CN111423454B - 哌嗪类化合物及其在制备趋化因子受体ccr2拮抗剂中的应用 - Google Patents

哌嗪类化合物及其在制备趋化因子受体ccr2拮抗剂中的应用 Download PDF

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CN111423454B
CN111423454B CN202010333530.3A CN202010333530A CN111423454B CN 111423454 B CN111423454 B CN 111423454B CN 202010333530 A CN202010333530 A CN 202010333530A CN 111423454 B CN111423454 B CN 111423454B
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龙亚秋
谢欣
秦立怀
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Abstract

本发明涉及一种式(I)所示的哌嗪类化合物及其在制备趋化因子受体CCR2拮抗剂或用于治疗由CCR2介导的疾病的药物中的应用。

Description

哌嗪类化合物及其在制备趋化因子受体CCR2拮抗剂中的应用
技术领域
本发明涉及生物医药领域，尤其涉及一种哌嗪类化合物及其在制备趋化因子受体CCR2拮抗剂中的应用。
背景技术
趋化因子也称化学性趋化细胞因子，属于细胞因子大家族的一员，含有340-380个氨基酸。目前已至少发现40多种亚型，根据结构和功能的不同分为两个大类(CC和CXC)与两个小类(CX3C和C)。趋化因子与趋化因子受体构成一个非常复杂的网络，一个趋化因子可以结合不同的受体，一个趋化因子受体可以识别不同的配体。它们可以在免疫反应中调控免疫细胞的活化和运动，因此在免疫疾病中起着重要的作用。正是由于它们在疾病中的重要作用，使它们成为一类许多大的制药公司竞相开放的一类药物靶标。
CCR2属于CC类趋化因子受体，在多种细胞中高表达，主要是单核细胞、巨噬细胞、嗜碱性粒细胞、不成熟树突状细胞、记忆T细胞等。CCR2根据mRNA选择性剪切碳端的不同，可以分为CCR2A与CCR2B两种亚型。CCR2A含有360个氨基酸，CCR2B含有374个氨基酸。它们的氨基酸序列在313位之前都是相同的，在生命体中发挥主要作用是CCR2B。CCL2同样可以由很多细胞产生，包括上皮细胞、内皮细胞、平滑肌细胞、成纤维细胞和单核细胞等。并且可以诱导单核细胞、记忆T细胞和树突状细胞到达炎症区域。在动物模型中已经证明阻断CCL2/CCR2生物轴可以用来治疗多种炎症相关疾病，包括：风湿性关节炎、动脉粥样硬化、哮喘、肥胖、二型糖尿病、慢性神经退行性疾病和肿瘤等。因此拮抗CCR2是一个非常有前景的治疗策略，在过去的十多年里，众多的跨国医药公司竞相研发其抑制剂。
发明内容
为解决上述技术问题，本发明的目的是提供一种哌嗪类化合物及其在制备趋化因子受体CCR2拮抗剂中的应用。
本发明的一种哌嗪类化合物，其结构式如式(I)所示：
Figure BDA0002465807210000011
其中，A为芳基、杂芳基、苯并杂芳基、取代芳基、取代杂芳基或取代苯并杂芳基；所述杂芳基、苯并杂芳基、取代杂芳基或取代苯并杂芳基上的杂原子选自N、O或S原子；所述取代芳基、取代杂芳基或取代苯并杂芳基含有1-3个取代基；所述取代基独立地选自卤素、芳基、杂芳基、氰基、羟基、硝基、三氟甲基、C1-C4直链或支链烷基和烷氧基中的一种或几种；
L选自
Figure BDA0002465807210000021
a＝0或1；
X和Y分别独立地选自羰基或亚甲基，或者X、Y和与其相连的哌嗪环上的N原子形成五元杂芳基，五元杂芳基中的杂原子选自N、O或S原子；
Z为C原子或N原子；当Z为N原子时，b＝0；当Z为C原子时，b＝1；
R1和R2各自独立地选自氢原子、卤素、氰基、羟基、硝基、氨基、C1-C6烷基、C1-C6烷氧基、C1-C6烷基取代的酰胺基或C1-C6酯基；
R3为氢、卤素、氰基、羟基、硝基、氨基、C1-C6烷基、C1-C6烷氧基或苯基；
R4和R5各自独立地选自氢原子、羟基、氨基、卤素、C1-C3直链或支链烷基、杂原子取代的C3-C8环烷基、芳基、杂芳基、苯并杂芳基、取代芳基、取代杂芳基或取代苯并杂芳基；所述取代芳基、取代杂芳基或取代苯并杂芳基含有一个或多个取代基；所述取代基独立地选自C1-C6烷基、C1-C6烷氧基、C3-C8环烷基、杂原子取代的C3-C8环烷基、-CF3、-CN、-NO2、-NO或卤素；所述杂原子取代的C3-C8环烷基中的杂原子为O、N或S原子；或者
R4和R5形成以下螺环结构：
Figure BDA0002465807210000022
其中W选自O原子、S原子、SO基、SO2基或羰基；k为1或2；R9为氢原子、C1-C6烷基、C1-C6烷氧基、卤素或氰基；
R6，R7和R8分别独立地选自氢原子、卤素、氰基、羟基、硝基、氨基、C1-C6烷基、C1-C6烷氧基、芳基、杂芳基、取代芳基或取代杂芳基，所述取代芳基或取代杂芳基含有1-3个取代基；所述取代基分别独立地选选自氢原子、卤素、氰基、羟基、硝基、氨基或C1-C6烷基；
或者R5、R6和与二者相连的原子连接成苯基或苯并杂芳基；所述苯基或苯并杂芳基上含有至少一个取代基，所述取代基包括氢原子、氨基、卤素、三氟甲基和氰基中的一种或几种；
或者R6、R7和与二者相连的原子连接成苯基或苯并杂芳基；所述苯基或苯并杂芳基上含有至少一个取代基，所述取代基包括氢原子、氨基、卤素、三氟甲基和氰基中的一种或几种；
m和n分别独立地选自0-2中任一整数，且m+n≤3。
优选地，A为芳基、杂芳基、苯并杂芳基、取代芳基、取代杂芳基或取代苯并杂芳基；所述杂芳基、苯并杂芳基、取代杂芳基或取代苯并杂芳基上的杂原子选自N、O或S原子；所述取代芳基、取代杂芳基或取代苯并杂芳基含有1-3个取代基；所述取代基独立地选自卤素、芳基、杂芳基和三氟甲基中的一种或几种。
优选地，A为取代苯基或取代苯并杂芳基，取代苯基或取代苯并杂芳基上含有1-3个取代基，所述取代基选自卤素或三氟甲基。
优选地，L选自
Figure BDA0002465807210000031
a＝0或1。
优选地，X和Y分别独立地选自羰基或亚甲基，或者X、Y和与其相连的哌嗪环上的N原子形成五元杂芳基，五元杂芳基中的杂原子选自N或O原子。
优选地，R1和R2均为氢原子；或R1和R2其中之一为氢原子，另一个为卤素、C1-C6烷基、C1-C6烷氧基、C1-C6烷基取代的酰胺基或C1-C6酯基。
优选地，W选自O原子、S原子、SO基或SO2基；R9为氢原子、C1-C6烷基、卤素或氰基。
本发明中，基团结构式中的“*”表示该基团与其他基团或原子的连接位点。
最优选地，本发明的哌嗪类化合物为S1-S45所示的结构式：
Figure BDA0002465807210000032
Figure BDA0002465807210000041
本发明还公开了式(I)所示的哌嗪类化合物在制备趋化因子受体CCR2拮抗剂中的应用。
本发明还公开了式(I)所示的哌嗪类化合物在制备用于治疗由CCR2介导的疾病的药物中的应用。
进一步地，由CCR2介导的疾病包括风湿性关节炎、动脉粥样硬化、哮喘、肥胖、二型糖尿病、慢性神经退行性疾病和肿瘤中的一种或几种。
借由上述方案，本发明至少具有以下优点：
本发明公开了一类新型哌嗪类化合物，该化合物可用于制备趋化因子受体CCR2拮抗剂或制备用于治疗由CCR2介导的疾病的药物。
上述说明仅是本发明技术方案的概述，为了能够更清楚了解本发明的技术手段，并可依照说明书的内容予以实施，以下以本发明的较佳实施例并配合详细说明如后。
具体实施方式
下面结合实施例，对本发明的具体实施方式作进一步详细描述。以下实施例用于说明本发明，但不用来限制本发明的范围。
本发明以下实施例中，所有原料可以从商业途径获得，或者通过本领域已知的方法制备，或根据本文所述方法制备。
化合物的结构通过核磁共振(1H-NMR)和/或质谱(MS)来确定。NMR测定是用Varian公司的Mercury-400型核磁共振仪，测定溶剂为氘代氯仿(CDCl3)、氘代甲醇(CD3OD)、氘代二甲亚砜(DMSO-d6)或氘代乙腈(CD3CN)，TMS为内标。MS的测定用Thermo Finnigan LCQ-DecaXP型(ESI)液相色谱-质谱联用仪。柱层析分离纯化产物使用的是ISCO
Figure BDA0002465807210000052
Rf 75快速制备色谱仪，载体采用青岛海洋化工厂的200-300目硅胶。
本发明以下实施例中，S1-S45的结构式与说明书上文中的S1-S45所示的结构式相对应。
实施例1：
Figure BDA0002465807210000051
以上反应路线中，所使用的试剂与条件如下：a)1,3-二溴丙烷，碳酸钾，丙酮；b)盐酸甲醇，3,5-二氟苯乙烯酸，1-羟基苯并三唑，N,N-二异丙基乙胺，1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐；c)硫脲，氢氧化钠，乙醇，微波140℃；d)正丁基锂，N-乙氧羰基-4-哌啶酮，四氢呋喃，-78℃；e)三氟乙酸，二氯甲烷；f)5N氢氧化钠溶液，乙醇，回流。反应步骤具体如下：
步骤1：合成叔-丁基4-(3-溴丙基)哌嗪-1-羧酸酯(式ii所示)
N-Boc-哌嗪(式i所示，186mg,1mmol)溶于10mL的丙酮中，依次加入1,3-二溴丙烷(0.85mL,5mmol)，碳酸钾(276mg,2mmol)，室温搅拌过夜，旋干丙酮，加入15mL的水，乙酸乙酯萃取3次，合并有机相，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离。得到淡黄色液体，放置变成固体(200mg,65％)。1H NMR(400MHz,CDCl3)δ3.46(t,J＝6.6Hz,2H),3.43–3.37(m,4H),2.47(t,J＝7.0Hz,2H),2.41–2.31(m,4H),2.02(p,J＝6.7Hz,2H),1.45(s,9H).
步骤2：合成(E)-1-(4-(3-溴丙基)哌嗪-1-基)-3-(3,5-二氟苯基)丙-2-烯-1-酮(式iii所示)
取叔-丁基4-(3-溴丙基)哌嗪-1-羧酸酯(2.2g,7.3mmol)溶于20mL盐酸甲醇中，反应6h后，停止反应，直接旋干，未处理投入下一步。取3,5-二氟苯乙烯酸(1.34g,7.3mmol)溶于50mL的二氯甲烷，依次加入1-羟基苯并三唑(1.18g,8.76mmol)，N,N-二异丙基乙胺(3.6mL,22mmol)，1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐(1.67g,8.76mmol)，室温搅拌1h后。加入脱Boc产物。继续反应3h后，停止反应。用饱和食盐水洗涤，无水硫酸钠干燥。硅胶柱分离得到淡黄色固体(1.8g,68％)。1H NMR(300MHz,CDCl3)δ7.53(d,J＝15.4Hz,1H),7.05–6.96(m,2H),6.85(d,J＝15.4Hz,1H),6.77(tt,J＝8.7,2.3Hz,1H),3.80–3.57(m,5H),3.47(t,J＝6.6Hz,1H),2.57–2.42(m,6H),2.07–1.91(m,2H).
步骤3：合成(2-溴苯基)甲硫醇(式II所示)
2-溴溴苄(式I所示，4.37g,17.5mmol)与硫脲(1.6g,21mmol)溶于8mL乙醇中，混合后140℃微波反应30min后，冷却到室温，加入氢氧化钠(2.1g,52.5mmol)和4mL水，继续用微波照射10min后，停止反应，用盐酸调成酸性，乙酸乙酯萃取3次，合并有机相，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到无色液体(2g,57％)。1H NMR(400MHz,CDCl3)δ7.55(dd,J＝8.0,1.2Hz,1H),7.39–7.35(m,1H),7.31–7.27(m,1H),7.14–7.08(m,1H),3.83(d,J＝8.1Hz,2H),2.00(t,J＝8.1Hz,1H).
