Development of a mass spectrometric screening assay for hepatitis B virus entry inhibitors

Goh, Byoungsook; Choi, Jieun; Kang, Jung‐Ah; Park, Sung‐Gyoo; Seo, Jiwon; Kim, Tae‐Young

doi:10.1016/j.jpba.2019.112959

Cited by 5 publications

(3 citation statements)

References 30 publications

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“…Blocking this process is an attractive means of controlling both HBV and HDV, and myrcludex B 9 , a synthetic analogue of the preS1 lipopeptide has been approved to treat HDV in Europe and Russia. In vitro screening assays have been used to identify other chemical entities that prevent HBV entry into hepatocytes 10 , 11 . Similar to myrcludex B, these compounds tend to inhibit the native function of the receptor in the uptake of bile salts.…”

Section: Mainmentioning

confidence: 99%

Structural insights into the HBV receptor and bile acid transporter NTCP

Park

Iwamoto

Yun

et al. 2022

Nature

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Around 250 million people are infected with hepatitis B virus (HBV) worldwide1, and 15 million may also carry the satellite virus hepatitis D virus (HDV), which confers even greater risk of severe liver disease2. The HBV receptor has been identified as sodium taurocholate co-transporting polypeptide (NTCP), which interacts directly with the first 48 amino acid residues of the N-myristoylated N-terminal preS1 domain of the viral large protein3. Despite the pressing need for therapeutic agents to counter HBV, the structure of NTCP remains unsolved. This 349-residue protein is closely related to human apical sodium-dependent bile acid transporter (ASBT), another member of the solute carrier family SLC10. Crystal structures have been reported of similar bile acid transporters from bacteria4,5, and these models are believed to resemble closely both NTCP and ASBT. Here we have used cryo-electron microscopy to solve the structure of NTCP bound to an antibody, clearly showing that the transporter has no equivalent of the first transmembrane helix found in other SLC10 proteins, and that the N terminus is exposed on the extracellular face. Comparison of our structure with those of related proteins indicates a common mechanism of bile acid transport, but the NTCP structure displays an additional pocket formed by residues that are known to interact with preS1, presenting new opportunities for structure-based drug design.

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Section: Mainmentioning

confidence: 99%

Structural insights into the HBV receptor and bile acid transporter NTCP

Park

Iwamoto

Yun

et al. 2022

Nature

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“…On the basis of the therapeutically used effects of bulevirtide, a promising second strategy for the development of HBV/HDV entry inhibitors targeting NTCP would be to (I) investigate small molecules with oral bioavailability which (II) block virus binding to NTCP, but (III) without tackling its physiologically important bile salt transporter activity at relevant therapeutic drug concentrations. Currently, several studies identified novel chemical entry inhibitors for HBV and HDV by screening for bile salt transport inhibitors 20 or by screening for inhibitors of myr-preS1 2-48 lipopeptide attachment and/or in vitro HBV/HDV infection 21 in appropriate cell culture models overexpressing NTCP. However, none of these studies addressed and fulfilled all of the criteria outlined above.…”

Section: Introductionmentioning

confidence: 99%

Selective hepatitis B and D virus entry inhibitors from the group of pentacyclic lupane-type betulin-derived triterpenoids

Kirstgen

Lowjaga

Müller

et al. 2020

Sci Rep

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Current treatment options against hepatitis B and D virus (HBV/HDV) infections have only limited curative effects. Identification of Na+/taurocholate co-transporting polypeptide (NTCP) as the high-affinity hepatic receptor for both viruses in 2012 enables target-based development of HBV/HDV cell-entry inhibitors. Many studies already identified appropriate NTCP inhibitors. However, most of them interfere with NTCP’s physiological function as a hepatic bile acid transporter. To overcome this drawback, the present study aimed to find compounds that specifically block HBV/HDV binding to NTCP without affecting its transporter function. A novel assay was conceptualized to screen for both in parallel; virus binding to NTCP (measured via binding of a preS1-derived peptide of the large HBV/HDV envelope protein) and bile acid transport via NTCP. Hits were subsequently validated by in vitro HDV infection studies using NTCP-HepG2 cells. Derivatives of the birch-derived pentacyclic lupane-type triterpenoid betulin revealed clear NTCP inhibitory potency and selectivity for the virus receptor function of NTCP. Best performing compounds in both aspects were 2, 6, 19, and 25. In conclusion, betulin derivatives show clear structure–activity relationships for potent and selective inhibition of the HBV/HDV virus receptor function of NTCP without tackling its physiological bile acid transport function and therefore are promising drug candidates.

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“…Based on that, a synthetic analogue of this lipopeptide (Hepcludex ® ) has been developed and was recently approved as first HDV entry inhibitor interacting with NTCP [ 10 ]. Furthermore, numerous studies identified novel chemical entry inhibitors for HBV and HDV by screening for bile salt transport inhibitors [ 11 ] or by screening for inhibitors of myr-preS1 2–48 lipopeptide attachment and/or in vitro HBV/HDV infection [ 12 ] in appropriate hepatoma cell culture models overexpressing NTCP. However, to date, none of these small molecules have been able to enter the clinical development phase so far.…”

Section: Introductionmentioning

confidence: 99%

Identification of Novel HBV/HDV Entry Inhibitors by Pharmacophore- and QSAR-Guided Virtual Screening

Kirstgen

Müller

Lowjaga

et al. 2021

Viruses

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The hepatic bile acid transporter Na+/taurocholate co-transporting polypeptide (NTCP) was identified in 2012 as the high-affinity hepatic receptor for the hepatitis B and D viruses (HBV/HDV). Since then, this carrier has emerged as promising drug target for HBV/HDV virus entry inhibitors, but the synthetic peptide Hepcludex® of high molecular weight is the only approved HDV entry inhibitor so far. The present study aimed to identify small molecules as novel NTCP inhibitors with anti-viral activity. A ligand-based bioinformatic approach was used to generate and validate appropriate pharmacophore and QSAR (quantitative structure–activity relationship) models. Half-maximal inhibitory concentrations (IC50) for binding inhibition of the HBV/HDV-derived preS1 peptide (as surrogate parameter for virus binding to NTCP) were determined in NTCP-expressing HEK293 cells for 150 compounds of different chemical classes. IC50 values ranged from 2 µM up to >1000 µM. The generated pharmacophore and QSAR models were used for virtual screening of drug-like chemicals from the ZINC15 database (~11 million compounds). The 20 best-performing compounds were then experimentally tested for preS1-peptide binding inhibition in NTCP-HEK293 cells. Among them, four compounds were active and revealed experimental IC50 values for preS1-peptide binding inhibition of 9, 19, 20, and 35 µM, which were comparable to the QSAR-based predictions. All these compounds also significantly inhibited in vitro HDV infection of NTCP-HepG2 cells, without showing any cytotoxicity. The best-performing compound in all assays was ZINC000253533654. In conclusion, the present study demonstrates that virtual compound screening based on NTCP-specific pharmacophore and QSAR models can predict novel active hit compounds for the development of HBV/HDV entry inhibitors.

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Development of a mass spectrometric screening assay for hepatitis B virus entry inhibitors

Cited by 5 publications

References 30 publications

Structural insights into the HBV receptor and bile acid transporter NTCP

Structural insights into the HBV receptor and bile acid transporter NTCP

Selective hepatitis B and D virus entry inhibitors from the group of pentacyclic lupane-type betulin-derived triterpenoids

Identification of Novel HBV/HDV Entry Inhibitors by Pharmacophore- and QSAR-Guided Virtual Screening

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