CN109810071B - 一种miRNA生物合成抑制剂 - Google Patents
一种miRNA生物合成抑制剂 Download PDFInfo
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Abstract
本发明提供了一种式I所示的化合物、或其构象异构体、或其旋光异构体、或其药学上可接受的盐。其能够与miRNA生物合成过程中的相关结合蛋白紧密地结合，并且能够有效抑制miRNA‑21的合成。本发明制备的活性化合物可用做miRNA‑21抑制剂，进一步作为治疗恶性肿瘤的潜在药物。
Description
技术领域
本发明属于化学医药领域，具体涉及一种miRNA生物合成抑制剂。
背景技术
癌症在中国城市已成为首位死因，在农村为第二位死因。虽然我国把抗癌药物的研发作为优先项目进行发展，但是目前对抗癌药物的研究主要存在以下两个困境：一是药物靶点过少：目前临床上市的两千多个药物的靶点仅为三百个左右，主要分布于激素受体和GPCR等七类基因家族。二是对疾病的发生以及蛋白质信号网络了解的不清楚，导致药物的临床开发失败，成本过高。
miRNA在体内具有重要的功能，miRNA的表达异常会导致诸多疾病的发生，因此，miRNA近年来成为在癌症治疗中日益关注的新药靶点以及诊断标志物。而TRBP由于其具有生物学功能多样性，且与多种恶性肿瘤细胞的增殖、分化和迁移有关，在动物细胞中TRBP作为Dicer伴侣，TRBP可通过影响Dicer和Ago2介导的miRNA成熟，进而影响癌细胞的恶性转移。
但是，目前已有的miRNA抑制剂，其抑制效果还远远不能达到临床要求。因此，进一步深入研究化合物对miRNA的抑制机理，研发出新的对miRNA的生成具有更佳抑制效果的抑制剂，在肿瘤治疗中十分必要。
发明内容
本发明的目的在于提供一种有效抑制miRNA-21生物合成的化合物。
本发明提供了式I所示的化合物、或其构象异构体、或其旋光异构体、或其药学上可接受的盐：
其中：
X选自O、S；Y选自N；
R1、R2、R3、R4、R5各自独立地选自氢、取代或未取代的C1-2烷基、取代或未取代的C1-2烷氧基、卤素、氰基、硝基、取代或未取代的苯基、-COOR9；
或，R1、R2、R5各自独立地选自氢、取代或未取代的C1-2烷基、取代或未取代的C1-2烷氧基、卤素、氰基、硝基、取代或未取代的苯基、-COOR9，R3、R4与其连接的两个碳原子一起形成苯环；所述取代基选自卤素、C1-4烷基、羟基、羧基、硝基、氨基、羧基、C2-C4烯基、C1-C4烷基、C2-C4炔基、碳环基、杂环基；
R6、R8各自独立地选自氢、羟基、卤素、C1-4烷基、C3-6环烷烃基、苯基、
--COOR9、-CONHR10，其中，R9、R10各自独立地选自C1-2烷基、
其中L0、L1、L2、L3各自独立地选自1-4个亚甲基，R11选自卤素或羟基；
且当X为O，Y为N，R1、R2、R4、R5为氢，R3为甲氧基，R6为甲基，R8
为-COOR9时，R9不为乙基。
进一步地，
X选自O、S；Y选自N；
R1、R2、R3、R4、R5各自独立地选自氢、取代或未取代的C1-2烷基、取代或未取代的C1-2烷氧基、卤素、取代或未取代的苯基；或，R1、R2、R5各自独立地选自氢、取代或未取代的C1-2烷基、取代或未取代的C1-2烷氧基、卤素、取代或未取代的苯基，R3、R4与其连接的两个碳原子一起形成苯环；所述取代基选自卤素；
R6、R8各自独立地选自C1-2烷基、-COOR9，其中，R9选自C1-2烷基、
其中L0选自1-4个亚甲基。
进一步地，
X选自O；Y选自N；
R1、R2、R3、R4、R5各自独立地选自氢、取代或未取代的C1-2烷基、取代或未取代的C1-2烷氧基、卤素、取代或未取代苯基；所述取代基选自卤素；
R6、R8各自独立地选自C1-2烷基、-COOR9，其中，R9选自C1-2烷基、
其中L0选自2个亚甲基。
进一步地，
所述化合物的结构如式I-1所示：
其中：
R6选自C1-2烷基；
R7选自C1-2烷基、
其中L0选自2个亚甲基；
R3选自甲基、甲氧基、苯基、卤素，优选甲基、甲氧基、苯基、氯、溴。
进一步地，
X选自S；Y选自N；
R1、R2、R3、R4、R5各自独立地选自氢、取代或未取代的C1-2烷基、取代或未取代的C1-2烷氧基、卤素、取代或未取代的苯基；或，R1、R2、R5各自独立地选自氢、取代或未取代的C1-2烷基、取代或未取代的C1-2烷氧基、卤素、取代或未取代的苯基，R3、R4与其连接的两个碳原子一起形成苯环；所述取代基选自卤素；
R6、R8各自独立地选自C1-2烷基、-COOR9，其中，R9选自C1-2烷基、
其中L0选自1-4个亚甲基。
进一步地，
X选自S；Y选自N；
R1、R2、R4、R5为氢，R3选自取代或未取代的C1-2烷基、取代或未取代的C1-2烷氧基、卤素、取代或未取代的苯基，所述取代基选自卤素；或，R1、R2、R5为氢，R3、R4与其连接的两个碳原子一起形成苯环；
R6、R8各自独立地选自C1-2烷基、-COOR9，其中，R9选自C1-2烷基、
L0为2个亚甲基。
进一步地，
其结构如式I-2：
其中：
R6、R8中，一个为C1-2烷基，另一个为-COOR9，其中，R9选自C1-2烷基、
L0为2个亚甲基；
R3选自卤代或未卤代的C1-2烷基、C1-2烷氧基、卤素、苯基，优选三氟甲基、甲氧基、乙氧基、甲基；R4选自氢、甲基；或，R3、R4与其连接的两个碳原子一起形成苯环。
进一步地，
所述化合物为：
本发明还提供了上述化合物或其药学上可接受的盐作为miRNA-21生物合成抑制剂的用途。
进一步地，所述抑制剂为治疗肿瘤的药物，优选为治疗恶性肿瘤的药物。。
通过试验证明，本发明提供了一种式Ⅰ所示的化合物或其药学上可接受的盐，其能够与miRNA生物合成过程中的相关结合蛋白紧密地结合，并且能够有效抑制miRNA-21的合成。本发明制备的活性化合物可用做miRNA-21抑制剂，进一步作为治疗恶性肿瘤的潜在药物。
“取代”是指分子中的氢原子被其它不同的原子或分子所替换。
碳氢基团中碳原子含量的最小值和最大值通过前缀表示，例如，C1-4烷基表明任何含1-4个碳原子的烷基。
显然，根据本发明的上述内容，按照本领域的普通技术知识和惯用手段，在不脱离本发明上述基本技术思想前提下，还可以做出其它多种形式的修改、替换或变更。
以下通过实施例形式的具体实施方式，对本发明的上述内容再作进一步的详细说明。但不应将此理解为本发明上述主题的范围仅限于以下的实例。凡基于本发明上述内容所实现的技术均属于本发明的范围。
附图说明
图1为活性筛选实验中本发明制备的化合物、对照化合物3b的相对荧光强度。
图2为本发明部分活性化合物、对照化合物3b的相对荧光强度。
图3为Hela-luciferase-miRNA-21细胞毒活性检测试验结果。
图4为本发明化合物与Dicer-TRBP分子对接的2D(左)和3D(右)图，2D图中的颜色代表：氨基酸残基外带着色晕圈，表示相互作用残基的溶剂可及表面积；其中，各编号对应的颜色分别表示：(1)：与化合物烷基形成烷基–π作用的氨基酸残基；(2)：与化合物形成分子间范德瓦尔斯作用的氨基酸残基；(3)：与化合物形成分子间氢键作用的氨基酸残基；(4)：与化合物形成分子间C-H氢键作用的氨基酸残基；(5)：与化合物形成Cation–π作用的氨基酸残基；(6)：与化合物形成分子间Halogen-H(Cl,Br,I)氢键作用的氨基酸残基；(7)：与化合物形成分子间π-π共轭作用的氨基酸残基。
图5为本发明化合物与TRBP(dsRBD2)分子对接的2D(左)和3D(右)图，2D图中的颜色代表：氨基酸残基外带着色晕圈，表示相互作用残基的溶剂可及表面积；其中，各编号对应的颜色分别表示：(1)：与化合物烷基形成烷基–π作用的氨基酸残基；(2)：与化合物形成分子间范德瓦尔斯作用的氨基酸残基；(3)：与化合物形成分子间氢键作用的氨基酸残基；(4)：与化合物形成分子间C-H氢键作用的氨基酸残基；(5)：与化合物形成Cation–π作用的氨基酸残基；(6)：与化合物形成分子间Halogen-H(Cl,Br,I)氢键作用的氨基酸残基；(7)：与化合物形成分子间π-π共轭作用的氨基酸残基。
图6为细胞筛选模型图。
具体实施方式
本发明具体实施方式中使用的原料、设备均为已知产品，通过购买市售产品获得。
实施例1、本发明化合物的合成
Ethyl 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylate(T2):在25mL的圆底烧瓶中加入4-甲氧基苯甲酰胺(152.2mg,1mmol)，劳森试剂(485.4mg,1.2mmol)和THF(20mL)。在常温条件下反应4h后除去劳森试剂与溶剂，经柱层析分离纯化得到中间体硫代苯甲酰胺，然后与3-溴-2-氧代丁酸乙酯(209mg,1mmol)在乙醇(10mL)作为溶剂，70℃反应条件下回流4h，反应完毕后处理除去溶剂，经柱层析分离纯化获得化合物T2。产率92％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝3:1),1H NMR(400MHz,CDCl3)7.91(d,J＝8.8Hz,2H),6.96(d,J＝8.8Hz,2H),4.37(q,J＝7.1Hz,2H),3.86(s,3H),2.76(s,3H),1.68(s,1H),1.40(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ169.9,162.4,161.9,161.0,128.4,125.9,120.9,114.4,61.1,55.5,17.6,14.4；HR-ESI-MS(positive mode)m/z:278.0851[M+H]+(Calcd for C14H16NO3S:278.0851),m/z:300.0668[M+Na]+(Calcd forC14H15NO3SNa:300.0670).
Ethyl 2-(4-methoxyphenyl)-5-methyl-1H-imidazole-4-carboxylate(T3):在25mL的圆底烧瓶中加入乙氧甲酰基亚甲基三苯基膦(CEMTPP)(1044.2mg,3mmol)，乙酰氯(474.5mg,6mmol)，N,N-二异丙基乙胺(DIPEA)(516.9mg,4mmol)，二氯甲烷(DCM)(20mL)反应1h得到3-三苯基磷乙酰乙酸乙酯，然后在THF中加入Oxone(614.7mg,1mmol)氧化，反应4h后得中间体2,3-二氧代丁酸乙酯，再与4-甲氧基苯甲醛(272.3mg,2mmol)在乙酸溶剂(10mL)，100℃温度下反应12h，反应完毕后处理除去溶剂，经柱层析分离纯化获得化合物T3。产率69％，白色固体，展开体系PE/EA10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)7.84(d,J＝8.8Hz,2H),6.94(d,J＝8.8Hz,2H),4.38(q,J＝7.1Hz,2H),3.84(s,3H),2.54(s,3H),1.39(t,J＝7.1Hz)；13C NMR(100MHz,CDCl3)δ160.8,127.3,121.8,114.3,60.6,55.4,14.5；HR-ESI-MS(positive mode)m/z:261.1236[M+H]+(Calcd for C14H17N2O3:261.1239),m/z:283.1052[M+Na]+(Calcd for C14H16N2O3Na:283.1059).
