Genomic instability and CCNE1 amplification as emerging biomarkers for stratifying high-grade serous ovarian cancer

Abstract

Introduction: Ovarian cancer (OC) is one of the leading causes of cancer-related death in women worldwide. Treatment with PARP-inhibitors has significantly improved survival in patients with high-grade serous cancer (HGSC) bearing BRCA1/2 mutations (~22% of the cases), and/or homologous recombination deficiency (HRD, ~50%). Unfortunately, limited therapeutic alternatives are available for BRCA1/2 wild type/HR proficient HGSC patients, who usually exhibit resistance to standard treatments and poor prognosis.

Methods: Herein, we present the results of a comprehensive genomic profiling (CGP) analysis using the Oncomine Comprehensive Assay® (OCA) Plus in a consecutive retrospective cohort of 102 HGSC patients characterized in our institution.

Results: Genomic instability, measured by Genomic Instability Metric (GIM) >16, was found in 40% of the cases and was significantly associated with BRCA1/2 mutations (p=0.009), with a better prognosis in terms of recurrence-free survival (p=0.01). CCNE1 amplification was observed in 29% of cases and was negatively correlated with BRCA1/2 mutations (p=0.001), without any association with GIM, supporting CCNE1 as a strong and independent driver of tumorigenesis. Additionally, CCNE1 amplification was validated with fluorescent in situ hybridization (FISH), supporting the analytical robustness of NGS data (rho=0.93), and investigated by immunohistochemistry (IHC), revealing that CCNE1 protein overexpression was observed in the absence of gene amplification in 45% of cases.

Discussion: Our real-world study supports the clinical utility of the GIM metric and the analytical validity of CCNE1 amplification, a new promising biomarker for personalizing treatment in HR proficient HGSC patients. The discordance between CCNE1 amplification and protein expression raises intriguing questions about the mechanisms of CCNE1-driven tumorigenesis and warrants further investigation.

Keywords: CCNE1 amplification; comprehensive genomic profiling (CGP); high-grade serous ovarian cancer (HGSC); homologous recombination deficiency (HRD) score; precision oncology cancer.

Figure 1

Oncoplots showing the top 50 altered genes by mutations. Only patients harboring alterations in selected genes are reported. Samples are represented by columns, and genes are represented by rows. Pathogenic/likely pathogenic alterations are marked in black. Indications about the Genomic Instability Metric (GIM) CCNE1 amplification status were also added. Amp, amplified; Not amp, not amplified; DEL, deletion; INS, insertion; SNV, single-nucleotide variation.

Figure 2

Oncoplots showing the top 50 altered genes by copy number variations. Only patients harboring alterations in selected genes are reported. Samples are represented by columns, and genes are represented by rows. Pathogenic/likely pathogenic alterations are marked in black. Indications about Genomic Instability Metric (GIM) and BRCA1/2 status were also added. Amp, amplified; Not amp, not amplified; DEL, deletion; INS, insertion; SNV, single-nucleotide variation.

Figure 3

Boxplot showing the inverse correlation between pathogenic BRCA1/2 mutation and CCNE1 amplification detected by NGS. NGS, next-generation sequencing.

Figure 4

NGS and FISH CCNE1 pattern in an amplified case. (A) Top panels showing log2 ratios of the copy number estimates by NGS assay for each amplicon (blue and brown points). Brown horizontal bars represent genomic segmentation resulting from amplicons with similar log2 ratios clustered together. The blue arrow indicates the segment localized on CytoBand 19q12, corresponding to CCNE1 gene amplicons. The image shows a case with focal amplification of CCNE1, confirmed by FISH in the picture below (B), where green and red signals refer to centromere and CCNE1 gene, respectively. NGS, next-generation sequencing; FISH, fluorescence in situ hybridization.

Figure 5

CCNE1 immunohistochemistry. (A) Boxplot showing the positive correlation between CCNE1 amplification results, detected by both NGS and FISH, and protein expression, detected by IHC. (B–D) Representative IHC staining of CCNE1 in three HGSC cases. (B) HGSC with CCNE1 amplification and positive IHC. (C) HGSC with no CCNE1 amplification and positive IHC. (D) HGSC with no CCNE1 amplification and a negative IHC. Original magnification: ×5 main image and ×20–×10 boxes. HGSC, high-grade serous ovarian carcinoma; IHC, immunohistochemistry; NGS, next-generation sequencing; FISH, fluorescence in situ hybridization.

Figure 6

Clinical outcomes. (A–C) Kaplan–Meier estimates of recurrence-free survival according to BRCA1/2 mutational status (A), homologous recombination status calculated with GIM score (B), and both (C). Panel (C) confirms that patients with GIM-positive scores had a similar outcome independent of BRCA1/2 mutation presence. wt, wild type; mut, mutated; BRCAm, BRCA1/2 mutated; BRCAwt, BRCA1/2 wild type; GIM, Genomic Instability Metric.