A Recombinant Protein Vaccine Based On Xenogeneic Endoglin Mediates Anti-Tumor Effects In Mouse Models

Angiogenesis facilitates tumor perfusion and play a critical role in tumor growth and metastasis. Increasing evidence suggests that endoglin is a powerful marker of angiogenesis in malignancies. Thus, breaking of immune tolerance of self-endoglin-associated angiogenesis is an attractive approach to cancer therapy by active immunization. In the first section of this study, we used endoglin as a model antigen to explore the feasibility of the active immunotherapy with a vaccine based on a xenogeneic protein, targeting endoglin associated tumor angiogenesis. To test this concept, we recombined the extracellular domain of porcine endoglin, and used it as the xenogeneic vaccine. At the same time, the extracellular domain of the murine endoglin was also recombined and used as the homologous control. The vaccines were used to test for the capability of inducing anti-tumor immunity in tumor models in mice. In this section of experiment, we found that active immunotherapy with porcine endoglin was effective at both protective and therapeutic anti-tumor immunization in three mouse tumor models. Autoantibodies against murine and porcine endoglin were identified by Western blot and ELISA assay. IgG1 and IgG2b were substantially increased in response to porcine endoglin vaccination. Anti-endoglin antibody-producing B cells were detectable by ELISPOT assay. There was endothelial deposition of self-immunoglobulins within tumors. The anti-tumor activity was also induced by the adoptive transfer of the purified immunoglobulins. The anti-tumor activity and production of autoantibodies against mouse endoglin could be abrogated by depletion of CD4+ T lymphocytes. Angiogenesis was apparently inhibited within the tumor tissues and on the alginate beads. Remarkably, no marked toxicity was found in the immunized mice.Although we have found that anti-angiogenic therapy with xenogeneic endoglin as a protein vaccine has shown effective in inhibiting tumor growth in three murine tumor models in the first section of this study, it still remains uncertain whether endoglin inducing anti-angiogenic therapy is tumoricidal. Recently, many studies have concluded that this therapeutic limitation may be overcome by using combination of angiogenic inhibitors with cytotoxic drugs, even when using them in a dose far below maximum tolerated dose (MTD) . So in the second section of this study, we explored the efficacy of a strategy combining low-dose cisplatin (0.3 mg/kg, once per three days) and the xenogeneic endoglin as a protein vaccine. We found that both low dosage cispaltin and xenogeneic endoglin vaccine individually resulted in effective suppression of tumor growth in two tumor models via the inhibition of tumor angiogenesis. Remarkably, the combination therapy resulted in not only significant anti-angiogenic effects but also additional promotion of tumor cell apoptosis and inhibition of tumor cell proliferation in the treated tumor tissues, without any ensuing increase in host toxicity during the course of treatment, which lasted for six months. In addition, thecombination of the two demonstrated a synergistic relationship, which was shown in all of the synergistic indexes calculated using values assayed from tumor volume, angiogenesis, apoptosis and proliferation, respectively. Both antibodies and antibody-producing B cells against mouse self-endoglin were observed in all mice immunized by the xengogeneic endoglin vaccine (alone and combination), which suggested that low-dose cisplatin did not suppress the host immune response, but potentiates the anti-tumor activity of the xenogeneic endoglin vaccine.The above observations may provide the basis for effective alternative strateges for cancer therapy in the near future.