Immune Intervention Of TC-1 Graft Tumor In Mice With Chimeric Virus-like Particles Presenting HPV16 CTLs Epitopes

Background Existing therapeutic HPV vaccines lack significant curative effects in human clinical trials of treatment for cervical cancer or cervical intraepithelial neoplasia.The important strategy of improving the vaccine effect is to enhance tumor antigen specific killing effects of cellular immune response. Virus-like particles (VLPs) has proved to be highly immunogenic in stimulating humoral immunity and is an ideal vaccine vector. Objective This study sought to explore the VLPs of hepatitis B core antigen (HBcAg) to be a potential vaccine carrier for eliciting specific cellular immune responses, and also provide a valuable candidate of therapeutic HPV vaccine, through presenting HPV CTLs antigen peptides on the surface of VLPs and then assessing elicited immune responses and intervention effects on tumor growth in a graft tumor model. Methods Four reported effective HPV16 E6 and E7 CTLs epitopes were selected and inserted into the immune dominant domain of HBcAg by genetic recombination method. The recombinant plasmids were tansformed into E.coli DHsa cells, and the expression of the recombinant proteins were induced by IPTG. The proteins were purified with a procedure consists of ammonium sulfate precipitation and sucrose density gradient centrifugation, and the presence of VLPs was detected with HPLC and electron microscopy. Mice were immunized with the VLPs through prevention and therapeutic strategies separately, and the immune effects of the VLPs on grafted TC-1 tumor were assessed. The dynamic changes of tumor size were examined. In addition, the splenocytes isolated from the experimental mice were stimulated in vitro with the synthetic antigenic peptides and the immune responses were analyzed.The expression of IFN-y was detected by ELISA, the level of IFN-y secreting lymphocyte was measured by ELISPOT, and the number of CD8+IFN-y+ cells were analyzed by flow cytometry. Moreover, The time course of the immune memory response stimulated by VLPs vaccine was investigated, and the influences of pre-existing anti-HBcAg antibody, the structure of VLPs or not and immunization route on VLPs-induced specific cellular immune responses were also assessed.Results The recombinant proteins were expressed efficiently in E.coli,and the purified protein exists mainly in the form of VLPs. Mice immunized with VLPs presenting E7(49-57)epitope showed completely suppressed growth of grafted TC-1 tumor using a preventive vaccination strategy.In a study with a therapeutic vaccination strategy, VLPs significantly suppressed the growth of tumor with 2-3mm size, and also showed obvious inhibition of larger tumor with the size of 5-6 mm or even 8-9mm. In addition, the effective immune memory responses produced by VLPs immunization maintained for at least 15 weeks, evidenced by significantly suppressed tumor growth and elevated IFN-yexpression of immune cells.The pre-existing anti-carrier antibodies didn’t produce any obvious affects on the immune reponses and protective effects of VLPs vaccine.In the immunized mice, the IFN-γ expression of splenocytes, the level of IFN-γ secreting lymphocyte, and CD8+IFN-γ+ cells were displayed to be significantly increased. Conclusion VLPs vaccine presenting a HPV E7 CTLs epitope could elicit a sustained and effective E7 specific cellular immune response and significantly inhibited tumor growth. The results indicated that the VLPs vaccine can be used as a candidate of HPV therapeutic vaccine, and HBcAg VLPs is a potential therapeutic carrier.