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Structure-based discovery and in vitro validation of inhibitors of chloride intracellular channel 4 protein

Olotu, Fisayo, Medina-Carmona, Encarnacion, Serrano Sanchez, Angela, Ossa, Felipe, El-Hamdaoui, Abdelaziz, Bishop, Özlem Tastan, Ortega-Roldan, Jose L., Abdul-Salam, Vahitha B. (2023) Structure-based discovery and in vitro validation of inhibitors of chloride intracellular channel 4 protein. Computational and Structural Biotechnology Journal, 21 . pp. 688-701. ISSN 2001-0370. (doi:10.1016/j.csbj.2022.12.040) (KAR id:99877)

Abstract

The use of computer-aided methods have continued to propel accelerated drug discovery across various disease models, interestingly allowing the specific inhibition of pathogenic targets. Chloride Intracellular Channel Protein 4 (CLIC4) is a novel class of intracellular ion channel highly implicated in tumor and vascular biology. It regulates cell proliferation, apoptosis and angiogenesis; and is involved in multiple pathologic signaling pathways. Absence of specific inhibitors however impedes its advancement to translational research. Here, we integrate structural bioinformatics and experimental research approaches for the discovery and validation of small-molecule inhibitors of CLIC4. High-affinity allosteric binders were identified from a library of 1615 Food and Drug Administration (FDA)-approved drugs via a high-performance computing-powered blind-docking approach, resulting in the selection of amphotericin B and rapamycin. NMR assays confirmed the binding and conformational disruptive effects of both drugs while they also reversed stress-induced membrane translocation of CLIC4 and inhibited endothelial cell migration. Structural and dynamics simulation studies further revealed that the inhibitory mechanisms of these compounds were hinged on the allosteric modulation of the catalytic glutathione (GSH)-like site loop and the extended catalytic β loop which may elicit interference with the catalytic activities of CLIC4. Structure-based insights from this study provide the basis for the selective targeting of CLIC4 to treat the associated pathologies.

Item Type: Article
DOI/Identification number: 10.1016/j.csbj.2022.12.040
Uncontrolled keywords: HUVEC, Human umbilical vein endothelial cells, IKKβ, Inhibitor of nuclear kappa-B-kinase subunit beta, Chloride intracellular channel protein 4, Nuclear magnetic resonance, MKK6, Mitogen-activated protein kinase kinase-6, PAH, Pulmonary arterial hypertension, p38, Mitogen activated protein kinases, HSQC, Heteronuclear single quantum coherence spectroscopy, DAPI, 4′,6-diamidino-2-phenylindole, Allosteric inhibition, DOPE, Discrete optimized protein energy, TIP3P, Transferable intermolecular potential 3 P, GSH-like catalytic site, NPT, The constant-temperature, constant-pressure ensemble, GPU, Graphics Processing Unit, HEPES, (4-(2-hydroxyethyl)− 1-piperazineethanesulfonic acid;, JNK, c-Jun N-terminal kinase, SASA, Solvent accessible surface area, Computational high-throughput screening, TEV, Tobacco etch virus, DMSO, Dimethyl sulfoxide, NF-κB, Nuclear factor kappa-light-chain-enhancer of activated B cells, NaCL, Sodium chloride, MOI, Multiplicity of infection, SEK1, Dual specificity mitogen-activated protein kinase kinase 4, DIDS, 4,4′-Diisothiocyano-2,2′-stilbenedisulfonic acid, CLIC, Chloride intracellular channel protein, Bcl-2, B-cell lymphoma 2, CDK, Cyclin-dependent kinases, Ad, Adenovirus, TROSY, Transverse relaxation optimized spectroscopy, Bcl-xL, B-cell lymphoma-extra large, Bad, BCL2 associated agonist of cell death, VEGF, Vascular endothelial growth factor, UCSF, University of California, San Francisco, RAPA, Rapamycin, AMPhB, Amphotericin B, GST, glutathione S-transferases, NMR, Nuclear magnetic resonance, Smad, Suppressor of Mothers against Decapentaplegic, Structure-based drug discovery, A9C, 9-Anthracenecarboxylic acid, GUI, Graphical User Interface, HIF, Hypoxia-inducible factor
Subjects: Q Science > QH Natural history > QH301 Biology
Q Science > QH Natural history > QH581.2 Cell Biology
Divisions: Divisions > Division of Natural Sciences > Biosciences
Funders: University of Kent (https://ror.org/00xkeyj56)
SWORD Depositor: JISC Publications Router
Depositing User: JISC Publications Router
Date Deposited: 24 Feb 2023 15:49 UTC
Last Modified: 27 Feb 2023 12:45 UTC
Resource URI: https://kar.kent.ac.uk/id/eprint/99877 (The current URI for this page, for reference purposes)

University of Kent Author Information

Serrano Sanchez, Angela.

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Ortega-Roldan, Jose L..

Creator's ORCID: https://orcid.org/0000-0002-6316-4390
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