Engineered herpes simplex virus expressing IL-12 in the treatment of experimental murine brain tumors

Genetically engineered, neuroattenuated herpes simplex viruses (HSVs) expre ssing various cytokines can improve survival when used in the treatment of experimental brain tumors. These attenuated viruses have both copies of gam ma(1)34.5 deleted. Recently, we demonstrated increased survival of C57BL/6 mice bearing syngeneic GL-261 gliomas when treated with an engineered HSV e xpressing IL-4, as compared with treatment with the parent construct (gamma (1)34.5(-)) alone or with a virus expressing IL-10. Herein, we report const ruction of a conditionally replication-competent mutant expressing both sub units of mIL-12 (M002) and its evaluation in a syngeneic neuroblastoma muri ne model. IL-12 induces a helper T cell subset type 1 response, which may i nduce more durable antitumor effects. In vitro studies showed that, when in fected with M002, both Vero cells and murine Neuro-2a neuroblastoma cells p roduced physiologically relevant levels of IL-12 heterodimers, as determine d by ELISA. M002 was cytotoxic for Neuro-Za cells and human glioma cell lin es U251MG and D54MG. Neurotoxicity studies, as defined by plaque-forming un its/LD50, performed in HSV-1-sensitive A/J strain mice found that M002 was not toxic even at high doses. When evaluated in an intracranial syngeneic n euroblastoma murine model, median survival of M002-treated animals was sign ificantly longer than the median survival of animals treated with R3659, th e parent gamma(1)34.5(-) mutant lacking any cytokine gene insert. Immunohis tochemical analysis of M002-treated tumors identified a pronounced influx o f CD4(+) T cells and macrophages as well as CD8(+) cells when compared with an analysis of R3659-treated tumors. We conclude that M002 produced a surv ival benefit via oncolytic effects combined with immunologic effects medita ted by helper T cells of subset type 1.