591 Comparison of two oHSV Vectors for the treatment of glioblastoma

Background Glioblastoma multiforme (GBM) is the most common human brain cancer. Despite a well-established standard of care, the 5-year mortality rate of GBM patients is 95%, highlighting the need for innovative therapeutic interventions. A variety of oncolytic viruses, including those derived from herpes simplex virus (oHSV), have been designed for GBM therapy, but early-phase clinical trials have reported few complete responses and no evidence of durable anti-tumor immunity. Potential reasons for the lack of efficacy are limited vector potency (i.e., virulence) and the presence of a highly immunosuppressive tumor microenvironment (TME) comprised of few activated lymphocytes, large numbers of immunosuppressive myeloid cells (macrophages, myeloid derived suppressor cells [MDSCs], microglia), and an agglomerate of immunosuppressive cytokines (IL-10, VEGF, MIF, etc.).1 Herein we explore these obstacles by comparing the anti-tumor activity two different oHSV designs, an HSV-1 KOS strain derivative designated KG4:T124, and an F strain derivative designated rQNestin34.5v.1 (a similar oHSV, rQNestin34.5v.2, is currently in a phase I clinical trial for GBM).2 Methods Using the murine syngeneic GBM models, GL261N4 and CT2A, we compared the anti-tumor activity of KG4:T124 and rQNestin34.5v1. In vitro, we evaluated the viral entry, replication capacity, and cytotoxicity of both oHSVs. In vivo, we measured the impact of both vectors on tumor progression, TME immune cell composition, and animal survival. Results Virus entry into cancer cells of KG4:T124 or rQNestin34.5v1 was relatively similar, but rQNestin34.5v.1 replicated more effectively and generally induced greater viral mediated cytotoxicity. In syngeneic mice, rQNestin34.5v.1 reduced orthotopic GL261N4 tumor burden and enhanced animal survival compared to KG4:T124. However, preliminary data indicate that multiple injections of KG4:T124 but not rQNestin34.5 enhance GL261N4 survival outcome. Neither oHSV impacted survival outcomes in the more pernicious CT2A model. Analysis, of either the GL261N4 or CT2A TME two days post virus administration revealed that both viruses had reduced microglia cell frequency, induced the influx of tumor associated macrophages and polymorphonuclear cells, but did not alter the frequency of monocytic MDSCs, natural killer cells, CD8+ or CD4+ T-cells. Conclusions rQNestin34.5 had greater oncolytic activity In vivo and in vitro, but did not benefit from multiple oHSV injections. Both viruses induced similar changes in the TME immune cell composition. However, the presence of vital adaptive immune cell types within the TME was not observed at 2 days post oHSV treatment. Acknowledgements Research reported in this abstract was supported by the National Institutes of Health grants P01CA163205 (EAC & JCG) and 5T32CA082084-19 (JWJ). References Muller S, Kohanbash G, Liu SJ, Alvarado B, Carrera D, Bhaduri A, Watchmaker PB, Yagnik G, Di Lullo E, Malatesta M, Amankulor NM, Kriegstein AR, Lim DA, Aghi M, Okada H, Diaz A. Single-cell profiling of human gliomas reveals macrophage ontogeny as a basis for regional differences in macrophage activation in the tumor microenvironment. Genome Biol. 2017;18(1):234. Epub 2017/12/22. doi: 10.1186/s13059-017-1362-4. PubMed PMID: 29262845; PMCID: PMC5738907. Chiocca EA, Nakashima H, Kasai K, Fernandez SA, Oglesbee M. Preclinical Toxicology of rQNestin34.5v.2: An Oncolytic Herpes Virus with Transcriptional Regulation of the ICP34.5 Neurovirulence Gene. Mol Ther Methods Clin Dev 2020;17:871–93. Epub 2020/05/07. doi: 10.1016/j.omtm.2020.03.028. PubMed PMID: 32373649; PMCID: PMC7195500.