Cancer immunotherapy localized to the tumor microenvironment holds great potential to promote innate and adaptive immune responses against tumors, while avoiding toxicities related to systemic administration of immuno-modulatory therapeutics. Current strategies for tumor-targeted, gene-based delivery of immune therapies face limitations in the clinic due to suboptimal target expression, anti-vector immunity, potential for unwanted genomic rearrangements and other off target effects. We developed a highly potent synthetic mRNA-based platform for in vivo transfection and sustained intratumoral expression of immuno-modulatory molecules that is capable of inducing immunity to tumor specific antigens while avoiding anti-vector immunity and distal tissue expression often associated with viral gene therapy or oncolytic viruses. Using iterative rounds of in vivo screening in murine syngeneic tumor models, we identified a combination of four synthetic mRNAs encoding potent cytokines that mediate high frequency of complete tumor regressions across different tumor models. Mechanistically, anti-tumor activity of cytokine mRNAs relied on the action of T cells as well as NK cells and was accompanied by robust intratumoral induction of interferon gamma, systemic expansion of antigen-specific T cells and increased granzyme B positive CD8+ T cell infiltration. Immunological memory to both dominant and subdominant antigens was formed that protected long-term survivors from re-challenge with autologous tumors. Importantly, although cytokine mRNAs were administered intratumorally, resulting in local target expression, anti-tumor activity extended beyond the injected lesion to effectively control the growth of distant tumors in both a dual tumor model and in an experimental lung metastasis model. Finally, the combination of four cytokine mRNAs with immunomodulatory antibodies further enhanced the abscopal effect leading to improved overall survival and higher incidence of complete tumor regressions across several preclinical models. In summary, the robust and versatile synthetic mRNA platform reported herein was used to identify multi-modal localized cancer immunotherapy with broad anti-tumor activity against treated and untreated lesions in a range of preclinical cancer models.

Citation Format: Timothy R. Wagenaar, Christian Hotz, Friederike Gieseke, Hui Cao, Jan Diekmann, Mustafa Diken, Christian Grunwitz, Andy Hebert, Karl Hsu, Marie Jordan, Katalin Kariko, Sebastian Kreiter, Andreas N. Kuhn, Mikhail Levit, Natalia Malkova, Serena Masciari, Jack Pollard, Hui Qu, Abderaouf Selmi, Julia Schlereth, Fangxian Sun, Bodo Tillmann, Tatiana Tolstykh, Lena Wicke, Sonja Witzel, Qunyan Yu, Yu-An Zhang, Gang Zheng, Gary Nabel, Joanne Lager, Ugur Sahin, Dmitri Wiederschain. Combinatorial treatment with intratumoral cytokine mRNAs results in high frequency of tumor rejection and development of anti-tumor immunity across a range of preclinical cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-130.