Targeting Bacterial Signaling Cascades as a Novel Antibiotic Strategy

背景和退伍军人意义：病原体如分枝杆菌和金黄色葡萄球菌， 不仅本身具有抗生素耐药性，而且正在迅速获得多重耐药性特征。二十亿人 目前感染结核分枝杆菌，越来越多的比例是耐药的临床 治疗。耐甲氧西林金黄色葡萄球菌（MRSA）是医疗保健的巨大负担， 万古霉素抗性菌株越来越成问题。这些耐药感染是流行于 发展中国家和地区的常规军事存在，增加了退伍军人的负担 行政保健系统。此外，MRSA感染是VA医院的主要问题， 是时间、资源和生命的巨大代价。需要新的有效抗生素， 治疗由这些耐药生物体引起的感染。 新的抗生素靶点：蛋白激酶和磷酸酶在转导细胞和 环境信号引发生长和分裂或对压力和环境变化作出反应。 人类的磷酸化信号已经被很好地研究过了，但是细菌的信号却鲜为人知，也没有抗生素 针对这些途径存在。青霉素结合和丝氨酸/苏氨酸相关（PASTA）激酶是 存在于放线菌和厚壁菌门中的独特跨膜激酶。PASTA激酶是必需的， 毒力，使它们成为有吸引力的药物靶点。MRSA的基因敲除和药物抑制 MRSA或结核增加β-内酰胺类药物的敏感性。此外，同源S/T的遗传缺失 MRSA中的磷酸酶增加这种协同作用并降低体内毒力。抑制激酶和 磷酸酶增强β-内酰胺协同作用是抗生素开发的新方法。 该奖项的目标：该奖项的目标是1）提供时间和资源，以促进博士。 Wlodarchak从一个受指导的科学家转变为VA的独立翻译研究员 系统和2）开发针对MRSA和结核病中新抗生素靶标的先导化合物。的 这里测试的中心假设是对丝氨酸/苏氨酸的几个节点的协同攻击 磷酸化信号级联将是一种有效的药理学策略， 抵抗力的发展。这一假设将通过表征来自生物化学物质的命中来测试。 磷酸酶筛选，开发MRSA激酶抑制剂，并进行第一次全球转录组学和 磷酸化蛋白质组学筛选在该途径上的药物应激下的MRSA。 预期成果和影响：在完成这项奖励后，预计Wlodarchak博士将 过渡到一个完全独立的VA研究者与几个良好表征的先导化合物， MRSA和结核病，并导致在这一途径中的其他潜在目标。这项研究将提供 翻译药物开发项目的竞争性VA Merit奖的初步数据。这将具有 不仅对沃达查克博士的职业生涯产生了积极的影响，而且通过提供机械的 深入了解磷酸化信号如何控制毒力，并提供有形的化合物， 成为临床药物。这将有助于提供遏制当前抗生素耐药性危机的选择， 提高患者生存率和生活质量。

Background and Veteran significance: Pathogens such as mycobacteria and Staphylococcus aureus, are not only intrinsically antibiotic resistant, but are rapidly acquiring multi-drug resistant traits. Two billion people are currently infected with Mycobacterium tuberculosis, and an increasing proportion are resistant to clinical treatments. Methicillin resistant Staphylococcus aureus (MRSA) is a massive burden in healthcare, and vancomycin resistant strains are increasingly problematic. These resistant infections are endemic to developing countries and areas with a regular military presence, causing an increased burden for the Veterans Administration healthcare system. Furthermore, MRSA infections are a major concern in VA hospitals and are a significant cost in time, resources, and lives. New effective antibiotics are needed, and development against novel targets is needed to treat infections caused by these resistant organisms. Novel antibiotic targets: Protein kinases and phosphatases are critical in transducing cellular and environmental signals to trigger growth and division or to respond to stress and environmental changes. Human phosphorylation signaling is well studied, but bacterial signaling is less known and no antibiotics targeting these pathways exist. Penicillin-binding And Serine/Threonine Associated (PASTA) kinases are unique transmembrane kinases present in Actinobacteria and Firmicutes. PASTA kinases are necessary for virulence, making them attractive drug targets. Genetic knockouts in MRSA and pharmacological inhibition in MRSA or tuberculosis increase β-lactam susceptibility. Furthermore, genetic deletion of the cognate S/T phosphatase in MRSA increases this synergy and decreases in vivo virulence. Inhibiting both kinases and phosphatases to enhance β-lactam synergy is a novel approach to antibiotic development. Goals of this award: The goals of this award are to 1) provide time and resources to facilitate Dr. Wlodarchak’s transition from a mentored scientist to an independent translational investigator in the VA system and to 2) develop lead compounds against novel antibiotic targets in MRSA and tuberculosis. The central hypothesis tested here is that a coordinated attack on several nodes of the serine/threonine phosphorylation signaling cascade will be an effective pharmacological strategy with low likelihood of resistance development. This hypothesis will be tested by characterizing hits from a biochemical phosphatase screen, developing MRSA kinase inhibitors, and performing the first global transcriptomic and phosphoproteomic screen on MRSA under pharmacologic stress on this pathway. Expected outcomes and impacts: Upon completion of this award, it is expected that Dr. Wlodarchak will transition to a fully independent VA investigator with several well-characterized lead compounds against MRSA and tuberculosis and leads on other potential targets in this pathway. This research will provide preliminary data for a competitive VA Merit award for a translational drug development project. This will have a positive impact on not only Dr. Wlodarchak’s career but also on Veterans’ health by providing mechanistic insight on how phosphorylation signaling controls virulence and by providing tangible compounds that may become clinical drugs. This will help provide options to stifle the current antibiotic resistance crisis and thus improve patient survival and quality of life.