步骤4：合成乙基4-羟基-4-(2-(巯基甲基)苯基)哌啶-1-羧酸酯(式III所示)
取(2-溴苯基)甲硫醇(式II所示，1g,4.9mmol)溶于10mL的四氢呋喃中，冷却到-78℃中，加入正丁基锂(1.6M in hexane,6.5mL)，搅拌1h后，加入N-乙氧羰基-4-哌啶酮(1.5g,9mmol)的四氢呋喃溶液，继续搅拌2h后，停止反应，加入饱和氯化铵溶液淬灭，乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到无色液体。
步骤5：合成乙基2H-螺[苯并[b]噻吩-3,4'-哌啶]-1'-羧酸酯(式IV所示)
取乙基4-羟基-4-(2-(巯基甲基)苯基)哌啶-1-羧酸酯(式III所示，100mg,0.34mmol)溶于2mL的二氯甲烷中，冰浴条件下加入1mL的三氟乙酸，室温搅拌2h后，停止反应，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到无色液体(70mg,75％)。1H NMR(300MHz,CDCl3)δ7.31–7.21(m,3H),7.20–7.12(m,1H),4.36–4.15(m,6H),3.12(t,J＝13.3Hz,2H),2.18–2.01(m,2H),1.99–1.86(m,2H),1.31(t,J＝7.1Hz,3H).
步骤6：合成2H-螺[苯并[b]噻吩-3,4'-哌啶](式V所示)
取乙基2H-螺[苯并[b]噻吩-3,4'-哌啶]-1'-羧酸酯(式IV所示，450mg,1.6mmol)溶于乙醇中，加入5N的氢氧化钠溶液(4.5mL,21mmol)，回流反应16h后，停止反应，旋掉有机溶剂，用水稀释，二氯甲烷萃取4次，合并有机相，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到淡红色液体(200mg,61％)。1H NMR(400MHz,CDCl3)δ7.29–7.21(m,4H),4.17(s,2H),3.18–3.10(m,2H),2.95(td,J＝12.3,2.2Hz,2H),2.11(td,J＝13.0,4.2Hz,2H),1.95–1.87(m,2H).
步骤7：合成(E)-1-(4-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)哌嗪-1-基)-3-(3,5-二氟苯基)丙-2-烯-1-酮(S1)
取2H-螺[苯并[b]噻吩-3,4'-哌啶](式V所示，110mg,0.54mmol)、(E)-1-(4-(3-溴丙基)哌嗪-1-基)-3-(3,5-二氟苯基)丙-2-烯-1-酮(式III所示199mg,0.54mmol)和碳酸钾(111mg,0.8mmol)溶于15mLN,N-二甲基甲酰胺。室温反应过夜，停止反应，倒入冰水中，乙酸乙酯萃取3次，合并有机相，饱和食盐水洗涤2次，无水硫酸钠干燥，硅胶柱分离。得到白色泡沫状固体(150mg,57％)。1H NMR(400MHz,CDCl3)δ7.54(d,J＝15.4Hz,1H),7.25–7.19(m,4H),7.04–6.98(m,2H),6.86(d,J＝15.4Hz,1H),6.78(tt,J＝8.7,2.2Hz,1H),4.16(s,2H),3.79–3.60(m,4H),3.04–2.96(m,2H),2.52–2.37(m,8H),2.30–2.17(m,4H),1.96–1.88(m,2H),1.79–1.68(m,2H).13C NMR(151MHz,CDCl3)δ164.51,163.20(dd,J＝248.9,12.8Hz,2×C),148.05,140.30,140.17(s),138.60(t,J＝9.5Hz),127.39,127.05,125.15,123.15,119.82,110.39(dd,J＝20.3,5.1Hz,2×C),104.74(t,J＝25.6Hz),63.63,56.75,56.58,53.54,52.84,52.17,45.91,42.26,40.94,35.24,24.46.HRMS(ESI):calcd forC28H34F2N3OS[M+H]+:498.2385；found 498.2390.
实施例2：
Figure BDA0002465807210000071
试剂与条件：a)三乙胺，二碳酸二叔丁酯，二氯甲烷；b)0.5M高碘酸钠水溶液。
合成步骤：
步骤1：合成叔-丁基3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-羧酸酯(式VI所示)
取2H-螺[苯并[b]噻吩-3,4'-哌啶](式V所示，210mg,1mmol)溶于10mL的二氯甲烷中，0℃下加入三乙胺(208μL,1.5mmol)和二碳酸二叔丁酯(261mg,1.2mmol)，室温搅拌2h后，停止反应，饱和氯化铵溶液洗涤，无水硫酸钠干燥，硅胶柱分离，得到白色固体(240mg,79％)。1H NMR(400MHz,CDCl3)δ7.30–7.23(m,4H),7.18–7.14(m,1H),4.31–4.12(m,4H),3.09–2.93(m,2H),2.06(td,J＝13.0,4.4Hz,2H),1.91–1.84(m,2H),1.49(s,9H).
步骤2：叔-丁基3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-羧酸酯2-氧化(式VII所示)
取叔-丁基3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-羧酸酯(240mg,0.78mmol)溶于4mL的乙醇中，在0℃下加入事先配好的0.5M高碘酸钠水溶液(0.81mmol)，室温搅拌过夜后，旋掉有机溶剂，用水稀释后，乙酸乙酯萃取，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到白色固体(式VII所示，232mg,93％)。1H NMR(400MHz,CDCl3)δ7.38–7.31(m,3H),7.25–7.22(m,1H),4.37(d,J＝16.7Hz,1H),4.29–4.09(m,2H),4.05(d,J＝16.8Hz,1H),3.29–3.12(m,2H),2.47–2.39(m,1H),2.26–2.14(m,1H),1.95–1.83(m,1H),1.62–1.55(m,1H),1.50(s,9H).
步骤3：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(2-羟基-3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S2)
按照实施例1步骤7中的方法合成化合物S2，不同之处在于，将其中的式V所示化合物替换为等量的式VII所示化合物。1H NMR(400MHz,CDCl3)δ7.55(d,J＝15.3Hz,1H),7.37–7.28(m,4H),7.05–6.99(m,2H),6.87(d,J＝15.4Hz,1H),6.80(tt,J＝8.6,2.1Hz,1H),4.34(d,J＝16.7Hz,1H),4.02(d,J＝16.8Hz,1H),3.79–3.61(m,4H),3.10–2.93(m,2H),2.54–2.39(m,10H),1.80–1.71(m,2H),1.67–1.62(m,2H).13C NMR(126MHz,CDCl3)δ164.52,163.16(dd,J＝248.9,12.9Hz,2×C),144.02,140.23,138.59(t,J＝9.5Hz),135.13,128.73,128.61,126.94,124.80,119.86,110.36(dd,J＝19.7,6.0Hz,2×C),104.67(t,J＝25.5Hz),72.79,56.69,56.42,54.95,53.51,52.79,52.05,50.96,45.85,42.21,33.93,27.53,24.32.HRMS(ESI):calcd for C28H34F2N3O2S[M+H]+:514.2334；found 514.2336.
实施例3：
步骤1：取实施例1步骤5所得式IV所示化合物溶于10mL的二氯甲烷中，加入间氯过氧苯甲酸，室温搅拌1h后，停止反应，1N NaOH溶液洗涤，无水硫酸钠干燥，硅胶柱分离。得到白色固体，结构式为：
Figure BDA0002465807210000081
1H NMR(400MHz,CDCl3)δ7.43–7.32(m,2H),7.30–7.26(m,1H),7.22(d,J＝7.7Hz,1H),4.37(s,2H),4.24–4.08(m,4H),3.48(t,J＝13.4Hz,2H),2.42–2.34(m,2H),2.06–1.95(m,2H),1.29(t,J＝7.1Hz,3H).
步骤2：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(2,2-二羟基-3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S3)
以本实施例步骤1产物为起始原料，将其与实施例1中式iii所示化合物反应，操作步骤参考实施例1步骤7，得到S3。1H NMR(400MHz,CDCl3)δ7.54(d,J＝15.4Hz,1H),7.41–7.22(m,4H),7.05–6.97(m,2H),6.86(d,J＝15.4Hz,1H),6.82–6.73(m,1H),4.33(s,2H),3.79–3.60(m,4H),2.94–2.84(m,2H),2.63(t,J＝11.3Hz,2H),2.54–2.37(m,10H),2.20–2.07(m,2H),1.80–1.68(m,2H).13C NMR(126MHz,CDCl3)δ164.49,163.19(dd,J＝249.0,12.8Hz,2×C),141.49,140.26,138.61(t,J＝9.5Hz),129.12,128.80,128.45,125.66,124.20,119.85,110.37(dd,J＝19.7,6.0Hz,2×C),104.71(t,J＝25.6Hz),65.34,56.59,56.52,54.11,53.56,52.83,49.81,45.91,42.25,32.69,24.42.HRMS(ESI):calcd forC28H34F2N3O3S[M+H]+:530.2283；found 530.2268.
实施例4：
Figure BDA0002465807210000082
a)1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐，1-羟基苯并三唑，N,N-二异丙基乙胺，二氯甲烷；b)氢化钠，1,3-二溴丙烷，N,N-二甲基甲酰胺，冰浴。
步骤1：合成(E)-4-(3-(3,5-二氟苯基)丙烯酰)哌嗪-2-酮(式②所示)
将3,5-二氟苯乙烯酸(式①所示，200mg,1.04mmol)溶于5mL的二氯甲烷中，加入1-羟基苯并三唑(167mg,1.24mmol)，N,N-二异丙基乙胺(255μL,1.56mmol)，1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐(238mg,1.24mmol)，2-哌嗪酮(109mg,1.09mmol)，室温搅拌过夜，停止反应，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离。得到白色固体(150mg,52％)。1H NMR(400MHz,CDCl3)δ7.57(d,J＝15.0Hz,1H),7.11–6.92(m,2H),6.89–6.71(m,2H),4.30(s,2H),3.97–3.74(m,2H),3.56–3.29(m,2H).
步骤2：合成(E)-1-(3-溴丙基)-4-(3-(3,5-二氟苯基)丙烯酰)哌嗪-2-酮(式③所示)
取(E)-4-(3-(3,5-二氟苯基)丙烯酰)哌嗪-2-酮(50mg,0.19mmol)溶于干燥的N,N-二甲基甲酰胺中，冰浴条件下加入氢化钠(15mg,0.38mmol)，维持此温度反应0.5h后，加入1,3-二溴丙烷(100μL,0.94mmol)，保持温度反应1h后，升到室温反应0.5h后，停止反应，加入冰水淬灭，乙酸乙酯萃取3次，合并有机相，用饱和食盐水洗涤2次，无水硫酸钠干燥，硅胶柱分离，得到无色液体(50mg,72％)。1H NMR(300MHz,CDCl3)δ7.61(d,J＝15.5Hz,1H),7.09–6.98(m,2H),6.90–6.73(m,2H),4.38–4.27(m,2H),3.93(s,2H),3.64–3.35(m,6H),2.27–2.09(m,2H).
步骤3：合成(E)-1-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-4-(3-(3,5-二氟苯基)丙烯酰)哌嗪-2-酮(S4)
按照实施例1步骤7的方法合成化合物S4，不同之处在于，将实施例1步骤7中式iii所示化合物替换为式③所示化合物。1H NMR(400MHz,CDCl3)δ7.61(d,J＝15.4Hz,1H),7.25–7.19(m,4H),7.05–6.99(m,2H),6.85–6.76(m,2H),4.32(s,2H),4.15(s,2H),3.99–3.86(m,2H),3.55–3.41(m,4H),3.00–2.92(m,2H),2.47–2.40(m,2H),2.31–2.17(m,4H),1.96–1.89(m,2H),1.87–1.78(m,2H).13C NMR(126MHz,CDCl3)δ164.33,164.09,163.23(dd,J＝249.4,12.8Hz,2×C),148.00,141.72,140.19,138.03(t,J＝8.2Hz),127.38,127.01,125.14,123.10,118.65,110.58(dd,J＝20.0,5.8Hz,2×C),105.22(t,J＝25.1Hz),63.54,55.80,52.10,49.04,46.01,45.76,40.93,39.38,35.22,24.69.HRMS(ESI):calcd forC28H32F2N3O2S[M+H]+:512.2178；found 512.2170.
实施例5：合成(E)-4-(3-(3,5-二氟苯基)丙烯酰)-1-(3-(螺[异硫代色烷-1,4'-哌啶]-1'-基)丙基)哌嗪-2-酮(S5)
按照实施例1步骤3-6的方法，将步骤3中原料式I所示的化合物替换为
Figure BDA0002465807210000091
得到
Figure BDA0002465807210000092
然后按照实施例4步骤3的方法，将其与式③所示化合物，得到化合物S5。1H NMR(400MHz,CDCl3)δ7.60(d,J＝15.4Hz,1H),7.44(d,J＝7.8Hz,1H),7.24–7.18(m,1H),7.16–7.07(m,2H),7.05–6.98(m,2H),6.87–6.74(m,2H),4.31(s,2H),3.98–3.84(m,2H),3.55–3.39(m,4H),3.10(t,J＝6.1Hz,2H),2.81–2.70(m,4H),2.64–2.52(m,2H),2.49–2.39(m,2H),2.31(td,J＝13.6,4.0Hz,2H),1.94–1.87(m,2H),1.87–1.76(m,2H).13C NMR(126MHz,CDCl3)δ164.31,164.09,163.23(dd,J＝249.4,12.8Hz,2×C),142.65,141.66,138.07(t,J＝9.9Hz),136.40,130.29,126.47,126.19,125.59,118.71,110.57(dd,J＝19.8,5.9Hz,2×C),105.18(t,J＝25.4Hz),55.75,49.87,49.05,46.57,46.02,45.78,39.39,39.18,32.42,24.67,23.39.HRMS(ESI):calcd for C29H34F2N3O2S[M+H]+:526.2334；found 526.2347.
