In vitro induction of T cells that are resistant to A2 adenosine receptor-mediated immunosuppression

The increased levels of extracellular adenosine in inflamed tissues down-regulate activated immune cells via the A(2A) adenosine receptor. This A(2A) adenosine receptor-mediated immunosuppression is a disqualifying obstacle in cancer immunotherapy as it protects cancerous tissues from adoptively transferred anti-tumour T cells. The aim of this study was to test whether the negative selection of T cells will produce T cells that are resistant to inhibition by extracellular adenosine. Cytotoxic T lymphocytes (CTL) were developed by mixed lymphocyte culture in the presence or absence of the adenosine receptor agonist 5'-N-ethylcarboxamidoadenosine (NECA). The sensitivity of CTL to adenosine analogues was characterized by cAMP induction, interferon-gamma production and cytotoxicity. CTL that could proliferate even in the presence of NECA were less susceptible to inhibition by A(2A) adenosine receptor agonists, as shown by a much smaller accumulation of cAMP and less inhibition of interferon-gamma production compared with control CTL. The successful protocol to produce CTL that are both resistant to adenosine-mediated immunosuppression and maintain strong cytotoxicity and interferon-gamma secretion required NECA to be added only during the expansion stage after the establishment of CTL. In contrast, the priming of resting T cells in the presence of NECA resulted in T cells with impaired effector functions. Adenosine-resistant effector T cells were successfully obtained by exposure of activated T cells to NECA. These in vitro studies form the basis for future attempts to produce anti-tumour T cells that are more effective in adoptive immunotherapy.