Development of nanobody immunotherapeutics that prevent and treat gonorrhea

开发预防和治疗淋病的纳米抗体免疫疗法

• 批准号: 10753164
• 负责人: SANJAY RAM
• 金额: $ 26.57万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-02 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10753164
• 关键词: Affect; Affinity; Alpaca; Antibiotic Resistance; Antibiotics; Antibodies; Antibody Therapy; Antigens; Antimicrobial Cationic Peptides; Attenuated; B-Lymphocytes; Bacteria; Binding; Binding Proteins; Biotechnology; Ceftriaxone; Cell division; Cells; Characteristics; Chlamydia; Clinical; Collaborations; Complement; Complement Factor H; Complement Membrane Attack Complex; Cytidine Monophosphate N-Acetylneuraminic Acid; Development; Disease; Drug Delivery Systems; Drug-resistant Neisseria Gonorrhoeae; Ectopic Pregnancy; Enzymes; Epitopes; Flow Cytometry; Genes; Goals; Gonorrhea; HIV; Human; IgG1; Immune; Immune Evasion; Immunization; Immunize; Immunoglobulin G; Immunotherapeutic agent; In Vitro; Incidence; Individual; Infection; Infertility; Inflammatory Response; Lead; Libraries; Link; Mediating; Messenger RNA; Microbe; Modeling; Molecular Conformation; Multi-Drug Resistance; Mus; Mutation; Neisseria gonorrhoeae; Organism; Pathogenesis; Pathway interactions; Persons; Phage Display; Predisposition; Prevention; Preventive; Property; Proteins; Reaction; Recombinants; Recommendation; Reporting; Resistance; Sexually Transmitted Diseases; Sialic Acids; Sialyltransferases; Specificity; Surface; Technology; Testing; Therapeutic; Therapeutic Agents; Therapeutic antibodies; Topical application; United States; Vagina; Vaginal Ring; Variant; Woman; Work; antitoxin; bactericide; chronic pelvic pain; drug-resistant gonorrhea; efficacy testing; hyperimmunization; immune clearance; improved; in silico; in vivo; infection burden; inhibitor; lipooligosaccharide; mucosal site; mutant; nanobodies; novel; prevent; public health priorities; receptor; screening; sialic acid binding Ig-like lectin; sialylation; synergism; technology platform; transmission process; trend; vaccine candidate

PROJECT SUMMARY/ABSTRACT About 87 million new cases of gonorrhea occur worldwide annually. In 2020, 677,769 cases were reported to the CDC, a 111% increase in annual incidence since the historic low in 2009. Neisseria gonorrhoeae (Ng) has become resistant to almost every antibiotic in clinical use. Reports of resistance to ceftriaxone – the only currently recommended first-line of treatment – from almost every continent portends an era of untreatable gonorrhea. Development of novel treatments and preventives against Ng is a global public health priority. Our group, in collaboration with Evaxion Biotech, has identified two putative cell-division proteins (NGO0265 and NGO1549 (FtsN)) as lead vaccine candidates. NGO0265 and FtsN are expressed by all Ng isolates and FtsN is essential for bacterial viability. Immunization of mice with NGO0265 and FtsN elicits bactericidal antibodies (Abs) and significantly reduces the duration and burden of gonococcal colonization of mouse vaginas. Ng deploy a unique immune evasion strategy wherein it caps its lipooligosaccharide (LOS) with sialic acid using its surface LOS sialyltransferase (Lst) and host- derived CMP-sialic acid. LOS sialylation enables Ng to evade complement, cationic antimicrobial peptides (CAMPs) and down-regulate the inflammatory response by engaging host Siglec receptors. Gonococcal lst deletion mutants are attenuated in mice. Supporting its key role in pathogenesis, all gonococcal disease isolates sequenced thus far possess a functional Lst gene. Camelid single-domain antibodies called VHHs or nanobodies, have been found to possess unique properties that offer enormous versatility to facilitate the rapid development of simple and economical antibody-based therapeutics. VHH- based products can exploit the small size and high stability of VHH components, their propensity to bind conformation- dependent neutralizing epitopes, and their tractability for functional expression as heteromultimers. These characteristics make possible commercially favorable antibody therapeutic agents having ultrahigh target affinities, broad natural variant specificities and the ability to bind multiple different targets. Here, we aim to develop nanobody-based therapeutics against NGO0265, FtsN and Lst for use as an adjunctive treatment and as a preventive against multidrug-resistant gonorrhea. In Aim 1, we will immunize alpacas with purified recombinant chimeric NGO0265-NGO1549 and Lst, create a nanobody-display phage library from immune B cells and identify nanobodies that recognize non-overlapping epitopes on their targets with high-affinity. The ability of the nanobodies to block enzyme function or to activate complement and kill Ng when linked to IgG Fc will be evaluated in Aim 2. In Aim 3, we will test the ability of the lead anti-Ng nanobodies against each of the targets to clear gonococcal colonization in a well-established mouse vaginal colonization model. Successful completion of the work will identify and characterize nanobodies with activity against multidrug-resistant Ng, which can then be adapted to a variety of formats and drug delivery platforms.

