Ultra-high throughput-based screening for the discovery of antiplatelet drugs affecting receptor dependent calcium signaling dynamics
Abstract Distinct platelet activation patterns are elicited by the tyrosine kinase-linked collagen receptor glycoprotein VI (GPVI) and the G-protein coupled protease-activated receptors (PAR1/4) for thrombin. This is reflected in the different platelet Ca2+ responses induced by the GPVI agonist coll...
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| Format: | Article |
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Nature Portfolio
2024-03-01
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| Series: | Scientific Reports |
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| Online Access: | https://doi.org/10.1038/s41598-024-56799-4 |
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| author | Delia I. Fernández Sara Troitiño Vladimír Sobota Bibian M. E. Tullemans Jinmi Zou Helma van den Hurk Ángel García Saman Honarnejad Marijke J. E. Kuijpers Johan W. M. Heemskerk |
| author_facet | Delia I. Fernández Sara Troitiño Vladimír Sobota Bibian M. E. Tullemans Jinmi Zou Helma van den Hurk Ángel García Saman Honarnejad Marijke J. E. Kuijpers Johan W. M. Heemskerk |
| author_sort | Delia I. Fernández |
| collection | DOAJ |
| description | Abstract Distinct platelet activation patterns are elicited by the tyrosine kinase-linked collagen receptor glycoprotein VI (GPVI) and the G-protein coupled protease-activated receptors (PAR1/4) for thrombin. This is reflected in the different platelet Ca2+ responses induced by the GPVI agonist collagen-related peptide (CRP) and the PAR1/4 agonist thrombin. Using a 96 well-plate assay with human Calcium-6-loaded platelets and a panel of 22 pharmacological inhibitors, we assessed the cytosolic Ca2+ signaling domains of these receptors and developed an automated Ca2+ curve algorithm. The algorithm was used to evaluate an ultra-high throughput (UHT) based screening of 16,635 chemically diverse small molecules with orally active physicochemical properties for effects on platelets stimulated with CRP or thrombin. Stringent agonist-specific selection criteria resulted in the identification of 151 drug-like molecules, of which three hit compounds were further characterized. The dibenzyl formamide derivative ANO61 selectively modulated thrombin-induced Ca2+ responses, whereas the aromatic sulfonyl imidazole AF299 and the phenothiazine ethopropazine affected CRP-induced responses. Platelet functional assays confirmed selectivity of these hits. Ethopropazine retained its inhibitory potential in the presence of plasma, and suppressed collagen-dependent thrombus buildup at arterial shear rate. In conclusion, targeting of platelet Ca2+ signaling dynamics in a screening campaign has the potential of identifying novel platelet-inhibiting molecules. |
| first_indexed | 2024-04-24T23:08:38Z |
| format | Article |
| id | doaj.art-e0e4cce4e1b944baa6e8d33649209870 |
| institution | Directory Open Access Journal |
| issn | 2045-2322 |
| language | English |
| last_indexed | 2024-04-24T23:08:38Z |
| publishDate | 2024-03-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj.art-e0e4cce4e1b944baa6e8d336492098702024-03-17T12:22:38ZengNature PortfolioScientific Reports2045-23222024-03-0114111510.1038/s41598-024-56799-4Ultra-high throughput-based screening for the discovery of antiplatelet drugs affecting receptor dependent calcium signaling dynamicsDelia I. Fernández0Sara Troitiño1Vladimír Sobota2Bibian M. E. Tullemans3Jinmi Zou4Helma van den Hurk5Ángel García6Saman Honarnejad7Marijke J. E. Kuijpers8Johan W. M. Heemskerk9The Department of Biochemistry, CARIM, Maastricht UniversityPlatelet Proteomics Group, CiMUS, Universidade de Santiago de CompostelaIHU-LIRYC, Electrophysiology and Heart Modeling Institute, Fondation Bordeaux UniversitéThe Department of Biochemistry, CARIM, Maastricht UniversityThe Department of Biochemistry, CARIM, Maastricht UniversityPivot Park Screening CentrePlatelet Proteomics Group, CiMUS, Universidade de Santiago de CompostelaPivot Park Screening CentreThe Department of Biochemistry, CARIM, Maastricht UniversityThe Department of Biochemistry, CARIM, Maastricht UniversityAbstract Distinct platelet activation patterns are elicited by the tyrosine kinase-linked collagen receptor glycoprotein VI (GPVI) and the G-protein coupled protease-activated receptors (PAR1/4) for thrombin. This is reflected in the different platelet Ca2+ responses induced by the GPVI agonist collagen-related peptide (CRP) and the PAR1/4 agonist thrombin. Using a 96 well-plate assay with human Calcium-6-loaded platelets and a panel of 22 pharmacological inhibitors, we assessed the cytosolic Ca2+ signaling domains of these receptors and developed an automated Ca2+ curve algorithm. The algorithm was used to evaluate an ultra-high throughput (UHT) based screening of 16,635 chemically diverse small molecules with orally active physicochemical properties for effects on platelets stimulated with CRP or thrombin. Stringent agonist-specific selection criteria resulted in the identification of 151 drug-like molecules, of which three hit compounds were further characterized. The dibenzyl formamide derivative ANO61 selectively modulated thrombin-induced Ca2+ responses, whereas the aromatic sulfonyl imidazole AF299 and the phenothiazine ethopropazine affected CRP-induced responses. Platelet functional assays confirmed selectivity of these hits. Ethopropazine retained its inhibitory potential in the presence of plasma, and suppressed collagen-dependent thrombus buildup at arterial shear rate. In conclusion, targeting of platelet Ca2+ signaling dynamics in a screening campaign has the potential of identifying novel platelet-inhibiting molecules.https://doi.org/10.1038/s41598-024-56799-4CollagenCytosolic calciumGlycoprotein VIPrestwick libraryThrombinThrombosis |
| spellingShingle | Delia I. Fernández Sara Troitiño Vladimír Sobota Bibian M. E. Tullemans Jinmi Zou Helma van den Hurk Ángel García Saman Honarnejad Marijke J. E. Kuijpers Johan W. M. Heemskerk Ultra-high throughput-based screening for the discovery of antiplatelet drugs affecting receptor dependent calcium signaling dynamics Scientific Reports Collagen Cytosolic calcium Glycoprotein VI Prestwick library Thrombin Thrombosis |
| title | Ultra-high throughput-based screening for the discovery of antiplatelet drugs affecting receptor dependent calcium signaling dynamics |
| title_full | Ultra-high throughput-based screening for the discovery of antiplatelet drugs affecting receptor dependent calcium signaling dynamics |
| title_fullStr | Ultra-high throughput-based screening for the discovery of antiplatelet drugs affecting receptor dependent calcium signaling dynamics |
| title_full_unstemmed | Ultra-high throughput-based screening for the discovery of antiplatelet drugs affecting receptor dependent calcium signaling dynamics |
| title_short | Ultra-high throughput-based screening for the discovery of antiplatelet drugs affecting receptor dependent calcium signaling dynamics |
| title_sort | ultra high throughput based screening for the discovery of antiplatelet drugs affecting receptor dependent calcium signaling dynamics |
| topic | Collagen Cytosolic calcium Glycoprotein VI Prestwick library Thrombin Thrombosis |
| url | https://doi.org/10.1038/s41598-024-56799-4 |
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