WO2022257568A1 - 氮杂吲哚嘧啶胺杂环化合物及其制备方法和应用 - Google Patents

氮杂吲哚嘧啶胺杂环化合物及其制备方法和应用 Download PDF

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WO2022257568A1
WO2022257568A1 PCT/CN2022/084057 CN2022084057W WO2022257568A1 WO 2022257568 A1 WO2022257568 A1 WO 2022257568A1 CN 2022084057 W CN2022084057 W CN 2022084057W WO 2022257568 A1 WO2022257568 A1 WO 2022257568A1
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cancer
hydrogen
compound
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杨鹏
袁凯
陈玮娇
邝文彬
姬明慧
王晓
王丽萍
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China Pharmaceutical University
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia

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  • the invention relates to the field of medicinal chemistry, in particular to an azaindopyrimidine amine heterocyclic compound and a preparation method and application thereof.
  • CDKs Cyclin-dependent kinases
  • the present invention provides an azaindopyrimidine amine heterocyclic compound, which can inhibit CDK6.
  • the invention also provides the preparation method and application of the compound.
  • the X is selected from C(O) or (CH 2 ) n ; n is 0-8;
  • R 1 is selected from hydrogen or halogen
  • the R 2 is selected from hydrogen, C 1 -C 8 alkyl or C 3 -C 6 cycloalkyl;
  • the R 3 is selected from hydrogen, halogen or C 1 -C 3 alkoxy
  • the R 4 is selected from hydrogen, C 1 -C 8 alkyl, -S(O) 2 C 1 -C 3 alkyl or -C(O)OC 1 -C 6 alkyl.
  • the X is C(O) or (CH 2 ) n ; n is 0-4.
  • said R 1 is hydrogen or fluorine.
  • the R 2 is hydrogen, C 1 -C 4 alkyl, cyclopentyl or cyclohexane; in some specific embodiments, the R 2 is hydrogen, methyl radical, ethyl, isopropyl, isobutyl or cyclopentyl.
  • the R 3 is hydrogen, fluoro or methoxy.
  • the R 4 is hydrogen, C 1 -C 3 alkyl, -S(O) 2 C 1 -C 3 alkyl or -C(O)OC 1 -C 4 alkyl ; In some more specific embodiments, said R 4 is hydrogen, methyl, ethyl, isopropyl or tert-butoxycarbonyl.
  • said X is selected from: (CH 2 ) n or C(O), n is 0 or 1; said R 1 is selected from: hydrogen, fluorine; said R 2 Selected from: hydrogen, methyl, ethyl, isopropyl, isobutyl, cyclopentyl; said R3 is selected from: hydrogen, fluorine or methoxy ; said R4 is selected from: hydrogen, methyl , ethyl, isopropyl or tert-butoxycarbonyl.
  • the present invention also provides compounds selected from S-1 to S-44 or pharmaceutically acceptable salts thereof:
  • compositions wherein the acid used for salt formation includes inorganic acid and organic acid, and described inorganic acid comprises: hydrochloric acid, sulfuric acid, phosphoric acid and methanesulfonic acid, Organic acids include acetic acid, trichloroacetic acid, propionic acid, butyric acid, maleic acid, p-toluenesulfonic acid, malic acid, malonic acid, cinnamic acid, citric acid, fumaric acid, camphoric acid, digluconic acid, aspartic acid and tartaric acid.
  • organic acids include acetic acid, trichloroacetic acid, propionic acid, butyric acid, maleic acid, p-toluenesulfonic acid, malic acid, malonic acid, cinnamic acid, citric acid, fumaric acid, camphoric acid, digluconic acid, aspartic acid and tartaric acid.
  • the pharmaceutically acceptable salt described in the present invention is hydrochloride.
  • the preparation method of the compound of general formula (S) of the present invention prepares compound (S) through coupling reaction by compound (A) and compound (B) under the effect of palladium catalyst:
  • the X is selected from C(O) or (CH 2 ) n ; n is 0-8;
  • R 1 is selected from hydrogen or halogen
  • the R 2 is selected from hydrogen, C 1 -C 8 alkyl or C 3 -C 6 cycloalkyl;
  • the R 3 is selected from hydrogen, halogen or C 1 -C 3 alkoxy
  • the R 4 is selected from hydrogen, C 1 -C 8 alkyl, -S(O) 2 C 1 -C 3 alkyl or -C(O)OC 1 -C 6 alkyl.
  • the reaction system of compound (A) and compound (B) is as follows: the reaction reagents are Pd 2 (dba) 3 , BINAP and cesium carbonate, under an inert gas atmosphere, the reaction temperature is 95-105°C, and the reaction time is 10-24h.
  • the reaction reagents are Pd 2 (dba) 3 , BINAP and cesium carbonate, under an inert gas atmosphere, the reaction temperature is 95-105°C, and the reaction time is 10-24h.
  • the present invention also provides a pharmaceutical composition of the compound as described in formula (S) or its pharmaceutically acceptable salt, ester, stereoisomer, solvate or prodrug, and a pharmaceutically acceptable carrier.
  • a pharmaceutically acceptable carrier refers to an excipient or diluent that does not cause significant irritation to the organism and does not interfere with the biological activity and properties of the administered compound.
  • the present invention also provides the use of a compound of formula (S) or its pharmaceutically acceptable salt, ester, stereoisomer, solvate or prodrug or the composition described in the present invention in the preparation of medicine.
  • the present invention also provides a compound of formula (S) or its pharmaceutically acceptable salt, ester, stereoisomer, solvate or prodrug or the use of the composition described in the present invention in the preparation of CDK6 inhibitors;
  • the CDK6 kinase inhibitor is used for treating cancer or tumor-related diseases.
  • Cancer or tumor related diseases including but not limited to multiple myeloma, leukemia, breast cancer, prostate cancer, lung cancer, liver cancer, stomach cancer, bone cancer, brain cancer, head and neck cancer, bowel cancer, pancreatic cancer, bladder cancer, testicular cancer, ovarian cancer cancer and endometrial cancer.
  • the compound of the general formula (S) or the pharmaceutically acceptable salt thereof in the present invention has CDK6 target inhibitory activity, and has a therapeutic effect on cell malignant proliferation tumors.
  • Me is methyl
  • Et is ethyl
  • Boc is tert-butoxycarbonyl
  • alkyl means a straight or branched chain saturated hydrocarbon group having the stated number of carbon atoms.
  • C 1 -C 8 alkyl refers to a linear or branched saturated hydrocarbon group having 1 to 8 carbon atoms.
  • C 1 -C 8 Alkyl groups include but are not limited to methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl base, isohexyl, 2,2-dimethylbutyl and 2,3-dimethylbutyl, etc.
  • C 3 -C 6 cycloalkane includes cyclopropanyl, cyclobutanyl, cyclopentyl and cyclohexyl.
  • C 1 -C 3 alkyl refers to a linear or branched saturated hydrocarbon group having 1 to 3 carbon atoms.
  • alkoxy means O-alkyl.
  • C 1 -C 3 alkoxy means having an O C 1 -C 3 alkyl group.
  • halogen means fluorine, chlorine, bromine or iodine. Preferred are fluorine, chlorine, bromine.
  • the compound represented by the general formula (S) of the present invention can inhibit the malignant proliferation of cancer, has good therapeutic effect, low toxicity, has good drug metabolism characteristics, and is not easy to produce drug resistance, and can be used to prepare and treat cancer or tumor-related diseases drug.
  • Reactants (ZA) and reactants (ZB) can be purchased directly or developed independently, which can significantly reduce costs.
  • the specific preparation methods of the self-developed reactant (ZA) and reactant (ZB) are as follows:
  • 3-Bromo-7-azaindole (3.94g, 20mmol) was dissolved in DMF (100mL), and NaH (1.6g, 40mmol) was added under ice-bath conditions, and after 30 minutes of reaction, iodoisopropane (1.36 g, 80 mmol), reacted at room temperature for 8 hours, quenched with water in an ice bath, extracted with dichloromethane, filtered and concentrated the solvent, and purified by flash silica gel column to obtain compound ZA9 (4.21 g, yield 88%).
  • Step 1 the synthesis of tert-butyl 4-(6-nitropyridin-3-yl)piperazine-1-carboxylate: Weigh 5-bromo-2-nitropyridine (0.41g, 2.0mmol), tert Butylpiperazine-1-carboxylate (0.48g, 2.6mmol) and triethylamine (0.41g, 4.0mmol) were dissolved in DMSO (5mL), heated to 60°C, and reacted for 18 hours. Concentrated by cold filtration and flash silica gel column purification to obtain compound tert-butyl 4-(6-nitropyridin-3-yl)piperazine-1-carboxylate (0.49 g, yield 80%).
  • Step 2 the synthesis of tert-butyl 4-(6-aminopyridin-3-yl)piperazine-1-carboxylate: Weigh tert-butyl 4-(6-nitropyridin-3-yl)piperazine -1-carboxylate (0.31g, 1.0mmol), reduced iron powder (0.17g, 3.0mmol) and ammonium chloride (0.49g, 9.0mmol) were dissolved in 70% ethanol (10mL), heated to 70°C, and reacted 6 hours. Concentrated by cold filtration and flash silica gel column purification to obtain compound ZB8 (0.24 g, yield 85%).
  • Example 3 The compound of Example 3 (120 mg) was dissolved in dichloromethane (40 mL), and HCl gas was passed through at 0° C. for 2 h. After the reaction was completed, it was concentrated to obtain compound S4 with a yield of 100% as a pale yellow solid.
  • Eu can transfer energy to the adjacent fluorescent substance Ulight acceptor, and then detect the emitted light.
  • the compound was dissolved and diluted to 2 mM with DMSO, and then 3-fold diluted to 10 concentrations, so that the compound concentration in the final experimental plate was 10 ⁇ M to 0.508 nM.
  • CDK6/cyclinD1 Life Technologies
  • Ulight-4E-BPI-peptide substrate PerkinElmer
  • reaction buffer 50mM Hepes pH 7.5, 10mM MgCl 2 , 0.01% Birj-35, 1mM EDTA, 2mM DTT
  • 5 ⁇ l of polypeptide and kinase mixture was mixed with 100 ⁇ L of compounds of different concentrations in an OptiPlate-384 microwell plate. After incubation at room temperature for 15 minutes, 5 ⁇ L of 350 ⁇ M ATP solution was added and incubated at room temperature for 90 min.
