Computational design of novel antigens targeting mature and germline b12

针对成熟和种系 b12 的新型抗原的计算设计

• 批准号: 8117981
• 负责人: WILLIAM R. SCHIEF
• 金额: $ 52.24万
• 依托单位: SEATTLE BIOMEDICAL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-05-01 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8117981
• 关键词: Address; Affect; Affinity; Antibodies; Antibody Binding Sites; Antigen Targeting; Antigens; B-Lymphocytes; Binding; Binding Sites; Complex; Dissociation; Engineering; Epitopes; Feedback; Grant; HIV; HIV Envelope Protein gp120; HIV vaccine; Human; Immunization; In Vitro; Infusion procedures; Kinetics; Libraries; Macaca; Mannose; Membrane; Mus; Mutagenesis; Names; Oryctolagus cuniculus; Polysaccharides; Proteins; Receptors, Antigen, B-Cell; Scaffolding Protein; Screening procedure; Serum; Specificity; Structure; Testing; Transplantation; V3 Loop; Variant; Yeasts; antigen binding; base; crosslink; design; directed evolution; env Gene Products; human monoclonal antibodies; in vivo; neutralizing antibody; novel; particle; scaffold; simian human immunodeficiency virus; vaccine development

Here we focus on immunogen design to elicit b12-like antibodies against the conserved cd4 binding site (cd4bs) of HIV Envelope. Many different engineered variants of HIV Envelope have failed to elicit such antibodies, but here we develop two novel types of b12 antigen that have not previously been tested - non-HIV protein scaffolds onto which the b12 epitope has been transplanted, and minimized, stabilized variants of core gp120. Further, we employ a combination of computational protein design and yeast display directed evolution that has not been employed for b12 antigen design previously. We will test the following hypotheses: (i) to induce b12-like antibodies rather than non- or narrowly-neutralizing antibodies against the cd4bs, it will be necessary to design antigens that bind b12 but not other cd4bs antibodies (ii) to optimally stimulate b12 B-cells and elicit b12-like antibodies will require antigens that stabilize the structure of the b12 epitope and optimize the affinity and kinetics of the antigen-b12 interaction (iii) to stimulate naive B cells to develop into those producing the mature broadly-neutralizing form of b12, antigens that bind both germline b12 and mature b12 will be required; (iv) to maximally stimulate b12 B cells by crosslinking B cell receptors, it will be necessary to multimerize b12-antigens on particles in an oriented fashion with the epitope facing outward. Our (Project 1) design efforts will be informed by structural and biophysical analysis of b12 antigens and their interactions with b12 and with mouse MAbs elicited by designed antigens (Project 2), by analysis of b12 antigen stimulation of B cells in vitro and in vivo (Project 3), and by binding specificity and neutralization analysis of rabbit sera elicited by selected b12 antigens (Core B).

在这里，我们着重于免疫原料设计，以引发针对HIV包膜的保守CD4结合位点（CD4B）的B12样抗体。 HIV包膜的许多不同工程的变体未能引起此类抗体，但是在这里，我们开发了两种新型的B12抗原类型，这些抗原以前尚未进行过测试 - 非HIV蛋白支架已将B12表位已移植并最小化，并最小化，稳定的Core Core core gp120的变体。此外，我们采用了以前尚未用于B12抗原设计的计算蛋白设计和酵母显示的定向进化。 We will test the following hypotheses: (i) to induce b12-like antibodies rather than non- or narrowly-neutralizing antibodies against the cd4bs, it will be necessary to design antigens that bind b12 but not other cd4bs antibodies (ii) to optimally stimulate b12 B-cells and elicit b12-like antibodies will require antigens that stabilize the structure of the b12 epitope and优化抗原-B12相互作用（III）的亲和力和动力学，以刺激幼稚的B细胞发展成产生B12成熟的宽中和形式的抗原，这是结合两种生殖线的抗原 需要B12和成熟的B12； （iv）通过交联B细胞受体最大程度地刺激B12 B细胞，必须以面向向外的表位以面向颗粒的方式对B12抗原进行多合中的B12抗原。我们的（项目1）设计工作将通过对B12抗原的结构和生物物理分析及其与B12的相互作用以及与设计抗原引起的小鼠MAB的相互作用（项目2），通过分析B12抗原刺激B12抗原在体外和体内对B细胞的刺激（Project 3），以及通过对Rabbit sera Alabitige bbity Sera Anallation（Project）的b122222222。