Discovery and Functional Validation of EP3 Receptor Ligands with Therapeutic Potential in Cardiovascular Disease
Abstract
1. Introduction
2. Results
2.1. Blind Docking Revealed Two Distinct Potential Binding Sites on the EP3 Receptor
2.2. Prediction of Ligands by Virtual Screening
2.2.1. Selection of Libraries
2.2.2. Ligand-Based Virtual Screening with LigandScout
2.2.3. Structure-Based Virtual Screening with LigandScout
2.2.4. Structure-Based Virtual Screening with AutoDock Vina
2.3. Consensus, QikProp, and Compound Selection
2.4. Cell Viability Assay
2.5. Cell Migration Assay
2.6. Platelet Activation and Leukocyte–Platelet Aggregates Measurement Assay
3. Discussion
3.1. Ligand Identification via Computational Methods
3.2. Biological Evaluation of Candidate Compounds
4. Materials and Methods
4.1. Blind Docking
4.2. Virtual Screenings
- Using LigandScout, ligand-based screening (using the pharmacophoric model of misoprostol) and structure-based screening (using interactions between misoprostol and EP3) were performed.
- Using AutoDock Vina, structure-based screening was performed using restricted docking (scoring different poses of the ligand in the chosen region and comparing the best scores of all molecules).
4.2.1. LigandScout
4.2.2. AutoDock Vina
4.3. Consensus and Other Metrics
4.4. Cell Culture and Drugs
4.5. Cell Viability Assay
4.6. Cell Migration Assay (Wound Healing Assay)
4.7. Ex Vivo Assessment of Platelet Activation and Platelet–Leukocyte Aggregates (PLAs)
4.8. Data Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
ADME/Tox | Absorption, Distribution, Metabolism, Excretion / Toxicity |
GUI | Graphical User Interface |
TUCA | Taurocholic Acid |
CVD | Cardiovascular Disease |
PGE2 | Prostaglandin E2 |
NOGA | Nordihydroguaiaretic acid (Masoprocol) |
MTT | Thiazolyl Blue Tetrazolium Bromide |
PLA | Leukocyte–Platelet Aggregate |
EDTA | Ethylenediaminetetraacetic Acid |
PGI2 | Prostaglandin I2 |
HMG-CoA | Hydroxymethylglutaryl-CoA |
FBS | Fetal Bovine Serum |
DMSO | Dimethyl Sulfoxide |
PBS | Phosphate-Buffered Saline |
PMN | Polymorphonuclear Leukocytes |
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Code | AD | SB-0 | SB-1 | SB-2 | LB-1 | LB-2 | Name |
---|---|---|---|---|---|---|---|
FDB023044 | −8.12 | 0.98 | 0.98 | 0.98 | 0.86 | 0.86 | Prostaglandin F1a |
FDB022853 | −8.00 | 0.98 | 0.98 | 0.98 | 0.86 | 0.86 | Prostaglandin F3a |
FDB023346 | −7.90 | 0.98 | 0.98 | 0.98 | 0.87 | 0.87 | 6-KetoPG 1 E1 |
FDB022647 | −8.27 | 0.97 | 0.97 | 0.97 | 0.76 | Prostaglandin F2b | |
FDB022851 | −7.99 | 0.98 | 0.98 | 0.98 | 0.76 | 8-isoPG 1 F2b | |
DB00770 | −7.76 | 0.89 | 0.97 | 0.87 | 0.87 | Alprostadil | |
FDB022625 | −7.71 | 0.89 | 0.97 | 0.87 | 0.87 | Prostaglandin E1 | |
DB12708 | −9.34 | 0.89 | 0.78 | Sulprostone | |||
DB01240 | −8.14 | 0.97 | 0.97 | 0.97 | 0.77 | Epoprostenol | |
FDB022560 | −8.06 | 0.97 | 0.97 | 0.97 | 0.77 | Prostaglandin I2 | |
DB11507 | −9.22 | 0.81 | 0.87 | 0.87 | Cloprostenol | ||
DB00179 | −8.76 | 0.86 | 0.86 | Masoprocol (NOGA) | |||
FDB022448 | −8.11 | 0.98 | 0.98 | 0.76 | Prostaglandin F2a | ||
DB14544 | −8.66 | 0.86 | 0.86 | HC 2 valerate | |||
DB01088 | −7.99 | 0.97 | 0.76 | Iloprost | |||
DB06555 | −13.08 | Siramesine | |||||
FDB011293 | −13.00 | Sesamolinol | |||||
DB08512 | −12.65 | ||||||
DB07538 | −12.55 | ||||||
DB03583 | −12.48 | ||||||
DB00175 | −8.25 | 0.81 | Pravastatin | ||||
DB08964 | −8.22 | 0.86 | 0.96 | Gemeprost | |||
DB00929 | −8.19 | 0.87 | 0.87 | Misoprostol | |||
DB07528 | −12.42 | ||||||
DB06925 | −12.36 | ||||||
DB15345 | −12.26 | GSK-945237 | |||||
DB07545 | −12.17 | ||||||
FDB000425 | −12.15 | ||||||
FDB023593 | −7.36 | 0.81 | Pravastatin | ||||
FDB023130 | −8.22 | 0.90 | 0.90 | Thromboxane B2 | |||
DB12043 | −9.55 | 0.86 | 0.86 | Sepetaprost | |||
DB04348 | −6.53 | 0.81 | Taurocholic acid (TUCA) |
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Alonso-Fernández, J.-R.; Montoro-García, S.; Cruz, A.-F.; Ponce-Valencia, A.; Carmena-Bargueño, M.; Pérez-Sánchez, H. Discovery and Functional Validation of EP3 Receptor Ligands with Therapeutic Potential in Cardiovascular Disease. Int. J. Mol. Sci. 2025, 26, 4879. https://doi.org/10.3390/ijms26104879
Alonso-Fernández J-R, Montoro-García S, Cruz A-F, Ponce-Valencia A, Carmena-Bargueño M, Pérez-Sánchez H. Discovery and Functional Validation of EP3 Receptor Ligands with Therapeutic Potential in Cardiovascular Disease. International Journal of Molecular Sciences. 2025; 26(10):4879. https://doi.org/10.3390/ijms26104879
Chicago/Turabian StyleAlonso-Fernández, Jorge-Ricardo, Silvia Montoro-García, Andreia-Filipa Cruz, Alicia Ponce-Valencia, Miguel Carmena-Bargueño, and Horacio Pérez-Sánchez. 2025. "Discovery and Functional Validation of EP3 Receptor Ligands with Therapeutic Potential in Cardiovascular Disease" International Journal of Molecular Sciences 26, no. 10: 4879. https://doi.org/10.3390/ijms26104879
APA StyleAlonso-Fernández, J.-R., Montoro-García, S., Cruz, A.-F., Ponce-Valencia, A., Carmena-Bargueño, M., & Pérez-Sánchez, H. (2025). Discovery and Functional Validation of EP3 Receptor Ligands with Therapeutic Potential in Cardiovascular Disease. International Journal of Molecular Sciences, 26(10), 4879. https://doi.org/10.3390/ijms26104879