reaction was complete, the mixture was concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol = 9 / 1) to obtain compound 3E (1.4 g).

[0113] MS: m / z = 241.2 [M+H] + .

[0114] Step 6: Preparation of 4-(4-methylpiperazine-1-yl-3,3,5,5-d4)benzene-1,2-diamine (Compound 3F)

[0115] Compound 3E (1.4 g), zinc powder (1.9 g), ammonium chloride (0.31 g), ethanol (30 mL) and water (5 mL) were added to a 100 mL reaction bottle in sequence. The mixture was stirred at 80°C for reaction. After the reaction was complete, the mixture was filtered and the filtrate was concentrated under reduced pressure to obtain compound 3F (1.2 g).

[0116] MS: m / z = 211.3 [M+H] + .

[0117] Step 7: Preparation of ethyl 2-(5-(4-methylpiperazin-1-yl-3,3,5,5-d4)-1H-benzo[d]imidazol-2-yl)acetate (Compound 3G)

[0118] In a 100 mL reaction bottle, compound 3F (1.2 g), ethyl 3-ethoxy-3-iminopropionate hydrochloride (2.23 g) and ethanol (30 mL) were added in sequence. The mixture was stirred at 80°C for reaction. After the reaction was complete, the mixture was concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol=30 / 1) to obtain compound 3G (0.5 g).

[0119] MS: m / z = 306.9 [M+H] + .

[0120] Step 8: Preparation of 4-amino-5-(5-(4-methylpiperazin-1-yl-3,3,5,5-d4)-1H-benzo[d]imidazol-2-yl)thiophene[2,3-b]pyridin-6(7H)-one (Compound I-3)

[0121] Compound 3G (0.5 g), 2-aminothiophene-3-carbonitrile (0.32 g), tetrahydrofuran (20 ml), lithium diisopropylamide (1.75 g) were added to a 100 mL reaction bottle in sequence, and stirred at 60 ° C under nitrogen protection. After the reaction was complete, saturated ammonium chloride was used to quench the reaction, and ethyl acetate was used for extraction (30 mL*3). The organic phases were combined, dried, filtered, and concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol = 9 / 1) to obtain compound I-3 (0.1 g).

[0122] MS: m / z = 385.5 [M+H] + .

[0123] 1H NMR (500MHz, DMSO-d6) δ12.62 (s, 1H), 12.10 (s, 1H), 10.67 (d, J = 47.3Hz, 1H),8.01(s,1H),7.60(d,J=5.6Hz,1H),7.48(dd,J=32.3,8.4Hz,1H),7.17(dd,J=24.0,18.4Hz,2H),6.90(s,1H),3.13(s,4H),2.33(s,3H).

[0124] Example 4: Preparation of 4-amino-5-(5-(4-methylpiperazin-1-yl-2,2,6,6-d4)-1H-benzo[d]imidazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one (Compound I-4)

[0125]

[0126] Step 1: Preparation of 1-methyl-4-nitrosopiperazine (Compound 4A)

[0127] In a 100 mL reaction bottle, 1-methylpiperazine (10 g), water (20 mL) and concentrated hydrochloric acid (25 mL) were added in sequence. At 0°C, a solution of sodium nitrite (8.27 g) dissolved in water (20 mL) was added to the reaction solution. The reaction was stirred at 25°C. After the reaction was complete, the reaction solution was adjusted to alkaline. The reaction solution was extracted with dichloromethane (100 mL*3), the organic phases were combined, dried, filtered, and concentrated under reduced pressure to obtain compound 4A (10.8 g).

[0128] MS: m / z = 130.2 [M+H] + .

[0129] Step 2: Preparation of 1-methyl-4-nitrosopiperazine-3,3,5,5-d4 (Compound 4B)

[0130] To a 100 mL reaction bottle, heavy water (30 mL), compound 4A (5 g) and deuterated sodium hydroxide (5.29 g) were added in sequence. Under nitrogen protection, the mixture was stirred at 100 °C for reaction. After completion of the reaction, the reaction solution was extracted with dichloromethane (50 mL*3). The organic phases were combined, dried, filtered, and concentrated under reduced pressure to obtain compound 4B (5.3 g).

[0131] 1 H NMR (500MHz, CDCl3) δ2.57(s,2H),2.33(s,2H),2.31(s,3H).

[0132] Step 3: Preparation of 1-methylpiperazine-3,3,5,5-d4 (Compound 4C)

[0133] In a 100 mL reaction bottle, compound 4B (5 g), sodium methoxide (15.2 g), deuterated ethanol (23 mL), heavy water (23 mL) and nickel aluminum alloy (15 g) were added in sequence and stirred at 30 ° C. After the reaction was complete, the mixture was filtered and the filtrate was concentrated under reduced pressure. The residue was extracted with dichloromethane (100 mL*3), and the organic phases were combined, dried, filtered, and concentrated under reduced pressure to obtain compound 4C (4 g).

[0134] MS: m / z = 105.5 [M + H] + .