  • the measured IC 50 values are shown in Table 1 below. From the experimental results, it can be seen that the example compounds of the present invention have strong inhibitory activity on CDK6 kinase activity.
  • Example IC 50 (nM) Example IC 50 (nM) Example IC 50 (nM) Example IC 50 (nM) 1 130 16 30 31 32 2 120 17 10 32 42 3 >10000 18 >10000 33 27 4 251 19 33 34 45 5 >10000 20 35 35 36 6 56 twenty one 25 36 2622 7 46 twenty two 38 37 2200 8 440 twenty three >1000 38 1869 9 >10000 twenty four 18 39 36 10 905 25 >10000 40 46 11 >10000 26 86 41 25 12 445 27 53 42 >1000 13 113 28 45 43 69 14 126 29 40 44 twenty four 15 87 30 >1000 / /
  • the compounds of the present invention have very good inhibitory activity on CDK6 kinase, among which Example 17 has the strongest inhibitory activity on CDK6, and it will be further evaluated for in vivo activity.
  • Test animals ICR mice (provided by Shanghai Slack Experimental Animal Co., Ltd.); 18-22 g; female; 20 in total.
  • Group dose setting There are four groups in total, control group; 1000mg/kg group; Group 5 mice, observed for 14 days;
  • Test sample Example 17 and positive drug Palbociclib
  • embodiment 17 dosage is 100mg/kg, every day is given intragastric administration 1 time, when administration 23 days, to human multiple myeloma cell PRMI8226 nude mouse transplanted tumor T/C (%) is 58.3%, The tumor inhibition rate is 34.1%; the dose of embodiment 17 is 200mg/kg, administered by intragastric administration 1 time every day, and when administered for 23 days, the T/C (%) to human multiple myeloma cell PRMI8226 nude mouse transplanted tumor is 33.8%, tumor inhibition rate was 61.5%.
  • the dose of the positive drug Palbociclib is 100mg/kg, administered once a day by intragastric administration, and when administered for 23 days, the T/C (%) of the human multiple myeloma cell PRMI8226 nude mouse transplanted tumor was 37.9%, and the tumor inhibition rate was 55.6%.
  • the experimental results showed that the body weight of the nude mice in each group did not decrease significantly during the administration period.
  • Example 17 has a significant inhibitory effect on the growth of human multiple myeloma cells PRMI8226 xenograft tumors in nude mice, and the high dose group of Example 17 has strong anti-tumor activity on human multiple myeloma PRMI8226 xenograft tumors in nude mice In the positive drug Palbociclib group.

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Abstract

本发明公开了一种氮杂吲哚嘧啶胺杂环化合物及其制备方法和应用，所述化合物如式(S)所示。本发明的化合物能抑制癌症的恶性增殖，治疗效果好、毒性低，具有良好药物代谢特性、不易产生耐药性，可用于制备治疗癌症或肿瘤相关疾病药物，所述癌症或肿瘤相关疾病包括多发性骨髓瘤、白血病、乳腺癌、前列腺癌、肺癌、肝癌、胃癌、骨癌、脑癌、头颈癌、肠癌、胰腺癌、膀胱癌、睾丸癌、卵巢癌以及子宫内膜癌。

Description

氮杂吲哚嘧啶胺杂环化合物及其制备方法和应用 技术领域
本发明涉及药物化学领域，特别涉及一种氮杂吲哚嘧啶胺杂环化合物及其制备方法和应用。
背景技术
不受控制的细胞连续分裂导致细胞不断增殖是癌症的一个显著的特征，因此阻止细胞不断分裂和增殖是治疗癌症的一个非常有效的方式。细胞周期蛋白依赖性激酶(Cyclin-dependent kinases，CDKs)在细胞分裂和细胞增殖中发挥着至关重要的调节作用。选择性的抑制CDK6将细胞周期阻滞在G1期，从而抑制细胞增殖。并且当CDK6缺失时，在CDK1的代偿作用下，不会对正常成纤维细胞的增殖产生影响，因此，选择性的抑制CDK6是治疗癌症的一个安全有效的治疗策略。目前，选择性CDK6抑制剂帕博西尼(Palbociclib)，瑞博西尼(Ribociclib)和阿贝西利(Abemaciclib)已经被FDA批准用于乳腺癌的治疗。目前，对于CDK6抑制剂的耐药性已经出现并且日益严重，因此，找寻全新骨架的CDK6抑制剂是非常有必要的。
发明内容
发明目的：本发明提供了一种氮杂吲哚嘧啶胺杂环化合物，对CDK6产生抑制作用。本发明还提供了该化合物的制备方法和应用。
技术方案：本发明所述的如式(S)所示的化合物或其药学上可接受的盐：
Figure PCTCN2022084057-appb-000001
其中，
所述X选自C(O)或(CH 2) n；n为0-8；
所述R 1选自氢或卤素；
所述R 2选自氢、C 1-C 8烷基或C 3-C 6环烷基；
所述R 3选自氢、卤素或C 1-C 3烷氧基；
所述R 4选自氢、C 1-C 8烷基、-S(O) 2C 1-C 3烷基或-C(O)OC 1-C 6烷基。
在本发明的一些实施例中，所述X为C(O)或(CH 2) n；n为0-4。
在本发明的一些实施例中，所述R 1为氢或氟。
在本发明的一些实施例中，所述R 2为氢、C 1-C 4烷基、环戊烷基或环己烷基；在一些具体的实施例中，所述R 2为氢、甲基、乙基、异丙基、异丁基或环戊烷基。
在本发明的一些实施例中，所述R 3为氢、氟或甲氧基。
在本发明的一些实施例中，所述R 4为氢、C 1-C 3烷基、-S(O) 2C 1-C 3烷基或-C(O)OC 1-C 4烷基；在一些更具体的实施例中，所述R 4为氢、甲基、乙基、异丙基或叔丁氧羰基。
在本发明的一些实施例中，优选地：所述X选自：(CH 2) n或C(O)，n为0或1；所述R 1选自：氢、氟；所述R 2选自：氢、甲基、乙基、异丙基、异丁基、环戊烷基；所述R 3选自：氢、氟或甲氧基；所述R 4选自：氢、甲基、乙基、异丙基或叔丁氧羰基。
在本发明的一些具体的实施例中，本发明还提供选自S-1至S-44所示的化合物或其 药学上可接受的盐：
Figure PCTCN2022084057-appb-000002
Figure PCTCN2022084057-appb-000003
Figure PCTCN2022084057-appb-000004
Figure PCTCN2022084057-appb-000005
Figure PCTCN2022084057-appb-000006
Figure PCTCN2022084057-appb-000007
Figure PCTCN2022084057-appb-000008
上述药学上可接受的盐为通式(S)化合物的酸加成盐，其中用于成盐的酸包括无机酸及有机酸，所述无机酸包括：盐酸、硫酸、磷酸和甲磺酸，有机酸包括乙酸、三氯乙酸、丙酸、丁酸、马来酸、对甲苯磺酸、苹果酸、丙二酸、肉桂酸、柠檬酸、富马酸、樟脑酸、二葡糖酸、天冬氨酸和酒石酸。
优选地，本发明中所述的药学上可接受的盐为盐酸盐。
本发明通式(S)化合物的制备方法，由化合物(A)与化合物(B)在钯催化剂的作用下经偶联反应制备化合物(S)：
Figure PCTCN2022084057-appb-000009
其中，
所述X选自C(O)或(CH 2) n；n为0-8；
所述R 1选自氢或卤素；
所述R 2选自氢、C 1-C 8烷基或C 3-C 6环烷基；
所述R 3选自氢、卤素或C 1-C 3烷氧基；
所述R 4选自氢、C 1-C 8烷基、-S(O) 2C 1-C 3烷基或-C(O)OC 1-C 6烷基。
优选地，所述化合物(A)与化合物(B)的反应体系为：反应试剂为Pd 2(dba) 3、BINAP以及碳酸铯，在惰性气体氛围下，反应温度为95-105℃，反应时间为10-24h。
本发明还提供了如式(S)所述的化合物或其药学上可接受的盐、酯、立体异构体、溶剂化合物或前药，以及药学上可接受的载体的药物组合物。
药学上可接受的载体指的是对有机体不引起明显的刺激性和不干扰所给予化合物的生物活性和性质的赋形剂或稀释剂。
本发明还提供了一种式(S)化合物或其药学上可接受的盐、酯、立体异构体、溶剂化合物或前药或本发明所述的组合物在制备药物中的用途。
本发明还提供了一种式(S)化合物或其药学上可接受的盐、酯、立体异构体、溶剂化合物或前药或本发明所述的组合物在制备CDK6抑制剂中的应用；所述CDK6激酶抑制剂用于治疗癌症或肿瘤相关疾病。
本发明所述的化合物或其药学上可接受的盐、酯、立体异构体、溶剂化合物或前药或本发明所述的组合物在制备治疗癌症或肿瘤相关疾病药物中的应用。癌症或肿瘤相关疾病包括但不限于多发性骨髓瘤、白血病、乳腺癌、前列腺癌、肺癌、肝癌、胃癌、骨癌、脑癌、头颈癌、肠癌、胰腺癌、膀胱癌、睾丸癌、卵巢癌以及子宫内膜癌等。
本发明所述的通式(S)化合物或其药学上可接受的盐，具有CDK6靶点抑制活性，对细胞恶性增值肿瘤具有治疗效果。
本发明中的术语除特别说明外，一般具有如下的含义。
Me为甲基，Et为乙基，Boc为叔丁氧羰基。
术语“烷基”表示具有所述数目之碳原子的直链或支链饱和烃基。
术语“C 1-C 8烷基”是指具有1-8个碳原子的直链或支链饱和烃基。C 1-C 8烷基包括但不限于甲基、乙基、正丙基、异丙基、正丁基、异丁基、叔丁基、正戊基、异戊基、新戊基、正己基、异己基、2,2-二甲基丁基和2,3-二甲基丁基等。
术语“C 3-C 6环烷烃”包括环丙烷基、环丁烷基、环戊烷基以及环己烷基。
术语“C 1-C 3烷基”是指具有1-3个碳原子的直链或支链饱和烃基。
术语“烷氧基”表示O-烷基。术语“C 1-C 3烷氧基”是指具有O-C 1-C 3烷基。
术语“卤素”为氟、氯、溴或碘。优选为氟、氯、溴。
有益效果：本发明通式(S)所示的化合物，能抑制癌症的恶性增殖，治疗效果好、毒性低，具有良好药物代谢特性、不易产生耐药性，可用于制备治疗癌症或肿瘤相关疾病药物。
具体实施方式
以下的实施例便于更好地理解本发明，但并不限定本发明。下述实施例中的实验方法，如无特殊说明，均为常规方法。下述实施例中所用的试验材料，如无特殊说明，均为自常规生化试剂商店购买得到的。下面结合具体实施例对本申请作出详细说明。
一、中间反应物的合成
反应物(ZA)和反应物(ZB)可以直接购买或者自主研发，自主研发能够显著降低成本。自主研发反应物(ZA)和反应物(ZB)的具体制备方法如下：
(1)1-BOC-3-溴-7-氮杂吲哚(ZA1)的合成
Figure PCTCN2022084057-appb-000010
将3-溴-7-氮杂吲哚(3.94g，20mmol)，二碳酸二叔丁酯(4.80g，22mmol)溶于二氯甲烷(150mL)，然后加入三乙胺(3.06g，30mmol)，4-二甲氨基吡啶(0.49g，4mmol)，室温反应12个小时，快速硅胶柱纯化得到化合物ZA1(5.53g，产率为93％)。 1H NMR(300MHz,Chloroform-d)δ8.55(dd,J＝4.8,1.7Hz,1H),7.84(dd,J＝7.9,1.7Hz,1H),7.71(s,1H),7.30–7.27(m,1H),1.67(s,9H).
