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Controlling genes via tetracyclines:
Is the approach applicable in doping?
Tetracycline regulatable transcription control systems are most widely and successfullyused tools für controlling gene expression in many eukaryotic cells and organisms (1, 2).
The main reasons fo their broad applicability (3) are the exceptional properties of the tetrepressor/operator/inducer system. Particularly the fact that inducers like tetracycline andmany of its derivatives are widely used antibiotics with a superb specificity provides awealth of information on the chemistry and pharmacology of this class of substances.
Accordingly, a panel of non-toxic and weIl characterized effector compounds is available.
More recently, our laboratory has developed a family of tetracycline controlled transcrip-tional activators and silencers with new specificities which can be used individually or incombination. Together, these elements enhance our capabilities of controlling one or sev-eral target genes upon transfer into cells or whole o!ganisms (4).
Transfer of the Tet regulatory systems to mammals including mice, rats and non-humanprimates hag allowed to control gene expression tightly and reversibly over long periods oftime. Using proper promoters driving tetracycline controlled transactivators, Tet regulationmay be restricted to single cell types such as hepatocytes or muscle cells (5). Moreover, thekinetics of gene activation and inactivation are fast (6). Results obtained with rodents sug-gest that the Tet systems may also be suitable für applications in gene therapy. Thus - asshown in the mouse model - the activity of therapeutic genes may be adjusted to relevantlevels which can be modulated over long periods of time (7, 8). In this context, the panelof tetracycline controlled transcriptional regulators available and the choice of differenteffector substances appears useful for future developments.
Approaches being developed für gene therapy may well qualify für misuse in the field ofdoping. The goal to control quantitatively and reversibly EPO in respective patients mayappear particularly attractive and results obtained in animal models suggest that indeedsuch approaches hold promise (7). As the transfer of such controlled expression units mayoccur at the level of DNA, e.g. into skeletal muscle via single injections, it may be difficultto develop diagnostic means which unequivocally identify respective mistige.
Controlling Genes via Tetracyclines: Is the approach applicable in Doping? Literature
1. AURISICCHIO, L.; BUJARD, H.; HILLEN, W. et al.: Regulated and prolonged expression of mIFN(alpha) in immunocompetent mice. Gene Ther 24 (2002) 8, 1817-1825 2. BARON, U.; BUJARD, H.: Tet repressor-based system for regulated gene expression in eukaryotic cells: principles and advances. Methods Enzymol (2000) 327, 401-421 3. BERGER, S.; BUJARD, H.: In: Handbook of Exp. Pharmacology. Berlin, Heidelberg, 4. GOSSEN, M.; BUJARD, H.: Tight control of gene expression in mammalian cells by tetracycline-responsive promoters. Proc Natl Acad Sci 89 (1992) 12, 5547-5551 5. GOSSEN, M.; FREUNDLIEB, S.; BENDER, G.; MÜLLER, G.; HILLEN W.; BUJARD, H.: Transcriptional activation by tetracyclines in mammalian cells. Science(1995) 268 (5218), 1766-1769 6. GOSSEN, M.; BUJARD, H.: Studying gene function in eukaryotes by conditional gene inactivation. Ann Rev Genet (2002) 36, 153-173 7. LAMARTINA, S. SILVI, L.; ROSCILLI, G. ET AL. et al.: Construction of an rtTA2(s)-m2/tts(kid)-based transcription regulatory switch that displays no basalactivity, good inducibility, and high responsiveness to doxycycline in mice and Non-Human primates. Mol Ther 7 (2003) 2, 271-280 8. SCHÖNIG, K.; BUJARD, H.: Generating conditional mouse mutants via tetracycline- controlled gene expression. Methods Mol Biol (2003) 209, 69-104 Prof. Dr. Hermann BujardZentrum für Molekulare Biologie der Universität Heidelberg(ZMBH)Im Neuenheimer Feld 282D-69120 Heidelberg

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