实施例6：合成(E)-4-(3-(3,5-二氟苯基)丙烯酰)-1-(3-(5-甲氧基-3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)哌嗪-2-酮(S6)
按照实施例1步骤3-6的方法，将步骤3中原料式I所示的化合物替换为
Figure BDA0002465807210000101
得到
Figure BDA0002465807210000102
然后按照实施例4步骤3的方法，将其与式③所示化合物，得到化合物S6。1H NMR(400MHz,CDCl3)δ7.60(d,J＝15.3Hz,1H),7.09(d,J＝8.5Hz,1H),7.06–6.98(m,2H),6.85–6.76(m,3H),6.75–6.70(m,1H),4.31(s,2H),4.11(s,2H),3.99–3.84(m,2H),3.78(s,3H),3.55–3.41(m,4H),2.99–2.89(m,2H),2.47–2.37(m,2H),2.30–2.09(m,4H),1.92–1.75(m,4H).13C NMR(126MHz,CDCl3)δ164.33,164.10,163.23(dd,J＝249.4,12.7Hz,2×C),159.24,141.68,141.57,140.34,138.06(t,J＝9.6Hz),123.47,118.68,113.26,110.57(dd,J＝19.9,6.0Hz,2×C),109.94,105.19(t,J＝25.5Hz),62.82,55.79,55.47,52.23,49.04,46.01,45.76,41.13,39.38,35.20,24.68.HRMS(ESI):calcdfor C29H34F2N3O3S[M+H]+:542.2283；found 542.2290.
实施例7：合成(E)-4-(3-(3,5-二氟苯基)丙烯酰)-1-(3-(5-氟-3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)哌嗪-2-酮(S7)
按照实施例1步骤3-6的方法，将步骤3中原料式I所示的化合物替换为
Figure BDA0002465807210000103
得到
Figure BDA0002465807210000104
然后按照实施例4步骤3的方法，将其与式③所示化合物，得到化合物S7。1H NMR(400MHz,CDCl3)δ7.60(d,J＝15.3Hz,1H),7.16–7.10(m,1H),7.06–6.98(m,2H),6.97–6.87(m,2H),6.84–6.75(m,2H),4.31(s,2H),4.12(s,2H),3.99–3.81(m,2H),3.55–3.40(m,4H),2.99–2.90(m,2H),2.47–2.36(m,2H),2.28–2.19(m,2H),2.18–2.09(m,2H),1.94–1.88(m,2H),1.85–1.76(m,2H).13C NMR(126MHz,CDCl3)δ164.10,163.24(dd,J＝249.4,12.8Hz,2×C),162.19(d,J＝245.7Hz),143.79(d,J＝2.2Hz),142.23(d,J＝7.8Hz),141.71,138.05(t,J＝9.2Hz),123.89(d,J＝9.0Hz),118.65,114.15(d,J＝22.5Hz),111.86(d,J＝22.2Hz),110.57(dd,J＝19.8,6.0Hz,2×C),105.21(t,J＝25.4Hz),62.63,55.75,52.08,49.03,46.01,45.74,41.03,39.41,34.84,24.67.HRMS(ESI):calcd for C28H31F3N3O2S[M+H]+:530.2084；found 530.2074.
实施例8：合成(E)-4-(3-(3,5-二氟苯基)丙烯酰)-1-(3-(4-氟-3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)哌嗪-2-酮(S8)
按照实施例1步骤3-6的方法，将步骤3中原料式I所示的化合物替换为
Figure BDA0002465807210000111
得到
Figure BDA0002465807210000112
然后按照实施例4步骤3的方法，将其与式③所示化合物，得到化合物S8。1H NMR(400MHz,CDCl3)δ7.59(d,J＝15.3Hz,1H),7.25–7.17(m,1H),7.05–6.94(m,3H),6.90(t,J＝8.6Hz,1H),6.84–6.75(m,2H),4.37–4.27(m,2H),4.11(s,2H),3.93(s,2H),3.53–3.40(m,4H),2.99–2.90(m,2H),2.42(t,J＝7.3Hz,2H),2.28–2.10(m,4H),1.94–1.87(m,2H),1.85–1.75(m,2H).13C NMR(126MHz,CDCl3)δ164.30,163.21(dd,J＝249.4,12.8Hz,2×C),164.12,158.87(d,J＝246.7Hz),151.76(d,J＝3.8Hz),141.66,138.06(t,J＝9.1Hz),129.12(d,J＝7.3Hz),127.15(d,J＝18.5Hz),118.69,118.58(d,J＝3.1Hz),113.74(d,J＝20.1Hz),110.57(dd,J＝19.9,5.8Hz,2×C),105.18(t,J＝25.4Hz),64.06,58.39,55.74,52.01,49.02,46.64,46.00,45.72,40.90,39.38,30.94,24.65.HRMS(ESI):calcd for C28H31F3N3O2S[M+H]+:530.2084；found 530.2071.
实施例9：合成(E)-4-(3-(3,5-二氟苯基)丙烯酰)-1-(3-(7-氟-3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)哌嗪-2-酮(S9)
按照实施例1步骤3-6的方法，将步骤3中原料式I所示的化合物替换为
Figure BDA0002465807210000113
得到
Figure BDA0002465807210000114
然后按照实施例4步骤3的方法，将其与式③所示化合物，得到化合物S9。1H NMR(400MHz,CDCl3)δ7.60(d,J＝15.3Hz,1H),7.22–7.15(m,1H),7.07–6.96(m,3H),6.92–6.75(m,3H),4.31(s,2H),4.18(s,2H),3.98–3.83(m,2H),3.56–3.41(m,4H),3.02–2.88(m,2H),2.60(td,J＝12.9,3.7Hz,2H),2.42(t,J＝7.2Hz,2H),2.23(t,J＝12.1Hz,2H),1.92–1.74(m,4H).13C NMR(126MHz,CDCl3)δ164.32,164.11,163.23(dd,J＝249.3,12.8Hz,2×C),159.19(d,J＝249.1Hz),143.17(d,J＝5.7Hz),141.60,138.10(t,J＝8.9Hz),133.81(d,J＝12.8Hz),129.04(d,J＝8.4Hz),120.94(d,J＝2.8Hz),118.75,114.44(d,J＝22.0Hz),110.57(dd,J＝19.9,6.0Hz,2×C),105.16(t,J＝25.4Hz),63.95(d,J＝4.1Hz),55.74,51.95,49.04,46.75,46.09,45.85,39.41,38.48,35.66,29.69,24.62.HRMS(ESI):calcd for C28H31F3N3O2S[M+H]+:530.2084；found 530.2087.
实施例10：
Figure BDA0002465807210000121
试剂与条件：a)碳酸钾，N,N-二甲基甲酰胺；b)盐酸甲醇；c)乌洛托品，三氟乙酸，浓硫酸d)四丁基碘化铵，碳酸钾，氯乙酸甲酯，130℃。
步骤1：合成叔-丁基4-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-3-羰基哌嗪-1-羧酸酯(式(2)所示)
取叔-丁基4-(3-溴丙基)-3-羰基哌嗪-1-羧酸酯(式(ii)所示，1.68g,5.23mmol)和式V所示的2H-螺[苯并[b]噻吩-3,4'-哌啶](1g,4.76mmol)溶于10mL的N,N-二甲基甲酰胺中，加入碳酸钾(0.98g,7.14mmol)，70℃反应5h后，停止反应，倒入冰水中，乙酸乙酯萃取3次，合并有机相，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到淡黄色液体(1.8g,85％)。1H NMR(400MHz,CDCl3)δ7.26–7.20(m,4H),4.15(s,2H),4.06(s,2H),3.67–3.61(m,2H),3.49–3.43(m,2H),3.39–3.33(m,2H),2.99–2.93(m,2H),2.46–2.38(m,2H),2.29–2.16(m,4H),1.94–1.87(m,2H),1.84–1.76(m,2H),1.46(s,9H).
步骤2：合成1-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)哌嗪-2-酮(式(3)所示)
取本实施例步骤1所得产物(1.8g,4mmol)溶于15mL的甲醇中，加入盐酸甲醇溶液15mL，室温搅拌1h后，旋干溶剂，加入饱和碳酸氢钠溶液调节pH，二氯甲烷萃取6次，无水硫酸钠干燥，硅胶柱分离。得到淡黄色液体，放置变成固体(0.8g,58％)1H NMR(400MHz,CDCl3)δ7.28–7.22(m,2H),4.18(d,J＝3.6Hz,1H),3.54(d,J＝3.7Hz,1H),3.50–3.43(m,1H),3.38–3.33(m,1H),3.14–3.08(m,1H),3.04–2.98(m,1H),2.51–2.43(m,1H),2.34–2.20(m,2H),1.97–1.91(m,1H),1.89–1.80(m,1H).
步骤3：合成3,5-二氟-2-羟基苯甲醛(式(5)所示)
2,4-二氟苯酚(1g，7.7mmol)溶于6mL的TFA中，加入乌洛托品(2.16g,15.4mmol)，随后加热至80℃，回流反应20h后，冷却至室温加入12mL水，3mL的浓硫酸，搅拌反应2h后，乙酸乙酯萃取2次，合并有机相，3N盐酸洗涤4次，饱和碳酸氢钠溶液洗涤1次，硅胶柱分离，得到淡粉红色固体(0.6g,49％)。1H NMR(300MHz,CDCl3)δ10.72(s,1H),9.88(d,J＝1.3Hz,1H),7.21–7.08(m,2H).
步骤4：合成5,7-二氟苯并呋喃-2-羧酸(式(6)所示)
取3,5-二氟-2-羟基苯甲醛(式(5)所示，540mg,3.42mmol)和四丁基碘化铵(126mg,0.34mmol)、碳酸钾(1.9g,13.68mmol)、氯乙酸甲酯(650μL，7.18mmol)混合后，加热至130℃。反应4h后，冷却至0℃，加入5mL的THF、5mL的水、KOH(1.14g,20.52mmol)，室温搅拌过夜后，旋掉有机溶剂，加入水稀释，浓盐酸调节至酸性，乙酸乙酯萃取，无水硫酸钠干燥，用甲苯：乙酸乙酯：醋酸(35:5:1)体系过柱，得到棕色固体(175mg,26％)。1H NMR(400MHz,CD3OD)δ7.55(d,J＝2.7Hz,1H),7.26–7.21(m,1H),7.13–7.08(m,1H).
步骤5：合成1-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-4-(5,7-二氟苯并呋喃-2-羰基)哌嗪-2-酮(S10)
取本实施例步骤2所得产物(80mg,0.23mmol)和5,7-二氟苯并呋喃-2-羧酸(式(6)所示，46mg,0.23mmol)溶于10mL的二氯甲烷中，0℃加入HATU(36mg,0.27mmol)，继续搅拌2h后，停止反应，恢复室温搅拌0.5h后，停止反应，饱和氯化铵溶液洗涤，无水硫酸钠干燥，得到白色固体(40mg,50％)1H NMR(400MHz,CDCl3)δ7.44(s,1H),7.25–7.19(m,4H),7.17–7.12(m,1H),6.97(td,J＝10.2,2.3Hz,1H),4.67–4.36(m,2H),4.23–3.95(m,4H),3.62–3.44(m,4H),3.03–2.93(m,2H),2.45(t,J＝7.4Hz,2H),2.33–2.14(m,4H),1.96–1.78(m,4H).HRMS(ESI):calcd for C28H30F2N3O3S[M+H]+:526.1970；found 526.1956.
实施例11：合成1-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-4-(5-(3,5-二氟苯基)噁唑-2-羰基)哌嗪-2-酮(S11)
Figure BDA0002465807210000131
试剂与条件：a)碳酸钾，对甲苯磺酰甲基异腈，甲醇；b)正丁基锂，六氯乙烷，四氢呋喃。
步骤1：合成5-(3,5-二氟苯基)恶唑
3,5-二氟苯甲醛(2g,14mmol)和对甲苯磺酰甲基异腈(2.75g,14mmol)溶于30mL甲醇中，加入碳酸钾(1.9g,14mmol)，回流反应1h后，冷却到室温，蒸干溶剂，重新溶于乙酸乙酯中，用饱和碳酸氢钠溶液洗涤，无水硫酸钠干燥，硅胶柱分离，得到白色固体(2.4g,95％)。1H NMR(400MHz,CDCl3)δ7.94(s,1H),7.40(s,1H),7.20–7.13(m,2H),6.78(tt,J＝8.8,2.2Hz,1H).
步骤2：合成2-氯-5-(3,5-二氟苯基)恶唑
将5-(3,5-二氟苯基)恶唑(0.5g,2.76mmol)溶于10mL的THF中，-70℃下慢慢加入正丁基锂(3.3mmol)，维持温度搅拌30min后，加入六氯乙烷，随后慢慢恢复到室温，搅拌5h后，冷却到0℃，加入饱和氯化铵溶液淬灭，乙酸乙酯萃取3次，合并有机相，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到白色固体(220mg,37％)。1H NMR(400MHz,CDCl3)δ7.33(s,1H),7.15–7.08(m,2H),6.81(tt,J＝8.7,2.2Hz,1H).
