The identification of HLA-A*0201 restricted cytotoxic T lymphocyte epitope in HPV-16 E5 and The evaluation of immune response to different HPV-16 E7 tumor vaccines by a prime-boost strategy in C57BL/6 mice

• Main Authors: Mei-Fang Lin, 林梅芳
• Other Authors: Show-Lin Chen
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/27546754690926096459

碩士 === 國防醫學院 === 微生物及免疫學研究所 === 92 === Subject Ι : The identification of HLA-A*0201 restricted cytotoxic T lymphocyte epitope in HPV-16 E5 Previously, we have identified HPV-16 E5 as a tumor rejection antigen that can induce cytotoxic T lymphocyte(CTL) to protect tumor growth (44). And we have mapped the CTL epitope of E5 protein by analyzing E5-specific, CD8+ interferon-γ+ (IFN-γ+) double-positive cells in H-2b C57BL/6 mice with flow cytometry. The result showed the region spanning amino acid 25 to 33 contains the potential Db-restricted CTL epitop in H-2b C57BL/6 mice(76). So I tried to identify HLA-A* 0201 specific E5 CTL epitope in HLA-A* 0201 trnasgenic mice. My strategies include:《1》Using comupter program to scan the entire E5 protein sequence for peptides that match the motifs for HLA-A* 0201.《2》 Testing synthetic peptides by stabilization assay on the surface of T2 cells.《3》 Staining intracytoplasmic cytokine of CTL elicited by HLA-A*0201 E5 CTL epitope peptides in HLA-A*0201 transgenic mice.《4》Using the fluorometric assessment of T-lymphocyte antigen-specific lysis (FATAL) assay(47) to test CTL activity induced by HLA-A*0201 E5 CTL epitope peptides in HLA-A*0201 transgenic mice.《5》Performing tumor elimination assay by HLA-A*0201 E5 CTL epitope peptides in HLA-A*0201 transgenic mice.《6》 Generating of HPV-E5 specific cytotoxic T lymphocyte by using human peripheral blood monocyte transduced with rAd-E5(32)。 Subject Π : The evaluation of immune response to different HPV-16 E7 tumor vaccines by a prime-boost strategy in C57BL/6 mice From the previous project, we have established recombinant Listeria monocytogenes (40) and adeno-associated virus (43) encoding HPV-E7 as a tumor vaccine that successfully elicit CTL to eradicate tumor growth in syngeneic animal models. The prime-boost immunization strategy generates a high frequency and high avidity CD8+ cytotoxic T lymphocyte population. The prime and boost vaccines use different vectors but have the same antigen. We used two different vector systems encoding HPV-16 E7 gene as tumor vaccines for cervical cancer. The two vector systems are rAAV-E7CTL.hsp and rL.M.-E7. The purpose of this project is to evaluate the efficacy of these two systems in C57BL/6 mice individually or in different combinations by prime-boost immunization, and try to find the best vaccination protocol for future clinical trials.