A PI3Kδ inhibitor and its synthesis method and application

Abstract

The present invention discloses a small molecule compound having PI3Kδ inhibitory activity as shown in formula (I), a method for synthesizing the compound, and application of the compound in preparing a pharmaceutical preparation for preventing or treating a disease caused by abnormal B cells. The disease is lymphoma, including chronic lymphocytic leukemia, B cell lymphoma, mantle cell lymphoma, lymphoplasmacytic lymphoma, diffuse large B cell lymphoma, non-Hodgkin's lymphoma, follicle center lymphoma, marginal zone B cell lymphoma, or an autoimmune disease, including rheumatoid arthritis, systemic lupus erythematosus, ankylosing spondylitis, psoriasis, etc.

Description

Technical Field

[0001] The present invention relates to the field of small molecule drugs. Specifically, the present invention provides a compound with phosphatidylinositol-3-kinase delta subtype (PI3Kδ) inhibitory activity, as well as a method for synthesizing and using the compound. Background Art

[0002] B-cell lymphoma is a heterogeneous group of lymphocyte proliferative diseases and a malignant tumor with a high mortality rate. It can be divided into two categories: Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL). HL usually occurs in lymph nodes and is characterized by Reed-Sternberg tumor cells; NHL often occurs in extranodal lymphoid tissues and has a generally higher incidence rate, including mantle cell lymphoma (MCL), diffuse large B-cell lymphoma (DLBCL), chronic lymphocytic leukemia (CLL), follicular lymphoma (FL), marginal zone B-cell lymphoma (MZL), etc. At present, the conventional treatment for B-cell lymphoma is radiation and chemotherapy. The NCCN Clinical Drug Guidelines in the United States point out that the first-line treatment for B-cell lymphoma is R-CHOP therapy, which is a combination of rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone. However, for elderly patients or patients with other diseases, the toxicity of these therapies is difficult for them to tolerate.

[0003] The pathogenesis of B-cell lymphoma is closely related to the B-cell receptor (BCR). In many B-cell lymphomas, BCR is overexpressed and the BCR signaling pathway is overactive, resulting in the inhibition of normal differentiation and apoptosis of B cells and the promotion of abnormal proliferation. Therefore, the BCR signaling pathway is crucial for the growth and survival of a variety of tumor cells. Different targeted drugs targeting this pathway can inhibit the progression of B-cell-related diseases, including spleen tyrosine kinase (SYK), Bruton's tyrosine kinase (BTK) and phosphatidylinositol-3-kinase (PI3K).

[0004] PI3K is a type of lipid kinase that participates in the regulation of physiological processes related to the occurrence and development of tumors. It plays an important role in cell growth, development, division, differentiation and apoptosis, and is closely related to the occurrence and development of tumors. According to the different structures and substrate specificities, PI3K can be divided into three types: type I, type II and type III. Among them, type I PI3K is the most intensively studied and widely studied subtype, and its relationship with tumors is also the closest, while type I PI3K is divided into type IA and type IB. The catalytic subunits of type IA PI3K include three proteins, p110α, p110β, and p110δ. The catalytic subunit of type IB PI3K is only one, p110γ. According to the different catalytic subunits, these PI3Ks are called PI3Kα, PI3Kβ, PI3Kδ, and PI3Kγ, respectively. In terms of expression, PI3Kα and PI3Kβ are expressed in a variety of cells, while PI3Kδ and PI3Kγ are only expressed in the immune system.

[0005] Figure 1 Statistics on the development of pan-PI3K inhibitors in the early years show that in 2012, there were 11 pan-PI3K inhibitors in the late stage of development, most of which had advanced to Phase 1 clinical trials; by 2017, 10 of the 11 products had terminated their development, and only copanlisib made it to the end and was approved for the treatment of relapsed follicular lymphoma in 2017. Among these pan-PI3K inhibitors that were discontinued, the main reason was the strong toxic side effects. The above facts show that the development of pan-PI3K inhibitors has obviously reached a dead end.

[0006] Faced with the dilemma of adverse reactions, researchers have turned to the development of subtype-specific PI3K inhibitors.

