Previous studies conducted in our laboratories indicated that genetically engineered HSV-1 mutants display oncolytic activities against human gliomas. Currently, we present new data indicating that several mutant viruses (HSV-1) effectively killed in vitro human cell lines of various origins (i.e., gliomas, meningiomas, neuroblastomas, melanomas), as indicated by a semi-quantitative microassay used to measure cell viability. Additionally, cell infectivity by 7 different HSV-1 mutants was supported by replication assay results, immunohistochemical, in situ hybridization and Western blot analysis. Infection of normal human astrocytes and sensitivity to antiviral drugs such as Acyclovir were tested (E.C.50 =0.5u.g/mL) to address safety concerns for potential m vivo use. HSV-1 mutant R4009 (stop codon inserted in open reading frame of neurovirulence gene yi 34.5) significantly prolonged survival of C.B-17 seid mice bearing intracranial xenografts of two different human glioma cell lines, U251MG and D54MG. Median survival of U251MG-bearing seid mice increased from 33-34 days to 49-56 days (pO.005) at virus doses of 107 pfu added at, or 5 days after, tumor induction. Similarly, seid mice intracranially injected with DS4MG experienced increases in median survival from 19-22 days to 40-53 days (p<0.005). Based on evidence presented herein, genetically engineered HSV mutants appear likely candidates for treatment of malignant human gliomas in conjunction with adjunctive therapies.