Combination of Cy and AdIL-12 enhances in vivo antitumor immune response by reversion of Tregs and MDSC-driven immunosuppression

MALVICINI M; INGOLOTTI M; PICCIONI F; GARCIA MG; BAYO J; ATORRASAGASTI C; ALANIZ L; SCHAROVSKY G; MATAR P; MAZZOLINI G

Buenos Aires

Congreso; 1 Congreso frances-argentino de Inmunologia; 2010

Background: We have previously demonstrated the synergistic antitumor effect of the sequential administration of low-dose cyclophosphamide (Cy) followed by sub-therapeutic doses of adenovirus expressing IL-12 genes (AdIL-12) in a mouse colorectal carcinoma (CRC) model (CT26). Our results suggested that the combined treatment allowed the generation of strong antitumor immune responses. The aim of this work was to investigate the immunological mechanisms responsible for the therapeutic effect of the combination. Methods: Balb/c mice were s.c injected with CT26 cells (day 0), distributed in experimental groups (day 7) and treated with: saline; Cy 50 mg/Kg i.p (day 7); AdIL-12 109 TCID50 i.t (day 8) or Cy + AdIL-12. Tumor volume was measured and samples of peripheral blood, spleen and tumor were taken. CD4+CD25+ and CD4+CD25- T lymphocytes were isolated by magnetic separation. Results: We observed that mice non-responder to the combined treatment showed a proportion of regulatory T cells (Tregs) higher than in responder mice, both in peripheral blood and spleen, as well as in tumor samples (p<0.05). Also, the antitumor effect of combined therapy was reverted by in vivo administration of Tregs. The combination Cy+AdIL-12 inhibited Tregs ability to secrete IL-10 and TGF-beta and their capacity to inhibit dendritic cells maturation. We also observed that a subpopulation of immature cells with myeloid and suppressive phenotype (MDSCs) was recruited in the spleen of mice during tumor progression. Cy + AdIL-12 induced a significant decrease in the percentage of MDSCs with respect to untreated mice (4,5 % vs. 16 %; p<0,05). Importantly, depletion of Tregs and MDSCs by combined therapy lead to development of specific IFN-ã-secreting CD4+ T-lymphocytes response, able to eradicate CRC tumors followed to their adoptive transfer. Conclusion: Our results suggest that Cy + AdIL-12 induces a synergistic antitumor immune response against CRC by reversion of Tregs and MDSCs number and function.