[0007] Currently, five PI3K inhibitors have been approved by the FDA for marketing. In July 2014, the first selective PI3Kδ inhibitor Idelalisib was approved by the FDA for marketing. Its selectivity is more than 40 times that of other subtypes and is mainly used to treat relapsed CLL, FL or SLL. However, Idelalisib has a black box warning, with serious side effects such as hepatotoxicity, severe diarrhea, colitis, pneumonia and intestinal perforation. In view of the serious adverse reactions of Idelalisib, in September 2017, the FDA accelerated the approval of Copanlisib for the treatment of FL. Copanlisib is a PI3Kα / δ inhibitor with similar therapeutic effects to Idelalisib, but its safety has been greatly improved. Duvelisib, which was approved for marketing in October 2018, is a PI3Kδ / γ selective inhibitor with an overall remission rate of 78%. Alpelisib is a PI3Kα inhibitor with high selectivity. It is mainly used to treat advanced metastatic breast cancer with PIK3CA mutations, filling the gap of poor treatment effect for breast cancer patients with PIK3CA mutations. Compared with the placebo group, Alpelisib tripled the survival period. The newly approved PI3Kδ / CK1ε dual-target inhibitor Umbralisib has been granted orphan drug status by the FDA for the treatment of FL / MZL.

[0008] At present, the global research on PI3K inhibitors is still very hot, but there are many complex mechanisms in the PI3K target, and each subtype has its own unique function, so inhibitors targeting different subtypes inhibit tumors through different mechanisms, and the toxicity is also superimposed. Therefore, PI3K inhibitors with high subtype selectivity will have more development prospects in terms of safety.

[0009] PI3Kδ is only expressed in hematopoietic cells, and studies have shown that B cell function is severely impaired in mice with PI3Kδ knockout. Therefore, PI3Kδ is an effective target for the treatment of B cell lymphoma. At the same time, PI3Kδ is also related to immune diseases. It can regulate immune thresholds and limit the immunity of endogenous microbiota. It is expected to be used to treat immune cell-related diseases such as rheumatoid arthritis, multiple sclerosis, and Alzheimer's disease. The more eye-catching second-generation PI3Kδ inhibitor Umbralisib has recently been launched in the United States for use in previously treated MZL and FL patients. Umbralisib has better tolerability and safety, and is less likely to develop autoimmune toxicity. Therefore, second-generation PI3K inhibitors have higher selectivity and reduce side effects caused by superimposed toxicity to a greater extent. The structures of the newer representative PI3Kδ inhibitors are shown below. Figure 2 shown.

[0010] However, the subtype selectivity of current PI3K inhibitors still needs to be improved. Summary of the invention

[0011] The object of the present invention is to provide a 1H-pyrazolo[3,4-d]pyrimidine-4-amine structure compound and its pharmaceutically acceptable salt, a synthesis method thereof and its application in drugs for inhibiting B protein receptors in view of the deficiencies of the prior art.

[0012] The object of the present invention is achieved by the following technical scheme: a compound having a 1H-pyrazolo[3,4-d]pyrimidin-4-amine structure and a pharmaceutically acceptable salt thereof, characterized in that they have a structure as shown in formula (I):

[0013]

[0014] Wherein, R has the following structure: methyl, ethyl, propyl, cyclopropyl, phenyl, methoxyphenyl, halogenated phenyl, methoxyhalogenated phenyl, phenoxyphenyl, pyridyl, methoxypyridyl, halogenated pyridyl, methoxyhalogenated pyridyl, or amide pyridyl.

[0015] Furthermore, the present invention provides a compound having a 1H-pyrazolo[3,4-d]pyrimidine-4-amine structure, which has the following structure:

[0016]

[0017] The compound having a 1H-pyrazolo[3,4-d]pyrimidine-4-amine structure described in the present invention can be used alone, or can be prepared into a pharmaceutically acceptable salt by a conventional method for use. The pharmaceutically acceptable salt is hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate, phosphate, acetate, propionate, butyrate, oxalate, tartrate, methanesulfonate, p-toluenesulfonate, fumarate, taurine, citrate, succinate, or a mixed salt thereof.

[0018] The present invention also provides a compound having a 1H-pyrazolo[3,4-d]pyrimidine-4-amine structure and a pharmaceutically acceptable salt thereof for use in medicine, the specific use of which is: for preparing a pharmaceutical preparation for preventing or treating a disease caused by abnormal PI3Kδ. The disease is lymphoma, including chronic lymphocytic leukemia, B-cell lymphoma, mantle cell lymphoma, lymphoplasmacytic lymphoma, diffuse large B-cell lymphoma, non-Hodgkin's lymphoma, follicular center lymphoma, marginal zone B-cell lymphoma, or an autoimmune disease, including rheumatoid arthritis, systemic lupus erythematosus, ankylosing spondylitis, psoriasis, etc.

