Basic and Applied Studies in Development of an AIDS Vacc

我们正在寻求一种复制能力腺病毒(Ad)-重组启动/亚单位增强方法作为艾滋病毒/艾滋病的预防性疫苗。研究表明，疫苗方案可以诱导体液、细胞和粘膜免疫，并在非人类灵长类动物模型中提供显著保护，导致这种方法进入第一阶段人体试验。与此同时，进一步的临床前研究正在评估编码更多病毒基因的重组体的实用性，研究新的亚单位蛋白启动子，并探索免疫途径，以获得最佳的整体免疫反应和保护效果。最近的一项研究对复杂的Ad5hr-SIV重组启动方案进行了研究，其中包括一种编码非肉豆蔻酰化SIV nef的方案，以及由gp120或代表包膜蛋白CD4结合位点的多肽聚合物增强的方案。结果表明，用编码3个或3个以上SIV基因的重组体免疫并用gp120加强免疫的猕猴，或仅用Ad5hr-SIVenv/rev重组体免疫但用多肽聚合物加强免疫的猕猴，39%的免疫效果良好。在用高致病性SIVmac251分离株攻击后，病毒血症要么无法检测到，要么被清除或控制在检测阈值。在感染的慢性期，细胞免疫反应与病毒血症的控制显著相关，而抗包膜结合抗体与急性期病毒血症的减少显著相关。这些抗体随后被证明介导了抗体依赖的细胞毒性(ADCC)。持续的研究正在调查ADCC在预防艾滋病毒复制和疾病进展方面的作用。在后续研究中，73%的高度保护猕猴继续表现出对第二次挑战的有效保护，一年后在没有干预免疫的情况下注射SIVmac251。这种持久的保护与体内CD8+T细胞耗尽所显示的细胞免疫反应有关。在黑猩猩模型中，我们已经证明，与含有相同插入的HIV基因的非复制型重组体相比，具有复制能力的Ad-HIVenv重组体的粘膜免疫诱导了增强的细胞免疫应答和更好的初始抗体应答。我们认为，复制能力载体可能对疫苗设计最有利，因为它可以在较低剂量下实现插入基因的等量表达，能够更有效地靶向粘膜诱导部位，能够更好地诱导作为天然佐剂的细胞因子和共刺激分子，并具有更大的产生更持久免疫的潜力。对新型HIVnef免疫原的研究表明，编码非肉豆蔻酰化nef的重组体比野生型nef能诱导更有效的细胞免疫应答。肉豆蔻化位点的去除可以防止MHC-I类分子和表达该蛋白的细胞表面的CD4下调，这是HIV可以避免强烈的宿主细胞免疫反应的一种机制。最后，编码HIV野生型Tat或易显性Tat突变体的重组子在小鼠中被证明是免疫原性的。重要的是，任何一种Ad-HIVtat重组体的免疫都能增强对共同注射的SIV Gag抗原的细胞免疫反应。正在进行的研究正在调查这些TAT重组体在非人类灵长类动物中的免疫原性和保护效果。

We are pursuing a replication-competent Adenovirus (Ad)-recombinant priming/subunit boosting approach as a prophylactic vaccine for HIV/AIDS. Studies showing that the vaccine regimen can elicit humoral, cellular, and mucosal immunity together with significant protection in non-human primate models has resulted in movement of this approach towards Phase I human trials. At the same time, further pre-clinical studies are evaluating the utility of recombinants encoding additional viral genes, investigating novel subunit protein boosts, and exploring routes of immunization for elicitation of the best overall immune responses and protective efficacy. A recent study investigated complex Ad5hr-SIV recombinant priming regimens, including one encoding a non-myristoylated SIV nef, together with boosting by either gp120 or a peptide polymer representing the CD4 binding site of the envelope protein. Impressive results were obtained which showed that 39% of macaques immunized with recombinants encoding 3 or more SIV genes and boosted with gp120, or immunized with the Ad5hr-SIVenv/rev recombinant only but boosted with the peptide polymer, were highly protected. Following challenge with the highly pathogenic SIVmac251 isolate, viremia was either undetectable or it was cleared or controlled at the threshold of detection. Cellular immune responses were significantly correlated with control of viremia during the chronic phase of infection, while anti-envelope binding antibodies were significantly correlated with reduced acute phase viremia. These antibodies were subsequently shown to mediate antibody-dependent cellular cytotoxicity (ADCC). Continuing studies are investigating the role of ADCC in protection against HIV replication and disease progression. In follow-up studies, 73% of the highly protected macaques continued to exhibit potent protection against a second challenge with SIVmac251 administered one year later with no intervening immunization. This durable protection was associated with cellular immune responses as shown by in vivo depletion of CD8+ T cells. In the chimpanzee model, we have shown that mucosal immunization with replication-competent Ad-HIVenv recombinants elicit enhanced cellular immune responses and better prime antibody responses compared to non-replicating Ad-recombinants containing the same inserted HIV gene. We believe the replication competent vector may be most advantageous for vaccine design, as it can achieve equivalent expression of inserted genes at lower doses, can more effectively target mucosal inductive sites, can better elicit cytokines and co-stimulatory molecules which serve as natural adjuvants, and has a greater potential for generating more persistent immunity. Investigation of novel HIVnef immunogens has shown that Ad-recombinants encoding non-myristoylated nef elicit more potent cellular immune responses compared to wild-type nef. Removal of the myristoylation site prevents down-modulation of both MHC-class I molecules and CD4 from the surface of cells expressing the protein, a mechanism by which HIV can avoid a strong host cellular immune response. Finally, Ad-recombinants encoding HIV wild-type tat or a transdominant tat mutant were shown to be immunogenic in mice. Importantly, immunization with either Ad-HIVtat recombinant resulted in enhanced cellular immune responses to a co-administered SIV Gag antigen. On-going studies are investigating the immunogenicity and protective efficacy of these tat recombinants in non-human primates.