Investigating tissue-resident memory T cell differentiation of CAR T cells reveals an unexpected role for CD69

Abstract

Abstract Adoptive cell therapy using chimeric antigen receptor-expressing (CAR) CD8+ T cells has revolutionized the treatment of liquid tumors; however, CAR T cells have had more limited success against solid tumors. CD8+ tissue-resident memory T cells (Trm), which are characterized by surface expression of CD69 and CD103 and a unique transcription factor circuit, are found within solid tumors and associated with improved tumor control. However, it is unclear whether CAR T cells adopt Trm features in the tumor microenvironment and how enhancing Trm differentiation of CAR T cells might impact their efficacy against solid tumors. Using the murine tumor model MC38, we identified both CD69+CD103- and CD69+CD103+ subsets that express Trm signature genes. To examine if CAR T cells follow a similar differentiation pattern, human carcinoembryonic antigen (hCEA)-specific CAR T cells with either a CD28 or 4-1BB costimulatory domain were transferred into hosts bearing MC38-hCEA tumors. We found the CD28 domain drove expansion of the CAR T population while delaying Trm marker expression compared to 4-1BB and non-CAR T cells. The persistence of a CD69-CD103-subset of CD28 CAR T cells strongly correlated with the number of hCEA-CD28 CAR T cells within the tumor. CD69-CD103- hCEA-CD28 CAR T cells were more proliferative and expressed lower levels of surface markers associated with dysfunction, including PD-1 and LAG3. We hypothesized CD69 could negatively regulate T cell proliferation and function; indeed, deletion of CD69 in hCEA-CD28 CAR T cells enhanced their proliferation and accumulation within tumors. In summary, hCEA-CD28 CAR signaling delayed Trm differentiation and CD69 expression in solid tumors resulting in prolonged T cell proliferation. NIH 7R01AI153096