[0019] The present invention also provides a method for preparing a compound having a 1H-pyrazolo[3,4-d]pyrimidine-4-amine structure, comprising the following steps:

[0020] Using 3-fluoro-4-chlorophenol as the starting material, an acetyl group is introduced through Fries rearrangement; A-2 is then obtained through Williamson ether synthesis; intermediate A-3 is obtained after NaBH4 reduction; intermediate A-4 is obtained by Mitsunobu reaction with iodinated pyrazolopyrimidine fragments; and finally, the final product is obtained through Suzuki coupling reaction with boric esters or boric acids containing different groups. The reaction equation is as follows:

[0021]

[0022] Beneficial effects of the present invention:

[0023] The compound with PI3Kδ activity provided by the present invention has good subtype selectivity and can therefore be widely used in the preparation of pharmaceutical preparations for preventing or treating diseases caused by abnormal B cell receptors. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 The research and development of pan-PI3K inhibitors in the early years;

[0025] Figure 2 It is a representative PI3Kδ inhibitor;

[0026] Figure 3 HH COSY spectrum (a) and NOESY spectrum (b) of intermediate A-1. DETAILED DESCRIPTION

[0027] The present invention is further described below by way of examples, but the present invention is not limited to the scope of the examples. The experimental methods in the following examples without specifying specific conditions are carried out according to conventional methods and conditions, or selected according to the product specifications.

[0028] Example 1 Synthesis of Compound 1

[0029] Step a. Synthesis of intermediate 4-fluoro-5-chloro-2-hydroxyacetophenone (A-1)

[0030]

[0031] In a 500mL single-necked bottle equipped with a condenser and a drying tube, 3-fluoro-4-chlorophenol (30g, 206mmol) and acetic anhydride (29mL, 310mmol) were added in sequence, and concentrated sulfuric acid (1mL, 22mmol) was slowly added, and the temperature was raised to 70°C for 50min, and the reaction was monitored by TLC. After the reaction was completed, the reaction solution was cooled to room temperature, saturated sodium carbonate solution (400mL) and ethyl acetate (400mL) were added and stirred for 10min, and the organic layer was separated after the reaction solution was cooled, and the aqueous phase was extracted with ethyl acetate, the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and the solvent was concentrated to obtain 39g of colorless oily 3-fluoro-4-chlorophenol methyl ester.

[0032] In a 500mL three-necked flask equipped with a condenser and a drying tube, add the phenol ester intermediate (39g, 209mmol) and nitrobenzene (8mL, 65mmol) in sequence, slowly add aluminum chloride (33g, 248mmol) in three batches, each time with an interval of 5min, and heat and stir at 120℃ for 3h. After the reaction is completed by TLC monitoring, cool the reaction solution to 0℃, slowly add 140mL of ice water to the reaction flask to quench the aluminum chloride, and HCl gas is released during the process. Add dichloromethane (500mL) and 1N hydrochloric acid (300mL) to the reaction flask, stir at room temperature for 20min to dissolve the insoluble solid. Separate the organic layer and wash with 1N hydrochloric acid (200mL) and water (200mL) respectively. The organic phase was extracted with 1N sodium hydroxide (500 mL×3) and adjusted to pH>9. All aqueous phases were combined and the pH was adjusted to pH<2 with concentrated hydrochloric acid until no more precipitate was precipitated. Finally, the solid was filtered and washed with water twice to obtain 33 g of off-white solid with a yield of 84%. 1 H NMR (400 MHz, CDCl 3 )δ12.43(s,1H),7.78(d,J=8.1Hz,1H),6.77(d,J=10.2Hz,1H),2.61(s,3H); ESI-MS: m / z=189.46[M+H] + .

[0033] Fries rearrangement is dominated by kinetics at low temperatures, mainly rearranging to the para position, and thermodynamics at high temperatures, rearranging to the ortho position. The final product may have two isomers, namely intermediate A-1 and byproduct A-1b, so it is necessary to use 2D-NMR spectrum to analyze. The hydrogens on the benzene ring of intermediate A-1 will not couple with each other, while byproduct A-1b will couple. Figure 3 The HH COSY and NOESY spectra confirmed that the obtained product was A-1.