(2)叔丁基3-(4,4,5,5-四甲基-1,3,2-二噁硼戊烷-2-基)-1氢-吡咯并吡啶-1-羧酸酯(ZA2)的合成
Figure PCTCN2022084057-appb-000011
将1-BOC-3-溴-7-氮杂吲哚(4.46g，15mmol)溶于Dioxane(100mL)，然后加入联硼酸频那醇酯(4.57g，18mmol)，Pd 2(dba) 3(687mg，0.75mmol)，PCy 3(505mg，1.8mmol)和醋酸钾(4.41g，45mmol)，置换氩气三次，加热至100℃，反应8h。冷却过滤浓缩，快速硅胶柱纯化得到化合物ZA2(3.87g，产率为75％)。 1H NMR(400MHz,Chloroform-d)δ8.99(dd,J＝8.0,1.7Hz,1H),8.62(dd,J＝4.8,1.7Hz,1H),8.53(d,J＝2.2Hz,1H),8.49(d,J＝2.9Hz,1H),7.39(dd,J＝8.0,4.8Hz,1H),1.73(s,9H).
(3)叔丁基3-(2-氯-5-氟嘧啶-4-基)-1氢-吡咯并吡啶-1-羧酸酯(ZA3)的合成
Figure PCTCN2022084057-appb-000012
将2，4-二氯-5-氟嘧啶(2.34g，14mmol)加到三颈烧瓶中，再加入Pd(PPh 3) 2Cl 2(351mg，0.5mmol)，碳酸钠(2.65g，25mmol)，乙二醇二甲醚(100mL)和H 2O(12.5mL)，置换氩气三次，加热至80℃。化合物ZA2(3.44g，10mmol)溶于乙二醇二甲醚(50mL)，缓慢滴加到三颈烧瓶中，反应16h。冷却过滤浓缩，快速硅胶柱纯化得到化合物ZA3(1.67g，产率为48％)。 1H NMR(400MHz,Chloroform-d)δ8.99(dd,J＝8.0,1.7Hz,1H),8.62(dd,J＝4.8,1.7Hz,1H),8.53(d,J＝2.2Hz,1H),8.49(d,J＝2.9Hz,1H),7.39(dd,J＝8.0,4.8Hz,1H),1.73(s,9H).
(4)叔丁基3-(2-氯-5-氟嘧啶-4-基)-1-甲基-1氢-吡咯并吡啶(ZA4)的合成
Figure PCTCN2022084057-appb-000013
将化合物ZA3(1.395g，4mmol)溶于二氯甲烷(50mL)，通入现场制得的HCl气体，反应两个小时后，旋干溶剂，得到脱BOC的产物，无须纯化，直接溶于DMF(50mL)，冰浴的条件下加入NaH(320mg，8mmol)，反应30分钟后，加入碘甲烷(2.27g，16mmol)，室温反应8个小时，冰浴的条件下加水淬灭，二氯甲烷萃取，过滤浓缩溶剂，快速硅胶柱纯化得到化合物ZA4(600mg，产率为56％)。 1H NMR(300MHz,Chloroform-d)δ8.91(dd,J＝8.0,1.6Hz,1H),8.44(dd,J＝4.7,1.6Hz,1H),8.33(d,J＝3.2Hz,1H),8.06(d,J＝2.5Hz,1H),7.30–7.27(m,1H),3.98(s,3H).
(5)3-(2-氯-5-氟嘧啶-4-基)-1-乙基-1氢-吡咯并吡啶(ZA5)的合成
Figure PCTCN2022084057-appb-000014
参照化合物(ZA4)的合成方法，产率为40％。
(6)3-(2-氯-5-氟嘧啶-4-基)-1-异丙基-1氢-吡咯并吡啶(ZA6)的合成
Figure PCTCN2022084057-appb-000015
参照化合物(ZA4)的合成方法，产率为46％。 1H NMR(300MHz,Chloroform-d)δ8.95(dd,J＝8.0,1.6Hz,1H),8.45(dd,J＝4.7,1.7Hz,1H),8.36(d,J＝3.3Hz,1H),8.21(d,J＝2.2Hz,1H),7.32–7.28(m,1H),5.34–5.25(m,1H),1.63(d,J＝6.8Hz,6H).
(7)1-(仲丁基)-3-(2-氯-5-氟嘧啶-4-基)-1氢-吡咯并吡啶(ZA7)的合成
Figure PCTCN2022084057-appb-000016
参照化合物(ZA4)的合成方法，产率为39％。
(8)3-(2-氯-5-氟嘧啶-4-基)-1-环戊基-1氢-吡咯并吡啶(ZA8)的合成
Figure PCTCN2022084057-appb-000017
参照化合物(ZA4)的合成方法，产率为33％。
(9)3-溴-1-异丙基-1氢-吡咯并吡啶(ZA9)的合成
Figure PCTCN2022084057-appb-000018
将3-溴-7-氮杂吲哚(3.94g，20mmol)溶于DMF(100mL)，冰浴的条件下加入NaH(1.6g，40mmol)，反应30分钟后，加入碘代异丙烷(1.36g，80mmol)，室温反应8个小时，冰浴的条件下加水淬灭，二氯甲烷萃取，过滤浓缩溶剂，快速硅胶柱纯化得到化合物ZA9(4.21g，产率为88％)。 1H NMR(300MHz,Chloroform-d)δ8.34(dd,J＝4.7,1.6Hz,1H),7.84(dd,J＝7.8,1.6Hz,1H),7.33(s,1H),7.12(dd,J＝7.9,4.7Hz,1H),5.26–5.17(m,1H),1.51(d,J＝6.7Hz,6H).
(10)3-溴-5-氟-1-异丙基-1氢-吡咯并吡啶(ZA10)的合成
Figure PCTCN2022084057-appb-000019
参照化合物(ZA9)的合成方法，产率为90％。 1H NMR(300MHz,Chloroform-d)δ8.21(s,1H),7.51(dd,J＝8.5,2.6Hz,1H),7.37(s,1H),5.19–5.10(m,1H),1.50(d,J＝6.8Hz,6H).
(11)1-异丙基-3-(四甲基-1,3,2-二噁硼戊烷-2-基)-1氢-吡咯并吡啶(ZA11)的合成
Figure PCTCN2022084057-appb-000020
参照化合物(ZA2)的合成方法，产率为54％。 1H NMR(300MHz,Chloroform-d)δ8.31(d,J＝4.6Hz,1H),8.27(d,J＝7.8Hz,1H),7.81(s,1H),7.13–7.08(m,1H),5.23–5.14(m,1H),1.53(d,J＝6.8Hz,6H),1.37(s,12H).
(12)5-氟-1-异丙基-3-(四甲基-1,3,2-二噁硼戊烷-2-基)-1氢-吡咯并吡啶(ZA12)的合成
Figure PCTCN2022084057-appb-000021
参照化合物(ZA2)的合成方法，产率为68％。 1H NMR(300MHz,Chloroform-d)δ8.19–8.17(m,1H),7.96(dd,J＝9.2,2.7Hz,1H),7.84(s,1H),5.16–5.07(m,1H),1.52(d,J＝6.7Hz,6H),1.36(s,12H).
(13)3-(2-氯-5-氟嘧啶-4-基)-5-氟-1-异丙基-吡咯并吡啶(ZA13)的合成
Figure PCTCN2022084057-appb-000022
参照化合物(ZA3)的合成方法，产率为85％。 1H NMR(400MHz,Chloroform-d)δ8.65(dd,J＝9.3,2.6Hz,1H),8.38–8.36(m,1H),8.31–8.30(m,2H),8.24(s,1H),5.25–5.18(m,1H),1.61(d,J＝6.6Hz,6H).
(14)3-(2-氯-4-基)-1-异丙基-1氢-吡咯并吡啶(ZA14)的合成
Figure PCTCN2022084057-appb-000023
参照化合物(ZA3)的合成方法，产率为88％。 1H NMR(300MHz,Chloroform-d)δ8.65(d,J＝8.0Hz,1H),8.47(d,J＝5.4Hz,1H),8.42(d,J＝4.7Hz,1H),8.17(s,1H),7.48(d,J＝5.4Hz,1H),7.27(dd,J＝5.6,2.6Hz,1H),5.32–5.23(m,1H),1.60(d,J＝6.7Hz,6H).
(15)3-(2-氯-5-甲氧基嘧啶-4-基)-1-异丙基-1氢-吡咯并吡啶(ZA15)的合成
Figure PCTCN2022084057-appb-000024
参照化合物(ZA3)的合成方法，产率为86％。 1H NMR(300MHz,Chloroform-d)δ9.05(dd,J＝8.0,1.7Hz,1H),8.41(dd,J＝4.7,1.7Hz,1H),8.38(s,1H),8.14(s,1H),7.29–7.25(m,1H),5.31–5.22(m,1H),4.10(s,3H),1.61(d,J＝6.8Hz,6H).
