本发明属于医药化工领域，具体涉及一种烟酰二氢吡唑类化合物及其药物用途。
背景技术：
：肿瘤是机体在各种致瘤因素的作用下，局部组织的细胞异常增生而形成的新生物，常表现为局部肿块。肿瘤细胞具有异常的形态、代谢和功能。它生长旺盛且常呈持续性生长。近年来，肿瘤尤其是恶性肿瘤已经成为严重威胁人类健康的疾病之一。在全世界50多亿人口中平均每年死于恶性肿瘤者达690万人，新发病例为870万例，且数字在不断增长中。化疗、放疗、手术、生物治疗和中西药治疗等方法已经成为治疗肿瘤的最有效的方法。随着肿瘤的药物的发展，新型的抗肿瘤药物在不断地更新换代，在提高肿瘤患者的治愈率，延长患者生存时间、延缓疾病等方面发挥了巨大的作用。多年来，人们不断地开发新的抗肿瘤药物，并通过结构改造，试图开发出一系列地新的抗肿瘤药物衍生物。药物的化学结构改造是基于药物原有的基本化学结构，对其中某些官能团进行化学改造或者将两个或两个以上的具有相同或不同活性的化学结构进行结合，得到这类药物的新的衍生物。通过结构可能会改变原有的理化性质和药理活性，在临床应用上有极其重要的作用。QinYa-Juan等设计了一类具有抑制微管聚合活性的吡唑啉类化合物(Design,synthesisandbiologicalevaluationofnovelpyrazoline-containingderivativesaspotentialtubulinassemblinginhibitors.EuropeanJournalofMedicinalChemistry94(2015)447-457)，体外活性实验显示具有一定的抑制肿瘤细胞增殖活性。技术实现要素：本发明在现有技术基础上，对吡唑啉类化合物进行了结构改造，得到了一类新的具有良好抗肿瘤活性的烟酰二氢吡唑类化合物及药学上可成的盐，具有如下结构式：其中，R1为H、Br或OCH3；R2为H、F、Cl、Br或OCH3；R3为H、Cl、OCH3、CH3。上述化合物及药学上可成的盐，优选R2为H、4-F、4-Cl、4-Br、3-OCH3或4-OCH3、3,4-OCH3或3,4,5-OCH3，R3为H、2-Cl、6-Cl、6-CH3、6-OCH3。本发明的优选化合物如下：R1为H，R2为H、3,4,5-OCH3，R3为H、2-Cl、6-Cl、6-CH3、6-OCH3；或者，R1为Br，R2为H、4-F、4-Cl、4-Br、4-OCH3、3,4-OCH3或3,4,5-OCH3，R3为H、6-OCH3；或者，R1为OCH3，R2为H、4-F、4-Cl、4-Br、3-OCH3、4-OCH3、3,4-OCH3或3,4,5-OCH3，R3为H、6-CH3或6-OCH3。更优选化合物如下：本发明所述化合物及药学上可成的盐可以以单一药物的形式被给药或可以与其它药物联合给药。本发明的化合物的可药用盐包括各种无机或有机酸盐如盐酸盐、氢溴酸盐、磷酸盐、硫酸盐、柠檬酸盐、乳酸盐、酒石酸盐、马来酸盐、延胡索酸盐、扁桃酸盐和草酸盐；各种无机或有机碱盐如氢氧化钠、三羟甲基氨基甲烷和N-甲基-葡萄糖胺。本发明的还提供了本发明所述烟酰二氢吡唑类化合物及药学上可成的盐在制备抗肿瘤药物中的应用。所述肿瘤优选乳腺癌、肺癌、肝癌等。本发明还提供了一种抗肿瘤药物组合物，包括本发明所述烟酰二氢吡唑类化合物及药学上可成的盐以及药用载体。所述抗肿瘤药物组合物包括作为活性成分的本发明的化合物或其可药用盐及可药用载体。较佳的，本发明的药物组合物有0.1-99.9％重量百分比的作为活性成分的本发明的化合物或其可药用盐。“可药用载体”包括但不限于：离子交换材料、氧化铝、硬脂酸铝、卵磷脂、自乳化药物传递系统(SEDDS)如d-维生素E聚乙二醇1000琥珀酸酯、吐温或其他类似聚合介质等药物制剂用的表面活性剂、血清蛋白如人血清白蛋白、缓冲物质如磷酸盐、氨基乙酸、山梨酸、山梨酸钾、饱和植物脂肪酸部分甘油酯混合、水、盐、电解质如硫酸盐精蛋白、磷酸氢二钠、磷酸氢钾、氯化钠、锌盐、硅胶、硅酸镁等。聚乙烯吡咯酮、纤维素物质、聚乙烯醇、羧甲基纤维素钠、聚丙烯酸酯、乙烯-聚氧乙烯-嵌段聚合物和羊毛脂、环糊精如α-、β-、γ-环糊精或其经化学修饰的衍生物如2-和3-羟丙基-β-环糊精等羟烷基环糊精或其他可溶性衍生物等均可用于促进本发明的化合物、其药用盐或前药的药物传递。其他可药用辅料如填充剂(如无水乳糖、淀粉、乳糖珠粒和葡萄糖)、粘合剂(如微晶纤维素)、崩解剂(如交联羧甲基淀粉钠、交联羧甲基纤维素钠、低取代羟丙基纤维素和交联PVP)、润滑剂(如硬脂酸镁)、吸收促进剂、香味剂、甜味剂、稀释剂、赋形剂、润湿剂、溶剂、增溶剂和着色剂等也可加入本发明的药物组合物中。上述本发明的化合物或其可药用盐以及药物组合物可通过肠道或者非肠道途径给药。非肠道给药制剂包括注射剂、霜剂、软膏剂、贴剂、喷雾剂等。给药途径包括皮下、皮内、动脉内、静脉内、肌内、关节内、滑液内、胸骨内、鞘内、病灶内、颅内注射或输注，或者，口服、局部、直肠、经鼻、经颊、阴道、舌下、皮内、粘膜、气管、尿道给药，或者通过吸入气雾或植入蓄积或者针刺方式给药。上述本发明的化合物或其可药用盐以及药物组合物的治疗有效量为0.001-100mg/kg/d之间，可用于相关疾病的单一用药或联合用药治疗，为本领域技术人员能够理解的范围。