基于“shock and kill”策略研究两株深海链霉菌中HIV潜伏激活的活性成分

The “shock and kill” strategy works by two steps: the first “shock” step by giving a drug revives latent HIV in patients; then the second “kill” step to clear all activated HIV by Highly Active Antiretroviral Therapy (HAART). Since HAART is universally recognized as one of the most effective approach to treat AIDS, the key point of the strategy is to discover compounds from natural sources with latent HIV reactivation bioactivities, which is also the hotspot for research and development of the anti-AIDS drugs. On the basis of huge microorgamisn resources of the Marine Culture Collection of China (MCCC), all 104 deep-sea derived streptomycetes in MCCCC were selected and subjected to small-scale fermentation. Their crude extracts were tested for HIV latency reactivative activities and analysed for chemical compositions as well. Consequently, two strains of streptomyces sp. 1A09851 and 1A01793 were found to have diverse components and potent bioactivities (The reactivation rates were 23.7% and 21.1%, respectively, under the concentration of 10 μg/mL, which was about 50% compared to that of the positive control). Therefore, these two strains were chosen in this project to search for HIV latency reactivative compounds using bio-guided isolation. The most bioactive compounds will be accumulated by 100 L large-scale fermentation under optimized media and conditions. Then it will be subjected to in-depth bioactive mechanism study. In general, the project will afford some anti-AIDS lead compounds with the self-property right completely owned by our country.

“shock and kill”是通过“shock”药物激活潜伏HIV病毒，而后用“kill”药物（高效抗逆转录病毒疗法）清除激活的病毒，达到根除HIV的目的，是目前国际上抗AIDS研究的热点。其关键在于如何从天然产物中寻找发现HIV潜伏激活的活性成分，这也是当前抗AIDS药物研发的重点。前期，申请人选取中国海洋微生物菌株保藏管理中心所有的104株深海链霉菌，通过HIV潜伏激活活性测试及成分分析，发现1A09851和1A01793不仅次级代谢产物丰富，而且具有很强的活性（在10 μg/mL浓度下，其激活率分别为23.7%和21.1%，约为阳性对照药的50%）。因此本项目拟以该两株深海链霉菌为研究对象，以活性为导向，发现HIV潜伏激活的活性成分，对活性最好的目标化合物进行大量富集，深入研究其作用机制。通过本项目研究，有望为具有我国完全自主知识产权的抗AIDS药物的研发提供模式结构和药物前体。

海洋微生物是新颖结构和独特活性化合物发现的重要来源，是天然产物研究新的重点领域。本项目旨在前期研究的基础上，基于从具有抗HIV潜伏激活活性的天然产物中寻找抗艾滋病（AIDS）药物的思路，以前期筛选获得具有强潜伏激活活性的两株海洋微生物为研究对象，通过微生物发酵、次级代谢物提取、分离、结构鉴定、活性测试等工作，发现其中的活性成分单体。项目执行四年来，我们对五株海洋微生物Nesterenkonia halobia、Aspergillus ochraceus、Microbacterium sp.、Botryotinia fuckeliana和Chaetomium globosum进行了系统的化学成分研究，从中一共分离得到61个化合物，其中新化合物20个。经体外抗HIV活性测试，发现四个强活性化合物：fellutamide B、8-methoxy-1,2,3,4-tetrahydronaphthalene-1,2,4-triol、1,3-dihydro-4,5,6-trihydroxy-7-methyl-isobenzofuran和5,6,7-trihydroxy-4-methylisobenzofuran-1(3H)-one。尤其是fellutamide B，5 μM浓度下HIV潜伏病毒激活率为62.27 %，同时Jurkat 2D10细胞存活率为55.4%，因此具有良好的研发前景。为了拓宽活性，我们还对所获得的化合物分别进行了抗炎、抗肿瘤、抗过敏等活性的测试，发现了一批具有活性较好的化合物，基于以上的研究，本项目一共发表相关研究论文15篇，申请专利3项。综上所述，本项目的实施为从海洋微生物来源的抗AIDS药物的研发提供了模式结构和药物前体。为具有我国完全自主知识产权的海洋药物发现奠定了坚实的物质基础。