(16)5-((4-乙基哌嗪-1-基)甲基)吡啶-2-胺(ZB1)的合成
Figure PCTCN2022084057-appb-000025
将2-氨基-5-醛基吡啶(2.34g，10mmol)和N-乙基哌嗪(1.37g，12mmol)溶于1，2-二氯乙烷(50mL)，室温搅拌2个小时，随后加入三乙酰基硼氢化钠(3.18g，15mmol)，室温搅拌8个小时。加入1M NaOH(50mL)淬灭，DCM(50mLx3)萃取，无水硫酸钠干燥，浓缩后柱层析(DCM/MeOH＝10:1)得到化合物ZB1(1.85g，产率为84％)。 1H NMR(300MHz,DMSO-d 6)δ7.74(d,J＝2.3Hz,1H),7.26(dd,J＝8.4,2.3Hz,1H),6.40(d,J＝8.4Hz,1H),3.23(s,2H),2.31–2.24(m,10H),0.96(t,J＝7.1Hz,3H).
(17)5-((4-异丙基哌嗪-1-基)甲基)吡啶-2-胺(ZB2)的合成
Figure PCTCN2022084057-appb-000026
参考化合物(ZB1)的合成方法，产率为81％。 1H NMR(300MHz,CDCl 3)δ7.95(d,J＝2.3Hz,1H),7.42(dd,J＝8.4,2.3Hz,1H),6.48(dd,J＝8.5,0.8Hz,1H),3.39(s,2H),2.75(p,J＝6.5Hz,1H),2.59(d,J＝22.1Hz,8H),1.10(d,J＝6.5Hz,6H).
(18)叔丁基4-((6-胺基吡啶-3-基)甲基)哌嗪-1-羧酸酯(ZB3)的合成
Figure PCTCN2022084057-appb-000027
参考化合物(ZB1)的合成方法，产率为86％。 1H NMR(300MHz,CDCl 3)δ7.94–7.93(m,1H),7.43(dd,J＝8.4,2.3Hz,1H),6.50(dd,J＝8.3,0.8Hz,1H),3.43–3.37(m,6H),2.36(t,J＝5.0Hz,4H),1.45(s,9H).
(19)(6-氨基吡啶-3-基)(4-乙基哌嗪-1-基)甲酮(ZB4)的合成
Figure PCTCN2022084057-appb-000028
称取6-氨基烟酸(1.38g，10mmol)，N，N'-羰基二咪唑(3.24g，20mmol)溶于DMF(50mL)，70℃反应10分钟，室温再搅拌1个小时，加入N-乙基哌嗪(1.37g，12mmol)，室温反应过夜，浓缩，快速硅胶柱纯化得到化合物ZB4(1.92g，产率为82％)。 1H NMR(300MHz,CDCl 3)δ8.19(d,J＝2.3Hz,1H),7.54(dd,J＝8.5,2.3Hz,1H),6.50(d,J＝8.5Hz,1H),3.67(s,4H),2.49–2.42(m,6H),1.11(t,J＝7.2Hz,3H).
(20)叔丁基4-(6-胺基烟碱酰基)哌嗪-1-羧酸酯(ZB5)的合成。
Figure PCTCN2022084057-appb-000029
参考化合物(ZB4)的合成方法，产率为87％。 1H NMR(300MHz,CDCl3)δ8.18(d,J＝2.2Hz,1H),7.56(dd,J＝8.5,2.2Hz,1H),6.51(d,J＝8.5Hz,1H),4.76(s,2H),3.65–3.56(m,4H),3.48–3.42(m,4H),1.48(s,9H)。
(21)(6-氨基吡啶-3-基)(4-异丙基哌嗪-1-基)甲酮(ZB6)的合成
Figure PCTCN2022084057-appb-000030
参考化合物(ZB4)的合成方法，产率为85％。 1H NMR(400MHz,CDCl 3)δ8.19(s,1H),7.56(d,J＝2.5Hz,1H),6.50(d,J＝8.2Hz,1H),4.67(s,2H),3.65(s,4H),2.74(dt,J＝12.0,6.3Hz,1H),2.53(s,4H),1.05(td,J＝6.2,5.7,4.0Hz,6H).
(22)(6-氨基吡啶-3-基)(4-甲基哌嗪-1-基)甲酮(ZB7)的合成
Figure PCTCN2022084057-appb-000031
参考化合物(ZB4)的合成方法，产率为87％。 1H NMR(300MHz,Chloroform-d)δ8.19(dd,J＝2.4,0.8Hz,1H),7.55(dd,J＝8.5,2.3Hz,1H),6.50(dd,J＝8.5,0.8Hz,1H),4.82(s,2H),3.69–3.65(m,4H),2.46–2.42(m,4H),2.33(s,3H).
(23)叔丁基4-(6-胺基吡啶-3-基)哌嗪-1-羧酸酯(ZB8)的合成
Figure PCTCN2022084057-appb-000032
步骤一，叔丁基4-(6-硝基吡啶-3-基)哌嗪-1-羧酸酯的合成：称取5-溴-2-硝基吡啶(0.41g，2.0mmol)，叔丁基哌嗪-1-羧酸酯(0.48g，2.6mmol)和三乙胺(0.41g，4.0mmol)溶于DMSO(5mL)，加热至60℃，反应18个小时。冷却过滤浓缩，快速硅胶柱纯化得到化合物叔丁基4-(6-硝基吡啶-3-基)哌嗪-1-羧酸酯(0.49g，产率为80％)。 1H NMR(400MHz,CDCl 3)δ8.18(d,J＝9.2Hz,1H),8.13(d,J＝3.0Hz,1H),7.22(dd,J＝9.2,3.1Hz,1H),3.65(dd,J＝6.6,4.0Hz,4H),3.47(dd,J＝6.5,4.1Hz,4H),1.49(s,9H).
Figure PCTCN2022084057-appb-000033
步骤二，叔丁基4-(6-胺基吡啶-3-基)哌嗪-1-羧酸酯的合成：称取叔丁基4-(6-硝基吡啶-3-基)哌嗪-1-羧酸酯(0.31g，1.0mmol)，还原铁粉(0.17g，3.0mmol)和氯化铵(0.49g，9.0mmol)溶于70％乙醇(10mL)，加热至70℃，反应6个小时。冷却过滤浓缩，快速硅胶柱纯化得到化合物ZB8(0.24g，产率为85％)。 1H NMR(300MHz,CDCl 3)δ7.66(d,J＝2.8Hz,1H),7.24(dd,J＝8.9,2.8Hz,1H),6.63(d,J＝8.9Hz,1H),3.14(q,J＝7.3Hz,4H),2.95(t,J＝5.1Hz,4H),1.48(s,9H).
二、化合物S-1-S-44的合成
实施例1：N-(5-((4-乙基哌嗪-1-基)甲基)吡啶-2-基)-5-氟-4-(1氢-吡咯并吡啶-3-基)嘧啶-2-胺(S-1)的合成
Figure PCTCN2022084057-appb-000034
化合物ZA3(348.7mg，1.0mmol)和ZB1(264.4mg，1.2mmol)溶于二氧六环(10mL)，然后加入Pd 2(dba) 3(45.8mg，0.05mmol)，BINAP(62.3mg，0.1mmol)，碳酸铯(651.6mg，2.0mmol)，置换氩气三次，加热至100℃，反应12h。冷却过滤浓缩柱层析(DCM/MeOH＝20:1，v/v)得到化合物S-1(207.6mg，48％收率)，白色固体。 1H NMR(400MHz,CDCl 3)δ8.96(d,J＝2.3Hz,2H),8.91(s,1H),8.73(s,1H),8.60(d,J＝3.1Hz,1H),8.54(d,J＝8.8Hz,2H),8.49(d,J＝2.2Hz,1H),8.29(d,J＝8.9Hz,1H),7.86(dd,J＝8.7,2.4Hz,1H),3.65-3.50(m,8H),1.48(s,9H).
实施例2：(4-乙基哌嗪-1-基)(6-((5-氟-4-(1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)甲酮(S-2)的合成
Figure PCTCN2022084057-appb-000035
参考实施例1的合成方法，产率为42％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ12.55(s,1H),10.50(s,1H),9.33(dd,J＝8.0,1.6Hz,1H),8.57(d,J＝3.6Hz,1H),8.39(dd,J＝2.4,0.8Hz,1H),8.37(dd,J＝4.6,1.7Hz,1H),8.31(s,1H),8.27(d,J＝8.4Hz,1H),7.83(dd,J＝8.7,2.4Hz,1H),7.25(dd,J＝8.0,4.7Hz,1H),3.57–3.53(m,4H),2.41–2.34(m,6H),1.01(t,J＝7.2Hz,3H).
实施例3：叔丁基4-((6-((5-氟-4-(1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)甲酮)哌嗪-1-羧酸酯(S-3)的合成
Figure PCTCN2022084057-appb-000036
参考实施例1的合成方法，产率为45％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ12.53(s,1H),10.17(s,1H),9.30(dd,J＝8.0,1.6Hz,1H),8.52(d,J＝3.6Hz,1H),8.37(dd,J＝4.6,1.7Hz,1H),8.29–8.28(m,1H),8.22(d,J＝2.3Hz,1H),8.16(d,J＝8.6Hz,1H),7.68(dd,J＝8.6,2.3Hz,1H),7.23(dd,J＝8.0,4.6Hz,1H),3.47(s,2H),3.32(t,J＝5.0Hz,4H),2.34(t,J＝5.0Hz,4H),1.39(s,9H).
实施例4：5-氟-N-(5-(哌嗪-1-基甲基)吡啶-2-基)-4-(1氢-吡咯并吡啶-3-基)嘧啶-2-胺盐酸盐(S-4)的合成
Figure PCTCN2022084057-appb-000037
取实施例3化合物(120mg)溶于二氯甲烷(40mL)中，0℃条件下通入HCl气体反应2h。反应结束后，浓缩得到化合物S4，产率为100％，淡黄色固体。 1H NMR(400MHz,DMSO-d 6)δ12.98(s,1H),12.41(s,1H),10.07(s,2H),9.37(dd,J＝8.0,1.5Hz,1H),8.70–8.68(m,2H),8.52(dd,J＝9.0,2.1Hz,1H),8.46–8.43(m,2H),7.99(d,J＝9.0Hz,1H),7.38(dd,J＝8.0,4.8Hz,1H),4.58(s,2H),3.52–3.49(m,8H).
