VLP vaccines against HPV-induced tumors

人类乳头瘤病毒(HPV)是宫颈癌的必要原因，宫颈癌是世界上第三种最常见的女性癌症。几乎在所有宫颈癌中都能检测到HPVDNA，其中约50%-60%是由高危HPV16型引起的。目前还没有人在体外模拟有效的HPV复制条件，从而阻碍了经典疫苗的发展。然而，当病毒的主要病毒粒子蛋白(L1)在真核细胞中过度表达时，它会自组装成病毒样颗粒(VLP)。HPV VLP已经成为通过诱导中和抗体来预防未来HPV损害的疫苗试验的领先策略，但它们不太可能对已经感染的数百万女性产生影响，也不会对不再表达L1基因的现有宫颈癌产生任何影响。然而，它们确实表达早期基因产物E6和E7，这些基因产物在宫颈癌中选择性地保留和表达，是细胞转化状态所必需的。因此，将E6和E7蛋白整合到嵌合VLP(CVLP)中可以为基于VLP的疫苗提供治疗潜力。VLP在诱导T细胞反应、激活DC、在肿瘤预防环境中以及在初步的肿瘤治疗环境和基因传递方面都显示出巨大的潜力。基于这些研究，我们现在建议探索以下新的目标：1)优化使用HPV VLP作为抗原递送系统，2)使用HPV VLP作为工具来定位人内源性呈现的HPV16E6/E7蛋白的抗原表位，3)使用HPV VLP来探索HPV与朗格汉斯细胞的相互作用，以及4)使用HPV VLP作为基因递送系统。为了达到这些目的，将使用以下方法：1)通过不同的cVLP或不同的途径传递HPV16E6/E7蛋白，以避免针对cVLP的抗体活性，从而中和针对E6/E7蛋白的重复免疫；2)体外用负载表达E6/E7蛋白的自体DC免疫人外周血T细胞；3)HPV VLP与人和小鼠郎格汉斯细胞的相互作用和激活；以及4)将新的潜在肿瘤抑制基因或细胞因子基因传递给宫颈癌细胞，并将抗原编码基因或细胞因子基因传递给树突状细胞。这些联合的目的将增加HPV VLP作为治疗HPV诱导的宫颈癌的药物的潜力和通用性。

Human papillomaviruses (HPV are a necessary cause for cervical carcinoma, the third most common cancer among women worldwide. HPV DNA is detected in virtually every cervical carcinoma and about 50-60% of these are accounted for by the high risk HPV type 16. Efficient HPV replication conditions have not yet been mimicked in vitro thereby hampering the development of classical vaccines. However, when the major virion protein of the virus (L1) is over- expressed in eukaryotic cells, it self-assembles into virus-like particles (VLP). HPV VLP have become the leading strategy in vaccine trials for prevention of future HPV lesions by inducing neutralizing antibodies, but they are unlikely to have effects in the millions of women that are already infected and will not have any effect on existing cervical cancers that do no longer expression the L1 genes. They do, however, express the early gene products E6 and E7 that are selectively retained and expressed in cervical carcinoma and necessary for the transformed state of the cells. Therefore, incorporation of the E6 and E7 protein into chimeric VLP (cVLP) could confer a therapeutic potential to a VLP based vaccine. The tremendous potential of VLP has been shown in inducing T cell responses, in activating DC, in tumor preventive setting and, in preliminary ways also in tumor therapeutic settings and gene delivery. Based on these studies we now propose to explore the following new aims: 1) optimizing the use of HPV VLP as an antigen delivery system, 2) the use of HPV VLP as a tool for mapping human endogenously presented antigenic epitopes of HPV16 E6/E7 proteins, 3) the use of HPV VLP to explore the interaction of HPV with Langerhans cells, and 4) the use of HPV VLP as a gene delivery system. To achieve these aims the following methodology will be used: 1) delivery the of the HPV16 E6/E7 protein by different cVLP or different routes in order to avoid antibody activity against the cVLP that will neutralize repeated vaccination against the E6/E7 proteins, 2) in vitro immunization of human peripheral blood T cells with autologous DC loaded with cVLP expressing E6/E7 protein, 3) HPV VLP interaction and activation of human and mouse Langerhans cells, and 4) delivery of a new potential tumor suppressor gene or cytokine gene to cervical cancer cells and an antigen encoding gene or cytokine gene to dendritic cells. These combined aims will increase the potential and versatility of HPV VLP as therapeutic agents for HPV induced cervical cancer.