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 gen e therapy. Among the viral gene transfer systems, adenoassociated viru s vectors have recently gained attention. We studied the feasibility o f using adenoassociated virus-transduced human keratinocytes to provid e a long-term, high-level production of a therapeutic factor after imp lantation in mice. Results: Transduction of HeLa cells by an adenoasso ciated virus vector was ascertained by transfer of the beta-galactosid ase reporter gene, which was visualized by the blue staining of infect ed cells after fixation and coloring by X-Gal (the substrate of the re action for beta-galactosidase activity). In a second step, 2 HeLa cell lines transduced with an AAV harboring the erythropoietin complementa ry DNA and producing high amounts of erythropoietin in vitro were isol ated. After implantation in nude mice, a high-level and long-term incr ease in hematocrit (for the 1-month duration of the study) was found, which was correlated to the size of the induced tumor. Conclusions: Ad enoassociated virus-transduced HeLa keratinocytes provide high-level, stable, and longterm production of a therapeutic protein in mice. Thes e results must now be extended to human primary keratinocytes.