[0034]

[0035] Structure of intermediate A-1 and possible by-product A-1b Step b. Synthesis of intermediate 4-fluoro-5-chloro-2-ethoxyacetophenone (A-2)

[0036]

[0037] The intermediate A-1 (19 g, 100 mmol) obtained by the above reaction was put into a 500 mL single-necked flask, and N, N-dimethylformamide (100 mL) was used as a solvent. After dissolution, potassium carbonate (28 g, 199 mmol) and iodoethane (10 mL, 120 mmol) were added in sequence. The reaction solution was stirred at room temperature for 45 min, and then heated to 60 ° C for 3 h. After the reaction was completed by TLC monitoring, the heating was stopped, 300 mL of water was added to the reaction bottle, and stirred at room temperature for 1 h. The unreacted phenol salt was dissolved in water, and the product was precipitated in large quantities because it was insoluble in water. The solid was filtered, the filtrate was retained, the filter cake was rinsed with water twice, and then the filter cake was dissolved in ethyl acetate (400 mL), washed twice with 1N hydrochloric acid (400 mL × 2), washed once with saturated brine (400 mL), dried over anhydrous sodium sulfate, filtered, and the solvent was dried to obtain a solid product. The filtrate was extracted with ethyl acetate (400 mL x 2) and water (400 mL), the organic phases were combined, washed with saturated brine, and the solvent was concentrated to obtain a portion of the product. The combined A-2 crude product was separated by silica gel column chromatography to obtain 20 g of off-white solid, with a yield of 92%. 1 H NMR (400 MHz, CDCl 3 )δ7.86(d,J=8.8Hz,1H),6.75(d,J=10.7Hz,1H),4.11(q,J=7.0Hz,2H),2.61(s,3H),1.51(t,J=7.0Hz,3H); ESI-MS: m / z=216.12[M+H] + .

[0038] Step c. Synthesis of intermediate 4-fluoro-5-chloro-2-ethoxyphenylethanol (A-3)

[0039]

[0040] A-2 (11 g, 52 mmol) was added to a 500 mL single-necked bottle equipped with a balloon, dissolved in anhydrous methanol (100 mL), and sodium borohydride (4 g, 104 mmol) was slowly added under an ice bath, and stirred at room temperature for 30 min. After the reaction was completed by TLC monitoring, water was added under an ice bath to quench until no more bubbles were generated. Methanol was removed by rotary evaporation, and extracted with ethyl acetate (200 mL × 2) and water (100 mL), the organic phases were combined, washed with saturated brine (150 mL), and the solvent was concentrated and the product was separated by silica gel column chromatography to obtain 11 g of light yellow transparent oil with a yield of 96%. 1H NMR (400 MHz, CDCl 3 )δ7.30(d,J=8.5Hz,1H),6.59(d,J=10.9Hz,1H),4.99(q,J=6.5Hz,1H),3.96(q,J=7.0Hz,2H),1.39(t,J=7.0Hz,3H),1.38(d,J=12.0Hz,3H).

[0041] Step d. Synthesis of intermediate 1-(1-(2-ethoxy-4-fluoro-5-chlorophenyl)ethyl)-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (A-4)

[0042]

[0043] 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (7g, 27mmol) and anhydrous tetrahydrofuran (80mL) were added to a 250mL three-necked flask equipped with a rubber stopper, a 10mL dropping funnel, and a nitrogen balloon. The mixture was heated to dissolve as much as possible, and then intermediate A-3 (9g, 41mmol) and triphenylphosphine (14g, 55mmol) were added in sequence. The nitrogen atmosphere was replaced three times, and diisopropyl azodicarboxylate (11mL, 55mmol) was slowly added dropwise under an ice bath. After the addition was completed, the mixture was stirred at room temperature for 1h and the reaction was monitored. If the raw material was not completely converted, the same amount of intermediate A-3, TPP and DIAD were added continuously, and the mixture was stirred at room temperature for 1h. The reaction was monitored by TLC, and most of the reaction was completed. Then add an appropriate amount of water to quench the reaction, remove tetrahydrofuran by rotary evaporation, extract the reactant with dichloromethane (200mL×2) and water (200mL), combine the organic phases, wash with saturated sodium chloride solution (200mL), dry with anhydrous sodium sulfate, filter and concentrate the solvent. Since the product and the by-product triphenylphosphine oxide have similar polarity, the crude product is separated by silica gel column chromatography to obtain a mixture of the two. Add anhydrous ethanol (40mL) to the mixture to form a homogenate, stir at room temperature for 8h, filter the solid after beating, and place it in a vacuum drying oven to dry for 48h to obtain 7g of white solid, with a yield of 57%. 1 H NMR(400MHz,THF)δ8.03(s,1H),7.23(d,J=8.5Hz,1H),6.78(d,J=11.2Hz,1H),6.32(q,J=7.0Hz,1H), 3.92(dd,J=17.1,7.0Hz,2H),1.68(d,J=7.1Hz,3H),1.24(t,J=7.0Hz,3H); ESI-MS:m / z=462.00[M+H] + .

