A Universal Env Immunogen for all HIV-1 Subtypes

描述（由申请人提供）：HIV基因组是高度可变的，对于最流行的M组病毒，它被分为9个亚型。亚型间的遗传差异可达30%。这种高水平的遗传变异已成为艾滋病疫苗开发的主要障碍。为了克服这个问题，我们为所有亚型生成了M组共识（con/con）环境基因。这种con/con env基因与任何亚型env基因之间的遗传距离大约是不同亚型之间的遗传距离的一半。Western blot和ELISA检测显示con/con Env蛋白能与sCD、单克隆抗体和患者血清结合。Biacore实验表明，它不仅可以与中和抗体和非中和抗体结合，还可以发生构象变化，以天然gp120的形式与17b mAb结合。我们的初步结果表明con/con Env蛋白表达线性、不连续和多糖抗体表位，并含有血清抗体的交叉反应表位。理论预测表明，所有特征良好的T细胞表位及其切割位点都保留在该蛋白上。这些数据表明con/con Env蛋白上的抗体和CTL表位可以被加工并提呈给免疫系统。因此，con/con Env蛋白可能是所有HIV-1亚型的通用免疫原。在目前的提案中，我们将通过测量豚鼠和小鼠的跨枝中和活性和细胞免疫反应来确定这种人工衍生的蛋白质是否可以诱导比亚型特异性Env免疫原更广泛的免疫反应。这些动物将用裸DNA进行免疫，然后用表达Env蛋白的rgp120或重组痘苗病毒（rVV）进行增强。我们将收集豚鼠的血清样本，以确定con/con Env蛋白是否会引发更高滴度的交叉进化中和抗体，以及rgp120或rVV是否能进一步增强体液免疫反应。我们还将从免疫小鼠中分离脾细胞，通过比较亚型特异性Env免疫原，确定con/con Env蛋白是否能单独或与rVV增强一起诱导强的跨进化细胞免疫反应。这项研究的结果将确定con/con Env蛋白是否可以作为所有HIV-1亚型的通用免疫原，并将作为设计广泛反应性免疫策略的指导方针。

DESCRIPTION (provided by applicant): HIV genomes are highly variable and have been classified into nine subtypes for most prevalent group M viruses. The genetic divergence among the subtypes can be up to 30%. This high level of genetic variation has become a major hurdle for AIDS vaccine development. To overcome this problem, we have generated a group M consensus (con/con) env gene for all subtypes. The genetic distance between this con/con env gene and any subtype env gene is about the half of that between different subtypes. Western blot and ELISA assays show that the con/con Env protein can bind to the sCD, mAbs and patient sera. The Biacore assay shows that it not only can bind to the neutralizing antibodies and non-neutralizing antibodies but also can undergo conformational changes to bind to 17b mAb as native gp120s. Our preliminary results show that the con/con Env protein expresses linear, discontinuous and glycan antibody epitopes, and it contains cross-reactive epitopes for serum antibodies. Theoretical prediction shows that all well-characterized T cell epitopes and their cleavage sites are preserved on this protein. This data suggests that antibody and CTL epitopes on the con/con Env protein can be processed and presented to the immune system. Therefore, the con/con Env protein may serve as a universal immunogen for all HIV-1 subtypes. In the current proposal we will determine whether such artificially derived proteins can induce broader immune responses than subtype-specific Env immunogens by measuring both cross-clade neutralization activity and cellular immune responses in guinea pigs and mice. The animals will be immunized with naked DNA and then boosted with either rgp120 or recombinant vaccinia viruses (rVV) expressing Env proteins. We will collect serum samples from guinea pigs to determine if higher titers of cross-clade neutralizing antibodies will be elicited by the con/con Env protein, and if humoral immune responses can be further augmented by either rgp120 or rVV. We will also isolate the splenocytes from immunized mice to determine if the con/con Env protein can induce strong cross-clade cellular immune responses alone or with rVV boost by comparing to subtype specific Env immunogens. Results from this study will determine if con/con Env protein can serve as a universal immunogen for all HIV-1 subtypes and will be used as a guideline to design broadly reactive immunization strategies.