10月，国际学术杂志Retrovirology正式发表了上海巴斯德所研究人员关于有效捕获HIV-1包膜蛋白上瞬间暴露中和表位的研究成果。这项研究由周保罗研究员领导的“抗病毒免疫与遗传治疗”研究组与美国贝勒医学院合作完成。
如何设计免疫原使其诱导出针对HIV-1包膜蛋白的广谱中和抗体反应是当今HIV-1疫苗开发的难点。诱导出这些广谱中和抗体的包膜蛋白表位包括构象依赖的和构象非依赖两种方式。迄今为止发现的约10个广谱中和抗体识别的表位均是构象非依赖的。然而，HIV-1感染靶细胞时，包膜蛋白与靶细胞上的受体和辅助性受体作用后会发生一系列构象改变，从而暴露出构象诱导的瞬时性广谱中和表位。但这些构象诱导的瞬时暴露的广谱中和表位往往很难用常规方法发现和界定。
在本研究中，研究人员用糖基磷脂酰肌醇锚(GPI)将构象依赖的单链抗体锚定在HIV-1易感细胞表面介导病毒进入或者释放的脂筏区。研究发现其中4个单链抗体可不同程度地中和HIV-1，并发现一个识别CD4诱导的极为保守表位的GPI锚定的单链抗体X5，能完全抑制本研究中所用的多亚型HIV-1假病毒株以及5个野生型病毒株的感染。此外，该研究还发现细胞表面表达有GPI锚定的单链抗体X5的人T细胞能够长期完全性抑制HIV-1的复制、完全阻止gp120介导的细胞-细胞间融合以及抑制人树突状细胞通过DC-SIGN捕获的HIV-1的感染。更为重要的是，X5的表位对于保持HIV-1生物学活性具有重要作用，也证明了X5表位是抗HIV-1的潜在重要靶点。因此，GPI锚定的单链抗体有可能开发为一种通用且有效的方法来发现并界定HIV-1和其它包膜病毒中构象诱导的瞬时暴露的中和抗体及中和表位。GPI锚定的单链抗体X5，因其抗HIV-1的广谱性、强有力的中和活性及其中和表位不易逃逸的保守性，极有潜力被开发成为抗HIV-1的新型制剂，从而对艾滋病的预防及治疗起到关键作用。
本研究得到了国家自然科学基金委、国家科技部973项目、国家重大科技专项、上海巴斯德健康研究基金会项目以及阿海珐(AREVA)国际合作项目的资助。（生物谷Bioon.com）
生物谷推荐英文摘要：
Retrovirology doi:10.1186/1742-4690-7-79
GPI-anchored single chain Fv - an effective way to capture transiently-exposed neutralization epitopes on HIV-1 envelope spike
Michael Wen1 , Reetakshi Arora2 , Huiqiang Wang1 , Lihong Liu1 , Jason T Kimata2 and Paul Zhou1
1 The Unit of Anti-Viral Immunity and Genetic Therapy, the Key Laboratory of Molecular Virology and Immunology, the Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, 200025, China
2 Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas, 77030, USA
Background
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.
Results
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.
Conclusions
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.
