Structure-based discovery and development of HIV-1 gp41 fusion inhibitors

描述（由申请人提供）：本提案描述了对人类免疫缺陷病毒（HIV-1）融合有效的低分子量gp41抑制剂的发现和开发。融合抑制剂具有阻断HIV传播和预防疾病进展的优异特性，因为它们阻断健康细胞的初始感染和感染的细胞间扩散。然而，目前还没有小分子的融合抑制剂，而且肽融合抑制剂具有成本高、易被蛋白水解等不理想的特点。该应用程序的目的是开发高亲和力结合gp41的小分子先导化合物，防止N-和c -螺旋结构域的结合，这是融合反应的核心特征。该提案实现了一个系统的基于结构的融合抑制剂开发方法，有三个具体目标：(1)gp41 n -螺旋卷曲线圈的模块化亚段聚焦于或围绕已知疏水口袋将被检查小分子结合，通过开发荧光，核磁共振和生物分析来检测和定量结合。(2)将开发新的核磁共振方法来获得明确的结构约束，定义结合配体的构象和取向，为计算机建模和合成改进抑制剂提供定量的构效关系数据。(3)具有拟肽支架的化合物在疏水口袋中显示出有希望的结合特性，并将成为制备新型抑制剂的设计策略的基础，使用结合、结构和生物学分析进行评估和优化。在分析应用过程中获得的附加或相邻命中和结构数据将纳入先导物优化过程。细胞-细胞融合和病毒感染性试验将用于提供结合研究的确证，并确保主要候选药物具有所需的体外特性。这种方法将允许测试中心假设，即小分子结合到疏水口袋并可能延伸到疏水口袋附近的凹槽中，可以发展成高效的融合抑制剂，与肽相媲美。所描述的研究将提供有效的方法来检测新一代抑制剂，并获得其与gp41复合物的结构信息。因此，由于目前没有描述小分子gp41复合物细节的实验数据，它们将有助于填补gp41抑制机制知识的空白。这项提议的意义在于，它有可能为HIV-1融合抑制剂的开发做出特别贡献，也有助于开发适用于广泛的1类病毒的技术，这些病毒使用与HIV-1相似的融合机制，这与NIH增强基础知识以治疗人类疾病的使命有关。该项目的长期目标是产生类似药物的化合物，对HIV-1融合具有活性，适用于治疗或杀微生物剂，可用于对抗或预防全球每年发生的数百万例新感染。

DESCRIPTION (provided by applicant): This proposal describes the discovery and development of low molecular weight gp41 inhibitors effective against Human Immunodeficiency Virus (HIV-1) fusion. Fusion inhibitors possess excellent characteristics for interrupting HIV transmission and preventing disease progression, since they block initial infection of healthy cells and cell-to-cell spread of infection. Yet there are currently no small molecule inhibitors of fusion, and peptide fusion inhibitors possess some undesirable characteristics such as high cost and susceptibility to proteolysis. The objective of this application is to develop small molecule lead compounds which bind to gp41 with high affinity, preventing the association of N- and C-helical domains that is a central feature of the fusion reaction. The proposal implements a systematic structure-based approach to fusion inhibitor development in three Specific Aims: (1) Modular subsections of the gp41 N-helical coiled coil focused on or around a known hydrophobic pocket will be examined for small molecule binding, through the development of fluorescence, NMR and biological assays to detect and quantitate binding. (2) Novel NMR methods for obtaining explicit structural constraints defining bound ligand conformation and orientation will be developed, providing quantitative structure-activity relationship data for computer modeling and synthesis of improved inhibitors. (3) Compounds with peptidomimetic scaffolds have shown promising binding characteristics in the hydrophobic pocket and will form the basis of design strategies for preparing novel inhibitors, using binding, structural and biological assays for evaluation and optimization. Additional or adjacent hits and structural data obtained during assay application will be incorporated into the lead optimization process. Cell-cell fusion and viral infectivity assays will be applied to provide corroboration of binding studies and to ensure that lead candidates have the desired in vitro properties. This approach will allow testing of the central hypothesis that small molecules binding to the hydrophobic pocket and potentially extending into grooves adjacent to the hydrophobic pocket can be developed into highly potent fusion inhibitors, rivaling peptides. The studies described will provide efficient methods to detect new generations of inhibitors and obtain structural information on their complexes with gp41. Thus they will contribute to a gap in knowledge of the mechanism of gp41 inhibition, since there are currently no experimental data describing the details of small-molecule - gp41 complexes. The significance of this proposal lies in its potential to contribute specifically to HIV-1 fusion inhibitor development and also to the development of techniques applicable to a wide range of Class 1 viruses which use a similar fusion mechanism to HIV-1, a broader goal relevant to NIH's mission of enhancing fundamental knowledge to treat human disease. The long-term goal of the project is to generate drug-like compounds active against HIV-1 fusion, suitable for therapeutic or microbicide use, which could be used to counter or prevent the millions of new infections that occur annually worldwide. PUBLIC HEALTH RELEVANCE: The prevalence of HIV infection remains a significant public health problem due to the development of viral strains resistant to current treatments, and due to spread of the virus from individuals who are unaware they are infected. This proposal seeks to apply structure-aided drug design to develop low molecular weight HIV fusion inhibitors which prevent viral entry and the cell-to-cell spread that causes immune deficiency. Drugs in this class would be useful in microbicides to prevent infection, or could be given orally to treat infection as an alternative to the currently used fusion inhibitor Enfuvirtide(R), which must be given intravenously.