Novel immunotherapeutics against multi-drug resistant Neisseria gonorrhoeae

针对多重耐药淋病奈瑟菌的新型免疫疗法

• 批准号: 8703886
• 负责人: SANJAY RAM
• 金额: $ 20.89万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-01 至 2016-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8703886
• 关键词: Address; Affect; Amino Acids; Anti-Bacterial Agents; Antibiotic Resistance; Antibiotics; Attenuated; Bacteria; Binding; Binding Sites; Ceftriaxone; Cells; Chimeric Proteins; Complement; Complement Activation; Complement Factor H; Complement Inactivators; Complement component C5; Cytolysis; Development; Disease; Drug resistance; Ectopic Pregnancy; Epitopes; Fc Receptor; Fucose; Gonorrhea; Human; Human Factor H; IgG1; Immune; Immune system; Immunoglobulin G; Immunotherapeutic agent; In Vitro; Infection; Infertility; Injury; Kidney; Lead; Length; Ligands; Mediating; Membrane Proteins; Microbe; Modeling; Multi-Drug Resistance; Mus; Mutation; Natural Immunity; Neisseria gonorrhoeae; Organ; Pathway interactions; Pelvic Inflammatory Disease; Phagocytes; Phase; Point Mutation; Polysaccharides; Public Health; Resistance; Safety; Sexually Transmitted Diseases; Simulate; Testing; Therapeutic; Therapeutic Uses; Toxic effect; Transgenic Mice; Vagina; arm; bactericide; chronic pelvic pain; design; in vivo; inhibitor/antagonist; killings; lipooligosaccharide; macrophage; mouse model; neutrophil; novel; novel strategies; novel therapeutics; novel vaccines; pathogen; porin; product development; public health relevance; resistant strain; sialylation; uptake

DESCRIPTION (provided by applicant): Neisseria gonorrhoeae (Ng) is the causative agent of the sexually transmitted infection (STI), gonorrhea. Worldwide, >100 million new cases occur annually. Ng has become resistant to almost every antibiotic that has been used and has now achieved "superbug" status. The emergence and spread of ceftriaxone-resistant strains, in particular, has heralded an era of untreatable gonorrhea and represents a global public health crisis. Novel therapeutics against this pathogen are urgently needed. Complement (C2) is a critical arm of innate immune defenses against Ng. Ng evade C2 using several mechanisms, including scavenging the human C2 inhibitor, factor H (fH). fH comprises 20 domains - domains 18-20 interact with porin B and binding is augmented by lipooligosaccharide sialylation, while domains 6-7 bind to neisserial surface protein A. All Ng we have tested bind to one or both of these regions in fH. Chimeric molecules created by fusing fH domains 6-7 or 18-20 to the Fc region of IgG can effect C2-dependent killing of Ng. fH domains 18-20 in the context of full-length native fH binds to and serves to limit pathological injury to host cells, which limits the therapeutic use of 'unmodified' domains 18-20 in fH/Fc. We have overcome this limitation by introducing a point mutation in domain 19 (D1119G) of fH18-20/Fc that eliminates C2-mediated lysis of host cells without altering anti-Ng efficacy, thus providing us with a lead therapeutic molecule. In Aim 1, we will maximize the ability of the human IgG1 Fc region of fHD1119G/Fc to mediate opsonophagocytosis and C2-dependent killing of multi-drug resistant Ng isolates by introducing 3 amino acid point mutations in the Fc and eliminating the core fucose of Fc glycans. Similarly, in Aim 2a, we will optimize the effector functions of the Fc that is fused to human fH domains 6 and 7 (fH6,7/Fc) and evaluate its ability to mediate C2-dependent killing and opsonophagocytosis of drug-resistant Ng. The ability of fHD111G/Fc and fH67/Fc, which recognize distinct epitopes, to synergize and stimulate bactericidal activity against Ng will be evaluated in Aim 2b. The milestones achieved in Aims 1 and 2 (the R21 phase) will be followed by studies in vivo (R33 phase) to determine the safety, efficacy and mechanism of action of fH/Fc. Renal, ocular and hematologic toxicity (target organs for C2-mediated injury) will be assessed in Aim 3. Introducing the D1119G mutation could create a neo-epitope and elicit auto-Abs; this possibility will be addressed using our human fH transgenic (Tg) mice. In Aim 4 we will determine efficacy of the fH/Fc fusion proteins against a panel of drug-resistant Ng in the mouse model of Ng infection using our novel human fH/C4BP (C4BP; classical pathway C2 inhibitor) "double" Tg mice (Ng bind only human, but not mouse C2 inhibitors). Using mice that lack complement C3, C5, Fc receptors, PMNs or macrophages we will define how fH/Fc clear Ng in vivo. Many medically important microbes bind human fH through domains 6-7 and/or 18-20. While we will use Ng as a model pathogen, successful completion of these studies will provide a strong platform to assess these fH/Fc molecules against other emerging drug-resistant pathogens.

