Lentivirus-mediated gene transfer to human epidermis.
For long-term cutaneous gene therapy, the therapeutic gene must be targeted to stem cells and be stably transmitted to and expressed in descendant cells. Retroviral vectors are highly efficient in gene transfer to human keratinocyte stem cells in culture; however, they cannot transduce quiescent stem cells in vivo. As lentiviral vectors (LVV) transduce non-proliferating cells, their ability to target human epidermal stem cells was evaluated. LVV were highly efficient in gene transfer to clonogenic keratinocytes in vitro. Despite higher transgene DNA content and comparable levels of transgene mRNA, levels of transgene product directed by lentivectors were 3-folds lower than that of retrovectors. When transduced keratinocytes were grafted onto mice, transgene expression persisted for at least 20 wk; however, transgene product was detected primarily in the uppermost layers of epidermis. Inclusion of an element that is known to facilitate nuclear export of intron-less transcripts, resulted in enhanced transgene expression in keratinocytes. In vivo transduction of xenografted human skin with these vectors resulted in efficient gene transfer to epidermal progenitor cells. These results demonstrate stem cell transduction by LVV and point out the utility of using these vectors for direct gene transfer to and sustained expression in human epidermis.
Ghazizadeh
Soosan
S
Department of Oral Biology and Pathology, Stony Brook University, Stony Brook, New York, USA. sg2163@columbia.edu
Katz
Anne B
AB
Harrington
Robin
R
Taichman
Lorne B
LB
D000818:Animals; D002478:Cells, Cultured; D002999:Clone Cells; D004817:Epidermis / Q000502:physiology*; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics; D007763:Lac Operon; D016086:Lentivirus / Q000235:genetics*; D008297:Male; D051379:Mice; D008819:Mice, Nude; D009924:Organ Culture Techniques; D012313:RNA / Q000378:metabolism; D013234:Stem Cells / Q000502:physiology*; D019076:Transgenes; D014183:Transplantation, Heterologous
D049452:Green Fluorescent Proteins; D012313:RNA