[0044] Step e. Synthesis of target compound 1 (R = methyl)

[0045]

[0046] The intermediate A-4 (400 mg, 0.9 mmol) obtained in the previous step and potassium methyl fluoroborate (262 mg, 2 mmol) were put into a 100 mL three-necked flask, dissolved in a 1:1 mixed solvent (8 mL) of tetrahydrofuran and water, added cesium carbonate (840 mg, 2 mmol) to the reaction flask, the air in the reaction flask was purged with nitrogen, 1,1'-bis(diphenylphosphino)ferrocenepalladium dichloride (63 mg, 0.09 mmol) was added, the air was continued to be purged with nitrogen for 5 min, and heated to reflux at 100 ° C for 5 h. After the reaction was completed as monitored by TLC, the reaction solution was filtered through diatomaceous earth, rinsed with ethyl acetate, and the organic solvent was removed by rotary evaporation. The reaction solution was extracted with ethyl acetate (100 mL×2) and water (100 mL), the organic phases were combined, washed with saturated brine (200 mL), dried over anhydrous sodium sulfate, filtered, and the solvent was concentrated. The product was separated by silica gel column chromatography, and the product was separated again by silica gel preparation plate. 47 mg of white solid was obtained with a yield of 16%. 1 H NMR (400 MHz, CDCl 3 )δ8.31(s,1H),7.26–7.21(m,2H),6.62(d,J=10.8Hz,1H),6.37(q,J=6.6Hz,1H),5.56(s,1 H),3.97(dd,J=15.4,6.8Hz,2H),2.65(s,2H),1.83(d,J=6.9Hz,3H),1.35(t,J=6.8Hz,3H); 13 CNMR(101MHz,THF)δ158.97,158.02,156.52,155.99,155.57,153.73,128.29,127.69, 110.83,103.78,101.04,86.50,64.59,49.39,19.38,13.85; ESI-MS: m / z=350.12[M+H] + .

[0047] Example 2 Synthesis of Compound 2

[0048] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,dioxane:H 2 O=4:1,N 2 ,100℃. 1H NMR (400MHz, DMSO) δ8.12(s,1H),7.14(d,J=11.5Hz,1H),7.09(d,J=8.6Hz,1H),6.19(q,J=7.0H z,1H),4.06(dd,J=14.0,7.0Hz,2H),2.52(s,4H),1.69(d,J=7.0Hz,3H),1.29(t,J=6.9Hz,3H); 13 C NMR (101MHz, DMSO) δ158.69,156.12,155.49,154.13,146.35,128.85,127.86,110.11,1 02.36,102.11,99.54,65.03,48.78,20.30,14.75,9.09,8.25; ESI-MS: m / z=375.13[M+H] + .

[0049] Example 3 Synthesis of Compound 3

[0050] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,THF:H 2 O=1:1,N 2 ,80℃. 1 H NMR (400 MHz, CDCl 3 )δ8.35(s,1H),7.61(d,J=8.6Hz,2H),7.36(d,J=8.4Hz,1H),7.04(d,J=8.6Hz,2H),6.64(d,J=10.9Hz,1H),6.50(q,J =7.0Hz,1H),5.71(s,2H),4.69–4.55(m,1H),4.00(dd,J=12.9,6.9Hz,2H),1.90(d,J=7.0Hz,3H),1.42–1.34(m,9H); 13 C NMR (101 MHz, CDCl 3 )δ185.18,167.72,158.79,157.48,156.54,155.17,154.94,153.95,144.42,132.07,130.91,129.86,128.85, 127.37,125.23,116.56,111.49,100.61,70.10,65.57,64.55,22.01,20.16,14.54; ESI-MS: m / z=470.17[M+H] +.

[0051] Example 4 Synthesis of Compound 4

[0052] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,THF:H 2 O=1:1,N 2 ,80℃. 1 H NMR (400 MHz, CDCl 3 )δ8.36(s,1H),7.52–7.39(m,2H),7.33(d,J=8.4Hz,1H),7.13(d,J=8.5Hz,1H),6.65(d,J=10.8Hz,1H),6.50 (d,J=7.0Hz,1H),5.76(s,2H),4.07–3.98(m,2H),3.97(s,3H),1.89(d,J=7.0Hz,3H),1.38(t,J=6.9Hz,3H); 13 C NMR (101MHz, DMSO) δ158.76,158.41,156.33,155.74,155.65,154.10,137.50,132.50,132.00,131.91,129.28,129.16,128.28,128.11 ,125.78,114.71,114.59,110.30,110.12,109.88,102.49,102.24,99.20,65.09,57.14,49.24,20.29,14.67; ESI-MS: m / z=460.13[M+H] + .