本发明考察了本发明所述烟酰二氢吡唑类化合物体外对人肝癌细胞(HepG2)，人乳腺癌细胞(MCF-7)以及人肺腺癌细胞(A549)的抑制作用以及对人肾上皮细胞(293T)的细胞毒性结果显示本发明所述烟酰二氢吡唑类化合物具有良好的抗肿瘤作用，对正常细胞的毒性低，是安全、有效、低毒的候选抗肿瘤药物。具体实施方式以下通过实施例说明本发明的具体步骤，但不受实施例限制。在本发明中所使用的术语，除非另有说明，一般具有本领域普通技术人员通常理解的含义。下面结合具体实施例并参照数据进一步详细描述本发明。应理解，这些实施例只是为了举例说明本发明，而非以任何方式限制本发明的范围。在以下实施例中，未详细描述的各种过程和方法是本领域中公知的常规方法。实施例1本发明所述烟酰二氢吡唑类化合物的制备本发明所述烟酰二氢吡唑类化合物及药学上可成的盐可采用如下方法制备而成。具体的制备方法为：冰浴条件下，向反应容器中依次加入POCl3(20mL)与DMF(40mL)，化合物1(35mmol)，反应2h，倒入冰水中用NaOH调节PH至9.0，用乙酸乙酯萃取干燥得化合物2。在冰浴下，向反应容器中加入化合物2(25mmol)，THF(15mL)，NaH(62.5mmol)，搅拌15min，再加入CH3I(33mmol)室温反应24h。TLC跟踪，充分反应后用乙酸乙酯萃取干燥得化合物3。向反应容器中加入化合物3(2mmol)与苯乙酮(2mmol)，无水乙醇(20mL)以及KOH(6mmol)，室温反应24h，过滤后乙醇与二氯甲烷重结晶得化合物4。依次向反应容器中加入无水乙醇溶液(5mL)，化合物4(1mmol)，水合肼(2mmol)，回流2h，冷却转移至-20℃过夜，固体过滤用石油醚洗净得化合物5。向反应容器中加入取代烟酸(2mmol)、EDC·HCl(3mmol)、HOBt(1.2mmol)以及二氯甲烷(5mL)，在惰性气体保护下加入化合物5(1mmol)，反应24h后，过滤并用乙醇与水洗净，粗产物在乙醇与二氯甲烷中重结晶得到一系列本发明所述烟酰二氢吡唑类化合物(化合物6)，化合物如表1所示。表1化合物R1R2R3ZYL-1(式Ⅰ)HHHZYL-2(式Ⅰ)HH6-ClZYL-3(式Ⅱ)HHHZYL-4(式Ⅰ)HH6-CH3ZYL-5(式Ⅰ)HH2-ClZYL-6(式Ⅰ)HH6-OCH3ZYL-7(式Ⅰ)H3,4,5-OCH36-CH3ZYL-8(式Ⅰ)Br4-ClHZYL-9(式Ⅰ)Br3,4,5-OCH3HZYL-10(式Ⅱ)Br3,4-OCH3HZYL-11(式Ⅰ)Br4-FHZYL-12(式Ⅰ)Br4-BrHZYL-13(式Ⅰ)Br3,4-OCH3HZYL-14(式Ⅰ)Br4-OCH3HZYL-15(式Ⅰ)BrH6-OCH3ZYL-16(式Ⅰ)OCH3HHZYL-17(式Ⅰ)OCH3H6-OCH3ZYL-18(式Ⅰ)OCH34-F6-OCH3ZYL-19(式Ⅰ)OCH34-Cl6-OCH3ZYL-20(式Ⅰ)OCH34-Br6-OCH3ZYL-21(式Ⅰ)OCH34-OCH36-OCH3ZYL-22(式Ⅰ)OCH33,4,5-OCH36-OCH3ZYL-23(式Ⅰ)OCH33-OCH3HZYL-24(式Ⅰ)OCH33-OCH36-CH3ZYL-25(式Ⅰ)OCH33-OCH36-OCH3上述化合物结构鉴定数据如下：(5-(1-methyl-1H-indol-3-yl)-3-phenyl-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-1)Redcrystal.Yield:29.8％.m.p.165～166℃.1HNMR(DMSO-d6,400MHz)δ:9.00(s,1H),8.69(dd,J＝4.7,1.3Hz,1H),8.20(d,J＝7.8Hz,1H),7.78(dd,J＝6.5,2.9Hz,2H),7.59–7.34(m,7H),7.15(t,J＝7.6Hz,1H),7.01(t,J＝7.5Hz,1H),6.07(dd,J＝11.7,4.7Hz,1H),3.94(dd,J＝18.0,11.8Hz,1H),3.75(s,3H),3.35(dd,J＝18.0,4.8Hz,1H).MS(ESI)m/z:318.16(C24H20N4O,[M+H]+).Anal.CalcdforC24H20N4O:C,75.77；H,5.30；N,14.73.Found:C,75.87；H,5.31；N,14.76。(6-chloropyridin-3-yl)(5-(1-methyl-1H-indol-3-yl)-3-phenyl-4,5-dihydro-1H-pyrazol-1-yl)methanone(ZYL-2)Lightyellowpowder.Yield:22.1％.m.p.129～130℃.1HNMR(DMSO-d6,400MHz)δ:8.85(s,1H),8.27(d,J＝6.3Hz,1H),7.87–7.72(m,2H),7.65(d,J＝8.3Hz,1H),7.59–7.36(m,6H),7.15(t,J＝7.6Hz,1H),7.01(t,J＝7.6Hz,1H),6.06(dd,J＝11.7,4.7Hz,1H),3.95(dd,J＝18.1,11.7Hz,1H),3.75(s,3H),3.36(dd,J＝18.0,4.8Hz,1H).MS(ESI)m/z:415.12(C24H19ClN4O,[M+H]+).Anal.CalcdforC24H19ClN4O:C,69.48；H,4.62；N,13.50.Found:C,69.44；H,4.62；N,13.52。