Inhibitors of S. aureus bNOS for adjunctive therapy

用于辅助治疗的金黄色葡萄球菌 bNOS 抑制剂

基本信息

批准号：
8393335
负责人：
Donald T Moir
金额：
$ 29.05万
依托单位：
MICROBIOTIX, INC
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-07-01 至 2014-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8393335
关键词：
Animal Model Anti-Bacterial Agents Antibiotic Resistance Antibiotic Therapy Antibiotics Bacillus (bacterium)Bacillus anthracis Bacteria Bacterial Infections Biochemical Biological Assay Blood Circulation Cell Death Cells Cessation of life Chemicals Chemosensitization Communicable Diseases Community Hospitals Development Dose Drug Design Drug resistance Effectiveness Enzymes Fluorescent Probes Fluoroquinolones Gene Deletion Genes Genus staphylococcus Glycopeptide Antibiotics Glycopeptides Goals Growth Hospitals Host Defense Hydrogen Peroxide Hydroxyl Radical Immune In Vitro Infection Inhibitory Concentration 50 Isoenzymes Lactams Lead Libraries Mediating Membrane Monobactams Nitric Oxide Nitric Oxide Synthase Nosocomial Infections Oxidative Stress Parents Patients Pharmaceutical Preparations Phase Production Reaction Reactive Oxygen Species Reporter Research Resistance Resistance development Respiratory Burst Screening procedure Site Skin Skin Tissue Sodium Chloride Soft Tissue Infections Specificity Staphylococcus aureus Superoxide Dismutase System Therapeutic Toxic effect Toxicology Urban Hospitals bactericide base beta-Lactam Resistance catalase clinically relevant community setting cytotoxicity improved in vivo inhibitor/antagonist innovation killings methicillin resistant Staphylococcus aureus neutrophil novel pathogen preclinical study pressure resistant strain small molecule soft tissue

项目摘要

DESCRIPTION (provided by applicant): Staphylococcus aureus is a major cause of community and hospital-acquired infections of the skin, soft tissue, and bloodstream. The recent dramatic increase in occurrence of strains resistant to beta-lactam antibiotics (MRSA) has reduced therapeutic options significantly, but most MRSA strains remain sensitive to other bactericidal antibiotics in the fluoroquinolone (FQ) and glycopeptide (GP) classes. Recent research has demonstrated that bactericidal antibiotics stimulate the production of reactive oxygen species (ROS) in bacteria, which contribute to cell death in a manner similar to the host immune oxidative burst. Alleviation of this oxidative stress by the production of nitric oxide (NO) by bacterial NO synthase (bNOS) enhances the survival of several bacterial species, including staphylococci, to antibacterial therapy and to neutrophil killing. Consequently, small molecule bNOS inhibitors will provide an adjunctive therapeutic approach to bolster the effectiveness of antibiotics and neutrophils against MRSA and potentially reduce the selective pressure for development of drug-resistance by increasing the duration of effective circulating levels of antibiotic. Substantial differences between mammalian and bacterial NOS enzymes indicate that selective inhibition of bNOS is feasible. The overall goal of this project is to discover and develop drugs that increase the efficacy of clinically relevant bactericidal antibiotics against pathogenic staphylococci such as MRSA by specifically inhibiting bacterial NO production. Our strategy is to identify small molecule bNOS inhibitors and to develop them into innovative adjunctive therapies that increase the bactericidal activity of FQ, GP, and beta-lactam antibiotics. In preliminary studies, we established proof of concept for bNOS as a novel target for adjunctive therapy by demonstrating that the growth and viability of strains of S. aureus, B. subtilis, and B. anthracis carrying deletions of the bNOS gene were more sensitive to several bactericidal antibiotics than were their wild-type parents. NO in bacterial cells was shown to activate catalase, induce sodA (superoxide dismutase), suppress the Fenton reaction (production of reactive hydroxyl radicals from H2O2), and rescue cells from ROS generated by bactericidal drugs and by the immune oxidative burst. These results indicate that bactericidal drugs can be potentiated by targeting bacterial systems that reduce ROS damage, such as bNOS. In Phase I, we will construct and optimize luminescent, fluorescent, and biochemical primary and secondary screening assays for bNOS inhibitors, apply them to libraries representing >300,000 discrete chemical compounds, confirm the hits, and validate them as potent, selective potentiators of several antibiotics and neutrophils vs. MRSA and other drug- resistant species. The most potent, broadest acting bNOS inhibitors will be characterized to eliminate compounds with off-target activity vs. the 3 mammalian NOS isozymes and cytotoxicity. In Phase II, we will develop the most promising validated hits into lead compounds by optimizing their activity and specificity using rational drug design and evaluate them for efficacy and toxicity in animal models of infection. PUBLIC HEALTH RELEVANCE: This research is aimed at discovering new drugs that increase the effectiveness of existing antibiotics against pathogenic staphylococcal species, including MRSA. New drugs that inhibit the bacterial production of nitric oxide will cripple the pathogen's ability to survive antibiotic therapy. These new drugs will prolong the duration of effective concentrations of existing antibiotics, thus reducing selection for resistance and improving therapy.

