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NTCP (Na+-Taurocholate Cotransporting Polypeptide)

The Na+-Taurocholate Cotransporting Polypeptide (NTCP) transporter is involved in the disposition and recirculation of bile acids; bile salts secreted into the bile are often re-absorbed into the portal circulation and back into the liver via NTCP. Transport via this mechanism is sodium dependent and electrogenic. Regulation of NTCP copes with changes in bile salt load and expression levels decrease when challenged by a high concentration of bile salts. Although it is not a promiscuous drug transporter, NTCP is a transporter of the widely prescribed, hepatically acting, HMG-CoA reductase inhibitors, notably statins, hence increasing the risk of drug-drug interactions (DDI).

Sygnature’s NTCP inhibition assay utilises readily plated CHO-cells expressing the transporter. The endogenous substrate, Taurocholic acid (TCA) is incubated with a range of test compound concentrations. By measuring the uptake of TCA, we are able to measure an IC50 value.

The taurine conjugate of chenodeoxycholate, TCDC, and inhibitor of TCA uptake by NTCP is used as positive control. The experiment is performed in Na-supplemented buffer and sodium-depleted buffer is used to correct for passive diffusion and non-NTCP mediated transport.

Protocol

 

Compound requirements 10 mM DMSO, 50µL
Substrate 0.5 μM taurocholic acid (TCA) & 25kBq Radchem probe
Test Article Concentrations 9-point IC50 in duplicate
Incubation Time 5 min at 37°C CO2 incubator
Test System Sodium-dependent uptake into NTCP transfected CHO cells (other species available on request)
Analysis Method Radiometric Liquid Scintillation Counting
Controls 20mM TCDC (positive control compounds)

Na-negative buffer can be used for correction of passive transport.

Data Delivery IC50

 

Results

Figure 1. Inhibition of TCA uptake into CHO cells expressing the NTCP transporter by test compounds and TCDC.

 

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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.

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