Human Anti-Macaque CCR5 Mabs for Passive Immunotherapy

用于被动免疫治疗的人抗猕猴 CCR5 Mab

• 批准号: 6696386
• 负责人: Wayne A. Marasco
• 金额: $ 21.16万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-07-15 至 2007-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6696386
• 关键词: HIV infections; Macaca; T lymphocyte; antigen antibody reaction; antireceptor antibody; chimeric proteins; conformation; congenital infection; cytokine receptors; disease /disorder model; epitope mapping; flow cytometry; helper T lymphocyte; human immunodeficiency virus 1; immunoregulation; monoclonal antibody; neutralizing antibody; passive immunization; peptide library; protein structure function; receptor binding; simian immunodeficiency virus; transfection; vertical transmission

DESCRIPTION (provided by applicant): Infection of rhesus macaques with SIV has proven to be a valuable model for the testing of new therapeutic strategies (including vaccines) for the treatment of HIV-1 infection and AIDS. In this proposal we plan to investigate several important hypotheses as they relate to growing evidence that the HIV-1 co-receptor CCR5 exists in multiple conformational states on CD4+ T-cells, that these conformational states may effect the permissiveness of HIV-1 entry and that blocking CCR5 with high-affinity human anti-hCCR5 Mabs may be a cost effective and viable strategy to prevent the vertical transmission of HIV-l in certain clinical settings. We have chosen the well characterized macaque model of SIVmac239 infection for these studies. Specifically, we will use our 15 billion member human single chain antibody (sFv) phage display library to isolate high affinity human anti-RhCCR5 antibodies against different conformational states of rhesus macaque CCR5 (RhCCR5). We will perform epitope mapping studies and determine the sFv binding constants for the anti-RhCCR5 sFvs. We will use multiparameter FACS to investigate the existence of multiple conformational states of RhCCR5 on different subpopulations of macaque CD4+ T-cells by examining the anti-RhCCR5 sFv binding profiles under resting and different conditions of stimulation. We will also investigate whether certain conformational states of RhCCR5 may be more permissive for SIV entry by testing the anti-RhCCR5 sFvs both individually and in combinations for their ability to block in vitro SIV infection of macaque CD4+ T-cells. We will also determine if co-receptor adaptation by SIV occurs in vitro as a result of antibody treatment. We will also perform "in vitro" affinity maturation on the most promising SIV neutralizing anti-RhCCR5 sFv to test the hypothesis that increased anti-viral activity can be achieved by improving sFv binding affinity to the nM or sub-nanomolar range. Finally, we will use these high-affinity and SIV neutralizing anti-RhCCR5 sFvs to prepare chimeric macaque IgG1 Mabs to determine if whole anti-RhCCR5 Mabs have enhanced neutralizing activity in SIV challenge studies of macaque CD4+ T-cells. These macaque anti-RhCCR5 Mabs will be used in in vivo studies to test the hypothesis that passive immunotherapy with macaque anti-RhCCR5 Mabs can be used to block the vertical transmission of SIVmac239. These studies will supply important information that may be directly applicable to similar human clinical settings.

描述（由申请人提供）：恒河猴感染 SIV 已被证明是测试治疗 HIV-1 感染和艾滋病的新治疗策略（包括疫苗）的有价值的模型。在本提案中，我们计划研究几个重要的假设，因为它们涉及越来越多的证据，表明 HIV-1 辅助受体 CCR5 在 CD4+ T 细胞上以多种构象状态存在，这些构象状态可能会影响 HIV-1 进入的许可性，并且在某些临床环境中，用高亲和力人抗 hCCR5 Mab 阻断 CCR5 可能是预防 HIV-1 垂直传播的一种具有成本效益且可行的策略。我们为这些研究选择了特征明确的 SIVmac239 感染猕猴模型。具体来说，我们将使用我们的 150 亿成员人类单链抗体 (sFv) 噬菌体展示库来分离针对不同构象状态的恒河猴 CCR5 (RhCCR5) 的高亲和力人类抗 RhCCR5 抗体。我们将进行表位作图研究并确定抗 RhCCR5 sFv 的 sFv 结合常数。我们将使用多参数 FACS 通过检查静息和不同刺激条件下的抗 RhCCR5 sFv 结合谱来研究不同猕猴 CD4+ T 细胞亚群上 RhCCR5 的多种构象状态的存在。我们还将通过单独和组合测试抗 RhCCR5 sFv 阻断猕猴 CD4+ T 细胞体外 SIV 感染的能力，研究 RhCCR5 的某些构象状态是否更容易允许 SIV 进入。我们还将确定 SIV 的辅助受体适应是否因抗体治疗而在体外发生。我们还将对最有希望的 SIV 中和性抗 RhCCR5 sFv 进行“体外”亲和力成熟，以测试通过将 sFv 结合亲和力提高到 nM 或亚纳摩尔范围来提高抗病毒活性的假设。最后，我们将使用这些高亲和力和 SIV 中和抗 RhCCR5 sFv 来制备嵌合猕猴 IgG1 Mab，以确定整个抗 RhCCR5 Mab 在猕猴 CD4+ T 细胞的 SIV 攻击研究中是否具有增强的中和活性。这些猕猴抗RhCCR5 Mab将用于体内研究，以测试猕猴抗RhCCR5 Mab的被动免疫疗法可用于阻断SIVmac239的垂直传播的假设。这些研究将提供可能直接适用于类似人类临床环境的重要信息。