2-hydroxyethyl2-(4-methoxyphenyl)-5-methyloxazole-4-carboxylate(T4):在25mL的圆底烧瓶中加入化合物3b(5-甲基-2-(4-甲氧基苯基)-4-噁唑甲酸乙酯，261.3mg,1mmol)，EtOH/10％ NaOH＝1:1(20mL),在70℃下反应1小时，再用浓盐酸调节pH至1～2之间，二氯甲萃取，浓缩有机相得中间体3，3不用进一步纯化直接进行下一步反应。向中间体3中加入二氯甲烷20mL，1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐(EDCI)(383.4mg,2mmol)，4-二甲氨基吡啶(DMAP)(24.4mg,0.2mmol)，乙二醇(124.1mg,2mmol)，反应体系加热到80℃，回流6h。反应结束后加入20mL水淬灭反应，二氯甲烷萃取，合并有机相并浓缩，再经柱层析分离纯化(石油醚/乙酸乙酯/甲醇＝10/1/0.01)，得化合物T4。产率77％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1HNMR(400MHz,CDCl3)7.97(d,J＝8.9Hz,2H),6.97(d,J＝8.9Hz,2H),4.46(t,J＝9.2Hz,2H),3.97(t,J＝9.2Hz,2H),3.86(s,3H),2.67(s,3H),2.21(s,1H)；13C NMR(100MHz,CDCl3)δ162.6,161.8,159.9,156.3,128.3,128.0,119.0,114.3,66.8,61.0,55.4,12.1；HR-ESI-MS(positive mode)m/z:278.1023[M+H]+(Calcd for C14H16NO5:278.1028),m/z:300.0839[M+Na]+(Calcd for C14H15NO5Na:300.0848).
2-hydroxyethyl2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylate(T5):化合物T5是以T2为原料，并参考T4的合成方法获得化合物T5。产率80％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.91(d,J＝8.8Hz,2H),6.96(d,J＝8.9Hz,2H),4.44(t,J＝4.6Hz,2H),3.94(q,J＝4.1Hz,2H),3.86(s,3H),2.76(s,3H),1.67(s,1H)；13C NMR(100MHz,CDCl3)δ170.3,162.6,162.1,161.7,128.5,125.7,120.1,114.4,66.7,61.3,55.5,17.6；HR-ESI-MS(positive mode)m/z:294.0796[M+H]+(Calcd for C14H16NO4S:294.0800),m/z:316.0612[M+Na]+(Calcd forC14H15NO4SNa:316.0619).
ethyl 2-(4-methoxyphenyl)-4-methyloxazole-5-carboxylate(T6):在25mL的圆底烧瓶中加入4-甲氧基苯甲酰胺(152.2mg,1mmol)，2-氯代乙酰乙酸乙酯(329.2mg,2mmol)，以及溶剂乙醇(20mL)，在100℃的封管中反应24h得到化合物T6。产率87％，化合物T6，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝3:1),1H NMR(400MHz,CDCl3)8.06(d,J＝9.0Hz,2H),6.97(d,J＝9.0Hz,2H),4.42(q,J＝7.2Hz,2H),3.86(s,3H),2.51(s,3H),1.42(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.4,162.3,159.0,147.1,137.0,129.0,119.1,114.3,61.0,55.4,14.4,13.5；HR-ESI-MS(positive mode)m/z:262.1072[M+H]+(Calcd for C14H16NO4:262.1079),m/z:284.0895[M+Na]+(Calcd for C14H15NO4Na:284.0899).
Ethyl 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylate(T7):在25mL的圆底烧瓶中加入4-甲氧基苯甲酰胺(152.2mg,1mmol)，劳森试剂(485.4mg,1.2mmol)和THF(20mL)在常温条件下反应4h后除去劳森试剂与溶剂，经柱层析分离纯化得到中间体硫代苯甲酰胺，然后与2-氯代乙酰乙酸乙酯(329.2mg,2mmol)在溶剂乙醇(20mL)中70℃回流6小时，反应完毕后处理除去溶剂，经柱层析分离纯化获得化合物T7。产率92％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝3:1),1H NMR(400MHz,CDCl3)7.88(d,J＝8.8Hz,2H),6.95(d,J＝8.8Hz,2H),4.46(q,J＝7.1Hz,2H),3.85(s,3H),2.78(s,3H),1.46(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ163.7,162.7,161.3,143.7,142.0,128.2,125.9,114.2,61.1,55.4,14.4,13.3；HR-ESI-MS(positive mode)m/z:278.0850[M+H]+(Calcdfor C14H16NO3S:278.0851),m/z:300.0670[M+Na]+(Calcd for C14H15NO3SNa:300.0670).
Ethyl 2-(4-methoxyphenyl)-4-methyl-1H-imidazole-5-carboxylate(T8):在25mL的圆底烧瓶中加入DL-丙氨酸(540mg,6mmol)，10％的NaOH溶剂(10mL,pH＝11～12)，4-甲氧基苯甲酰氯(1026mg,6mmol),常温下反应，通过TLC监测待反应完毕，除去反应原料和溶剂，得到粗产品，再加入EDCI(223mg,1mmol)，DCM溶剂(20mL)常温下搅拌1小时后加入三丁基膦(202.3mg,0.5mmol)，氰基甲酸乙酯(79mg,0.8mmol)，TLC监测反应，待反应完毕，后处理除去溶剂与原料，经柱层析分离纯化获得化合物T8。产率64％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CD3OD)7.88(d,J＝8.8Hz,2H),7.03(d,J＝8.8Hz,2H),4.39(q,J＝7.1Hz,2H),3.86(s,3H),2.56(s,3H),1.43(t,J＝7.0Hz,3H)；13CNMR(100MHz,CDCl3)δ160.8,127.3,121.8,114.3,60.6,55.4,29.7,29.4,14.5；HR-ESI-MS(positive mode)m/z:261.1246[M+H]+(Calcd for C14H17N2O3:261.1239),m/z:283.1063[M+Na]+(Calcd for C14H16N2O3Na:283.1059).
2-hydroxyethyl 2-(4-methoxyphenyl)-4-methyloxazole-5-carboxylate(T9):化合物T9合成参考T4的合成方法。产率81％，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)8.06(d,J＝9.0Hz,2H),6.98(d,J＝9.0Hz,2H),4.49(t,J＝4.6Hz,2H),3.97(t,J＝4.6Hz,2H),3.88(s,3H),2.53(s,3H),1.69(s,1H)；13C NMR(100MHz,CDCl3)δ162.7,162.4,159.1,148.0,136.5,129.1,118.9,114.3,66.5,61.3,55.5,13.6；HR-ESI-MS(positive mode)m/z:278.1017[M+H]+(Calcd for C14H16NO5:278.1028),m/z:300.0832[M+Na]+(Calcd for C16H19NO4Na:300.0848).
2-hydroxyethyl 2-(4-methoxyphenyl)-4-methylthiazole-5-carboxylate(T10):化合物T10以T7为原料，并参考T4的合成方法获得化合物T10。产率84％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.81(d,J＝8.8Hz,2H),6.94(d,J＝8.8Hz,2H),4.45(t,J＝4.4Hz,2H),3.96(t,J＝4.4Hz,2H),3.85(s,3H),2.76(s,3H)；13C NMR(100MHz,CDCl3)δ164.1,162.6,161.5,144.7,141.2,128.2,125.5,114.3,67.0,60.9,55.4,13.3；HR-ESI-MS(positive mode)m/z:294.0800[M+H]+(Calcd for C14H16NO4S:294.0800),m/z:316.0618[M+Na]+(Calcd for C14H15NO4SNa:316.0619).
Ethyl 5-methyl-2-(pyridin-4-yl)oxazole-4-carboxylate(T11):化合物T11的合成参考3b的合成方法。产率61％，白色固体，展开体系PE/EA10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.74(d,J＝6.1Hz,2H),7.92(d,J＝6.2Hz,2H),4.46(q,J＝7.1Hz,2H),2.73(s,3H),1.44(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.0,157.4,157.3,150.6,133.5,129.5,120.1,61.3,14.4,12.3；HR-ESI-MS(positive mode)m/z:233.0920[M+H]+(Calcd for C11H12NO5:233.0926),m/z:255.0740[M+Na]+(Calcd for C11H11NO5Na:255.0746).
Ethyl 5-methyl-2-(pyridin-2-yl)oxazole-4-carboxylate(T12):化合物T12的合成参考3b的合成方法。产率67％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.71(d,J＝4.2Hz,1H),8.25(d,J＝8.0Hz,1H),7.83-7.79(m,1H),7.39-7.36(m,1H),4.44(q,J＝7.2Hz,2H),2.74(s,3H),1.42(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.2,158.3,157.5,149.9,145.4,137.0,129.1,125.1,122.5,61.2,14.4,12.4；HR-ESI-MS(positive mode)m/z:233.0925[M+H]+(Calcd for C11H12NO5:233.0926),m/z:255.0749[M+Na]+(Calcd for C11H11NO5Na:255.0746).
2-hydroxyethyl 5-methyl-2-(pyridin-2-yl)oxazole-4-carboxylate(T13):化合物T13以T12为原料，并参考T4合成方法获得。产率60％，白色固体，展开体系PE/EA/MeOH1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.70(dd,J＝1.1Hz,1H),7.46(dd,J＝1.1Hz,1H),7.12(q,J＝3.7Hz,1H),4.46(t,J＝4.6Hz,2H),3.97(t,J＝4.6Hz,2H),3.21(s,1H),2.66(s,3H)；13C NMR(100MHz,CDCl3)δ162.3,156.3,155.9,129.1,128.7,128.6,128.2,128.0,66.8,60.9,12.1；HR-ESI-MS(positive mode)m/z:247.1772[M-H]-(Calcd for C12H11N2O4:247.0719),m/z:271.0686[M+Na]+(Calcd for C12H12N2O4Na:271.0695).
Ethyl 2-(furan-2-yl)-5-methyloxazole-4-carboxylate(T14):化合物T14的合成参考3b的合成方法。产率66％，无色液体，展开体系PE/EA10:1(Rf＝0.5,PE/EA＝5:1),1HNMR(400MHz,CDCl3)7.54(dd,J＝0.6Hz,1H),7.08(dd,J＝0.5Hz,1H),6.53(q,J＝1.8Hz,1H),4.42(q,J＝7.1Hz,2H),2.68(s,3H),1.41(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.2,155.7,152.3,144.7,142.1,128.6,112.3,111.9,61.1,14.4,12.1；HR-ESI-MS(positive mode)m/z:222.0763[M+H]+(Calcd for C11H12NO4:222.0766),m/z:244.0590[M+Na]+(Calcd for C11H11NO4Na:244.0586).
2-hydroxyethyl 2-(furan-2-yl)-5-methyloxazole-4-carboxylate(T15):化合物T15合成线路是以T14为原料，并参考T4合成方法获得。产率60％，无色液体，展开体系PE/EA/MeOH1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.57(dd,J＝0.6Hz,1H),7.06(dd,J＝0.5Hz,1H),6.54(q,J＝1.8Hz,1H),4.45(t,J＝4.6Hz,2H),3.97(t,J＝4.7Hz,2H),3.59(s,1H),2.66(s,3H)；13C NMR(100MHz,CDCl3)δ162.2,156.2,152.3,144.9,141.8,128.1,112.6,112.0,66.8,60.7,12.0；HR-ESI-MS(positive mode)m/z:238.0718[M+H]+(Calcd for C11H12NO5:238.0715),m/z:260.0538[M+Na]+(Calcd forC11H11NO5Na:260.0535).
Ethyl 5-methyl-2-(thiophen-2-yl)oxazole-4-carboxylate(T16):化合物T16的合成参考3b的合成方法。产率70％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)7.72(dd,J＝1.2Hz,1H),7.44(dd,J＝1.2Hz,1H),7.11(q,J＝3.7Hz,1H),4.43(q,J＝7.1Hz,2H),2.67(s,3H),1.42(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.3,155.9,155.7,129.0,128.8,128.7,128.5,127.9,61.1,14.4,12.2；HR-ESI-MS(positive mode)m/z:238.0545[M-H]-(Calcd for C11H12NO3S:238.0538),m/z:260.0359[M+Na]+(Calcd for C11H11NO3SNa:260.0357).
