synNotch CAR-Macrophages promote persistent antitumour activity through enhanced adaptive immunity

Chimeric antigen receptor-macrophage (CAR-M) possess the ability to migrate into the immunosuppressive tumour microenvironment (TME) of solid tumours and mediate phagocytosis. However, their efficacy is limited by challenges in precisely controlling tumour-specific immune activation and sustaining adaptive antitumour responses due to systemic off-target risks and inefficient antigen presentation. To overcome these limitations, we engineered an antigen prime-and-kill circuit, wherein tumour-specific synthetic Notch (syN) receptors locally induce the production of a kill antigen. This circuit enhances the precision of CAR-M while establishing self-sustaining antigen-presentation networks, amplifying tumour-specific T-cell adaptive immunity without eliciting systemic cytokine toxicity. To enable in situ CAR-M generation, we developed a macrophage-derived exosome-based delivery system to introduce syN-CAR genes into tumour-associated macrophages (TAMs), thereby generating syN-CAR-M within the TME. These engineered CAR-M actively sought and engulfed cancer cells while promoting a robust adaptive antitumour immune response in the TME in a murine tumour model. Our findings highlight that in situ induction of syN-CAR-M triggers a potent, tumour-specific adaptive immune response, offering a novel strategy for CAR-M-based immunotherapy and advancing precision cancer treatment by enhancing efficacy while minimizing systemic toxicity.