实施例5：叔丁基4-(6-((5-氟-4-(1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)哌嗪-1-羧酸酯(S-5)的合成
Figure PCTCN2022084057-appb-000038
参考实施例1的合成方法，产率为44％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ12.50(s,1H),9.89(s,1H),9.24(d,J＝8.1Hz,1H),8.47(d,J＝3.6Hz,1H),8.36(d,J＝4.6Hz,1H),8.27–8.25(m,1H),8.06–8.01(m,2H),7.47(dd,J＝9.2,3.1Hz,1H),7.23(dd,J＝8.0,4.7Hz,1H),3.49(t,J＝5.0Hz,4H),3.09(t,J＝5.3Hz,4H),1.43(s,9H).
实施例6：5-氟-N-(5-(哌嗪-1-基)吡啶-2-基)-4-(1氢-吡咯并吡啶-3-基)嘧啶-2-胺盐酸盐(S-6)的合成
Figure PCTCN2022084057-appb-000039
参考实施例4的合成方法，产率为100％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ12.98(s,1H),12.27(s,1H),9.66(s,2H),9.30(dd,J＝8.0,1.6Hz,1H),8.67(d,J＝3.5Hz,1H),8.45–8.43(m,2H),8.27(dd,J＝9.7,2.9Hz,1H),7.94(d,J＝2.9Hz,1H),7.85(d,J＝9.6Hz,1H),7.36(dd,J＝8.0,4.8Hz,1H),3.48–3.46(m,4H),3.29–3.25(m,4H).
实施例7：(6-((5-氟-4-(1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)胺)吡啶-3-基)(4-异丙基哌嗪-1-基)甲酮(S-7)的合成
Figure PCTCN2022084057-appb-000040
参考实施例1的合成方法，产率为40％，白色固体。 1H NMR(300MHz,DMSO-d 6)δ12.56(s,1H),10.51(s,1H),9.33(dd,J＝8.0,1.7Hz,1H),8.57(d,J＝3.6Hz,1H),8.39–8.36(m,2H),8.32–8.30(m,1H),8.27(d,J＝8.7Hz,1H),7.83(dd,J＝8.7,2.4Hz,1H),7.25(dd,J＝8.0,4.7Hz,1H),3.55–3.51(m,4H),2.74–2.66(m,1H),2.48–2.46(m,4H),0.98(d,J＝6.5Hz,6H).
实施例8：N-(5-((4-乙基哌嗪-1-基)甲基)吡啶-2-基)-5-氟-4-(1-甲基-吡咯并吡啶-3-基)嘧啶-2-胺(S-8)的合成
Figure PCTCN2022084057-appb-000041
参考实施例1的合成方法，产率为48％，白色固体。 1H NMR(300MHz,Chloroform-d)δ8.92(dd,J＝8.0,1.7Hz,1H),8.44(dd,J＝4.7,1.6Hz,1H),8.33–8.30(m,2H),8.25(d,J＝2.2Hz,1H),8.22(s,1H),8.07(d,J＝2.4Hz,1H),7.68(dd,J＝8.6,2.3Hz,1H),7.22(dd,J＝8.0,4.7Hz,1H),3.99(s,3H),3.50(s,2H),2.56–2.42(m,10H),1.11(t,J＝7.2Hz,3H).
实施例9：叔丁基4-((6-((5-氟-4-(1-甲基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)甲酮)哌嗪-1-羧酸酯(S-9)的合成
Figure PCTCN2022084057-appb-000042
参考实施例1的合成方法，产率为47％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ10.18(s,1H),9.33(dd,J＝8.0,1.6Hz,1H),8.52(d,J＝3.5Hz,1H),8.43–8.40(m,2H),8.22(d,J＝2.3Hz,1H),8.15(d,J＝8.6Hz,1H),7.68(dd,J＝8.6,2.3Hz,1H),7.27(dd,J＝8.0,4.6Hz,1H),3.95(s,3H),3.47(s,2H),3.35–3.31(m,4H),2.35–2.33(m,4H),1.39(s,9H).
实施例10：5-氟-4-(1-甲基-1氢-吡咯并吡啶-3-基)-N-(5-(哌嗪-1-基甲基)吡啶-2-基)嘧啶-2-胺盐酸盐(S-10)的合成
Figure PCTCN2022084057-appb-000043
参考实施例4的合成方法，产率为100％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ12.37(s,1H),10.02(s,2H),9.32(dd,J＝8.0,1.6Hz,1H),8.68(d,J＝3.1Hz,2H),8.60(d,J＝2.8Hz,1H),8.51(dd,J＝9.0,2.2Hz,1H),8.46(dd,J＝4.7,1.6Hz,1H),7.98(d,J＝9.0Hz,1H),7.39(dd,J＝7.9,4.7Hz,1H),4.57(s,2H),3.98(s,3H),3.52–3.49(m,8H).
实施例11：叔丁基4-(6-((5-氟-4-(1-甲基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)哌嗪-1-羧酸酯(S-11)的合成
Figure PCTCN2022084057-appb-000044
参考实施例1的合成方法，产率为43％，白固体。 1H NMR(400MHz,Chloroform-d)δ8.94(dd,J＝8.0,1.6Hz,1H),8.44(dd,J＝4.7,1.6Hz,1H),8.28–8.25(m,2H),8.13(s,1H),8.08(d,J＝2.4Hz,1H),8.00(d,J＝2.9Hz,1H),7.36(dd,J＝9.1,3.0Hz,1H),7.24(dd,J＝8.0,4.7Hz,1H),4.00(s,3H),3.61(t,J＝5.1Hz,4H),3.09(t,J＝5.2Hz,4H),1.50(s,9H).
实施例12：5-氟-4-(1-甲基-1氢-吡咯并吡啶-3-基)-N-(5-(哌嗪-1-基)吡啶-2-基)嘧啶-2-胺盐酸盐(S-12)的合成
Figure PCTCN2022084057-appb-000045
参考实施例4的合成方法，产率为100％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ12.01(s,1H),9.48(s,2H),9.23(dd,J＝8.0,1.6Hz,1H),8.67(d,J＝3.6Hz,1H),8.60(d,J＝2.8Hz,1H),8.47(dd,J＝4.7,1.6Hz,1H),8.23(dd,J＝9.6,2.9Hz,1H),7.94(d,J＝2.9Hz,1H),7.81(d,J＝9.6Hz,1H),7.37(dd,J＝8.0,4.7Hz,1H),3.98(s,3H),3.46–3.44(m,4H),3.28–3.26(m,4H).
实施例13：(6-((1-乙基-1氢-吡咯并吡啶-3-基)-5-氟嘧啶-2-基)胺)吡啶-3-基)(4-乙基哌嗪-1-基)甲酮(S-13)的合成
Figure PCTCN2022084057-appb-000046
参考实施例1的合成方法，产率为42％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.92(dd,J＝8.0,1.6Hz,1H),8.46–8.44(m,3H),8.34(d,J＝3.4Hz,1H),8.25(s,1H),8.15(d,J＝2.3Hz,1H),7.80(dd,J＝8.6,2.4Hz,1H),7.25–7.23(m,1H),4.46(q,J＝7.3Hz,2H),3.80–3.57(m,4H),2.50–2.45(m,6H),1.58(t,J＝7.3Hz,3H),1.12(t,J＝7.2Hz,3H).
实施例14：(6-((1-乙基-1氢-吡咯并吡啶-3-基)-5-氟嘧啶-2-基)胺)吡啶-3-基)(4-异丙基哌嗪-1-基)甲酮(S-14)的合成
Figure PCTCN2022084057-appb-000047
参考实施例1的合成方法，产率为43％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.91(dd,J＝8.0,1.6Hz,1H),8.47–8.43(m,4H),8.35(d,J＝3.4Hz,1H),8.14(d,J＝2.3Hz,1H),7.81(dd,J＝8.6,2.4Hz,1H),7.24(dd,J＝8.0,4.7Hz,1H),4.46(q,J＝7.3Hz,2H),3.78–3.60(m,4H),2.79–2.72(m,1H),2.60–2.51(m,4H),1.57(t,J＝7.3Hz,3H),1.07(d,J＝6.5Hz,6H).
实施例15：4-(1-乙基-1氢-吡咯并吡啶-3-基)-5-氟-N-(5-((4-异丙基哌嗪-1-基)甲基)吡啶-2-基)嘧啶-2-胺(S-15)的合成
Figure PCTCN2022084057-appb-000048
参考实施例1的合成方法，产率为43％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.94(dd,J＝8.0,1.7Hz,1H),8.44(dd,J＝4.7,1.6Hz,1H),8.33–8.30(m,2H),8.22(d,J＝2.3Hz,1H),8.14(d,J＝2.4Hz,1H),7.92(s,1H),7.68(dd,J＝8.6,2.3Hz,1H),7.25–7.23(m,1H),4.46(q,J＝7.3Hz,2H),3.50(s,2H),2.71–2.51(m,8H),1.57(t,J＝7.3Hz,3H),1.07(d,J＝6.5Hz,6H).
实施例16：N-(5-((4-乙基哌嗪-1-基)甲基)吡啶-2-基)-5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-胺(S-16)的合成
Figure PCTCN2022084057-appb-000049
参考实施例1的合成方法，产率为47％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.93(dd,J＝8.0,1.6Hz,1H),8.42(dd,J＝4.6,1.7Hz,1H),8.34–8.31(m,2H),8.24(d,J＝2.2Hz,1H),8.18(d,J＝2.2Hz,1H),8.15(s,1H),7.68(dd,J＝8.6,2.3Hz,1H),7.23(dd,J＝8.0,4.7Hz,1H),5.33–5.26(m,1H),3.50(s,2H),2.56–2.43(m,10H),1.62(d,J＝6.8Hz,6H),1.10(t,J＝7.2Hz,3H).
实施例17：5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)-N-(5-((4-异丙基哌嗪-1-基)甲基)吡啶-2-基)嘧啶-2-胺(S-17)的合成
Figure PCTCN2022084057-appb-000050
参考实施例1的合成方法，产率为49％，白色固体。 1H NMR(300MHz,Chloroform-d)δ8.94(dd,J＝8.0,1.6Hz,1H),8.43(dd,J＝4.7,1.7Hz,1H),8.34–8.31(m,2H),8.23(dd,J＝2.4,0.8Hz,1H),8.19(d,J＝2.2Hz,1H),8.01(s,1H),7.67(dd,J＝8.6,2.3Hz,1H),7.23(dd,J＝8.0,4.7Hz,1H),5.34–5.25(m,1H),3.51(s,2H),2.73–2.56(m,9H),1.62(d,J＝6.8Hz,6H),1.09(d,J＝6.5Hz,6H).