描述（由申请人提供）：淋病奈瑟菌（Ng）是性传播感染（STI）淋病的病原体。全球每年新增病例超过 1 亿。吴恩达对几乎所有使用过的抗生素都产生了耐药性，现在已经达到了“超级细菌”的地位。尤其是头孢曲松耐药菌株的出现和传播，预示着淋病无法治愈的时代的到来，并代表着一场全球公共卫生危机。迫切需要针对这种病原体的新疗法。补体 (C2) 是针对 Ng 的先天免疫防御的重要组成部分。 Ng 使用多种机制逃避 C2，包括清除人类 C2 抑制剂 H 因子 (fH)。 fH 包含 20 个结构域 - 结构域 18-20 与孔蛋白 B 相互作用，并且通过脂寡糖唾液酸化增强结合，而结构域 6-7 与奈瑟球菌表面蛋白 A 结合。我们测试的所有 Ng 都与 fH 中的这些区域之一或两个结合。通过将 fH 结构域 6-7 或 18-20 与 IgG Fc 区融合而产生的嵌合分子可以实现 Ng 的 C2 依赖性杀伤。全长天然fH背景下的fH结构域18-20结合宿主细胞并用于限制对宿主细胞的病理损伤，这限制了fH/Fc中“未修饰”结构域18-20的治疗用途。我们通过在 fH18-20/Fc 的结构域 19 (D1119G) 中引入点突变克服了这一限制，该点突变消除了 C2 介导的宿主细胞裂解而不改变抗 Ng 功效，从而为我们提供了一种先导治疗分子。在目标 1 中，我们将通过在 Fc 中引入 3 个氨基酸点突变并消除 fHD1119G/Fc 的核心岩藻糖，最大限度地提高 fHD1119G/Fc 的人 IgG1 Fc 区介导调理吞噬作用和 C2 依赖性杀伤多重耐药 Ng 分离株的能力。 Fc 聚糖。同样，在目标 2a 中，我们将优化与人 fH 结构域 6 和 7 (fH6,7/Fc) 融合的 Fc 的效应器功能，并评估其介导耐药 Ng 的 C2 依赖性杀伤和调理吞噬作用的能力。 fHD111G/Fc 和 fH67/Fc 识别不同表位，协同并刺激针对 Ng 的杀菌活性的能力将在目标 2b 中进行评估。在目标 1 和 2（R21 阶段）中实现的里程碑之后，将进行体内研究（R33 阶段），以确定 fH/Fc 的安全性、有效性和作用机制。目标 3 将评估肾、眼和血液毒性（C2 介导损伤的靶器官）。引入 D1119G 突变可以产生新表位并引发自身抗体；我们将使用我们的人类 fH 转基因 (Tg) 小鼠来解决这种可能性。在目标 4 中，我们将使用我们的新型人 fH/C4BP（C4BP；经典途径 C2 抑制剂）“双”Tg 小鼠（ Ng 仅结合人类，但不结合小鼠 C2 抑制剂）。使用缺乏补体 C3、C5、Fc 受体、PMN 或巨噬细胞的小鼠，我们将定义 fH/Fc 如何在体内清除 Ng。许多医学上重要的微生物通过结构域 6-7 和/或 18-20 结合人类 fH。虽然我们将使用 Ng 作为模型病原体，但这些研究的成功完成将为评估这些 fH/Fc 分子对抗其他新兴耐药病原体提供强大的平台。