Analysis of the Initiation of an HIV Broadly Neutralizing Antibody Lineage in a Single Host

人类免疫缺陷病毒(Hiv)是一种快速进化的病原体，可以逃避免疫防御。 由大多数疫苗诱导的抗体提供。诱导广谱中和抗体的建议策略 通过接种疫苗(BNAbs)需要更深入地了解感染免疫反应的演变，因为 这些保护性抗体通常需要4-5年的时间才能形成。在感染艾滋病毒的个人中，病毒和 产生抗体的B细胞共同进化，与病毒种群一起形成病毒-抗体“军备竞赛” 抗体存在于整个感染过程中。建议的研究是分析关键的早期时间点 一名捐献者体内的军备竞赛，他开发出了广泛的抗体，以指导免疫原设计。 除了快速变异外，HIV还使用重糖基化和构象掩蔽来逃避 免疫系统。供体CH848产生了一种名为DH2 70的bNab谱系，它与糖链相互作用 在HIV包膜(Env)可变环路V3的底部的“SuperSite”。配合物的晶体结构分析 在HIV Env的成熟成员和片段之间，再加上结合数据，表明这是不可能的 抗体中的突变导致不同分支中和抗体的不同性质 在抗体副表位或抗原表位没有任何重大结构变化的情况下。虽然很多人 V3-葡聚糖“超位”bNAbs识别N332糖链，它们的实际表位在其他糖链和环境中不同 它们能识别的多肽。因此，是什么触发了DH2 70谱系的发展仍有待确定。 了解HIV的特性 环境以及与抗体的相互作用是DH2 70的关键 HIV的谱系发育、原子分辨结构 环境因子将通过低温电子显微镜进行测定。 和/或DH2 70谱系的早期成员DH270.IA4的X射线结晶学，并与合作 抗体谱系成员DH475和DH0022。CH848捐赠者也产生了合作抗体， 引发的病毒逃逸突变改善了与DH2 70谱系抗体的结合，并可能加速 DH2 70谱系的亲和力成熟。DH2 70系祖细胞抗体如何结合的假说 Env也可以从DH270.IA4的复杂结构中推导出来，因为DH270.IA4不同于前身 由五种氨基酸组成。假说将通过将突变引入Fabs和/或HIV Env和 用生物膜干涉法测定结合亲和力。络合物中协同抗体的结构 Env将确定HIV env的特性(即构象、糖基化模式等)。引发了这些非- 中和抗体，以及尽管它们的表位重叠，但它们如何帮助DH2 70谱系的发展。 这些数据将确定触发广谱中和糖链发展的机制(S)。 依赖抗体，并指导疫苗设计。这笔助学金资助的本科生研究生 提案将使用尖端技术探索对公共卫生至关重要的问题，成为合著者 并接受致力于长期职业发展的专家的指导。

Human immunodeficiency virus (HIV) is a rapidly evolving pathogen that escapes immune defenses provided by most vaccine-induced antibodies. Proposed strategies to elicit broadly neutralizing antibodies (bnAbs) by vaccination require a deeper understanding of evolution of the immune response to infection, since these protective antibodies typically take ~4-5 years to develop. In HIV infected individuals, viruses and antibody producing B-cells evolve together, creating a virus-antibody “arms race”, with populations of viruses and antibodies present throughout infection. The proposed research is to analyze critical early time-points of the arms race in a donor who developed antibodies of significant breadth, to guide immunogen design. In addition to rapid mutation, HIV also uses heavy glycosylation and conformational masking to evade the immune system. Donor CH848 produced a bnAb lineage, called DH270, which interacts with the glycan “supersite” at the base of the HIV envelope (Env) variable loop V3. Analysis of crystal structures of complexes between mature members and fragments of the HIV Env, together with binding data, suggest that improbable mutations in the antibodies led to the different neutralization properties of antibodies in the different branches of the lineage, without any major structural change in the antibody paratope or antigen epitope. While many V3-glycan “supersite” bnAbs recognize the N332 glycan, their actual epitopes differ in other glycans and Env peptides they recognize. Thus, it remains to be determined what triggered DH270 lineage development. To understand properties of HIV Env and interactions with antibodies that were critical for DH270 lineage development, atomic resolution structures of HIV Envs will be determined by cryo-electron microscopy and/or X-ray crystallography with an early member of the DH270 lineage, DH270.IA4, and with cooperating antibody lineage members, DH475 and DH0022. Cooperating antibodies, also produced in the CH848 donor, triggered virus escape mutations that improved binding to DH270 lineage antibodies and likely accelerated affinity maturation in the DH270 lineage. Hypotheses on how the DH270 lineage progenitor antibody could bind Env may also be deduced from the DH270.IA4 complex structure, since DH270.IA4 differs from the progenitor by five amino acids. Hypotheses will be tested by introducing mutations into the Fabs and/or HIV Env and determining binding affinities by biolayer interferometry. Structures of cooperating antibodies in complex with Env will identify properties of HIV Env (i.e., conformation, glycosylation patterns, etc.) that triggered these non- neutralizing antibodies, and despite their overlapping epitopes, how they aided DH270 lineage development. These data will identify mechanism(s) that triggered the development of broadly neutralizing glycan- dependent antibodies, and guide vaccine design. Undergraduate research students supported by this grant proposal will explore an issue of critical public health importance using cutting edge techniques, be co-authors on published work and be mentored by experts committed to their long-term career development.