SELECTING ENV TRIMERS IN HIV-VLP VACCINE PRESENTATIONS

描述（由申请人提供）：在HIV-1感染期间，包膜蛋白（Env）以多种形式呈现给免疫系统。在目前公认的模型中，Env的功能形式是gp120/gp41异二聚体的三聚体复合物。Env的非功能形式包括单体gp120、未裂解的gp160前体和gp120已解离的gp41。功能三聚体已经进化成一个紧凑的结构；事实上，虽然大多数单克隆抗体（mab）只能识别非功能形式的Env，但中和型mab似乎也能结合功能性的Env三聚体。从颗粒和受感染细胞中释放的Env片段被认为是导致HIV-1感染的中和反应质量普遍较差的原因。然而，新的证据表明，感染性HIV-1颗粒也携带非功能形式的Env。为了追求开发一种能够引发强效中和抗体的HIV-1疫苗的目标，我们选择了HIV-1假病毒粒子作为模型免疫原。在Specific Aim 1中，我们将使用一套全面的技术来尝试生成假病毒粒子，这些假病毒粒子只携带功能性三聚体，这些三聚体只能被中和的单克隆抗体识别。鉴于三聚体复合物紧凑、抗抗体的性质，我们将在Specific Aim 2中扩展我们的研究，将受体参与形式的Env作为替代中和靶标。env受体结合诱导暴露其他隐结构。虽然这些结构只在自然感染中短暂暴露，但它们是被特征良好的中和单克隆抗体识别的可信的中和靶点，例如2F5。为此，我们产生了一种假病毒体突变体，它附着在易感细胞上，但只以氧化还原依赖的方式融合。使用该模型，我们将研究HIV-I+血清中的附着后中和。在特异性目标3中，我们将测试i)只携带功能性三聚体的假病毒粒子和ii)附着于自体猕猴淋巴细胞的受体接合假病毒粒子的免疫原性。

DESCRIPTION (provided by applicant): During HIV-1 infection, the envelope protein (Env) is presented to the immune system in many forms. In the currently accepted model, the functional form of Env is a trimeric complex of gp120/gp41 heterodimers. Examples of non-functional forms of Env include monomeric gp120, uncleaved gp160 precursor and gp41 from which gp120 has dissociated. The functional trimer has evolved to be a compact structure; indeed, while most monoclonal antibodies (mAbs) recognize only non-functional forms of Env, neutralizing mAbs appear to also bind the functional Env trimer. Env fragments released from particles and infected cells have been considered responsible for the generally poor quality neutralizing response to HIV-1 infection. However, new evidence suggests that infectious HIV-1 particles also bear non-functional forms of Env. In pursuing the goal of developing an HIV-1 vaccine able to elicit potent neutralizing antibodies, we have chosen HIV-1 pseudovirions as model immunogens. In Specific Aim 1, we will use a comprehensive set of techniques in an attempt to generate pseudovirions that exclusively bear functional trimers that are only recognized by neutralizing mAbs. Given the compact, antibody-resistant nature of the trimeric complex, we will expand our studies in Specific Aim 2 to include the receptor-engaged form of Env as an alternative neutralization target. Env-receptor binding induces exposure of otherwise cryptic structures. Although these structures are only transiently exposed in natural infection, they are plausible neutralization targets recognized by well-characterized neutralizing mAbs, exemplified by 2F5. To this end, we have generated a pseudovirion mutant that attaches to susceptible cells but only fuses in a redox-dependent manner. Using this model, we will investigate post-attachment neutralization in HIV-I+ serum. In Specific Aim 3, we will test the immunogenicity of i) pseudovirions bearing only functional trimers and ii) receptor-engaged pseudovirions attached to autologous macaque lymphocytes.