[0053] Example 5 Synthesis of Compound 5

[0054] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,THF:H 2 O=1:1,N 2 ,80℃. 1 H NMR (400 MHz, CDCl 3)δ8.38(s,1H),7.35(d,J=8.2Hz,2H),6.65(d,J=10.8Hz,1H),6.53(q,J=6.9Hz,1H),5 .60(s,2H),3.99(dd,J=15.4,7.1Hz,2H),1.90(d,J=7.0Hz,3H),1.36(t,J=6.9Hz,3H); 13 C NMR (101 MHz, CDCl 3 )δ159.08,157.60,156.62,155.73,155.29,155.21,154.05,149.99,136.88,128.64,127.05,127.01,126.35,125.01,123. 55,123.44,118.42,118.25,111.56,111.39,100.90,100.65,99.78,64.57,49.60,20.09,14.47; ESI-MS: m / z=448.11[M+H] + .

[0055] Example 6 Synthesis of Compound 6

[0056] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,THF:H 2 O=1:1,N 2 ,80℃. 1 H NMR(400MHz,DMSO)δ8.23(s,1H),7.31–7.26(m,2H),7.20–7.12(m,2H),6.36(q,J=6.9Hz,1 H),4.06(dd,J=18.4,7.0Hz,2H),3.95(s,3H),1.79(d,J=7.0Hz,3H),1.25(t,J=7.0Hz,3H); 13C NMR (101MHz, DMSO) δ158.76,158.41,156.33,155.74,155.65,154.10,137.50,132.50,132.00,131.91,129.28,129.16,128.28,128.11 ,125.78,114.71,114.59,110.30,110.12,109.88,102.49,102.24,99.20,65.09,57.14,49.24,20.29,14.67; ESI-MS: m / z=478.12[M+H] + .

[0057] Example 7 Synthesis of Compound 7

[0058] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,THF:H 2 O=1:1,N 2 ,80℃. 1 H NMR (400 MHz, CDCl 3 )δ8.38(s,1H),7.34(d,J=8.3Hz,2H),7.22–7.10(m,1H),6.64(d,J=10.8Hz,1H),6.53(q,J=7.0 Hz,1H),5.65(s,2H),3.99(dd,J=16.3,7.0Hz,2H),1.89(d,J=7.0Hz,3H),1.36(t,J=7.0Hz,3H); 13 C NMR (101 MHz, CDCl 3 )δ159.10,157.57,156.64,155.73,155.30,155.22,154.05,150.57,139.33,136.23,131.95,128.62,126.93,125.20, 118.92,113.25,113.07,111.56,111.39,100.92,100.67,99.74,64.57,49.60,20.07,14.47; ESI-MS: m / z=466.11[M+H] + .

[0059] Example 8 Synthesis of Compound 8

[0060] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K2 CO 3 ,THF:H 2 O=1:1,N 2 ,80℃. 1 H NMR (400 MHz, CDCl 3 )δ8.39(s,1H),7.38(t,J=6.7Hz,2H),7.32(d,J=8.3Hz,1H),6.65(d,J=10.8Hz,1H),6.52(dd,J=13.5 ,6.6Hz,1H),5.74(s,2H),3.99(dd,J=15.7,7.3Hz,2H),1.89(d,J=6.9Hz,3H),1.37(t,J=6.8Hz,3H); 13 C NMR (101 MHz, CDCl 3 )δ159.13,157.49,156.66,155.85,155.34,155.25,154.40,152.91,150.44,141.19,128.61,126.77,112. 98,112.76,111.56,111.39,100.95,100.71,98.26,64.59,49.51,20.04,14.51; ESI-MS: m / z=466.10[M+H] + .

[0061] Example 9 Synthesis of Compound 9

[0062] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,THF:H 2 O=1:1,N 2 ,100℃. 1 HNMR (400MHz, DMSO) δ8.84(s,1H),8.68(d,J=3.3Hz,1H),8.27(s,1H),8.04(d,J=7.9Hz,1H),7.57(d,J=7.8Hz,1H),7.34(d,J=8.5 Hz,1H),7.16(d,J=11.5Hz,1H),6.38(q,J=7.0Hz,1H),4.05(dd,J=17.5,7.0Hz,2H),1.81(d,J=7.0Hz,3H),1.24(t,J=7.0Hz,4H); 13C NMR(101MHz,DMSO)δ158.71,156.31,155.82,155.73,154.60,149.90,149.23,141.48,136.25,129.33, 128.23,128.04,124.52,110.28,102.24,98.12,65.07,49.29,20.26,14.72; ESI-MS: m / z=413.12[M+H] + .

