Ori-TTV particles enable potent in vivo CAR-T cell engineering for the treatment of hematologic malignancies and autoimmune diseases 2256860

Abstract Introduction Chimeric antigen receptor (CAR) T cell therapy has significantly advanced treatment of hematologic malignancies, including B-cell acute lymphoblastic leukemia (B-ALL), non-Hodgkin lymphoma (NHL), and multiple myeloma (MM). However, conventional autologous CAR-T therapies are hampered by logistical complexity, high costs, and lengthy manufacturing timelines, limiting accessibility–particularly for patients with aggressive disease. To address these challenges, we developed an in vivo CAR-T platform that directly engineers functional CAR T cells within the body, eliminating the need for ex vivo manipulation. Methods Here we report the development and validation of Ori-TTV, a rationally designed T cell—targeting lentiviral vector optimized for safe and efficient in vivo CAR-T generation. Key innovations of Ori-TTV include: (1) an Ori Multiple Targeting Domain (Ori-MTD) for selective T cell transduction; (2) a synthetic T cell—specific promoter for lineage-restricted CAR expression; (3) a phagocytosis-resistant modification to evade macrophage clearance; and (4) a de-targeted fusogen to enhance safety and reduce off-target effects. These features enable selective in vivo reprogramming of T cells. Results Ori-TTV encodes a dual-target CAR recognizing both CD19 and BCMA, facilitating simultaneous elimination of malignant B and plasma cells. This expands therapeutic potential across relapsed/refractory B-ALL, NHL, and MM, and B/plasma cell—driven autoimmune disorders such as systemic lupus erythematosus (SLE), idiopathic inflammatory myopathy (IIM), and systemic sclerosis (SSc). In vitro, the T cell—specific promoter drives CAR expression at levels comparable to EF-1α, supporting robust and multi-round cytotoxicity against CD19+ and BCMA+ cells. In vivo, Ori-TTV administration induced significant and durable tumor regression in humanized PBMC and CD34+ mouse models. Conclusion In summary, Ori-TTV offers a safe, scalable, cost-effective platform to broaden access to CAR-T immunotherapy. Funding Source n/a Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)