Abstract 474: IL-12 immunotherapy using a novel gene delivery platform for RNA payloads achieves effective antitumor responses without systemic toxicity in an aggressive CT26 murine cancer model

Background: IL-12 exhibits potent anti-tumor properties, which enhance cytolytic activity and tumor infiltration of natural killer (NK) cells and cytotoxic T lymphocytes (CTLs) and activates Th-1-type immune responses [1-2]. However, systemic administration of recombinant IL-12 protein is still a challenge due to serious toxicity resulting from interferon (IFN)-γ overproduction [3-5]. Viral and non-viral vectors for IL-12 gene therapy represent an alternative therapeutic strategy, which can be combined with pro-drug activated cancer killing gene therapy approaches. Combined gene delivery of IL-12 and herpes simplex virus thymidine kinase (HSV-TK) significantly boosts anti-tumor efficacy in animal studies and has been shown to be safe in human clinical studies when administered intratumorally [6-12]. Here we present a novel gene therapy approach for in vivo gene delivery of IL-12 combined with an enhanced gain of function-variant of HSV-TK (v-eTK). Methods: Non-replicating retrovectors encoding murine IL-12 only (mGEN1018, encoding both p35 and p40 subunits, as described previously [13-14], or in combination with v-eTK (mGEN1013), were administered intratumorally or systemically to a subcutaneous tumor model of CT26 murine colorectal cancer in syngeneic BALB/c mice. Mice receiving mGEN1013 were subsequently treated with Ganciclovir (GCV) pro-drug. General health and body weight as well as tumor growth and survival were monitored over time, and immunophenotyping of peripheral blood, spleen, and tumor-infiltrating immune cells was performed. Results: Our data show that in subcutaneous CT26 tumor models, intratumoral and systemic gene delivery of IL-12 by the mGEN1018 vector induces both local and systemic anti-tumor immune responses primarily by stimulating production of IFN-γ from NK cells and CTLs. Gene delivery of IL-12 in combination with v-eTK by the mGEN1013 vector followed by GCV pro-drug showed enhanced tumor growth inhibition and increased survival compared to untreated controls. Systemic anti-tumor effects of IL-12 gene therapy were not associated with visible signs of toxicity or significant body weight loss. Conclusion: Gene delivery of v-eTK followed by GCV treatment results in tumor cell lysis and exposure of tumor antigens, while simultaneously, IL-12 gene delivery enhances anti-tumor immune responses though upregulation of inflammatory cytokines. These results show the potential of mGEN1013 as a novel RNA payload delivery platform for combined GCV pro-drug and more effective tumor-localized IL-12 immunotherapy without systemic toxicity. Citation Format: Laura Strauss, Carli Jones Burns, Akihito Inagaki, Stephanie Lees, Noriyuki Kasahara, Cecilia Roh, Robert G. Johnson. IL-12 immunotherapy using a novel gene delivery platform for RNA payloads achieves effective antitumor responses without systemic toxicity in an aggressive CT26 murine cancer model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 474.