Vδ2 T cell subsets, defined by PD-1 and TIM-3 expression, present varied cytokine responses in acute myeloid leukemia patients

•The levels of PD-1+TIM-3+ Vδ2 T cells were significantly higher in AML patients.•PD-1+TIM-3− subset presented the highest TNF-α and IFN-γ expression.•PD-1+TIM-3+ subset presented the lowest TNF-α and IFN-γ expression.•Anti-TIM-3 inhibition and anti-PD-1/TIM-3 dual inhibition elevated TNF-α and IFN-γ.•Anti-PD-1 blocking antibodies increased the frequency of TIM-3+ cells in Vδ2 T cells. Vδ2 T cells represent the major γδ T cell subset in humans and can serve as an important early source of TNF-α and IFN-γ during inflammatory responses. In acute myeloid leukemia (AML) patients receiving allogeneic stem cell transplantation, higher γδ T cell count predicted better prognosis. The impact of PD-1 and TIM-3 expression on the function of Vδ2 T cells is yet unclear. In this study, we showed that the frequencies of PD-1+TIM-3− Vδ2 T cells were comparable between healthy controls and AML patients, but the frequencies of PD-1−TIM-3+ Vδ2 T cells and of PD-1+TIM-3+ Vδ2 T cells were significantly higher in AML patients than in healthy controls. Both PD-1 and TIM-3 were upregulated upon phosphoantigen + IL-2 activation, but the relative differences in the frequencies of various PD-1 vs. TIM-3 subsets between AML patients and healthy controls remained. Interestingly, among all PD-1 vs. TIM-3 subsets, the PD-1+TIM-3− subset presented the highest TNF-α and IFN-γ expression, while the PD-1+TIM-3+ subset presented the lowest TNF-α and IFN-γ expression. Anti-PD-1 inhibition did not significantly affect the production of TNF-α or IFN-γ, but anti-TIM-3 inhibition and anti-PD-1/TIM-3 dual inhibition significantly elevated the production of TNF-α and IFN-γ. Interestingly, anti-PD-1 blocking antibodies had significantly increased the frequency of TIM-3+ cells in Vδ2 T cells, suggesting a compensatory TIM-3 upregulation. In addition, the levels of PD-L1 and HMGB-1 were significantly higher in AML patients than in healthy subjects. In summary, this study provides knowledge on the cytokine expression patterns by PD-1 and/or TIM-3-expressing Vδ2 T cells in AML patients, and indicates that the upregulation of PD-1 alone is insufficient to indicate functional impairment, and Vδ2 T cells may require anti-TIM-3 inhibition for functional revival.