{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T09:59:02Z","timestamp":1785405542029,"version":"3.56.0"},"reference-count":39,"publisher":"MDPI AG","issue":"14","license":[{"start":{"date-parts":[[2026,7,11]],"date-time":"2026-07-11T00:00:00Z","timestamp":1783728000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100003336","name":"National Science Fund of Bulgaria","doi-asserted-by":"crossref","award":["KP-06-COST\/30\/2024"],"award-info":[{"award-number":["KP-06-COST\/30\/2024"]}],"id":[{"id":"10.13039\/501100003336","id-type":"DOI","asserted-by":"crossref"}]},{"award":["KP-06-COST\/30\/2024"],"award-info":[{"award-number":["KP-06-COST\/30\/2024"]}],"id":[{"id":"https:\/\/ror.org\/05eg49r29","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Molecules"],"abstract":"<jats:p>Breast cancer remains one of the leading causes of cancer-related mortality among women worldwide, highlighting the urgent need for novel therapeutic agents with improved efficacy and selectivity. In this study, we investigated the anticancer potential of selected thieno[2,3-d]pyrimidine-based hybrids in breast cancer cell models with different metastatic potential. The compounds were evaluated for their effects on cell viability, clonogenic survival, migration, cell-cycle progression, cytoskeletal organization, and plasma membrane organization. Several derivatives exhibited concentration-dependent antiproliferative activity, inhibited colony formation and cell migration, disrupted cytoskeletal integrity, and modulated plasma membrane organization. Cell-cycle analysis revealed compound- and cell line-dependent alterations in the distribution of cells across the G1, S, and G2\/M phases. Among the tested compounds, derivatives 1, 5, and 6 displayed the most favorable biological profiles, combining antiproliferative activity with moderate selectivity toward breast cancer cells, whereas compound 2 exerted the most pronounced effects on plasma membrane organization and cell migration. Collectively, these findings identify selected thieno[2,3-d]pyrimidine-based hybrids as promising lead compounds for the development of novel anticancer agents targeting both tumor cell proliferation and plasma membrane organization.<\/jats:p>","DOI":"10.3390\/molecules31142435","type":"journal-article","created":{"date-parts":[[2026,7,13]],"date-time":"2026-07-13T08:05:31Z","timestamp":1783929931000},"page":"2435","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Selected Thieno[2,3-d] Pyrimidine Derivatives Target Breast Cancer Cell Proliferation and Membrane Organization"],"prefix":"10.3390","volume":"31","author":[{"given":"Aleksandrina","family":"Nesheva","sequence":"first","affiliation":[{"name":"Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2480-5116","authenticated-orcid":false,"given":"Ivan","family":"Iliev","sequence":"additional","affiliation":[{"name":"Institute of Experimental Morphology, Pathology and Anthropology with Museum, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 25, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0455-4321","authenticated-orcid":false,"given":"Anelia","family":"Mavrova","sequence":"additional","affiliation":[{"name":"Department of Organic Synthesis, Faculty of Chemical Technologies, University of Chemical Technology and Metallurgy, S8 Kliment Ohridski Blvd., 1756 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1580-9681","authenticated-orcid":false,"given":"Denitsa","family":"Yancheva","sequence":"additional","affiliation":[{"name":"Institute of Organic Chemistry with Centre of Phytochemistry, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 9, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8097-3608","authenticated-orcid":false,"given":"Aneliya","family":"Kostadinova","sequence":"additional","affiliation":[{"name":"Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Severina","family":"Semkova","sequence":"additional","affiliation":[{"name":"Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Albena","family":"Momchilova","sequence":"additional","affiliation":[{"name":"Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6908-6082","authenticated-orcid":false,"given":"Iana","family":"Tsoneva","sequence":"additional","affiliation":[{"name":"Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9742-9812","authenticated-orcid":false,"given":"Biliana","family":"Nikolova","sequence":"additional","affiliation":[{"name":"Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2089-3603","authenticated-orcid":false,"given":"Galya","family":"Staneva","sequence":"additional","affiliation":[{"name":"Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, 