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BSEP inhibition

The bile salt export pump, BSEP (ABCB11), is located at the canalicular (apical) membrane of hepatocytes and is involved in the secretion of bilirubin/bile salts from the liver. Since bile formation and secretion are key functions of the liver, any persistent accumulation of bilirubin/bile salts in the liver through inhibition of BSEP activity by drugs can lead to drug-induced liver injury (DILI).

Sygnature’s human BSEP or rat bsep inhibition assay measures the inhibition of uptake of taurocholic acid (TCA) into inverted vesicles containing the BSEP or bsep transporter which have been extracted from transformed Sf9 insect cells expressing the associated protein. TCA and test compound are incubated with the vesicles in the presence of ATP. AMP is included to correct for passive (non-active) transport and vehicle incubations are included as negative controls.

Ketoconazole, troglitazone, cyclosporin A and/or glibenclamide have been used in the assay validation phase and are included as positive control inhibitors.

 

Protocol

Compound requirements 10 mM DMSO, 50µL
Substrate 5 μM taurocholic acid (TCA) + [H3]-TCA
Test Article Concentrations 8-point IC50 in duplicate
Incubation Time 30 min at 37°C
Test System ATP-dependent uptake into inside-out human or rat BSEP/bsep vesicles (other species available on request)
Analysis Method Radiometric: Liquid Scintillation Counting
Controls 5 mM AMP (to correct for non-active uptake)
Ketoconazole, troglitazone, cyclosporine A and/or glibenclamide (positive control compounds)
Data Delivery IC50

 

Results

Figure 1 Comparison between Sygnature Discovery and literature published (1) IC50 (µM) values for a number of known human BSEP inhibitors. Sygnature Discovery data shown are the mean of 6 independent experiments.

 

Figure 2 Inhibition of TCA uptake into inside-out vesicles expressing human BSEP transporter by a number of human BSEP inhibitors.


About Us

The DMPK & Physical Sciences department at Sygnature Discovery is dedicated to understanding and optimising the absorption, distribution, metabolism and excretion of drug candidates by working in close partnership with clients and other departments within Sygnature to provide successful optimisation strategies.

We have extensive know-how and expertise to provide well validated, state-of-the-art assays and a comprehensive applied consultancy service for interpretation of the in vitro ADME and in vivo PK data.

Our corporate vision is to accelerate the discovery of new medicines, from the laboratory into development to treat patients.

Our DMPK mission is to deliver tailored DMPK expertise through innovation, quality and commitment.

 

References

 

  • Dawson, et al. 2012. Drug Metab Dispos 40(1):130-8

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