Development of Drugs to Target Arginine Biosythesis in Mycobacterium tuberculosis

摘要 结核病(TB)仍然是一个重大的全球健康问题，导致1000万新病例和120万 根据世界卫生组织(WHO)的数据，仅在去年就有死亡人数。结核病的单一最大障碍 控制是它形成持久者的能力，结核分枝杆菌(Mtb)细胞的亚群是 对杀菌药物或免疫效应物的表型抵抗。我们发现精氨酸 通过将结核分枝杆菌细胞的突变体接种到无精氨酸的培养液或小鼠中而引起的饥饿， 在体外和体内都能杀死活跃生长的结核分枝杆菌细胞和结核分枝杆菌持久细胞的绝育。此外， 我们已经成功地确定了Mtb ArgB酶的结构，并使用了基于片段的药物 发现(FBDD)发现两种结合和抑制ArgB酶活性的支架化合物。这个 人类和我们的精氨酸生物合成途径酶Arg A、B、C、D和J的缺失 证明Mtb不能绕过argB或argF缺失使精氨酸生物合成酶具有吸引力 结核病药物开发的药物靶点。转录、代谢组学和流式细胞术分析 精氨酸饥饿细胞为绝育过程提供了新的见解。通过比较精氨酸 饥饿与其他三种绝育方案相比，我们确定了一组提供绝育的六个基因 签名。我们计划利用这些基因来改进药物发现，通过识别抑制 利用基于片段的药物设计，从基因上确认Mtb ArgB的任何额外靶点 精氨酸生物合成途径拓展药物靶向空间，并确定相关或致病因素 常见的不孕标志性基因在结核分枝杆菌不育症中发挥作用。这些研究将共同提供 一种快速消毒结核分枝杆菌培养的新方法，并为缩短结核病提供新的药物可能性和新的见解 化疗和治疗耐药结核病。

SUMMARY Tuberculosis (TB) remains a significant global health problem causing 10 million new cases and 1.2 million deaths just last year according to the World Health Organization (WHO). The single greatest impediment to TB control is its ability to form persisters, the subpopulation of Mycobacterium tuberculosis (Mtb) cells that are phenotypically resistant to killing by bactericidal drugs or immune effectors. We have discovered that arginine starvation, induced by inoculating mutants of Mtb cells into arginine-free media or mice, mediates rapid sterilization that kills both actively growing Mtb cells and Mtb persister cells both in vitro and in vivo. Moreover, we have successfully determined the structure of the Mtb ArgB enzyme and used Fragment-Based Drug Discovery (FBDD) to discover two scaffold compounds that bind and inhibit the enzymatic activity of ArgB. The absence of the de novo arginine biosynthesis pathway enzymes Arg A, B, C, D, and J in humans and our demonstration that Mtb cannot bypass argB or argF deletions make Arginine Biosynthetic Enzymes attractive drug targets for TB drug development. Transcriptomic, metabolomics and flow-cytometric analyses on the arginine starving cells have provided new insights into the sterilization process. By comparing arginine starvation to three other sterilizing regimens, we identified a set of six genes that provide a sterilization signature. We plan to exploit these genes for improved drug discovery, by identifying novel drugs to inhibit the activity of Mtb ArgB using fragment-based drug design, confirm genetically any additional targets of the arginine biosynthetic pathway to expand the drug target space, and determining the correlative or causative role the common sterilizing signature genes play in Mtb sterilization. Together these studies will make available a new way to rapidly sterilize cultures of Mtb and provide new drug possibilities and new insights to shorten TB chemotherapy and treat drug-resistant TB.