Local Delivery of mRNA Encoding Cytokines Promotes Antitumor Immunity and Tumor Eradication Across Multiple Preclinical Tumor Models

Local immunotherapy ideally stimulates immune responses against tumors while avoiding toxicities associated with systemic administration. Current strategies for tumor-targeted, gene-based delivery, however, are limited by adverse effects such as off-targeting or anti-vector immunity. We investigated the intratumoral administration of saline-formulated messenger (m)RNA encoding four cytokines that were identified as mediators of tumor regression across different tumor models: interleukin (IL)-12 single chain, interferon (IFN)-α, granulocyte-macrophage colony-stimulating factor, and IL-15 sushi. Effective antitumor activity of these cytokines relied on multiple immune cell populations and was accompanied by intratumoral IFN-γ induction, systemic antigen-specific T cell expansion, increased granzyme B+ T cell infiltration, and formation of immune memory. Antitumor activity extended beyond the treated lesions and inhibited growth of distant tumors as well as disseminated tumors. Combining the mRNAs with immunomodulatory antibodies enhanced antitumor responses in both injected and uninjected tumors, thus improving survival and tumor regression. Consequently, clinical testing of this cytokine-encoding mRNA mixture is now underway. Subcutaneous B16F10 tumors in C57BL6 mice (n=5/group) were treated with a single intratumoral injection of cytokine mRNA mixture (80 µg total, 20 µg per mRNA component) encoding a murine single chain IL-12, murine IL-15sushi, murine GM-CSF and murine IFNalpha or control mRNA (80 µg luciferase mRNA) . Tumors were collected 7 days following intratumoral mRNA injection and subjected to global gene expression to assess changes between cytokine mRNA and control treated tumors.