步骤3：合成1-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-4-(5-(3,5-二氟苯基)噁唑-2-羰基)哌嗪-2-酮(S11)
取实施例10合成的式(3)所示化合物(80mg,0.23mmol)和2-氯-5-(3,5-二氟苯基)恶唑(58mg,0.27mmol)溶于1mL的N,N-二甲基甲酰胺中，加入N,N-二异丙基乙胺(70mg,0.54mmol)，微波160℃反应30min后，停止反应，倒入冰水中，乙酸乙酯萃取3次，饱和食盐水洗涤2次，无水硫酸钠干燥，得到白色固体(80mg,57％)。1H NMR(400MHz,CDCl3)δ7.45–7.30(m,5H),7.10(d,J＝6.4Hz,2H),6.79(t,J＝8.7Hz,1H),4.38(s,2H),4.29(s,2H),4.04–3.96(m,2H),3.73–3.62(m,4H),3.15–3.07(m,2H),2.58(t,J＝7.3Hz,2H),2.44–2.29(m,4H),2.11–1.93(m,4H).13C NMR(126MHz,CDCl3)δ164.49,163.44(dd,J＝248.5,13.2Hz,2×C),,159.80,148.03,144.25(t,J＝3.4Hz),140.20,131.08(t,J＝10.8Hz),127.36,127.00,125.13,124.46,123.08,105.42(dd,J＝21.0,6.8Hz,2×C),102.16(t,J＝25.5Hz),63.55,55.81,52.10,48.98,45.99,45.68,42.85,40.95,35.21,24.68.HRMS(ESI):calcd for C28H31F2N4O2S[M+H]+:525.2130；found 525.2120.
实施例12：合成N-(2-(4-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-3-羰基哌嗪-1-基)-2-羰基乙基)-3-(三氟甲基)苯酰胺(S12)
按照实施例10步骤5的方法合成S12，区别在于，将5,7-二氟苯并呋喃-2-羧酸替换为间三氟甲基马尿酸。1H NMR(400MHz,CDCl3)δ8.19–7.94(m,2H),7.83–7.72(m,1H),7.64–7.34(m,2H),7.27–7.15(m,3H),4.38–4.06(m,6H),3.84(d,J＝56.6Hz,2H),3.62–3.39(m,4H),2.98(d,J＝7.0Hz,2H),2.53–2.38(m,2H),2.36–2.12(m,4H),2.03–1.74(m,4H).
实施例13：合成1-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-4-(3,5-二氟苯甲酰)哌嗪-2-酮(S13)
按照实施例10步骤5的方法合成S13，区别在于，将5,7-二氟苯并呋喃-2-羧酸替换为3,5-二氟苯甲酸。1H NMR(400MHz,CDCl3)δ7.25–7.18(m,4H),7.01–6.88(m,3H),4.45–3.77(m,6H),3.54–3.38(m,4H),3.00–2.92(m,2H),2.43(t,J＝7.3Hz,2H),2.31–2.15(m,4H),1.96–1.88(m,2H),1.87–1.77(m,2H).13C NMR(126MHz,CDCl3)δ167.35,162.97(dd,J＝252.0,12.1Hz,2×C),147.98,140.19,127.39,127.02,125.15,123.09,110.64(m,2×C),106.07(t,J＝22.6Hz),63.52,55.74,52.08,45.59,40.92,35.23,24.67.HRMS(ESI):calcdfor C26H30F2N3O2S[M+H]+:486.2021；found 486.2020.
实施例14：合成1-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-4-(3,5-二氟苯基)哌嗪-2-酮(S14)
取1-溴-3,5-二氟苯(48mg,0.25mmol)、实施例10合成的式(3)所示化合物(80mg,0.23mmol)、叔丁醇钠(33mg,0.34mmol)、1,1'-联萘-2,2'-双二苯膦(44mg)和8mg的醋酸钯溶于10mL的甲苯中，氮气保护下85℃反应4h后，停止反应，加入水，乙酸乙酯萃取3次，合并有机相，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到白色固体。1H NMR(400MHz,CDCl3)δ7.34–7.12(m,4H),6.41–6.20(m,3H),4.16(s,2H),3.88(s,2H),3.68–3.39(m,6H),3.07–2.89(m,2H),2.53–2.38(m,2H),2.35–2.11(m,4H),2.04–1.72(m,4H).13C NMR(126MHz,CDCl3)δ165.42,164.12(dd,J＝245.0,15.8Hz,2×C),150.61(t,J＝12.5Hz),148.03,140.19,127.36,127.00,125.14,123.09,96.96(dd,J＝21.9,7.7Hz,2×C),94.39(t,J＝26.1Hz),63.55,55.87,52.12,51.66,46.14,45.34,44.71,40.96,35.22,24.68.HRMS(ESI):calcd for C25H30F2N3OS[M+H]+:458.2072；found 458.2061.
实施例15：合成4-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-N-(3,5-二氟苯基)-3-羰基哌嗪-1-甲酰胺(S15)
取实施例10合成的式(3)所示化合物(80mg，0.23mmol)和3,5-二氟苯异肼酸酯(39mg,0.25mmol)溶于二氯甲烷中，室温搅拌2h后，停止反应，直接用制备板分离，得到白色固体(70mg,61％)。1H NMR(400MHz,DMSO-d6)δ8.94(s,1H),7.36–7.17(m,6H),6.77(t,J＝9.4Hz,1H),4.20–4.03(m,4H),3.76–3.65(m,2H),3.48–3.37(m,4H),2.99–2.88(m,2H),2.33(t,J＝6.9Hz,2H),2.20–2.03(m,4H),1.83–1.61(m,4H).13C NMR(126MHz,DMSO-d6)δ164.93,162.80(dd,J＝241.6,15.7Hz,2×C),154.06,148.38,143.56(t,J＝14.0Hz),140.54,127.68,127.33,125.53,123.49,102.59–101.97(dd,J＝22.4,7.1Hz,2×C),97.23(t,J＝26.2Hz),63.75,55.90,52.05,48.14,46.16,45.13,41.08,40.76,34.74,24.45.HRMS(ESI):calcd for C26H31F2N4O2S[M+H]+:501.2130；found 501.2132.
实施例16：合成(E)-1-(3-(3H-螺[苯并[c]噻吩-1,4'-哌啶]-1'-基)丙基)-4-(3-(3,4,5-三氟苯基)丙烯酰)哌嗪-2-酮(S16)
按照实施例10步骤3的方法合成S13，区别在于，将5,7-二氟苯并呋喃-2-羧酸替换为3,4,5-三氟苯乙烯酸。1H NMR(400MHz,CDCl3)δ7.55(d,J＝15.2Hz,1H),7.25–7.07(m,6H),6.73(d,J＝15.0Hz,1H),4.31(s,2H),4.15(s,2H),3.99–3.82(m,2H),3.56–3.40(m,4H),3.01–2.89(m,2H),2.43(t,J＝7.1Hz,2H),2.31–2.13(m,4H),1.94–1.88(m,2H),1.87–1.77(m,2H).HRMS(ESI):calcd for C28H31F3N3O2S[M+H]+:530.2084；found 530.2070.
实施例17：
Figure BDA0002465807210000151
试剂与条件：a)正丁基锂，N-叔丁氧羰基-4-哌啶酮，四氢呋喃，乙醚，-70℃；b)甲基磺酰氯，三乙胺，二氯甲烷；c)盐酸甲醇。
步骤1：合成叔-丁基4-羟基-4-(2-(2-羟基乙基)苯基)哌啶-1-羧酸酯(式2所示)
将2-溴苯乙醇(式1所示，0.67mL,5mmol)溶于5mL乙醚和5mL四氢呋喃中，冷却到-70℃，将2.5M正丁基锂正己烷溶液(4mL,10mmol)慢慢加入其中，反应3h后，将1-Boc-4-哌啶酮(1.11g,5.6mmol)溶于5mL的四氢呋喃中，慢慢加入其中。并将反应体系缓慢升到室温，室温反应18h后，加入氯化铵溶液淬灭，乙酸乙酯萃取3次，无水硫酸钠干燥。旋干，硅胶柱纯化。得到无色液体(740mg,46％)。1H NMR(300MHz,CDCl3)δ7.31–7.13(m,4H),4.02–3.85(m,4H),3.28(s,4H),2.02–1.71(m,5H),1.44(s,9H).
步骤2：合成叔-丁基螺[异色烷-1,4'-哌啶]-1'-羧酸酯(式3所示)
将甲基磺酰氯(209μL,2.7mmol)溶于2mL二氯甲烷中，冰浴条件下慢慢加入到含有叔-丁基4-羟基-4-(2-(2-羟基乙基)苯基)哌啶-1-羧酸酯(740mg,2.44mmol)，三乙胺(707μL,5.1mmol)的二氯甲烷溶液中，恢复到室温，室温搅拌过夜后，加入冰水淬灭，依次用饱和氯化铵溶液，饱和碳酸氢钠溶液，饱和食盐水溶液洗涤。无水硫酸钠干燥，得到无色液体(500mg,67％)。1H NMR(300MHz,CDCl3)δ7.23–7.05(m,4H),4.08–3.87(m,4H),3.27–3.01(m,2H),2.84(t,J＝5.4Hz,2H),1.95–1.78(m,4H),1.50(s,9H).
步骤3：合成螺[异色烷-1,4'-哌啶](式4所示)
操作步骤同实施例10步骤2，区别在于，将式(2)所示化合物替换为本实施例合成的式3所示化合物。1H NMR(300MHz,CDCl3)δ7.22–7.05(m,4H),3.91(t,J＝5.5Hz,2H),3.16–3.02(m,2H),2.98–2.87(m,2H),2.83(t,J＝5.5Hz,2H),2.42(s,1H),1.95–1.81(m,4H).
步骤4：合成N-(2-羰基-2-(4-(3-(螺[异色烷-1,4'-哌啶]-1'-基)丙基)哌嗪-1-基)乙基)-3-(三氟甲基)苯酰胺(S17)
以步骤3所得式4所示化合物为原料，将其与间三氟甲基马尿酸反应，操作步骤参考实施例10步骤5。1H NMR(400MHz,CDCl3)δ8.12(s,1H),8.01(d,J＝7.9Hz,1H),7.77(d,J＝7.8Hz,1H),7.58(t,J＝7.8Hz,1H),7.41(t,J＝3.5Hz,1H),7.21–7.18(m,2H),7.17–7.12(m,1H),7.08(d,J＝7.0Hz,1H),4.26(d,J＝3.9Hz,2H),3.90(t,J＝5.5Hz,2H),3.73–3.67(m,2H),3.51–3.46(m,2H),2.83(t,J＝5.4Hz,2H),2.78(d,J＝11.7Hz,2H),2.53–2.35(m,10H),2.10–1.99(m,2H),1.95–1.87(m,2H),1.81–1.75(m,2H).
实施例18：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(螺[异色烷-1,4'-哌啶]-1'-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S18)
将实施例17步骤3所得原料与3，5-二氟苯乙烯酸反应得到S18，操作步骤参考实施例10。1H NMR(400MHz,CDCl3)δ7.55(d,J＝15.4Hz,1H),7.21–7.12(m,3H),7.10–7.06(m,1H),7.04–6.99(m,2H),6.87(d,J＝15.4Hz,1H),6.79(tt,J＝8.7,2.3Hz,1H),3.90(t,J＝5.5Hz,2H),3.79–3.61(m,4H),2.83(t,J＝5.4Hz,2H),2.80–2.73(m,2H),2.53–2.35(m,10H),2.10–1.98(m,2H),1.94–1.87(m,2H),1.81–1.74(m,2H).13C NMR(126MHz,CDCl3)δ164.20,162.87(dd,J＝248.9,12.9Hz,2×C),141.66,139.92,138.31(t,J＝9.4Hz),133.45,128.49,125.90,125.80,125.08,119.56,110.04(dd,J＝19.7,6.0Hz,2×C),104.37(t,J＝25.5Hz),72.82,58.41,56.49,56.29,53.19,52.50,49.06,45.59,41.94,36.54,29.36,24.14.HRMS(ESI):calcd for C29H36F2N3O2[M+H]+:496.2770；found493.2757.
