The Unit of Anti-Viral Immunity and Genetic Therapy, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, 200025, China.
Retrovirology. 2010 Oct 6;7:79. doi: 10.1186/1742-4690-7-79.
Identification of broad neutralization epitopes in HIV-1 envelope spikes is paramount for HIV-1 vaccine development. A few broad neutralization epitopes identified so far are present on the surface of native HIV-1 envelope spikes whose recognition by antibodies does not depend on conformational changes of the envelope spikes. However, HIV-1 envelope spikes also contain transiently-exposed neutralization epitopes, which are more difficult to identify.
In this study, we constructed single chain Fvs (scFvs) derived from seven human monoclonal antibodies and genetically linked them with or without a glycosyl-phosphatidylinositol (GPI) attachment signal. We show that with a GPI attachment signal the scFvs are targeted to lipid rafts of plasma membranes. In addition, we demonstrate that four of the GPI-anchored scFvs, but not their secreted counterparts, neutralize HIV-1 with various degrees of breadth and potency. Among them, GPI-anchored scFv (X5) exhibits extremely potent and broad neutralization activity against multiple clades of HIV-1 strains tested. Moreover, we show that GPI-anchored scFv (4E10) also exhibited more potent neutralization activity than its secretory counterpart. Finally, we demonstrate that expression of GPI-anchored scFv (X5) in the lipid raft of plasma membrane of human CD4+ T cells confers long-term resistance to HIV-1 infection, HIV-1 envelope-mediated cell-cell fusion, and the infection of HIV-1 captured and transferred by human DCs.
Thus GPI-anchored scFv could be used as a general and effective way to identify antibodies that react with transiently-exposed neutralization epitopes in envelope proteins of HIV-1 and other enveloped viruses. The GPI-anchored scFv (X5), because of its breadth and potency, should have a great potential to be developed into anti-viral agent for HIV-1 prevention and therapy.
鉴定 HIV-1 包膜刺突中的广谱中和表位对于 HIV-1 疫苗的开发至关重要。迄今为止已鉴定出的少数几个广谱中和表位存在于天然 HIV-1 包膜刺突的表面，这些表位的识别不依赖于包膜刺突的构象变化。然而，HIV-1 包膜刺突还包含瞬时暴露的中和表位，这些表位更难识别。
在这项研究中，我们构建了来自七个人源单克隆抗体的单链 Fv（scFv），并将它们与或不与糖基磷脂酰肌醇（GPI）连接信号基因相连。我们表明，带有 GPI 连接信号的 scFv 被靶向到质膜的脂筏。此外，我们证明，四种 GPI 锚定的 scFv，但不是它们的分泌型对应物，以不同程度的广度和效力中和 HIV-1。其中，GPI 锚定的 scFv（X5）对测试的多种 HIV-1 株表现出极其强大和广谱的中和活性。此外，我们表明 GPI 锚定的 scFv（4E10）也表现出比其分泌型对应物更强的中和活性。最后，我们证明 GPI 锚定的 scFv（X5）在人 CD4+T 细胞质膜的脂筏中的表达赋予了对 HIV-1 感染、HIV-1 包膜介导的细胞-细胞融合以及被人类 DC 捕获和转移的 HIV-1 感染的长期抗性。
因此，GPI 锚定的 scFv 可用于鉴定与 HIV-1 和其他包膜病毒包膜蛋白中瞬时暴露的中和表位反应的抗体的通用有效方法。由于其广度和效力，GPI 锚定的 scFv（X5）具有很大的潜力被开发为用于 HIV-1 预防和治疗的抗病毒药物。
