喜树碱类肝靶向衍生物的设计合成及其肝靶向机制

Camptothecin is an important natural product with anti-tumor activity，which show good inhibitory activity against liver tumor cells in vitro , but show the bad effect of treatment of liver cancer in vivo and toxic side effects mainly due to poor liver targeting. In this project, Bile acid and its analogues are selected as targeting ligands because of its own biological bodyderived liver targeting substances , and a series of camptothecin-bile acid conjugates are designed and synthesized. The topoisomerase inhibitory activity of these conjugates are evaluated to examine the original anti-tumor targets topoisomerase of camptothecin derivatives after a bile acid modification; The antitumor targeting effect of these conjugates are checked by competitive parallel experiment using the bile acid receptor expression of positive and negative tumor cells, and bile acid molecular mechanisms as targets are investigated. Molecular techniques are using to examine the characteristics of apoptosis protein induced by bile acid and camptothecin and protein changes induced by the new conjugates in liver cancer cells; The new conjugates of liver targeting metabolism in vivo and tissue distribution experiments are performed.The liver targeting camptothecin derivatives are designed and synthesized to provide new ideas for targeting drug, to perform practical basis for bile acid as a target for liver targeting substances, and to reveal its mechanism as a liver targeting molecules.

喜树碱是重要的具有抗肿瘤活性的天然产物，对肝肿瘤细胞的体外抑制活性很好，但在体内治疗肝癌效果差和毒副作用大，主要原因是肝靶向差，项目选择生物体内源性的肝靶向物质胆酸及其类似物作为靶向配基，与喜树碱进行偶联，设计合成一系列喜树碱-胆酸偶合物，并通过分子生物学手段对偶合物进行拓扑异构酶抑制活性的评价，考察喜树碱衍生物经过胆酸修饰后是否保持了原有的抗肿瘤作用靶点拓扑异构酶；同时通过胆酸受体表达阳性和阴性的肿瘤细胞进行竞争性平行实验，揭示新偶合物的抗肿瘤效果以及胆酸作为靶标的分子机制，利用分子手段考察新偶合物作用后的肿瘤细胞中喜树碱诱导调亡的特征调亡蛋白量和胆酸诱导的蛋白量的变化，进一步揭示新衍生物的肝靶向机制；通过动物体内代谢和组织分布等实验进一步验证新偶合物的肝靶向性。该项目实施将为肝靶向性的喜树碱衍生物的设计合成提供新思路，为胆酸作为肝靶向物质的靶标提实践依据，并揭示其作为肝靶标的分子机制

项目利用人体内源性物质胆酸具有肝肠特异吸收的生物学特性这一生理学现象，选择胆酸及其类似物脱氧胆酸、脱氢胆酸、饿脱氧胆酸作为靶向配基与喜树碱偶联，设计合成了31个胆酸-喜树碱新化合物，并通过质谱、核磁、红外等手段确证了新化合物的结构和理化性质。所用合成路线简单、反应条件温和、速率快、产率高、产物易纯化，成本低，易于产业化。从这些胆酸－喜树碱新衍生物中找到了肝肿瘤靶向选择性好且对正常组织毒性低的化合物，这些化合物仍具有很强的和抗肿瘤靶点拓扑异构酶I和DNA形成三元复合物的能力，并且内酯环稳定性得到大大提升，肝肿瘤靶向性的分子机制研究实验揭示出对肝肿瘤SMMC-7721细胞表象出明显选择性抑制的化合物进入细胞的主动转运载体与胆酸相同，从细胞水平和体内动物水平证实肝肿瘤靶向好的胆酸－喜树碱新衍生物是借助胆酸的选择性细胞转运过程实现肝癌靶向性的。这预示着胆酸作为结构修饰的靶向配基的确发挥了靶标的导向作用。为进一步证实上述推测的科学性，课题又合成了系列小檗碱--胆酸新衍生物（此部分内容不在申请书计划内），从中也找到了肝肿瘤靶向选择性好毒性低的小檗碱-胆酸新衍生物，该化合物在动物体内代谢过程中显示非常好的肝肠靶向富集能力，进一步证实了胆酸作为靶向前药设计的靶向配基的可行性和巨大的应用推广潜力。