[0063] Example 10 Synthesis of Compound 10

[0064] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,THF:H 2 O=1:1,N 2 ,100℃. 1 HNMR (400MHz, DMSO) δ8.38(s,1H),8.24(s,1H),8.09(s,1H),7.51(s,1H),7.28(d,J=8.4Hz,1H),7.17(d,J=11.4Hz, 1H), 6.37 (q, J = 6.4Hz, 1H), 4.05 (dd, J = 19.0, 7.0Hz, 2H), 3.37 (s, 2H), 1.79 (d, J = 6.8Hz, 3H), 1.24 (t, J = 6.7Hz, 3H); 13 C NMR (101MHz, DMSO) δ161.79,158.46,157.53,156.40,155.77,154.22,148.43,143.01,137.20,128.09,122 .92,116.38,116.08,110.29,102.48,102.23,99.21,65.06,49.37,20.23,14.67; ESI-MS: m / z=431.11[M+H] + .

[0065] Example 11 Synthesis of Compound 11

[0066] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,THF:H 2 O=1:1,N 2 ,80℃. 1H NMR (400MHz, DMSO) δ8.37(d,J=5.1Hz,1H),8.29(s,1H),7.61(d,J=3.7Hz,1H),7.37(s,1H),7.33(d,J=8.5Hz,1H),7.1 6(d,J=11.5Hz,1H),6.39(q,J=6.9Hz,1H),4.03(dd,J=18.8,7.0Hz,2H),1.82(d,J=7.0Hz,3H),1.22(t,J=6.9Hz,4H); 13 C NMR (101MHz, DMSO) δ165.13,158.54,156.40,155.88,154.75,148.57,146.17,141.02,131.91,128.25, 127.72,121.66,110.31,109.52,102.48,98.08,65.05,49.53,20.14,14.68; ESI-MS: m / z=431.11[M+H] + .

[0067] Example 12 Synthesis of Compound 12

[0068] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,THF:H 2 O=1:1,N 2 ,80℃. 1 H NMR (400 MHz, CDCl 3 )δ8.43(s,1H),8.30(s,1H),7.85(d,J=10.3Hz,1H),7.30(d,J=8.5Hz,1H),6.85(d,J=8.5Hz,1H),6.57(d,J=10.8Hz,1 H),6.45(dd,J=13.8,6.8Hz,1H),5.73(s,2H),4.24(s,2H),3.94(s,3H),1.82(d,J=7.0Hz,3H),0.89(t,J=7.3Hz,11H); 13 CNMR (101MHz, CDCl 3)δ166.70,163.62,156.32,154.27,153.83,153.00,145.54,140.38,137.78,131.31,129.89,127 .82,123.45,121.50,110.71,99.90,64.55,52.79,29.56,18.17,12.71; ESI-MS:m / z=443.13[M+H] + .

[0069] Example 13 Synthesis of Compound 13

[0070] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3 ,dioxane:H 2 O=2:1,N 2 ,80℃. 1 H NMR (400 MHz, CDCl 3 )δ8.21(s,1H),7.57(d,J=8.7Hz,2H),7.34–7.30(m,2H),7.28(d,J=6.3Hz,1H),7.12–7.08(m,2H),7.06(s, 1H),7.01(dd,J=8.6,1.0Hz,2H),3.91(dd,J=14.2,7.0Hz,3H),1.82(d,J=7.1Hz,4H),1.30(t,J=7.0Hz,3H); 13 C NMR (101 MHz, CDCl 3 )δ158.03,157.59,156.54,155.56,155.29,154.21,153.05,152.66,143.28,131.02,129.01,128.95,127.66,127.46,126 .58,126.02,123.03,118.53,118.10,110.48,99.85,99.61,97.24,63.52,48.37,19.07,13.50; ESI-MS: m / z=504.15[M+H] + .

[0071] Example 14 Synthesis of Compound 14

[0072] The operation was the same as that of compound 1. Conditions: Pd(dppf)Cl 2 ,K 2 CO 3,dioxane:H 2 O=2:1,N 2 ,80℃. 1 H NMR (400 MHz, CDCl 3 )δ9.74(s,1H),8.67(s,1H),8.43(d,J=2.5Hz,1H),8.39(s,1H),8.31(s,1H), 8.12(d,J=8.2Hz,2H),7.90(d,J=8.2Hz,2H),7.71–7.64(m,1H),7.46(ddd,J= 7.3,5.3,2.3Hz,1H),7.37(d,J=8.4Hz,1H),6.55(q,J=7.0Hz,1H),5.67(s,2H ), 4.01 (dd, J = 13.4, 6.9 Hz, 2H), 1.92 ( d, J = 7.0 Hz, 3H), 1.38 ( t, J = 7.0 Hz, 3H); 13 C NMR (101 MHz, CDCl 3 )δ164.76,159.10,157.51,156.63,155.55,155.22,154.29,148.17,142.90,142.13,140.63,137.58,137.30,133.67,132. 14,132.04,129.06,128.64,128.40,126.88,100.95,100.70,98.52,64.59,49.55,20.29,14.50; ESI-MS: m / z=532.16[M+H] + .

