ENHANCING HPV-16 E6-SPECIFIC ANTITUMOR IMMUNITY

描述（由申请人提供）：针对人乳头瘤病毒（HPV） E6蛋白的免疫疗法可能提供控制HPV相关宫颈恶性肿瘤的机会。已经确定T细胞介导的免疫是防御hpv相关病变的最重要组成部分之一，树突状细胞（dc）是体内最有效的专业抗原呈递细胞（APCs），主要是辅助和杀伤T细胞。此外，研究表明，通过基因枪皮内给药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装甲运兵车的策略。我们已经成功地测试了几种细胞内靶向策略，增强了MHC I类和II类的加工和呈现，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疫苗联合使用时提高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修饰DC，我们已经能够将延长DC寿命的抗凋亡策略与增强DC的MHC I类和II类抗原呈递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与编码抗凋亡蛋白的DNA共同给药可以显著提高DNA疫苗的效力，但使用编码抗凋亡蛋白的DNA引起了与致癌性相关的重大关注。一种相对较新的技术，RNA干扰（RNAi）使用小干扰RNA （siRNA）靶向促凋亡蛋白，可能提供类似的效果，同时减轻对致癌性的担忧。因此，在目前的提案中，我们计划验证这样一种假设，即将编码HPV-16 E6的DNA疫苗与靶向关键促凋亡蛋白的siRNA结合皮内递送到表达抗原的树突状细胞，可以延长转染的DC寿命，并增强E6特异性T细胞介导的免疫反应和体内抗肿瘤作用。

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.