1113 Sofia, Bulgaria"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,7,11]]},"reference":[{"key":"ref_1","unstructured":"World Health Organization (2026, April 17). Breast Cancer. Available online: https:\/\/www.who.int\/news-room\/fact-sheets\/detail\/breast-cancer."},{"key":"ref_2","first-page":"3147","article-title":"The Story of MCF-7 Breast Cancer Cell Line: 40 years of Experience in Research","volume":"35","author":"Raica","year":"2015","journal-title":"Anticancer Res."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"117","DOI":"10.4236\/jct.2022.133009","article-title":"Breast Cancer MCF-7 Cell Spheroid Culture for Drug Discovery and Development","volume":"13","author":"Chen","year":"2022","journal-title":"J. Cancer Ther."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"105846","DOI":"10.1016\/j.tiv.2024.105846","article-title":"Comprehensive studies on the biological activities of human metastatic (MDA-MB-231) and non-metastatic (MCF-7) breast cancer cell lines, directly or combinedly treated using non-thermal plasma-based approaches","volume":"98","author":"Terefinko","year":"2024","journal-title":"Toxicol. In Vitro"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"43","DOI":"10.1186\/s12935-018-0534-y","article-title":"Characterization of MCF-12A cell phenotype, response to estrogens, and growth in 3D","volume":"18","author":"Sweeney","year":"2018","journal-title":"Cancer Cell Int."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"201","DOI":"10.1038\/s41392-024-01899-w","article-title":"Therapeutic advances of targeting receptor tyrosine kinases in cancer","volume":"9","author":"Tomuleasa","year":"2024","journal-title":"Signal Transduct. Target. Ther."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"341","DOI":"10.1016\/S0002-9440(10)64545-2","article-title":"Aberrant ALK Tyrosine Kinase Signaling","volume":"157","author":"Ladanyi","year":"2000","journal-title":"Am. J. Pathol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"267","DOI":"10.1007\/s10549-020-06060-z","article-title":"Triple-negative and HER2-positive breast cancers found by mammography screening show excellent prognosis","volume":"187","author":"Alanko","year":"2021","journal-title":"Breast Cancer Res. Treat."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"45455","DOI":"10.1021\/acsomega.2c06078","article-title":"Synthesis, Antitumor Activity, and In Silico Drug Design of New Thieno[2,3-d]Pyrimidine-4-One Derivatives as Nonclassical Lipophilic Dihydrofolate Reductase Inhibitors","volume":"7","author":"Abdelaziz","year":"2022","journal-title":"ACS Omega"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"5606","DOI":"10.1039\/D5RA09678B","article-title":"Thienopyrimidine scaffolds as promising antimicrobial agents: Synthesis, biological evaluation, DFT analysis and molecular docking","volume":"16","author":"Sayed","year":"2026","journal-title":"RSC Adv."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"105208","DOI":"10.1016\/j.bioorg.2021.105208","article-title":"New thieno[3,2-d]pyrimidine-based derivatives: Design, synthesis and biological evaluation as antiproliferative agents, EGFR and ARO inhibitors inducing apoptosis in breast cancer cells","volume":"115","author":"Farghaly","year":"2021","journal-title":"Bioorg. Chem."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"21079","DOI":"10.1039\/D5RA09370H","article-title":"Discovery of thieno[2,3- d ]pyrimidine-based dual Aurora B\/VEGFR-2 inhibitors with potent anticancer activity: Molecular docking, mechanistic studies, and in vivo validation in a breast cancer model","volume":"16","author":"Hamdi","year":"2026","journal-title":"RSC Adv."