Evaluation of Tet-on system to avoid transgene down-regulation in ex vivo gene transfer to the CNS.

摘要: Ex vivo gene transfer to the CNS has so far been hampered by instability of transgene expression. To avoid phenomenon down-regulation, we have employed strong, constitutive promoters and compared this expression system with inducible Tet incorporated in a single plasmid vector or lentiviral vectors. Plasmid-based directed constitutive, human ubiquitin promoter, UbC, was stable transfected HiB5 cells vitro comparable strength CMV promoter. However, after transplantation UbC clones rat striatum, silencing occurred most soon implantation cells. The Tet-on elements were generated. Inducible displayed varying basal activity, which could not be ascribed an effect cis-elements vector, but rather due, at least part, intrinsic activity minimal Basal blocked majority expressing transrepressor tTS. Fully induced levels promoters. Similar down-regulated grafting striatum. Lentiviral vectors can direct long-term take advantage quality two followed transduction Hib5 Interestingly, all established showed very similar patterns tight regulatability that apparently independent copy number integration site. Nevertheless, shortly These results confirm general down-regulation. Moreover, suggest considerable advantages offered for delivery cannot necessarily transferred directly ex delivery. This emphasizes need alternative strategies transfer.