Inhibitors of Nitro Drug Targets as Antimicrobials against Trichomonas Vaginalis

 描述(申请人提供)：单细胞寄生虫阴道毛滴虫是全球最流行的非病毒性性传播感染，每年约有1/4亿人感染滴虫病。滴虫病是美国最常见的寄生虫感染，每年的发病率估计为500万例。阴道毛滴虫感染的频率和抗药性临床分离株数量的增加突显了开发新的化疗策略和药物来消除这种寄生虫的必要性。滴虫病患者中约有5%对5-硝基咪唑(5NI)类药物甲硝唑(Mz)和替硝唑(Tz)产生抗药性。5NI是一种前药，在厌氧微生物内通过选择性还原激活，这一步骤对杀死细菌至关重要，但也是微生物产生耐药性的原因 失去了减少5NIS的能力。在激活时，5NI通过与重要的靶分子形成共价、灭活的加合物来杀死细胞，这些加合物的定义很差。这项研究提出的长期目标是确定对阴道毛滴虫生存至关重要的5NI药物的微生物靶点，并利用这些信息开发对治疗耐药滴虫病有效的蛋白质靶点的抑制剂。拟议的研究有五个具体目标。前两个目标将在R21阶段实现，后三个目标将在R33阶段实现。在目标1中，我们将使用点击化学和质谱学的策略在阴道毛滴虫模型菌株中识别硝基药物靶标。在目标2中，我们将使用反向遗传学来确定哪些蛋白质靶标对阴道毛滴虫的生存是必不可少的，以测试该基因的消除是否具有致命性。在过渡到R33阶段后，在目标3中，我们将确定在广泛的地理和遗传多样性的阴道毛滴虫临床分离株中常见的硝基药物靶点。在目标4中，我们将确定目标3中确定的选定的共同目标是否需要寄生虫生存。在目标5中，我们将为最有希望的硝基药物靶点开发抑制剂，并在体外和在 活着。最后，我们的目标是验证新的药物靶点并开发针对这些靶点的新抑制剂，作为有效治疗耐甲氧西林滴虫病的候选药物。拟议的研究还将为使用相同的方法识别和验证新的抗菌素来治疗其他厌氧寄生虫病铺平道路，例如贾第鞭毛虫病和阿米巴病，Mz是治疗这些感染的主要药物。

 DESCRIPTION (provided by applicant): The unicellular parasite Trichomonas vaginalis is responsible for the most prevalent, non-viral, sexually-transmitted infection worldwide, with approximately 1/4 billion people contracting trichomoniasis annually. Trichomoniasis is the most common parasitic infection in the US and has an annual incidence estimated at 5 million cases. The frequency of infection and an increase in the number of drug resistant clinical isolates of T. vaginalis underscore the need to develop new chemotherapeutic strategies and drugs that eliminate the parasite. Resistance to the only drugs licensed for therapy, the 5-nitroimidazole (5NI) drugs metronidazole (Mz) and tinidazole (Tz), occurs in approximately 5% of cases of trichomoniasis. 5NIs are prodrugs that are activated by selective reduction inside anaerobic microbes, a step that is critical for killing, but also responsible for resistance when the microbe loses its capacity to reduce 5NIs. Upon activation, 5NIs kill the cell by forming covalent, inactivating adducts with essential target molecules, which are poorly defined. The long-term goal of the research proposed here is to define microbial targets of 5NI drugs that are essential for the survival of T. vaginalis and leverage this information to develop inhibitors of protein targets that are effective in the treatment of drug resistant trichomoniasis. The proposed studies have five Specific Aims. The first two aims will be achieved during the R21 phase and the latter three during the R33 phase. In Aim 1, we will identify nitro drug targets in model T. vaginalis strains using a strategy that employs click chemistry and mass spectrometry. In Aim 2, we will determine which protein targets are essential for T. vaginalis viability using reverse genetics to test whether elimination of the gene is lethal. After transitioning to the R33 phase, in Aim 3 we will identify nitro drug targets that are common to a broad range of geographically and genetically diverse T. vaginalis clinical isolates. In Aim 4, we will determine whether selected, common targets identified in Aim 3 are required for parasite viability. In Aim 5 we will develop inhibitors for the most promising nitro drug targets and examine inhibitor activity in vitro and in vivo. In the end, we aim to have validated novel drug targets and developed new inhibitors against these targets as candidates for effective treatment of Mz resistant trichomoniasis. The proposed research will also pave the way for using the same approach to identify and validate new antimicrobials to treat other anaerobic parasitic infections, such as giardiasis and amoebiasis, for which Mz is the primary drug used for therapy.