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1159","DOI":"10.1016\/j.bmc.2019.02.044","article-title":"Thieno[2,3-d]pyrimidine as a promising scaffold in medicinal chemistry: Recent advances","volume":"27","author":"Ali","year":"2019","journal-title":"Bioorg. Med. Chem."},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"El-Gamal, R., Elfarrash, S., El-Nablaway, M., Salem, A.A., Zaraei, S.-O., Anbar, H.S., Shoma, A., and El-Gamal, M.I. (2023). Anti-proliferative activity of RIHMS-Qi-23 against MCF-7 breast cancer cell line is through inhibition of cell proliferation and senescence but not inhibition of targeted kinases. BMC Cancer, 23.","DOI":"10.1186\/s12885-023-11547-1"},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Iacob, S., \u015etefan, C., Nechita, A., Matei, M.-N., Lis\u0103, E.-L., Tutunaru, D., Fulga, I., Fulga, A., Cristea, A.-G., and Dragostin, O.-M. (2025). Hybrid Molecules with Purine and Pyrimidine Derivatives for Antitumor Therapy: News, Perspectives, and Future Directions. Molecules, 30.","DOI":"10.3390\/molecules30132707"},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Elkaeed, E.B., Salam, H.A.A.E., Sabt, A., Al-Ansary, G.H., and Eldehna, W.M. (2021). Recent Advancements in the Development of Anti-Breast Cancer Synthetic Small Molecules. Molecules, 26.","DOI":"10.3390\/molecules26247611"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"4354","DOI":"10.1016\/j.bmc.2015.06.025","article-title":"Antiproliferative activities of halogenated pyrrolo[3,2-d]pyrimidines","volume":"23","author":"Temburnikar","year":"2015","journal-title":"Bioorg. Med. Chem."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"659","DOI":"10.1007\/s00044-023-03040-y","article-title":"Recent updates on thienopyrimidine derivatives as anticancer agents","volume":"32","author":"Sayed","year":"2023","journal-title":"Med. Chem. Res."},{"key":"ref_19","first-page":"e-20240198","article-title":"Synthesis and Evaluation of Biological Activities of Thieno-[2,3-d]-pyrimidine Derivatives","volume":"36","author":"Guo","year":"2024","journal-title":"J. Braz. Chem. Soc."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"803","DOI":"10.1080\/17568919.2025.2479422","article-title":"Anti-breast cancer potential of thieno-pyrimidine derivatives as VEGFR-2 inhibitors","volume":"17","author":"Alsfouk","year":"2025","journal-title":"Future Med. Chem."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"507","DOI":"10.1080\/00304948.2019.1666635","article-title":"Facile Synthesis and Anticancer Activity of Novel 4-Aminothieno[2,3-d]pyrimidines and Triazolothienopyrimidines","volume":"51","author":"Ali","year":"2019","journal-title":"Org. Prep. Proced. Int."},{"key":"ref_22","first-page":"103492","article-title":"Design, molecular modeling and anticancer evaluation of thieno[2,3-d]pyrimidine derivatives as inhibitors of topoisomerase II","volume":"94","author":"Khalil","year":"2019","journal-title":"Bioorg. Chem."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Mavrova, A., Dimov, S., Sulikovska, I., Yancheva, D., Iliev, I., Tsoneva, I., Staneva, G., and Nikolova, B. (2022). Design, Cytotoxicity and Antiproliferative Activity of 4-Amino-5-methyl-thieno[2,3-d]pyrimidine-6-carboxylates against MFC-7 and MDA-MB-231 Breast Cancer Cell Lines. Molecules, 27.","DOI":"10.3390\/molecules27103314"},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Iliev, I., Mavrova, A., Yancheva, D., Dimov, S., Staneva, G., Nesheva, A., Tsoneva, I., and Nikolova, B. (2023). 2-Alkyl-Substituted-4-Amino-Thieno[2,3-d]Pyrimidines: Anti-Proliferative Properties to In Vitro Breast Cancer Models. Molecules, 28.","DOI":"10.3390\/molecules28176347"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"142586","DOI":"10.1016\/j.molstruc.2025.142586","article-title":"Discovery of new thieno[2,3-d]pyrimidine-based dual VEGFR-2 and EGFR inhibitors for enhanced therapeutic efficacy in breast cancer","volume":"1341","author":"Yousef","year":"2025","journal-title":"J. Mol. Struct."