Hypoxia reduces adenoviral replication in cancer cells by downregulation of viral protein expression

Successful cancer therapy using replicating viral vectors relies on the spread of virus from infected to uninfected cells. To date, there has been limited clinical success in the use of replicating adenoviruses. In animal models, established xenograft tumors are rarely eliminated despite the persistence of high viral titers within the tumor. Hypoxia is a prevalent characteristic of solid tumors. Adenovirus naturally infects tissues exposed to ambient oxygen concentrations. In addition, the proximity of the virus to blood vessels within persistent xenograft tumors suggests that the hypoxic environment plays an important role in limiting adenoviral replication. We have previously established that hypoxia (1% oxygen) limits adenoviral replication in H1299 and A549 lung cancer cells (Mol Ther 2003; 7: 5 part 2). However, hypoxia did not reduce cell viability or restrict S-phase entry. To further evaluate the mechanism by which hypoxia reduces viral production, the transcription and translation of viral proteins were evaluated in hypoxic and ambient conditions. H1299 and A549 lung cancer cells were infected with wild type adenovirus Ad309 (MOI 10) and incubated for 24h or 48h in ambient (21% oxygen) or hypoxic (1% oxygen) environments. Immunoblotting was performed for two viral proteins, E1a and fiber. For both H1299 and A549 cells the production of E1a and fiber under hypoxic conditions was substantially decreased at 24h and 48h compared to room air controls. In contrast, northern analysis for A549 and H1299 cells showed similar levels of E1a mRNA in room air and hypoxic conditions at 24h. In conclusion, the level of hypoxia similar to that found within tumors reduces the replication of adenoviral vectors by reduction of viral protein expression without a reduction in mRNA levels.