ENHANCING HPV-16 E6-SPECIFIC ANTITUMOR IMMUNITY

描述(申请人提供)：针对人乳头瘤病毒(HPV)E6蛋白的免疫疗法可能为控制HPV相关的宫颈恶性肿瘤提供机会。已证实，T细胞介导的免疫是抵御HPV相关病变的最重要的组成部分之一，树突状细胞(DC)是体内最强大的专职抗原提呈细胞(APC)，在体内启动T细胞的辅助和杀伤。此外，已有研究表明，通过基因枪皮内注射DNA疫苗是将DNA疫苗输送到体内专业APC的有效手段。因此，我们使用基因枪递送系统来测试需要将DNA疫苗直接递送到专业APC的策略。我们已经成功地测试了几种细胞内靶向策略，这些策略增强了MHC I类和II类的处理和呈现，并产生了令人印象深刻的结果。最近，我们测试了多种抗凋亡因子在与DNA疫苗联合应用时提高DC存活和抗原特异性CD8+T细胞免疫反应的能力。由于细胞内靶向和抗凋亡策略通过不同的机制改变DC，我们已经能够将延长DC寿命的抗凋亡策略与增强DC提呈MHC I类和II类抗原的细胞内靶向策略结合起来，以提高DNA疫苗的效力。虽然编码抗原的DNA和编码抗凋亡蛋白的DNA共同接种可以显著增强DNA疫苗的效力，但编码抗凋亡蛋白的DNA的使用引起了与致瘤性相关的重大问题。一种相对较新的技术，使用针对促凋亡蛋白的小干扰RNA(SiRNA)的RNA干扰(RNAi)可能会提供类似的效果，同时缓解人们对致癌性的担忧。因此，在目前的提案中，我们计划测试这样一个假设，即皮内将编码HPV-16E6的DNA疫苗与靶向关键促凋亡蛋白的siRNA一起递送到抗原表达的树突状细胞将延长转基因DC的寿命，并导致体内增强E6特异性T细胞介导的免疫反应和抗肿瘤效应。

DESCRIPTION (provided by applicant): Immunotherapy that targets the human papillomavirus (HPV) E6 protein may provide an opportunity to control HPV-associated cervical malignancies. It has been established that T cell-mediated immunity is one of the most crucial components of defense against HPV-associated lesions and that dendritic cells (DCs) are the most potent professional antigen presenting cells (APCs) that prime helper and killer T cells in vivo. Furthermore, it has been shown that intradermal administration of DNA vaccines via gene gun represents an efficient means of delivering DNA vaccines into professional APCs in vivo. We have therefore used the gene gun delivery system to test strategies that require direct delivery of DNA vaccines to professional APCs. We have successfully tested several intracellular targeting strategies that enhance MHC class I and class II processing and presentation and have generated impressive results. Recently, we tested a variety of anti-apoptotic factors for their ability to enhance DC survival and antigen-specific CD8+ T cell immune responses when co-administered with DNA vaccines. Because intracellular targeting and anti-apoptotic strategies modify DCs via different mechanisms, we have been able to combine anti-apoptotic strategies for prolonging DC life with intracellular targeting strategies for enhancing MHC class I and II presentation of antigen by DCs to improve DNA vaccine potency. While coadministration of DNA encoding antigen with DNA encoding anti-apoptotic proteins can significantly enhance DNA vaccine potency, the use of DNA encoding anti-apoptotic proteins raises significant concerns related to oncogenicity. A relatively new technology, RNA interference (RNAi) using small interfering RNA (siRNA) targeting pro-apoptotic proteins may provide similar effects while alleviating concerns for oncogenicity. Thus, in the current proposal we plan to test the hypothesis that intradermal delivery of a DNA vaccine encoding HPV-16 E6 in conjunction with siRNA targeting key pro-apoptotic proteins to antigen-expressing dendritic cells would prolong transfected DC life and lead to enhanced E6-specific T cell-mediated immune responses and antitumor effects in vivo.