An engineered α1β1 integrin-mediated FcγRI signaling component to control enhanced CAR macrophage activation and phagocytosis

Treatment of solid tumors remains difficult, and therefore there has been increased focus on chimeric antigen receptor macrophages (CAR-M) to challenge solid tumors. However, CAR domain design of of adoptive cell therapy, which leads to differences in antitumor activity and triggered antitumor potential, remains poorly understood for macrophages. We developed an α1β1 integrin-mediated Fc-gamma receptor I (FcγRI) signaling component for CAR-M specific activation and its antitumor potential. We evaluated CAR-M effects with α1β1 integrin-mediated FcγRI signaling (ACT CAR-M) on the activation and antitumor phagocytic response of macrophages in vitro. Subcutaneous tumor model in BALB/c mice and carcinomatosis model in immunodeficient mice were used to test the antitumor effect of ACT CAR-M compared with CD3ζ CAR-M. The α1β1 integrin-mediated FcγRI signaling engagement of CAR-M was associated with enhanced macrophage activation and specific phagocytosis in primary human macrophages, and significantly improved tumor control and survival in multiple cancer models when compared to CD3ζ CAR-M. RNA-sequencing suggested that α1β1 integrin-mediated FcγRI engagement increased antitumor immunity by enhancing pro-inflammatory M1 phenotype-associated pathways, such as Toll-like receptor signaling, tumor necrosis factor signaling, and IL-17 signaling. α1β1 integrin-mediated FcγRI signaling engagement markedly enhanced antitumor effects of CAR-M immunotherapy, which is proposed as an advanced engineering CAR domain material to expand the clinical application of CAR-M. Immune suppression usually causes immune cells switched into dysfunctional states. There is increasing interest in chimeric antigen receptor macrophages (CAR-M) to challenge solid tumors. However, the optimal CAR for CAR-M is not clear. Here we designed an α1β1-mediated Fc-gamma receptor I (FcγRI) signaling component for CAR-M specific activation and phagocytosis. Using stringent models, we demonstrate that α1β1-mediated FcγRI signaling more strongly than CD3ζ signaling influences the anti-tumor effects of CAR-M immunotherapy in vivo. These studies highlight CAR-guided engineering of intracellular domains for macrophage and broad the potential of clinical application on CAR-M. [Display omitted] •α1β1 integrin-mediated FcγRI signaling component contribute to ACT CAR-M enhanced activation and phagocytosis.•ACT CAR-M have superior antitumor control compared to those of CD3ζ CAR-M in vivo.•α1β1 integrin-mediated Fc