(5-(1-methyl-1H-indol-3-yl)-3-phenyl-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanone(ZYL-3)Lightpurplepowder.Yield:23.0％.m.p.175～176℃.1HNMR(DMSO-d6,400MHz)δ:8.72(d,J＝5.6Hz,2H),7.86–7.76(m,2H),7.70(d,J＝5.6Hz,2H),7.59–7.36(m,6H),7.15(t,J＝7.5Hz,1H),7.01(t,J＝7.5Hz,1H),6.05(dd,J＝11.7,4.6Hz,1H),3.95(dd,J＝18.1,11.8Hz,1H),3.76(s,3H),3.44–3.31(m,1H).MS(ESI)m/z:381.16(C24H20N4O,[M+H]+).Anal.CalcdforC24H20N4O:C,75.77；H,5.30；N,14.73.Found:C,75.85；H,5.28；N,14.74。(5-(1-methyl-1H-indol-3-yl)-3-phenyl-4,5-dihydro-1H-pyrazol-1-yl)(6-methylpyridin-3-yl)methanone(ZYL-4)Lightyellowpowder.Yield:21.2％.m.p.158～160℃.1HNMR(DMSO-d6,400MHz)δ:8.90(s,1H),8.10(d,J＝6.6Hz,1H),7.79(dd,J＝6.6,2.9Hz,2H),7.58–7.33(m,7H),7.15(t,J＝7.9Hz,1H),7.01(t,J＝7.7Hz,1H),6.06(dd,J＝11.7,4.8Hz,1H),3.95(dd,J＝18.0,11.8Hz,1H),3.76(s,3H),3.35(dd,J＝17.9,4.9Hz,1H),2.54(s,3H).MS(ESI)m/z:395.18(C25H22N4O,[M+H]+).Anal.CalcdforC25H22N4O:C,76.12；H,5.62；N,14.20.Found:C,75.89；H,5.60；N,14.21。(2-chloropyridin-3-yl)(5-(1-methyl-1H-indol-3-yl)-3-phenyl-4,5-dihydro-1H-pyrazol-1-yl)methanone(ZYL-5)Lightyellowsolid.Yield:48.2％.m.p.191～193℃.1HNMR(DMSO-d6,400MHz)δ:8.51(dd,J＝4.8,1.9Hz,1H),7.89(dd,J＝7.5,1.9Hz,1H),7.65(d,J＝6.3Hz,2H),7.54(dd,J＝7.5,4.8Hz,1H),7.51–7.37(m,6H),7.18(t,J＝7.3Hz,1H),7.02(t,J＝7.3Hz,1H),6.04(dd,J＝11.7,4.7Hz,1H),4.00(dd,J＝18.2,11.7Hz,1H),3.79(s,3H),3.38(dd,J＝18.2,4.7Hz,1H).MS(ESI)m/z:415.12(C24H19ClN4O,[M+H]+).Anal.CalcdforC24H19ClN4O:C,69.48；H,4.62；N,13.50.Found:C,69.67；H,4.63；N,13.49。(6-methoxypyridin-3-yl)(5-(1-methyl-1H-indol-3-yl)-3-phenyl-4,5-dihydro-1H-pyrazol-1-yl)methanone(ZYL-6)Orangesolid.Yield:36.4％.m.p.174～176℃.1HNMR(DMSO-d6,400MHz)δ:8.75(s,1H),8.18(dd,J＝8.7,2.2Hz,1H),7.81(dd,J＝6.5,3.0Hz,2H),7.58–7.46(m,3H),7.42(d,J＝8.5Hz,2H),7.37(s,1H),7.14(t,J＝7.8Hz,1H),6.99(t,J＝7.4Hz,1H),6.91(d,J＝8.7Hz,1H),6.04(dd,J＝11.8,4.8Hz,1H),4.07–3.84(m,4H),3.75(s,3H),3.33–3.30(m,1H).MS(ESI)m/z:411.17(C25H22N4O2,[M+H]+).Anal.CalcdforC25H22N4O2:C,73.15；H,5.40；N,13.65.Found:C,73.34；H,5.39；N,13.64。