实施例19：合成(E)-1-(4-(3-(4-(1H-吲哚-3-基)哌啶-1-基)丙基)哌嗪-1-基)-3-(3,5-二氟苯基)丙-2-烯-1-酮(S19)
将实施例1中步骤7的2H-螺[苯并[b]噻吩-3,4'-哌啶]换成3-(哌啶-4-基)-1H-吲哚，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ7.63(d,J＝7.8Hz,1H),7.54(d,J＝15.4Hz,1H),7.35–7.29(m,1H),7.16(t,J＝7.5Hz,1H),7.08(t,J＝7.4Hz,1H),7.04–6.96(m,2H),6.95–6.90(m,1H),6.87(d,J＝15.4Hz,1H),6.79(tt,J＝8.7,2.1Hz,1H),3.81–3.54(m,4H),3.07(d,J＝11.4Hz,2H),2.93–2.78(m,1H),2.52–2.35(m,8H),2.19–2.01(m,4H),1.89–1.71(m,4H).13C NMR(126MHz,CDCl3)δ164.60,163.20(dd,J＝248.9,12.9Hz,2×C),140.29,138.60(t,J＝9.5Hz),136.47,126.68,121.79,121.19,119.88,119.79,119.04,118.95,111.30,110.39(dd,J＝19.7,6.0Hz,2×C),104.73(t,J＝25.5Hz),57.01,56.58,54.52,53.47,52.81,45.90,42.28,33.54,33.01,24.44.HRMS(ESI):calcd for C29H35F2N4O[M+H]+:493.2773；found 493.2775.
实施例20：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(3,4-二氢-1H-吡啶并[3,4-b]吲哚-2(9H)-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S20)
将实施例1中步骤7中的2H-螺[苯并[b]噻吩-3,4'-哌啶]换成2,3,4,5-四氢-1H-吡啶并[4,3-b]吲哚，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ8.46(br,J＝7.7Hz,1H),7.54(d,J＝15.4Hz,1H),7.41–7.35(m,1H),7.24–7.19(m,1H),7.12–6.96(m,4H),6.86(d,J＝15.4Hz,1H),6.78(tt,J＝8.7,2.2Hz,1H),3.80–3.57(m,6H),2.91–2.71(m,4H),2.71–2.60(m,2H),2.52–2.36(m,6H),1.90–1.79(m,2H).13C NMR(126MHz,CDCl3)δ164.13,162.77(dd,J＝248.9,12.8Hz,2×C),139.87,138.16(t,J＝9.5Hz),135.72,131.68,125.69,120.70,119.42,118.80,117.00,110.28,109.95(dd,J＝19.6,6.0Hz,2×C),107.85,104.31(t,J＝25.5Hz),56.01,55.49,53.11,52.41,50.36,49.21,45.48,41.85,24.44,23.27.HRMS(ESI):calcd for C27H33F2N4O2[M+H]+:465.2460；found465.2454.
实施例21：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(4-甲基-4-苯基哌啶-1-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S21)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成4-甲基-4-苯基哌啶，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ7.53(d,J＝15.4Hz,1H),7.36–7.27(m,4H),7.19–7.14(m,1H),7.04–6.96(m,2H),6.85(d,J＝15.4Hz,1H),6.80–6.73(m,1H),3.77–3.55(m,4H),2.56–2.27(m,13H),2.20–2.09(m,2H),1.82–1.63(m,4H),1.21(s,3H).13C NMR(126MHz,CDCl3)δ164.09,162.76(dd,J＝248.9,12.9Hz,2×C),139.84,138.17(t,J＝9.5Hz),127.90,125.37,125.16,119.40,109.94(dd,J＝19.7,6.0Hz,2×C),104.28(t,J＝25.5Hz),56.32,56.14,53.06,52.36,49.92,45.45,41.80,36.57,35.76,23.97.HRMS(ESI):calcd for C28H36F2N3O[M+H]+:468.2821；found 468.2826.
实施例22：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(哌啶-1-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S22)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成哌啶，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ7.48(d,J＝15.4Hz,1H),7.00–6.92(m,2H),6.82(d,J＝15.4Hz,1H),6.73(tt,J＝8.7,2.3Hz,1H),3.72–3.53(m,4H),2.47–2.20(m,12H),1.71–1.59(m,2H),1.56–1.47(m,4H),1.41–1.30(m,2H).13C NMR(126MHz,CDCl3)δ164.47,163.18(dd,J＝248.9,12.8Hz,2×C),140.19,138.62(t,J＝9.5Hz),119.87,110.33(dd,J＝19.7,6.0Hz,2×C),104.66(t,J＝25.5Hz),57.31,56.61,54.65,53.47,52.80,45.89,42.24,25.94,24.42,24.33.HRMS(ESI):calcd for C21H30F2N3O[M+H]+:378.2351；found378.2343.
实施例23：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(4-苯基哌啶-1-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S23)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成4-苯基哌啶，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ7.55(d,J＝15.4Hz,1H),7.33–7.27(m,2H),7.25–7.17(m,3H),7.05–6.98(m,2H),6.87(d,J＝15.4Hz,1H),6.79(tt,J＝8.7,2.3Hz,1H),3.80–3.60(m,4H),3.14–3.05(m,2H),2.55–2.39(m,9H),2.16–2.02(m,2H),1.91–1.73(m,8H).13C NMR(126MHz,CDCl3)δ164.52,163.16(dd,J＝248.9,12.9Hz,2×C),145.95,140.21,138.61(t,J＝9.5Hz),128.43,126.82,126.22,119.89,110.36(dd,J＝19.7,6.0Hz,2×C),104.67(t,J＝25.5Hz),56.74,56.37,54.32,53.43,52.80,45.86,42.46,42.22,33.07,24.12.HRMS(ESI):calcd for C27H34F2N3O[M+H]+:454.2664；found454.2667.
实施例24：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(4-羟基-4-苯基哌啶-1-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S24)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成4-苯基哌啶-4-醇，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ7.50–7.43(m,3H),7.33–7.26(m,2H),7.23–7.17(m,1H),7.00–6.92(m,2H),6.82(d,J＝15.4Hz,1H),6.75(tt,J＝8.7,2.2Hz,1H),3.70–3.54(m,4H),2.83–2.71(m,2H),2.49–2.31(m,10H),2.18–2.05(m,2H),1.77–1.64(m,4H).13C NMR(126MHz,CDCl3)δ164.52,163.17(dd,J＝248.9,12.8Hz,2×C),148.50,140.30,138.57(t,J＝9.5Hz),128.29,126.93,124.58,119.78,110.36(dd,J＝19.7,6.0Hz,2×C),104.70(t,J＝25.5Hz),71.06,56.67,56.52,53.47,52.76,49.61,45.85,42.22,38.36,24.37.HRMS(ESI):calcd for C27H34F2N3O2[M+H]+:470.2614；found470.2601.
实施例25：合成(E)-1-(4-(3-(4-(1H-吲哚-4-基)哌啶-1-基)丙基)哌嗪-1-基)-3-(3,5-二氟苯基)丙-2-烯-1-酮(S25)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成4-(哌啶-4-基)-1H-吲哚，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ7.63(d,J＝7.8Hz,1H),7.54(d,J＝15.4Hz,1H),7.35–7.29(m,1H),7.16(t,J＝7.5Hz,1H),7.08(t,J＝7.4Hz,1H),7.04–6.96(m,2H),6.95–6.90(m,1H),6.87(d,J＝15.4Hz,1H),6.79(tt,J＝8.7,2.1Hz,1H),3.81–3.54(m,4H),3.07(d,J＝11.4Hz,2H),2.93–2.78(m,1H),2.52–2.35(m,8H),2.19–2.01(m,4H),1.89–1.71(m,4H).13C NMR(126MHz,CDCl3)δ164.60,163.20(dd,J＝248.9,12.9Hz,2×C),140.29,138.60(t,J＝9.5Hz),136.47,126.68,121.79,121.19,119.88,119.79,119.04,118.95,111.30,110.39(dd,J＝19.7,6.0Hz,2×C),104.73(t,J＝25.5Hz),57.01,56.58,54.52,53.47,52.81,45.90,42.28,33.54,33.01,24.44.HRMS(ESI):calcd for C29H35F2N4O[M+H]+:493.2773；found 493.2775.
实施例26：合成(4,6-二氟-1H-吲哚-2-基)(4-(3-(4-苯基哌啶-1-基)丙基)哌嗪-1-基)甲酮(S26)
按照实施例1步骤2的方法合成
Figure BDA0002465807210000191
区别在于将3,5-二氟苯乙烯酸替换为4，6-二氟吲哚-2-羧酸。接着将其作为原料与4-苯基哌啶反应得到S26，1H NMR(400MHz,CDCl3)δ7.33–7.27(m,2H),7.25–7.17(m,3H),6.91(dd,J＝8.8,1.7Hz,1H),6.81(d,J＝0.7Hz,1H),6.63(td,J＝10.0,2.0Hz,1H),4.01–3.87(m,3H),3.07(d,J＝11.5Hz,2H),2.59–2.53(m,4H),2.48–2.40(m,4H),2.06(td,J＝11.4,3.3Hz,2H),1.89–1.70(m,10H).HRMS(ESI):calcd for C27H33F2N4O[M+H]+:467.2617；found 467.2611.
实施例27：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(3,4-二氢异喹啉-2(1H)-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S27)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成1,2,3,4-四氢异喹啉，其它操作步骤同实施例1步骤7。1H NMR(300MHz,CDCl3)δ7.54(d,J＝15.4Hz,1H),7.16–7.05(m,4H),7.06–6.96(m,3H),6.91–6.74(m,2H),3.81–3.58(m,6H),2.95–2.85(m,2H),2.83–2.69(m,2H),2.61–2.41(m,7H),1.93–1.74(m,3H).HRMS(ESI):calcd for C25H30F2N3O[M+H]+:426.2351；found 426.2354.
实施例28：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(4-(吡咯烷-1-基)哌啶-1-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S28)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成4-(吡咯烷-1-基)哌啶，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ7.54(d,J＝15.4Hz,1H),7.05–6.97(m,2H),6.86(d,J＝15.4Hz,1H),6.79(tt,J＝8.8,2.1Hz,1H),3.79–3.56(m,4H),2.94–2.85(m,2H),2.60–2.43(m,8H),2.42–2.28(m,4H),2.05–1.82(m,6H),1.81–1.73(m,4H),1.73–1.63(m,2H),1.60–1.48(m,2H).13C NMR(126MHz,CDCl3)δ164.04,162.74(dd,J＝248.9,12.9Hz,2×C),139.78,138.17(t,J＝9.4Hz),119.42,109.91(dd,J＝19.7,6.0Hz,2×C),104.24(t,J＝25.5Hz),61.64,56.12,56.04,53.08,52.31,50.94,45.45,41.80,30.90,24.12,22.81.HRMS(ESI):calcd for C25H37F2N4O[M+H]+:447.2930；found447.2934.
实施例29：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(3,4-二氢喹啉-1(2H)-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S29)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成1,2,3,4-四氢喹啉，其它操作步骤同实施例1步骤7。1H NMR(300MHz,CDCl3)δ7.55(d,J＝15.4Hz,1H),7.08–6.98(m,3H),6.97–6.75(m,3H),6.66–6.51(m,2H),3.82–3.60(m,4H),3.39–3.24(m,4H),2.75(t,J＝6.3Hz,2H),2.57–2.38(m,6H),2.01–1.85(m,2H),1.84–1.72(m,2H).
实施例30：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(4-苯基哌嗪-1-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S30)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成1-苯基哌嗪，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ7.55(d,J＝15.4Hz,1H),7.29–7.26(m,1H),7.05–6.98(m,2H),6.93(d,J＝7.9Hz,2H),6.89–6.86(m,1H),6.84–6.77(m,1H),6.37–6.30(m,1H),6.18–6.08(m,1H),5.80–5.75(m,1H),3.80–3.62(m,4H),3.31–3.25(m,4H),2.86–2.77(m,4H),2.66–2.59(m,2H),2.57–2.49(m,4H),2.46(t,J＝7.3Hz,2H),1.90–1.79(m,2H).13C NMR(126MHz,CDCl3)δ169.96,164.55,163.21(dd,J＝249.0,12.8Hz,2×C),151.03,140.38,138.58(t,J＝9.4Hz),131.18,129.16,128.80,120.01,119.75,116.21,110.37(dd,J＝19.7,6.0Hz,2×C),104.74(t,J＝25.5Hz),56.15,53.36,52.90,52.69,48.70,45.73,42.09,29.69,23.45.HRMS(ESI):calcd for C26H33F2N4O[M+H]+:455.2617；found 455.2604.
实施例31：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(4-(吡啶-4-基)哌啶-1-基)丙基)哌嗪-1-基)丙-2-烯-1-酮(S31)
将实施例1中步骤7中2H-螺[苯并[b]噻吩-3,4'-哌啶]换成4-(哌啶-4-基)吡啶，其它操作步骤同实施例1步骤7。1H NMR(400MHz,CDCl3)δ8.50(dd,J＝4.5,1.6Hz,2H),7.55(d,J＝15.4Hz,1H),7.14(dd,J＝4.6,1.5Hz,2H),7.05–6.98(m,2H),6.87(d,J＝15.4Hz,1H),6.79(tt,J＝8.8,2.3Hz,1H),3.79–3.60(m,4H),3.10–3.02(m,2H),2.54–2.45(m,5H),2.42(td,J＝7.7,1.9Hz,4H),2.04(td,J＝11.4,2.2Hz,2H),1.89–1.69(m,6H).13C NMR(126MHz,CDCl3)δ164.49,163.17(dd,J＝248.9,12.8Hz,2×C),154.94,149.82,140.23,138.60(t,J＝9.5Hz),122.30,119.84,110.34(dd,J＝19.7,6.0Hz,2×C),104.68(t,J＝25.5Hz),56.80,56.50,54.03,53.52,52.79,45.88,42.23,41.94,32.55,24.46.HRMS(ESI):calcd for C26H33F2N4O[M+H]+:455.2617；found 455.2609.
