Structures of Amphotericin-Sterol Complexes

描述（由申请人提供）：该项目旨在促进对临床重要但剧毒抗真菌药物两性霉素B （AmB）的作用方式的理解。与经典的离子通道模型不同，本提案的初步研究表明，AmB形成大的膜外聚集体，从脂质双层中提取甾醇，从而杀死细胞。这种新颖的“甾醇海绵”模型为改善治疗指标提供了一个新的、更可行的路线图，即最大化AmB甾醇海绵对真菌（麦角甾醇）和人类（胆固醇）中发现的甾醇的相对结合亲和力。目的1是确定在生理相关脂质双分子层存在下组装的AmB甾醇海绵的结构。目的2是确定AmB海绵与麦角甾醇和胆固醇复合物的结构。目的3是确定从AmB的新衍生物AmBMU中提取的甾醇海绵和相应的麦角甾醇复合物的结构，AmBMU最近被发现并被证明可以结合麦角甾醇而不是胆固醇，对人体细胞无毒，并且在体外和小鼠中保持有效的抗真菌活性。总的来说，这些研究将为AmB生物活性基础上的原子相互作用提供高分辨率的图像，从而有力地促进这种临床重要天然产物的毒性较低的衍生物的合理开发。这些研究也将进一步阐明如何实现临床验证的耐药抗微生物作用的基本特征，并为其他生物学相关的小分子-小分子的前沿探索奠定基础

DESCRIPTION (provided by applicant): This project aims to advance understanding of the mode of action of the clinically vital but also highly toxic antifungal drug amphotericin B (AmB). Alternative to the classic ion channel model, preliminary studies in this proposal show that AmB forms large extramembranous aggregates that extract sterols from lipid bilayers and thereby kill cells. This novel "sterol sponge" model illuminates a new and more actionable roadmap to an improved therapeutic index, i.e., maximize the relative binding affinity of the AmB sterol sponge for the sterol found in fungi (ergosterol) vs. humans (cholesterol). Aim 1 is to determine the structure of the AmB sterol sponge, assembled in the presence of physiologically relevant lipid bilayers. Aim 2 is to determine the structures of the complexes of the AmB sponge with ergosterol and cholesterol. Aim 3 is to determine the structure of the sterol sponge and corresponding ergosterol complex derived from a new derivative of AmB, AmBMU, which was recently discovered and shown to bind ergosterol but not cholesterol, to be non-toxic to human cells, and to retain potent antifungal activity in vitro and in mice. Collectively, these studies wll provide a high-resolution picture of the atomistic interactions that underlie the biological activities of AmB and thus powerfully enable the rational development of less toxic derivatives of this clinically vital natural product. These studies will also further illuminate the fundamental features of how clinically validated resistance-refractory antimicrobial action can be achieved and lay the foundation for the frontier pursuit of other biologically relevant small molecule-small molecule interactions. Relevance to Human Health. Amphotericin B is the powerful but unfortunately highly toxic gold standard therapy for treatment of systemic fungal infections, and this drug has uniquely evaded the emergence of microbial resistance despite more than half a century of widespread clinical utilization. Better understanding how AmB exerts its biological activities is thus critical for guiding the rational development of derivatives with an improved therapeutic index as well as other resistance-refractory antimicrobial agents.