实施例18：叔丁基4-((6-((5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)甲酮)哌嗪-1-羧酸酯(S-18)的合成
Figure PCTCN2022084057-appb-000051
参考实施例1的合成方法，产率为42％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ10.17(s,1H),9.32(dd,J＝7.9,1.6Hz,1H),8.54(d,J＝3.6Hz,1H),8.41(d,J＝2.7Hz,2H),8.22(s,1H),8.15(d,J＝8.5Hz,1H),7.68(dd,J＝8.6,2.3Hz,1H),7.27(dd,J＝8.0,4.6Hz,1H),5.23–5.16(m,1H),3.47(s,2H),3.35–3.31(m,4H),2.33(t,J＝5.1Hz,4H),1.58(d,J＝6.7Hz,6H),1.39(s,9H).
实施例19：5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)-N-(5-(哌嗪-1-基甲基)吡啶-2-基)嘧啶-2-胺盐酸盐(S-19)的合成
Figure PCTCN2022084057-appb-000052
参考实施例4的合成方法，产率为100％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ12.00(s,1H),9.85(s,2H),9.30(dd,J＝8.0,1.7Hz,1H),8.69(d,J＝3.6Hz,1H),8.65(d,J＝2.2Hz,1H),8.55(d,J＝2.3Hz,1H),8.47–8.42(m,2H),8.00(d,J＝9.0Hz,1H),7.39(dd,J＝8.0,4.6Hz,1H),5.25–5.18(m,1H),4.51(s,2H),3.49–3.46(m,10H),1.60(d,J＝6.7Hz,6H).
实施例20：(4-乙基哌嗪-1-基)(6-((5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)甲酮(S-20)的合成
Figure PCTCN2022084057-appb-000053
参考实施例1的合成方法，产率为52％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.90(dd,J＝8.0,1.7Hz,1H),8.66(s,1H),8.50–8.46(m,2H),8.43(dd,J＝4.7,1.6Hz,1H),8.37(d,J＝3.5Hz,1H),8.20(d,J＝2.2Hz,1H),7.81(dd,J＝8.7,2.4Hz,1H),7.23(dd,J＝8.0,4.7Hz,1H),5.33–5.26(m,1H),3.88–3.83(m,4H),2.70–2.64(m,4H),1.62(d,J＝6.8Hz,6H),1.24–1.21(m,3H).
实施例21：(6-((5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)(4-异丙基哌嗪-1-基)甲酮(S-21)的合成
Figure PCTCN2022084057-appb-000054
参考实施例1的合成方法，产率为50％，白色固体。 1H NMR(300MHz,Chloroform-d)δ8.92(dd,J＝8.0,1.6Hz,1H),8.51–8.43(m,1H),8.46–8.42(m,2H),8.40(s,1H),8.35(d,J＝3.5Hz,1H),8.20(d,J＝2.2Hz,1H),7.81(ddd,J＝8.8,2.2,0.5Hz,1H),7.25–7.22(m,1H),5.34–5.25(m,1H),3.74–3.65(m,4H),2.81–2.72(m,1H),2.58–2.55(m,4H),1.62(d,J＝6.8Hz,6H),1.07(d,J＝6.5Hz,6H).
实施例22：(6-((5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)(4-甲基哌嗪-1-基)甲酮(S-22)的合成
Figure PCTCN2022084057-appb-000055
参考实施例1的合成方法，产率为48％。 1H NMR(300MHz,Chloroform-d)δ8.91(dd,J＝8.0,1.7Hz,1H),8.48–8.46(m,1H),8.46–8.42(m,3H),8.35(d,J＝3.5Hz,1H),8.20(d,J＝2.3Hz,1H),7.82–7.78(m,1H),7.24(dd,J＝8.0,4.7Hz,1H),5.34–5.25(m,1H),3.73–3.63(m,4H),2.48–2.46(m,4H),2.35(s,3H),1.62(d,J＝6.8Hz,6H).
实施例23：叔丁基4-(6-((5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)烟酰)哌嗪-1-羧酸酯(S-23)的合成
Figure PCTCN2022084057-appb-000056
参考实施例1的合成方法，产率为52％。 1H NMR(300MHz,Chloroform-d)δ8.91(dd,J＝8.0,1.6Hz,1H),8.51–8.45(m,3H),8.43(dd,J＝4.7,1.6Hz,1H),8.36(d,J＝3.5Hz,1H),8.20(d,J＝2.2Hz,1H),7.80(dd,J＝8.7,2.4Hz,1H),7.28–7.18(m,1H),5.34–5.25(m,1H),3.66–3.62(m,4H),3.51–3.47(m,4H),1.62(d,J＝6.8Hz,6H),1.48(s,9H).
实施例24：(6-((5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)(哌嗪-1-基)甲酮(S-24)的合成
Figure PCTCN2022084057-appb-000057
参考实施例4的合成方法，产率为100％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ11.81(s,1H),9.58(s,2H),9.33(dd,J＝8.0,1.6Hz,1H),8.69(d,J＝3.6Hz,1H),8.54(d,J＝2.2Hz,2H),8.45(dd,J＝4.7,1.6Hz,1H),8.18–8.15(m,1H),8.08(d,J＝8.9Hz,1H),7.36(dd,J＝8.0,4.6Hz,1H),5.25–5.18(m,1H),3.80–3.77(m,4H),3.20–3.17(m,4H),1.60(d,J＝6.8Hz,6H).
实施例25：叔丁基4-(6-((5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)哌嗪-1-羧酸酯(S-25)的合成
Figure PCTCN2022084057-appb-000058
参考实施例1的合成方法，产率为47％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.93(dd,J＝8.0,1.6Hz,1H),8.52–8.41(m,1H),8.29–8.26(m,2H),8.17(d,J＝2.2Hz,1H),8.13(s,0H),8.03(dd,J＝3.0,0.7Hz,1H),7.35(dd,J＝9.1,3.0Hz,1H),7.22(dd,J＝8.0,4.6Hz,1H),5.32–5.26(m,1H),3.61(t,J＝5.1Hz,4H),3.09(t,J＝5.1Hz,4H),1.61(d,J＝6.8Hz,6H),1.50(s,9H).
实施例26：5-氟-4-(1-异丙基-1氢-吡咯并吡啶-3-基)-N-(5-(哌嗪-1-基)吡啶-2-基)嘧啶-2-胺盐酸盐(S-26)的合成：
Figure PCTCN2022084057-appb-000059
参考实施例4的合成方法，产率为100％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ12.67(s,1H),10.17(s,2H),9.38(d,J＝7.9Hz,1H),8.63(d,J＝3.5Hz,1H),8.48(d,J＝2.1Hz,1H),8.43(d,J＝4.5Hz,1H),8.34(d,J＝9.6Hz,1H),8.04–8.02(m,2H),7.32(dd, J＝8.0,4.6Hz,1H),5.22–5.15(m,1H),3.56–3.53(m,4H),3.25–3.23(m,4H),1.59(d,J＝6.6Hz,6H).
实施例27：N-(5-((4-乙基哌嗪-1-基)甲基)吡啶-2-基)-5-氟-4-(5-氟-1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-胺(S-27)的合成
Figure PCTCN2022084057-appb-000060
参考实施例1的合成方法，产率为43％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.67(dd,J＝9.5,2.8Hz,1H),8.32(d,J＝3.5Hz,1H),8.29(dd,J＝2.8,1.3Hz,1H),8.26–8.24(m,2H),8.21(d,J＝2.1Hz,1H),8.02(s,1H),7.70(dd,J＝8.7,2.2Hz,1H),5.25–5.18(m,1H),3.51(s,2H),2.54–2.42(m,10H),1.61(d,J＝6.8Hz,6H),1.10(t,J＝7.2Hz,3H).
实施例28：(4-乙基哌嗪-1-基)(6-((5-氟-4-(5-氟-1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)甲酮(S-28)的合成
Figure PCTCN2022084057-appb-000061
参考实施例1的合成方法，产率为46％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.66(dd,J＝9.5,2.8Hz,1H),8.47(dd,J＝2.3,0.8Hz,1H),8.37(dd,J＝8.7,0.8Hz,1H),8.35(d,J＝3.5Hz,1H),8.30–8.29(m,2H),8.23(d,J＝2.1Hz,1H),7.81(dd,J＝8.7,2.3Hz,1H),5.26–5.19(m,1H),3.75–3.65(m,4H),2.51–2.45(m,6H),1.61(d,J＝6.8Hz,6H),1.12(t,J＝7.2Hz,3H).
实施例29：(6-((5-氟-4-(5-氟-1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)(4-异丙基哌嗪-1-基)甲酮(S-29)的合成
Figure PCTCN2022084057-appb-000062
参考实施例1的合成方法，产率为43％，白色固体。 1H NMR(300MHz,Chloroform-d)δ8.66(dd,J＝9.4,2.8Hz,1H),8.48(dd,J＝2.3,0.8Hz,1H),8.39–8.35(m,3H),8.30(dd,J＝2.8,1.3Hz,1H),8.23(d,J＝2.1Hz,1H),7.81(ddd,J＝8.6,2.3,0.5Hz,1H),5.27–5.18(m,1H),3.80–3.56(m,4H),2.80–2.71(m,1H),2.61–2.53(m,4H),1.61(d,J＝6.8Hz,6H),1.07(d,J＝6.6Hz,6H).
实施例30：叔丁基4-(6-((5-氟-4-(5-氟-1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)烟酰)哌嗪-1-羧酸酯(S-30)的合成
Figure PCTCN2022084057-appb-000063
参考实施例1的合成方法，产率为45％。 1H NMR(300MHz,Chloroform-d)δ8.67–8.63(m,2H),8.51(dd,J＝2.3,0.8Hz,1H),8.40–8.37(m,2H),8.30(dd,J＝2.8,1.3Hz,1H),8.23(d,J＝2.1Hz,1H),7.80(dd,J＝8.7,2.4Hz,1H),5.27–5.18(m,1H),3.71–3.61(m,4H),3.54–3.49(m,4H),1.62(d,J＝6.8Hz,6H),1.49(s,9H).
实施例31：(6-((5-氟-4-(5-氟-1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)(哌嗪-1-基)甲酮(S-31)的合成
Figure PCTCN2022084057-appb-000064
参考实施例4的合成方法，产率为100％，白色固体。 1H NMR(300MHz,DMSO-d 6)δ12.75(s,1H),9.93(s,2H),9.22–9.17(m,1H),8.69(d,J＝3.4Hz,1H),8.62–8.59(m,2H),8.44(s,1H),8.34(dd,J＝8.9,2.2Hz,1H),8.04(d,J＝9.0Hz,1H),5.19–5.10(m,1H),3.88–3.78(m,4H),3.24–3.16(m,4H),1.59(d,J＝6.7Hz,6H).
