Activated tumor-infiltrating CD4+ regulatory T cells restrain antitumor immunity in patients with primary or metastatic liver cancer

Abstract

The mechanisms that enable liver cancer to escape elimination by the immune system remain unclear, but their elucidation may provide novel therapeutic interventions. We investigated the influence of tumor-infiltrating regulatory T cells on tumor-specific T cell responses in patients with liver cancer, using ex vivo isolated cells from individuals with hepatocellular carcinoma (HCC) or liver metastases from colorectal cancer (LM-CRC). Here we report that in both HCC and LM-CRC, CD41CD251Foxp31 regulatory T cells (Tregs) accumulate in the tumor milieu and are potent suppressors of autologous tumorspecific T cell responses. Especially in LM-CRC, where Treg accumulation is more prominent, there is good evidence for local proliferation of Tregs at the cancer site. We show that tumor Tregs up-regulate the expression of glucocorticoid-induced tumor necrosis factor receptor (GITR) compared with Tregs in tumor-free liver tissue and blood. Importantly, treatment with soluble GITR ligand (GITRL) induces a decrease in the suppression mediated by the activated tumor-infiltrating Tregs and restores the proliferative capacity and cytokine production of CD41CD252 T cells. Conclusion: Our results show that tumor-associated Tregs are critical for immune evasion in liver cancer, and we propose that GITRL constitutes a rational treatment for this disease. (HEPATOLOGY 2013;57:183-194) T he two most common types of cancer affecting the liver are hepatocellular carcinoma (HCC) and liver metastases from colorectal cancer (LM-CRC). 1,2 The current therapeutic options for both malignancies are limited to liver surgery and local (ablative) therapy. At the time of diagnosis, the majority of HCC patients are not candidates for curative treatment, and in patients with LM-CRC there is a high rate of recurrence after treatment. 1,2 Consequently, there is a pressing need for novel therapeutic strategies. Immunotherapy is attractive because of the exquisite specificity of the immune response and may thus avoid many of the side effects associated with currently available clinical options. 3 However, immunological tolerance of the liver or immunoregulatory mechanisms present in the tumor microenvironment, such as those described by us and others in breast, 4-6 ovarian, 7 and renal 8 cancer, may contribute to tumor outgrowth and limit immunotherapeutic strategies by suppressing the local antitumor response.There is accumulating evidence that CD4þFoxP3þ regulatory T cells (Tregs) hamper the development of effective tumor immunity in individuals with cancer. 9,10

Blockade of LAG3 enhances responses of tumor-infiltrating T cells in mismatch repair-proficient liver metastases of colorectal cancer

“…As a consequence, tumor-infiltrating Treg, 40 , 41 type 1 regulatory T cells 52 and probably other types of immune suppressor cells were also present in these assays. Therefore, the reported functional effects of checkpoint inhibitors on effector T cells in these assays may be partly indirect, mediated by effects of these antibodies on suppressor cells.…”

“…35 Moreover, the unique immune environment in the liver 36 favors immunological tolerance, 36 and one of the mechanisms used by the liver to resist immune responses is the induction of expression of inhibitory receptors on hepatic T cells 37 , 38 and their ligands on hepatocytes and other liver tissue cells. 39 Previously we have observed that intra-tumoral CD8 + cytotoxic T cells (CTL) and CD4 + T helper cells (Th) are functionally compromised in LM-CRC, 40 and we also demonstrated that the suppression mediated by intra-tumoral regulatory T cells (Treg) in LM-CRC can be alleviated by blocking the inhibitory receptor CTLA4 and activating the stimulatory receptor GITR. 41 However, the expression and functional roles of inhibitory receptor-ligand pathways in regulating tumor-infiltrating effector T cell responses have not been studied yet in LM-CRC.…”

“…40 , 41 Myeloid dendritic cells (mDC) were isolated from patient autologous PBMC by depletion of CD19 + B cells followed by positive selection for BDCA-1 (BDCA-1 DC isolation kit, Miltenyi Biotec). mDC were cultured overnight with or without 20 µg/mL autologous tumor lysates or normal liver lysates, in the presence of 10 ng/mL granulocyte-macrophage colony-stimulating factor (Miltenyi Biotec) and 0.5 µg/mL polyinosinic: polycytidylic acid (InvivoGen, San Diego, CA).…”

GITR engagement in combination with CTLA-4 blockade completely abrogates immunosuppression mediated by human liver tumor-derived regulatory T cellsex vivo

“…10,11 In support of this we and others have described the presence of regulatory CD4 C Foxp3 C T cells in liver tumors, that suppress local antitumor immunity and that are associated with poor patient prognosis. [12][13][14][15][16] Liver TiTreg are characterized by high expression of glucocorticoidinduced tumor necrosis factor receptor related gene (GITR), CTLA-4 and inducible T cell co-stimulator (ICOS). 12 These molecules are important for the immunosuppressive function of Treg and can be targets for immunotherapeutic interventions.…”

“…[12][13][14][15][16] Liver TiTreg are characterized by high expression of glucocorticoidinduced tumor necrosis factor receptor related gene (GITR), CTLA-4 and inducible T cell co-stimulator (ICOS). 12 These molecules are important for the immunosuppressive function of Treg and can be targets for immunotherapeutic interventions. Abrogation of Ti-Treg function may allow the induction of antitumor immunity at the tumor site and improve the outcome of immunotherapy aimed to activate antitumor responses.…”