近日，国际知名杂志《病毒学期刊》（Journal of Virology）在线发表了中国科学院上海巴斯德研究所周保罗研究组关于艾滋病病毒I型（HIV-1）疫苗的最新成果：一种基于果蝇S2细胞的新型HIV-1 病毒样颗粒（VLP）表达系统。这是世界上首次报道采用果蝇S2细胞表达系统制备HIV-1 VLP作为艾滋病疫苗组分。
2009年在泰国完成的RV144 HIV疫苗实验虽然获得了31.2%的保护率，但是仍然不能用于人群接种，该疫苗采用痘病毒载体初免，GP120蛋白加强。一个成功的HIV-1疫苗应既能够诱导出HIV-1胞膜蛋白特异性的广谱中和抗体，又能够诱导出多种蛋白特异性的T细胞免疫反应。在众多的HIV-1免疫原设计中，VLP由于能够将天然构象的胞膜蛋白刺突展示于其表面，并且能够同时诱导体液和细胞免疫反应而被认为优于GP120蛋白。但是，目前的HIV-1 VLP表达系统在某些方面还存在缺陷。
为了克服这些缺陷，博士研究生杨立飞、宋宇峰等在周保罗研究员的指导下，开发出一种基于果蝇S2细胞的新型HIV-1 VLP表达系统。首先，他们构建了表达HIV-1 VLP的S2细胞稳转克隆；然后研究了该系统表达的VLP胞膜蛋白的表达、剪切、组装以及糖基化等理化特征，并且采用冷冻电镜（Croy-EM）和冷冻电子断层成像（Croy Electron Tomography）对VLP的形态学及其表面刺突的数量进行了仔细分析；最后研究了VLP的抗原性和免疫原性。
研究表明，采用这一新型VLP表达系统制备的HIV-1 VLP其表面膜蛋白能够正确剪切、糖基化并且和gag蛋白一起组装成病毒样颗粒；采用该系统制备的HIV-1 VLP产量与杆状病毒表达系统的产量相当，甚至更高。Cryo-EM和Tomography研究发现，纯化后的VLP表面平均包含17个膜蛋白刺突，并且保留了多个已知的广谱中和抗体表位。研究人员采用DNA初免-VLP加强，并伴以CpG佐剂的接种策略，可以在小鼠体内诱导出胞膜蛋白特异性的抗体反应：包括ELISA结合抗体、中和抗体、ADCC和ADCVI特异性抗体反应，以及胞膜蛋白和gag特异性的CD8 T细胞免疫反应等，具有更好的免疫保护效果。综上所述，这一新型VLP表达系统制备的HIV-1 VLP具有很多优良的特性，可以开发成为HIV-1疫苗的有效组分。
该研究是与中科院生物物理研究所朱平研究组合作完成的，得到了国家科技重大专项的资助。（生物谷Bioon.com）
doi:10.1128/?JVI.07164-11
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HIV-1 Virus-Like Particles Produced by Stably Transfected Drosophila S2 Cells: a Desirable Vaccine Component
Lifei Yanga, Yufeng Songa, Xiaomin Lib, Xiaoxing Huangb, Jingjing Liua, Heng Dinga, Ping Zhub and Paul Zhoua
The development of a successful vaccine against human immunodeficiency virus type 1 (HIV-1) likely requires immunogens that elicit both broadly neutralizing antibodies against envelope spikes and T cell responses that recognize multiple viral proteins. HIV-1 virus-like particles (VLP), because they display authentic envelope spikes on the particle surface, may be developed into such immunogens. However, in one way or the other current systems for HIV-1 VLP production have many limitations. To overcome these, in the present study we developed a novel strategy to produce HIV-1 VLP using stably transfected Drosophila S2 cells. We cotransfected S2 cells with plasmids encoding HIV-1 envelope, Gag, and Rev proteins and a selection marker. After stably transfected S2 clones were established, HIV-1 VLP and their immunogenicity in mice were carefully evaluated. Here, we report that HIV-1 envelope proteins are properly cleaved, glycosylated, and incorporated into VLP with Gag. The amount of VLP released into culture supernatants is comparable to those produced by insect cells infected with recombinant baculoviruses. Moreover, cryo-electron microscopy tomography revealed average 17 spikes per purified VLP, and antigenic epitopes on the spikes were recognized by the broadly neutralizing antibodies 2G12, b12, VRC01, and 4E10 but not by PG16. Finally, mice primed with DNA and boosted with VLP in the presence of CpG exhibited anti-envelope antibody responses, including ELISA-binding, neutralizing, antibody-dependent cell-mediated cytotoxicity and antibody-dependent cell-mediated viral inhibition, as well as envelope and Gag-specific CD8 T cell responses. Thus, we conclude that HIV-1 VLP produced by the S2 expression system has many desirable features to be developed into a vaccine component against HIV-1.
