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A new procedure is described for the generation of high-titer, helper-free retrovirus vectors employing receptor-mediated, adenovirus-augmented transfection into a standard packaging cell line. Viral titers are increased 30-fold to 100-fold in transiently (> 10(5) infectious units per mL) and stable (> 10(7) infectious units per mL) transfected cells as compared with either CaPO4-mediated transfection or retroviral infection of a packaging cell line. Further, expression of the transduced genes was drastically increased in the transfected cells, but, as expected, there was no difference in transduction efficiency and gene expression in the infected target cells. The increases in viral titers were most likely due to the high number of stable, integrated copies of the vector plasmid DNA in the resulting packaging lines following G418 selection. In addition, experiments generating recombinant retroviruses from non-packaging cell lines are presented. The results suggest that this procedure may be of use to generate high-titer retrovirus vectors in packaging cell lines as well as in primary cells, thus providing a technical basis for in vivo gene transfer upon transplantation of these cells into various organs.
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