2-hydroxyethyl 5-methyl-2-(thiophen-2-yl)oxazole-4-carboxylate(T17):化合物T17合成线路是以T16为原料，并参考T4合成方法获得。产率74％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.70(dd,J＝1.2Hz,1H),7.46(dd,J＝1.2Hz,1H),7.12(q,J＝3.7Hz,1H),4.45(t,J＝4.6Hz,2H),3.97(t,J＝4.6Hz,2H),3.11(s,1H),2.66(s,3H)；13C NMR(100MHz,CDCl3)δ162.3,156.3,155.9,129.1,128.7,128.6,128.2,128.0,66.8,60.9,12.1；HR-ESI-MS(positivemode)m/z:254.0465[M-H]-(Calcd for C11H12NO4S:254.0487),m/z:276.0288[M+Na]+(Calcd for C11H11NO4SNa:276.0306).
Ethyl 5-methyl-2-phenyloxazole-4-carboxylate(T18):化合物T18的合成参考3b的合成方法。产率64％，白色固体，展开体系PE/EA 10:1(Rf＝0.6,PE/EA＝3:1),1H NMR(400MHz,CDCl3)8.06-8.03(m,3H),7.74(d,J＝2.6Hz,2H),4.43(q,J＝7.1Hz,2H),2.68(s,3H),1.41(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.5,159.7,156.2,130.7,128.8,128.7,126.6,61.1,14.4,12.2；HR-ESI-MS(positive mode)m/z:232.0965[M+H]+(Calcdfor C13H14NO3:232.0974),m/z:254.0787[M+Na]+(Calcd for C13H13NO3Na:254.0793).
2-hydroxyethyl 5-methyl-2-phenyloxazole-4-carboxylate(T19):化合物T19的合成参考T4的合成方法。产率61％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)8.02-8.00(m,2H),7.46-7.45(m,3H),4.45(t,J＝4.5,2H),3.97(t,J＝4.6,2H),2.69(s,3H)；13C NMR(100MHz,CDCl3)δ162.4,159.8,156.8,131.0,128.9,128.3,126.5,126.3,66.8,60.9,12.1；HR-ESI-MS(positive mode)m/z:248.0917[M+H]+(Calcd for C13H14NO4:248.0923),m/z:270.0723[M+Na]+(Calcd forC13H13NO4Na:270.0742).
Ethyl 2-(2-methoxyphenyl)-5-methyloxazole-4-carboxylate(T20):化合物T20的合成参考3b的合成方法。产率69％，无色液体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)7.93(dd,J＝1.7Hz,1H),7.42-7.38(m,1H),7.01-6.97(m,2H),4.41(q,J＝7.1Hz,2H),3.91(s,3H),2.68(s,3H),1.40(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.6,158.3,157.7,156.0,132.1,130.7,128.5,120.5,115.8,111.7,69.9,55.9,14.4,12.2；HR-ESI-MS(positive mode)m/z:262.1082[M+H]+(Calcd forC14H16NO4:262.1079),m/z:284.0920[M+Na]+(Calcd for C14H15NO4Na:284.0899).
2-hydroxyethyl 2-(2-methoxyphenyl)-5-methyloxazole-4-carboxylate(T21):化合物T21的合成参考T4的合成方法。产率66％，无色液体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.90(dd,J＝1.7Hz,1H),7.46-7.41(m,1H),7.04-7.00(m,2H),4.43(t,J＝4.5Hz,2H),3.93-3.91(m,5H),3.33(s,1H),2.68(s,3H)；13C NMR(100MHz,CDCl3)δ162.6,158.3,157.7,156.6,132.3,130.5,128.0,120.6,115.5,111.8,66.7,60.8,55.9,12.1；HR-ESI-MS(positive mode)m/z:278.1027[M+H]+(Calcd for C14H16NO5:278.1028),m/z:300.0848[M+Na]+(Calcd forC14H15NO5Na:300.0848).
Ethyl 2-(3-methoxyphenyl)-5-methyloxazole-4-carboxylate(T22):化合物T22的合成参考3b的合成方法。产率70％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)7.63(d,J＝7.2Hz,1H),7.58(t,J＝2.4Hz,1H),7.34(t,J＝8.0Hz,1H),6.99(dd,J＝2.5Hz,1H),4.43(q,J＝7.1Hz,2H),3.85(s,3H),2.68(s,3H),1.42(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.5,159.8,159.6,156.2,129.8,128.8,127.8,119.0,117.5,111.0,61.1,55.5,14.4,12.3；HR-ESI-MS(positive mode)m/z:262.1073[M+H]+(Calcd for C14H16NO4:262.1079),m/z:284.0899[M+Na]+(Calcd forC14H15NO4Na:284.0899).
2-hydroxyethyl 2-(3-methoxyphenyl)-5-methyloxazole-4-carboxylate(T23):化合物T23的合成参考T4的合成方法。产率73％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.61(d,J＝7.2Hz,1H),7.55(t,J＝2.4Hz,1H),7.37(t,J＝8.0Hz,1H),7.02(dd,J＝2.5Hz,1H),4.47(t,J＝4.6Hz,2H),3.98(t,J＝4.6Hz,2H),3.86(s,3H),3.27(s,1H),2.66(s,3H)；13C NMR(100MHz,CDCl3)δ162.5,159.9,159.6,156.7,130.0,128.3,127.5,119.0,117.5,111.1,66.8,60.9,55.5,12.2；HR-ESI-MS(positive mode)m/z:278.1017[M+H]+(Calcd for C14H16NO5:278.1028),m/z:300.0841[M+Na]+(Calcd for C14H15NO5Na:300.0848).
Ethyl 5-methyl-2-(p-tolyl)oxazole-4-carboxylate(T24):化合物T24的合成参考3b的合成方法。产率76％，无色液体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1HNMR(400MHz,CDCl3)7.99(d,J＝8.2Hz,2H),7.28(d,J＝8.0Hz,2H),4.47(q,J＝7.1Hz,2H),2.71(s,3H),2.41(s,3H),1.45(t,J＝7.2Hz,3H)；13C NMR(100MHz,CDCl3)δ162.6,159.9,155.9,141.1,129.4,128.7,126.6,123.9,61.0,21.5,14.4,12.2；HR-ESI-MS(positivemode)m/z:246.1122[M+H]+(Calcd for C14H16NO3:246.1130),m/z:268.0949[M+Na]+(Calcdfor C14H15NO3Na:268.0950).
2-hydroxyethyl 5-methyl-2-(p-tolyl)oxazole-4-carboxylate(T25):化合物T25的合成参考T4的合成方法。产率71％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.92(d,J＝8.2Hz,2H),7.28(d,J＝8.0Hz,2H),4.48(t,J＝4.6Hz,2H),3.99(t,J＝4.7Hz,2H),3.89(s,1H),2.66(s,3H),2.41(s,3H)；13C NMR(100MHz,CDCl3)δ162.5,159.9,156.4,141.4,129.5,128.1,126.5,123,6,66.7,60.8,21.6,12.1；HR-ESI-MS(positive mode)m/z:262.1082[M+H]+(Calcd forC14H16NO4:261.1079),m/z:284.0905[M+Na]+(Calcd for C14H15NO4Na:284.0899).
Ethyl 2-(4-(fluorooxy)phenyl)-5-methyloxazole-4-carboxylate(T26):化合物T26的合成线路参考3b的合成方法。产率80％，白色固体，展开体系PE/EA10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.13(d,J＝8.8Hz,2H),7.31(d,J＝9.2Hz,2H),4.47(q,J＝7.2Hz,2H),2.72(s,3H),1.45(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.3,158.4,156.5,150.9(d,J＝1.9Hz),129.0,128.3,125.2,124.2 121.6,121.0,119.1,61.1,14.4,12.2；HR-ESI-MS(positive mode)m/z:316.0792[M+H]+(Calcd for C14H13F3NO4:316.0797),m/z:338.0612[M+Na]+(Calcd for C14H12F3NO4Na:338.0616).
2-hydroxyethyl5-methyl-2-(4-(trifluoromethoxy)phenyl)oxazole-4-carboxylate(T27):化合物T27的合成参考T4的合成方法。产率84％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)8.08(d,J＝8.9Hz,2H),7.32(d,J＝8.1Hz,2H),4.49(t,J＝4.6Hz,2H),4.00(t,J＝4.7Hz,2H),3.47(s,1H),2.70(s,3H)；13C NMR(100MHz,CDCl3)δ162.3,158.5,157.0,151.0(d,J＝1.9Hz),128.5,128.2,124.9,121.6,121.1,119.0,66.7,60.9,12.1；HR-ESI-MS(positive mode)m/z:332.0740[M+H]+(Calcd for C14H13F3NO5:332.0746),m/z:354.0563[M+Na]+(Calcd forC14H12F3NO5Na:354.0565).
Ethyl 2-(4-fluorophenyl)-5-methyloxazole-4-carboxylate(T28):化合物T28的合成参考3b的合成方法。产率83％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.07-8.04(m,2H),7.15-7.11(m,2H),4.44(q,J＝7.1Hz,2H),2.69(s,3H),1.43(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ165.5,163.0,162.4,158.8,156.2,128.8(d,J＝8.8Hz),123.0(d,J＝3.4Hz),116.1(d,J＝22.1Hz),61.1,14.4,12.2；HR-ESI-MS(positive mode)m/z:250.0877[M+H]+(Calcd for C13H13FNO3:250.0879),m/z:272.0698[M+Na]+(Calcd for C13H12FNO3Na:272.0699).
2-hydroxyethyl 2-(4-fluorophenyl)-5-methyloxazole-4-carboxylate(T29):化合物T29的合成参考T4的合成方法。产率86％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)8.02-7.97(m,2H),7.16-7.10(m,2H),4.45(t,J＝4.6Hz,2H),3.96(t,J＝4.7Hz,2H),3.71(s,1H),2.64(s,3H)；13C NMR(100MHz,CDCl3)δ165.6,163.1,162.3,158.9,156.7,128.8(d,J＝8.8Hz),128.3,122.7(d,J＝3.4Hz),116.2(d,J＝22.3Hz),66.7,60.8,12.1；HR-ESI-MS(positive mode)m/z:266.0829[M+H]+(Calcd for C13H13FNO4:266.0829),m/z:288.0651[M+Na]+(Calcd forC13H12FNO4Na:288.0648).
Ethyl 2-(4-chlorophenyl)-5-methyloxazole-4-carboxylate(T30):化合物T30的合成参考3b的合成方法。产率80％，白色固体，展开体系PE/EA10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.01-7.97(m,2H),7.43-7.40(m,2H),4.44(q,J＝7.1Hz,2H),2.69(s,3H),1.42(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.3,158.7,156.4,136.9,129.1,127.9,125.1,61.1,14.4,12.2；HR-ESI-MS(positive mode)m/z:266.0580[M+H]+(Calcd for C13H13ClNO3:266.0584),m/z:288.0409[M+Na]+(Calcd for C13H12ClNO3Na:288.0403).
2-hydroxyethyl 2-(4-chlorophenyl)-5-methyloxazole-4-carboxylate(T31):化合物T31的合成参考T4的合成方法。产率87％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.95(d,J＝8.6Hz,2H),7.43(d,J＝8.6Hz,2H),4.47(t,J＝4.5Hz,2H),3.97(t,J＝4.6Hz,2H),3.20(s,1H),2.67(s,3H)；13CNMR(100MHz,CDCl3)δ162.3,158.8,156.9,137.2,129.2,128.5,127.8,124.8,66.8,60.9,12.2；HR-ESI-MS(positive mode)m/z:282.0532[M+H]+(Calcd for C13H13ClNO4:282.0533),m/z:304.0348[M+Na]+(Calcd for C13H12ClNO4Na:304.0353).
Ethyl 2-(4-bromophenyl)-5-methyloxazole-4-carboxylate(T32):化合物T32的合成参考3b的合成方法。产率90％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)7.92(d,J＝8.6Hz,2H),7.58(d,J＝8.6Hz,2H),4.43(q,J＝7.2Hz,2H),2.68(s,3H),1.42(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.3,158.8,156.4,132.0,130.0,128.0,125.5,125.3,61.1,14.4,12.3；HR-ESI-MS(positive mode)m/z:310.0087[M+H]+(Calcd for C13H13brNO3:310.0079),m/z:331.9902[M+Na]+(Calcd forC13H12BrNO3Na:331.9898).
