Downmodulation of E1A protein expression as a novel strategy to design cancer-selective adenoviruses.

Oncolytic adenoviruses are being tested as potential therapies for human malignant tumors, including gliomas. Here we report for the first time that a mutation in the E1A gene results in low levels of E1A protein, conditioning the replication of mutant adenoviruses specifically to cancer cells. In this study, we compared the oncolytic potencies of three mutant adenoviruses encompassing deletions within the CR1 (Delta-39), CR2 (Delta-24) regions, or both regions (Delta-24/39) of the E1A protein. Delta-39 and Delta-24 induced a cytopathic effect with similar efficiency in glioma cells and a comparable capacity for replication. Importantly, the activity of Delta-39 was significantly attenuated compared to Delta-24 in proliferating normal human astrocytes. Direct analyses of the activation of E2F-1 promoter demonstrated the inability of Delta-39 to induce S-phase-related transcriptional activity in normal cells. Interestingly, E1A protein levels in cells infected with Delta-39 were remarkably downmodulated. Furthermore, protein stability studies revealed enhanced degradation of CR1 mutant E1A proteins, and inhibition of the proteasome activity resulted in the striking rescue of E1A levels. We conclude that the level of E1A protein is a critical determinant of oncolytic phenotype and we propose a completely novel strategy for the design and construction of conditionally replicative adenoviruses.