项目摘要/摘要 每年大约有8700万例新的淋病病例发生。在2020年，疾病预防控制中心报告了677,769例 自2009年历史低年度以来，每年发病率增加了111％。淋病奈瑟氏菌（NG）已经对 几乎每种抗生素在临床用途中。对头孢曲松抵抗的报告 - 目前唯一推荐的一线 治疗 - 几乎每个大陆都预示着一个无法治疗的淋病时代。新疗法的发展 反对NG的预防措施是全球公共卫生优先事项。 我们的小组与Evaxion Biotech合作，已经确定了两种推定的细胞分割蛋白（NGO0265和 NGO1549（FTSN））为候选铅疫苗。 NGO0265和FTSN均由所有NG分离株表达，FTSN对于 细菌生存能力。用NGO0265和FTSN对小鼠的免疫接种，引起细菌抗体（ABS），并显着显着 减少了小鼠阴道的淋球菌定植的持续时间和燃烧。 NG部署独特的免疫进化 其中，它使用其表面LOS siAllyltransferas（LST）和宿主 - 将其lipoligosacacacy（LOS）用唾液酸限制 衍生的CMP-Sialic酸。 Los lylation使NG能够逃避完整的阳离子抗菌胡椒（CAMP）和 通过使宿主SIGLEC受体接合来下调炎症反应。淋球菌LST缺失突变体是 在小鼠中衰减。支持其在发病机理中的关键作用，迄今为止测序的所有淋球菌疾病分离株均具有 功能性LST基因。 骆驼层单域抗体称为VHHS或纳米型，已被发现具有提供的独特特性 巨大的多功能性促进了基于简单和经济的抗体治疗的快速发展。 VHH- 基于的产品可以探索VHH组件的小尺寸和高稳定性，它们的承诺结合构象 - 依赖性中和表位及其作为异栽培者功能表达的易干性。这些特征 使可能具有超高目标亲和力的商业有利的抗体治疗剂，广泛的自然变体 特异性和结合多个不同目标的能力。 在这里，我们旨在开发针对NGO0265，FTSN和LST作为辅助治疗的基于纳米的治疗 并作为预防性抗多药的淋病。在AIM 1中，我们将用纯化的重组对羊驼免疫 Chimeric NGO0265-NGO1549和LST，从免疫B细胞中创建一个纳米型 - 播放噬菌体图书馆，并识别 识别具有高亲和力的靶标的非重叠表位的纳米体。纳米体的能力 将酶函数或激活与IgG FC链接时杀死NG的块功能将在AIM 2中评估。在AIM 3中， 我们将测试铅抗NG纳米型针对每个靶标的清除A中淋球菌定植的能力 公认的小鼠阴道定植模型。 成功完成工作将识别并表征纳米体，并具有针对多种耐药性NG的活动， 然后可以适应各种格式和药物输送平台。