Copy Number Variations (CNVs) Account for 10.8% of Pathogenic Variants in Patients Referred for Hereditary Cancer Testing

Abstract

Background/aim: Germline copy number variation (CNV) is a type of genetic variant that predisposes significantly to inherited cancers. Today, next-generation sequencing (NGS) technologies have contributed to multi gene panel analysis in clinical practice.

Materials and methods: A total of 2,163 patients were screened for cancer susceptibility, using a solution-based capture method. A panel of 52 genes was used for targeted NGS. The capture-based approach enables computational analysis of CNVs from NGS data. We studied the performance of the CNV module of the commercial software suite SeqPilot (JSI Medical Systems) and of the non-commercial tool panelcn.MOPS. Additionally, we tested the performance of digital multiplex ligation-dependent probe amplification (digitalMLPA).

Results: Pathogenic/likely pathogenic variants (P/LP) were identified in 464 samples (21.5%). CNV accounts for 10.8% (50/464) of pathogenic variants, referring to deletion/duplication of one or more exons of a gene. In patients with breast and ovarian cancer, CNVs accounted for 10.2% and 6.8% of pathogenic variants, respectively. In colorectal cancer patients, CNV accounted for 28.6% of pathogenic/likely pathogenic variants.

Conclusion: In silico CNV detection tools provide a viable and cost-effective method to identify CNVs from NGS experiments. CNVs constitute a substantial percentage of P/LP variants, since they represent up to one of every ten P/LP findings identified by NGS multigene analysis; therefore, their evaluation is highly recommended to improve the diagnostic yield of hereditary cancer analysis.

Keywords: CNVs; Hereditary cancer; MLPA; computational CNV analysis; digital MLPA.

Figure 1. Schematic of the study’s workflow. The capture-based method enabled computational analysis of copy number variations (CNVs) in nextgeneration sequencing (NGS) data.

Figure 2. Variant types identified in the 52 genes tested in this study. A) Distribution of variant types for the pathogenic/likely pathogenic variants identified in 464 individuals with positive findings. B) Distribution of genes with copy number variations (CNVs).

Figure 3. Percentage of pathogenic/ likely pathogenic (P/LP) copy number variations (CNVs) compared to all identified P/LP variants in the different types of cancer.

Figure 4. A case of a patient with a pathogenic copy number variation (CNV). A) Patient pedigree with a deletion of exons 2-9 in the PMS2 gene. B) Detection of CNVs using next-generation sequencing utilizing JSI software was used to visualize the deletion of PMS2 germline exons 2-9. The bar plot depicts the relative coverage of each target ROI of the patient sample in green and the average relative target coverage of control samples in blue. Below each couple of bars is the ratio of the coverage of target ROIs of the patient sample versus the controls. Reds lines correspond to the 75% deletion limit (lower red dotted line) and the 135% duplication limit (upper red dotted line) for the calculated ratio. The average relative target coverage data are reported as means±standard deviations of control samples. C) Sample plots from Coffalyser.Net displaying probe ratios with 95% confidence intervals as error bars for all PMS2 exons. The proband’s MLPA analysis revealed heterozygous deletions of exons 2-9 in the PMS2 gene (SALSA MLPA probemix P008, MRC Holland).