Discovery of therapeutics against Cryptococcosis by Repurposing Pharmaceutical Libraries

摘要：由隐球菌属感染引起的隐球菌病。真菌。新生革兰氏菌和加蒂亚革兰氏菌遗骸 艾滋病毒携带者和器官移植后状态的头号杀手。最近暴发的C. 具有免疫能力的人群中的Gattii引起了对隐球菌种对 公共卫生。隐球菌病是最难治疗的感染之一，死亡率超过50%。新的 迫切需要具有新作用机制(MOA)的治疗方法来治疗感染者。高吞吐量 筛选靶向药物文库是发现抗真菌药物的一种非常有前途的策略 具有新的MOA，因为这些库中含有定义明确的分子/化合物 物理化学性质，通常与其他治疗适应症的现有临床前数据一起。因此，一旦 一旦确定，它们就可以快速开发用于抗真菌等新应用。隐球菌含有 必需基因Prp8中的蛋白质自剪接元件，称为内含素。因为内含子不存在于多个 细胞生物体，如人类，经常扰乱关键微生物基因的功能，它们很有吸引力 毒品目标。我们已经开发了高通量筛选(HTS)策略来识别Prp8 内含素剪接抑制药。在我们的初步结果中，我们确定了几种小分子抑制剂，它们可以抑制 Prp8内含子在体外剪接，体内抑制隐球菌病。在本应用程序中，我们将使用我们的油井- 开发了HTS检测方法，以筛选ICCB的预定义和靶向药剂库- 哈佛医学院的朗伍德筛选设施，用于识别Prp8内含子剪接抑制物。这个 已确定的候选抑制剂将在体外和体内进行表征和优化，最终目标是 开发它们作为治疗隐球菌病的新疗法。

Abstract: Cryptococcosis caused by infections of Cryptococcus spp. fungi. C. neoformans and C. gattii remains a leading killer of people living with HIV and post-organ transplant status. Recent outbreaks of infections of C. gattii in immune competent people raise significant concerns about the overall threat of cryptococcal species to public health. Cryptococcosis is one of the most difficult infections to treat with a mortality rate over 50%. New therapies with novel mechanism of actions (MOAs) are urgently needed to treat the infected. High throughput screening of target-based pharmaceutical libraries represents a very promising strategy to discover antifungals with novel MOAs because these libraries harbor well-defined molecules/compounds with desirable physicochemical properties, often with existing preclinical data for other therapeutic indications. Therefore, once identified, they can be quickly developed for novel applications such as antifungal. Cryptococcal fungi contain protein self-splicing elements, called inteins, in the essential gene, Prp8. Because inteins do not exist in multi- cellular organisms, such as humans, and often disrupt the functions of critical microbial genes, they are attractive drug targets. We have developed high throughput screening (HTS) strategies to allow identification of the Prp8 intein splicing inhibitors. In our preliminary results, we identified a few small molecule inhibitors that can inhibit the Prp8 intein splicing in vitro and inhibit the cryptococcosis in vivo. In this application, we will use our well- developed HTS assays to screen the predefined and target-based pharmaceutical libraries at the ICCB- Longwood screening facilities at the Harvard Medical School to identify the Prp8 intein splicing inhibitors. The identified candidate inhibitors will be characterized and optimized both in vitro and in vivo with the final aim to develop them as novel therapeutics for cryptococcosis.