Conserved Element DNA Vaccine

一种理想的艾滋病毒疫苗应该能够预防所有的艾滋病毒-1变异。因此，艾滋病毒疫苗开发的一个重要方面是免疫原的选择，这必须考虑到不同艾滋病毒进化支的多样性和能够诱导相关免疫反应的关键表位的鉴定，避免潜在的免疫优势“诱饵”表位。事实上，我们报道了Env对诱导Gag细胞反应的有效影响。为了解决这些问题，我们正在探索利用马赛克和共识分子最大化免疫强度和广度的方法，并专注于HIV的高度保守区域，以诱导对几乎不变的蛋白质组片段的免疫反应，这对病毒的功能至关重要，同时排除对可变和潜在的免疫优势“诱饵”表位的反应。利用后者，我们开发了针对HIV Gag和Env高度保守区域的原型疫苗。根据严格的保守性、功能重要性、广泛的hla覆盖范围和与病毒控制的关联，设计了编码HIV-1 Gag保守元件（CE）的免疫原。在小鼠和猕猴的概念验证研究中，我们证明了用Gag CE DNA免疫可以引起针对CE的强大的细胞和体液免疫反应，这是用全长p55gag DNA接种无法实现的。该疫苗可诱导针对亚显性表位的强效细胞毒性T细胞反应。重要的是，我们证明了CE DNA的引物和p55gag DNA的增强极大地增强了CE特异性反应，而p24CE+gag DNA的增强在幅度和广度上都最大化。因此，我们确定了一种新的有效策略来最大限度地提高对Gag的反应，并提供了一种新的策略来改变免疫优势等级，并诱导对亚显性表位的强大免疫反应，有效地靶向病毒的阿喀琉斯之脚跟。我们发现这种DNA疫苗方案诱导了强有力的记忆反应，可以在2年后通过CE DNA加强疫苗快速回忆起来。表达HIV Gag/Poland Env的重组修饰安卡拉牛苗（rMVA）加强疫苗也能有效地唤起CE记忆反应。这些在猕猴身上进行的研究表明，这种疫苗方案可以应用于人类，其中DNA prime - rMVA增强已经诱导了有希望的免疫原性。我们进一步开发了CE的SIV同源物，并证明了CE DNA的引物和CE+gag DNA增强疫苗接种显著增加了细胞毒性T细胞对亚显性高度保守的gag表位的反应，并最大化了反应广度。这些数据反映了我们在HIV p24CE疫苗中的发现，并为我们探索Gag CE DNA疫苗在猕猴模型中的功能应用提供了工具。SIV Gag ce特异性T细胞能够在自体体外病毒抑制试验中减少病毒感染，这一重要特征已被证明与受感染动物体内病毒控制相关。疫苗诱导的CE特异性T细胞在SIV感染后扩增，表明纳入DNA疫苗的预测CE表位即使在其自然环境中也会被感染细胞处理和暴露。因此，我们人工生成的免疫原在CE之间含有连接物，以优化CE肽的蛋白水解过程和MHC结合，产生与病毒感染细胞相似的肽。我们发现CE反应有助于控制病毒血症。这种CE DNA疫苗的应用扩展到我们在猕猴模型中的治疗性疫苗试验。我们已将环境行政长官的概念进一步扩展至爱滋病环境。正如在HIV Gag CE中发现的那样，我们报道了HIV Env中鉴定的CE是亚优势的，并且我们的新型DNA疫苗方案通过包含CE DNA引物来改变免疫层次，并诱导识别这些表位的强大的细胞毒性T细胞反应。与Gag CE DNA疫苗一起，这为我们提供了一种疫苗，以最大限度地提高Gag和Env的强度和广度。我们正在HIV疫苗试验网络(HVTN)/DAIDS/ niaid支持的临床试验（HVTN 119；于2017年10月4日开放；NCT03181789）中翻译新的HIV Gag CE DNA疫苗概念（CE DNA引物-CE+全长Gag DNA boost）到临床，目的是测试我们的p24CE DNA疫苗概念与包含完整p55Gag蛋白的优化免疫原相比是否能引起更好的呼吸和Gag反应。该疫苗包括Profectus的GENEVAX IL-12 DNA作为分子佐剂和体内电穿孔作为DNA递送方法，我们的研究表明，这两种疫苗成分对在猕猴中诱导有效的T细胞反应最为重要。HVTN 119将结合我们多年来在疫苗开发中取得的几个里程碑（DNA表达载体、佐剂和递送）。我们还在艾滋病临床试验组的支持下，在I/IIb期试验中翻译Gag CE DNA疫苗（ACTG A5369; NCT03560258，计划于2018年8月17日开始）。在这项试验中，CE DNA疫苗将在HAART下的HIV感染者中进行测试，目的是测试HIV CE疫苗方案与p55gag DNA疫苗相比的安全性和免疫原性，以T细胞反应的广度和幅度作为关键终点。尽管抗逆转录病毒疗法（ART）降低了感染者与艾滋病毒相关的死亡率和发病率，但根除艾滋病毒的治疗方案尚未实现。我们的目标是开发和测试能够减少或消除病毒库的免疫治疗方法。诱导针对病毒亚优势高度保守区域的广泛T细胞反应提供了一种独特的可能性，可以探索我们的假设，即这些高细胞毒性T细胞反应具有靶向HIV致命弱点的弹药。HVTN119和A5369试验将使我们能够探索将我们的数据从小鼠和猕猴转化为人类。

An ideal HIV vaccine should provide protection against all HIV-1 variants. Thus, an important aspect of HIV vaccine development is the selection of immunogens, which has to take into consideration the diversity of the different HIV clades and the identification of the critical epitopes able to induce relevant immune responses, avoiding potential immunodominant "decoy" epitopes. Indeed, we reported a potent impact of Env on the induction of Gag cellular responses. To address these problems, we are exploring approaches to maximize immunological strength and breadth using mosaic and consensus molecules as well as focusing on highly conserved regions of HIV to induce immune responses to nearly invariable proteome segments, essential for the function of the virus, while excluding responses to variable and potentially immunodominant "decoy" epitopes. Using the latter, we have developed prototype vaccines targeting highly conserved regions within HIV Gag and Env. Immunogens were engineered encoding conserved elements (CE) of HIV-1 Gag selected on the basis of stringent conservation, functional importance, broad HLA-coverage, and association with viral control. In proof-of-concept studies in mice and macaques, we demonstrated that immunization with Gag CE DNA elicits robust cellular and humoral immune responses against CE, which cannot be achieved by vaccination with the full-length p55gag DNA. This vaccine induces robust cytotoxic T cell responses targeting subdominant epitopes. Importantly, we demonstrated that priming with CE DNA and boosting with p55gag DNA greatly augments the CE-specific responses and that inclusion of the p24CE+gag DNA in the boost maximizes both magnitude and breadth. Thus, we identified a novel and effective strategy to maximize responses against Gag and provide a novel strategy to shift the immunodominance hierarchy and to induce robust immune responses to subdominant epitopes, effectively targeting the Achilles' heel of the virus. We showed that