(5-(1-methyl-1H-indol-3-yl)-3-(3,4,5-trimethoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(6-methylpyridin-3-yl)methanone(ZYL-7)Browncrystal.Yield:52.1％.m.p.217～219℃.1HNMR(DMSO-d6,400MHz)δ:8.91(s,1H),8.16(d,J＝7.6Hz,1H),7.53–7.30(m,4H),7.15(t,J＝7.6Hz,1H),7.06(s,2H),7.02(t,J＝7.5Hz,1H),6.06(dd,J＝11.7,4.6Hz,1H),3.90(dd,J＝18.1,11.7Hz,1H),3.81(s,6H),3.75(s,3H),3.71(s,3H),3.40(dd,J＝18.1,4.7Hz,1H),2.53(s,3H).MS(ESI)m/z:485.21(C28H28N4O4,[M+H]+).Anal.CalcdforC28H28N4O4:C,69.41；H,5.82；N,11.56.Found:C,69.19；H,5.83；N,11.53。(5-(5-bromo-1-methyl-1H-indol-3-yl)-3-(4-chlorophenyl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-8)Whitesolid.Yield:21.4％.m.p.202～203℃.1HNMR(DMSO-d6,400MHz)δ:9.00(s,1H),8.70(d,J＝3.5Hz,1H),8.19(d,J＝7.8Hz,1H),7.78(d,J＝8.5Hz,2H),7.71(s,1H),7.55(d,J＝8.6Hz,3H),7.46(s,1H),7.42(d,J＝8.7Hz,1H),7.28(dd,J＝8.7,1.7Hz,1H),6.08(dd,J＝11.6,4.6Hz,1H),3.94(dd,J＝18.1,11.7Hz,1H),3.74(s,3H),3.35(dd,J＝18.1,4.7Hz,1H).MS(ESI)m/z:493.04(C24H18BrClN4O,[M+H]+).Anal.CalcdforC24H18BrClN4O:C,58.38；H,3.67；N,11.35.Found:C,58.46；H,3.66；N,11.37。(5-(5-bromo-1-methyl-1H-indol-3-yl)-3-(3,4,5-trimethoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-9)Whitepowder.Yield:19.2％.m.p.194～195℃.1HNMR(DMSO-d6,400MHz)δ:9.03(s,1H),8.70(dd,J＝4.7,1.3Hz,1H),8.24(d,J＝7.6Hz,1H),7.75(s,1H),7.53(dd,J＝7.7,5.0Hz,1H),7.45–7.38(m,2H),7.29(dd,J＝8.7,1.8Hz,1H),7.05(s,2H),6.09(dd,J＝11.4,4.2Hz,1H),3.92(dd,J＝18.0,11.5Hz,1H),3.81(s,6H),3.73(d,J＝12.5Hz,6H),3.41(dd,J＝18.0,4.4Hz,1H).MS(ESI)m/z:549.11(C27H25BrN4O4,[M+H]+).Anal.CalcdforC27H25BrN4O4:C,59.02；H,4.59；N,10.20.Found:C,59.19；H,4.58；N,10.21。(5-(5-bromo-1-methyl-1H-indol-3-yl)-3-(3,4-dimethoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanone(ZYL-10)Graypowder.Yield:18.9％.m.p.168～169℃.1HNMR(DMSO-d6,400MHz)δ:8.74(d,J＝5.7Hz,2H),7.75(d,J＝5.8Hz,2H),7.71(s,1H),7.42(d,J＝7.3Hz,2H),7.40–7.23(m,3H),7.04(d,J＝8.5Hz,1H),6.05(dd,J＝11.4,4.2Hz,1H),3.90(dd,J＝18.0,11.5Hz,1H),3.81(s,3H),3.78(s,3H),3.75(s,3H),3.40–3.32(m,1H).MS(ESI)m/z:519.10(C26H23BrN4O3,[M+H]+).Anal.CalcdforC26H23BrN4O3:C,60.12；H,4.46；N,10.79.Found:C,60.02；H,4.47；N,10.81。(5-(5-bromo-1-methyl-1H-indol-3-yl)-3-(4-fluorophenyl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-11)Lightyellowpowder.Yield:19.9％.m.p.166～168℃.1HNMR(DMSO-d6,400MHz)δ:8.99(s,1H),8.69(dd,J＝4.8,1.6Hz,1H),8.20(d,J＝7.9Hz,1H),7.83(dd,J＝8.8,5.5Hz,2H),7.71(s,1H),7.53(dd,J＝7.8,4.9Hz,1H),7.49–7.39(m,2H),7.39–7.24(m,3H),6.07(dd,J＝11.6,4.6Hz,1H),3.94(dd,J＝18.1,11.6Hz,1H),3.75(s,3H),3.44–3.31(m,1H).13CNMR(DMSO-d6,101MHz)δ:165.12,163.91,162.65,156.24,151.67,150.39,137.42,136.04,131.10,129.67,129.27,128.02,127.32,124.24,123.55,121.38,116.57,116.35,114.46,112.67,112.27,54.27,41.10,33.02.MS(ESI)m/z:477.06(C24H18BrFN4O,[M+H]+).Anal.CalcdforC24H18BrFN4O:C,60.39；H,3.80；N,11.74.Found:C,60.50；H,3.79；N,11.74。