Selecting HIV recombination libraries for stable Env trimer immunogens

描述(由申请人提供)：一种有效的疫苗是应对艾滋病毒/艾滋病全球流行的更好的解决方案。开发HIV-1疫苗的一个主要优先事项是确定一种能够诱导针对循环中的主要病毒的包膜糖蛋白(Env)尖峰的高滴度中和抗体(NAB)的免疫原。病毒粒子和感染细胞表面的功能(天然)形式是gp120-gp41异源三聚体。在诱导HIV-1NAB过程中的一个关键障碍并不是一般地诱导抗Env抗体，而是抗Env反应不成比例地被诱导到无关形式的Env及其亚单位，而不是天然的Env三聚体。可能导致对HIV免疫原的弱和/或分离的特异性NAB反应的因素是环境中的序列可变性和异质性，以及膜表面发现的三聚体天然形式的不稳定性和低拷贝数。 该项目的目标是识别、表征和开发病毒样颗粒(VP或VLP免疫原)，这些免疫原显示比野生型HIV-1更稳定、更同质、拷贝数更多的环境三聚体。从用DNA重组方法建立的不同的Env文库中，Env VP将受到各种不稳定条件的影响，然后被选择与天然构象和受体激活构象中的三聚体特异性NAB和NAB前体结合，同时对非中和抗体进行反筛选。为了鉴定产量更高和天然环境三聚体含量更高的VP，我们将再次使用NABS，从不同的gp41细胞质Tail和GAG文库中进行选择。我们将选择具有稳定、高拷贝数环境三聚体的VLP免疫兔子，并在HIV-1中和试验中测定针对相关毒株和异源毒株的血清NAB效价。我们将使用血清抗体和一组单抗在各种结合和中和试验中探索Lead Env免疫原，以确定野生型Env三聚体的抗原模拟程度。稳定的环境三聚体和抗血清的详细分子特征应该允许进一步优化铅免疫原和免疫方案，以引发针对不同HIV-1初级分离株的交叉反应NAB反应。我们希望拟议的研究将对基于环境三聚体的疫苗的开发做出积极贡献。

DESCRIPTION (provided by applicant): A preferable solution to the HIV/AIDS global pandemic is an effective vaccine. A major priority for HIV-1 vaccine development is to identify an immunogen that can elicit high titers of neutralizing antibody (nAb) against the envelope glycoprotein (Env) spikes on circulating primary viruses. The functional (native) form of Env on the surface of virions and infected cells is a gp120-gp41 heterotrimer. A critical barrier in eliciing HIV-1 nAb is not one of eliciting anti-Env Ab in general, but rather that anti-Env responses are disproportionately elicited to irrelevant forms of Env and its subunits, rather than to the native Env trimer. Factors that likely contribute to the elicitation of weak and/or isolate specific nAb responses to HIV immunogens are sequence variability and heterogeneity in Env, as well as lability and low copy number of the trimeric, native form that is found on membrane surfaces. The goal of this project is to identify, characterize and develop virus-like particles (VPs or VLPs immunogens that display Env trimers that are more stable, more homogeneously native and in higher copy number than that of wild-type HIV-1. From diverse Env libraries created using DNA recombination methods Env VPs will be subjected to a variety of destabilizing conditions and then selected for binding to trimer-specific nAbs and nAb precursors in native and receptor-activated conformations while counter-screening against non-neutralizing Abs. To identify VPs with improved yield and native Env trimer content, we will engineer and select from diverse gp41 cytoplasmic tail and Gag libraries, again using nAbs. We will immunize rabbits using selected VLPs with stable, high copy number Env trimers and determine serum nAb titers against related strains and heterologous isolates in HIV-1 neutralization assays. We will probe lead Env immunogens using serum Ab and a panel of mAbs in a variety of binding and neutralization assays to ascertain the extent of antigenic mimicry of wildtype Env trimers. Detailed molecular characterization of the stable Env trimers and antisera should allow further optimization of lead immunogens and immunization regimens to elicit crossreactive nAb responses against different primary isolates of HIV-1. We expect the proposed studies to positively contribute to the development of an Env trimer-based vaccine.