this DNA vaccine regimen induces potent memory responses that can be rapidly recalled 2 years later by CE DNA booster vaccinations. The CE memory response can also be potently recalled by recombinant Modified Vaccinia Ankara (rMVA) booster vaccination, expression HIV Gag/Poland Env. These studies in macaques suggest application of this vaccine regimen in humans where DNA prime - rMVA boost has induced promising immunogenicity. We further developed an SIV homolog of the CE and demonstrated that priming with CE DNA followed by CE+gag DNA booster vaccination significantly increased cytotoxic T cell responses to subdominant highly conserved Gag epitopes and maximized response breadth. These data mirror our findings from the HIV p24CE vaccine and provide us with a tool to explore the functional applications of the Gag CE DNA vaccine in the macaque model. SIV Gag CE-specific T cells are able to reduce viral infection in an autologous in vitro virus inhibition assay, an important feature that has been shown to correlate with in vivo viral control in infected animals. The vaccine-induced CE-specific T cells were expanded upon SIV infection, indicating that the predicted CE epitopes incorporated in the DNA vaccine are processed and exposed by infected cells even in their natural context. Thus, our artificially generated immunogen containing linkers between the CE to optimize proteolytic processing and MHC association of the CE peptides, generates similar peptides as the virus infected cells. We found that CE responses contribute to control of viremia. Application of this CE DNA vaccine is expanded to our therapeutic vaccine trials in the macaque model. We have further expanded the CE concept to HIV Env. As found for HIV Gag CE, we reported that the identified CE in HIV Env are subdominant and that our novel DNA vaccine regimen by including a CE DNA prime alters the immune hierarchy and enable induction of robust cytotoxic T cell responses recognizing these epitopes. Together with the Gag CE DNA vaccine, this provides us with a vaccine to maximize magnitude and breadth of both Gag and Env. We are translating the novel HIV Gag CE DNA vaccine concept (CE DNA prime-CE+full length gag DNA boost) to the clinic in the HIV Vaccine Trial Network (HVTN)/DAIDS/NIAID-supported clinical trial (HVTN 119; opened October 4, 2017; NCT03181789) with the aim to test whether our p24CE DNA vaccine concept elicits superior breath and magnitude of Gag responses compared to the optimized immunogen comprising the complete p55Gag protein. This vaccine includes Profectus' GENEVAX IL-12 DNA as molecular adjuvant and in vivo electroporation as DNA delivery method, two vaccine components our research had shown to be of outmost importance to induce potent T cell responses in macaques. HVTN 119 will combine several of milestones (DNA expression vectors, adjuvants and delivery) we have achieved over many years in vaccine development. We are also translating the Gag CE DNA vaccine in a phase I/IIb trial (ACTG A5369; NCT03560258; planned opening August 17, 2018) supported by the AIDS Clinical Trial Group. In this trial, the CE DNA vaccine will be tested in HIV-infected persons under HAART with the aim to test safety and the immunogenicity of the HIV CE vaccine regimen in comparison to p55gag DNA vaccine, with breadth and magnitude of T cell responses as key endpoints. Although Antiretroviral Therapy (ART) has reduced HIV-related mortality and morbidity for infected individuals, a therapeutic regimen to eradicate HIV has not been achieved. Our goal is to develop and test immunotherapeutic methods that can lead to virus reservoir reduction or elimination. The induction of broad T cell responses targeting the subdominant highly conserved regions of the virus provides a unique possibility to explore our hypothesis that these highly cytotoxic T cell responses have bear the ammunition to target the Achilles's heel of HIV. Together, HVTN119 and A5369 trial will allow us to explore the translation of our data from mice and macaques to humans.