T regulatory type 1 cells in squamous cell carcinoma of the head and neck: Mechanisms of suppression and expansion in advanced disease

Purpose: Regulatory T cells play a major role in tumor escape from immunosurveillance. T regulatory cells type 1 (Tr1), a subset of regulatory T cells present in the tumor and peripheral circulation of patients with head and neck squamous cell carcinoma (HNSCC), mediate immune suppression and might contribute to tumor progression. Experimental Design: CD4(+)CD25(-)T cells were isolated from peripheral blood mononuclear cells (PBMC) or tumor-infiltrating lymphocytes (TIL) of 26 HNSCC patients and 10 normal controls. The Tr1 cell phenotype was determined before and after culture in the presence of interleukin (IL)-2, IL-10, and IL-15, each at 10 to 20 IU/mL. Suppression was measured in carboxyfluorescein diacetate succinimidyl ester - based proliferation assays with or without neutralizing anti-IL-10 or anti-transforming growth factor-beta(1) (TGF-beta(1)) monoclonal antibodies in Transwell systems. ELISA was used to define the Tr1 cytokine profile. Results: TO cells originate from CD4(+)CD25(-) precursors present in TIL and PBMC of HNSCC patients. Cytokine-driven ex vivo expansion of TO precursors yielded CD4(+)CD25(-)Foxp3(low)CD 132(+)IL-10(+)TGF-beta(+)(1) populations that mediated higher suppression than TO cells of normal controls (P < 0.0001). Tr1 cells suppressed proliferation of autologous responders via IL-10 and TGF-beta(1) secretion. Expression of these cytokines was higher in TIL-derived than PBMC-derived TO cells (P < 0.0001). The Tr1 cell frequency and suppressor function were significantly higher in patients presenting with advanced than early disease stages and in patients "cured" by oncologic therapies than in those with active disease. Conclusions: In HNSCC, Tr1 cell generation is promoted at the tumor site. Tr1 cells use TGF-beta and IL-10 to mediate suppression. They expand during disease progression and also following cancer therapy in patients with no evident disease.