Metronomic dosing of ovarian cancer cells with the ATR inhibitor AZD6738 leads to loss of CDC25A expression and resistance to ATRi treatment

Wei Ao; Hong Im Kim; Domenic Tommarello; Kelly A Conrads; Brian L Hood; Tracy Litzi; Tamara Abulez; Pang-Ning Teng; Clifton L Dalgard; Xijun Zhang; Matthew D Wilkerson; Kathleen M Darcy; Christopher M Tarney; Neil T Phippen; Christopher J Bakkenist; G Larry Maxwell; Thomas P Conrads; John I Risinger; Nicholas W Bateman

doi:10.1016/j.ygyno.2023.08.004

Metronomic dosing of ovarian cancer cells with the ATR inhibitor AZD6738 leads to loss of CDC25A expression and resistance to ATRi treatment

Gynecol Oncol. 2023 Oct:177:60-71. doi: 10.1016/j.ygyno.2023.08.004. Epub 2023 Aug 26.

Authors

Wei Ao¹, Hong Im Kim², Domenic Tommarello¹, Kelly A Conrads¹, Brian L Hood¹, Tracy Litzi¹, Tamara Abulez¹, Pang-Ning Teng¹, Clifton L Dalgard³, Xijun Zhang³, Matthew D Wilkerson³, Kathleen M Darcy⁴, Christopher M Tarney⁵, Neil T Phippen⁵, Christopher J Bakkenist⁶, G Larry Maxwell⁷, Thomas P Conrads⁷, John I Risinger², Nicholas W Bateman⁸

Affiliations

¹ Gynecologic Cancer Center of Excellence, Department of Gynecologic Surgery and Obstetrics, Uniformed Services University and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda 20889, MD, USA; Henry M. Jackson Foundation for the Advancement of Military Medicine, Inc. Bethesda, MD 20817, USA.
² Department of Obstetrics, Gynecology and Reproductive Biology, College of Human Medicine, Michigan State University Grand Rapids, MI, USA.
³ The American Genome Center, Department of Anatomy Physiology and Genetics, Collaborative Health Initiative Research Program, Uniformed Services University, 4301 Jones Bridge Road, Bethesda, MD 20814, USA.
⁴ Gynecologic Cancer Center of Excellence, Department of Gynecologic Surgery and Obstetrics, Uniformed Services University and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda 20889, MD, USA; Henry M. Jackson Foundation for the Advancement of Military Medicine, Inc. Bethesda, MD 20817, USA; The John P. Murtha Cancer Center, Uniformed Services University and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda 20889, MD, USA.
⁵ Gynecologic Cancer Center of Excellence, Department of Gynecologic Surgery and Obstetrics, Uniformed Services University and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda 20889, MD, USA; The John P. Murtha Cancer Center, Uniformed Services University and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda 20889, MD, USA.
⁶ Departments of Radiation Biology and Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA.
⁷ Gynecologic Cancer Center of Excellence, Department of Gynecologic Surgery and Obstetrics, Uniformed Services University and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda 20889, MD, USA; The John P. Murtha Cancer Center, Uniformed Services University and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda 20889, MD, USA; Department of Obstetrics and Gynecology, Inova Fairfax Medical Campus, 3300 Gallows Rd. Falls Church, VA 22042, USA.
⁸ Gynecologic Cancer Center of Excellence, Department of Gynecologic Surgery and Obstetrics, Uniformed Services University and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda 20889, MD, USA; Henry M. Jackson Foundation for the Advancement of Military Medicine, Inc. Bethesda, MD 20817, USA; The John P. Murtha Cancer Center, Uniformed Services University and Walter Reed National Military Medical Center, 8901 Wisconsin Avenue, Bethesda 20889, MD, USA. Electronic address: batemann@whirc.org.

PMID: 37639904
DOI: 10.1016/j.ygyno.2023.08.004

Abstract

Objective: ATR kinase inhibitors promote cell killing by inducing replication stress and through potentiation of genotoxic agents in gynecologic cancer cells. To explore mechanisms of acquired resistance to ATRi in ovarian cancer, we characterized ATRi-resistant ovarian cancer cells generated by metronomic dosing with the clinical ATR inhibitor AZD6738.

Methods: ATRi-resistant ovarian cancer cells (OVCAR3 and OV90) were generated by dosing with AZD6738 and assessed for sensitivity to Chk1i (LY2603618), PARPi (Olaparib) and combination with cisplatin or a CDK4/6 inhibitor (Palbociclib). Models were characterized by diverse methods including silencing CDC25A in OV90 cells and assessing impact on ATRi response. Serum proteomic analysis of ATRi-resistant OV90 xenografts was performed to identify circulating biomarker candidates of ATRi-resistance.

Results: AZD6738-resistant cell lines are refractory to LY2603618, but not to Olaparib or combinations with cisplatin. Cell cycle analyses showed ATRi-resistant cells exhibit G1/S arrest following AZD6738 treatment. Accordingly, combination with Palbociclib confers resistance to AZD6738. AZD6738-resistant cells exhibit altered abundances of G1/S phase regulatory proteins, including loss of CDC25A in AZD6738-resistant OV90 cells. Silencing of CDC25A in OV90 cells confers resistance to AZD6738. Serum proteomics from AZD6738-resistant OV90 xenografts identified Vitamin D-Binding Protein (GC), Apolipoprotein E (APOE) and A1 (APOA1) as significantly elevated in AZD6738-resistant backgrounds.

Conclusions: We show that metronomic dosing of ovarian cancer cells with AZD6738 results in resistance to ATR/ Chk1 inhibitors, that loss of CDC25A expression represents a mechanism of resistance to ATRi treatment in ovarian cancer cells and identify several circulating biomarker candidates of CDC25A low, AZD6738-resistant ovarian cancer cells.

Keywords: (CDC25A); ATR; Ataxia telangiectasia and Rad3-related; Cell division cycle 25A; Checkpoint kinase 1; Chk1; Drug resistance; Ovarian cancer.