Fragment to small molecule hit discovery targeting Mycobacterium tuberculosis FtsZ

结核病是发展中国家和发达国家发病率和死亡率的主要原因。2021年，有1060万人患上了这种疾病，160万人死于结核病。大约45万例报告病例被归类为多药耐药结核病(MDR-TB)，部分患者患有广泛耐药结核病(XDR-TB)。世界上约四分之一的人口患有潜伏性结核病，约10%-15%的人继续发展和传播感染。患者能否获得目前的治疗以及遵守漫长的剂量方案是一个全球性的挑战。耐多药结核病更难治疗，新出现的广泛耐药结核病的选择更加有限。迫切需要开发新型抗结核药物，以成功对抗新出现的耐药性，这是一种尚未得到满足的医疗需求。已知的抗结核药物的再利用和再利用取得了最近的进展，但在临床试验中还没有新的药物类别通过新的作用机制发挥作用。“丝化温度敏感突变体Z”(FtsZ)在所有类型的细菌中都高度保守，包括引起结核病的分枝杆菌物种。它是一种重要的细胞分裂蛋白，具有GTP酶和聚合活性。FtsZ亚基聚合成原丝，形成一种称为Z环的动态环状结构，作为分裂体组装的支架，分裂体是一种多蛋白质复合体，导致Z环收缩，最终导致隔膜形成和细胞分裂。FtsZ的异常阻断了Z环的功能，导致细长的丝状表型，抑制细胞分裂和随后的细胞死亡。虽然FtsZ抑制剂已被鉴定，但大多数来自天然产物，结构信息普遍缺乏，许多抑制剂被归类为假阳性。我们已经大量提纯了结核分枝杆菌(MTB)FtsZ，用于生化、生物物理和结构研究。在与Monash核磁共振筛选设施的合作下，我们确定了38个针对FtsZ的片段。我们建立了生物物理(DSF，MST)和生化(GTPase，直角散射)分析来表征其GTPase活性和聚合/解聚动力学。独一无二的是，我们确定了FtsZ与碎片HITS形成的络合物的晶体结构，并建立了它们的作用机制。我们已经就片段的生长和合并评估了片段结合位点，并准备启动逐个目录的SAR和定制的HITS优化，以开发更有效的类似物。在这个项目中，我们的目标是开发和表征有效的结核分枝杆菌FtsZ抑制剂，作为一种治疗结核病感染的新型抗生素。我们预计，这些化合物将是结核病及其耐药变种现有治疗选择的一个受欢迎的补充。这一合作的药物发现项目解决了一个明确的未得到满足的医疗需求。该项目的成功完成将为包括UCL其他合作伙伴在内的更广泛的合作药物发现计划奠定基础。

Tuberculosis (TB) is a major cause of morbidity and mortality in developing and developed countries. 10.6 million people developed the disease and 1.6 million people died from TB in 2021. Approximately 450,000 reported cases are classified as multidrug-resistant tuberculosis (MDR-TB), with a subset of patients developing extensively-drug-resistant tuberculosis (XDR-TB).About a quarter of the world's population have latent TB, with ~10-15% going on to develop and spread the infection. Patient access to current treatments, and compliance with the lengthy dosing regimens is a global challenge. MDR-TB is more difficult to treat, and options for the emerging XDR-TB are even more limited. There is an urgent unmet medical need to develop novel anti-TB agents to successfully combat emerging resistance. Recent progress has been achieved by the repurposing and redosing of known anti-TB drugs, but there are no new drug classes in clinical trials that act by novel mechanisms of action.The protein "Filamenting temperature-sensitive mutant Z" (FtsZ) is highly conserved among all types of bacteria, including Mycobacterium species that cause TB. It is an essential cell division protein with both GTPase and polymerisation activities. FtsZ subunits polymerise into protofilaments to form a dynamic ring-like structure called the Z-ring that functions as a scaffold for the assembly of the divisome, a multiprotein complex that causes contraction of the Z-ring, finally resulting in septum formation and cell division. Abnormalities in FtsZ block the Z-ring functions, leading to an elongated, filamentous phenotype, inhibition of cell division and subsequent cell death. Although FtsZ inhibitors have been identified, most are derived from natural products and there is a general lack of structural information, with many of the inhibitors classified as "false positives".We have purified M. tuberculosis (MTb) FtsZ in large amounts for biochemical, biophysical and structural studies. In collaboration with the Monash NMR screening facility, we identified 38 fragment hits against FtsZ. We established biophysical (DSF, MST) and biochemical (GTPase, right-angle scattering) assays to characterise its GTPase activity and polymerisation/depolymerisation dynamics. Uniquely, we have determined the crystal structure of FtsZ in complex with fragment hits and established their mechanism of action. We have evaluated the fragment binding sites with respect to fragment growth and merging and we are ready to initiate SAR-by-catalogue and custom-made optimisation of the hits to develop more potent analogues.In this project we aim to develop and characterise potent inhibitors of MTb FtsZ as a new class of antibiotic for the treatment of tuberculosis infections. We anticipate that such compounds will be a welcome addition to the treatment options available for TB and its drug-resistant variants. This collaborative drug discovery project addresses a clear unmet medical need. Successful completion of this project will underpin a broader collaborative drug discovery program including other UCL partners.