SELECTING ENV TRIMERS IN HIV-VLP VACCINE PRESENTATIONS

描述(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杂二聚体的三聚体复合体。非功能形式的env的例子包括单体gp120、未切割的gp160前体和gp41，其中gp120已经解离。功能三聚体已经进化成一个紧凑的结构；事实上，虽然大多数单抗(MAbs)只识别非功能性形式的Env，但中和性mAb似乎也结合了功能性Env三聚体。从颗粒和受感染细胞中释放的Env片段被认为是导致对HIV-1感染的中和反应普遍较差的原因。然而，新的证据表明，具有传染性的HIV-1颗粒也带有非功能性形式的环境病毒。在追求开发一种能够诱导有效中和抗体的HIV-1疫苗的目标时，我们选择了HIV-1假病毒粒子作为模型免疫原。在特定的目标1中，我们将使用一套全面的技术，试图产生专门携带只有中和单抗才能识别的功能三聚体的假病毒粒子。鉴于三聚体复合体的致密、抗抗体的性质，我们将扩大我们在特定目标2的研究，以包括与受体结合的形式的Env作为替代中和靶点。Env受体结合会导致原本隐秘的结构暴露。虽然这些结构只在自然感染下短暂暴露，但它们是被特征良好的中和单抗识别的看似合理的中和靶点，以2F5为例。为此，我们产生了一个假病毒突变体，它附着在敏感细胞上，但只以氧化还原依赖的方式融合。利用这个模型，我们将研究HIV-I+血清中的附着后中和。在特定的目标3中，我们将测试I)仅带有功能三聚体的假病毒粒子和II)与自体猕猴淋巴细胞结合的受体参与的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.