(5-(5-bromo-1-methyl-1H-indol-3-yl)-3-(4-bromophenyl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-12)Whitepowder.Yield:26.5％.m.p.213～214℃.1HNMR(DMSO-d6,400MHz)δ:8.99(s,1H),8.70(dd,J＝4.7,1.2Hz,1H),8.19(d,J＝7.8Hz,1H),7.89–7.64(m,5H),7.61–7.52(m,1H),7.45(dd,J＝26.9,16.2Hz,2H),7.28(dd,J＝8.7,1.8Hz,1H),6.08(dd,J＝11.6,4.6Hz,1H),3.94(dd,J＝18.1,11.7Hz,1H),3.74(s,3H),3.36–3.28(m,1H).MS(ESI)m/z:536.98(C24H18Br2N4O,[M+H]+).Anal.CalcdforC24H18Br2N4O:C,53.56；H,3.37；N,10.41.Found:C,53.44；H,3.38；N,10.43。(5-(5-bromo-1-methyl-1H-indol-3-yl)-3-(3,4-dimethoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-13)Pinkpowder.Yield:32.3％.m.p.112～114℃.1HNMR(DMSO-d6,400MHz)δ:9.02(s,1H),8.69(d,J＝4.1Hz,1H),8.22(d,J＝7.6Hz,1H),7.72(s,1H),7.53(dd,J＝7.7,5.0Hz,1H),7.42(d,J＝8.4Hz,2H),7.39–7.24(m,3H),7.05(d,J＝8.4Hz,1H),6.05(dd,J＝11.4,4.2Hz,1H),3.91(dd,J＝18.0,11.5Hz,1H),3.81(s,3H),3.78(s,3H),3.75(s,3H),3.39–3.31(m,1H).MS(ESI)m/z:519.10(C26H23BrN4O3,[M+H]+).Anal.CalcdforC26H23BrN4O3:C,60.12；H,4.46；N,10.79.Found:C,59.99；H,4.47；N,10.78。(5-(5-bromo-1-methyl-1H-indol-3-yl)-3-(4-methoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-14)Whitecrystal.Yield:25.4％.m.p.180～181℃.1HNMR(DMSO-d6,400MHz)δ:9.00(s,1H),8.69(dd,J＝4.8,1.5Hz,1H),8.20(d,J＝7.9Hz,1H),7.79–7.64(m,3H),7.53(dd,J＝7.8,4.9Hz,1H),7.48–7.38(m,2H),7.27(dd,J＝8.7,1.8Hz,1H),7.03(d,J＝8.9Hz,2H),6.04(dd,J＝11.5,4.4Hz,1H),3.91(dd,J＝18.0,11.6Hz,1H),3.81(s,3H),3.74(s,3H),3.34–3.29(m,1H).MS(ESI)m/z:489.08(C25H21BrN4O2,[M+H]+).Anal.CalcdforC25H21BrN4O2:C,61.36；H,4.33；N,11.45.Found:C,61.49；H,4.32；N,11.43。(5-(5-bromo-1-methyl-1H-indol-3-yl)-3-phenyl-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-15)Orangecrystal.Yield:40.1％.m.p.185～186℃.1HNMR(DMSO-d6,400MHz)δ:9.00(s,1H),8.70(dd,J＝4.8,1.5Hz,1H),8.20(d,J＝7.8Hz,1H),7.87–7.73(m,2H),7.71(s,1H),7.64–7.38(m,6H),7.28(dd,J＝8.7,1.8Hz,1H),6.07(dd,J＝11.6,4.6Hz,1H),3.95(dd,J＝18.0,11.6Hz,1H),3.75(s,3H),3.40–3.30(m,1H).MS(ESI)m/z:459.07(C24H19BrN4O,[M+H]+).Anal.CalcdforC24H19BrN4O:C,62.76；H,4.17；N,12.20.Found:C,62.88；H,4.18；N,12.21。