VLP vaccines against HPV-induced tumors

人类乳头瘤病毒（HPV）是导致宫颈癌的必要原因，宫颈癌是全世界妇女中第三大常见癌症。几乎在所有宫颈癌中都检测到HPV DNA，其中约50-60%是由高风险的HPV 16型引起的。有效的HPV复制条件尚未在体外模拟，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已成为疫苗试验中的主要策略，通过诱导中和抗体来预防未来的HPV病变，但它们不太可能对数百万已经感染的妇女产生影响，并且对不再表达L1基因的现有宫颈癌也没有任何影响。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，并且是细胞转化状态所必需的。因此，将E6和E7蛋白掺入嵌合VLP （cVLP）中可以赋予基于VLP的疫苗治疗潜力。VLP在诱导T细胞反应、激活DC、肿瘤预防以及肿瘤治疗和基因传递方面的巨大潜力已经得到证实。基于这些研究，我们现在提出以下新的探索目标：1)优化HPV VLP作为抗原递送系统的使用，2)使用HPV VLP作为绘制人内源性HPV16 E6/E7蛋白抗原表位的工具，3)使用HPV VLP探索HPV与朗格汉斯细胞的相互作用，4)使用HPV VLP作为基因递送系统。为实现这些目标，将采用以下方法：1)通过不同的cVLP或不同的途径递送HPV16 E6/E7蛋白，以避免cVLP对E6/E7蛋白的抗体活性，从而中和重复接种E6/E7蛋白；2)用自体DC负载表达E6/E7蛋白的cVLP体外免疫人外周血T细胞；3)HPV VLP与人和小鼠朗格汉斯细胞的相互作用和活化。4)向宫颈癌细胞传递新的潜在肿瘤抑制基因或细胞因子基因，向树突状细胞传递抗原编码基因或细胞因子基因。这些综合目标将增加HPV VLP作为HPV诱导宫颈癌治疗剂的潜力和多功能性。

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.