Targeting of LOS for Treatment of Antibiotic-Resistant Neisseria gonorrhoeae

LOS 靶向治疗抗生素耐药性淋病奈瑟菌

• 批准号: 10363529
• 负责人: Gary A Jarvis
• 金额: --
• 依托单位: VETERANS AFFAIRS MED CTR SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10363529
• 关键词: Amino Acids; Anabolism; Anti-Bacterial Agents; Antibiotic Therapy; Antibiotics; Antimicrobial Cationic Peptides; Azithromycin; Bacteria; Binding Proteins; Biological Assay; Canada; Cardiovascular system; Cations; Cefixime; Ceftriaxone; Cells; Centers for Disease Control and Prevention (U.S.); Cephalosporins; Cervical; Clinical; Complement; Country; Cytolysis; DNA Binding; Data; Denmark; Drug Kinetics; Drug-resistant Neisseria Gonorrhoeae; Ectopic Pregnancy; Environment; Enzymes; Epithelial Cells; Evaluation; Exposure to; Female; Generations; Gonorrhea; Gram-Negative Bacteria; HIV; Healthcare; Healthcare Systems; Hemolysis; Human; Immune response; In Vitro; Incidence; Infection; Infertility; Inflammation; Inflammatory; Innate Immune System; Investigation; Japan; Lipid A; Measurable; Mediating; Membrane; Military Personnel; Mission; Modeling; Morbidity - disease rate; Multi-Drug Resistance; Neisseria gonorrhoeae; Outcome; Patient Care; Pelvic Inflammatory Disease; Peptides; Predisposition; Proteolysis; Public Health; Reporting; Research; Resistance; Resistance development; Risk Behaviors; Sexually Transmitted Diseases; Testing; Therapeutic; Time; Toxic effect; Vaccines; Variant; Veterans; Woman; Work; active duty; antimicrobial; bactericide; base; chronic pelvic pain; cytokine; cytotoxicity; design; efficacy testing; efflux pump; high risk; in vitro Assay; in vitro testing; in vivo; in vivo evaluation; inhibitor; interest; lead candidate; lipid biosynthesis; lipooligosaccharide; men; military service; military veteran; mouse model; neutrophil; novel; novel therapeutics; prevent; reproductive tract; resistant strain; transmission process

Infections due to N. gonorrhoeae are a major cause of morbidity with an estimated 850,000 cases in the U.S. and 87 million cases worldwide annually. Within the VA Health Care System, cases of gonorrhea increased between 2013 and 2017 with the total number in that time period at 10,587. The most serious sequelae are suffered by infected women as gonococci ascend to the upper reproductive tract and cause pelvic inflammatory disease in 10-20% of women with infections, which encompasses a wide range of inflammatory conditions and often leads to chronic pelvic pain, infertility, and ectopic pregnancy. There is no vaccine to N. gonorrhoeae and a great need for new antibiotics due to the alarming rise in multidrug-resistance (MDR), which is making emergence of untreatable gonococcal infections a real prospect. Currently only ceftriaxone and azithromycin are recommended for first-line therapy, and clinical isolates resistant to both of those antibiotics have been reported in countries including Denmark, Canada, and Japan. Thus, there is a compelling need for new antimicrobials for gonococcal infections. Our studies to date of N. gonorrhoeae lipooligosaccharide (LOS) and the human innate immune system have shown that the lipid A portion of LOS is the primary inducer of cytokine-mediated inflammation and investigations by others have shown that the lipid A also facilitates gonococcal infection. These data led us to the concept that targeting lipid A biosynthesis would be an effective approach to combating N. gonorrhoeae infections. We recently reported that inhibition of LpxC, the enzyme that catalyzes the second step of lipid A biosynthesis, was bactericidal for nine multidrug-resistant and human challenge strains of gonococci and reduced cytokine induction without apparent human cell cytotoxicity. From the LpxC inhibitor data, we postulated that membrane disruption due to the inhibition of LOS biosynthesis was lethal for gonococci. To investigate this, we recently evaluated the bactericidal potential of a 12 amino acid cell-penetrating peptide (CPP) for MDR and human challenge strains of N. gonorrhoeae and found that the CPP penetrated the bacterial membrane and was bactericidal for all nine MDR and human challenge strains of gonococci tested. Importantly, no apparent resistance to the CPP developed in surviving bacteria as susceptibility was the same in bacteria from colonies after exposure to CPP and then retreated. Further, the CPP reduced inflammatory cytokine induction and prevented bacterial cell invasion of cervical epithelial cells in the absence of measurable cell cytotoxicity. These novel data highlight LpxC and CPP as promising antimicrobials for N. gonorrhoeae and strongly support the hypothesis of this application that inhibiting the biosynthesis of lipid A components with LpxC inhibitors and disrupting outer membrane integrity with CPP will impact bacterial viability and host response to N. gonorrhoeae infection in vitro and in vivo, which will have a therapeutic impact on infection outcomes. This project is focused on optimizing and testing the efficacy of the CPP and LpxC inhibitor in relevant in vitro assays of bactericidal activity, cytokine induction, hemolysis, and cell cytotoxicity. Mechanistic studies will include investigations of DNA binding, cell permeabilization, proteolysis resistance, protein binding and the effect of the MtrCDE, MacAB and NorM gonococcal efflux pumps. The lead candidate CPP and LpxC inhibitor identified in vitro will be tested for in vivo efficacy, pharmacokinetics and cardiovascular toxicity in an established female mouse model of gonococcal genital tract infection that has been increasingly used for evaluation of candidate antimicrobials for treatment of gonorrhea. We expect that the results from our studies will demonstrate the efficacy of these two antimicrobials as new therapeutics for N. gonorrhoeae infection, which are urgently needed given the rise in MDR gonococcal strains. This will be the first study of its kind to test these two classes of antimicrobials for efficacy against gonorrhea.

