Abstract LB381: Preclinical toxicology and efficacy of NG34C: A novel oncolytic HSV-1 expressing truncated GADD34 gene for the treatment of glioblastoma

Abstract Glioblastoma (GBM) remains a formidable medical challenge with limited therapeutic progress. Oncolytic viruses (OVs) offer promise by triggering immunogenic cell death that potentially activates anti-tumor immunity. We recently completed a Phase I clinical trial of rQNestin34.5v2 (CAN-3110), an oncolytic virus encoding the viral neurovirulence gene ICP34.5 under the nestin promoter, demonstrating safety at doses up to 1 x 1010 PFU (Ling et al., Nature, 2023). The human GADD34 gene, a stress-response protein homologous to the C-terminal region of ICP34.5, has been used to replace ICP34.5 in oHSV (NG34). This modified virus showed comparable efficacy to rQNestin34.5 with reduced toxicity (Nakashima et al., Clinical Cancer Research, 2018). However, GADD34 is inherently unstable due to rapid proteasomal degradation. To enhance GADD34 stability, we deleted its N-terminal region responsible for proteasomal degradation, generating NG34C. This modified oncolytic virus maintained stable GADD34 expression in gliomas, sustained eIF2α dephosphorylation, and improved replication under hypoxia by mitigating elevated eIF2α phosphorylation. NG34C produced viral yields comparable to rQNestin34.5 and NG34 under normoxia while outperforming them in hypoxia. Toxicology and efficacy studies in mice demonstrated NG34C’s safety, with no neurotoxicity at doses up to 3 × 107 PFU and minimal toxicity at 108 PFU. Unlike wild-type HSV-1 (F strain), which caused significant lethality, NG34C did not persist in normal brain tissue, as viral DNA and RNA levels were comparable to PBS by day 3. In a 1694 GBM model (human EGFR overexpression with Cdkn2a and Pten loss), NG34C significantly improved survival at 3 × 105 and 3 × 107 PFU doses (P < 0.0001 vs. PBS). These results establish NG34C as a safe, hypoxia-adapted, and effective oncolytic virus for GBM. NG34C is now being produced under GMP for an upcoming INS-clinical study. Citation Format: Min J. Kim, Silja Schule, Tran Nguyen, Kazue Kasai, Michal O. Nowicki, Alexander L. Ling, William F. Goins, Hiroshi Nakashima, E. Antonio Chiocca. Preclinical toxicology and efficacy of NG34C: A novel oncolytic HSV-1 expressing truncated GADD34 gene for the treatment of glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2):Abstract nr LB381.