In Vivo generation of anti-CD19 CAR-T cells for the treatment of B-cell mediated autoimmune diseases

Abstract

Abstract Emerging clinical data demonstrate the transformative potential of B cell-depleting CAR-T therapies for patients with autoimmune diseases. However, despite their promise, the broad clinical deployment of ex vivo CAR-T therapies remains limited by several key challenges including manufacturing complexity, high cost, the need for lymphodepleting chemotherapy, and safety risks related to cytokine release syndrome. In addition, the persistence of administered CAR-T cells can lead to prolonged B cell aplasia, leaving patients immunocompromised long after the clearance of pathogenic B cell clones. To overcome these limitations while preserving efficacy, we have developed a novel, transient in vivo T cell engineering platform that utilizes our targeted lipid nanoparticle (tLNP) to deliver mRNA encoding an anti-CD19 CAR specifically to cytotoxic CD8+ T cells. This targeted delivery system drives rapid and precise in vivo engineering of T cells without the need for ex vivo manipulation or pre-conditioning. The resulting CAR-T cells exhibit potent cytolytic activity and high specificity across in vitro assays and preclinical in vivo models. Following a single two-dose treatment cycle in studies of CD34+ humanized mice and non-human primates, we observe potent and durable B cell depletion from the periphery and B cell rich tissues, including the spleen, bone marrow and lymph nodes at doses as low as 0.3 mg/kg. Importantly, this depletion encompasses differentiated B cell populations while sparing early progenitors. In the weeks after the transient depletion phase, B cell recovery is observed, marked by the emergence of a predominantly naïve B cell compartment. This profile is consistent with a clinically relevant immune reset, thought to be a benchmark for successful treatment in autoimmune patients. Together, these data demonstrate that transient, in vivo-generated CAR-T cells, enabled by our proprietary tLNP platform, can recapitulate the benefits of ex vivo CAR-T therapies while minimizing associated risks. This approach offers a scalable, repeatable, and safer alternative to ex vivo CAR T therapies in autoimmune disease, with broad applicability across diverse patient populations.