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

• 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
• 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

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(申请人提供)：金黄色葡萄球菌是社区和医院获得性皮肤、软组织和血流感染的主要原因。最近对β-内酰胺类抗生素(MRSA)耐药的菌株急剧增加，大大减少了治疗选择，但大多数MRSA菌株对氟喹诺酮(FQ)和糖肽(GP)类其他杀菌抗生素仍然敏感。最近的研究表明，杀菌抗生素能刺激细菌中活性氧物种(ROS)的产生，从而导致细胞死亡，其方式类似于宿主免疫氧化爆发。通过产生一氧化氮(NO)来缓解这种氧化应激 通过细菌一氧化氮合酶(Bnos)增强包括葡萄球菌在内的几种细菌对抗菌治疗和中性粒细胞的杀伤作用。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，并可能通过延长抗生素有效循环水平的持续时间来降低耐药发展的选择压力。哺乳动物和细菌的一氧化氮合酶的显著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通过特别抑制细菌NO产生来提高临床相关杀菌抗生素对致病葡萄球菌(如MRSA)的疗效的药物。我们的战略是确定小分子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，我们证明了携带bnos基因缺失的金黄色葡萄球菌、枯草杆菌和炭疽杆菌的生长和活力对几种杀菌抗生素比它们的野生型亲本更敏感，从而证明了bnos作为辅助治疗的新靶点的概念。细菌细胞中的NO可以激活过氧化氢酶，诱导超氧化物歧化酶(SODA)，抑制Fenton反应(从过氧化氢中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，杀菌药物可以通过靶向减少ROS损害的细菌系统来增强，例如BNOS。在第一阶段，我们将构建和优化BNOS抑制剂的发光、荧光和生化一次和二次筛选方法，将它们应用于代表&gt；300,000种离散化合物的库，确认HITS，并验证它们作为几种抗生素和中性粒细胞与MRSA和其他耐药物种的有效、选择性增强剂。最有效、最广泛的作用最广的bnos抑制剂的特点是消除具有非靶标活性的化合物，而不是哺乳动物的3种一氧化氮合酶同工酶和细胞毒性。在第二阶段，我们将通过合理的药物设计优化它们的活性和特异性，将最有希望的有效HITS开发成先导化合物，并在动物感染模型中评估它们的有效性和毒性。 公共卫生相关性：这项研究的目的是发现新的药物，以提高现有抗生素对包括MRSA在内的致病葡萄球菌的有效性。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，从而减少对耐药性的选择，并改善治疗。