几种黔产民族药用植物内生真菌中新颖P-gp抑制剂的研究

Multidrug resistance (MDR) is the leading cause of treatment failure in cancer therapy. One of the most common mechanisms of MDR is drug efflux mediated by transporters such as P-glycoprotein (P-gp), one of the most typical and the most widely investigated ATP-binding cassette (ABC) transporters. To address this problem, P-gp inhibitors such as verapamil, quinine, and cyclosporin A have been introduced to reverse MDR since the early 1980s. However, clinical trials have been disappointing due to the high inherent toxicities of P-gp inhibitors and pharmacokinetic interactions between anticancer drugs and inhibitors. Therefore, it is imperative to discover a new kind of natural, efficient and safe inhibitors of P-gp in cancer treatment. Guizhou Provience is rich in ethnic medicinal plant. The secondary metabolites with novel structure and activities could be important resources to discover new effective compounds, which produced by endophytic fungi associated with Guizhou ethnic medicinal plants. Our group has recently first discovered the endophytic fungi in the ethnomedicine “Ainaxiang”, “Jixiangcao”, “Qinghao” and “Muhao” with the activation of inhibiting P-gp. The fungi can also produce a series of different-structured-compounds, which probably produce an active secondary metabolite from the fungi with the activity of inhibiting P-gp potentially. Therefore, this project will continue to screen the endophytic fungi from several Guizhou ethnic medicinal plants in accordance with the thoughts of the new sources, new compounds and the new activity. The combination of activity, chemistry and gene method was used, for the first time, as an effective strategy to screen the bioactive endophytic fungi. The secondary metabolites produced by the active strains with high quality were separated and purified, and then their structures were identified. The activity of inhibiting P-gp and the mechanism were evaluated. Finally we can gain the novel, active P-gp inhibitors. The present study will provide the scientific basis for the research of P-gp inhibitors, and also utilize microbial sources for drug development.

肿瘤细胞的多药耐药性(MDR)是化疗失败的主要原因，P-糖蛋白(P-gp)的过度表达是导致MDR的主要机制，临床上仍缺乏高效低毒的P-gp抑制剂。寻找天然高效安全的P-gp抑制剂是恶性肿瘤治疗的当务之急。贵州省民族药用植物资源丰富，而植物内生真菌次生代谢产物具有结构新颖、活性独特等特点，是寻找先导化合物的重要资源。我们在前期对内生真菌的研究中，首次发现四株分别来源于艾纳香、吉祥草、青蒿和牡蒿的内生真菌均具有较强P-gp抑制活性，能产生系列多结构类型化合物，提示菌株次生代谢产物可能是其发挥P-gp抑制活性的主体。因此，本项目按照新来源、新化合物和新活性的思路，首次采用活性、化学和基因三种技术结合的方法对优质(具丰富化学成分和功能基因)活性菌株进行筛选，对其次生代谢产物进行分离纯化和结构鉴定，最终获得结构新颖活性显著的P-gp抑制剂，为P-gp抑制剂和微生物源药物的研究提供科学依据和方法借鉴。

肿瘤细胞的多药耐药性(MDR)是化疗失败的主要原因，P-糖蛋白(P-gp)的过度表达是导致MDR的主要机制，临床上仍缺乏高效低毒的P-gp抑制剂。寻找天然高效安全的P-gp抑制剂是恶性肿瘤治疗的当务之急。贵州省民族药用植物资源丰富，而植物内生真菌次生代谢产物具有结构新颖、活性独特等特点，是寻找先导化合物的重要资源。我们在对内生真菌的研究中，建立了黔产药用植物内生真菌菌种资源库，共有菌种1730株，筛选获得具有P-gp抑制活性的菌株21株，优质(即具丰富化学成分和功能基因)活性菌株6株，获得富含新型P-gp 抑制剂的资源；共获得优质活性菌株次生代谢产物70个，其中1个为新化合物，结构涉及环肽（对称环四肽，二肽类化合物）、聚酮、脂肪链糖苷类等化合物类型。对这些化合物进行抗肿瘤逆转活性筛选发现，槲皮素、山奈酚、小白菊内脂、木犀草素和甜菜碱对MCF-7/ADR具有较高的逆转活性，其中小白菊内脂逆转倍数最高为10.35；山奈酚对A2780/Taxol的逆转倍数为5.04。此外，筛选获得的菌株中PF02-2粗提物对MCF-7/ADR的逆转倍数达3.79，具有较高的逆转活性潜力。进一步研究发现，PF02-2粗提物通过介导P-gp相关基因的表达，抑制了MCF-7/ADR的P-gp的活性，从而实现逆转。PF02-2中共分离获得30个单体化合物，对MCF-7/ADR的逆转活性尚在进行当中。