2-hydroxyethyl 2-(4-bromophenyl)-5-methyloxazole-4-carboxylate(T33):化合物T33的合成参考T4的合成方法。产率87％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.90(d,J＝8.6Hz,2H),7.60(d,J＝8.5Hz,2H),4.48(t,J＝4.5Hz,2H),3.97(t,J＝4.7Hz,2H),3.03(s,1H),2.68(s,3H)；13CNMR(100MHz,CDCl3)δ162.4,158.9,157.0,132.2,128.5,128.0,125.5,125.3,66.8,61.0,12.2；HR-ESI-MS(positive mode)m/z:326.0027[M+H]+(Calcd for C13H13brNO3:326.0028),m/z:347.9847[M+Na]+(Calcd for C13H12BrNO3Na:347.9847).
Ethyl 2-([1,1'-biphenyl]-4-yl)-5-methyloxazole-4-carboxylate(T34):化合物T34的合成参考3b的合成方法。产率87％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1HNMR(400MHz,CDCl3)8.15(d,J＝8.3Hz,2H),7.69(d,J＝8.3Hz,2H),7.64(d,J＝7.3Hz,2H),7.48(t,J＝7.2Hz,2H),7.40(t,J＝7.3Hz,1H),4.46(q,J＝7.1Hz,2H),2.72(s,3H),1.45(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.5,159.5,156.2,143.4,140.0,128.9,128.0,127.4,127.1,127.1,125.5,61.1,14.4,12.3；HR-ESI-MS(positivemode)m/z:308.1284[M+H]+(Calcd for C19H18NO3:308.1287),m/z:330.1097[M+Na]+(Calcdfor C19H17NO3Na:330.1106).
2-hydroxyethyl 2-([1,1'-biphenyl]-4-yl)-5-methyloxazole-4-carboxylate(T35):化合物T35的合成参考T4的合成方法。产率87％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)8.09(d,J＝8.4Hz,2H),7.69(d,J＝8.7Hz,2H),7.64(d,J＝7.2Hz,2H),7.48(t,J＝7.2Hz,2H),7.40(t,J＝7.3Hz,1H),4.49(t,J＝4.5Hz,2H),4.00(t,J＝4.6Hz,2H),3.20(s,1H),2.69(s,3H)；13C NMR(100MHz,CDCl3)δ162.5,159.6,156.8,143.6,139.9,129.0,128.4,128.0,127.5,127.1,125.2,66.8,61.0,12.2；HR-ESI-MS(positive mode)m/z:324.1242[M+H]+(Calcd forC19H18NO4:324.1236),m/z:346.1058[M+Na]+(Calcd for C19H17NO4Na:346.1055).
Ethyl2-(3,5-difluorophenyl)-5-methyloxazole-4-carboxylate(T36):化合物T36的合成参考3b的合成方法。产率87％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1HNMR(400MHz,CDCl3)7.61-7.59(m,2H),6.93-6.88(m,1H),4.46(q,J＝7.1Hz,2H),2.71(s,3H),1.44(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ164.5(d,J＝12.6Hz),162.1,162.0(d,J＝12.4Hz),155.9,129.4(t,J＝13.8Hz),109.8(d,J＝11.8Hz),109.7(d,J＝11.8Hz),106.4(t,J＝25.4Hz),61.3,14.4,12.3；HR-ESI-MS(positive mode)m/z:268.0773[M+H]+(Calcd for C13H12F2NO3:268.0785),m/z:290.0592[M+Na]+(Calcd forC13H11F2NO3Na:290.0605).
2-hydroxyethyl 2-(3,5-difluorophenyl)-5-methyloxazole-4-carboxylate(T37):化合物T37的合成参考T4的合成方法。产率86％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.58(m,2H),6.94(m,H),4.49(t,J＝4.6Hz,2H),3.98(t,J＝4.7Hz,2H),2.71(s,3H),2.55(s,1H)；13C NMR(100MHz,CDCl3)δ164.5(d,J＝12.6Hz),162.3,162.0(d,J＝12.4Hz),157.4,129.2(t,J＝13.8Hz),109.8(d,J＝11.8Hz),109.6(d,J＝11.8Hz),106.3(t,J＝25.4Hz),66.8,61.1,12.2；HR-ESI-MS(positive mode)m/z:284.0727[M+H]+(Calcd for C13H12F2NO4:284.0734),m/z:306.0549[M+Na]+(Calcd for C13H11F2NO4Na:306.0554).
Ethyl 2-(3,5-bis(trifluoromethyl)phenyl)-5-methyloxazole-4-carboxylate(T38):化合物T38的合成参考3b的合成方法。产率89％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.53(s,2H),7.95(s,1H),4.47(q,J＝7.1Hz,2H),2.75(s,3H),1.45(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ161.9,157.4,156.8,133.0(q,J＝34.0Hz),129.5,128.6,127.0,126.6(d,J＝3.1Hz),124.3,124.0(q,J＝7.4Hz),121.5,118.8,61.4,14.4,12.3；HR-ESI-MS(positive mode)m/z:368.0703[M+H]+(Calcd for C15H12F6NO3:368.0721),m/z:390.0532[M+Na]+(Calcd forC15H11F6NO3Na:390.0541).
2-hydroxyethyl 2-(3,5-bis(trifluoromethyl)phenyl)-5-methyloxazole-4-carboxylate(T39):化合物T39的合成参考T4的合成方法。产率83％，白色固体，展开体系PE/EA/MeOH1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)8.49(s,2H),7.96(s,1H),4.50(t,J＝4.5Hz,2H),3.99(t,J＝4.6Hz,2H),2.75(s,3H),2.52(s,1H)；13C NMR(100MHz,CDCl3)δ162.0,158.0,156.9,133.1(q,J＝34.0Hz),129.1,128.4,126.9,126.5(d,J＝3.2Hz),124.2(q,J＝7.6Hz),121.5,118.8,66.9,60.9,12.3；HR-ESI-MS(positive mode)m/z:384.0667[M+H]+(Calcd for C15H12F6NO4:384.0671),m/z:406.0487[M+Na]+(Calcd for C15H11F6NO4Na:406.0490).
Ethyl 5-ethyl-2-(4-methoxyphenyl)oxazole-4-carboxylate(T40):化合物T40的合成参考3b的合成方法。产率94％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.01(d,J＝8.9Hz,2H),6.96(d,J＝8.9Hz,2H),4.43(q,J＝7.1Hz,2H),3.84(s,3H),3.13(q,J＝7.7Hz,2H),1.42(t,J＝7.1Hz,3H),1.34(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.6,161.6,160.5,159.7,128.3,127.7,119.5,114.1,61.0,55.4,19.8,14.4,12.2；HR-ESI-MS(positive mode)m/z:276.1221[M+H]+(Calcd forC15H18NO4:276.1236),m/z:298.1038[M+Na]+(Calcd for C15H17NO4Na:298.1055).
2-hydroxyethyl 5-ethyl-2-(4-methoxyphenyl)oxazole-4-carboxylate(T41):化合物T41的合成参考T4的合成方法。产率91％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.96(d,J＝8.7Hz,2H),6.95(d,J＝8.7Hz,2H),4.45(t,J＝4.3Hz,2H),3.96(t,J＝4.5Hz,2H),3.84(s,3H),3.10(q,J＝7.6Hz,2H),1.32(t,J＝7.6Hz,3H),1.24(s,1H)；13C NMR(100MHz,CDCl3)δ162.5,161.7,161.0,159.8,128.3,127.2,119.2,114.2,66.6,60.8,55.4,19.7,12.1；HR-ESI-MS(positive mode)m/z:292.1174[M+H]+(Calcd for C15H18NO5:292.1185),m/z:314.1002[M+Na]+(Calcd for C15H17NO5Na:314.1004).
Ethyl 2-(4-methoxyphenyl)-5-propyloxazole-4-carboxylate(T42):化合物T42的合成参考3b的合成方法。产率89％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.01(d,J＝8.8Hz,2H),6.95(d,J＝8.9Hz,2H),4.43(q,J＝7.1Hz,2H),3.84(t,J＝7.4Hz,2H),1.81-1.72(m,2H),1.42(t,J＝7.1Hz,3H),1.02(t,J＝7.4Hz,3H)；13C NMR(100MHz,CDCl3)δ162.6,161.6,159.8,159.4,128.3,119.5,114.1,60.9,55.4,28.0,21.4,14.4,13.7；HR-ESI-MS(positive mode)m/z:290.1382[M+H]+(Calcd for C16H20NO4:290.1392),m/z:312.1202[M+Na]+(Calcd for C16H19NO4Na:312.1212).
2-hydroxyethyl 2-(4-methoxyphenyl)-5-propyloxazole-4-carboxylate(T43):化合物T43的合成参考T4的合成方法。产率87％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.98(d,J＝8.8Hz,2H),6.97(d,J＝8.8Hz,2H),4.46(t,J＝4.5Hz,2H),3.97(t,J＝4.7Hz,2H),3.85(s,3H),3.10(s,1H),3.07(t,J＝7.4Hz,2H),1.81-1.72(m,2H),1.03(t,J＝7.4Hz,3H)；13C NMR(100MHz,CDCl3)δ162.7,161.7,160.0,159.9,128.3,127.8,119.2,114.2,66.7,61.0,55.4,27.9,21.3,13.7；HR-ESI-MS(positive mode)m/z:306.1341[M+H]+(Calcd for C16H20NO5:306.1347),m/z:328.1153[M+Na]+(Calcd for C16H19NO5Na:328.1161).
Ethyl 5-isopropyl-2-(4-methoxyphenyl)oxazole-4-carboxylate(T44):化合物T44的合成参考3b的合成方法。产率89％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1HNMR(400MHz,CDCl3)8.02(d,J＝8.8Hz,2H),6.96(d,J＝8.8Hz,2H),4.44(q,J＝7.1Hz,2H),3.85(s,3H),3.84(q,7.0Hz,1H),1.43(t,J＝7.1Hz,3H),1.36(d,J＝7.0Hz,6H)；13CNMR(100MHz,CDCl3)δ163.7,162.6,161.6,159.5,128.3,126.7,119.5,114.1,60.9,55.4,26.2,20.8,14.4；HR-ESI-MS(positive mode)m/z:290.1380[M+H]+(Calcd forC16H20NO4:290.1392),m/z:312.1197[M+Na]+(Calcd for C16H19NO4Na:312.1212).
2-hydroxyethyl 5-isopropyl-2-(4-methoxyphenyl)oxazole-4-carboxylate(T45):化合物T45的合成参考T4的合成方法。产率87％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.97(d,J＝8.8Hz,2H),6.96(d,J＝8.8Hz,2H),4.45(t,J＝4.5Hz,2H),3.96(t,J＝4.7Hz,2H),3.84(s,3H),3.83(q,J＝7Hz,1H),3.67(s,1H),1.34(d,J＝7.0Hz,6H)；13C NMR(100MHz,CDCl3)δ164.3,162.6,161.7,159.6,128.3,126.1,119.2,114.2,66.7,60.8,55.4,26.2,20.7；HR-ESI-MS(positive mode)m/z:306.1337[M+H]+(Calcd for C16H20NO5:306.1341),m/z:328.1153[M+Na]+(Calcd for C16H19NO5Na:328.1161).
Ethyl 5-(tert-butyl)-2-(4-methoxyphenyl)oxazole-4-carboxylate(T46):化合物T46的合成参考3b的合成方法。产率85％，白色固体，展开体系Compound wt-3-7wasobtained as awhite solid in 62％yield；isolated by column chromatography PE/EA10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.00(d,J＝8.8Hz,2H),6.96(d,J＝8.8Hz,2H),4.45(q,J＝7.1Hz,2H),3.86(s,3H),1.50(s,9H),1.44(t,J＝7.1Hz,3H)；13CNMR(100MHz,CDCl3)δ165.1,162.6,161.5,157.9,128.3,127.3,119.5,114.1,61.2,55.4,33.5,28.3,14.4；HR-ESI-MS(positive mode)m/z:304.1550[M+H]+(Calcd for C17H22NO4:306.1341),m/z:326.1374[M+Na]+(Calcd for C17H21NO4Na:326.1368).
