Genetic Testing in Hereditary Breast and Ovarian Cancer Using Massive Parallel Sequencing

Overview

Journal Biomed Res Int

Publisher Wiley

Specialties Biomedical Engineering
Biotechnology
General Medicine

Date 2014 Aug 20

PMID 25136594

Citations 8

Authors

Anna Ruiz

Gemma Llort

Carmen Yague

Neus Baena

Marina Vinas

Montse Torra

Anna Brunet

Miquel A Segui

Eugeni Saigi

Miriam Guitart

Affiliations

Soon will be listed here.

Abstract

High throughput methods such as next generation sequencing are increasingly used in molecular diagnosis. The aim of this study was to develop a workflow for the detection of BRCA1 and BRCA2 mutations using massive parallel sequencing in a 454 GS Junior bench top sequencer. Our approach was first validated in a panel of 23 patients containing 62 unique variants that had been previously Sanger sequenced. Subsequently, 101 patients with familial breast and ovarian cancer were studied. BRCA1 and BRCA2 exon enrichment has been performed by PCR amplification using the BRCA MASTR kit (Multiplicom). Bioinformatic analysis of reads is performed with the AVA software v2.7 (Roche). In total, all 62 variants were detected resulting in a sensitivity of 100%. 71 false positives were called resulting in a specificity of 97.35%. All of them correspond to deletions located in homopolymeric stretches. The analysis of the homopolymers stretches of 6 bp or longer using the BRCA HP kit (Multiplicom) increased the specificity of the detection of BRCA1 and BRCA2 mutations to 99.99%. We show here that massive parallel pyrosequencing can be used as a diagnostic strategy to test for BRCA1 and BRCA2 mutations meeting very stringent sensitivity and specificity parameters replacing traditional Sanger sequencing with a lower cost.

Citing Articles

Prevalence of BRCA homopolymeric indels in an ION Torrent-based tumour-to-germline testing workflow in high-grade ovarian carcinoma.

Azzollini J, Agnelli L, Conca E, Torelli T, Busico A, Capone I Sci Rep. 2023; 13(1):7781.

PMID: 37179432 PMC: 10182972. DOI: 10.1038/s41598-023-33857-x.

The Progress of the Specific and Rapid Genetic Detection Methods for Ovarian Cancer Diagnosis and Treatment.

Dong K, Zhang W, Cheng S, Shu W, Zhao R, Wang H Technol Cancer Res Treat. 2022; 21:15330338221114497.

PMID: 36062718 PMC: 9446467. DOI: 10.1177/15330338221114497.

Management of BRCA Tumour Testing in an Integrated Molecular Tumour Board Multidisciplinary Model.

Azzollini J, Vingiani A, Agnelli L, Tamborini E, Perrone F, Conca E Front Oncol. 2022; 12:857515.

PMID: 35463374 PMC: 9026437. DOI: 10.3389/fonc.2022.857515.

Rapid detection of copy number variations and point mutations in genes using a single workflow by ion semiconductor sequencing pipeline.

Germani A, Libi F, Maggi S, Stanzani G, Lombardi A, Pellegrini P Oncotarget. 2018; 9(72):33648-33655.

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Targeted resequencing and variant validation using pxlence PCR assays.

Coppieters F, Verniers K, De Leeneer K, Vandesompele J, Lefever S Biomol Detect Quantif. 2016; 6:22-6.

PMID: 27077044 PMC: 4822215. DOI: 10.1016/j.bdq.2015.09.001.

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