An automated microfluidic platform for the screening and characterization of novel hepatitis B virus capsid assembly modulators

Vermes, Tamás; Kielpinski, Mark; Henkel, Thomas; Pericàs, Miquel À.; Alza, Esther; Corcuera, Angelica; Buschmann, Helmut; Goldner, Thomas; Urban, Andreas

doi:10.1039/d1ay01227d

Cited by 5 publications

(4 citation statements)

References 47 publications

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“…This atomic-level fingerprint presents an interesting complement to kinetic methods (e.g. microfluidic screening 54 ) used to distinguish CAM-A from CAM-E compounds.…”

Section: Resultsmentioning

confidence: 99%

Molecular elucidation of drug-induced abnormal assemblies of the hepatitis B virus capsid protein by solid-state NMR

et al. 2023

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Hepatitis B virus (HBV) capsid assembly modulators (CAMs) represent a recent class of anti-HBV antivirals. CAMs disturb proper nucleocapsid assembly, by inducing formation of either aberrant assemblies (CAM-A) or of apparently normal but genome-less empty capsids (CAM-E). Classical structural approaches have revealed the CAM binding sites on the capsid protein (Cp), but conformational information on the CAM-induced off-path aberrant assemblies is lacking. Here we show that solid-state NMR can provide such information, including for wild-type full-length Cp183, and we find that in these assemblies, the asymmetric unit comprises a single Cp molecule rather than the four quasi-equivalent conformers typical for the icosahedral T = 4 symmetry of the normal HBV capsids. Furthermore, while in contrast to truncated Cp149, full-length Cp183 assemblies appear, on the mesoscopic level, unaffected by CAM-A, NMR reveals that on the molecular level, Cp183 assemblies are equally aberrant. Finally, we use a eukaryotic cell-free system to reveal how CAMs modulate capsid-RNA interactions and capsid phosphorylation. Our results establish a structural view on assembly modulation of the HBV capsid, and they provide a rationale for recently observed differences between in-cell versus in vitro capsid assembly modulation.

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“…This atomic-level fingerprint presents an interesting complement to kinetic methods (e.g. microfluidic screening 54 ) used to distinguish CAM-A from CAM-E compounds.…”

Section: Resultsmentioning

confidence: 99%

Molecular elucidation of drug-induced abnormal assemblies of the hepatitis B virus capsid protein by solid-state NMR

et al. 2023

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“…This atomic-level fingerprint presents an interesting complement to kinetic methods (e.g. microfluidic screening 49 ) used to distinguish CAM-A from CAM-E compounds. S5).…”

Section: Nmr Fingerprints Directly Pinpoint Cam-proximal Residuesmentioning

confidence: 99%

Molecular elucidation of drug-induced abnormal assemblies of the Hepatitis B Virus capsid protein by solid-state NMR

Lecoq

Brigandat

Huber

et al. 2022

Preprint

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Hepatitis B virus (HBV) capsid assembly modulators (CAMs) represent a new class of anti-HBV antivirals. CAMs disturb proper nucleocapsid assembly, by inducing formation of either aberrant assemblies (CAM-A) or of apparently normal but genome-less empty capsids (CAM-E). Classical structural approaches have revealed the CAM binding sites on the capsid protein (Cp), but conformational information on the CAM-induced off-path aberrant assemblies is lacking. We show that solid-state NMR can provide such information, including for wild-type full-length Cp183, and we find that in these assemblies, the asymmetric unit comprises a single Cp molecule rather than the four quasi-equivalent conformers typical for the icosahedral T=4 symmetry of the normal HBV capsids. Furthermore, while in contrast to truncated Cp149, fulllength Cp183 assemblies appear, on the mesoscopic level, unaffected by CAM-A, NMR reveals that on the molecular level, Cp183 assemblies are equally aberrant. Finally, we use a eukaryotic cell-free system to reveal how CAMs modulate capsid-RNA interactions and capsid phosphorylation. Our results establish a structural view on assembly modulation of the HBV capsid, and they provide a rationale for recently observed differences between in-cell versus in vitro capsid assembly modulation.

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“…Further inhibitors of HBcAg have also issued with promising results both in vivo and in vitro. These are included Yhhu6669 [22], pyrazolyl-thiazole (PT), glyoxamide-pyrrolamide (GPA), and dibenzo-thiazepin-2-one (DBT) [23], JNJ-64530440 [24], Compound A and B [25], Ciclopirox [26], NVR 3-778 [27], HEC72702 [28], JNJ-827 and JNJ-890 [29], Cetylpyridinium chloride (CPC) [30], Bay 41-4109 [18,31], isothiafludine (NZ-4) [5], and other derivatives [3,32]. These interests in developing direct HBcAg-binders show the novelty of such approaches, and repurposing drugs with same or enhanced direct-action on HBV core protein still holds much promises.…”

Section: Introductionmentioning

confidence: 99%

Computational-Aided Reproposing Drug(s) and Discovery for Potential Antivirals Targeting Hepatitis B Virus Capsid Protein

Mohebbi,

Naderi,

Sharifian

et al. 2023

Preprint

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Background: Chronic Hepatitis B Virus (HBV) infection is a global health concern, associated with severe liver diseases, necessitating ongoing research on novel drug candidates. This study aims to identify potential drug candidates targeting HBV core protein (HBcAg) and disrupting capsid assembly, a critical step in the virus's life cycle. Methods: HBcAg in complex with HBV inhibitors were obtained from the Protein Data Bank (PDB). CavityPlus server was used for analysis of druggable cavity. Structure-based pharmacophores were extracted from identified cavities, and potential allosteric ligand binding sites were assessed using CavPharmer, CorrSite, and CovCys. LigandScout was employed for ligand-based pharmacophore screening against an FDA-approved library. The ZINC database was screened with features extracted from CavPharmer. Molecular docking studies were conducted using Autodock Vina. Lead compounds were selected based on docking scores, binding modes, and interactions within the druggable cavity. Results: Strong druggable pockets were found for Ciclopirox, while Compound 24, NVR10-001E2, and others showed medium to weak pockets. Ligand-based pharmacophores varied in size and complexity. Screening revealed potential hits matching these pharmacophores, including Ciclopirox olamine, Voriconazole, Enasidenib, and Statins. A large compound database search yielded additional hits like ZINC86859997 and ZINC63280172. Docking analyses confirmed these hits' potential, highlighting their interactions with critical HBc protein residues, offering promising leads for hepatitis B drug development. Conclusions: Voriconazole, Enasidenib, and Lovastatin have shown promises. These hits displayed favorable interactions with crucial HBc protein residues, indicating their potential as lead compounds The mechanism of action of statins with anti-HBV activities also highlighted. This comprehensive approach offers valuable insights into targeting HBc protein for antiviral drug discovery.

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An automated microfluidic platform for the screening and characterization of novel hepatitis B virus capsid assembly modulators

Abstract: A fully automated microfluidic system was developed to screen for novel anti-HBV capsid assembly modulators. High-resolution dose–response curves were generated using convection-dominated Taylor–Aris dispersion of the screening compounds.

Cited by 5 publications

References 47 publications

Molecular elucidation of drug-induced abnormal assemblies of the hepatitis B virus capsid protein by solid-state NMR

Molecular elucidation of drug-induced abnormal assemblies of the hepatitis B virus capsid protein by solid-state NMR

Molecular elucidation of drug-induced abnormal assemblies of the Hepatitis B Virus capsid protein by solid-state NMR

Computational-Aided Reproposing Drug(s) and Discovery for Potential Antivirals Targeting Hepatitis B Virus Capsid Protein

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