2-hydroxyethyl 5-(tert-butyl)-2-(4-methoxyphenyl)oxazole-4-carboxylate(T47):化合物T47的合成参考T4的合成方法。产率88％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.97(d,J＝8.8Hz,2H),6.98(d,J＝8.8Hz,2H),4.46(t,J＝4.4Hz,2H),3.97(t,J＝4.6Hz,2H),3.86(s,3H),2.36(s,1H),1.50(s,9H)；13C NMR(100MHz,CDCl3)δ166.0,162.5,161.7,158.0,128.2,126.7,119.2,114.3,67.1,61.0,55.4,33.5,28.1；HR-ESI-MS(positive mode)m/z:320.1501[M+H]+(Calcd for C17H22NO5:320.1498),m/z:342.1322[M+Na]+(Calcd forC17H21NO5Na:342.1317).
Ethyl 5-cyclopropyl-2-(4-methoxyphenyl)oxazole-4-carboxylate(T48):化合物T48的合成参考3b的合成方法。产率88％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)7.93(d,J＝8.9Hz,2H),6.93(d,J＝d,J＝8.9Hz,2H),4.45(q,J＝7.1Hz,2H),3.83(s,3H),2.83-2.76(m,1H),1.43(t,J＝7.1Hz,3H),1.18-1.15(m,4H)；13C NMR(100MHz,CDCl3)δ162.9,161.5,160.3,158.3,128.2,128.0,119.4,114.1,60.9,55.4,14.5,9.2,8.1；HR-ESI-MS(positive mode)m/z:288.1244[M+H]+(Calcd forC16H18NO4:288.1236),m/z:310.1059[M+Na]+(Calcd for C16H17NO4Na:310.1055).
2-hydroxyethyl 5-cyclopropyl-2-(4-methoxyphenyl)oxazole-4-carboxylate(T49):化合物T49的合成参考T4的合成方法。产率86％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.84(d,J＝8.6Hz,2H),6.91(d,J＝8.6Hz,2H),4.45(t,J＝4.5Hz,2H),4.27(s,1H),3.96(t,J＝4.5Hz,2H),3.81(s,3H),2.75-2.71(m,1H),1.13-1.09(m,4H)；13C NMR(100MHz,CDCl3)δ162.8,161.6,160.8,158.3,128.1,127.5,119.0,114.2,66.6,60.8,55.4,9.4,8.0；HR-ESI-MS(positivemode)m/z:304.1179[M+H]+(Calcd for C16H18NO5:304.1185),m/z:326.0999[M+Na]+(Calcdfor C16H17NO5Na:326.1004).
Ethyl 2-(4-methoxyphenyl)-5-phenyloxazole-4-carboxylate(T50):化合物T50的合成参考3b的合成方法。产率92％，白色固体，展开体系PE/EA 10:1(Rf＝0.5,PE/EA＝5:1),1H NMR(400MHz,CDCl3)8.10-8.06(m,4H),7.50-7.42(m,3H),6.98(d,J＝8.9Hz,2H),4.47(q,J＝7.1Hz,2H),3.84(s,3H),1.43(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ162.4,161.9,159.9,154.6,130.1,128.6,128.5,128.4,128.1,127.3,119.1,114.2,61.4,55.4,14.3；HR-ESI-MS(positive mode)m/z:324.1237[M+H]+(Calcd for C19H18NO4:324.1236),m/z:346.1060[M+Na]+(Calcd for C19H17NO4Na:346.1055).
2-hydroxyethyl 2-(4-methoxyphenyl)-5-phenyloxazole-4-carboxylate(T51):化合物T51的合成参考T4的合成方法。产率91％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)8.04(dd,J＝4.0Hz,2.2Hz,2H),7.97(d,J＝8.8Hz,2H),7.46(dd,J＝0.9Hz,2.3Hz,3H),6.92(d,J＝8.9Hz,2H),4.47(t,J＝4.4Hz,2H),3.98(t,J＝4.6Hz,3H),3.81(s,3H)；13C NMR(100MHz,CDCl3)δ162.2,162.0,159.9,154.8,130.3,128.5,128.4,128.4,127.5,126.9,118.7,114.3,67.2,60.7,55.4；HR-ESI-MS(positive mode)m/z:340.1175[M+H]+(Calcd for C19H18NO5:340.1185),m/z:362.0995[M+Na]+(Calcd for C19H17NO5Na:362.1004).
4-hydroxybutyl2-(4-methoxyphenyl)-5-methyloxazole-4-carboxylate(T52):在25mL的圆底烧瓶中加入化合物3b(261.3mg,1mmol)，EtOH/10％ NaOH＝1:1(20mL),在70℃下反应1h，再用浓盐酸调节pH至1～2之间，二氯甲萃取，浓缩有机相得中间体化合物3不用进一步纯化；向中间体3中加入二氯甲烷20mL，二环己基碳二亚胺(DCC)(412.6mg,2mmol)，4-二甲氨基吡啶(DMAP)(24.4mg,0.2mmol)，以及醇(2mmol)，反应体系加热到80℃，回流6个小时。反应结束后加入20mL水淬灭反应，二氯甲烷萃取，合并有机相并浓缩，再经柱层析分离纯化(石油醚/乙酸乙酯/甲醇＝10/1/0.01)，得化合物T52。产率82％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)8.00(d,J＝9.0Hz,2H),6.96(d,J＝9.0Hz,2H),4.41(t,J＝6.5Hz,2H),3.85(s,3H),3.74(t,J＝6.3Hz,2H)2.67(s,3H),1.93-1.86(m,3H),1.75-1.69(m,2H)；13C NMR(100MHz,CDCl3)δ162.5,161.7,159.8,155.6,128.4,128.3,119.3,114.2,64.8,62.2,55.4,29.4,25.2,12.2；HR-ESI-MS(positive mode)m/z:306.1341[M+H]+(Calcd for C16H20NO4:306.1333),m/z:328.1155[M+Na]+(Calcd for C16H19NO4Na:328.1161).
N-(2-hydroxyethyl)-2-(4-methoxyphenyl)-5-methyloxazole-4-carboxamide(T53):化合物T53合成方法参考T52。产率71％，白色固体，展开体系PE/EA/MeOH 1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.91(d,J＝8.9Hz,2H),7.47(s,1H),6.96(d,J＝8.9Hz,2H),3.85(s,3H),3.83(t,J＝4.8Hz,2H),3.61(q,J＝5.6Hz,2H),2.67(s,3H)；13C NMR(100MHz,CDCl3)δ163.4,161.6,158.8,152.6,129.8,128.0,119.5,114.3,62.6,55.4,42.2,11.8；HR-ESI-MS(positive mode)m/z:277.1189[M+H]+(Calcdfor C14H17N2O4:277.1188),m/z:299.1011[M+Na]+(Calcd for C14H16N2O4Na:299.1008).
2-(2-hydroxyethoxy)ethyl 2-(4-methoxyphenyl)-5-methyloxazole-4-carboxylate(T54):化合物T54的合成参考T52的合成方法。产率80％，白色固体，展开体系PE/EA/MeOH1:1:0.01(Rf＝0.5,PE/EA/MeOH＝1:1:0.01),1H NMR(400MHz,CDCl3)7.97(d,J＝8.9Hz,2H),6.93(d,J＝8.9Hz,2H),4.47(t,J＝4.7Hz,2H),3.83-3.81(m,5H),3.75(t,J＝4.2Hz,2H),3.64(t,J＝4.9Hz,2H),3.05(s,1H),2.64(s,3H)；13C NMR(100MHz,CDCl3)δ162.4,161.7,159.8,155.9,128.3,128.2,119.2,114.2,72.6,68.8,63.7,61.6,55.4,12.1；HR-ESI-MS(positive mode)m/z:322.1295[M+H]+(Calcd for C16H20NO6:322.1291),m/z:344.1114[M+Na]+(Calcd for C16H19NO6Na:344.1110).
5-chloro-2-(4-methoxyphenyl)-6-methylpyrimidin-4-ol(T55):在氮气保护下，在25mL的圆底烧瓶中加入4-甲氧基苯甲脒(330.4mg,2mmol)，NaOH(374mg,2mmol)，甲醇(20mL)，60℃反应2h，反应完毕后除去溶剂，经柱层析分离纯化获得化合物T55。产率54％，白色固体，展开体系PE/EA4:1(Rf＝0.5,PE/EA＝2:1),1H NMR(400MHz,DMSO-d6)13.01(s,1H),8.11(d,J＝8.8Hz,2H),7.09(d,J＝8.9Hz,2H),3.85(s,3H),2.42(s,3H)；13C NMR(100MHz,DMSO-d6)δ162.7,130.2,114.6,56.0,48.0,33.8,25.8,24.9,22.6；HR-ESI-MS(positive mode)m/z:251.0588[M+H]+(Calcd for C12H12ClN2O2:251.0587),m/z:273.0403[M+Na]+(Calcd for C14H16N2O3Na:273.0407).
2-(chloromethyl)-5-phenyl-1,3,4-oxadiazole(T56):在25mL的圆底烧瓶中加入苯甲酰肼(272.3mg,2mmol)，三氯氧磷(306.7mg,2mmol)以及氯乙酸(20mL)回流制备T57。产率96％，白色固体，展开体系PE/EA1:5(Rf＝0.5,PE/EA＝1:2),1H NMR(400MHz,CDCl3)8.05(d,J＝6.9Hz,2H),7.56-7.47(m,3H),4.77(s,2H)；13C NMR(150MHz,CDCl3)δ166.0,162.2,132.2,129.1,127.1,123.3,33.0；HR-ESI-MS(positive mode)m/z:195.0315[M+H]+(Calcd for C9H8N2OCl:195.0325),m/z:217.0139[M+Na]+(Calcd for C9H7N2OClNa:217.0145).
2-(2-chloroethyl)-5-phenyl-1,3,4-oxadiazole(T57):化合物T57的合成参考T56的合成方法。产率95％，白色固体，展开体系PE/EA1:5(Rf＝0.5,PE/EA＝1:2),1H NMR(400MHz,CDCl3)8.04(d,J＝7.3Hz,2H),7.55-7.48(m,3H),3.97(t,J＝6.9Hz,2H),3.43(t,J＝6.9Hz,2H)；13C NMR(150MHz,CDCl3)δ165.1,163.5,131.8,129.1,126.9,123.7,39.6,29.2；HR-ESI-MS(positive mode)m/z:209.0470[M+H]+(Calcd for C10H10N2OCl:209.0482),m/z:231.0290[M+Na]+(Calcd for C10H9N2OClNa:231.0301).
2-phenylthiazole(S1):化合物S1的合成参考T7。产率97％，无色液体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)7.99-7.96(m,2H),7.88(d,J＝3.2Hz,1H),7.46-7.41(m,3H),7.33-7.31(m,1H)；13C NMR(100MHz,CDCl3)δ168.5,143.6,133.5,130.1,129.0,126.6,118.9；HR-ESI-MS(positive mode)m/z:162.0365[M+H]+(Calcd for C9H8NS:162.0377),m/z:184.0199[M+Na]+(Calcd for C9H7NSNa:184.0197).
ethyl 2-(4-methoxyphenyl)thiazole-5-carboxylate(S2):化合物S2的合成参考T7。产率91％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)8.06(s,1H),7.93(d,J＝8.8Hz,2H),6.93(d,J＝8.8Hz,2H),4.44(q,J＝7.1Hz,2H),3.82(s,3H),1.41(t,J＝7.1Hz,3H)；13C NMR(150MHz,CDCl3)δ168.7,161.6,161.5,147.8,128.5,126.2,125.8,114.3,61.4,55.4,14.3；HR-ESI-MS(positive mode)m/z:264.0689[M+H]+(Calcd for C13H14NO3S:264.0694),m/z:286.0509[M+Na]+(Calcd for C13H13NO3SNa:286.0514).
