Keratinocytes as a target for gene therapy - Sustained production of erythropoietin in mice by human keratinocytes transduced with an adenoassociated virus vector

Background and Design: Keratinocytes are ideal targets for somatic gene therapy. Among the viral gene transfer systems, adenoassociated virus vectors have recently gained attention. We studied the feasibility of using adenoassociated virus-transduced human keratinocytes to provide a long-term, high-level production of a therapeutic factor after implantation in mice. Results: Transduction of HeLa cells by an adenoassociated virus vector was ascertained by transfer of the beta-galactosidase reporter gene, which was visualized by the blue staining of infected cells after fixation and coloring by X-Gal (the substrate of the reaction for beta-galactosidase activity). In a second step, 2 HeLa cell lines transduced with an AAV harboring the erythropoietin complementary DNA and producing high amounts of erythropoietin in vitro were isolated. After implantation in nude mice, a high-level and long-term increase in hematocrit (for the 1-month duration of the study) was found, which was correlated to the size of the induced tumor. Conclusions: Adenoassociated virus-transduced HeLa keratinocytes provide high-level, stable, and longterm production of a therapeutic protein in mice. These results must now be extended to human primary keratinocytes.