Exploiting metabolic toxicities to identify compounds that inhibit cholesterol metabolism in M. Tuberculosis

 描述（由申请人提供）：结核病（TB）是一种持续的全球流行病，每年夺去约150万人的生命。结核分枝杆菌（Mtb）是TB的病原体，并且该细菌通过在巨噬细胞内存活并操纵宿主免疫应答来建立长期感染。在肺组织内，结核分枝杆菌可以持续数十年，远离宿主免疫反应的压力。在这种持续性感染期间，Mtb仅利用宿主来源的营养物存活。结核分枝杆菌具有在感染期间利用宿主衍生的胆固醇来提供能量产生和/或生物合成途径的能力。因此，胆固醇催化剂的抑制可能是一个弱点，可以用于发现新的抗结核药物。在这个项目中，我们将开发一种适合HTS的基于细胞的检测方法，该方法可以识别结核分枝杆菌中胆固醇催化剂的抑制剂。目的1：我们将开发一个强大的化学抑制HTS检测384孔格式使用Mtb Icl 1菌株。目标2：我们将完成一个试点HTS并验证屏幕上的命中。目标3：将重点放在 鉴定HTS命中的酶靶标。这些研究将产生HTS就绪测定，并提供“原理证明”，即我们的方法可以鉴定Mtb中胆固醇催化剂的选择性抑制剂。

 DESCRIPTION (provided by applicant): Tuberculosis (TB) is a lasting global epidemic that claims ~1.5 million human lives annually. Mycobacterium tuberculosis (Mtb) is the causative agent of TB and this bacterium establishes long term infections by surviving within macrophages and manipulating the host immune response. Within lung tissue Mtb can persist for decades sequestered away from the pressures of the host immune response. During this persistent infection Mtb solely utilizes host-derived nutrients for survival. Mtb has the ability t utilize host-derived cholesterol during an infection to supply energy producing and/or biosynthetic pathways. Therefore, inhibition of cholesterol catabolism may be a weakness that can be exploited for discovery of new anti-Mtb drugs. For this project we will develop a cell-based assay suitable for HTS that can identify inhibitors of cholesterol catabolism in Mtb. Aim 1: we will develop a robust chemical-suppressor HTS assay 384-well format using the Mtb Icl1 strain. Aim 2: we will complete a pilot HTS and validate hits from the screen. Aim 3: will focus on identifying the enzyme targets of the HTS hits. These studies will yield a HTS ready assay and provide "proof of principle" that our approach can identify selective inhibitors of cholesterol catabolism in Mtb.