B-myb promoter retargeting of herpes simplex virus γ34.5 gene-mediated virulence toward tumor and cycling cells

Deletion of the gamma 34.5 gene coding for virulence markedly reduces cytotoxicity mediated by herpes simplex virus type 1 (HSV-1) (J. M. Markert et al., Neurosurgery 32:597-603, 1993; N. S. Markovitz et at, J. Virol. 71:5560-5569, 1997), To target lytic virulence to tumors, we have created a novel HSV-1 mutant, designated Myb34.5. This viral mutant is characterized by a deletion of the gene for infected cell polypeptide 6 (ICP6; also known as UL39 or ribonucleotide reductase) and of the two endogenous copies of the gamma 34.5 gene (RL1) and by reintroduction of one copy of gamma 34,5 under control of the E2F-responsive, cellular B-myb promoter. On direct intracerebral inoculation in BALB/c mice, the 50% lethal dose (LD50) for Myb34,5 was 2.7 x 10(7) PFU while that for HSVs with mutations in the gamma 34,5 gene could not be technically achieved with available viral stocks and it was estimated as >1 x 10(7) PFU. The LD50 for an HSV with a single defect in ICP6 function was 1.3 x 10(6) PFU. Conversely, Myb34.5's oncolytic efficacy against a variety of human glioma cells in culture and in vivo was enhanced compared to that of HSVs with gamma 34.5 mutations, and in fact, it was comparable to that of the wild-type F strain and of viral mutants that possess a wild-type gamma 34.5 gene. The characteristic shutoff of host protein synthesis, occurring after infection of human SK-N-SH neuroblastoma cells by gamma 34,5 mutant viruses (J. Chou and B. Roizman, Proc. Natl. Acad. Sci. USA 89:3266-3270, 1992), was not present after infection with Myb34.5. There was an increase of almost 3 logarithmic units in the production of progeny virus in arrested fibroblasts compared to that in cycling fibroblasts infected with Myb34.5. These results suggest that transcriptional regulation of gamma 34.5 by cell cycle-regulated promoters can be used to target HSV-1 virulence toward tumors while maintaining the desirable neuroattenuated phenotype of a gamma 34.5 mutant.