{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T13:55:29Z","timestamp":1790344529200,"version":"4.1.0"},"reference-count":40,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2022,8,3]],"date-time":"2022-08-03T00:00:00Z","timestamp":1659484800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Deanship of Scientific Research, King Saud University"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Antioxidants"],"abstract":"<jats:p>Centaurea calcitrapa has been intensively utilized in ethnomedicinal practices as a natural therapeutic recipe to cure various ailments. The current study aimed to chemically characterize ethanolic extract of C. calcitrapa (EECC) aerial parts (leaves and shoots) by use of gas chromatography-mass spectrometry analyses (GC-MS) and investigate its antioxidant and in vitro anticancer activities, elucidating the underlying molecular mechanism by use of flow cytometry-based fluorescence-activated cell sorting (FACS) and conducting in silico assessment of binding inhibitory activities of EECC major compounds docked to caspase-3. CG-MS profiling of EECC identified a total of 26 major flavonoids and polyphenolic compounds. DPPH and ABTS assays revealed that EECC exhibits potent antioxidant activity comparable to standard reducing agents. Results of the proliferation assay revealed that EECC exhibit potent, dose-dependent cytotoxic activities against triple-positive (MCF-7) and triple-negative (MDA-MB-231) breast cancer cell models, with IC50 values of 1.3 \u00d7 102 and 8.7 \u00d7 101 \u00b5g\/mL, respectively. The observed cytotoxic effect was specific to studied cancer cells since EECC exhibited minimal (~&lt;10%) cytotoxicity against MCF-12, a normal breast cell line. FACS analysis employing annexin V-FITC\/propidium iodide double labeling demonstrated that the observed anti-proliferative activity against MCF-7 and MDA-MB-231 was mediated via apoptotic as well as necrotic signaling transduction processes. The increase in fluorescence intensity associated with DCFH oxidation to DCF, as reported by FACS, indicated that apoptosis is caused by generation of ROS. The use of caspase-3-specific fluorogenic substrate revealed a dose-dependent elevation in caspase-3 substrate-cleavage activity, which further supports EECC-mediated apoptosis in MCF-7 cells. The major EECC compounds were examined for their inhibitory activity against caspase-3 receptor (1HD2) using molecular docking. Three compounds exhibited the highest glide score energy of \u22125.156, \u22124.691 and \u22124.551 kcal\/mol, respectively. Phenol, 2,6-dimethoxy established strong binding in caspase-3 receptor of hydrogenic type, with residue ARG 207 and of PI-PI stacking type with residue HIS 121. By contract, hexadecenoic acid showed 3 H-bond with the following residues: ASN 615, ASN 616a and THR 646. Taken together, the current findings reveal that EECC exhibits significant and specific cytotoxicity against breast cancer cells mediated by the generation of ROS and culminating into necrosis and apoptosis. Further investigations of the phytoconstituents-rich C. calcitrapa are therefore warranted against breast as well as other human cancer cell models.<\/jats:p>","DOI":"10.3390\/antiox11081514","type":"journal-article","created":{"date-parts":[[2022,8,3]],"date-time":"2022-08-03T20:52:01Z","timestamp":1659559921000},"page":"1514","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":48,"title":["Antioxidant, Anti-Proliferative Activity and Chemical Fingerprinting of Centaurea calcitrapa against Breast Cancer Cells and Molecular Docking of Caspase-3"],"prefix":"10.3390","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9395-0563","authenticated-orcid":false,"given":"Mourad A. M.","family":"Aboul-Soud","sequence":"first","affiliation":[{"name":"Chair of Medical and Molecular Genetics Research, Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, King Saud University, P.O. Box 10219, Riyadh 11433, Saudi Arabia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hanane","family":"Ennaji","sequence":"additional","affiliation":[{"name":"Laboratory of Chemistry-Biochemistry, Environment, Nutrition and Health, Faculty of Medicine and Pharmacy, Hassan II University of Casablanca, Casablanca B.P. 5696, Morocco"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ashok","family":"Kumar","sequence":"additional","affiliation":[{"name":"Vitiligo Research Chair, Department of Dermatology, College of Medicine, King Saud University, Riyadh 11451, Saudi Arabia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3095-9154","authenticated-orcid":false,"given":"Mohammad A.","family":"Alfhili","sequence":"additional","affiliation":[{"name":"Chair of Medical and Molecular Genetics Research, Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, King Saud University, P.O. Box 10219, Riyadh 11433, Saudi Arabia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9447-2361","authenticated-orcid":false,"given":"Ahmed","family":"Bari","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University, P.O. Box 2457, Riyadh 11451, Saudi Arabia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6025-1950","authenticated-orcid":false,"given":"Maqusood","family":"Ahamed","sequence":"additional","affiliation":[{"name":"King Abdullah Institute for Nanotechnology, King Saud University, Riyadh 11451, Saudi Arabia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6614-9594","authenticated-orcid":false,"given":"Mohamed","family":"Chebaibi","sequence":"additional","affiliation":[{"name":"Biomedical and Translational Research Laboratory, Faculty of Medicine and Pharmacy, University of Sidi Mohamed Ben Abdellah, BP 1893, Km 22, Road of Sidi Harazem, Fez B.F. 1893, Morocco"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3707-8461","authenticated-orcid":false,"given":"Mohammed","family":"Bourhia","sequence":"additional","affiliation":[{"name":"Higher Institute of Nursing Professions and Technical Health, Laayoune 70000, Morocco"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Farid","family":"Khallouki","sequence":"additional","affiliation":[{"name":"Biology Department, FTSE, Moulay Ismail University of Meknes, BP 609, Errachidia 52000, Morocco"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Khalid M.","family":"Alghamdi","sequence":"additional","affiliation":[{"name":"Vitiligo Research Chair, Department of Dermatology, College of Medicine, King Saud University, Riyadh 11451, Saudi Arabia"},{"name":"Department of Dermatology, College of Medicine, King Saud University, P.O. Box 240997, Riyadh 11322, Saudi Arabia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"John P.","family":"Giesy","sequence":"additional","affiliation":[{"name":"Toxicology Centre, University of Saskatchewan, Saskatoon, SK S7N 5B3, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,8,3]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Aboul-Soud, M.A.M., Ashour, A.E., Challis, J.K., Ahmed, A.F., Kumar, A., Nassrallah, A., Alahmari, T.A., Saquib, Q., Siddiqui, M.A., and Al-Sheikh, Y. (2020). Biochemical and Molecular Investigation of In Vitro Antioxidant and Anticancer Activity Spectrum of Crude Extracts of Willow Leaves Salix safsaf. 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