The impact of pyrazinamide on metabolism in Mycobacterium tuberculosis

项目摘要 吡津酰胺(PZA)是结核病(TB)一线治疗的关键成分，因为它具有显著的 减少复发率和治疗持续时间。PZA在体外活性较弱，但在体内具有很强的杀菌作用 由于其对缓慢和非复制的结核分枝杆菌种群具有出色的活性 对大多数其他结核病药物具有表型耐受性。了解PZA体内杀菌活性的基础 代表了结核病药物研发中最重要的未得到满足的需求之一。而PZA的行动模式 仍在调查中，我们对控制易感性和耐药性的因素的了解有所进展 这在最近几年是相当大的。PZA是一种前体药物，必须转化为吡嗪酸的活性形式 (POA)是由结核分枝杆菌酰胺酶pncA引起的，而pncA功能丧失突变至少占70% 临床耐药。其他与抗药性有关的遗传变异也已被描述，但大多数 没有在同基因菌株中进行评估，以确定它们在耐药性中的作用，也没有进行过评估 在感染动物模型中与PZA耐药性的相关性。重要的是，PZA缺乏抗结核活性 裸鼠，表明T细胞介导的免疫在疗效中起关键作用。与这些一致 我们的实验室和其他人的观察表明，PZA在休息时对结核分枝杆菌没有活性 巨噬细胞，但对干扰素-ɣ激活的巨噬细胞的细菌杀灭有贡献。主机- 与干扰素-ɣ刺激相关的依赖应激源，如暴露在低pH值、营养限制 和活性氧物种，被认为是PZA作用的贡献者。我们最近的研究表明 揭示了特定结核分枝杆菌应激反应在PZA条件性易感性中的核心作用。穿过 在这些研究中，我们已经确定了一个调节PZA易感性的功能网络，当表达 相应的基因会发生改变。我们还确定了有助于PZA行动的特定宿主因素 在巨噬细胞和感染的小鼠体内。基于这些发现，我们已经确定了支持药物作用的方法 在PZA通常缺乏活动的情况下。通过这一提议，我们的目标是将小说 PZA抗性的分子机制，确定调节PZA敏感性的宿主因素，以及 评估以宿主和微生物为靶点的PZA增强的机会。这些研究将推动我们的 了解结核分枝杆菌对PZA的敏感性和耐药机制。作为一名 相当大比例的结核病是由T细胞反应受损引起的，这是至关重要的 了解这些反应如何与PZA疗效相关，以及我们如何使用这些信息来优化PZA 在免疫力受损的情况下采取行动。

Project Summary Pyrazinamide (PZA) is a critical component of first-line tuberculosis (TB) therapy because it has dramatically reduced relapse rates and treatment duration. PZA is weakly active in vitro, but potently sterilizing in vivo, due to its outstanding activity against slow and non-replicating populations of Mycobacterium tuberculosis that are phenotypically tolerant to most other TB drugs. Understanding the basis for the in vivo sterilizing activity of PZA represents one of the most important unmet needs in TB drug discovery. While the mode of action of PZA remains under investigation, our knowledge of factors that govern susceptibility and resistance has advanced considerably in recent years. PZA is a pro-drug that must be converted to the active form of pyrazinoic acid (POA) by the M. tuberculosis amidase PncA, and loss-of-function mutations in pncA account for at least 70% of clinical resistance. Other genetic variations associated with resistance have been described, but most have not been evaluated in isogenic strains to determine their role in resistance, and have not been assessed for association with PZA resistance in animal models of infection. Importantly, PZA lacks antitubercular activity in athymic nude mice, indicating a key role for T cell-mediated immunity in efficacy. Consistent with these observations, our lab and others have shown that PZA is inactive against M. tuberculosis in resting macrophages, but contributes to bacterial killing in macrophages activated by interferon-gamma (IFN-ɣ). Host- dependent stressors that are associated with IFN-ɣ stimulation, such as exposure to low pH, nutrient limitation and reactive oxygen species, have been implicated as contributors to PZA action. Our recent studies have revealed a central role for specific M. tuberculosis stress responses in PZA conditional susceptibility. Through these studies, we have identified a network of functions that modulates PZA susceptibility when expression of the corresponding genes is altered. We have also identified specific host factors that contribute to PZA action in macrophages and in infected mice. Based on these findings, we have identified means to bolster drug action under conditions where PZA typically lacks activity. Through this proposal, we aim to characterize novel molecular mechanisms for PZA resistance, determine host factors that modulate PZA susceptibility, and assess opportunities in host- and microbe-targeted PZA potentiation. These studies will advance our understanding of mechanisms that govern PZA susceptibility and resistance of M. tuberculosis. As a substantial proportion of TB disease results from impaired T cell responses, it is of fundamental importance to understand how these responses relate to PZA efficacy and how we can use this information to optimize PZA action in the context of impaired immunity.