[0135] Step 4: Preparation of 5-(4-methylpiperazine-1-yl-2,2,6,6-d4)-2-nitroaniline (Compound 4D)

[0136] Compound 4C (3.677 g), 5-chloro-2-nitroaniline (5.413 g), potassium carbonate (10.847 g), and N-methylpyrrolidone (50 mL) were added to a 100 mL reaction bottle in sequence, and stirred at 160° C. After the reaction was complete, the mixture was concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol=9 / 1) to obtain compound 4D (1.410 g).

[0137] MS: m / z = 241.2 [M+H] + .

[0138] Step 5: Preparation of 4-(4-methylpiperazine-1-yl-2,2,6,6-d4)benzene-1,2-diamine (Compound 4E)

[0139] In a 100 mL reaction bottle, 4D (1.400 g), ethanol (30 mL), water (5 mL), zinc powder (2.245 g) and ammonium chloride (0.400 g) were added in sequence. The mixture was stirred at 80°C for reaction. After the reaction was complete, the mixture was filtered and the filtrate was concentrated under reduced pressure to obtain compound 4E (1.403 g).

[0140] MS: m / z = 211.3 [M+H] + .

[0141] Step 6: Preparation of ethyl 2-(5-(4-methylpiperazin-1-yl-2,2,6,6-d4)-1H-benzo[d]imidazol-2-yl)acetate (Compound 4F)

[0142] To a 100 mL reaction bottle, ethyl 3-ethoxy-3-iminopropionate hydrochloride (2.230 g), compound 4E (1.400 g), and ethanol (50 mL) were added in sequence, and the mixture was stirred at 80° C. under nitrogen protection. After the reaction was complete, the mixture was concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol=4 / 1) to obtain compound 4F (1.241 g).

[0143] MS: m / z = 307.3 [M+H] + .

[0144] Step 7: Preparation of 4-amino-5-(5-(4-methylpiperazin-1-yl-2,2,6,6-d4)-1H-benzo[d]imidazol-2-yl)thieno[2,3-b]pyridin-6(7H)-one (Compound I-4)

[0145] Compound 4F (1.241 g), 2-aminothiophene-3-carbonitrile (0.600 g), tetrahydrofuran (40 mL), and 2M tetrahydrofuran solution of lithium diisopropylamide (20 mL) were added to a 100 mL reaction bottle in sequence. The mixture was stirred at 60°C under nitrogen protection. After the reaction was complete, the reaction was quenched with saturated ammonium chloride, extracted with ethyl acetate, and the organic phases were combined, dried, filtered, and concentrated under reduced pressure. The residue was separated by column chromatography (dichloromethane / methanol = 9 / 1) to obtain compound I-4 (70 mg).

[0146] MS: m / z = 385.5 [M+H] + .

[0147] 1 H NMR (500MHz, DMSO-d6) δ12.61(s,1H),12.10(s,1H),10.69(d,J=51.0Hz,1H),7.99(s,1H),7.60(d,J=5.5H z,1H),7.47(dd,J=9.0,32.5Hz,1H),7.19-7.10(m,2H),6.90-6.87(m,1H),2.51-2.48(m,4H),2.24(s,3H).

[0148] Example 5: Preparation of 4-amino-5-(5-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl-4,6-d2)thieno[2,3-b]pyridin-6(7H)-one-2-d (Compound I-11)

[0149]

[0150] Step 1: Preparation of 4-amino-5-(5-(4-methylpiperazin-1-yl)-1H-benzo[d]imidazol-2-yl-4,6-d2)thieno[2,3-b]pyridin-6(7H)-one-2-d (Compound I-11)

[0151] In a 25 mL microwave tube, add compound 4-amino-5-(5-(4-methylpiperazine-1-yl)-1H-benzo[d]imidazol-2-yl-4,6)thieno[2,3-b]pyridine-6(7H)-one (0.3 g), heavy water (8 mL) and trifluoroacetic anhydride (0.11 mL) in sequence. After the addition, heat the reaction solution to 140°C with microwave and stir to react. After the reaction is complete, adjust the reaction solution to alkalinity, extract with ethyl acetate (30 mL*3), combine the organic phases, dry, filter, and concentrate under reduced pressure. The residue is separated by column chromatography (dichloromethane / methanol=9 / 1) to obtain compound I-11 (0.197 g).

[0152] MS: m / z = 384.5 [M+H] + .

[0153] 1 H NMR (500MHz, DMSO-d6) δ12.64(s,1H),12.12(s,1H),10.68(d,J=48.1Hz,1H),8.04(s,1H),7.62( s,1H),7.49(d,J=32.3Hz,1H),3.21–3.12(m,4H),2.69(d,J=12.0Hz,4H),2.37(d,J=8.3Hz,3H).

