Mechanisms of Suppression Used by Regulatory T Cells in Patients Newly Diagnosed with Acute Myeloid Leukemia

An elevated frequency of CD4+CD25^high regulatory T cells (Treg) has been reported in the peripheral blood in patients with various solid tumors and hematologic malignancies. Although the increase in Treg seems to be a characteristic feature of most tumors, the functional role of Treg and the mechanisms of suppression, especially in patients with hematologic malignancies, have been less well defined. We investigated Treg-mediated suppression and the responsible mechanisms in thirty newly diagnosed acute myeloid leukemia (AML) patients prior to any treatment and twenty five healthy donors (NC). The percentage of circulating CD4+ CD25^high Treg was higher (p <0.0001) in the AML patients (4.5 ±0.2%, range 1.7–8.2%) compared to NC (1.5 ± 0.08%, range 0.9–3.1 %). To evaluate the suppressive function, CD4+CD25^high T cells (S) were co-cultured with sorted, CFSE-labeled autologous CD4⁺CD25^high T cells (R) at different S/R ratios. Suppression mediated by Treg co-incubated with proliferating autologous responders was significantly higher (p<0.001) in AML than that mediated by control Treg. To evaluate the role of cytokines produced by Treg in suppression and a need for cell-to- cell contact, transwell analysis of S/R co-cultures was performed. Co-incubation in the presence of transwell inserts (TRI) resulted in significant reduction of suppression (p<0.05), and the addition of neutralizing antibodies to IL-10 and TGF-β1 in the presence of TRI further decreased suppression mediated by Treg. These data suggest that both immunoinhibitory cytokine production and cell-to-cell contact are necessary for suppression. To explore other potential mechanisms of Treg suppression, we evaluated the expression by Treg of ectonucleotidases CD39 and CD73 and the capability of Treg to produce adenosine. CD4+CD25^high T cells of AML patients had higher expression (p<0.01) of CD39 and more efficiently hydrolyzed ATP to adenosine relative to Treg in NC. These data indicate that various mechanisms of suppression may be utilized by Treg in patients with AML. The increase frequency of Treg mediating potent suppression by various mechanisms is likely to play a role in host anti-tumor immune responses. Therefore, modulation of the frequency and functions of Treg might provide new immunotherapeutic approaches in AML.