Systemic and localized reversible regulation of transgene expression by tetracycline with tetR-mediated transcription repression switch.
BACKGROUND
We recently developed a new tetracycline-inducible gene switch employing the tetracycline operator-containing hCMV major immediate-early promoter and the tetracycline repressor, tetR, rather than the previously used tetR-mammalian cell transcription factor fusion derivatives.


METHODS
The present study demonstrates that this tetR-mediated transcription repression system can function as a powerful gene switch for On-and-Off regulation of therapeutic gene expression in ex vivo gene transfer protocols. Firstly, for achieving regulated gene expression in a localized tissue environment, R11/OEGF cells, a stable line that expresses hEGF under the control of the tetR-mediated transcription repression switch, were transplanted into porcine full-thickness wounds enclosed by wound chambers.


RESULTS
By topically applying tetracycline in wound chambers at various concentrations or at different time points post-transplantation, the levels and timing of hEGF expression in transplanted wounds could be reversibly regulated by tetracycline. Over 3000-fold induction in hEGF expression was achieved in the local wound microenvironment. Secondly, R11/OEGF cells were intramuscularly injected into NCr outbread nude mice to test the efficacy of intermittent systemic gene delivery of a soluble peptide(s).


CONCLUSIONS
Basal circulating hEGF was undetectable and induced up to at least 1,500-fold after administration of tetracycline. Furthermore, the timing and duration of hEGF expression could be finely adjusted by the presence or the absence of tetracycline in the drinking water.
Yao
Feng
F
Laboratory of Tissue Repair and Gene Transfer, Plastic Surgery, Brigham and Women's Hospital, and Department of Surgery, Harvard Medical School, Boston, Massachusetts 02115, USA. FYao@RICS.BWH.Harvard.Edu
Pomahac
Bohdan
B
Visovatti
Scott
S
Chen
Michael
M
Johnson
Starck
S
Augustinova
Hanka
H
Svensjo
Tor
T
Eriksson
Elof
E
D000818:Animals; D000900:Anti-Bacterial Agents; D045744:Cell Line, Tumor; D003587:Cytomegalovirus / Q000235:genetics*; D004815:Epidermal Growth Factor / Q000235:genetics; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D018014:Gene Transfer Techniques; D006801:Humans; D008297:Male; D051379:Mice; D008819:Mice, Nude; D012516:Osteosarcoma; D011401:Promoter Regions, Genetic / Q000235:genetics; D012867:Skin / Q000293:injuries; D034421:Sus scrofa; D013752:Tetracycline; D019076:Transgenes / Q000235:genetics*; D014947:Wounds and Injuries / Q000503:physiopathology
D000900:Anti-Bacterial Agents; D004815:Epidermal Growth Factor; D013752:Tetracycline