实施例32：N-(5-((4-乙基哌嗪-1-基)甲基)吡啶-2-基)-4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-胺(S-32)的合成
Figure PCTCN2022084057-appb-000065
参考实施例1的合成方法，产率为41％。 1H NMR(300MHz,Chloroform-d)δ8.70(dd,J＝8.0,1.6Hz,1H),8.46(s,1H),8.43(d,J＝2.5Hz,1H),8.40(dd,J＝4.7,1.6Hz,1H),8.25(d,J＝2.2Hz,1H),8.17(s,1H),8.05(s,1H),7.69(dd,J＝8.6,2.3Hz,1H),7.22(dd,J＝8.0,4.7Hz,1H),7.11(d,J＝5.4Hz,1H),5.33–5.24(m,1H),3.51(s,2H),2.56–2.44(m,10H),1.61(d,J＝6.8Hz,6H),1.12(t,J＝7.2Hz,3H).
实施例33：4-(1-异丙基-1氢-吡咯并吡啶-3-基)-N-(5-((4-异丙基哌嗪-1-基)甲基)吡啶-2-基)嘧啶-2-胺(S-33)的合成
Figure PCTCN2022084057-appb-000066
参考实施例1的合成方法，产率为42％。 1H NMR(300MHz,Chloroform-d)δ8.70(dd,J＝8.0,1.6Hz,1H),8.45(d,J＝3.7Hz,1H),8.43(s,1H),8.41(dd,J＝4.7,1.6Hz,1H),8.24(d,J＝2.2Hz,1H),8.06(s,1H),8.04(s,1H),7.67(dd,J＝8.6,2.3Hz,1H),7.23(dd,J＝8.0,4.7Hz,1H),7.12(d,J＝5.3Hz,1H),5.33–5.24(m,1H),3.54(s,2H),2.95–2.61(m,9H),1.61(d,J＝6.8Hz,6H),1.20(d,J＝6.5Hz,6H).
实施例34：(4-乙基哌嗪-1-基)(6-((4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)甲酮(S-34)的合成
Figure PCTCN2022084057-appb-000067
参考实施例1的合成方法，产率为47％，白色固体。 1H NMR(300MHz,Chloroform-d)δ9.19(s,1H),8.67(dd,J＝8.0,1.6Hz,1H),8.60(d,J＝8.7Hz,1H),8.55–8.52(m,2H),8.41(dd,J＝4.6,1.5Hz,1H),7.84(dd,J＝8.6,2.3Hz,1H),7.21(dd,J＝8.0,4.7Hz,1H),7.16(d,J＝5.3Hz,1H),5.33–5.24(m,1H),3.88–3.55(m,4H),2.49–2.43(m,6H),1.61(d,J＝6.7Hz,6H),1.11(t,J＝7.1Hz,3H).
实施例35：(6-((4-(1-异丙基-1氢-吡咯并吡啶-3-基)嘧啶-2-基)胺)吡啶-3-基)(4-异丙基哌嗪-1-基)甲酮(S-35)的合成
Figure PCTCN2022084057-appb-000068
参考实施例1的合成方法，产率为45％，白色固体。 1H NMR(300MHz,Chloroform-d)δ8.69(dd,J＝8.0,1.6Hz,1H),8.57(dd,J＝8.7,0.9Hz,1H),8.47(d,J＝5.4Hz,1H),8.44(dd,J＝2.3,0.8Hz,1H),8.41(dd,J＝4.7,1.6Hz,1H),8.25(s,1H),8.07(s,1H),7.82(dd,J＝8.7,2.3Hz,1H),7.24(dd,J＝8.0,4.7Hz,1H),7.17(d,J＝5.4Hz,1H),5.33–5.24(m,1H),3.83–3.53(m,4H),2.79–2.71(m,1H),2.57–2.54(m,4H),1.62(d,J＝6.8Hz,6H),1.07(d,J＝6.5Hz,6H).
实施例36：N-(5-((4-乙基哌嗪-1-基)甲基)吡啶-2-基)-4-(1-异丙基-1氢-吡咯并吡啶-3-基)-5-甲氧基嘧啶-2-胺(S-36)的合成
Figure PCTCN2022084057-appb-000069
参考实施例1的合成方法，产率为47％。 1H NMR(300MHz,Chloroform-d)δ9.01(dd,J＝8.0,1.7Hz,1H),8.40–8.38(m,2H),8.34(d,J＝8.6Hz,1H),8.22(d,J＝2.2Hz,1H),8.17(s,1H),8.06(s,1H),7.65(dd,J＝8.6,2.3Hz,1H),7.19(dd,J＝8.0,4.7Hz,1H),5.32–5.23(m,1H),4.02(s,3H),3.48(s,2H),2.52–2.40(m,10H),1.61(d,J＝6.7Hz,6H),1.09(t,J＝7.2Hz,3H).
实施例37：(4-乙基哌嗪-1-基)(6-((4-(1-异丙基-1氢-吡咯并吡啶-3-基)-5-甲氧基嘧啶-2-基)胺)吡啶-3-基)甲酮(S-37)的合成
Figure PCTCN2022084057-appb-000070
参考实施例1的合成方法，产率为43％，白色固体。 1H NMR(400MHz,Chloroform-d)δ9.00(dd,J＝8.0,1.7Hz,1H),8.46(dd,J＝8.7,0.8Hz,1H),8.44–8.43(m,1H),8.41–8.40(m,2H),8.28(s,1H),8.20(s,1H),7.77(dd,J＝8.7,2.4Hz,1H),7.21(dd,J＝8.0,4.6Hz,1H),5.32–5.25(m,1H),4.04(s,3H),3.82–3.59(m,4H),2.49–2.44(m,6H),1.61(d,J＝6.8Hz,6H),1.11(t,J＝7.2Hz,3H).
实施例38：(6-((4-(1-异丙基-1氢-吡咯并吡啶-3-基)-5-甲氧基嘧啶-2-基)胺)吡啶-3-基)(4-异丙基哌嗪-1-基)甲酮(S-38)的合成
Figure PCTCN2022084057-appb-000071
参考实施例1的合成方法，产率为42％。 1H NMR(300MHz,Chloroform-d)δ8.98(dd,J＝8.0,1.7Hz,1H),8.70(s,1H),8.48–8.45(m,2H),8.40–8.39(m,2H),8.22(s,1H),7.78(dd,J＝8.6,2.4Hz,1H),7.19(dd,J＝8.0,4.7Hz,1H),5.32–5.23(m,1H),4.01(s,3H),3.77–3.60(m,4H),2.78–2.70(m,1H),2.57–2.50(m,4H),1.61(d,J＝6.8Hz,6H),1.06(d,J＝6.5Hz,6H).
实施例39：4-(1-(仲丁基)-1氢-吡咯并吡啶-3-基)-5-氟-N-(5-((4-异丙基哌嗪-1-基)甲基)吡啶-2-基)嘧啶-2-胺(S-39)的合成
Figure PCTCN2022084057-appb-000072
参考实施例1的合成方法，产率为45％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.94(dd,J＝8.0,1.6Hz,1H),8.41(dd,J＝4.6,1.6Hz,1H),8.34–8.32(m,2H),8.25–8.21(m,2H),8.14(d,J＝2.2Hz,1H),7.68(dd,J＝8.6,2.3Hz,1H),7.24–7.21(m,1H),5.12–5.03(m,1H),3.49(s,2H),2.70–2.53(m,9H),2.02–1.93(m,2H),1.59(d,J＝6.8Hz,3H),1.06(d,J＝6.5Hz,6H),0.89(t,J＝7.4Hz,3H).
实施例40：(6-((4-(1-(仲丁基)-1氢-吡咯并吡啶-3-基)-5-氟嘧啶-2-基)胺)吡啶-3-基)(4-乙基哌嗪-1-基)甲酮(S-40)的合成
Figure PCTCN2022084057-appb-000073
参考实施例1的合成方法，产率为51％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.92(dd,J＝8.0,1.7Hz,1H),8.60(s,1H),8.48–8.45(m,2H),8.42(dd,J＝4.6,1.6Hz,1H),8.37(d,J＝3.5Hz,1H),8.16(d,J＝2.2Hz,1H),7.81(dd,J＝8.6,2.4Hz,1H),7.23(dd,J＝8.0,4.7Hz,1H),5.11–5.05(m,1H),3.78–3.61(m,4H),2.50–2.45(m,6H),2.03–1.94(m,2H),1.59(d,J＝6.9Hz,3H),1.12(t,J＝7.2Hz,3H),0.89(t,J＝7.3Hz,3H).
实施例41：(6-((4-(1-(仲丁基)-1氢-吡咯并吡啶-3-基)-5-氟嘧啶-2-基)胺)吡啶-3-基)(4-异丙基哌嗪-1-基)甲酮(S-41)的合成
Figure PCTCN2022084057-appb-000074
参考实施例1的合成方法，产率为52％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.93(dd,J＝8.0,1.7Hz,1H),8.46–8.42(m,3H),8.34(d,J＝3.5Hz,1H),8.20(s,1H),8.16(d,J＝2.2Hz,1H),7.80(dd,J＝8.6,2.4Hz,1H),7.25–7.23(m,1H),5.12–5.04(m,1H),3.81–3.54(m,4H),2.78–2.72(m,1H),2.62–2.49(m,4H),2.03–1.94(m,2H),1.59(d,J＝6.9Hz,3H),1.07(d,J＝6.5Hz,6H),0.89(t,J＝7.4Hz,3H).
实施例42：叔丁基4-(6-((4-(1-(仲丁基)-1氢-吡咯并吡啶-3-基)-5-氟嘧啶-2-基)胺)烟酰)哌嗪-1-羧酸酯(S-42)的合成
Figure PCTCN2022084057-appb-000075
参考实施例1的合成方法，产率为50％。 1H NMR(400MHz,Chloroform-d)δ8.92(dd,J＝8.0,1.6Hz,1H),8.48(d,J＝8.7Hz,1H),8.45–8.42(m,2H),8.34(d,J＝3.5Hz,1H),8.28(s,1H),8.16(d,J＝2.2Hz,1H),7.80(dd,J＝8.7,2.4Hz,1H),7.24(dd,J＝8.1,4.8Hz,1H),5.13–5.04(m,1H),3.70–3.47(m,8H),2.03–1.94(m,2H),1.59(d,J＝6.8Hz,3H),1.48(s,9H),0.89(t,J＝7.4Hz,3H).
