马蹄金素氟代二肽模拟物的设计合成及抗HBV活性研究

Amide bond exists widely in pharmaceuticals. Nevertheless, amide bond-containing biomolecules use as therapeutic agents is still limited by a few factors, for example, low cellular uptake and fast metabolization in vivo. One way to overcome these drawbacks and retain its biological activities is replaceing the amide bond with different bioisosteres to obtain corresponding mimetics. Matijin-Su [MTS] was isolated from ethnic drug Matijin (Dichondra repens Forst.) by our research group. In our previous studies, more than 800 of MTS derivatives were synthesized by structural modification, and their biological evaluation showed that some of the synthetic derivatives gave significant inhibition on HBV DNA replication, as well as high selectivity index (SI). In addition, bioactivity test of some derivatives showed significant effect of anti-duck hepatitis B virus without the effect of “back up”. One of the derivatives, named Tifentai (Y101), has been finished its preclinical studies and phase 1 clinical trial. The study on metabolism and pharmacokinetics of Y101 showed that its elimination half-life were short both in rat and human due to the amide bond of Y101 was easily hydrolyzed in vivo to produce a hydrolysis product M8 as major metabolite. The elimination half-life time of Y101 in rats after intragastric administration and in human body after single administration were 2.5-3.1 hours and 1.6-2.1 hours respectively. In this project, to improve the metabolic stability of MTS derivatives in vivo, enhance their efficacy, reduce the dosage, more than 70 fluorinated dipeptidomimetics of MTS will be designed and synthesized by using fluorine substituted bioisostere trifluoromethyl substituted amine unit or fluoroolefin unit to replace the amide bond of the MTS derivatives which have been found with higher anti-HBV activity in previous studies. In addition, the anti-HBV activity of all the synthetic dipeptidomimetics of MTS in vitro will be tested. The anti-DHBV activity of the effective derivatives in vivo and their mechanisms of action will be systematically studied. The results of this study will provide a scientific basis for the further studies of amide bond-containing biomolecules.

酰胺键结构广泛存在于药物中，然而酰胺类药物也存在着代谢不稳定、膜渗透性差等缺陷。克服这些缺陷的有效办法之一是利用生物电子等排体替换酰胺键，设计合成新型酰胺模拟物。本项目组前期设计合成了800多个马蹄金素（MTS）二肽衍生物，并进行了抗乙肝病毒（HBV）活性研究。其中一个衍生物替芬泰（Y101）已完成临床前研究和1期临床试验，发现其在大鼠和人的体内消除半衰期较短，主要代谢产物均为酰胺键水解产物M8。为探索利用生物电子等排体替换酰胺键，提高该类化合物的代谢稳定性，本项目将选取体外抗HBV活性较高、毒性较低的MTS衍生物，将其肽（酰胺）键结构单元替换为三氟甲基取代单元或氟代烯烃结构，设计合成70个以上MTS衍生物的氟代二肽模拟物，并测试其抗HBV活性，选择高活性MTS模拟物开展体内代谢稳定性和作用机制研究，争取发现高活性的目标分子，也为具有酰胺结构药物的深入研究开发供科学依据。

克服酰胺类药物代谢不稳定、膜渗透性差等缺陷的有效方法之一是利用生物电子等排体替换酰胺键，设计合成新型酰胺模拟物。临床新药替芬泰（Y101）是由先导化合物马蹄金素（MTS）经结构优化而得，鉴于Y101在大鼠和人的体内消除半衰期较短，主要代谢产物均为酰胺键水解产物M8。为探索利用生物电子等排体替换酰胺键，提高MTS类化合物的代谢稳定性，本项目选取前期发现的体外抗乙肝病毒（hepatitis B virus，HBV）活性较高、毒性较低的马蹄金素（MTS）衍生物，将其中肽键（酰胺）结构单元替换为生物电子等排体三氟甲基取代的甲氨结构单元，设计合成MTS肽键模拟物共计144个和含噁唑环结构的A系列目标分子共计21个。并对合成所得目标化合物进行了抗HBV活性测试，结果显示约有40%的目标化合物对2.2.15细胞的HBV DNA的复制均有一定的抑制作用，大多数化合物的抗HBV活性较原型化合物抗HBV活性有较大提高。其中化合物C20、C24、D28、D29、D61、D62、D75对HBV DNA的半数抑制浓度（IC50值）分别为0.14、0.42、0.29、0.37、0.29、0.42、0.34、0.31µM，抗HBV活性高于MTS原型化合物。经肝微粒体体外代谢测试结果显示肽键模拟物D28较原型化合物Y101的代谢稳定性有了显著提高，克服了MTS类化合物代谢不稳定的不足。该结果为研发新型抗乙肝病毒药物打下了研究基础。