2-hydroxyethyl2-(4-methoxyphenyl)thiazole-5-carboxylate(S3):化合物S3的合成参考T10。产率88％。白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)8.12(s,1H),7.91(d,J＝8.8Hz,2H),6.96(d,J＝8.8Hz,2H),4.47(t,J＝4.5Hz,2H),3.95(t,J＝4.6Hz,2H),3.85(s,3H),3.38(s,1H)；13C NMR(150MHz,CDCl3)δ169.1,161.8,161.6,147.2,128.6,127.0,126.5,125.5,114.4,114.3,67.1,60.9,55.4,52.4,29.7；HR-ESI-MS(positive mode)m/z:280.0634[M+H]+(Calcd for C13H14NO4S:280.0644),m/z:302.0451[M+Na]+(Calcd for C13H13NO4SNa:302.0463).
(2-(4-methoxyphenyl)-4-methylthiazol-5-yl)methanol(S4):以T7为起始原料，在氢化铝锂的THF溶剂中使用氮气保护，0℃下反应4～6h而获得化合物S4。产率85％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)7.82(d,J＝8.8Hz,2H),6.93(d,J＝8.8Hz,2H),4.78(s,2H),3.83(s,3H),2.39(s,3H),2.24(s,1H)；13CNMR(100MHz,CDCl3)δ166.2,161.1,150.1,130.1,127.9,126.5,114.2,55.8,55.4,15.1；HR-ESI-MS(positive mode)m/z:236.0751[M+H]+(Calcd for C12H14NO2S:236.0745),m/z:258.0569[M+Na]+(Calcd for C12H13NO2SNa:258.0565).
Diethyl 2-(4-methoxyphenyl)thiazole-4,5-dicarboxylate(S5):化合物S5的合成是以氯代草酰乙酸二乙酯为起始原料，并参考T7的合成方法。产率78％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)7.94(d,J＝8.9Hz,2H),6.96(d,J＝8.9Hz,2H),4.50(q,J＝7.2Hz,2H),4.39(q,J＝7.2Hz,2H),3.87(s,3H),1.44(t,J＝7.2Hz,3H),1.40(t,J＝7.2Hz,3H)；13C NMR(150MHz,CDCl3)δ171.4,163.4,162.4,160.3,150.9,128.8,127.0,125.1,114.5,62.3,62.1,55.5,14.1,14.0；HR-ESI-MS(positive mode)m/z:336.0916[M+H]+(Calcd for C16H18NO5S:336.0906),m/z:358.0734[M+Na]+(Calcd for C16H17NO5SNa:358.0725).
Ethyl 2-(4-chlorophenyl)-4-methylthiazole-5-carboxylate(S6):化合物S6的合成参考T7。产率87％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)7.78(d,J＝8.5Hz,2H),7.32(d,J＝8.5Hz,2H),4.32(q,J＝7.1Hz,2H),2.70(s,3H),1.36(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ168.1,161.9,161.0,136.9,131.3,129.1,127.8,122.0,61.2,17.5,14.3；HR-ESI-MS(positive mode)m/z:282.0360[M+H]+(Calcd for C13H13ClNO2S:282.0356),m/z:304.0182[M+Na]+(Calcd forC13H12ClNO2SNa:304.0175).
2-hydroxyethyl 2-(4-chlorophenyl)-4-methylthiazole-5-carboxylate(S7):化合物S7的合成参考T10。产率88％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)7.81(d,J＝8.5Hz,2H),7.37(d,J＝8.5Hz,2H),4.40(t,J＝4.6Hz,2H),3.92(t,J＝4.6Hz,2H),2.79(S,1H),2.72(s,3H)；13C NMR(100MHz,CDCl3)δ168.9,162.2,161.7,137.2,131.1,129.3,128.0,121.4,66.9,61.0,17.5；HR-ESI-MS(positivemode)m/z:298.0308[M+H]+(Calcd for C13H13ClNO3S:298.0305),m/z:320.0132[M+Na]+(Calcd for C13H12ClNO3SNa:320.0124).
Ethyl 2-(4-bromophenyl)-4-methylthiazole-5-carboxylate(S8):化合物S8的合成参考T7。产率98％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)7.79(d,J＝8.5Hz,2H),7.55(d,J＝8.5Hz,2H),4.37(q,J＝7.1Hz,2H),2.76(s,3H),1.40(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ168.3,162.1,161.1,132.2,131.8,128.1,125.4,122.1,61.3,17.5,14.4；HR-ESI-MS(positive mode)m/z:325.9831[M+H]+(Calcd for C13H13brNO2S:325.9850),m/z:347.9676[M+Na]+(Calcd forC13H12BrNO2SNa:347.9670).
2-hydroxyethyl 2-(4-bromophenyl)-4-methylthiazole-5-carboxylate(S9):化合物S9的合成参考T10。产率94％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)7.82(d,J＝8.5Hz,2H),7.59(d,J＝8.5Hz,2H),4.44(t,J＝4.6Hz,2H),3.95(t,J＝4.7Hz,2H),2.77(s,3H),2.01(s,1H)；13C NMR(100MHz,CDCl3)δ168.9,162.3,161.8,132.3,131.7,128.2,125.6,121.4,66.9,61.2,17.6；HR-ESI-MS(positivemode)m/z:341.9807[M+H]+(Calcd for C13H13brNO3S:341.9800),m/z:363.9624[M+Na]+(Calcd for C13H12BrNO3SNa:363.9619).
Ethyl 4-methyl-2-(4-(trifluoromethyl)phenyl)thiazole-5-carboxylate(S10):化合物S10的合成参考T7。产率87％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)8.03(d,J＝8.1Hz,2H),7.67(d,J＝8.2Hz,2H),4.36(q,J＝7.1Hz,2H),2.76(s,3H),1.39(t,J＝7.2Hz,3H)；13C NMR(100MHz,CDCl3)δ167.6,161.9(d,J＝74.1Hz),136.0,132.9(q,J＝32.7Hz),127.0,126.0(d,J＝3.7Hz),125.1,122.9,122.4,61.4,17.5,14.3；HR-ESI-MS(positive mode)m/z:316.0600[M+H]+(Calcd forC14H13F3NO2S:316.0619),m/z:338.0425[M+Na]+(Calcd for C14H12F3NO2SNa:338.0439).
2-hydroxyethyl4-methyl-2-(4-(trifluoromethyl)phenyl)thiazole-5-carboxylate(S11):化合物S11的合成参考T10。产率86％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)7.91(d,J＝8.2Hz,2H),7.59(d,J＝8.3Hz,2H),4.38(t,J＝4.6Hz,2H),3.90(t,J＝4.8Hz,2H),3.38(s,1H),2.68(s,3H)；13C NMR(100MHz,CDCl3)δ168.1,162.0(d,J＝38.1Hz),135.6,133.0(q,J＝32.8Hz),130.0,126.9,126.0(d,J＝3.8Hz),125.0,122.3,122.2,66.9,60.7,17.4；HR-ESI-MS(positive mode)m/z:332.0560[M+H]+(Calcd for C14H13F3NO3S:332.0568),m/z:354.0386[M+Na]+(Calcd forC14H12F3NO3SNa:354.0388).
Ethyl 4-methyl-2-(4-nitrophenyl)thiazole-5-carboxylate(S12):化合物S12的合成参考T7。产率83％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)8.31(d,J＝8.9Hz,2H),8.14(d,J＝8.9Hz,2H),4.40(q,J＝7.1Hz,2H),2.79(s,3H),1.41(t,J＝7.1Hz,3H)；13C NMR(150MHz,CDCl3)δ166.4,161.8,161.5,149.0,138.4,127.5,124.3,124.0,61.6,17.5,14.3；HR-ESI-MS(positive mode)m/z:293.0586[M+H]+(Calcd for C13H13N2O4S:293.0596),m/z:315.0418[M+Na]+(Calcd for C13H12N2O4SNa:315.0415).
2-hydroxyethyl4-methyl-2-(4-nitrophenyl)thiazole-5-carboxylate(S13):化合物S13的合成参考T10。产率79％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)8.32(d,J＝8.9Hz,2H),8.14(d,J＝8.9Hz,2H),4.48(t,J＝4.6Hz,2H),3.98(t,J＝4.7Hz,2H),2.81(s,3H),1.85(s,1H)；13C NMR(150MHz,CDCl3)δ166.8,162.2,162.0,149.1,138.2,127.5,124.4,123.2,67.0,61.2,17.6；HR-ESI-MS(positive mode)m/z:309.0531[M+H]+(Calcd for C13H13N2O5S:309.0545),m/z:331.0352[M+Na]+(Calcd for C13H12N2O5SNa:331.0365).
Ethyl 2-(4-isocyanophenyl)-4-methylthiazole-5-carboxylate(S14):化合物S14的合成参考T7。产率88％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1HNMR(400MHz,CDCl3)8.07(d,J＝8.4Hz,2H),7.74(d,J＝8.4Hz,2H),4.39(q,J＝7.1Hz,2H),2.78(s,3H),1.40(t,J＝7.2Hz,3H)；13C NMR(100MHz,CDCl3)δ166.9,161.9,161.4,136.7,132.8,127.2,123.6,118.2,114.1,61.6,17.5,14.3；HR-ESI-MS(positive mode)m/z:273.0692[M+H]+(Calcd for C14H13N2O2S:273.0698),m/z:295.0505[M+Na]+(Calcd forC14H12N2O2SNa:295.0517).
2-hydroxyethyl 2-(4-((2-hydroxyethoxy)carbonyl)phenyl)-4-methylthiazole-5-carboxylate(S15):4-氰基取代衍生物S14在10％NaOH溶剂中70℃加热1h，TLC检测，反应完成后使用浓HCl调节Ph至1～2，夺氯甲烷萃取并浓缩；然后加入二氯甲烷20mL，1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐(EDCI)(383.4mg,2mmol)，4-二甲氨基吡啶(DMAP)(24.4mg,0.2mmol)，乙二醇(124.1mg,2mmol)，反应体系加热到80℃，回流6个小时。待反应结束后加入20mL水淬灭反应，二氯甲烷萃取，合并有机相并浓缩，再经柱层析分离纯化，从而获得化合物S15。产率77％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)8.14(d,J＝8.4Hz,2H),8.05(d,J＝8.4Hz,2H),4.51(t,J＝4.6Hz,2H),4.47(t,J＝4.6Hz,2H),4.00(t,J＝4.6Hz,2H),3.97(t,J＝4.6Hz,2H),2.81(s,3H),1.86(s,2H)；13C NMR(150MHz,CDCl3)δ168.6,166.1,162.2,162.0,136.8,131.9,130.4,129.7,126.8,67.0,66.9,61.4,61.2,17.6；HR-ESI-MS(positive mode)m/z:352.0847[M+H]+(Calcd for C16H18NO6S:352.0855),m/z:374.0657[M+Na]+(Calcd forC16H17NO6SNa:374.0674).
Ethyl 4-methyl-2-(naphthalen-2-yl)thiazole-5-carboxylate(S16):化合物S16的合成参考T7。产率80％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1HNMR(400MHz,CDCl3)8.44(s,1H)8.01(dd,J＝1.8Hz,1.8Hz,1H),7.90-7.81(m,3H),7.53-7.48(m,2H),4.38(q,J＝7.2Hz,2H),2.81(s,3H),1.41(t,J＝7.1Hz,3H)；13C NMR(100MHz,CDCl3)δ169.9,162.3,161.1,134.5,133.1,130.2,128.9,128.8,127.9,127.5,126.9,126.6,123.8,121.9,61.3,17.6,14.4；HR-ESI-MS(positive mode)m/z:298.0894[M+H]+(Calcd for C17H16NO2S:298.0902),m/z:320.0706[M+Na]+(Calcd for C17H15NO2SNa:320.0721).
2-hydroxyethyl 4-methyl-2-(naphthalen-2-yl)thiazole-5-carboxylate(S17):化合物S17的合成参考T10。产率83％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)8.41(s,1H),7.96(dd,J＝1.6Hz,1.6Hz,1H),7.89-7.80(m,3H),7.54-7.48(m,2H),4.43(t,J＝4.5Hz,2H),3.95(t,J＝4.7Hz,2H),2.79(s,3H),2.55(s,1H)；13C NMR(100MHz,CDCl3)δ170.4,162.4,161.8,134.6,133.1,130.0,129.0,128.9,127.9,127.6,127.0,126.7,123.7,121.1,66.9,61.1,17.6；HR-ESI-MS(positive mode)m/z:314.0846[M+H]+(Calcd for C17H16NO3S:314.0851),m/z:336.0664[M+Na]+(Calcd forC17H15NO3SNa:336.0670).
