Applying High-Performance Protein Engineering Tools to HIV Immunogen Design

描述（由申请人提供）：被动转移中和抗体（nab）对HIV-1感染提供了明确的保护，但对这些nab的鉴定和机制研究尚未导致保护性疫苗的开发。我们假设先前的免疫原设计工作由于其有限的范围和局限的焦点而取得了有限的成功。缺乏高性能筛选平台严重限制了免疫原设计，并阻碍了将基础科学发现转化为疫苗开发。当试图在自然发生的保护性免疫反应很大程度上缺失的疾病背景下引发针对高度多样化病毒的保护性抗体时，我们假设所使用的技术和方法的规模必须与任务的规模充分匹配。组合的高通量蛋白质工程平台可以实现成功开发预防性HIV疫苗所必需的景观覆盖，并使我们能够将我们对nab的理解转化为诱导保护性抗体反应。因此，我们建议开发高性能的蛋白质工程工具，根据定义的标准筛选数十亿个HIV包膜三聚体序列变体，使我们能够进化出包膜免疫原，以有效地呈现功能相关和免疫原性的表位，能够驱动保护性抗体的产生。我们描述的多管齐下的策略既利用了传统免疫原设计的不断增长的试剂工具包，包括十几种新的广谱nab，也利用了改进的方法来功能解析存在于多种多克隆样品中的抗体；并进一步探索了一种基于天然和种系抗体库选择性参与的互补策略，我们认为这解决了以前免疫原设计工作的基本限制，并代表了一种创新和公正的正向工程策略，以诱导产生中和抗体。最终，这些研究开发的工具代表了适应性强的平台，能够根据灵活的设计标准从大量序列多样性中快速选择包膜变体，并可能通过弥合被称为“nAb问题”的翻译差距，为免疫原设计的根本突破开辟道路。

DESCRIPTION (provided by applicant): Passively transferred neutralizing antibodies (nAbs) have provided definitive protection from HIV-1 infection, yet identification and mechanistic study of these nAbs has not resulted in the development of a protective vaccine. We hypothesize that previous immunogen design efforts have achieved limited success due to their limited scope and restricted focus. The lack of a high-performance screening platform places severe constraints on immunogen design and has impeded the translation of findings from basic science into vaccine development. When attempting to elicit protective antibodies against a highly diverse virus in the context of a disease in which naturally occurring protective immune responses are largely absent, we posit that the scale of technology and methods used must be adequately matched to the scale of the task. Combinatorial, high-throughput protein engineering platforms can achieve the landscape coverage that may be necessary for successful development of a preventative HIV vaccine, and allow us to translate our understanding of nAbs into the induction of protective antibody responses. We therefore propose to develop high-performance protein engineering tools to screen billions of HIV envelope trimer sequence variants according to defined criteria, allowing us to evolve envelope immunogens tailored to effectively present functionally relevant and immunogenic epitopes capable of driving the generation of protective antibodies. The multi-pronged strategy we describe both leverages the growing reagent toolkit for traditional immunogen design, including more than a dozen new broad nAbs, and improved means to functionally parse Abs present in diverse, polyclonal samples; and further explores a complementary strategy based on selective engagement of na¿ve and germline antibody repertoires which we believe addresses the fundamental limitation of previous immunogen design efforts and represents an innovative and unbiased forward engineering strategy to induce the generation of neutralizing antibodies. Ultimately, the tools developed by these studies represent adaptable platforms capable of rapidly selecting envelope variants from vast sequence diversity according to flexible design criteria, and may open a path to a fundamental breakthrough in immunogen design by bridging the translational gap that has come to be known as "the nAb problem".