茄科植物中靶向HIF2α和TRAIL-DR5抗肾癌的withanolide类特色成分的发现及作用机制研究

Kidney cancer, known as renal cell carcinoma (RCC) is inherently multidrug-resistant tumor, with the highly drug- and radiation-resistant nature when compared with other neoplasms such as lung or breast cancer. It is very significant for us to find out some new drugs for the treatment of RCC. The Solanaceae plants are a rich source of natural withanolides. In our previous study, the extract of the aerial parts of Physalis pubescens L. (Solanaceae) and its major constituent physapubescin showed marked antiproliferative activity against human renal cell carcinoma 786-O cells. Physapubescin could not only reverse the tolerability and adaptability of renal cell carcinoma 786-O cells to hypoxia, but also sensitize TRAIL-induced apoptosis through targeting VHL-HIF2α-VEGF and TRAIL-DR5 pathway, resulting in the improvement of drug-resistance of RCC. However, its underlying action mechanisms have not yet been fully elucidated. Therefore, further investigation will be done in this project based on our previous work, including: a) isolate and identify more withanolides from three medicinal and edible plants of Solanaceae family, Physalis peruviana L., Tubocapsicum anomalum and Nicandra physaloides (L.) Gaertn under the guidance sensitive TLC, NMR and MS analyses; b) examine the antiproliferative activity of the isolated withanolides against human renal cell carcinoma 786-O cells, and clarify the structure-activity relationship against RCC combined with molecular docking virtual experiments, microscale thermophoresis method (MST) technique, enzymatic activity assay, cocrystallization, etc.; c) elucidate the underlying anticancer mechanisms of the active withanolides by using flow cytometry, Western blot and ELISA analysis, etc. The purpose of this project is to discover novel lead compounds of withanolides or drug candidates against renal cell carcinoma from the medicinal and edible plants of Solanaceae family and to provide scientific supports for the interpretation of traditional use of these plants.

肾癌是典型的多药耐药肿瘤，对放化疗高度不敏感。骨架类型丰富的withanolide类化合物是茄科种属植物中的特色成分，前期研究首次发现毛酸浆中的该类成分physapubescin能够通过靶向HIF2α和TRAIL-DR5，逆转肾癌细胞对缺氧的耐受性和增敏TRAIL诱导的细胞凋亡，从而有效对抗肾癌细胞耐药。本项目拟在上述研究基础上，采用TLC结合NMR、MS快速识别方法，对茄科三种富含withanolide类成分的植物，假酸浆、灯笼果和龙珠进行分离，利用基于靶点蛋白晶体结构的生物学筛选方法快速发现靶向性强的活性withanolide类化合物，确定不同亚型结构与靶点的作用，并采用系统生物学方法深入研究其抗肾癌作用机制。本课题的实施完成不仅可以发现不同类型的withanolide成分作为多靶点抗肾癌药物分子的新用途，也为阐明茄科药用植物发挥传统功效的物质基础提供科学依据，对肾癌治疗具有重要意义。

骨架类型丰富的withanolide类化合物是茄科种属植物中的特色成分，前期研究首次发现毛酸浆中的该类成分physapubescin能够通过靶向HIF2α和TRAIL-DR5，逆转肾癌细胞对缺氧的耐受性和增敏TRAIL诱导的细胞凋亡，从而有效对抗肾癌细胞耐药。本项目在上述研究基础上，采用TLC结合NMR、MS快速识别方法，对茄科三种富含withanolide类成分的植物，假酸浆、灯笼果和龙珠进行分离，共得到182个化合物，其中119个为该类型的成分；评价了其对人肾癌细胞（786-O、A-498），人前列腺癌细胞（C4-2B和22Rvl），人乳腺癌细胞（MDA-MB-231）以及人胰腺癌细胞（SW1990）的增殖抑制活性，发现了多个化合物具有显著的肿瘤细胞生长抑制作用；在上述活性研究的基础上，初步明确了该类化合物的抗肿瘤构效关系；采用流式细胞术、Western blot、计算机虚拟筛选等方法深入研究了活性化合物nicaphysalin I、ixocarpalactone A、tubocapsanolide A抗肿瘤作用机制及作用靶点。上述研究成果已发表相关SCI收录论文11篇，中文核心期刊文章1篇，国际会议报告1次，沈阳市自然科学学术成果奖三等奖1项，授权专利4项，培养研究生5名。本课题的实施完成不仅发现了不同类型的withanolide成分作为多靶点抗肿瘤药物分子的新用途，也为阐明茄科药用植物发挥传统功效的物质基础提供科学依据，对肿瘤治疗具有重要意义。