TH17 Pathway Promotes Tumor Cell Growth and Suppresses Immune Function in Myeloma: Potential for Therapeutic Application

TH17 cells are T helper cells generated from naive CD4 population in the presence of TGF-beta and IL-6 or IL-1-beta; and expanded in presence of IL-23. TH17 cells predominantly produce IL-17 and/or IL-22 and participate in both development of immunity and autoimmunity. Since TH17 cells are involved in modulation of immune response and we haveobserved both B and T cell immune dysfunction in multiple myeloma (MM), we have further evaluated the role of TH17 cells and associated cytokines in tumor cell growth, as well as, the imbalance of immune homeostasis in myeloma. We first observed that the baseline frequency of TH17 cells was significantly higher in mononuclear cells from MM patient peripheral blood (n=11) (2 folds) and bone marrow (n=4) (50%) compared to normal donor samples. Furthermore, under polarizing conditions with cytokine cocktail, significantly higher number of TH17 cells were induced in presence of TGF-beta in mononuclear cells from MM patients in both blood (4 fold) and bone-marrow (9 fold) compared to normal donor samples. These results indicate that the cytokines present in the BM microenvironment favor the development of TH17 cell subset. We next evaluated the serum levels of TH17-associated cytokines in sera from patients with MM, as well as, normal donors. We observed significant increase in levels of IL-17 (2 fold), IL-21 (5 fold), IL-22 (8 fold) and IL-23 (6 fold) in myeloma (n=17) compared with normal donor (n=6). Importantly, IL-23 levels were 10 fold higher in myeloma BM samples compared with matching blood samples. Next, we evaluated the effect of TH17-associated pro-inflammatory cytokines on MM cell growth and survival. We observed induction of proliferation of 6 MM cell lines, as measured by thymidine incorporation, by IL-17 (30%), IL-21 (35%) and IL-22 (25%). Additionally, IL-17 significantly induced growth of MM cells in clonogenic assay. These growth promoting effects are not observed in the presence of anti-IL-17 antibody. In an effort to evaluate the cellular basis for this growth promoting effect of these cytokines, we investigated production of MM growth promoting cytokines in the BM milieu. We observed induction of IL-6 production by IL-17 from BM stromal cells alone and in the presence of MM cells. Production of IL-6 induced by IL-17 was abrogated (40–50%) by transduction of siRNA targeting IRF-4 into co-culture components (stromal and myeloma cells). The inhibition of IL-17-induced IL-6 production in stromal cells was also observed with JAK-2 (70%) and MEK (76%) inhibitors suggesting role of multiple signalling pathways in induction of IL-6. Finally, we have also observed negative effects of TH17- related cytokines, especially IL-17 and IL-22, on development of IFN-gamma producing TH1 cells suggesting immunosuppressive effect. Taken together these results suggest significant role of IL-17 and related cytokines in myeloma cell growth, as well as, immune dysfunction observed in these patients. This data provides the preclinical rationale to target IL-17 in myeloma to suppress growth as well as improve immune responsiveness.