Abstract 5205: VG2025: Transcriptional and translational dually regulated (TTDR) oncolytic virus for cancer immunotherapy

Abstract

Abstract VG2025 is a genetically modified HSV-1 oncolytic virus (OV) wherein the transcription and translation of critical viral genes are regulated to enhance virus safety and promote tumor specific virus replication without attenuating the virus. The TTDR design is based on transcriptional control of the essential HSV-1 immediate-early protein ICP27 using a tumor specific CEA promoter, coupled with translational control of the key neurovirulence factor ICP34.5 accomplished by introducing multiple tandem copies of miRNA binding sites for miR-124 and miR-143 which are downregulated in tumors but abundantly expressed in normal brain tissues. The anti-tumor and immune stimulatory properties of VG2025 are also enhanced by expressing a pair of cytokines (IL-12 and IL-15/IL-15Rα complex) that are linked by 2A cleavage peptides and controlled by the tumor specific promoter CXCR4. The dual regulation strategy was verified in vitro and shown to be highly selective. Growth of VG2025 virus correlated with CEA expression in a panel of cell lines including normal cells and a selection of tumor cells with varying CEA levels. miRNA-mediated regulation of ICP34.5 expression was validated by transfecting HEK293t cells with miR-124 and miR-143 precursors, followed by infection with VG2025. RT-qPCR testing revealed a significant decrease in the levels of viral ICP34.5 mRNA when the miRNA was present. VG2025 virus replication was evaluated in a variety of human tumor cell lines, and two cell lines representing NSCLC and pancreatic cancer (A549 and BxPC3, respectively) were selected for testing in a humanized mouse model, where VG2025 displayed strong in vivo anti-tumor efficacy. The immune stimulatory characteristics of VG2025, and the bioactivity of expressed cytokines, were demonstrated in a syngeneic mouse colorectal tumor model (CT26) and lymphoma model (A20) using VG2026, which is an identical mouse specific variant of VG2025 that expresses murine IL-12 in place of human IL-12. In addition to a significant reduction in the growth of injected tumors, we observed a clear abscopal effect on contralateral tumors, indicating a robust anti-tumor immune response with an immune memory effect. Due to the tendency of HSV-1 to establish latency which may lead to neurotoxicity and potential safety concerns, we performed a safety pharmacology study by subcutaneously inoculating immunocompetent DBA mice with VG2025. Absence of neurotoxicity was further verified by more aggressive routes of administration, including via the nose and by corneal scarification. The risk of VG2025 reactivation from a latent state was evaluated in a two-step study: latent infection in the CNS was first established and confirmed, then the trigeminal ganglia were removed and co-cultured with Vero cells to detect reactivation-related cytopathic effects. All studies showed that VG2025 was safe with no reactivation events detected. Citation Format: Yanal Murad, Guoyu Liu, I-Fang Lee, Zahid Delwar, Dmitry Chouljenko, Will Liu, Jun Ding, Luke Bu, Ronghua Zhao, William Jia. VG2025: Transcriptional and translational dually regulated (TTDR) oncolytic virus for cancer immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5205.