[0154] Experimental Example 1: HPK1 in vitro kinase inhibitory activity assay

[0155] Dilute the kinase buffer Enzymatic buffer 5× to 1×, and add 10mM MgCl2, 1mM DTT and 0.005% Tween 20. Prepare 100ng / μL HPK1 (Life technology) stock solution into 1.67× 1.67ng / μL working solution (final concentration is 1ng / μL) with kinase buffer, and plate 6μL per well (384-well plate). Add different compounds dissolved in DMSO to the wells using a nanoliter pipette, so that the final concentration of the compound is 1000nM-0.244nM, 4-fold gradient, a total of 7 concentrations, and set up blank control empty (no enzyme) and negative control wells (enzyme, solvent DMSO), and set up 2 duplicate wells. After the enzyme and compound were incubated at room temperature for 1 hour, 5× 0.5mM ATP (final concentration 0.1mM) and 5× 2.5μM substrate (Cisbio, STK Substrate 1-biotin, final concentration 500nM) diluted with kinase buffer were mixed in equal volumes, 4μL was added to each well, the plate was sealed with a sealing film, and incubated at room temperature for 2 hours. The detection antibody was prepared by mixing equal volumes of antibody STK Antibody-cryptate (Cisbio, 5μl / test) and 4× 500nM Streptavidin-XL665 (Cisbio, final concentration 125nM), 10μL was added to each well, and incubated at room temperature for 1 hour. The PE Envision multi-function plate reader was used to detect the signal value (excitation 665nm, emission 620nm), and the IC50 was calculated by four-parameter fitting. The results are shown in Table 1:

[0156] Table 1

[0157] Example In vitro enzyme inhibitory activity HPK1 IC50 (nM) 2 9.7 3 7.5 4 9.2 5 8.8

[0158] Experimental Example 2 Detection of p-SLP76 phosphorylation inhibitory activity in Jurkat cells

[0159] Take Jurkat cells that are in good growth state, collect them into a centrifuge tube, resuspend them after centrifugation, and adjust the cell density to 6.25×10 6 / mL, inoculated in a 384-well small volume white plate (8μL / well); use a nanoliter sampler to add compounds to a final concentration of 2500nM-10.29nM, 2 replicates, and set up controls at the same time. After 1 hour of cell culture, add stimulator CD3CD28 (manufacturer: stemcell, 4μL), incubate at 37℃ for 30 minutes; add 3μL lysis solution (manufacturer: BioAuxilium) to each well, shake and lyse at room temperature for 30 minutes, after lysis and mixing, add 5μL of pre-mixed antibodies (manufacturer: BioAuxilium) prepared in detection buffer, incubate at room temperature overnight, and detect with PerkinElmer Envision multi-function microplate reader (excitation 320nm, emission 615nm / 665nm), and calculate IC50 using four-parameter fitting. The test results are shown in Table 2.

[0160] Table 2

[0161]

[0162] Test Example 3 In vitro liver microsome stability

[0163] Preparation of liver microsome incubation samples: PBS buffer (pH 7.4), liver microsome solution (0.5 mg / mL, HLM and MLM, respectively), test compound and NADPH+MgCl2 solution were mixed and incubated at 37°C and 300 rpm for 1 hour. Preparation of 0 hour samples: PBS buffer (pH 7.4), liver microsome solution (0.5 mg / mL, HLM and MLM, respectively) and test compound were mixed. Acetonitrile solution containing internal standard was added to the sample to prepare supernatant by protein precipitation, and diluted for LC / MS / MS determination. The results are shown in Table 3.

[0164] Table 3

[0165]

Claims

1. A compound of formula I or a pharmaceutically acceptable salt thereof, , in, R 4 , R 5 , R 10 and R 11 Selected from deuterium, R 1 , R 2 , R 3 , R 6 , R 7 , R 8 and R 9 Selected from hydrogen, R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium; or, R 6 , R 7 , R 8 and R 9 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 10 and R 11 Selected from hydrogen, R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium; or, R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 and R 11 Selected from deuterium, R 1 , R 2 and R 3 Selected from hydrogen, R 12 , R 13 , R 14 , R 15 and R 16 are each independently selected from hydrogen or deuterium; or, R 12 , R 14 and R 16 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 13 and R 15 are each independently selected from hydrogen or deuterium.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein R 4 , R 5 , R 10 , R 11 , R 12 , R 14 and R 16 Selected from deuterium, R 1 , R 2 , R 3 , R 6 , R 7 , R 8 and R 9 Selected from hydrogen, R 13 and R 15 are each independently selected from hydrogen or deuterium.

3. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein R 6 , R 7 , R 8 , R 9 , R 12 , R 14 and R 16 Selected from deuterium, R 1 , R 2 , R 3 , R 4 , R 5 , R 10 and R 11 Selected from hydrogen, R 13 and R 15 are each independently selected from hydrogen or deuterium.

4. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein The R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 10 , R 11 , R 12 , R 14 and R 16 Selected from deuterium, R 1 , R 2 and R 3 Selected from hydrogen, R 13 and R 15 are each independently selected from hydrogen or deuterium.

5. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein the compound is selected from: 。 6. A pharmaceutical composition comprising the compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof, optionally further comprising a pharmaceutically acceptable excipient.

7. Use of the compound according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 6, in the preparation of a medicament for treating a disease that benefits from the inhibition of HPK1 kinase activity.

8. The use according to claim 7, wherein The disease that benefits from the inhibition of HPK1 kinase activity is selected from tumors or cancers.

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