Increase of CD4⁺CD25⁺ regulatory T cells in the peripheral blood of patients with metastatic carcinoma: A Phase I clinical trial using cyclophosphamide and immunotherapy to eliminate CD4⁺CD25 ⁺ T lymphocytes

We determined the number and functional status of CD4⁺CD25 ^high regulatory T cells (T_reg) in blood samples from patients with metastatic carcinoma, and evaluated their sensitivity to a single intravenous infusion of cyclophosphamide. T_reg numbers were significantly higher in 49 patients with metastatic cancer (9.2% of CD4 ⁺ T cells) compared to 24 healthy donors (7.1%). These cells expressed the transcription factor forkhead box P3 (FoxP3), glucocorticoid- induced tumour necrosis factor receptor family-related protein (GITR) and intracellular CD152, and demonstrated a suppressive activity in vitro against CD4⁺CD25^- autologous proliferation. At a single intravenous infusion, cyclophosphamide failed, in association with a non-specific immunotherapy by intratumoral bacille Calmette-Guérin (BCG), to modulate significantly T_reg numbers or function. Metastatic cancer is associated with an expansion of peripheral blood CD4 ⁺CD25^highFoxP3⁺GITR⁺CD152 ⁺T_reg cells whose immunosuppressive properties do not differ from those of healthy subjects. Moreover, cyclophosphamide administration may not represent an optimal therapy to eliminate T_reg, which further underlines the need to identify specific agents that would selectively deplete these cells.