Transcriptional reporter assays

For the investigation of androgen effects on target genes cellular in vitro assay systems are implemented. This includes a suitable cell line either expressing the AR or, if the AR is not present, its expression is accomplished by cell transfection techniques. In addition, reporter genes (e.g. luciferase or S-galactosidase) under the control of an androgen responsive promoter such as MMTV (mouse mammary tumour virus), PSA (prostate specific antigen) or probasin employing AREs are transfected into the cells. After androgen treatment of the cells, the measurement of the reporter gene is corresponded with the activity of the AR-dependent transactivation and the potency of the hormone. If in addition comodulators are transfected into the cells, the activity of the AR-dependent reporter signal should either be enhanced or decreased. Importantly, when internal comodulators are expressed in the cells already that activate the AR-dependent reporter signal, an overexpression of these proteins may not modulate the reporter signal. It is therefore essential to know the expression level of the corresponding comodulators in the cells investigated.

A simple however straightforward tool to identify tissue-selective androgens is the comparison of androgen action as measured by reporter gene expression under the control of the same androgen responsible promoter employing different cells, representing the corresponding organs. In this system, the involved cell- or tissue-specific comodulators need not be known in detail. However a possible pitfall may be the alteration of comodulator gene expression in cell lines in comparison to tissue cells in vivo due to changes in the cellular properties, gene regulation and ultimately also AR signalling.

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