Authors

Discussion

The accumulation of functionally exhausted T cells in the microenvironment of developing tumours is key to the inability of the immune system to generate an efficient cytotoxic response to cancer cells. Exhausted T cells are characterised by high levels of inhibitory receptor expression, reduced inflammatory cytokine production and low proliferation rates. A range of transcriptional and metabolic alterations have also been reported.

In recent years, antibody-based therapies and small molecules that target inhibitory checkpoint receptors have been developed that can reverse T cell exhaustion. In vitro models that recapitulate this complex phenomenon can expedite the development of new therapeutic approaches in this area.

Here, we show that repeated in vitro stimulation of purified human T cells from multiple independent donors with CD3/CD28 Dynabeads^tm leads to the progressive and sustained expression of PD-1, Lag-3, and Tim-3, three key checkpoint markers of T cell exhaustion. We further show that T cells exhausted in vitro show reduced production of pro-inflammatory IL-2 in response to T cell activation. Preliminary data also indicate that treatment with commercially available checkpoint inhibitors reverses the exhaustion phenotype. Using this setup, we show the ability to reproducibly generate large numbers of exhausted T cells for downstream phenotypic and functional assays, including mixed-lymphocyte reactions. This constitutes a key-step towards the development of large-scale screening cascades for the identification of novel anti-tumoural therapeutic agents.