(5-(5-methoxy-1-methyl-1H-indol-3-yl)-3-phenyl-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-16)Pinkpowder.Yield:24.3％.m.p.134～136℃.1HNMR(DMSO-d6,400MHz)δ:8.99(s,1H),8.68(dd,J＝4.8,1.6Hz,1H),8.19(d,J＝7.8Hz,1H),7.80(dd,J＝6.5,3.0Hz,2H),7.63–7.44(m,4H),7.44–7.29(m,2H),6.88(d,J＝1.9Hz,1H),6.80(dd,J＝8.9,2.4Hz,1H),6.05(dd,J＝11.6,4.1Hz,1H),3.94(dd,J＝18.0,11.7Hz,1H),3.71(s,3H),3.56(s,3H),3.35–3.31(m,1H).MS(ESI)m/z:411.17(C25H22N4O2,[M+H]+).Anal.CalcdforC25H22N4O2:C,75.77；H,5.30；N,14.73.Found:C,75.89；H,5.30；N,14.68。(5-(5-methoxy-1-methyl-1H-indol-3-yl)-3-phenyl-4,5-dihydro-1H-pyrazol-1-yl)(6-methoxypyridin-3-yl)methanone(ZYL-17)Whitesolid.Yield:17.3％.m.p.148～150℃.1HNMR(DMSO-d6,400MHz)δ:8.76(s,1H),8.19(dd,J＝8.7,2.2Hz,1H),7.92–7.77(m,2H),7.59–7.42(m,3H),7.31(d,J＝8.6Hz,2H),6.92(d,J＝8.7Hz,1H),6.86(d,J＝2.3Hz,1H),6.78(dd,J＝8.8,2.4Hz,1H),6.02(dd,J＝11.7,4.3Hz,1H),4.04–3.82(m,4H),3.71(s,3H),3.55(s,3H),3.32(dd,J＝18.0,4.3Hz,1H).MS(ESI)m/z:441.18(C26H24N4O3,[M+H]+).Anal.CalcdforC26H24N4O3:C,70.89；H,5.49；N,12.72.Found:C,70.91；H,5.48；N,12.74。(3-(4-fluorophenyl)-5-(5-methoxy-1-methyl-1H-indol-3-yl)-4,5-dihydro-1H-pyrazol-1-yl)(6-methoxypyridin-3-yl)methanone(ZYL-18)Brownsolid.Yield:14.7％.m.p.123～126℃.1HNMR(DMSO-d6,400MHz)δ:8.76(s,1H),8.19(d,J＝8.7Hz,1H),7.89(dd,J＝8.7,5.5Hz,2H),7.40–7.28(m,4H),6.92(d,J＝8.7Hz,1H),6.87(d,J＝2.1Hz,1H),6.79(dd,J＝8.8,2.3Hz,1H),6.03(dd,J＝11.7,4.3Hz,1H),3.97–3.85(m,4H),3.71(s,3H),3.58(s,3H),3.33(dd,J＝18.0,4.3Hz,1H).MS(ESI)m/z:459.18(C26H23FN4O3,[M+H]+).Anal.CalcdforC26H23FN4O3:C,68.11；H,5.06；N,12.22.Found:C,68.29；H,5.07；N,12.26。(3-(4-chlorophenyl)-5-(5-methoxy-1-methyl-1H-indol-3-yl)-4,5-dihydro-1H-pyrazol-1-yl)(6-methoxypyridin-3-yl)methanone(ZYL-19)Yellowsolid.Yield:18.6％.m.p.157～159℃.1HNMR(DMSO-d6,400MHz)δ:8.75(s,1H),8.17(dd,J＝8.6,2.1Hz,1H),7.84(d,J＝8.6Hz,2H),7.57(d,J＝8.6Hz,2H),7.38–7.29(m,2H),6.92(d,J＝8.7Hz,1H),6.86(d,J＝2.1Hz,1H),6.80(dd,J＝8.9,2.3Hz,1H),6.03(dd,J＝11.7,4.4Hz,1H),4.00–3.84(m,4H),3.71(s,3H),3.59(s,3H),3.32(dd,J＝18.1,4.4Hz,1H).MS(ESI)m/z:475.15(C26H23ClN4O3,[M+H]+).Anal.CalcdforC26H23ClN4O3:C,65.75；H,4.88；N,11.80.Found:C,65.81；H,4.88；N,11.78。