Ethyl 2-(3,5-dichlorophenyl)-4-methylthiazole-5-carboxylate(S18):化合物S18的合成参考T7。产率90％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1HNMR(400MHz,CDCl3)7.82(d,J＝1.8Hz,2H),7.42(t,J＝1.8Hz,1H),4.38(q,J＝7.1Hz,2H),2.76(s,3H),1.40(t,J＝7.1Hz,3H)；13C NMR(150MHz,CDCl3)δ166.3,161.8,161.1,135.8,135.5,130.5,125.0,123.1,61.5,17.4,14.3；HR-ESI-MS(positive mode)m/z:315.9970[M+H]+(Calcd for C13H12Cl2NO2S:315.9966),m/z:337.9777[M+Na]+(Calcd forC13H11Cl2NO2SNa:337.9785).
2-hydroxyethyl 2-(3,5-dichlorophenyl)-4-methylthiazole-5-carboxylate(S19):化合物S19的合成参考T10。产率88％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)7.85(d,J＝1.8Hz,2H),7.45(t,J＝1.8Hz,1H),4.46(t,J＝4.6Hz,2H),3.96(t,J＝4.7Hz,2H),2.79(s,3H),1.81(s,1H)；13C NMR(150MHz,CDCl3)δ166.7,162.1,161.9,135.9,135.4,130.7,125.1,122.3,66.9,61.2,17.5；HR-ESI-MS(positive mode)m/z:331.9906[M+H]+(Calcd for C13H12Cl2NO3S:331.9915),m/z:353.9720[M+Na]+(Calcd for C13H11Cl2NO3SNa:353.9734).
Ethyl 2-(4-ethoxyphenyl)-4-methylthiazole-5-carboxylate(S20):化合物S20的合成参考T7。产率89％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1HNMR(400MHz,CDCl3)7.90(d,J＝8.8Hz,2H),6.94(d,J＝8.8Hz,2H),4.37(q,J＝7.1Hz,2H),4.11(q,J＝7.1Hz,2H),2.76(s,3H),1.46(t,J＝7.0Hz,3H),1.40(t,J＝7.0Hz,3H)；13C NMR(150MHz,CDCl3)δ169.9,162.4,161.4,160.9,128.4,125.7,120.8,114.9,63.7,61.1,17.5,14.7,14.3；HR-ESI-MS(positive mode)m/z:292.1003[M+H]+(Calcd for C15H18NO3S:292.1007),m/z:314.0816[M+Na]+(Calcd for C15H17NO3SNa:314.0827).
2-hydroxyethyl2-(4-ethoxyphenyl)-4-methylthiazole-5-carboxylate(S21):化合物S21的合成参考T10。产率90％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)7.90(d,J＝8.8Hz,2H),6.95(d,J＝8.8Hz,2H),4.44(t,J＝4.6Hz,2H),4.12(q,J＝7.0Hz,2H),3.95(t,J＝4.7Hz,2H),2.76(s,3H),2.10(s,1H),1.46(t,J＝7.0Hz,3H)；13CNMR(150MHz,CDCl3)δ170.4,162.6,161.7,161.5,128.5,125.5,120.0,114.9,66.7,63.7,61.3,17.6,14.7；HR-ESI-MS(positive mode)m/z:308.0955[M+H]+(Calcd for C15H18NO4S:308.0957),m/z:330.0770[M+Na]+(Calcd for C15H17NO4SNa:330.0776).
Ethyl 4-methhyl-2-(m-tolyl)thiazole-5-carboxylate(S22):化合物S22的合成参考T7。产率86％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)7.76(s,1H),7.71(d,J＝7.6Hz,1H),7.29(t,J＝7.6Hz,1H),7.23(d,J＝7.6Hz,1H),4.34(q,J＝7.1Hz,2H),2.76(s,3H),2.37(s,3H),1.38(t,J＝7.1Hz,3H)；13CNMR(100MHz,CDCl3)δ170.0,162.2,160.9,138.8,132.8,131.7,128.8,127.2,124.0,121.6,61.1,21.3,17.5,14.3；HR-ESI-MS(positive mode)m/z:262.0900[M+H]+(Calcdfor C14H16NO2S:262.0902),m/z:284.0718[M+Na]+(Calcd for C14H15NO2SNa:284.0721).
2-hydroxyethhyl 4-methyl-2-(m-tolyl)thiazole-5-carboxylate(S23):化合物S23的合成参考T10。产率84％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1HNMR(400MHz,CDCl3)7.75(s,1H),7.71(d,J＝7.4Hz,1H),7.34-7.28(m,2H),4.42(t,J＝4.4Hz,2H),3.94(t,J＝4.7Hz,2H),3.01(s,1H),2.76(s,3H),2.40(s,3H)；13C NMR(100MHz,CDCl3)δ170.7,162.4,161.5,138.9,132.6,129.0,127.3,124.1,121.0,66.8,61.0,21.3,17.5；HR-ESI-MS(positive mode)m/z:278.0845[M+H]+(Calcd for C14H16NO3S:278.0851),m/z:300.0661[M+Na]+(Calcd for C14H15NO3SNa:300.0670).
Ethyyll 2-(3,4-dimethylphenyl)-4-methylthiazole-5-carboxylate(S24):化合物S24的合成参考T7。产率88％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)7.73(s,1H),7.66(dd,J＝1.6Hz,1.6Hz,1H),7.17(d,J＝7.8Hz,1H),4.36(q,J＝7.1Hz,2H),2.76(s,3H),2.29(d,J＝7.5Hz,6H),1.39(t,J＝6.4Hz,3H)；13CNMR(100MHz,CDCl3)δ170.3,162.4,160.9,140.2,137.4,130.6,130.3,127.7,124.4,121.2,61.4,19.9,19.7,17.6,14.4；HR-ESI-MS(positive mode)m/z:276.1046[M+H]+(Calcd for C15H18NO2S:276.1058),m/z:298.0863[M+Na]+(Calcd for C15H17NO2SNa:298.0878).
2-hyddroxyethyl 2-(3,4-dimethylphenyl)-4-methylthiazole-5-carboxylate(S25):化合物S25的合成参考T10。产率82％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1H NMR(400MHz,CDCl3)7.54(s,1H),7.48(d,J＝7.6Hz,1H),7.04(d,J＝7.8Hz,1H),4.30(t,J＝5.9Hz,2H),3.92(s,1H),3.84(t,J＝4.5Hz,2H),2.63(s,3H),2.18(d,J＝6.9Hz,6H)；13C NMR(100MHz,CDCl3)δ170.8,162.3,161.2,140.4,137.4,130.2,127.7,124.3,120.6,66.7,60.6,19.8,19.7,17.4；HR-ESI-MS(positive mode)m/z:292.1001[M+H]+(Calcd for C15H18NO3S:292.1007),m/z:314.0817[M+Na]+(Calcd for C15H17NO3SNa:314.0827).
Ethyl 2-(3,5-dimethoxyphenyl)-4-methylthiazole-5-carboxylate(S26):化合物S26的合成参考T7。产率84％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)7.11(d,J＝2.2Hz,2H),6.56(t,J＝2.2Hz,2H),4.37(q,J＝7.1Hz,2H),3.85(s,6H),2.77(s,3H),1.40(t,J＝7.1Hz,3H)；13C NMR(150MHz,CDCl3)δ169.7,162.2,161.2,160.8,134.7,121.9,104.7,103.5,61.2,55.6,17.5,14.3；HR-ESI-MS(positive mode)m/z:308.0951[M+H]+(Calcd for C15H18NO4S:308.0957),m/z:330.0769[M+Na]+(Calcd for C15H17NO4SNa:330.0776).
2-hydroxyethyl 2-(3,5-dimethoxyphenyl)-4-methylthiazole-5-carboxylate(S27):化合物S27的合成参考T10。产率78％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1HNMR(400MHz,CDCl3)7.08(d,J＝2.2Hz,2H),6.55(t,J＝2.2Hz,1H),4.42(t,J＝4.6Hz,2H),3.94(t,J＝4.6Hz,2H),3.84(s,6H),2.76(s,3H),2.36(s,1H)；13C NMR(150MHz,CDCl3)δ170.2,162.4,161.5,161.2,134.4,121.2,104.7,103.7,66.8,61.1,55.6,17.5；HR-ESI-MS(positive mode)m/z:324.0894[M+H]+(Calcd for C15H18NO5S:324.0906),m/z:346.0707[M+Na]+(Calcd for C15H17NO5SNa:346.0725).
Ethyl 4-methyl-2-(3,4,5-trimethoxyphenyl)thiazole-5-carboxylate(S28):化合物S28的合成参考T7。产率79％，白色固体，展开体系PE/EA20:1(Rf＝0.5,PE/EA＝10:1),1H NMR(400MHz,CDCl3)7.17(s,2H),4.35(q,J＝7.1Hz,2H),3.92(s,6H),3.88(s,3H),2.74(s,3H),1.38(t,J＝7.1Hz,3H)；13C NMR(150MHz,CDCl3)δ169.6,162.2,160.9,153.6,140.7,128.4,121.5,104.0,61.2,60.9,56.3,17.5,14.3；HR-ESI-MS(positive mode)m/z:338.1048[M+H]+(Calcd for C16H20NO5S:338.1062),m/z:360.0868[M+Na]+(Calcd forC16H19NO5SNa:360.0882).
2-hydroxyethyl 4-methyl-2-(3,4,5-trimethoxyphenyl)thiazole-5-carboxylate(S29):化合物S29的合成参考T10。产率83％，白色固体，展开体系PE/EA1:1(Rf＝0.5,PE/EA＝1:1),1HNMR(400MHz,CDCl3)7.18(s,2H),4.44(t,J＝4.6Hz,2H),3.95-3.93(m,8H),3.90(s,3H),2.77(s,3H),2.12(s,1H)；13C NMR(150MHz,CDCl3)δ170.1,162.4,161.6,153.6,140.9,128.2,120.8,104.1,66.8,61.2,61.0,56.3,17.6；HR-ESI-MS(positive mode)m/z:354.1003[M+H]+(Calcd for C16H20NO6S:354.1011),m/z:376.0821[M+Na]+(Calcd for C16H19NO6SNa:376.0831).
2-((5-((3aS,4S,6aR)-2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl)pentanoyl)oxy)ethyl2-(4-methoxyphenyl)-5-methyloxazole-4-carboxylate(T58):在25mL的圆底烧瓶中加入D-生物素(D-Biotin)(244.3mg,1mmol),化合物3b(261.3mg,1mmol)，二环己基碳二亚胺(DCC)(412.6mg,2mmol)，4-二甲氨基吡啶(DMAP)(24.4mg,0.2mmol)和DMF(20mL)，在常温下搅拌24h，反应完毕后除去溶剂，经柱层析分离纯化，获得化合物T56。产率49％，白色固体，展开体系DCM/EtOH 30:1(Rf＝0.5,DCM/EtOH＝20:1),1HNMR(600MHz,DMSO-d6)7.90(d,J＝8.8Hz,2H),7.08(d,J＝8.8Hz,2H),5.55(d,J＝7.9Hz,2H),4.89(t,J＝5.6Hz,1H),4.26(t,J＝4.9Hz,2H),3.82(s,3H),3.69(q,J＝4.7Hz,2H),2.64(s,3H),1.71-1.69(m,4H),1.61-1.58(m,4H),1.50-1.47(m,2H)；13C NMR(150MHz,DMSO-d6)δ162.2,161.8,159.2,157.1,156.4,128.4,128.3,119.2,115.1,66.5,59.5,55.9,48.0,33.8,25.8,24.9,12.4；HR-ESI-MS(positive mode)m/z:504.1804[M+H]+(Calcd for C24H30N3O7S:504.1810),m/z:526.1622[M+Na]+(Calcd for C24H29N3O7SNa:526.1624).对照例1、对照化合物3b的合成
Ethyl 2-(4-methoxyphenyl)-5-methyloxazole-4-carboxylate(3b):在25mL的圆底烧瓶中加入4-甲氧基苯甲胺(274.4mg,2mmol)和DMF(10mL)，然后依次加入I2(304.6mg,1.2mmol)，乙酰乙酸乙酯(130.1mg,1mmol)，TBHP(180.2mg,2mmol)，醋酸铜(39.9mg,0.2mmol)；反应体系在常温下搅拌4小时。反应停止后加入10mL水淬灭反应，加入饱和硫代硫酸钠(20mL)除去未反应的碘，以乙酸乙酯萃取，有机相用饱和食盐水清洗，合并有机相，无水硫酸钠干燥并浓缩，粗品再经硅胶柱层析分离纯化(石油醚/乙酸乙酯＝10/1)得到目标化合物3b。产率87％，白色固体，展开体系PE/EA10:1(Rf＝0.6,PE/EA＝3:1),1HNMR(400MHz,CDCl3)8.00(d,J＝9.0Hz,2H),6.96(d,J＝9.0Hz,2H),4.43(q,J＝7.2Hz,2H),3.85(s,3H),2.67(s,3H),1.42(t,J＝7.1Hz,2H)；13C NMR(100MHz,CDCl3)δ162.6,161.6,159.8,155.6,128.6,128.3,119.4,114.1,61.0,55.4,14.4,12.2；HR-ESI-MS(positivemode)m/z:262.1068[M+H]+(Calcd for C14H16NO4:262.1079),m/z:284.0890[M+Na]+(Calcdfor C14H15NO4Na:284.0899).
