肝脏代谢处置导向的虫草素前药发现及其药动-药效学协同研究

Nonalcoholic fatty liver diseases (NAFLD) probably cause severe and progressive liver dysfunction, prompting the research and development of novel therapeutic drugs. Cordycepin, a biologically active natural product with confirmed alleviation of metaflammation, was considered as a desired lead compound for promising drug candidates for the treatment of NAFLD. Focusing on the distinctive hepatic metabolism and disposition of nucleoside drug, we described a novel liver-targeting nucleotide prodrug, bearing the characteristic group of ProTide strategy, to achieve the accumulation of active metabolite cordycepin-triphosphate in target tissue. We have smoothly synthesized one derivative named WJK8732 and validated its safety properties in toxicology evaluation (NOAEL > 1000 mg/kg). In the following, prodrug structure-pharmacokinetic-pharmacodynamic relationships will be described and the optimized prodrugs will be picked out through several rounds of followed two evaluation: the in vitro level of metabolic activation is determined by cellular pharmacokinetics, and the lipid-lowering and anti-inflammation activity are evaluated with NAFLD model. Notably, the hepatic metabolism and disposition of ProTide strategy has not reported in the state of NAFLD. Therefore, we attempt to discuss whether the difference of hepatic metabolism and disposition of ProTide strategy between normal and NAFLD mice do exist, and describe the underlying mechanism and the impact on the pharmacological activity. Based on the pharmacokinetic research platform in our lab, the following emphasis in our project is the achievement of collaborative innovation between nucleoside medicinal chemistry and cellular pharmacokinetics. Most of all, as the core issue in our project, the feasibility of this novel strategy will be confirmed, the key technology in the practicability of NAFLD evaluation will be developed and some candidate compounds will stand out with efficient anti-metaflammatory activity for further preclinical exploration.

非酒精性脂肪性肝病（NAFLD）导致患者肝功能恶化，药物研发刻不容缓。前期研究表明，虫草素能够有效缓解代谢性炎症，有望被开发为结构新颖、药效理想的NAFLD治疗药物。申请人聚焦核苷药物肝内代谢处置特征，拟采用ProTide策略将虫草素设计为单磷酸前药，增加虫草素活性代谢物的肝内富集。目前已成功合成WJK8732，其安全性理想（NOAEL＞1000mg/kg）。本课题将借助细胞药代动力学技术探究正常/疾病细胞模型中前药衍生物的代谢激活水平，并在体内外药效模型中揭示衍生物降脂抗炎活性；阐明结构-药动-药效关系，遴选优选前药分子。本课题还将首次关注ProTide前药在NAFLD状态下的代谢处置，探讨代谢酶表达异常对药动、药效的影响。课题将实现药代动力学与药物化学间的深度协同创新，验证虫草素前药设计概念，揭示ProTide前药代谢处置规律，为NAFLD的药物干预提供创新性基础研究及源头创新分子。

氧化应激损伤是众多疾病的核心病理环节，而抗氧化活性分子也在多种疾病的治疗及药物研发中发挥重要作用。我们首先聚焦天然活性分子虫草素，其具有丰富的药理学功能，包括免疫调节、抗炎、清除自由基、降脂及抗肿瘤等。虫草素的多重药效有望针对代谢性炎症的多重环节发挥治疗作用。虫草素为核苷类似物，其成药性需要进一步的优化。为此，我们希望通过前药策略优化虫草素的体内行为，进而改善药效。我们分别选用ProTide单磷酸前药策略及经典的酯前药策略，设计了两类虫草素衍生物。在化合物的合成中，我们实现了一类手性前药中间体的拆分，并通过培养单晶完成了其绝对构型的确证；这也帮助我们制备了光学纯手性单磷酸前药终产物。在前药的体外代谢研究中，我们基于口服吸收的多个环节，探究了一系列前药衍生物在人工胃液、肠液、大鼠肠S9、大鼠肝S9及大鼠血浆中的前药稳定性，并比较了其关键代谢物的生成。最终我们在两个系列的前药中均获得了优选分子，并计划开展深入的体内研究。核苷类前药存在代谢途径多元、代谢物结构多样且极性差异显著等诸多挑战。由于课题组前期已自主合成了一类核苷类药物ZL-01及其潜在的代谢物，为此，我们在开展上述研究内容的同时，亦针对ZL-01进行了体内LC-MS/MS分析方法的构建及代谢研究，以更好的胜任后续对虫草素前药的代谢探索。该体内分析方法符合生物分析方法要求；借助该方法，我们探究了ZL-01的细胞药代动力学，以及代谢物在肝脏及血浆中的代谢行为。. 我们在开展上述研究时，掌握了前药体内外代谢评价的一系列方法。基于此，我们希望通过前药策略，优化氢过硫化物（一类内源性含硫反应性代谢物）的稳定性与成药性。经过前药的设计、合成与释放性质的探索，我们成功发现了一类新型氢过硫化物供体，验证了该类供体在氧化应激状态下可释放氢过硫化物，并初步探索了其抗氧化活性。此外，本项目还拓展了一类内源性活性分子吡咯喹啉醌（PQQ）的应用，并对其进行了前药改造。