CircRNA-based, non-integrated, in vivo panCAR-mediated B cell immune resetting in mouse models and non-human primates

Abstract

B cell immune resetting, defined as the comprehensive depletion and subsequent reconstitution of B cells, represents a pivotal therapeutic approach for B cell-associated disorders. Recently, chimeric antigen receptor T cell (CAR-T) therapy has demonstrated substantial clinical efficacy in treating B cell malignancies and autoimmune diseases. However, this approach remains constrained by its reliance on lymphodepleting conditioning regimens, high invasiveness, high costs, and the risk of severe cytokine release syndrome (CRS). Antibody-based therapies, including monoclonal antibodies and T cell-engager (TCE) bispecific antibodies, suffer from suboptimal tissue penetration, resulting in inadequate B cell depletion in lymphoid and peripheral tissues, and subsequent rapid resurgence of pathogenic B cells. Here, we engineered highly stable circular RNAs (circRNAs) encoding anti-CD19 CAR (circRNA-CAR), which demonstrated prolonged and enhanced CAR protein expression compared to conventional linear mRNAs. By employing immune cell-tropic lipid nanoparticles (LNPs) for in vivo delivery, circRNA-CAR facilitated the generation of panCAR immune cells (including CAR-T, CAR-NK, and CAR-macrophages) in both murine models and cynomolgus monkeys. Compared to rituximab or teclistamab, in vivo panCAR induced more extensive B cell depletion across multiple tissue compartments in mice. Moreover, in vivo panCAR mediated robust B cell depletion, eliminated autoreactive antibodies, and restored renal function by alleviating proteinuria in a systemic lupus erythematosus (SLE) murine model. It also effectively reduced IgE antibody levels, decreased eosinophil activity, and improved lung pathology in an asthma murine model. Notably, in vivo panCAR-mediated B cell depletion significantly attenuated aging-associated phenotypes in 17- month-old aging mice, remarkably outperforming both rituximab and teclistamab. In cynomolgus monkeys, in vivo panCAR achieved sustained B cell depletion lasting approximately 40 to 50 days, followed by reconstitution of a naive B cell repertoire. Collectively, circRNA-based in vivo panCAR enables complete B cell immune resetting, offering a versatile and promising therapeutic platform for a broad spectrum of B cell- and antibody-mediated disorders.