实施例43：(6-((4-(1-(仲丁基)-1氢-吡咯并吡啶-3-基)-5-氟嘧啶-2-基)胺)吡啶-3-基)(哌嗪-1-基)甲酮(S-43)的合成：
Figure PCTCN2022084057-appb-000076
参考实施例4的合成方法，产率为100％，白色固体。 1H NMR(400MHz,DMSO-d 6)δ12.43(s,1H),9.82(s,2H),9.38(dd,J＝8.0,1.6Hz,1H),8.71(d,J＝3.5Hz,1H),8.57(dd,J＝16.0,2.2Hz,2H),8.45(dd,J＝4.7,1.6Hz,1H),8.28(dd,J＝8.9,2.2Hz,1H),8.08(d,J＝9.0Hz,1H),7.36(dd,J＝8.0,4.6Hz,1H),5.04–4.98(m,1H),3.86–3.77(m,4H),3.21–3.17(m,4H),2.11–1.92(m,2H),1.59(d,J＝6.8Hz,3H),0.76(t,J＝7.3Hz,3H).
实施例44：4-(1-环戊基-1氢-吡咯并吡啶-3-基)-5-氟-N-(5-((4-异丙基哌嗪-1-基)甲基)吡啶-2-基)嘧啶-2-胺(S-44)的合成
Figure PCTCN2022084057-appb-000077
参考实施例1的合成方法，产率为43％，白色固体。 1H NMR(400MHz,Chloroform-d)δ8.92(dd,J＝8.0,1.7Hz,1H),8.43(dd,J＝4.7,1.6Hz,1H),8.35(d,J＝8.6Hz,1H),8.31(d,J＝3.5Hz,1H),8.24(d,J＝2.3Hz,1H),8.18(d,J＝2.1Hz,1H),8.07(s,1H),7.65–7.62(m,1H),7.24(dd,J＝8.0,4.7Hz,1H),5.39–5.35(m,1H),3.60(s,2H),3.38–2.97(m,9H),2.37–2.30(m,2H),2.00–1.94(m,4H),1.87–1.83(m,2H),1.43(d,J＝6.6Hz,6H).
三、生物学评价实验
(1)CDK6激酶活性分析检测方法
为考察实施例化合物S1-S44对CDK6/cyclinD1蛋白激酶的抑制作用，采用PerkinElmer公司的Lance Ultra方法进行检测。在检测板中将蛋白激酶、Ulight标记的多肽底物、ATP及化合物混合，并孵育反应。之后，加入EDTA终止反映，并加入铕(Eu)螯合物标记的抗体进行检测。本实验采用PerkinElmer公司的Envision仪器进行分析，模式为TR-FRET。在320/340nm波长激发后，可以发射665nm和615nm波长的荧光信号。Eu可以将能量转移至相邻的荧光物质Ulight受体上，再对该发射光进行检测。首先用DMSO将化合物溶解并稀释至2mM，然后3倍稀释10个浓度，使最终实验板中化合物浓度为10μM至0.508nM。再将CDK6/cyclinD1(Life Technologies)和Ulight-4E-BPI-peptide substrate(珀金埃尔默)分别用反应缓冲液(50mM Hepes pH 7.5，10mM MgCl 2，0.01％Birj-35，1mM EDTA，2mM DTT)稀释至1nM和50nM，取5μl多肽和激酶混合液与100μL不同浓度的化合物在OptiPlate-384微孔板中混合，在室温下孵育15分中后加入5μL的350μM ATP溶液，室温孵育90min，最后加入10μL检测溶液(2nM Eu-labeled antibody，10mM EDTA以及1×检测缓冲液)室温孵育60min后检测。IC50结果用IDBS公司的XLfit5进行分析。
测得的IC 50值如下表1，从实验结果可以看出，本发明实例化合物对CDK6激酶活性具有很强的抑制活性。
表1 本发明化合物对CDK6激酶活性的IC 50测量值
实施例 IC 50(nM) 实施例 IC 50(nM) 实施例 IC 50(nM)
1 130 16 30 31 32
2 120 17 10 32 42
3 >10000 18 >10000 33 27
4 251 19 33 34 45
5 >10000 20 35 35 36
6 56 21 25 36 2622
7 46 22 38 37 2200
8 440 23 >1000 38 1869
9 >10000 24 18 39 36
10 905 25 >10000 40 46
11 >10000 26 86 41 25
12 445 27 53 42 >1000
13 113 28 45 43 69
14 126 29 40 44 24
15 87 30 >1000 / /
由表可见，本发明化合物对CDK6激酶用非常良好的抑制活性，其中实施例17对CDK6的抑制活性最强，将其进行进一步的体内活性评估。
(2)化合物急性毒性测定
受试动物：ICR小鼠(由上海斯莱克实验动物有限责任公司提供)；18-22g；雌性；共20只。
受试样品：实施例17
组别剂量设置：一共设置四组，对照组；1000mg/kg组；5000mg/kg组：10000mg/kg组：灌胃药物，给药组每组给药1次，对照组给予等量溶媒，每组5只小鼠，观察14天；
实验结果：各组小鼠给药后12个小时之内，动物未见异常。给药24个小时内未见动物死亡，给药第14天后动物未见死亡。未见其他明显异常。因此，受试药物灌胃给予1000mg/kg，5000mg/kg，10000mg/kg未见毒性反应，实施例17的安全性良好。
(3)化合物抗多发性骨髓瘤活性测定
受试样品：实施例17和阳性药Palbociclib
实验方法：取生长旺盛期的肿瘤，在无菌条件下，人多发性骨髓瘤PRMI8226细胞接种48只BALB/c裸小鼠右侧腋窝皮下，细胞接种量为5×10 6。裸小鼠移植瘤用游标卡尺测量移植瘤直径，待肿瘤生长至85mm 3左右时挑选生长状态良好且肿瘤大小均一性较好的荷瘤裸鼠24只，分成4组，每组6只，即(1)模型对照组(给予等量溶媒)；(2)阳性药Palbociclib组(给药量为100mg/kg)；(3)实施例17低剂量组(给药量为100mg/kg)；(4)实施例17高剂量组(给药量为200mg/kg)。使用测量瘤径的方法，动态观察受试物的抗肿瘤效应。肿瘤直径的测量次数为隔天一次，测量肿瘤直径的同时称量裸鼠体重。给药23天后脱颈处死裸鼠，手术剥取瘤块称重。
实验结果显示：实施例17剂量为100mg/kg，每天灌胃给药1次，给药23天时，对人多发性骨髓瘤细胞PRMI8226裸小鼠移植瘤的T/C(％)为58.3％，抑瘤率为34.1％；实施例17剂量为200mg/kg，每天灌胃给药1次，给药23天时，对人多发性骨髓瘤细胞PRMI8226裸小鼠移植瘤的T/C(％)为33.8％，抑瘤率为61.5％。阳性药Palbociclib剂量为100mg/kg，每天灌胃给药1次，给药23天时，对人多发性骨髓瘤细胞PRMI8226裸小鼠移植瘤的T/C(％)为37.9％，抑瘤率为55.6％。实验结果显示，各组裸鼠给药期间体重未见明显下降。因此，实施例17对人多发性骨髓瘤细胞PRMI8226裸鼠异种移植瘤生长有显著的抑制作用，并且实施例17高剂量组对人多发性骨髓瘤PRMI8226异种移植瘤荷瘤裸鼠抗肿瘤活性强于阳性药Palbociclib组。

Claims (10)

  1. 一种如式(S)所示的化合物或其药学上可接受的盐，其特征在于：
    Figure PCTCN2022084057-appb-100001
    其中，
    所述X选自C(O)或(CH 2) n；n为0-8；
    所述R 1选自氢或卤素；
    所述R 2选自氢、C 1-C 8烷基或C 3-C 6环烷基；
    所述R 3选自氢或卤素；
    所述R 4选自氢或C 1-C 8烷基。
  2. 根据权利要求1所述的化合物或其药学上可接受的盐，其特征在于：所述X为C(O)或(CH 2) n；n为0-4。
  3. 根据权利要求1所述的化合物或其药学上可接受的盐，其特征在于：所述R 1为氢或氟。
  4. 根据权利要求1所述的化合物或其药学上可接受的盐，其特征在于：所述R 2为氢、C 1-C 4烷基、环戊烷基或环己烷基。
  5. 根据权利要求1所述的化合物或其药学上可接受的盐，其特征在于：所述R 3为氢或氟。
  6. 根据权利要求1所述的化合物或其药学上可接受的盐，其特征在于：所述R 4为氢或C 1-C 3烷基。
  7. 根据权利要求1所述的化合物或其药学上可接受的盐，其特征在于：所述X选自C(O)或(CH 2) n；n为0或1；
    所述R 1选自氢或氟；
    所述R 2选自氢、甲基、乙基、异丙基、异丁基或环戊烷基；
    所述R 3选自氢或氟；
    所述R 4选自：氢、甲基、乙基或异丙基。
  8. 根据权利要求1所述的化合物或其药学上可接受的盐，其特征在于：选自以下化合物：
    Figure PCTCN2022084057-appb-100002
    Figure PCTCN2022084057-appb-100003
    Figure PCTCN2022084057-appb-100004
    Figure PCTCN2022084057-appb-100005
  9. 一种权利要求1所述的化合物的制备方法，其特征在于：由化合物(A)与化合物(B)在钯催化剂的作用下经偶联反应制备化合物(S)：
    Figure PCTCN2022084057-appb-100006
    所述X选自C(O)或(CH 2) n；n为0-8；
    所述R 1选自氢或卤素；
    所述R 2选自氢、C 1-C 8烷基或C 3-C 6环烷基；
    所述R 3选自氢或卤素；
    所述R 4选自氢或C 1-C 8烷基。
  10. 权利要求1～8任一项所述的化合物或其药学上可接受的盐或权利要求9制备的化合物在制备CDK6激酶抑制剂中的应用；所述CDK6激酶抑制剂用于治疗癌症或肿瘤相关疾病；所述癌症或肿瘤相关疾病包括多发性骨髓瘤、白血病、乳腺癌、前列腺癌、肺癌、肝癌、胃癌、骨癌、脑癌、头颈癌、肠癌、胰腺癌、膀胱癌、睾丸癌、卵巢癌以及子宫内膜癌。
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