VACCINE REGIMENS TO INDUCE CD4+ AND CD8+ T CELLS AGAINST SIV EPITOPES

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 目的：比较6种新的疫苗方案诱导针对SIV表位的CD4+和CD8+T细胞。 最近临床艾滋病毒疫苗的失败突显了更彻底地评估艾滋病毒基础科学以及测试新的疫苗方案和媒介的重要性。为了克服更多传统载体疫苗的局限性，我们的实验室开发了几种新的免疫策略。这些新方法将使我们能够直接启动特定的T细胞反应，这是以前使用其他疫苗载体和方案所不可能的。这应该使我们能够剖析特定的T细胞反应在SIV复制控制中的作用--对于占主导地位的CD8+T细胞和病毒特异性的CD4+T细胞的作用。 在这项拨款的R21阶段，我们将比较六种新的疫苗接种方案：多肽冲击的树突状细胞、多肽结合的纳米棒、多肽冲击的PBMC、融合到乙肝核心抗原(HBcAg)载体基因的SIV多肽、电穿孔DNA+IL-12和腺病毒5。 进度： 我们已经完成了R21阶段第一组和第二组动物的疫苗接种。这些动物接受三个Mamu-DRbw*606限制性的CD4表位冲击的自体树突状细胞，并用相同表位冲击的自体PBMC或多肽结合的纳米棒启动。在最后一次强化试验的两周后，没有任何动物产生Elispot或ICS检测到的任何可检测到的表位特异性CD4+T细胞反应。 HBcAg载体由Deborah Fuller博士完成，每个选定的Mamu-A*01限制性CD8表位插入单独的载体中。6只MAMU-A*01+动物，第3组和第4组，接受5剂HBcAg载体注射。在第五次接种后，没有动物产生任何可检测到的表位特异性CD8+T细胞反应；然而，它们对富勒博士提供的肝炎多肽确实有很好的反应，这表明它们确实接种了疫苗。在最终的启动一个月后，一组接受每一个A*01表位冲击的自体PBMC，而另一组接受与纳米表位结合的A*01表位。在增强这些动物后，我们仍然没有看到可检测到的表位特异性CD8+T细胞反应。 此外，我们还制备了含有3个CD4表位和5个CD8表位的DNA和AdenoVirus5载体。我们用这些结构免疫动物，希望能激发表位特异性的CD4+和CD8+T细胞反应。在Ad5加强免疫两周后，我们检测到针对疫苗中所有三个表位的高频SIV特异性CD4+T细胞反应。三只动物中有两只对所有三个表位做出了反应，而第三只动物对三个表位中的两个做出了反应。所有三只接受CD8表位的动物都对Env：Env TL9中的一个表位产生了高频反应。其中一只动物额外产生了Pol LV10特异性的CD8+T细胞反应。虽然表位特异性的CD4疫苗接种是成功的，但测试的疫苗方案中没有一种激发出几种高频表位特异性CD8+T细胞反应。 一份出版物正在筹备中。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Objective: To compare six novel vaccine regimens to induce CD4+ and CD8+ T Cells against SIV epitopes. Recent failures in clinical HIV vaccines have underscored the importance of more thoroughly evaluating basic science of HIV as well as testing new vaccine regimens and vectors. In an effort to overcome the limitations of more traditional vector-based vaccines, our laboratory has developed several novel immunization strategies. These new methods will allow us to directly prime specific T cell responses in a manner that previously has been impossible with other vaccine vectors and regimens. This should allow us to dissect the contributions of specific T cell responses in the control of SIV replication -- for the roles of both subdominant CD8+ T cells and virus-specific CD4+ T cells. In the R21 phase of this grant, we will compare six novel vaccination regimens: peptide-pulsed dendritic cells, peptide-conjugated nanobeads, peptide-pulsed PBMC, SIV peptides fused to a Hepatitis B core antigen (HBcAg) carrier gene, electroporated DNA+IL-12 and Adenovirus5. PROGRESS: We have completed vaccinating animals in groups one and two of the R21 phase. These animals received autologous dendritic cells pulsed with three Mamu-DRbw*606-restricted CD4 epitopes and primed with either autologous PBMC pulsed with the same epitopes or peptide-conjugated nanobeads. Two weeks after the final boosts, no animals made any detectable epitope-specific CD4+ T cell responses as detected by ELISPOT or ICS. The HBcAg vectors were completed by Dr. Deborah Fuller with each of the selected Mamu-A*01-restricted CD8 epitopes inserted into individual vectors. Six Mamu-A*01+ animals, groups three and four, received five doses of the HBcAg vector. After the fifth dose, no animals made any detectable epitope-specific CD8+ T cell response; however, they did respond well to Hepatitis peptide provided by Dr. Fuller indicating that they did receive doses of the vaccine. One month after the final prime, one group received autologous PBMC pulsed with each of the A*01 epitopes, while the other group received the A*01 epitopes conjugated to nanobeads. After boosting these animals, we still saw no detectable epitope-specific CD8+ T cell responses. Additionally, we made DNA and Adenovirus5 vectors containing the three CD4 epitopes and the five CD8 epitopes. We vaccinated animals with these constructs in the hope of eliciting epitope-specific CD4+ and CD8+ T cell responses. Two weeks after the Ad5 boost, we detected high frequency SIV-specific CD4+ T cell responses against all three epitopes in the vaccine. Two of the three animals made responses to all three epitopes, while the third animal made responses to two of the three epitopes. All three animals that received the CD8 epitopes made a high frequency response to one epitope in Env: Env TL9. One animal made an additional Pol LV10-specific CD8+ T cell response. While the epitope-specific CD4 vaccination was successful, none of the tested vaccine regimens elicited several high frequency epitope-specific CD8+ T cell responses. A publication is in preparation.