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.

感染N.淋病是发病的主要原因， 美国全球每年有8700万例在VA卫生保健系统内，淋病病例 在2013年至2017年期间增加，该期间的总数为10，587。最严重的 受感染的妇女遭受后遗症，因为淋球菌上升到上生殖道， 10 - 20%的女性感染盆腔炎，其中包括广泛的 炎症条件，并经常导致慢性盆腔疼痛，不孕症和异位妊娠。没有 疫苗接种到N.淋病和大量需要新的抗生素，由于多药耐药性的惊人增长 (MDR)，这使得无法治疗的淋球菌感染的出现成为一个真实的前景。目前只 头孢曲松和阿奇霉素被推荐作为一线治疗，临床分离株对这两种药物都有耐药性， 这些抗生素在包括丹麦、加拿大和日本在内的国家都有报道。由此可见，有一 迫切需要新的抗微生物剂治疗淋球菌感染。 我们的研究到N.淋球菌脂寡糖（LOS）与人类先天免疫系统 已经表明LOS的脂质A部分是精氨酸介导的炎症的主要诱导物， 其他人的研究表明，脂质A也促进淋球菌感染。这些数据使我们 认为靶向脂质A生物合成将是对抗N.淋病 感染.我们最近报道，抑制LpxC，这种酶催化脂质A的第二步， 生物合成，对9种多重耐药淋球菌和人类挑战菌株具有杀菌作用， 减少细胞因子诱导而无明显的人细胞毒性。 从LpxC抑制剂的数据，我们推测，由于LOS的抑制， 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 12个氨基酸的细胞穿透肽（CPP）对耐药和人类挑战的N。淋病和 发现CPP穿透细菌膜，对所有9种MDR和人类具有杀菌作用。 测试淋球菌的挑战菌株。重要的是，在存活的小鼠中没有对CPP产生明显的抗性。 暴露于CPP后再消退的菌落中的细菌敏感性相同。 此外，CPP减少了炎症细胞因子的诱导，并阻止了宫颈癌细菌细胞的侵袭。 上皮细胞，在没有可测量的细胞毒性。 这些新的数据突出了LpxC和CPP作为N.淋病和强烈 支持本申请的假设，即用LpxC抑制脂质A组分的生物合成 抑制剂和用CPP破坏外膜完整性将影响细菌活力和宿主对 n.淋病感染的体外和体内，这将有感染的结果治疗的影响。 该项目的重点是优化和测试CPP和LpxC抑制剂在相关治疗中的疗效。 杀菌活性、细胞因子诱导、溶血和细胞毒性的体外测定。机械研究将 包括DNA结合、细胞透化、蛋白质水解抗性、蛋白质结合和 MtrCDE、MacAB和NorM淋球菌外排泵的作用。主要候选CPP和LpxC抑制剂 将在体外试验中测试体内功效、药代动力学和心血管毒性。 已建立淋球菌生殖道感染的雌性小鼠模型， 评价治疗淋病的候选抗菌药物。 我们希望我们的研究结果将证明这两种抗菌剂作为新的抗菌剂的有效性。 N.淋病感染，这是迫切需要的耐多药淋球菌菌株的上升。 这将是同类研究中第一次测试这两类抗生素对淋病的有效性。