1. Human CTL epitopes encoded by human papillomavirus type 16 E6 and E7 identified through in vivo and in vitro immunogenicity studies of HLA-A*0201-binding peptides
M E Ressing, A Sette, R M Brandt, J Ruppert, P A Wentworth, M Hartman, C Oseroff, H M Grey, C J Melief, W M Kast J Immunol. 1995 Jun 1;154(11):5934-43.
Human papillomavirus type 16 (HPV16) is strongly associated with cervical carcinogenesis. The HPV16 E6 and E7 oncoproteins are constitutively expressed in the majority of cervical tumor cells and are, therefore, attractive targets for CTL-mediated immunotherapy. In mice, the outgrowth of a lethal dose of HPV16-induced tumor cells has been prevented by vaccination with a CTL epitope encoded by HPV16 E7, indicating the feasibility of peptide immunization to obtain antitumor CTL responses. In the present study, the immunogenicity of 9 HLA-A*0201-binding peptides encoded by HPV16 E6 and E7 was analyzed in vivo in HLA-A*0201Kb transgenic mice and in vitro in CTL cultures induced from PBMC of HLA-A*0201+ healthy donors. Four peptides with a good binding affinity were immunogenic in HLA-A*0201Kb transgenic mice, and three of them were also highly immunogenic in CTL induction experiments with PBMC of HLA-A*0201+ healthy donors. Human CTL clones specific for these three peptides were capable of lysing the HPV16 E7-containing HLA-A*0201+ cervical carcinoma cell line CaSki. These E7-derived peptides (11-20, YMLDLQPETT; 82-90, LLMGTLGIV; 86-93, TLGIVCPI), therefore, are likely to represent naturally processed human CTL epitopes of HPV16. Additionally, these three HPV16-encoded peptides have the highest affinity of binding to the HLA-A*0201 molecule. In this study, peptides with a lower binding affinity were less immunogenic. Therefore, our data illustrate that the HLA-binding affinity of a peptide has a major impact on its immunogenicity. In conclusion, we have identified immunogenic peptides encoded by HPV16 E6 and E7 that could be used in vaccines for the prevention and treatment of cervical carcinoma.
2. HPV type 16 protein E7 HLA-A2 binding peptides are immunogenic but not processed and presented
M Bauer, H Wagner, G B Lipford Immunol Lett. 2000 Jan 10;71(1):55-9. doi: 10.1016/s0165-2478(99)00170-4.
Human papillomaviruses (HPV) have been implicated in the etiology of cervical malignancies and a high percentage of cervical carcinoma cells express HPV-16 E6 and E7 oncoproteins. These proteins are attractive targets for cytolytic T lymphocyte (CTL) mediated immunotherapy. We screened peptides derived from the HPV-16 E7 protein for binding to HLA-A2 and tested their potential to induce specific CTL responses in chimeric HLA-A2/H2-Kb transgenic mice. From eight potential binding peptides four displayed binding and were tested for immunogenicity. CTL activity was tested using target cells pulsed with peptide or expressing E7 protein. While there was no CTL induction observed with the peptides 7-15, 66-74 and 82-90, CTL from mice immunized with 86-93 lysed targets presenting the peptide in the context of the HLA-A2/H2-Kb molecule or wild-type HLA-A2. In contrast, 86-93 induced CTL showed no cytolytic activity against cells expressing the protein E7 and vaccination with the E7 protein did not lead to cytotoxicity against targets pulsed with the 86-93 peptide. Therefore the peptide 86-93, which binds to HLA-A2, is able to induce CTL responses in context of HLA-A2, but the peptide appears not to be processed or presented by HPV type 16 infected cells.
3. Induction of human papillomavirus type 16-specific immunologic responses in a normal and an human papillomavirus-infected populations
Wen-Fang Cheng, Chien-Nan Lee, Yi-Ning Su, Ming-Cheng Chang, Wen-Chun Hsiao, Chi-An Chen, Chang-Yao Hsieh Immunology. 2005 May;115(1):136-49. doi: 10.1111/j.1365-2567.2005.02126.x.
Human papillomavirus (HPV) infection, especially with the oncogenic genotypes, is the most important risk factor for developing cervical cancer. We focused on generating HPV16 E7-specific cytotoxic CD8(+) T lymphocytes and evaluating HPV16 E7-specific immune responses in HPV16-infected and uninfected populations. Peripheral blood mononuclear cells (PBMCs) were first collected from an uninfected group with an human lymphocyte antigen (HLA) A2 haplotype (four volunteers). Mature monocyte-derived dendritic cells (DCs) were generated from the PBMCs and pulsed with one of two HLA-A2-restricted E7 peptides, aa 11-20 [YMLDLQPETT] and aa 86-93 [TLGIVCPI], as antigen presenting cells. The autologous naive or cultured PBMCs were then cultured with peptide-pulsed DCs to detect the HPV16 E7-specific immune responses by a variety of techniques such as enzyme-linked immunosorbent assay (ELISA), enzyme-linked immunospot (ELISPOT) assay and cytotoxic T lymphocyte assay. Interferon-gamma (IFN-gamma) from E7-specific cytotoxic CD8(+) T lymphocytes stimulated with the respective peptide was detected by ELISA. Using ELISPOT analysis, a marked increase in the number of IFN-gamma-secreting CD8(+) E7-specific lymphocytes was observed following peptide stimulation. Cultured CD8(+) T lymphocytes were highly cytotoxic against the CaSki cells. PBMCs were then collected from an HPV16-infected population of the HLA-A2 haplotype, including four persons of HPV16 infection only, four with cervical intraepithelial neoplasia (CIN) lesions, and four cervical cancer patients. We then compared the immunologic responses to E7 between HPV16-infected and uninfected populations by ELISA and ELISPOT assay. The E7-specific immunologic responses of the HPV16-infected populations were significantly higher than those of the uninfected population. In addition, persons with an HPV16 infection only or those with CIN lesions generated higher E7-specific immunologic responses than cervical cancer patients. Our results demonstrate methods for evaluating E7-specific immunologic responses and reflect the biological responses of HPV16-infected people during different periods of cervical disease.
