Detection of Germline Variants in 450 Breast/Ovarian Cancer Families with a Multi-Gene Panel Including Coding and Regulatory Regions.

Chiara Guglielmi,Enrico Tagliafico,Rosa Scarpitta,Eleonora Conti,Cinzia Cosini,Alvaro Galli,Stefania Tommasi,Matteo Ghilli,Tiziana Cervelli,Caterina Vivanet,Laura Cortesi,Margherita Patruno,Gaetana Gambino,Paolo Aretini,Luisa Balestrino,Maria Grazia Tibiletti,Ileana Carnevali,Brunella Pilato,Elisabetta Falaschi,Mariella Tancredi,Francesca Spina,Caterina Congregati,Elena Tenedini,Francesca Bellè,Marco Marino,Maria A Caligo

doi:10.3390/ijms22147693

Abstract

With the progress of sequencing technologies, an ever-increasing number of variants of unknown functional and clinical significance (VUS) have been identified in both coding and non-coding regions of the main Breast Cancer (BC) predisposition genes. The aim of this study is to identify a mutational profile of coding and intron-exon junction regions of 12 moderate penetrance genes (ATM, BRIP1, CDH1, CHEK2, NBN, PALB2, PTEN, RAD50, RAD51C, RAD51D, STK11, TP53) in a cohort of 450 Italian patients with Hereditary Breast/Ovarian Cancer Syndrome, wild type for germline mutation in BRCA1/2 genes. The analysis was extended to 5′UTR and 3′UTR of all the genes listed above and to the BRCA1 and BRCA2 known regulatory regions in a subset of 120 patients. The screening was performed through NGS target resequencing on the Illumina platform MiSeq. 8.7% of the patients analyzed is carriers of class 5/4 coding variants in the ATM (3.6%), BRIP1 (1.6%), CHEK2 (1.8%), PALB2 (0.7%), RAD51C (0.4%), RAD51D (0.4%), and TP53 (0.2%) genes, while variants of uncertain pathological significance (VUSs)/class 3 were identified in 9.1% of the samples. In intron-exon junctions and in regulatory regions, variants were detected respectively in 5.1% and in 32.5% of the cases analyzed. The average age of disease onset of 44.4 in non-coding variant carriers is absolutely similar to the average age of disease onset in coding variant carriers for each proband’s group with the same cancer type. Furthermore, there is not a statistically significant difference in the proportion of cases with a tumor onset under age of 40 between the two groups, but the presence of multiple non-coding variants in the same patient may affect the aggressiveness of the tumor and it is worth underlining that 25% of patients with an aggressive tumor are carriers of a PTEN 3′UTR-variant. This data provides initial information on how important it might be to extend mutational screening to the regulatory regions in clinical practice.

Highlights

Breast cancer (BC) is the most common malignant tumor in women representing 25%of the total number of cancer cases in women with an annual increase of 793,700 new cases and 197,600 deaths only in developed countries [1,2].The presence of germline mutations in the BRCA1 and BRCA2 genes significantly increases the risk of developing breast and ovarian cancer [3,4,5], only 25% of the risk of familial cancer can be attributed to pathogenetic mutations localized in the coding regions of these two genes
Mutational screening was performed for 450 patients in the coding regions and 50 bp flanking intronic sequences of 10 genes: ATM, BRIP1, CDH1, CHECK2, PALB2, PTEN, RAD51C, RAD51D, STK11, TP53
Among the four cases of patients with ovarian cancer, we found a Pathogenic variant in RAD51C, 2 VUSs, respectively in ATM and RAD50 genes, and a Likely-benign variant as Likely-benign and distributed as follows: 6 in ATM, 3 in RAD51D, 2 in CDH1 and 3 respectively in BRIP1, RAD50, and RAD51C genes

Summary

Introduction

Breast cancer (BC) is the most common malignant tumor in women representing 25%of the total number of cancer cases in women with an annual increase of 793,700 new cases and 197,600 deaths only in developed countries [1,2].The presence of germline mutations in the BRCA1 and BRCA2 genes significantly increases the risk of developing breast and ovarian cancer [3,4,5], only 25% of the risk of familial cancer can be attributed to pathogenetic mutations localized in the coding regions of these two genes. Hereditary Breast and Ovarian Cancer (HBOC) has been mostly limited to the coding regions and to the intron-exon junctions of BRCA1 and BRCA2, precluding the identification of mutations in the non-coding and/or regulatory regions and in other genes that can confer a high or moderate risk to the disease. Under these conditions, mutation screening is negative in 80% of the cases analyzed [6]. In recent years, the interest in investigating the effect that regulatory variants could have on cancer risk has increased [10]

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