Paper Details

ABSTRACT

Unlike conventional single-chain chimeric antigen receptor (CAR) T cell therapy, multi-chain killer immunoglobulin-like receptor (KIR)-based CAR T incorporates separate antigen binding and activation signals derived from natural killer (NK) cells. Recently, we demonstrated that KIR-CAR T cells have reduced tonic signaling and eliminate off-target activity in solid tumors in vitro. In vivo, KIR-CAR T cells also demonstrated tumor-restricted homing and accumulation a solid tumor model, of particular importance due to severe off-tumor toxicity reported in patients receiving single-chain CD3-based CAR T cells in clinical trials.

We have compared our novel CD19-targeting DS191-KIR-CAR T cells to single-chain CD3-based FMC63-41BBζ CAR T, analogous to tisagenlecleucel, and FMC63-CD28ζ, analogous to axicabtagene ciloleucel, in vivo in two different B-cell tumor xenograft NOD-SCID-IL2Rγc^–/– (NSG) mouse models. CD19 multi-chain KIR-CAR T cells worked faster and achieved deeper tumor control in a NALM6 leukemia xenograft model than single-chain 41BBζ CAR T cells, with significantly reduced cytokine levels (37-fold less IFNγ, 27-fold less TNFα and 17-fold less IL-2).  In a Burkitt lymphoma xenograft model, KIR-CAR T cells effectively controlled tumors and significantly improved survival, with reduced cytokine production, while single-chain CD28ζ CAR T cells failed to treat tumors despite increased cytokine secretion.

To better assess the benefit-risk profile of KIR-CAR we sought to characterize the tissue distribution of our CD19-targeting KIR-CAR T cells in vivo. NSG mice engrafted with NALM6 leukemia were treated with DS191-KIR-CAR, CD19 single-chain 41BBζ CAR, or negative control untransduced (UTD) T cells. Fourteen days after T cell infusion, serum and tissues were collected for serum toxicology, histopathology, and immunofluorescent histologic imaging. Overall, serum biochemistry results were not significantly different between KIR-CAR-treated and benchmark single-chain CAR-treated mice, and there were no major differences between either treatment group compared to the UTD T cell group. Histopathology revealed that in both negative control mice and 41BBζ single-chain CAR T-treated mice tumor had spread into the spleen, while in KIR-CAR treated mice this did not occur. This result is in keeping with the deeper tumor control seen in KIR-CAR treated mice by in vivo bioluminescent tumor imaging. Notably, despite histopathologic confirmation that no tumors were present in the lungs of any mice, human CD3 immunofluorescent staining demonstrated that single-chain CAR T-treated mice had significantly increased T cell infiltration and accumulation in lungs compared to both the UTD T cell group and to KIR-CAR T-treated mice, suggesting antigen-independent single-chain CAR T cell accumulation in normal lung tissue.

These results suggest that treatment with multi-chain DS191-KIR-CAR may translate into improved anti-tumor activity, reduced risk of toxicity via reduced cytokine secretion, and improved on- versus off-target healthy tissue infiltration and accumulation. These preclinical results continue to support our ongoing FIH Phase 1 clinical trial for patients with CAR-treated or CAR-naive relapsed/refractory B-cell non-Hodgkin lymphomas (NCT06544265), and clinical data will be shared.