描述（由申请人提供）：金黄色葡萄球菌是社区和医院获得性皮肤、软组织和血液感染的主要原因。最近，对 β-内酰胺抗生素 (MRSA) 耐药的菌株的出现急剧增加，显着减少了治疗选择，但大多数 MRSA 菌株仍然对氟喹诺酮 (FQ) 和糖肽 (GP) 类其他杀菌抗生素敏感。最近的研究表明，杀菌抗生素会刺激细菌中活性氧（ROS）的产生，从而以类似于宿主免疫氧化爆发的方式导致细胞死亡。通过产生一氧化氮 (NO) 缓解这种氧化应激细菌一氧化氮合酶（bNOS）可增强多种细菌（包括葡萄球菌）的存活率，从而实现抗菌治疗和杀灭中性粒细胞。因此，小分子 bNOS 抑制剂将提供一种辅助治疗方法，以增强抗生素和中性粒细胞对抗 MRSA 的有效性，并可能通过增加抗生素有效循环水平的持续时间来降低耐药性发展的选择性压力。哺乳动物和细菌 NOS 酶之间的显着差异表明选择性抑制 bNOS 是可行的。该项目的总体目标是发现和开发药物，通过特异性抑制细菌一氧化氮的产生，提高临床相关杀菌抗生素针对 MRSA 等致病性葡萄球菌的功效。我们的策略是识别小分子 bNOS 抑制剂，并将其开发为创新的辅助疗法，以提高 FQ、GP 和 β-内酰胺抗生素的杀菌活性。在初步研究中，我们通过证明携带 bNOS 基因缺失的金黄色葡萄球菌、枯草芽孢杆菌和炭疽芽孢杆菌菌株的生长和活力对 bNOS 更敏感，建立了 bNOS 作为辅助治疗新靶点的概念验证。比其野生型亲本具有多种杀菌抗生素。细菌细胞中的一氧化氮被证明可以激活过氧化氢酶，诱导 sodA（超氧化物歧化酶），抑制芬顿反应（从 H2O2 产生活性羟基自由基），并从杀菌药物和免疫氧化爆发产生的 ROS 中拯救细胞。这些结果表明，可以通过针对减少 ROS 损伤的细菌系统（例如 bNOS）来增强杀菌药物的效果。在第一阶段，我们将构建和优化 bNOS 抑制剂的发光、荧光和生化初级和二级筛选测定，将它们应用于代表超过 300,000 种离散化合物的文库，确认命中，并验证它们作为多种抗生素的有效、选择性增强剂以及中性粒细胞与 MRSA 和其他耐药物种的比较。最有效、最广泛作用的 bNOS 抑制剂将消除与 3 种哺乳动物 NOS 同工酶相比具有脱靶活性和细胞毒性的化合物。在第二阶段，我们将通过合理的药物设计优化其活性和特异性，开发最有希望的经过验证的先导化合物，并评估它们在感染动物模型中的功效和毒性。公共健康相关性：本研究旨在发现新药，提高现有抗生素对抗致病性葡萄球菌（包括 MRSA）的有效性。抑制细菌产生一氧化氮的新药将削弱病原体在抗生素治疗中生存的能力。这些新药将延长现有抗生素的有效浓度持续时间，从而减少耐药性的选择并改善治疗。