(3-(4-bromophenyl)-5-(5-methoxy-1-methyl-1H-indol-3-yl)-4,5-dihydro-1H-pyrazol-1-yl)(6-methoxypyridin-3-yl)methanone(ZYL-20)Orangecrystal.Yield:21.7％.m.p.163～164℃.1HNMR(DMSO-d6,400MHz)δ:8.75(s,1H),8.17(dd,J＝8.7,2.0Hz,1H),7.74(dd,J＝23.5,8.6Hz,4H),7.37–7.29(m,2H),6.92(d,J＝8.7Hz,1H),6.86(d,J＝2.1Hz,1H),6.80(dd,J＝8.8,2.3Hz,1H),6.03(dd,J＝11.7,4.4Hz,1H),3.96–3.81(m,4H),3.71(s,3H),3.59(s,3H),3.31(dd,J＝18.1,4.5Hz,1H).13CNMR(DMSO-d6,101MHz)δ:165.12,163.37,155.70,153.79,149.49,140.93,132.75,132.40,130.84,129.09,128.68,125.56,124.48,124.34,114.08,111.48,111.34,110.12,101.01,55.52,54.91,54.12,32.91.MS(ESI)m/z:519.10(C26H23BrN4O3,[M+H]+).Anal.CalcdforC26H23BrN4O3:C,60.12；H,4.46；N,10.79.Found:C,60.19；H,4.47；N,10.78。(5-(5-methoxy-1-methyl-1H-indol-3-yl)-3-(4-methoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(6-methoxypyridin-3-yl)methanone(ZYL-21)Lightyellowsolid.Yield:17.8％.m.p.90～93℃.1HNMR(DMSO-d6,400MHz)δ:8.76(s,1H),8.23–8.17(m,1H),7.80–7.73(m,4H),7.32(d,J＝8.6Hz,2H),6.92(d,J＝8.7Hz,1H),6.87(d,J＝2.1Hz,1H),6.79(dd,J＝8.9,2.3Hz,1H),6.00(dd,J＝11.5,4.2Hz,1H),3.92–3.79(m,7H),3.71(s,3H),3.57(s,3H),3.29(dd,J＝18.1,4.2Hz,1H).MS(ESI)m/z:471.20(C27H26N4O4,[M+H]+).Anal.CalcdforC27H26N4O4:C,68.92；H,5.57；N,11.91.Found:C,69.04；H,5.56；N,11.94。(5-(5-methoxy-1-methyl-1H-indol-3-yl)-3-(3,4,5-trimethoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(6-methoxypyridin-3-yl)methanone(ZYL-22)Yellowcrystal.Yield:18.9％.m.p.175～177℃.1HNMR(DMSO-d6,400MHz)δ:8.71(s,1H),8.23(d,J＝8.2Hz,1H),7.37–7.23(m,2H),7.09(s,2H),6.93(d,J＝9.1Hz,2H),6.80(dd,J＝8.9,2.3Hz,1H),6.03(dd,J＝11.6,4.0Hz,1H),3.92(s,3H),3.90–3.76(m,7H),3.72(d,J＝2.2Hz,6H),3.61(s,3H),3.43–3.38(m,1H).MS(ESI)m/z:531.22(C29H30N4O6,[M+H]+).Anal.CalcdforC29H30N4O6:C,65.65；H,5.70；N,10.56.Found:C,65.75；H,5.71；N,10.58。(5-(5-methoxy-1-methyl-1H-indol-3-yl)-3-(3-methoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-3-yl)methanone(ZYL-23)Orangepowder.Yield:21.2％.m.p.145～148℃.1HNMR(DMSO-d6,400MHz)δ:8.99(s,1H),8.69(dd,J＝4.8,1.4Hz,1H),8.20(d,J＝7.8Hz,1H),7.54(dd,J＝7.7,4.9Hz,1H),7.48–7.26(m,5H),7.22–7.02(m,1H),6.89(d,J＝1.5Hz,1H),6.81(dd,J＝8.9,2.3Hz,1H),6.05(dd,J＝11.6,4.1Hz,1H),3.93(dd,J＝18.0,11.7Hz,1H),3.79(s,3H),3.72(s,3H),3.59(s,3H),3.47–3.27(m,1H).13CNMR(DMSO-d6,101MHz)δ:163.73,159.94,157.01,153.82,151.63,150.41,137.46,132.82,132.79,131.19,130.56,128.70,125.55,123.51,119.54,116.54,113.95,112.64,111.53,111.36,101.03,55.68,55.50,54.72,32.92.MS(ESI)m/z:441.18(C26H24N4O3,[M+H]+).Anal.CalcdforC26H24N4O3:C,70.89；H,5.49；N,12.72.Found:C,70.86；H,5.51；N,12.75。