Human CD 4 + CD 39 + regulatory T cells produce adenosine upon co‐expression of surface CD 73 or contact with CD 73 + exosomes or CD 73 + cells

Summary While murine CD 4 + CD 39 + regulatory T cells (T reg ) co‐express CD 73 and hydrolyze exogenous (e) adenosine triphosphate ( ATP ) to immunosuppressive adenosine ( ADO ), surface co‐expression of CD 73 on human circulating CD 4 + CD 39 + T reg is rare. Therefore, the ability of human T reg to produce and utilize ADO for suppression remains unclear. Using mass spectrometry, we measured nucleoside production by subsets of human CD 4 + CD 39 + and CD 4 + CD 39(–) CD 73 + T cells or CD 19 + B cells isolated from blood of 30 volunteers and 14 cancer patients. CD 39 and CD 73 expression was evaluated by flow cytometry, Western blots, confocal microscopy or reverse transcription–polymerase chain reaction ( RT–PCR ). Circulating CD 4 + CD 39 + T reg which hydrolyzed e ATP to 5′‐ AMP contained few intracytoplasmic granules and had low CD 73 m RNA levels. Only ∼1% of these T reg were CD 39 + CD 73 + . In contrast, CD 4 + CD 39 neg CD 73 + T cells contained numerous CD 73 + granules in the cytoplasm and strongly expressed surface CD 73. In vitro‐ generated T reg ( T r1) and most B cells were CD 39 + CD 73 + . All these CD 73 + T cell subsets and B cells hydrolyzed 5′‐ AMP to ADO . Exosomes isolated from plasma of normal control ( NC ) or cancer patients carried enzymatically active CD 39 and CD 73 + and, when supplied with e ATP , hydrolyzed it to ADO . Only CD 4 + CD 39 + T reg co‐incubated with CD 4 + CD 73 + T cells, B cells or CD 39 + CD 73 + exosomes produced ADO . Thus, contact with membrane‐tethered CD 73 was sufficient for ADO production by CD 4 + CD 39 + T reg . In microenvironments containing CD 4 + CD 73 + T cells, B cells or CD 39 + CD 73 + exosomes, CD 73 is readily available to CD 4 + CD 39 + CD 73 neg T reg for the production of immunosuppressive ADO .