实施例32：合成(E)-4-(3-(3,5-二氟苯基)丙烯酰)-1-(3-(4-苯基哌啶-1-基)丙基)哌嗪-2-酮(S32)
参考实施例4合成S32，区别在于，将实施例4步骤3中的式V所示化合物替换为4-苯基哌啶。1H NMR(400MHz,CDCl3)δ7.60(d,J＝15.5Hz,1H),7.32–7.26(m,2H),7.24–7.16(m,3H),7.06–6.98(m,2H),6.86–6.75(m,2H),4.31(s,2H),4.01–3.82(m,2H),3.54–3.41(m,4H),3.08–2.97(m,2H),2.54–2.45(m,1H),2.45–2.35(m,2H),2.11–1.91(m,5H),1.88–1.80(m,4H).HRMS(ESI):calcd for C27H32F2N3O2[M+H]+:468.2457；found 468.2467.
实施例33：
Figure BDA0002465807210000201
试剂与条件：a)无水哌嗪，卡特缩合剂，二氯甲烷；b)溴丙酰氯，N,N-二异丙基乙胺，二氯甲烷；c)[1,1’-双(二苯基膦基)二茂铁]二氯化钯，碳酸钠，1,4-二氧六环，水；d)苄基氯，丙酮，硼氢化钠，甲醇；e)氢氧化钯，甲酸铵，甲醇。
步骤1：合成(E)-3-(3,5-二氟苯基)-1-(哌嗪-1-基)丙-2-烯-1-酮(式B所示)
将肉桂酸(0.92g,5mmol)、无水哌嗪(1.29g,15mmol)，卡特缩合剂(4.42g,10mmol)溶于60mL二氯甲烷，室温搅拌过夜，加入饱和碳酸氢钠溶液洗涤，无水硫酸钠干燥，旋干过柱，得到黄色液体，放置得到固体。1H NMR(400MHz,CDCl3)δ7.51(d,J＝15.4Hz,1H),7.03–6.93(m,2H),6.85(d,J＝15.4Hz,1H),6.75(tt,J＝8.8,2.0Hz,1H),3.76–3.55(m,4H),2.93–2.81(m,4H).
步骤2：合成(E)-1-(4-(3-溴丙酰)哌嗪-1-基)-3-(3,5-二氟苯基)丙-2-烯-1-酮(式C所示)
将(E)-3-(3,5-二氟苯基)-1-(哌嗪-1-基)丙-2-烯-1-酮(0.25g,1mmol)溶于干燥的二氯甲烷中，随后加入N,N-二异丙基乙胺(347μL,2.1mmol)，冰浴条件下加入溴丙酰氯(204μL,2mmol)，室温搅拌。反应3h后，停止反应。用饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱纯化，得到白色粉末(164mg，42％)1H NMR(300MHz,CDCl3)δ7.58(d,J＝15.3Hz,1H),7.07–6.95(m,2H),6.90–6.74(m,2H),3.83–3.60(m,8H),3.59–3.46(m,2H),2.95(t,J＝7.0Hz,2H).
步骤3：4-(苯并[d][1,3]二氧杂-5-基)吡啶(式E所示)
3,4-亚甲基苯硼酸(146mg,0.89mmol)和对碘吡啶(180mg,0.89mmol)溶于7.5mL甲苯和1.5mL的1,4二氧六环和0.5mL水，加入18mg的Pd(dppf)Cl2，碳酸钠(137mg,1.3mmol)，在氮气保护下，80℃反应反应6h后，停止反应，过滤除掉固体，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到棕色固体(85mg,48％)。1H NMR(300MHz,CDCl3)δ8.62(d,J＝6.0Hz,2H),7.43(d,J＝6.0Hz,2H),7.19–7.09(m,2H),6.92(d,J＝7.9Hz,1H),6.04(s,2H).
步骤4：4-(苯并[d][1,3]二氧杂-5-基)-1-苄基-1,2,3,6-四氢吡啶(式F所示)
取4-(苯并[d][1,3]二氧杂-5-基)吡啶(式E所示，75mg,0.38mmol)于1mL的丙酮中，加入苄溴(53μL,0.45mmol)室温搅拌36h后，把固体收集过滤，将固体重新溶到1.2mL的甲醇中，0℃条件下加入NaBH4(19mg,0.5mmol)，维持此温度搅拌1h后，恢复室温搅拌3h后，停止反应加入冰水淬灭反应，乙酸乙酯萃取3次，合并有机相，饱和食盐水洗涤3次，无水硫酸钠干燥，硅胶柱纯化，得到棕色固体(70mg,63％)。1H NMR(400MHz,CDCl3)δ7.40–7.27(m,5H),6.89(d,J＝1.7Hz,1H),6.85(dd,J＝8.1,1.8Hz,1H),6.75(d,J＝8.1Hz,1H),5.97–5.92(m,3H),3.63(s,2H),3.17–3.13(m,2H),2.69(t,J＝5.7Hz,2H),2.54–2.47(m,2H).
步骤5：4-(苯并[d][1,3]二氧杂-5-基)哌啶(式G所示)
取4-(苯并[d][1,3]二氧杂-5-基)-1-苄基-1,2,3,6-四氢吡啶(式F所示，70mg,0.24mmol)溶于2mL甲醇中，加入甲酸铵(120mg,2mmol)，加入7mg的Pd(OH)2，80℃反应2h后，停止反应，过滤旋干，溶于10mL的DCM中，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱纯化，得到白色固体(30mg,61％)。1H NMR(300MHz,CDCl3)δ6.77–6.59(m,3H),5.89(s,2H),3.15(d,J＝11.8Hz,2H),2.69(t,J＝12.0Hz,2H),2.52(t,J＝11.8Hz,1H),1.97(s,1H),1.77(d,J＝13.1Hz,2H),1.65–1.44(m,2H).
步骤6：合成(E)-1-(4-(3-(4-(苯并[d][1,3]二噁唑-5-基)哌啶-1-基)丙酰)哌嗪-1-基)-3-(3,5-二氟苯基)丙-2-烯-1-酮(S33)
以本实施例步骤2所得产物为起始原料，将其与本实施例步骤5所得产物反应，具体操作步骤参考实施例1步骤7。1H NMR(300MHz,CDCl3)δ7.59(d,J＝15.4Hz,1H),7.03(d,J＝6.7Hz,2H),6.90–6.64(m,5H),5.92(s,2H),3.82–3.52(m,8H),3.08–2.98(m,2H),2.81–2.73(m,2H),2.66–2.58(m,2H),2.51–2.37(m,1H),2.19–2.07(m,2H),1.88–1.66(m,4H).13CNMR(126MHz,CDCl3)δ170.74,164.86,163.18(dd,J＝249.1,12.8Hz,2×C),147.57,145.74,141.07,140.20,138.27(t,J＝9.5Hz),119.55,119.22,110.47(dd,J＝19.7,6.0Hz,2×C),108.15,107.24,104.96(t,J＝25.5Hz),100.77,54.47,54.33,45.58,45.23,42.24,41.61,41.31,33.68,31.24.HRMS(ESI):calcd for C28H32F2N3O4[M+H]+:512.2355；found 512.2354.
实施例34：合成(R,E)-3-(3,5-二氟苯基)-1-(3-甲基-4-(3-(4-苯基哌啶-1-基)丙酰)哌嗪-1-基)丙-2-烯-1-酮(S34)
采用(R)-1-N-Boc-2-甲基哌嗪为原料，操作步骤参考实施例33步骤2，得到
Figure BDA0002465807210000221
将其与4-苯基哌啶反应，得到S34。1H NMR(400MHz,CDCl3)δ7.59(d,J＝15.4Hz,1H),7.30–7.24(m,2H),7.22–7.14(m,3H),7.05–6.97(m,2H),6.92–6.75(m,2H),4.96–4.39(m,2H),4.26–3.67(m,2H),3.53–3.17(m,1H),3.13–2.44(m,9H),2.14(t,J＝11.6Hz,2H),1.90–1.68(m,4H),1.36–1.06(m,3H).13C NMR(126MHz,CDCl3)δ170.65,165.30,163.20(dd,J＝249.2,12.8Hz,2×C),146.09,141.15,138.29(t,J＝9.4Hz),128.44,126.80,126.19,119.01,110.48(dd,J＝19.8,6.0Hz,2×C),104.97(t,J＝25.5Hz),54.58,50.00,49.12,46.23,45.53,44.66,42.49,40.29,36.44,33.40,31.61,31.05.HRMS(ESI):calcd for C28H33F2N3O2[M+H]+:482.2614；found 482.2602.
实施例35：
Figure BDA0002465807210000222
试剂与条件：a)碳酸钠，二碳酸二叔丁酯，四氢呋喃；b)碘甲烷，碳酸钾，N,N-二甲基甲酰胺；c)盐酸甲醇；三乙胺，氯甲酸苄酯，二氯甲烷；d)N,N-二异丙基乙胺，溴丙酰氯，二氯甲烷；e)碳酸钾，4-苯基哌啶，N,N-二甲基甲酰胺；f)Pd/C，乙醇；g)1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐，1-羟基苯并三唑，N,N-二异丙基乙胺，二氯甲烷。
步骤1：合成1,4-二(叔-丁氧基羰基)哌嗪-2-羧酸(式1-2所示)
将Na2CO3(8g,76mmol)溶于50mL的水中，哌嗪-2-羧酸二盐酸盐(2g,10mmol)慢慢加入其中，随后加入Boc2O(7.9g,36.6mmol)和20mL的THF，室温继续搅拌20h后，停止反应，旋掉THF，乙醚萃取1次，取水相调节pH至3，乙酸乙酯萃取3次，合并有机相，无水硫酸钠干燥，过滤旋干，得到白色固体，未分离直接投入下一步。1H NMR(300MHz,CDCl3)δ4.66–4.48(m,2H),4.09–3.72(m,2H),3.31–3.04(m,2H),2.97–2.75(m,1H),1.45(d,J＝10.8Hz,18H).
步骤2：合成1,4-二-叔-丁基2-甲基哌嗪-1,2,4-三羧酸酯(式1-3所示)
将1,4-二(叔-丁氧基羰基)哌嗪-2-羧酸(2.2g,6.67mmol)溶于10mL的N,N-二甲基甲酰胺中，冰浴条件下加入碳酸钾(1.5g,10.6mmol)和碘甲烷(460μL,7.3mmol)，然后恢复室温，搅拌6h后，停止反应，倒入冰水混合物中，乙酸乙酯萃取3次，合并有机相，用饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱纯化，得到白色固体(2.2g,95％)。1H NMR(300MHz,CDCl3)δ4.75–4.66(m,1H),4.60–4.42(m,2H),4.13–3.78(m,2H),3.73(s,3H),3.32–2.98(m,2H),2.94–2.65(m,1H),1.50–1.35(m,18H).
步骤3：合成1-苯甲基3-甲基哌嗪-1,3-二羧酸酯(式1-4所示)
取1,4-二-叔-丁基2-甲基哌嗪-1,2,4-三羧酸酯溶于盐酸甲醇中，室温搅拌过夜后，未分离，旋干得到白色固体，直接投入下一步。取脱Boc的白色固体(217mg,1mmol)溶于10mL的二氯甲烷中，加入三乙胺(555μL,4mmol)，冷却到冰浴条件下，加入氯甲酸苄酯(141μL,1mmol)，恢复室温搅拌3h后，停止反应，加入水淬灭反应，乙酸乙酯萃取3次，合并有机相，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱分离，得到无色液体(218mg,79％)。1H NMR(300MHz,CDCl3)δ7.41–7.27(m,5H),5.12(s,2H),3.83–3.64(m,4H),3.54–3.41(m,1H),3.38–2.94(m,3H),2.82–2.66(m,1H),2.22–2.05(m,1H).
步骤4：合成1-苯甲基3-甲基4-(3-溴丙酰)哌嗪-1,3-二羧酸酯(式1-5所示)
将步骤3所得化合物为原料与溴丙酰氯反应，操作步骤参考实施例33步骤2。1HNMR(300MHz,CDCl3)δ7.42–7.29(m,5H),5.30(s,2H),5.25–5.03(m,3H),4.76–4.62(m,1H),3.83–3.43(m,7H),3.21–2.88(m,4H).ESI-MS m/z:435([M+Na]+),437([M+2+Na]+).
步骤5：合成1-苯甲基3-甲基4-(3-(4-苯基哌啶-1-基)丙酰)哌嗪-1,3-二羧酸酯(式1-6所示)
将步骤4所得化合物为原料与4-苯基哌啶反应，操作步骤参考实施例1步骤7。1HNMR(300MHz,CDCl3)δ7.41–7.16(m,11H),5.26–5.03(m,3H),3.81–3.44(m,5H),3.18–2.87(m,4H),2.82–2.43(m,5H),2.22–2.06(m,2H),1.91–1.65(m,5H).
