{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,18]],"date-time":"2026-06-18T07:12:49Z","timestamp":1781766769538,"version":"3.54.5"},"reference-count":62,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2026,1,19]],"date-time":"2026-01-19T00:00:00Z","timestamp":1768780800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100009326","name":"Cancer Research Society","doi-asserted-by":"publisher","award":["837782"],"award-info":[{"award-number":["837782"]}],"id":[{"id":"10.13039\/100009326","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100019711","name":"London Health Sciences Foundation","doi-asserted-by":"crossref","id":[{"id":"10.13039\/100019711","id-type":"DOI","asserted-by":"crossref"}]},{"name":"Department of Obstetrics and Gynecology at the Schulich School of Medicine and Dentistry, Western University"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cancers"],"abstract":"<jats:p>Background\/Objectives: Epithelial ovarian cancer (EOC) is often diagnosed at advanced stages, with metastasis driven by spheroid dissemination within the peritoneal cavity. We previously demonstrated that autophagy supports spheroid cell survival and suggest that it contributes to chemoresistance. Unc-51-like autophagy activating kinase 1 (ULK1), a key regulator of autophagy, has emerged as a promising therapeutic target. Here, we evaluated the effects of ULK1 inhibition via MRT68921, alone and in combination with afatinib\u2014a tyrosine kinase inhibitor (TKI) known to induce pro-survival autophagy\u2014in EOC. Methods: High-grade serous (HGSOC) and ovarian clear cell carcinoma (OCCC) cell lines were cultured under adherent and spheroid conditions. Immunoblotting confirmed on-target effects and modulation of autophagy. Autophagic flux was assessed using mCherry-eGFP-LC3 reporter assays. We assessed 96 dose combinations of MRT68921 and afatinib using drug combination matrices, with synergy evaluated via Synergy Finder. Promising combinations were evaluated across multiple EOC spheroid models and patient ascites-derived organoids. Results: MRT68921 inhibited ULK1 activity and reduced autophagic flux in a context-dependent manner while afatinib alone induced autophagy. Their combination produced synergistic effects at select concentrations, impairing spheroid reattachment and viability. However, MRT68921 alone significantly reduced viability across multiple EOC models, including patient ascites-derived organoids. Conclusions: This study is the first to evaluate the combined effects of MRT68921 and afatinib in epithelial ovarian cancer. Our findings demonstrate that ULK1 inhibition via MRT68921 consistently reduces cell viability across multiple ovarian cancer models, supporting ULK1 as a promising therapeutic target. In contrast, combination with afatinib produced limited and context-dependent effects, indicating that further investigation is needed to identify optimal combination strategies for ULK1-targeted therapies.<\/jats:p>","DOI":"10.3390\/cancers18020307","type":"journal-article","created":{"date-parts":[[2026,1,20]],"date-time":"2026-01-20T08:36:33Z","timestamp":1768898193000},"page":"307","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Evaluating the Therapeutic Potential of MRT68921 and Afatinib in Three-Dimensional Models of Epithelial Ovarian Cancer"],"prefix":"10.3390","volume":"18","author":[{"ORCID":"https:\/\/orcid.org\/0009-0003-2391-5182","authenticated-orcid":false,"given":"Tiffany P. A.","family":"Johnston","sequence":"first","affiliation":[{"name":"The Mary and John Knight Translational Ovarian Cancer Research Unit, Verspeeten Family Cancer Centre, London, ON N6A 5W9, Canada"},{"name":"Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada"},{"name":"Department of Obstetrics and Gynecology, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0007-0365-6880","authenticated-orcid":false,"given":"Jack D.","family":"Webb","sequence":"additional","affiliation":[{"name":"The Mary and John Knight Translational Ovarian Cancer Research Unit, Verspeeten Family Cancer Centre, London, ON N6A 5W9, Canada"},{"name":"Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8996-4436","authenticated-orcid":false,"given":"Matthew J.","family":"Borrelli","sequence":"additional","affiliation":[{"name":"The Mary and John Knight Translational Ovarian Cancer Research Unit, Verspeeten Family Cancer Centre, London, ON N6A 5W9, Canada"},{"name":"Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4528-3713","authenticated-orcid":false,"given":"Emily J.","family":"Tomas","sequence":"additional","affiliation":[{"name":"The Mary and John Knight Translational Ovarian Cancer Research Unit, Verspeeten Family Cancer Centre, London, ON N6A 5W9, Canada"},{"name":"Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0009-6306-5673","authenticated-orcid":false,"given":"\u00c1ine C.","family":"Pucchio","sequence":"additional","affiliation":[{"name":"The Mary and John Knight Translational Ovarian Cancer Research Unit, Verspeeten Family Cancer Centre, London, ON N6A 5W9, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yudith Ramos","family":"Vald\u00e9s","sequence":"additional","affiliation":[{"name":"The Mary and John Knight Translational Ovarian Cancer Research Unit, Verspeeten Family Cancer Centre, London, ON N6A 5W9, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Trevor G.","family":"Shepherd","sequence":"additional","affiliation":[{"name":"The Mary and John Knight Translational Ovarian Cancer Research Unit, Verspeeten Family Cancer Centre, London, ON N6A 5W9, Canada"},{"name":"Department of Anatomy and Cell Biology, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada"},{"name":"Department of Obstetrics and Gynecology, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada"},{"name":"Department of Oncology, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,1,19]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1240","DOI":"10.1016\/S0140-6736(18)32552-2","article-title":"Epithelial Ovarian Cancer","volume":"393","author":"Lheureux","year":"2019","journal-title":"Lancet"},{"key":"ref_2","first-page":"10","article-title":"Cancer statistics, 2025","volume":"75","author":"Siegel","year":"2025","journal-title":"CA Cancer J. 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