LAUSR

Noncanonical NF-κB signaling enhances the survival of CAR T cells expressing the 4-1BB costimulatory domain. Ensuring the persistence of CAR T cells Optimal antitumor responses in patients treated with chimeric antigen receptor (CAR) T cells correlates with the persistence of these cells after treatment. Confirming patient data, Philipson et al. found that 4-1BB–expressing CAR T cells survived longer and increased in number to a greater extent ex vivo compared to CD28-expressing CAR T cells. The 4-1BB–expressing CAR T cells exhibited enhanced noncanonical NF-κB signaling and decreased expression of apoptotic factors compared to their CD28-expressing counterparts. These findings suggest that manipulation of the noncanonical NF-κB pathway in future CAR T cell designs may provide clinical benefit. Clinical response to chimeric antigen receptor (CAR) T cell therapy is correlated with CAR T cell persistence, especially for CAR T cells that target CD19+ hematologic malignancies. 4-1BB–costimulated CAR (BBζ) T cells exhibit longer persistence after adoptive transfer than do CD28-costimulated CAR (28ζ) T cells. 4-1BB signaling improves T cell persistence even in the context of 28ζ CAR activation, which indicates distinct prosurvival signals mediated by the 4-1BB cytoplasmic domain. To specifically study signal transduction by CARs, we developed a cell-free, ligand-based activation and ex vivo culture system for CD19-specific CAR T cells. We observed greater ex vivo survival and subsequent expansion of BBζ CAR T cells when compared to 28ζ CAR T cells. We showed that only BBζ CARs activated noncanonical nuclear factor κB (ncNF-κB) signaling in T cells basally and that the anti-CD19 BBζ CAR further enhanced ncNF-κB signaling after ligand engagement. Reducing ncNF-κB signaling reduced the expansion and survival of anti-CD19 BBζ T cells and was associated with a substantial increase in the abundance of the most pro-apoptotic isoforms of Bim. Although our findings do not exclude the importance of other signaling differences between BBζ and 28ζ CARs, they demonstrate the necessary and nonredundant role of ncNF-κB signaling in promoting the survival of BBζ CAR T cells, which likely underlies the engraftment persistence observed with this CAR design.