步骤6：合成甲基1-(3-(4-苯基哌啶-1-基)丙酰)哌嗪-2-羧酸酯(式1-7所示)
取步骤5所得产物(330mg,0.67mmol)溶于15mL的乙醇中，加入30mg的Pd/C，室温常压反应6h后，停止反应，直接过滤，得到无色油状液体220mg，未分离直接投入下一步。1HNMR(400MHz,CDCl3)δ7.33–7.28(m,2H),7.25–7.18(m,3H),3.76(s,2H),3.74–3.67(m,1H),3.63–3.53(m,1H),3.48(s,2H),3.41(td,J＝12.5,3.6Hz,1H),3.25–3.14(m,2H),3.06–2.69(m,7H),2.63–2.52(m,1H),2.40–2.25(m,2H),2.05–1.85(m,4H).
步骤7：合成(E)-甲基4-(3-(3,5-二氟苯基)丙烯酰)-1-(3-(4-苯基哌啶-1-基)丙酰)哌嗪-2-羧酸酯(S35)
将式1-7所示化合物与3，5-二氟苯乙烯酸反应，得化合物S35，反应条件参考实施例4步骤4。1H NMR(400MHz,CDCl3)δ7.57(d,J＝15.4Hz,1H),7.33–7.27(m,2H),7.24–7.17(m,3H),7.07–6.91(m,3H),6.82(tt,J＝8.7,2.3Hz,1H),4.74–4.60(m,2H),3.89–3.79(m,1H),3.78–3.67(m,3H),3.45–3.28(m,2H),3.09–2.99(m,2H),2.90–2.62(m,5H),2.56–2.46(m,1H),2.24–2.09(m,2H),1.91–1.70(m,5H).HRMS(ESI):calcd for C29H34F2N3O4[M+H]+:526.2512；found 526.2505.
实施例36
Figure BDA0002465807210000241
试剂与条件：a)氢化铝锂，四氢呋喃；b)原乙酸三甲酯，一水合对甲苯磺酸，二氯甲烷；c)Jones’reagent，丙酮；d)2N氢氧化钠溶液，甲醇；e)1H-苯并三唑-1-基氧三吡咯烷基六氟磷酸盐，二氯甲烷；f)Dess-Martin试剂，二氯甲烷；g)4-苯基哌啶，三乙酰氧基硼氢化钠，1,2-二氯乙烷。
步骤1：合成2-异丙基丙烷-1,3-二醇(式19-1所示)
异丙基丙二酸二乙酯(2g,10mmol)溶于30mL的干燥四氢呋喃中，冰浴条件下加入氢化铝锂(570mg,15mmol)，室温反应3h后，停止反应，冰浴条件下加入10％氢氧化钠溶液淬灭，硅藻土过滤，滤液用乙酸乙酯萃取，合并有机相，有机相用饱和食盐水洗涤，无水硫酸钠干燥，过滤旋干，得到无色液体(1.1g,92％)。1H NMR(300MHz,CDCl3)δ3.92–3.71(m,4H),2.52(t,J＝4.4Hz,2H),1.80–1.67(m,1H),1.62–1.49(m,1H),0.93(d,J＝6.8Hz,6H).
步骤2：合成2-(羟甲基)-3-甲基丁基乙酸酯(式19-2所示)
取2-异丙基丙烷-1,3-二醇(1g,8.5mmol)溶于50mL的二氯甲烷中，加入原乙酸三甲酯(1.12g,9.35mmol)，一水合对甲苯磺酸(161mg,0.85mmol)，室温搅拌1h后，直接旋干，硅胶柱纯化，得到无色液体(860mg,64％)1H NMR(300MHz,CDCl3)δ4.29(dd,J＝11.3,4.4Hz,1H),4.14(dd,J＝11.3,6.7Hz,1H),3.74–3.53(m,2H),2.07(s,3H),1.96(t,J＝5.3Hz,1H),1.84–1.70(m,1H),0.96(d,J＝6.8Hz,6H).
步骤3：合成2-(乙酰氧基甲基)-3-甲基丁酸(式19-3所示)
取2-(羟甲基)-3-甲基丁基乙酸酯(0.86g,5.4mmol)溶于15mL的丙酮中，冰浴条件下，慢慢加入新鲜配置的Jones’reagent，直到反应体系维持棕色20min后，加入异丙醇慢慢使反应体系澄清，变成绿色，过滤，滤液浓缩后，用乙酸乙酯溶解，饱和碳酸氢钠洗涤，取水相，用3N盐酸调节pH至酸性，DCM萃取3次，合并有机相，无水硫酸钠干燥，过滤旋干，得到无色液体(0.635g,68％)。1H NMR(400MHz,CDCl3)δ4.31(dd,J＝10.9,4.6Hz,1H),4.22(dd,J＝10.9,9.7Hz,1H),2.59–2.50(m,1H),2.05(s,3H),2.00(dd,J＝13.7,6.8Hz,1H),1.01(d,J＝6.9Hz,6H).
步骤4：合成2-(羟甲基)-3-甲基丁酸(式19-4所示)
取2-(乙酰氧基甲基)-3-甲基丁酸(0.63g,3.6mmol)溶于10mL的甲醇中，加入2N氢氧化钠溶液3.6mL，回流反应4h后，停止反应，冷却至室温，旋去有机溶剂，用3N盐酸调节pH至酸性，DCM萃取3次，合并有机相，无水硫酸钠干燥，过滤旋干得到，白色固体(250mg,53％)1HNMR(300MHz,CDCl3)δ3.93–3.72(m,2H),2.49–2.36(m,1H),2.09–1.97(m,1H),0.99(dd,J＝6.7,3.3Hz,6H).
步骤5：合成(E)-1-(4-(3-(3,5-二氟苯基)丙烯酰)哌嗪-1-基)-2-(羟甲基)-3-甲基丁烷-1-酮(式19-5所示)
操作步骤参考实施例4步骤4。1H NMR(400MHz,CDCl3)δ7.50(d,J＝15.4Hz,1H),7.02–6.94(m,2H),6.84(d,J＝15.4Hz,1H),6.76–6.69(m,1H),3.85–3.47(m,11H),2.02–1.86(m,1H),0.88(dd,J＝21.8,6.7Hz,6H).
步骤6：合成(E)-2-(4-(3-(3,5-二氟苯基)丙烯酰)哌嗪-1-羰基)-3-甲基丁醛(式19-6所示)
取步骤5所得产物(100mg,0.27mmol)溶于15mL的二氯甲烷中，冰浴条件下加入Dess-Martin试剂(139mg,0.33mmol)，室温搅拌1h后，停止反应，体系出现浑浊，加入饱和碳酸氢钠溶液，DCM萃取3次，合并有机相，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱纯化，得到白色固体(90mg,91％)。1H NMR(400MHz,CDCl3)δ9.62(d,J＝3.8Hz,1H),7.59(d,J＝15.3Hz,1H),7.07–6.99(m,2H),6.88–6.79(m,2H),3.92–3.79(m,2H),3.79–3.51(m,6H),3.24–3.13(m,1H),2.64–2.55(m,1H),1.01(dd,J＝6.7,3.2Hz,6H).
步骤7：合成(E)-1-(4-(3-(3,5-二氟苯基)丙烯酰)哌嗪-1-基)-3-甲基-2-((4-苯基哌啶-1-基)甲基)丁烷-1-酮(S36)
取(E)-2-(4-(3-(3,5-二氟苯基)丙烯酰)哌嗪-1-羰基)-3-甲基丁醛(106mg,0.3mmol)溶于5mL的1,2-二氯乙烷中，加入4-苯基哌啶(58mg,0.36mmol)，三乙酰氧基硼氢化钠(95mg,0.45mmol)，室温搅拌0.5h后，停止反应，加入饱和碳酸氢钠溶液淬灭反应，二氯甲烷萃取3次，合并有机相，饱和食盐水洗涤，无水硫酸钠干燥，硅胶柱纯化，得到白色固体(80mg,53％)。1H NMR(400MHz,CDCl3)δ7.59(d,J＝15.5Hz,1H),7.31–7.27(m,2H),7.21–7.16(m,3H),7.06–7.00(m,2H),6.89–6.78(m,2H),3.95–3.59(m,8H),3.07–2.98(m,1H),2.91–2.82(m,2H),2.74–2.63(m,1H),2.54–2.41(m,2H),2.22–2.13(m,1H),2.11–2.01(m,1H),1.95–1.65(m,5H),0.98(d,J＝6.7Hz,3H),0.92(d,J＝6.7Hz,3H).HRMS(ESI):calcdfor C30H37F2N3O2[M+H]+:510.2927；found 510.2922.
实施例37：合成(S,E)-3-(3,5-二氟苯基)-1-(2-甲基-4-(3-(4-苯基哌啶-1-基)丙酰)哌嗪-1-基)丙-2-烯-1-酮(S37)
采用(S)-4-N-Boc-2-甲基哌嗪为原料为原料，操作步骤参考实施例33步骤2，得到
Figure BDA0002465807210000261
将其与4-苯基哌啶反应，得到S37。1H NMR(400MHz,CDCl3)δ7.58(d,J＝15.4Hz,1H),7.32–7.25(m,2H),7.23–7.15(m,3H),7.06–6.97(m,2H),6.88–6.74(m,2H),4.67–4.24(m,2H),3.80(dd,J＝67.0,12.6Hz,1H),3.47–3.18(m,2H),3.09–2.90(m,3H),2.85–2.45(m,6H),2.24–2.06(m,3H),1.90–1.70(m,4H),1.41–1.23(m,3H).HRMS(ESI):calcd for C28H34F2N3O2[M+H]+:482.2614；found 482.2620.
实施例38：合成(E)-3-(3,5-二氟苯基)-1-(4-(2-苯基-3-(4-苯基哌啶-1-基)丙酰)哌嗪-1-基)丙-2-烯-1-酮(38)
将实施例36步骤1的原料替换为
Figure BDA0002465807210000262
其他步骤相同，得到产物S38。1HNMR(400MHz,CDCl3)δ7.53(d,J＝15.3Hz,1H),7.40–7.13(m,10H),7.06–6.93(m,2H),6.88–6.68(m,2H),4.10–3.89(m,2H),3.85–3.26(m,7H),3.10–2.84(m,3H),2.61–2.40(m,2H),2.29–2.08(m,2H),1.84–1.62(m,4H).13C NMR(126MHz,CDCl3)δ171.45,164.77,163.20(dd,J＝249.2,12.8Hz,2×C),146.35,140.93,138.77,138.32(t,J＝9.4Hz),129.05,128.39,127.83,127.32,126.83,126.11,119.30,110.46(dd,J＝20.3,5.3Hz,2×C),104.93(t,J＝25.4Hz),62.69,54.85,47.63,47.24,45.51,45.37,42.47,42.28,41.93,33.48,33.44.HRMS(ESI):calcd for C33H36F2N3O2[M+H]+:544.2770；found 544.2759.
实施例39：合成(S,E)-3-(3,5-二氟苯基)-1-(3-甲基-4-(3-(4-苯基哌啶-1-基)丙酰)哌嗪-1-基)丙-2-烯-1-酮(S39)
采用(S)-1-N-Boc-2-甲基哌嗪为原料，按照实施例实施例33步骤2的方法合成
Figure BDA0002465807210000263
然后将其与4-苯基哌啶反应，得到S39。1H NMR(300MHz,CDCl3)δ7.60(d,J＝15.4Hz,1H),7.33–7.25(m,2H),7.24–7.14(m,3H),7.07–6.96(m,2H),6.94–6.73(m,2H),4.97–4.39(m,2H),4.28–3.64(m,2H),3.53–3.15(m,2H),3.11–2.42(m,8H),2.14(t,J＝11.3Hz,2H),1.92–1.68(m,4H),1.32–1.09(m,3H).HRMS(ESI):calcd forC28H34F2N3O2[M+H]+:482.2614；found 482.2626.
实施例40：合成(E)-1-(4-(3-(4-(1H-吲哚-4-基)哌啶-1-基)丙酰)哌嗪-1-基)-3-(3,5-二氟苯基)丙-2-烯-1-酮(S40)
将实施例33制备的式C所示的化合物与4-(哌啶-4-基)-1H-吲哚反应，得到S40。1HNMR(400MHz,CDCl3)δ8.59(s,1H),7.57(dd,J＝15.4,3.6Hz,1H),7.25–7.13(m,2H),7.08–6.97(m,3H),6.90–6.76(m,2H),6.58(dd,J＝33.6,7.7Hz,1H),3.82–3.40(m,9H),3.14–2.90(m,2H),2.79–2.51(m,4H),2.49–2.37(m,1H),2.13–1.62(m,6H).HRMS(ESI):calcdfor C29H33F2N4O2[M+H]+:507.2566；found 507.2577.
实施例41
Figure BDA0002465807210000271
试剂与条件：a)丙烯酸甲酯；b)水合肼，乙醇；c)三甲基氧鎓四氟硼酸，二氯甲烷；d)甲醇，回流。
步骤1：合成甲基3-(4-苯基哌啶-1-基)丙酸酯(式41-1所示)
取4-苯基哌啶(161mg,1mmol)溶于丙烯酸甲酯(0.3mL,3.3mmol)，室温搅拌2.5h后，停止反应，硅胶过滤，用少量乙醇洗涤，通风橱旋干，未分离直接投入下一步。得到白色固体(230mg,93％)。1H NMR(300MHz,CDCl3)δ7.34–7.25(m,2H),7.25–7.16(m,3H),3.69(s,3H),3.09–3.00(m,2H),2.77(t,J＝7.5Hz,2H),2.58(t,J＝7.4Hz,2H),2.54–2.44(m,1H),2.21–2.09(m,2H),1.89–1.80(m,4H).
