A novel "shock and kill" strategy for eliminating Mtb persisters in the CD4 T-cell-deficient host

一种新的“电击杀灭”策略，用于消除 CD4 T 细胞缺陷宿主中的 Mtb 持续存在

基本信息

批准号：
9208740
负责人：
Petros C Karakousis
金额：
$ 20.25万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-02-01 至 2019-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9208740
关键词：
Adherence Antibiotics Antitubercular Agents Bacillus (bacterium)Bacteria Biological Assay CD4 Positive T Lymphocytes Cells Cellular Immunity Cessation of life Chronic Chronic Phase Collaborations Complement Data Disease Dose Drug Interactions Drug Tolerance Drug resistance in tuberculosis Enzymes Evaluation Exhibits Extreme drug resistant tuberculosis Goals Growth Guanosine HIV Hypoxia Immune In Vitro Infection Knockout Mice Knowledge Libraries Lung Medical Metabolism Microbiology Molecular Molecular Biology Molecular Genetics Multi-Drug Resistance Mus Mycobacterium tuberculosis Nutrient Patients Persons Pharmaceutical Preparations Phenotype Play Predisposition Pyrazinamide RELA gene Recombinants Regimen Resistance development Rifampin Role Safety Shock Solubility Starvation Stress Testing Time Toxic effect Tuberculosis animal model development bactericide cell killing co-infection cytotoxicity drug development extensive drug resistance improved in vitro Assay in vivo inhibitor/antagonist isoniazid killings mouse model multidisciplinary mutant novel overexpression prevent public health relevance reconstitution relapse risk response scaffold small molecule inhibitor stability testing synergism treatment duration tuberculosis drugs tuberculosis treatment

项目摘要

DESCRIPTION (provided by applicant) The lengthy and complicated multidrug therapy currently available to treat active tuberculosis (TB) infection has contributed to medical non-adherence and the emerging problems of multidrug-resistant (MDR)- and extensively drug-resistant (XDR)-TB, which are particularly deadly in the setting of HIV co-infection. In addition, persons with HIV are at increased risk for relapse even upon successful completion of anti-TB treatment. The prolonged therapy required to eradicate TB infection is believed to reflect the ability of Mycobacterium tuberculosis (Mtb) to persist within the host in a non-replicating state characterized by antibiotic tolerance to bactericidal drugs, which predominantly target actively dividing tubercle bacilli. Hyperphosphorylated guanosine ((p)ppGpp), which is synthesized by the stringent response enzyme RelA, plays an important role in bacterial growth restriction and antibiotic tolerance. RelA is essential for long-term Mtb survival during various in vitro and in vivo stress conditions. However, whether RelA is required for Mtb persistence in the host lacking cell-mediated immunity represents an important knowledge gap. Recently, we found that relA-deficient Mtb exhibits continued replication and metabolism during stress conditions, ultimately leading to bacillary death, as well as enhanced susceptibility to isoniazid during nutrient starvation and in the lungs of chronically infected mice. In collaboration with GlaxoSmithKline's Tres Cantos Open Lab for Diseases of the Developing World (GSK-DDW), we have screened a library of over 2 million compounds in RelA-inhibition and whole-cell killing assays, and performed preliminary cytotoxicity studies. A total of 178 RelA inhibitor candidates, representing 18 unique scaffolds, were identified and 12 of 39 compounds tested so far exhibit RelA-specific activity and synergy with isoniazid against nutrient-starved Mtb. The central hypothesis of this proposal is that releasing the molecular "brakes" on Mtb growth and replication through inhibition of the stringent response enzyme RelA is a safe and effective strategy to prevent Mtb persistence, decrease drug tolerance, and enhance susceptibility to bactericidal drugs in the CD4 T-cell-deficient host. Specifically, we will screen hits using whole-cell nutrient starvation assays. Target-specific candidates fulfilling physicochemical and toxicity criteria will be prioritized for safety/PK studis in immune-competent C57BL/6J mice. Up to 3 favorable RelA inhibitor candidates will be tested in dose-ranging studies alone and in combination with the first-line regimen in this mouse model, which is the background strain for the Cd4tm1Mak mice, which will be used to validate RelA as a target for Mtb persisters in the CD4 T-cell-deficient host. Our novel "shock and kill" strategy is expected to shorten the duration of treatment for drug-susceptible and drug-resistant TB in the HIV-infected and HIV-uninfected host. In addition to improving medical adherence and reducing the potential for the development of drug resistance, an abbreviated drug regimen to treat active TB would be particularly useful in HIV co-infected patients, since drug- drug interactions and immune reconstitution may complicate the concurrent management of both infections.

描述（由申请人提供）目前用于治疗活动性结核病 (TB) 感染的漫长而复杂的多药治疗导致了医疗不依从性以及新出现的耐多药 (MDR) 和广泛耐药 (XDR) 结核病问题，这些问题尤其致命此外，即使成功完成抗结核治疗，艾滋病毒感染者的复发风险也增加，根除结核感染所需的长期治疗被认为反映了其能力。结核分枝杆菌 (Mtb) 其在宿主体内以非复制状态持续存在，其特征是对杀菌药物的抗生素耐受性，其主要针对活跃分裂的结核杆菌（由严格反应酶 RelA 合成），在其中发挥着重要作用。细菌生长限制和抗生素耐受性对于 Mtb 在各种体外和体内应激条件下的长期生存至关重要。然而，在缺乏细胞介导的免疫的宿主中，RelA 是否是 Mtb 持久存在所必需的，这代表了一个重要的知识空白。最近，我们发现 relA 缺陷的 Mtb 在应激条件下表现出持续的复制和代谢，最终导致细菌死亡。与葛兰素史克 (GlaxoSmithKline) 的 Tres Cantos 发展中国家疾病开放实验室合作，在营养饥饿期间和慢性感染小鼠的肺部对异烟肼的敏感性增强。 (GSK-DDW)，我们在 RelA 抑制和全细胞杀伤试验中筛选了超过 200 万种化合物，并进行了初步细胞毒性研究，鉴定出总共 178 种 RelA 抑制剂候选物，代表 18 种独特的支架。迄今为止测试的 39 种化合物表现出 RelA 特异性活性，并与异烟肼对抗营养匮乏的 Mtb 具有协同作用。该提案的中心假设是释放分子。通过抑制严格反应酶 RelA 来抑制 Mtb 的生长和复制是一种安全有效的策略，可以防止 Mtb 持续存在，降低药物耐受性，并增强 CD4 T 细胞缺陷宿主对杀菌药物的敏感性。将使用全细胞营养饥饿测定筛选符合理化和毒性标准的靶标候选物，将优先在免疫能力强的 C57BL/6J 小鼠中进行安全/药代动力学研究。 3 种有利的 RelA 候选抑制剂将在剂量范围研究中单独进行测试，并与该小鼠模型中的一线方案相结合，该小鼠模型是 Cd4tm1Mak 小鼠的背景品系，将用于验证 RelA 作为治疗的靶点Mtb 在 CD4 T 细胞缺陷宿主中持续存在，我们的新型“休克和杀伤”策略有望缩短 HIV 感染者和耐药结核病的治疗时间。未感染艾滋病毒的宿主除了提高医疗依从性和减少产生耐药性的可能性之外，治疗活动性结核病的简化药物治疗方案对于艾滋病毒合并感染的患者特别有用，因为药物间相互作用和免疫重建可能会发生。使两种感染的同时管理变得复杂。