[0073] Example 15 Preparation of hydrochloride (taking the compound of Example 12 as an example)

[0074] Take an appropriate amount of the compound, dissolve it in ethanol, slowly introduce HCl gas at room temperature until saturation, cool it in an ice bath, and slowly precipitate white crystals, which are the hydrochloride of the compound.

[0075] Example 16 PI3K kinase inhibition experiment

[0076] The inhibitory activity of PI3K kinase was detected by ADP-Glo ​​method:

[0077] Experimental process: Add 2.5μL of kinase solution at 2 times the final concentration to each well, centrifuge at 1000rpm for 30s, shake and mix, and incubate at room temperature for 10min. Then add 2.5μL of a mixed solution of ATP and substrate at 2 times the final concentration, centrifuge for 30s, shake and mix, and start the reaction at room temperature. Add 5μL ADP-Glo ​​Reagent, centrifuge for 30s, shake and mix, and incubate at room temperature for 180min. Add 10μL Kinase Detection Reagent, centrifuge, shake and mix, and incubate at room temperature for 30min. Finally, use an enzyme reader to read the luminescence value RLU. The results are as follows:

[0078] Table. In vitro inhibitory activity of compounds 1-14 (IC 50 ,nM))

[0079]

[0080]

[0081] It can be found that: the inhibitory activity of this series of compounds for PI3Kδ is generally good, among which compounds 4, 6, and 12 have good activity, up to 34.9nM. More importantly, this type of compound has good subtype selectivity, and does not show activity at the highest experimental concentration (10000nM) for the other three subtypes of PI3K α, β, and γ. Therefore, the compounds provided by the present invention may reduce toxic side effects, thereby being used in the preparation of pharmaceutical preparations for preventing or treating diseases caused by abnormal B cells. The disease is lymphoma, including chronic lymphocytic leukemia, B cell lymphoma, mantle cell lymphoma, lymphoplasmacytic lymphoma, diffuse large B cell lymphoma, non-Hodgkin's lymphoma, follicular center lymphoma, marginal zone B cell lymphoma, or autoimmune diseases, including rheumatoid arthritis, systemic lupus erythematosus, ankylosing spondylitis, psoriasis, etc.

Claims

1. A PI3Kδ inhibitor, characterized in that It is a compound having a 1H-pyrazolo[3,4-d]pyrimidin-4-amine structure, having the following structures:

2. The pharmaceutically acceptable salt of the PI3Kδ inhibitor according to claim 1, characterized in that: The pharmaceutically acceptable salt is hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate, phosphate, acetate, propionate, butyrate, oxalate, tartrate, methanesulfonate, p-toluenesulfonate, fumarate, taurate, citrate, succinate, or a mixed salt thereof.

3. The method for synthesizing a PI3Kδ inhibitor according to claim 1, characterized in that: The following steps are involved: Using 3-fluoro-4-chlorophenol as the starting material, an acetyl group is introduced through Fries rearrangement to obtain A-1; A-2 is then obtained through Williamson ether synthesis; intermediate A-3 is obtained after NaBH4 reduction; intermediate A-4 is obtained by Mitsunobu reaction with iodinated pyrazolopyrimidine fragments; and finally, a compound having a 1H-pyrazolo[3,4-d]pyrimidin-4-amine structure is obtained through Suzuki coupling reaction with boric esters or boric acids containing different groups.

4. Use of the PI3Kδ inhibitor according to claim 1 in the preparation of a pharmaceutical preparation for preventing or treating diseases caused by abnormal PI3Kδ protein.

5. The use according to claim 4, characterized in that: The disease is lymphoma or an autoimmune disease.

6. The use according to claim 5, characterized in that: The lymphoma is chronic lymphocytic leukemia, B-cell lymphoma, mantle cell lymphoma, lymphoplasmacytic lymphoma, non-Hodgkin's lymphoma or follicular center lymphoma; The autoimmune disease is rheumatoid arthritis, systemic lupus erythematosus, ankylosing spondylitis or psoriasis.

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