步骤2：合成3-(4-苯基哌啶-1-基)丙酰肼(式41-2所示)
取甲基3-(4-苯基哌啶-1-基)丙酸酯(230mg,0.93mmol)溶于5mL的乙醇中，加入水合肼200μL，回流反应20h后，停止反应，旋干有机溶剂，残余物直接上样过柱，得到白色固体(122mg,91％)。1H NMR(300MHz,CDCl3)δ9.39(s,1H),7.36–7.27(m,2H),7.25–7.17(m,3H),3.90(s,2H),3.08(d,J＝11.5Hz,2H),2.68–2.41(m,5H),2.16–2.03(m,2H),1.95–1.68(m,6H).
步骤3：合成(E)-3-(3,5-二氟苯基)-1-(3-(2-(4-苯基哌啶-1-基)乙基)-5,6-二氢-[1,2,4]三唑并[4,3-a]吡嗪-7(8H)-基)丙-2-烯-1-酮(S41)
取(E)-4-(3-(3,5-二氟苯基)丙烯酰)哌嗪-2-酮(式41-3所示，即实施例4步骤1所合成的化合物②，290mg,1.09mmol)溶于5mL干燥的二氯甲烷中，加入三甲基氧鎓四氟硼酸(161mg,1.09mmol)，室温搅拌过夜后，体系变成白色浑浊，将3-(4-苯基哌啶-1-基)丙酰肼(269mg,1.09mmol)用二氯甲烷溶解后加入，继续室温搅拌24h后，颜色变成澄清，停止反应，用饱和食盐水洗涤，无水硫酸钠干燥，过滤，得到白色固体(式41-4)，未分离直接投入下一步，用甲醇溶解，回流反应3h后，停止反应，旋干溶剂，硅胶柱分离，得到白色固体(式41-5，210mg,44％)。1H NMR(400MHz,CDCl3)δ7.63(d,J＝15.3Hz,1H),7.33–7.27(m,2H),7.24–7.17(m,3H),7.07–7.01(m,2H),6.92–6.80(m,2H),5.06(s,2H),4.21–3.99(m,4H),3.10–3.04(m,2H),2.98–2.93(m,2H),2.88–2.82(m,2H),2.57–2.47(m,1H),2.18(td,J＝11.8,2.3Hz,2H),1.91–1.85(m,2H),1.80–1.68(m,2H).13C NMR(126MHz,CDCl3)δ165.3,163.23(dd,J＝249.5,12.8Hz,2×C),152.76,146.95,145.97,142.34,137.85(t,J＝9.4Hz),128.58,126.77,126.25,118.50,110.69(dd,J＝19.9,6.0Hz,2×C),105.37(t,J＝25.5Hz),56.18,54.45,43.11,42.40,41.57,39.33,33.42,22.96.HRMS(ESI):calcd forC27H30F2N5O[M+H]+:478.2413；found 478.2413.
实施例42：合成(E)-3-(3,5-二氟苯基)-1-(4-(2-(4-苯基哌啶-1-基)乙酰基)哌嗪-1-基)丙-2-烯-1-酮(S42)
将实施例33步骤2中的溴丙酰氯替换为溴乙酰氯，得到
Figure BDA0002465807210000281
将其与4-苯基哌啶反应，得到S42。1H NMR(400MHz,CDCl3)δ7.60(d,J＝15.3Hz,1H),7.34–7.27(m,2H),7.24–7.17(m,3H),7.07–7.00(m,2H),6.91–6.76(m,2H),3.86–3.56(m,8H),3.25(s,2H),3.03–2.95(m,2H),2.59–2.46(m,1H),2.23(t,J＝11.6Hz,2H),1.93–1.83(m,2H),1.82–1.68(m,2H).HRMS(ESI):calcd for C26H30F2N3O2[M+H]+:454.2301；found454.2290.
实施例43：合成(E)-3-(3,5-二氟苯基)-1-(4-(2-(4-苯基哌啶-1-基)乙基)哌嗪-1-基)丙-2-烯-1-酮(S43)
按照实施例1步骤1-2的方法，将实施例1步骤1中的1,3-二溴丙烷替换为1,2-二溴乙烷，得到
Figure BDA0002465807210000282
将其与4-苯基哌啶反应，得到S43。1H NMR(400MHz,CDCl3)δ7.55(d,J＝15.4Hz,1H),7.33–7.27(m,2H),7.25–7.17(m,3H),7.05–6.98(m,2H),6.87(d,J＝15.4Hz,1H),6.80(tt,J＝8.8,2.3Hz,1H),3.80–3.62(m,4H),3.14–3.04(m,2H),2.68–2.45(m,9H),2.18–2.08(m,2H),1.89–1.79(m,4H).13C NMR(126MHz,CDCl3)δ164.08,162.77(dd,J＝248.9,12.8Hz,2×C),145.73,139.86,138.18(t,J＝9.5Hz),128.02,126.42,125.78,119.39,109.94(dd,J＝19.7,6.0Hz,2×C),104.30(t,J＝25.6Hz),55.77,55.49,54.45,53.54,52.79,45.47,42.14,41.81,32.92.HRMS(ESI):calcdfor C28H34F2O2[M+H]+:440.2521；found 440.2525.
实施例44：合成(E)-3-(3,5-二氟苯基)-1-(4-(3-(3-苯基哌啶-1-基)丙基)哌嗪-1-基)丙-2-烯-1-酮
将实施例1中步骤7的2H-螺[苯并[b]噻吩-3,4'-哌啶]换成3-苯基哌啶，其它操作步骤同实施例1。1H NMR(400MHz,CDCl3)δ7.54(d,J＝15.4Hz,1H),7.34–7.27(m,2H),7.25–7.18(m,3H),7.06–6.98(m,2H),6.86(d,J＝15.4Hz,1H),6.79(tt,J＝8.7,2.2Hz,1H),3.79–3.59(m,4H),3.04–2.95(m,2H),2.88–2.77(m,1H),2.50–2.44(m,4H),2.43–2.36(m,4H),2.04–1.90(m,3H),1.84–1.66(m,5H),1.52–1.40(m,1H).13C NMR(126MHz,CDCl3)δ164.49,163.19(dd,J＝248.9,12.8Hz,2×C),144.66,140.21,138.63(t,J＝9.5Hz),128.39,127.20,126.37,119.89,110.35(dd,J＝19.7,6.0Hz,2×C),104.68(t,J＝25.5Hz),61.36,56.99,56.54,54.00,53.48,52.80,45.90,42.91,42.25,31.57,25.75,24.31.HRMS(ESI):calcd for C27H34F2N3O[M+H]+:454.2664；found 454.2658.
实施例45：合成(E)-1-(4-(3-(4-苯甲酰哌嗪-1-基)丙基)哌嗪-1-基)-3-(3,5-二氟苯基)丙-2-烯-1-酮(S45)
将实施例1中步骤7的2H-螺[苯并[b]噻吩-3,4'-哌啶]换成1-苯甲酰哌嗪，其它操作步骤同实施例1。1H NMR(400MHz,CDCl3)δ7.54(d,J＝15.4Hz,1H),7.43–7.37(m,5H),7.04–6.97(m,2H),6.85(d,J＝15.4Hz,1H),6.79(tt,J＝8.8,2.3Hz,1H),3.85–3.60(m,6H),3.50–3.35(m,2H),2.58–2.31(m,12H),1.75–1.69(m,3H).13C NMR(126MHz,CDCl3)δ170.17,164.45,163.11(dd,J＝248.8,12.9Hz,2×C),140.11,138.61(t,J＝9.5Hz),135.80,129.61,128.42,126.98,119.93,110.33(dd,J＝19.7,6.0Hz,2×C),104.60(t,J＝25.5Hz),56.30,56.24,53.51,52.75,47.67,45.81,42.17,24.15.HRMS(ESI):calcd forC27H33F2N4O2[M+H]+:483.2566；found 483.2572.
实施例46：
通过建立了共转CCR2和Ga16的细胞系，使得受体被激活后能引起Gα16蛋白的活化，进而激活磷脂酶C(PLC)产生IP3和DAG，IP3可与细胞内内质网和线粒体上的IP3受体结合，从而引起胞内钙的释放。因此，测定胞内钙的变化可以作为检测CCR2活化状态的方法。Fluo-4/AM是一种钙荧光探针指示剂用来测量钙离子，作为非极性脂溶性的化合物，进入细胞后在细胞脂解酶的作用下，AM基团解离，释出Fluo-4；由于Fluo-4是极性分子，不易通过脂质双分子膜，它可使Fluo-4长时间保留在细胞内。最终可以通过测量被激发的荧光强度来反映Ga蛋白被激活的水平。如果筛选的化合物能够激动CCR2受体，则可以使钙流反应大大升高。反之，如果筛选的化合物能够拮抗CCR2，则可以使钙流反应大大降低。实验步骤如下：
将稳定表达CCR2/Ga16的CHO细胞种于96孔板，培养过夜。然后吸去种有细胞的孔内的培液，加入新鲜配制的染料40μL/孔，37℃培养箱内恒温孵育40分钟。再用钙缓冲液将待测的药物稀释并混匀。
(1)激动模式：
将染料吸尽弃去，用新鲜配制的钙缓冲液洗一遍后，换上50μL钙缓冲液。用FlexStation II仪检测，第15秒开始由仪器自动加入25μL溶解有实施例1-45中所制备S1-S45及TAK-779(阳性对照)的钙缓冲液，最终读取525nm处荧光值。
(2)拮抗模式：
将染料吸尽弃去，用新鲜配制的钙缓冲液洗一遍后，换上50μL溶解有实施例1-45中所制备S1-S45及TAK-779(阳性对照)的钙缓冲液。用FlexStation II仪检测，第15秒开始由仪器自动加入25μL溶解有CCL2的钙缓冲液，最终读取525nm处荧光值。结果如表1所示。
表1：不同化合物对CCR2的抑制活性
化合物 IC<sub>50</sub>(nM) 化合物 IC<sub>50</sub>(nM) 化合物 IC<sub>50</sub>(nM) 化合物 IC<sub>50</sub>(nM)
TAK-779 12 S13 NA S26 330 S39 7540
S1 114 S14 929 S27 NA S40 NA
S2 2549 S15 250 S28 NA S41 1600
S3 2077 S16 46 S29 NA S42 NA
S4 12 S17 NA S30 147 S43 860
S5 15 S18 120 S31 628 S44 686
S6 15 S19 365 S32 31 S45 NA
S7 12 S20 NA S33 1300
S8 111 S21 NA S34 3266
S9 66 S22 699 S35 2444
S10 4655 S23 135 S36 5966
S11 25 S24 365 S37 1077
S12 NA S25 560 S38 5926
表1中，NA代表没有抑制活性。结果表明，表中部分化合物表现出与阳性对照相当的抑制活性。
以上所述仅是本发明的优选实施方式，并不用于限制本发明，应当指出，对于本技术领域的普通技术人员来说，在不脱离本发明技术原理的前提下，还可以做出若干改进和变型，这些改进和变型也应视为本发明的保护范围。

Claims (5)

1.一种哌嗪类化合物，其特征在于，其结构式如式(I)所示：
Figure FDA0003065498880000011
其中，A为取代苯基；所述取代苯基上含有1-3个取代基；所述取代基选自卤素、三氟甲基；
L选自
Figure FDA0003065498880000012
a＝0或1；
X和Y分别独立地选自羰基或亚甲基，或者X、Y和与其相连的哌嗪环上的N原子形成五元杂芳基，五元杂芳基中的杂原子选自N原子；
Z为C原子，b＝1；
R1和R2均为氢原子，或R1和R2其中之一为氢原子，另一个为卤素、C1-C6烷基、C1-C6烷氧基、C1-C6烷基取代的酰胺基或C1-C6酯基；
R3为氢、C1-C6烷基或苯基；
R4和R5形成以下螺环结构：
Figure FDA0003065498880000013
其中W选自O原子、S原子、SO基、SO2基或羰基；k为1或2；R9为氢原子、C1-C6烷基、卤素或氰基；
R6，R7和R8分别独立地选自氢原子；
m和n分别独立地选自0-2中任一整数，且m+n≤3。
2.一种哌嗪类化合物，其特征在于，所述哌嗪类化合物的结构式如下：
Figure FDA0003065498880000021
3.如权利要求1-2中任一项所述的哌嗪类化合物在制备趋化因子受体CCR2拮抗剂中的应用。
4.如权利要求1-2中任一项所述的哌嗪类化合物在制备用于治疗由CCR2介导的疾病的药物中的应用。
5.根据权利要求4所述的应用，其特征在于：所述由CCR2介导的疾病包括风湿性关节炎、动脉粥样硬化、哮喘、肥胖、二型糖尿病、慢性神经退行性疾病和肿瘤中的一种或几种。
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