The Effects Of Active Immunization Against IL-4 And IL-13 On Grafted TC-1 Tumor Growth In Mouse And Efficacy Of An Experimental Therapeutic HPV Vaccine

Background Th2 cell and its cytokines play important roles in the process of cancer development. IL- 4 and IL - 13 can stimulate macrophage to differentiate into M2 type and express inhibitory factors including IL - 10, TGF - beta 1, and arginase 1, as well as tumor angiogenesis factors, such as EGF, VEGF etc; in addition, the M2 can express chemokines CCL17, CCL22 and CCL24 which promote infiltration of Tregs and MDSCs into tumor tissues, and then induce T cell incompetent and inhibit T cell-mediated cytotoxic effects. Therefore, targeting IL-4 and IL-13 holds the potentials of decreasing the responses of Th2, Treg, MDSCs and tumor associated macrophages, and enhancing anti-tumor CTLs/Thl responses.Objective To reveal the effects, of anti - IL-4 and IL-13 active immune against Th2 responses in tumor microenvironment, on tumor growth and efficacy of an experimental tumor antigen - specific therapeutic HPV vaccine, and also explore the immune mechanisms, from which to provide potential immunotherapeutic approaches and strategies.Methods By using gene recombinant techniques, HBcAg virus-like particles(VLPs) vaccines presenting IL - 4 or IL - 13 antigen epitope were constructed. A HPV infection-induced tumor model was established using TC-1 cells to graft into C57 mice. In one of the studies with the tumor model, a preventive strategy of vaccination was employed to investigate the effects of anti-IL-4 and IL-13 vaccine immunization on the growth of grafted TC - 1 tumor and involved immune responses. And, in another study, a treatment strategy of vaccine immunization was used. In this study,anti-IL-4 and IL-13 vaccine and/or therapeutic HPV vaccine carring a HPV16 E7 epitope were immunized after the tumor has been established. The purpose was to test whether anti-IL-4 and IL-13 immunzation are still suppress tumor growth and have a synerstic effect with the therapeutic HPV vaccine. For this intervention study, C57 mice were inoculated with TC-1 cells first. When the diameter of tumor mass reach to 1-2mm,anti-IL-4 and IL-13 vaccines were immunized. When the tumors grow to a size of 5-6mm, the therapeutic vaccine was immunized. The methods involved in the studies with the tumor model include: tumor size was measured every two or three days; at endpoint of the experiment with a model, flow cytometry was used to analyze the response level and function of immune cells(including Thl?Th2?Tregs and CTLs), ELISPOT was employed to analyze the antigen specific IFN - ? or IL - 4 -expressing splenocyes, and ELISA was used to detect the levels of IFN - gamma IFN-? , IL - 4 and IL-13 in serum and splenocyte culture supernatant.Results The recombinant proteins were expressed efficiently in E.coli. The purified proteins mainly exist in the form of VLPs. Anti- IL-4? IL-13 VLPs vaccine immunize mice and stimulate high levels of specific antibody response. In the study with preventive immunization, anti- IL-4 and IL-13 vaccine significantly inhibited tumor growth in mice; ELISPOT results showed that vaccine immunization significantly promoted the level of HPV16 E7 stimulated IFN - ? expressing splenocytes, while it suppressed the level of IL-4 -expressing cell; in immunized mice, the level of IFN - ?was increased while that of IL - 4 and IL - 13 was reduced in both serum and culture supernantant of isolated splenocytes;further, flow cytometry showed that vaccine significantly inhibited the reponses of Th2 and Tregs and enhanced the responses of Thl and CTLs. In the treatment study, anti-IL-4 and IL-13 immunization still showed significant inhibition of the tumor growth, increased IFN - ? level in serum, and promoted Thl/CTLs responses and inhibited Tregs response. While anti- IL-4 and IL-13 immunization was combined with the therapeutic vaccine immunization, significant synerstic effects were found, including tumor inhibition, enhancement of CTLs/Thl,and suppression of Treg cells.Conclusions Targeting IL - 4 and IL - 13 by active immune with a VLPs vaccine can induce high level of and persistent specific antibody response, significantly inhibiting tumor growth in mice, and has synerstic effects on the antitumor function of a tumor antigen - specific therapeutic vaccine. The current study indicates that anti- IL-4 and IL-13 active immunization is a potential tumor immune intervention strategy, and also may provide possible effective intervention approaches.