M yb34.5 is a herpes simplex virus 1 (HSV-1) mutant deleted in the gene for ribonucleotide reductase (ICP6). It also carries a version of γ₁34.5 (a viral gene product that promotes the dephosphorylation of eIF-2α) that is under control of the E2F-responsive cellular B-myb promoter, rather than of its endogenous promoter. Myb34.5 replication in tumor cells results in their destruction (oncolysis). γ₁34.5 expression by HSV-1 subverts an important cell defense mechanism against viral replication by preventing shutoff of protein synthesis after viral infection. Infection of colon carcinoma cells with Myb34.5 results in greater eIF-2α dephosphorylation and viral replication compared with infection with HSV-1 mutants completely defective in γ₁34.5 expression. In contrast, infection of normal hepatocytes with Myb34.5 results in low levels of eIF-2α dephosphorylation and viral replication that are similar to those observed with HSV-1 mutants completely defective in γ₁34.5 and ICP6. When administered intravascularly into mice with diffuse liver metastases, Myb34.5 has greater antineoplastic activity than HSV-1 mutants with completely defective γ₁34.5 expression and more restricted biodistribution compared with HSV-1 mutants with wild-type γ₁34.5 expression. Myb34.5 displays reduced virulence and toxicity compared to HSV-1 mutants with wild-type γ₁34.5 expression. Portal venous administration of Myb34.5 significantly reduces liver tumor burden in and prolongs the life of mice with diffuse liver metastases. Preexisting Ab’s to HSV-1 do not reduce the antitumor efficacy of Myb34.5 in vivo.