淋病奈瑟菌感染是发病的主要原因，估计有 850,000 例病例 美国和全球每年有 8700 万例病例。在 VA 医疗保健系统内，淋病病例 2013 年至 2017 年间有所增加，该时期总数达到 10,587 人。最严重的 当淋球菌上升到上生殖道并引起感染时，受感染的妇女会遭受后遗症 10-20% 的感染女性患有盆腔炎，其中包括多种疾病 炎症性疾病，常常导致慢性盆腔疼痛、不孕和宫外孕。没有 淋病奈瑟菌疫苗以及由于多重耐药性惊人上升而对新抗生素的巨大需求 （MDR），这使得无法治疗的淋球菌感染的出现成为现实。目前仅 推荐头孢曲松和阿奇霉素作为一线治疗，临床分离株对这两种药物均耐药 丹麦、加拿大和日本等国家已报告使用这些抗生素。因此，有一个 迫切需要新的抗菌药物来治疗淋球菌感染。 我们迄今为止对淋病奈瑟菌脂寡糖 (LOS) 和人类先天免疫系统的研究 研究表明，LOS 的脂质 A 部分是细胞因子介导的炎症的主要诱导物，并且 其他人的研究表明，脂质 A 也促进淋球菌感染。这些数据引导我们 以脂质 A 生物合成为目标将是对抗淋病奈瑟菌的有效方法的概念 感染。我们最近报道了对 LpxC 的抑制，LpxC 是催化脂质 A 第二步的酶 生物合成，对九种多重耐药性和人类挑战的淋球菌菌株具有杀菌作用 减少细胞因子诱导，而没有明显的人类细胞毒性。 根据 LpxC 抑制剂数据，我们假设由于 LOS 的抑制而导致膜破坏 生物合成对淋球菌是致命的。为了研究这一点，我们最近评估了一种细菌的杀菌潜力 12 个氨基酸的细胞穿透肽 (CPP)，用于治疗淋病奈瑟菌的 MDR 和人类攻击菌株 发现 CPP 能穿透细菌膜，对所有九种 MDR 和人类都有杀菌作用 测试了淋球菌的挑战菌株。重要的是，在幸存下来的过程中，并没有出现对菲律宾共产党的明显抵抗。 暴露于CPP然后撤退后的菌落中细菌的敏感性是相同的。 此外，CPP 减少炎症细胞因子的诱导并防止细菌细胞侵入宫颈。 上皮细胞在没有可测量的细胞毒性的情况下。 这些新数据强调 LpxC 和 CPP 作为治疗淋病奈瑟菌的有前景的抗菌剂，并强烈 支持本申请的假设：用LpxC抑制脂质A成分的生物合成 抑制剂和用 CPP 破坏外膜完整性将影响细菌活力和宿主对 体外和体内淋病奈瑟菌感染，这将对感染结果产生治疗影响。 该项目的重点是优化和测试 CPP 和 LpxC 抑制剂在相关领域的功效。 杀菌活性、细胞因子诱导、溶血和细胞毒性的体外测定。机理研究将 包括 DNA 结合、细胞透化、蛋白水解抗性、蛋白质结合和 MtrCDE、MacAB 和 NorM 淋球菌外排泵的效果。主要候选 CPP 和 LpxC 抑制剂 体外鉴定的药物将在体内药效、药代动力学和心血管毒性方面进行测试 建立了淋球菌生殖道感染的雌性小鼠模型，该模型已越来越多地用于 评估治疗淋病的候选抗菌药物。 我们期望我们的研究结果将证明这两种抗菌药物作为新抗菌药物的功效 鉴于耐多药淋球菌菌株的增加，迫切需要治疗淋病奈瑟菌感染的药物。 这将是第一项测试这两类抗菌药物对抗淋病疗效的研究。