Concomitant Radiation Therapy and TGFβ Neutralizing Antibodies Alters Tumor Microenvironment and Promotes Tumor Regression

Abstract

Transforming growth factor-β (TGFβ) has a broad range of pro-tumorigenic activities and immunosuppressive functions that have drawn considerable interest in cancer, especially since pharmaceutical inhibitors are in clinical trials. It is not clear how best to release the considerable potential of TGFβ inhibitors. Radiation induces TGFβ activity, which promotes effective response to DNA damage protecting cancer cells from radiation-induced cell death. RT is quite effective in eliciting immunogenic signals but is itself a source of immunosuppressive effects in the tumor microenvironment. Here we demonstrate that inhibition of TGFβ in pre-clinical a breast cancer model creates a tumor microenvironment that promotes immune response. Balb/c mice were inoculated with 105 non-metastatic TSA breast cancer cells into the flank and injected with 10mg/kg i.p. preclinical TGFβ neutralizing antibody 1D11 beginning on day 13 post tumor cell implantation. Twenty-four hours later, mice received 6 Gyx5 consecutive irradiation fractions. Tumor growth was caliper-measured thrice/week. Flow cytometry analysis and ex vivo cultures of primary cells were established to assess frequency and functional characteristics of immune cells. TGFβ neutralizing antibody treatment had no effect on tumor growth, however when combined with RT, tumor growth was substantially decreased relative to radiation treatment alone (p < 0.001). Notably, more than half of mice achieved complete tumor regression, suggesting activation of tumor immunity. TGFβ blockade decreases a splenic reservoir of CD11b+ myeloid-derived suppressor cells that mediate immunosuppression in the tumor microenvironment as well as CD11b+Gr1+/intF4/80+ precursors of tumor-associated macrophages. Immunoactivating IL12 was upregulated upon TGFβ inhibition while immunosuppressive IL10 and pro-angiogenic CXCL1 decreased. The number of splenic CD11c+ MHC Class I+ antigen presenting cells was dramatically reduced from irradiated tumor-bearing mice, but unaffected in mice receiving 1D11 antibody concomitant with RT. Tumor cell MHC Class I overexpression, which is induced by in vitro irradiation, was impaired after coculture with splenocytes from irradiated mice but not from mice receiving the dual treatment or 1D11 alone. TGFβ blockade sensitizes tumor cells to RT and generates a tumor microenvironment that promotes tumor control. TGFβ inhibition also alleviates tumor suppressive consequences of radiation and creates a pro-immunogenic signal. Together, these actions can achieve persistent tumor control.