以下通过实验例证明本发明制备的化合物的有益效果。
实验例1、化合物对miRNA-21的抑制活性分析
1、实验方法：
(1)实验材料与仪器
a.主要试剂：DMEM/高糖培养基、PBS缓冲液、EDTA-0.25％胰酶、青霉素链霉素(双抗)、BIOMYC-3Antibiotic Solution等购于Hyclone公司；胎牛血清购于Gibco公司；Coenzyme Asodium salt hydrate、D-Luciferin sodium salt购于Sigma公司；DMSO、G418购于Amresco公司。
b.主要仪器：CO2恒温细胞培养箱、超净工作台、全波长扫描式多功能读数仪、细胞操作台、光学倒置显微镜、压力蒸汽灭菌器、微型离心机、电子恒温水浴锅、水平摇床等。
(2)细胞模型的建立
通过荧光素酶报告基因构建miRNA-21小分子抑制剂的细胞筛选模型：化合物通过促进(或抑制)细胞内miRNA-21的表达，使转染的miRNA-21互补序列发生互补配对增加(或减少)，从而抑制(或增强)荧光素酶的表达，如图6所示。
(3)细胞培养
从液氮罐中取出冻存的HeLa-luciferase-miRNA-21细胞放进水浴锅中快速解冻，将细胞悬液吸入已提前配制好的培养基(DMEM高糖细胞培养基+10％胎牛血清+1％青霉素链霉素+1％ BIOMYC-3Antibiotic Solution+G418(300μg/mL))的培养皿中，然后放置5％CO2的37℃细胞培养箱中。
待细胞密度达到80-90％时，倒掉细胞培养基，用PBS缓冲液清洗细胞2次，倒掉PBS后加入1mL含有EDTA的0.25％胰酶，放入细胞培养箱中37℃消化3-5min，待观察到消化完全后加入新鲜细胞培养基，用吹打管轻柔反复吹打，将细胞从壁上吹打下来成细胞悬液，取100ul 1×104个细胞均匀接种到96孔板中，置于5％ CO2，37℃细胞培养箱中培养过夜。其中96孔板外围孔不用于化合物筛选，以排除边缘效应的影响。
(4)荧光素酶报告基因的检测
对96孔板细胞进行相应的加药处理24h后，吸去上清培养基用PBS缓冲液清洗，然后加入50μL cell lysis buffer裂解液并在摇床上震荡裂解15-20分钟后，取50μL裂解液到待测黑色96孔板中，再将提前解冻的荧光素酶底物加入到待测样品中(50μL/孔)，通过多功能读数仪检测每孔的荧光值。
(5)活性筛选
对Hela-miRNA-21稳定转染型细胞株进行复苏和传代培养；将对数期细胞接种于96孔板(1×104)中，待细胞贴壁，再在超净工作台内加入活性化合物(所有化合物的初筛浓度为10μM)。每个化合物三个复孔，并将96孔板放置细胞培养箱24小时，然后测定各个孔细胞裂解液的荧光信号强度：RFS(Relative Fluorescence Signal)，相对荧光强度大于前期已验证的miRNA-21抑制剂3b化合物则进行下一步实验。(RFS＝RFS sample/RFS control)
(6)细胞毒活性检测
采用AM-Blue细胞增殖与活性检测试剂盒来进行化合物对Hela-luciferase-miRNA-21细胞系活力的影响。
该试剂盒的检测原理：SunBio Am-Blue的主要成分是氧化型的靛青蓝色指示剂，该蓝色指示剂被还原之后会生成稳定的粉红色荧光物质。该荧光物质的激发光波长在530-560nm之间，发射光波长为590nM。增殖中的细胞其细胞内与细胞外相比处于还原状态，SunBio Am-Blue试剂的水溶性很好，比较容易进入细胞内部，最终在线粒体内被还原为粉红色荧光物质，然后释放到细胞外并溶于培养基中，使培养基从无荧光的靛青蓝变成有荧光的粉红色。通过用普通分光光度计或荧光光度计进行检测吸光度或荧光强度的变化从而分析细胞增殖的情况。将Hela-luciferase-miRNA-21稳定转染细胞接种100μL1×104个细胞于96孔板，设置8个浓度梯度：0，0.01μM，0.1μM，1μM，5μM，10μM，25μM，50μM，化合物处理24小时后，测定其荧光值，然后通过GraphPad Prism5软件计算所筛选化合物的EC50值。
2、实验结果：
本发明制备的化合物在活性筛选实验中的相对荧光强度如图1、2所示，以化合物3b为对照。
从图1、2看出，本发明制备的化合物T24、T30、T32、T35、T40、T2、T10、S11、S17、S20、S21、S25对miRNA-21生物合成具有明显的抑制活性，并且抑制活性高于3b。
我们进一步对上述高活性的化合物进行了Hela-luciferase-miRNA-21细胞毒活性检测试验，根据图3看出，S21的EC50达到了为0.093μM，其活性相比于3b(EC50＝1.38μM)提高了15.3倍，同样地，化合物T24、T30、T32、T35、T40、T2、T10、S11、S17、S20、S25的EC50也低于化合物3b，相比于化合物3b具有更高的活性。
实验例2、化合物与miRNA生物合成过程中TRBP相关结合蛋白对接的亲和力分析1、实验方法：
在我们之前的研究中，我们将7种已经报道的miRNA生物合成过程中TRBP相关结合蛋白(来源于PDB数据库，4种来源为哺乳动物和2种来源为果蝇基因以及1种来源于植物)分别与化合物3b进行了分子对接，结果发现只有来源于哺乳动物的两种蛋白晶体模型(4WYQ：Dicer-TRBP和3LLH：TRBP(dsRBD2))能够与3b形成活性口袋。
在动物细胞中TRBP作为Dicer伴侣，TRBP可通过影响Dicer和Ago2介导的miRNA成熟，进而影响癌细胞的恶性转移，TRBP被证实在RNA interference(RNAi)过程中也是不可或缺的；TRBP与Dicer或者Ago2相互作用并结合dsRNA组合形成诱导RISC复合体(RNAinduced silencing complex)发挥基因沉默功能，可加工Pre-miRNA形成成熟的miRNA。
为了进一步研究本发明化合物对miRNA的抑制作用，我们将这两种蛋白晶体模型分别与上述通过活性实验筛选得出的高活性小分子抑制剂，在计算机上进行分子亲和力对接。
(1)靶蛋白与仪器信息
读入的受体蛋白：4WYQ(Dicer-TRBP界面)、3LLH(TRBP的dsRBD2结构)、5N8M(与19bp siRNA结合的TRBP dsRBD 1和2复合物结构(配合物A))、5N8L(与19bp siRNA结合的TRBP dsRBD 1和2复合物结构(配合物B))、3ADL(TRBP2结构(植物源))、5NPA(果蝇dsRBD2结构)、5NPG(果蝇dsRBD1结构)；
受体蛋白来源于PDB数据库：https://www.rcsb.org/。
软件平台：Discovery Studio 4.5。
(2)实验操作
1)将从PDB数据库中下载得到的蛋白受体导入分子窗口，对蛋白受体赋予CHARMM立场，加氢，去除水分子，对不合理构象进行改正等。
2)在另一窗口中导入小分子，对小分子命名，排序，赋予ChARMM立场，以及能量最小化运算。
3)在蛋白受体中根据空腔选择可能的活性位点，共生成10组，取第一组位点为对接活性口袋。
4)用receptor-ligand interaction模块下的CDOCKER模块进行受体与小分子间的半柔性对接，单个分子构象集生成最多为10，分子动力学最大步数为1000。
5)对接完成后，对所有对接构象进行得分评价，选得分最高为最优构象，进行进一步的观察。
2、实验结果
本发明制备的活性化合物与Dicer-TRBP的蛋白晶体分子对接结果如图4所示。本发明制备的活性化合物与TRBP(dsRBD2)的蛋白晶体分子对接结果如图5所示。
本发明制备的化合物与两种蛋白的对接能量计算结构如表1所示。从小分子与蛋白上的氨基酸残基的结合度与构象结合能量来看，Dicer-TRBP蛋白上能够与抑制剂结合的氨基酸残基数量较多，并且最佳构象的对接能量比TRBP(dsRBD2)几乎低1倍(表1)，因此Dicer-TRBP interface蛋白模型中，亲和力远高于TRBP(dsRBD2)。通过计算机辅助分子对接能量计算可知，活性越高的化合物，其最佳构象能量值越低，说明化合物与靶蛋白的结合度越紧密。
从表1中看出，化合物T10、S21、S25与4WYQ蛋白对接的最佳构象能量值低于化合物3b与4WYQ蛋白的；化合物T10、T32、S21、S25与3LLH蛋白对接的最佳构象能量值低于化合物3b与3LLH蛋白的。也就是说，上述化合物与对应靶蛋白的结合度比3b更紧密。
表1本发明制备的活性化合物与蛋白的对接能量a
a最佳构象能量值；b总能量值评价函数为负，值越大，能量越小。
小分子通过与TRBP相关结合蛋白对接，能够抑制miRNA-21的生物合成，进而达到抑制恶性肿瘤的发生和阻断恶性肿瘤迁移的效果。而本发明制备的化合物，相比于对照化合物3b，与TRBP相关结合蛋白具有更紧密的亲和结合能力，对miRNA-21的生物合成具有更有强的抑制作用，因此，本发明的化合物对恶性肿瘤具有更优异的治疗效果。
综上所述，本发明提供了一种式Ⅰ所示的化合物或其药学上可接受的盐，其能够与miRNA生物合成过程中的相关结合蛋白紧密地结合，并且能够有效抑制miRNA-21的合成。本发明制备的活性化合物可用做miRNA-21抑制剂，进一步作为治疗恶性肿瘤的潜在药物。
Claims (4)
1.化合物或其药学上可接受的盐，其特征在于：所述化合物为：
2.权利要求1所述的化合物或其药学上可接受的盐在制备miRNA-21生物合成抑制剂的用途。
3.根据权利要求2所述的用途，其特征在于：所述抑制剂为治疗肿瘤的药物。
4.根据权利要求3所述的用途，其特征在于：所述抑制剂为治疗恶性肿瘤的药物。
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| CN107162982A (zh) * | 2017-06-19 | 2017-09-15 | 广东药科大学 | 一类具有抗癌活性的咪唑类化合物及其衍生物 |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
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| CN107162982A (zh) * | 2017-06-19 | 2017-09-15 | 广东药科大学 | 一类具有抗癌活性的咪唑类化合物及其衍生物 |
Non-Patent Citations (1)
| Title |
|---|
| Discovery of a Novel Small-Molecule Inhibitor Disrupting TRBP− Dicer Interaction against Hepatocellular Carcinoma via the Modulation of microRNA Biogenesis;Ting Peng，等;《J. Med. Chem.》;第第65卷卷(第第16期期);第11010–11033页 * |
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