Nutlin-3a: A Potential Therapeutic Opportunity for TP53 Wild-Type Ovarian Carcinomas.

Erin K Crane,Daisy I Izaguirre,Yvonne T M Tsang,Suet-Yan Kwan,Kwong-Kwok Wong,Joanne S Richards,Lisa K Mullany,David M Gershenson,Zhifei Zu,Annie Ny Cheung

doi:10.1371/journal.pone.0135101

Abstract

Epithelial ovarian cancer is a diverse molecular and clinical disease, yet standard treatment is the same for all subtypes. TP53 mutations represent a node of divergence in epithelial ovarian cancer histologic subtypes and may represent a therapeutic opportunity in subtypes expressing wild type, including most low-grade ovarian serous carcinomas, ovarian clear cell carcinomas and ovarian endometrioid carcinomas, which represent approximately 25% of all epithelial ovarian cancer. We therefore sought to investigate Nutlin-3a—a therapeutic which inhibits MDM2, activates wild-type p53, and induces apoptosis—as a therapeutic compound for TP53 wild-type ovarian carcinomas. Fifteen epithelial ovarian cancer cell lines of varying histologic subtypes were treated with Nutlin-3a with determination of IC50 values. Western Blot (WB) and quantitative real-time polymerase chain reaction (qRT-PCR) analyses quantified MDM2, p53, and p21 expression after Nutlin-3a treatment. DNA from 15 cell lines was then sequenced for TP53 mutations in exons 2-11 including intron-exon boundaries. Responses to Nutlin-3a were dependent upon TP53 mutation status. By qRT-PCR and WB, levels of MDM2 and p21 were upregulated in wild-type TP53 sensitive cell lines, and p21 induction was reduced or absent in mutant cell lines. Annexin V assays demonstrated apoptosis in sensitive cell lines treated with Nutlin-3a. Thus, Nutlin-3a could be a potential therapeutic agent for ovarian carcinomas expressing wild-type TP53 and warrants further investigation.

Highlights

While ovarian cancer is the ninth most common cancer in the United States, it is the sixth most deadly: approximately 22,000 new cases of ovarian cancer are diagnosed annually, with 14,270 attributable deaths [1]
The fluorescent sequencing chromatograms for the detected mutations for these nine cell lines are provided in the S1 Fig. All of these TP53 mutant cell lines were quite resistant to Nutlin-3a (IC50 = 20 to >70 μM)
TP53 mutations are virtually ubiquitous in high-grade serous ovarian carcinomas; this is not the case for other Epithelial ovarian cancer (EOC)

Summary

Introduction

While ovarian cancer is the ninth most common cancer in the United States, it is the sixth most deadly: approximately 22,000 new cases of ovarian cancer are diagnosed annually, with 14,270 attributable deaths [1]. High-grade serous ovarian carcinomas (SOCs) comprise the majority of epithelial ovarian cancer, less common subtypes account one third of all cases, many of which are chemoresistant and frequently with wild-type TP53. These histologic subtypes have distinct molecular origins, with correspondingly diverse and specific clinical behaviors; they are treated with the same regimen as high-grade SOC. Whereas high-grade SOCs typically affect postmenopausal patients and are chemo-sensitive, median overall survival is only 54 months (compared to 126 months for low-grade) [10] Those with advanced-stage clear cell carcinomas typically fare worse than those affected by high-grade SOC, partially due to their insensitivity to platinum-based chemotherapy [11]

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Conclusion