(5-(5-methoxy-1-methyl-1H-indol-3-yl)-3-(3-methoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(6-methylpyridin-3-yl)methanone(ZYL-24)Yellowsolid.Yield:36.4％.m.p.138～139℃.1HNMR(DMSO-d6,400MHz)δ:8.89(s,1H),8.10(d,J＝7.4Hz,1H),7.42–7.27(m,6H),7.08(s,1H),6.88(d,J＝1.8Hz,1H),6.79(dd,J＝8.9,2.3Hz,1H),6.02(dd,J＝11.6,4.1Hz,1H),3.90(dd,J＝18.0,11.8Hz,1H),3.79(s,3H),3.71(s,3H),3.58(s,3H),3.32–3.28(m,1H),2.53(s,3H).MS(ESI)m/z:455.20(C27H26N4O3,[M+H]+).Anal.CalcdforC27H26N4O3:C,71.35；H,5.77；N,12.33.Found:C,71.44；H,5.78；N,12.33。(5-(5-methoxy-1-methyl-1H-indol-3-yl)-3-(3-methoxyphenyl)-4,5-dihydro-1H-pyrazol-1-yl)(6-methoxypyridin-3-yl)methanone(ZYL-25)Graysolid.Yield:25.5％.m.p.133～135℃.1HNMR(DMSO-d6,400MHz)δ:8.75(s,1H),8.20(d,J＝8.4Hz,1H),7.43(d,J＝5.3Hz,2H),7.39–7.29(m,3H),7.23–6.97(m,1H),6.93(d,J＝8.7Hz,1H),6.87(d,J＝2.1Hz,1H),6.80(dd,J＝8.8,2.3Hz,1H),6.02(dd,J＝11.7,4.2Hz,1H),3.96–3.82(m,4H),3.81(s,3H),3.71(s,3H),3.58(s,3H),3.33–3.28(m,1H).MS(ESI)m/z:471.20(C27H26N4O4,[M+H]+).Anal.CalcdforC27H26N4O4:C,68.92；H,5.57；N,11.91.Found:C,69.04；H,5.56；N,11.93。实施例2体外抗肿瘤活性测定IC50：半数抑制率，为化合物杀伤癌细胞能力。人肝癌细胞(HepG2)，人乳腺癌细胞(MCF-7)，人肺腺癌细胞(A549)。CC50：半数毒性浓度，化合物对正常细胞的毒性。人肾上皮细胞(293T)。采用MTT[3-(4,5)-双甲基-2-噻唑-(2,5)-苯基溴化四氮唑蓝]法来测定实施例1制备得到的化合物对人肝癌细胞(HepG2)，人乳腺癌细胞(MCF-7)以及人肺腺癌细胞(A549)的抑制率达到50％时的药物浓度(halfmaximalinhibitoryconcentration,IC50)。以及对人肾上皮细胞(293T)的细胞毒性，每个化合物的毒性用抑制293T细胞存活率到50％时的浓度CC50来表示。(1)培养液的配制：DMEM(基础培养基)89％，胎牛血清10％，青霉素链霉素溶液(10000IU/mL，10000μg/mL)1％。(2)四种贴壁癌细胞的培养：利用上述培养液(培养液体积约为培养瓶容量的1/10)，在37℃，5％CO2培养箱中培养，根据癌细胞的生长状态判断传代时间。(3)不同浓度药物的配制：利用DMEM(少量DMSO助溶)配制储备液，加药后的每孔细胞悬液中DMSO的终浓度一般不超过0.05％-0.1％；用DMEM把储备液稀释至六个浓度梯度(100μM，50μM，10μM，1μM，0.1μM，0.01μM)；保存于-20℃冰箱中备用。(4)细胞孵育：取对数生长期肿瘤细胞，调细胞悬液浓度为1-1.5×105/mL，混匀后加入96孔培养板中(100μL/孔)，在37℃，5％CO2培养箱中培养24h。(5)加药：将稀释好的不同浓度梯度的药物分别加入到96孔培养板中，每个浓度梯度设3个复孔，继续培养48h。实验分为实验组(培养液、细胞、药物)、对照组(培养液和细胞)和空白组(只有培养液)。(6)存活细胞检测：在培养了48h后的96孔板中，加MTT(5mg/mL)10μL/孔；在37℃放置4h后，移除上清液，加DMSO150μL/孔，振荡至甲瓒结晶全部溶解；利用自动酶标仪在570nm波长处检测各孔的光密度(OD值)。抑制率的计算：生长抑制率＝(1－存活率)×100％＝[1－(OD实验－OD空白)/(OD对照－OD空白)]×100％(OD实验表示实验组的平均光密度，OD对照表示对照组的平均光密度，OD空白表示空白组的平均光密度)。根据药物浓度-细胞生长抑制率的标准曲线，求其IC50，单位μM。结果如表2所示。表2以上结果表明，本发明所述烟酰二氢吡唑类化合物具有良好的抗肿瘤作用，对正常细胞的毒性低。当前第1页1 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