A study of the immunomodulatory characteristics of FIP-Fve protein and its adjuvant effects in tumor immunotherapy

Abstract

Fve is a 12.7 kDa fungal protein isolated from the Flammulina velutipes mushroom and it has previously been reported to trigger immunological responses in both mouse and human lymphocytes. In the present study, the immunomodulatory effects of Fve on the T cells and dendritic cells (DCs) were investigated. In addition, the potential application as an adjuvant for tumor immunotherapy was explored. In vitro cell culture experiments showed that Fve stimulated full activation of both purified CD4+ and CD8+ T cells to proliferate and secrete high levels of IL-2, IFN-gamma, and IL-6 accompanied by up-regulation of CD69, OX-40 and 4-1BB in the presence of accessory cells such as DCs and B cells. Trans-well studies showed that accessory cell-T cell direct interaction was important for T cell's full activation. Moreover, in vitro experiments showed that Fve failed to drive bone marrow-derived dendritic cells (BM-DC) phenotypic maturation. In contrast, in vivo studies revealed that intraveneously injected Fve could drive splenic DC phenotypic and functional maturation as indicated by the up-regulation of MHC class II (MHC II) molecules and CD86 expression on DCs and the DC's capabilities of priming both the antigen-specific Th1-skewed CD4+ cells and CD8+ T cells. Notably, it was found that Fve-activated T cells could provide accessory help to induce phenotypic maturation of DC in cell contact-dependent manner. Taken together, these data demonstrated that Fve was capable of driving enhanced Th1-skewed polarization and CD8+ T cells activity in antigen-specific manner. In view of this, it was hypothesized that Fve could act as a vaccine adjuvant to enhance the immunogenicity of co-administered antigens. The proof of concept in vitro and in vivo studies were carried out with HPV type 16 E7 protein as a model antigen in tumor animal model induced by the cervical cancer related E7-expressing TC-1 tumor cells.<br>The results revealed that mice co-immunized with HPV-16 E7 and Fve showed increased production of HPV-16 E7-specific antibodies as well as enhanced expansion of HPV-16 E7-specific IFN-g-producing CD4+ and CD8+ T cells as compared to mice immunized with HPV-16 E7 alone. Tumor protection assays showed that 60% as compared to 20% of mice co-immunized with HPV-16 E7 plus Fve or immunized with HPV-16 E7 respectively remained tumor free for up to 167 days after the tumor cells challenge. Tumor therapeutic assays showed that HPV-16 E7 plus Fve treatments significantly prolonged the survival of tumor bearing mice as compared to those treated by HPV-16 E7. In vivo cell depletion and adoptive T cell transfer assays illustrated that CD4+, CD8+ T cells and IFN-gamma played critical roles in conferring the anti-tumor effects. Therefore, I conclude that the pleiotropic immunostimulatory effects of Fve on innate and adaptive immune cells leading to enhanced polarization of antigen specific CD4+ and CD8+ cells can be exploited to develop effective adjuvant for anti-cancer and anti-viral vaccines. <br>