Abstract 4100: Co-stimulatory signaling boosts CAR T cell efficacy against chronic lymphocytic leukemia
Authors: Zhao Ziran , Collins McKensie , Sun Clare , Wiestner Adrian , Melenhorst J. Joseph 
Published: April 2023

Abstract
Chimeric antigen receptor (CAR) T cell therapy has been promising in chronic lymphocytic leukemia (CLL) treatment showing durable remission. However, only a third of CLL patients can achieve complete remission with persistent effect featured by early memory T cells. This disparity is attributed to T cell-intrinsic defects or tumor-mediated immunosuppression. The mechanism how CLL cells impact CAR T cell potency still remains poorly understood. Here we used second-generation CD19- and ROR1-directed CAR T cells with a 4-1BB intracellular signaling domain. To recapitulate CLL-derived T cell defects, we developed an in vitro model of chronic antigen stimulation using CAR T cells recursively exposed to primary CLL cells. Upon each round of CLL stimulation, both types of CAR T cells were lack of proliferation and differentiation, accompanied with cytokine production failure in IL-2, suggesting hypofunction of CAR T cell induced by CLL. We next found that this lack of CAR T cell response is not permanent but can be rescued by strong antigen stimulus or IL-2 administration. On the other hand, IL-2 affects phenotype of CLL cells by enhancing expression of co-stimulatory molecules CD80 and CD86, suggesting that IL-2 supplementation or auxiliary co-stimulation via CD80/CD86 can rescue CAR T cell reactivity against these tumor cells. To test this hypothesis, we employed the immunomodulatory drugs (IMiDs) lenalidomide and pomalidomide that have previously been demonstrated to upregulate co-stimulatory molecules on CLL and restore T-cell function via IL-2 upregulation. We sought to test if IMiDs can improve CAR T cell response to CLL. CAR T cells repeatedly exposed to primary CLL cells were co-treated with IMiDs at each round, showing significantly increased proliferation compared to the group without drug treatment. Furthermore, IMiDs can enhance the functions of these CLL stimulated CAR T cells, including differentiation (higher frequency of effector memory T cells), activation (HLA-DR, CTLA-4), proliferation (Ki67), and effector function (Granzyme B). In addition, IMiDs can increase cytotoxic efficacy of CD8+ CAR19 T cells on CLL cells. Overall, our study unveiled that in CLL, activation defect of 4-1BB, CD3-signaling anti-CD19 CAR T cells was attributable to low levels of co-stimulatory molecules on CLL cells and can be effectively overcome via upregulation of co-stimulation. Our data suggested that CLL resistance to CAR T cell can be successfully rescued by concurrent treatment with IMiDs. This study provides a potential translational approach to overcome the bottleneck of CLL treatment by CAR T cells.
Citation Format: Ziran Zhao, McKensie Collins, Clare Sun, Adrian Wiestner, J. Joseph Melenhorst. Co-stimulatory signaling boosts CAR T cell efficacy against chronic lymphocytic leukemia. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4100.