High-Efficiency Stable Gene Transfer of Adenovirus into Mammalian Cells Using Ionizing Radiation

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Abstract
We report a novel method for targeting adenovirus-mediated gene delivery. By irradiating mammalian cells prior to adenoviral transduction, adenoviral gene transfer is greatly improved and the adenoviral genome integrates into cellular DNA. In this work, human and rodent cell lines were irradiated and subsequently transduced with the adenovirus vector Ad5CMVlacZ. Initial levels of transduction were as much as 40-fold higher in irradiated cells, and this improvement in transduction was radiation dose dependent. The duration of lacZ expression in irradiated cells was also much longer than in nonirradiated cells and reached a plateau after 21 days. At doses of 7 Gy, long-term (lacZ could be detected in 15% of cells by flow cytometry. This long-lasting expression of lacZ was due to viral DNA integration into the host genome. Thus, pretreatment of cells with ionizing radiation improves both immediate transduction efficiency and duration of transgene expression. This may lead to the development of new protocols combining radiation and gene therapy in treating human malignancy. We show that modest doses of ionizing radiation can greatly improve adenoviral transduction efficiency. This improvement of gene transfer was found in mouse (NIH-3T3) cells, human neoplastic cells (A549), and primary human cells (39F). This improvement is long lasting (more than 3 weeks). This long-lasting expression in irradiated cells is due to viral DNA integration into the host genome. The combination of radiation with adenoviral transduction increases both initial and long-term viral transgene expression, and may lead to new combined modality protocols.

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