Evaluation of Potential Human Sodium Taurocholate Co‐transporting Polypeptide Inhibitors

Abstract

ObjectiveChronic hepatitis B virus (HBV) infection is responsible for 50% of the cases of hepatocellular carcinoma. As currently available treatment is not able to eradicate the virus, new therapeutic approaches are highly needed. Sodium taurocholate co‐transporting polypeptide (NTCP), a bile acid transporter at the sinusoidal side of hepatocytes, has been identified as receptor for HBV entry into the cell. Since HBV shares a common docking site with NTCP substrates, inhibition of NTCP‐mediated transport is an attractive strategy to inhibit HBV entry and has already been proven successful (e.g. Myrcludex). The aim of this research was to identify novel NTCP inhibitors in vitro and develop a chemometric model that allows virtual screening.MethodsWild‐type and NTCP‐transfected Chinese hamster ovary cells were cultured and seeded in black 96‐well plates with transparent bottom. They were preincubated for 15 minutes with a potential inhibitor and the reaction was initiated by adding the fluorescent NTCP substrate tauro‐nor‐THCA‐24‐DBD. The final concentration of compound and substrate was 3 μM. Uptake was determined using fluorescence spectroscopy (excitation 440 nm and emission 580 nm). Potency of the hits was determined by incubating at least seven concentrations of each hit in presence of 3 μM substrate. IC50 values were estimated by fitting the sigmoid Emax model in R version 3.5.1. The inhibitors were subsequently docked into the taurocholate binding site of NTCP and a chemometric model was developed using rapid overlay of chemical structures.ResultsWe assessed more than 2500 compounds for their ability to inhibit NTCP. Forty‐one compounds were able to reduce the uptake to less than 50% (of the control, i.e. only substrate present) and were therefore classified as hits. The pIC50 of 30 compounds could be determined and ranged from 3.6 to 6.8. Several rings form the core of the 3D overlay model, surrounded by hydrophobic groups as well as hydrogen bond donor and acceptor groups. While the latter appear to be exposed to the solvent, many of the hydrophobic groups are buried inside the taurocholate binding site.ConclusionsWe identified NTCP inhibitors in vitro and developed a chemometric model that can be used for virtual screening. This model is currently being applied to the ChEMBL database for in silico identification of NTCP inhibitors. Future work also includes in vitro confirmation of the in silico hits.Support or Funding InformationThis research was supported by a grant of the Research Foundation Flanders (FWO grant G089515N).