Faithful Modeling of Terminal CD8 T Cell Dysfunction and Epigenetic Stabilization In Vitro [ATAC-seq]

Epigenetic scarring of terminally dysfunctional CD8 T cells hinders long-term protection and response to immune checkpoint blockade during chronic infections and cancer. We developed a faithful in vitro model for CD8 T cell terminal dysfunction as a platform to advance T cell immunotherapy. Using TCR-transgenic CD8 T cells, we found that 1-week peptide stimulation, mimicking conditions in previous models, failed to induce a stable exhaustion program. In contrast, prolonged stimulation for 2-3 weeks induced T cell dysfunction but triggered activation-induced cell death, precluding long-term investigation of exhaustion programs. To better mimic in vivo exhaustion, we provided post-effector, chronic TGFβ1 signals, enabling survival of chronically stimulated CD8 T cells for over 3 weeks. These conditions induced a stable state of terminal dysfunction (TDysf), marked by a stable loss of effector, cytotoxicity, and memory programs, along with mitochondrial stress and impaired protein translation. Importantly, transcriptomic and epigenetic analyses confirmed the development of terminal exhaustion-specific signatures in TDysf cells. Adoptive transfer of TDysf cells revealed their inability to recall effector functions or proliferate after acute LCMV rechallenge. This novel tractable model system enables investigation of molecular pathways driving T cell terminal dysfunction and discovery of new therapeutic targets for cancer or chronic infections.