用ATR抑制剂AZD6738对卵巢癌细胞进行节律性给药会导致细胞周期蛋白磷酸酶25A（CDC25A）表达缺失以及对ATR抑制剂治疗产生耐药性。

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

作者信息

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

机构信息

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.

Department of Obstetrics, Gynecology and Reproductive Biology, College of Human Medicine, Michigan State University Grand Rapids, MI, USA.

出版信息

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

DOI:10.1016/j.ygyno.2023.08.004

PMID:37639904

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.

摘要

目的

ATR激酶抑制剂通过诱导复制应激以及增强妇科癌细胞中基因毒性药物的作用来促进细胞杀伤。为了探究卵巢癌对ATR抑制剂（ATRi）获得性耐药的机制，我们对通过采用临床ATR抑制剂AZD6738进行小剂量持续给药产生的ATRi耐药卵巢癌细胞进行了特征分析。

方法

通过用AZD6738给药产生ATRi耐药卵巢癌细胞（OVCAR3和OV90），并评估其对Chk1抑制剂（LY2603618）、PARP抑制剂（奥拉帕利）以及与顺铂或CDK4/6抑制剂（帕博西尼）联合使用的敏感性。通过多种方法对模型进行特征分析，包括在OV90细胞中沉默细胞分裂周期蛋白25A（CDC25A）并评估其对ATRi反应的影响。对ATRi耐药的OV90异种移植瘤进行血清蛋白质组学分析，以鉴定ATRi耐药的循环生物标志物候选物。

结果

AZD6738耐药细胞系对LY2603618不敏感，但对奥拉帕利或与顺铂联合使用敏感。细胞周期分析表明，ATRi耐药细胞在AZD6738处理后表现出G1/S期阻滞。因此，与帕博西尼联合使用可使细胞对AZD6738产生耐药性。AZD6738耐药细胞表现出G1/S期调节蛋白丰度的改变，包括在AZD6738耐药的OV90细胞中CDC25A缺失。在OV90细胞中沉默CDC25A可使细胞对AZD6738产生耐药性。来自AZD6738耐药的OV90异种移植瘤的血清蛋白质组学分析确定，维生素D结合蛋白（GC）、载脂蛋白E（APOE）和载脂蛋白A1（APOA1）在AZD6738耐药背景下显著升高。