},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Malla, S., Nyinawabera, A., Neupane, R., Pathak, R., Lee, D., Abou-Dahech, M.S., Kumari, S., Sinha, S., Tang, Y., and Ray, A. (2024). Novel Thienopyrimidine-Hydrazinyl Compounds Induce DRP1-Mediated Non-Apoptotic Cell Death in Triple-Negative Breast Cancer Cells. Cancers, 16.","DOI":"10.3390\/cancers16152621"},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Nammalwar, B., and Bunce, R.A. (2024). Recent Advances in Pyrimidine-Based Drugs. Pharmaceuticals, 17.","DOI":"10.3390\/ph17010104"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"611","DOI":"10.1194\/jlr.TR119000439","article-title":"Lipid rafts as signaling hubs in cancer cell survival\/death and invasion: Implications in tumor progression and therapy","volume":"61","author":"Mollinedo","year":"2020","journal-title":"J. Lipid Res."},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Domenichini, M., Gogna, A., Maggi, C.A., Moreschi, E., Ventura, A., Codibue, M., Grillo, E., Corsini, M., and Mitola, S. (2025). Lipid\u2013Protein Interplay in the Regulation of Receptor Tyrosine Kinases. Cells, 14.","DOI":"10.3390\/cells14231836"},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Luck, M., Fischer, M., Werle, M., Scheidt, H.A., and M\u00fcller, P. (2021). Impact of Selected Small-Molecule Kinase Inhibitors on Lipid Membranes. Pharmaceuticals, 14.","DOI":"10.3390\/ph14080746"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"513","DOI":"10.1038\/s41580-019-0160-0","article-title":"Tying lipid rafts to oncogenic signalling","volume":"20","author":"Strzyz","year":"2019","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Bernardes, N., and Fialho, A.M. (2018). Perturbing the Dynamics and Organization of Cell Membrane Components: A New Paradigm for Cancer-Targeted Therapies. Int. J. Mol. Sci., 19.","DOI":"10.3390\/ijms19123871"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"321","DOI":"10.1007\/s10863-020-09846-4","article-title":"Lipid composition of the cancer cell membrane","volume":"52","author":"Szlasa","year":"2020","journal-title":"J. Bioenerg. Biomembr."},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Bansal, I., Pandey, A.K., and Ruwali, M. (2023). Small-molecule inhibitors of kinases in breast cancer therapy: Recent advances, opportunities, and challenges. Front. Pharmacol., 14.","DOI":"10.3389\/fphar.2023.1244597"},{"key":"ref_35","doi-asserted-by":"crossref","unstructured":"Fakhry, M.M., Mahmoud, K., Nafie, M.S., Noor, A.O., Hareeri, R.H., Salama, I., and Kishk, S.M. (2022). Rational Design, Synthesis and Biological Evaluation of Novel Pyrazoline-Based Antiproliferative Agents in MCF-7 Cancer Cells. Pharmaceuticals, 15.","DOI":"10.3390\/ph15101245"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1064","DOI":"10.55730\/1300-0527.3594","article-title":"Synthesis of new imine-\/amine-bearing imidazo[1,2-a]pyrimidine derivatives and screening of their cytotoxic activity","volume":"47","author":"Atalay","year":"2023","journal-title":"Turk. J. Chem."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"693","DOI":"10.1016\/j.bioorg.2018.07.009","article-title":"Synthesis and DNase I inhibitory properties of some 5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d]pyrimidines","volume":"80","author":"Mavrova","year":"2018","journal-title":"Bioorg. Chem."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"55","DOI":"10.1016\/0022-1759(83)90303-4","article-title":"Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays","volume":"65","author":"Mosmann","year":"1983","journal-title":"J. Immunol. Methods"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"2315","DOI":"10.1038\/nprot.2006.339","article-title":"Clonogenic assay of cells in vitro","volume":"1","author":"Franken","year":"2006","journal-title":"Nat. Protoc."}],"container-title":["Molecules"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1420-3049\/31\/14\/2435\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,14]],"date-time":"2026-07-14T04:11:46Z","timestamp":1784002306000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1420-3049\/31\/14\/2435"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,11]]},"references-count":39,"journal-issue":{"issue":"14","published-online":{"date-parts":[[2026,7]]}},"alternative-id":["molecules31142435"],"URL":"https:\/\/doi.org\/10.3390\/molecules31142435","relation":{},"ISSN":["1420-3049"],"issn-type":[{"value":"1420-3049","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,11]]}}}