A comprehensive  BRCA1/2  NGS pipeline for an immediate Copy Number Variation (CNV) detection in breast and ovarian cancer molecular diagnosis

Concolino, Paola; Rizza, Roberta; Mignone, Flavio; Costella, Alessandra; Guarino, Donatella; Carboni, Ilaria; Capoluongo, Ettore; Santonocito, Concetta; Urbani, Andrea; Minucci, Angelo

doi:10.1016/j.cca.2018.02.012

Cited by 32 publications

(26 citation statements)

References 15 publications

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“…10% of the overall BRCA1 mutations in Hungary. These frequencies are comparable with those detected in other Middle European populations [3,4,8], but fall short of the extreme ratios found in the Netherlands (36% of all BRCA1 mutants) [15], Italy (19% of all BRCA1 mutants) [16], and UK (20% of all BRCA1 mutants) [5], where frequently occurring founder mutations constitute the majority of LGR cases.…”

Section: Discussionsupporting

confidence: 84%

Complex Characterization of Germline Large Genomic Rearrangements of the BRCA1 and BRCA2 Genes in High-Risk Breast Cancer Patients—Novel Variants from a Large National Center

Bozsik

Pócza

Papp

et al. 2020

IJMS

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Large genomic rearrangements (LGRs) affecting one or more exons of BRCA1 and BRCA2 constitute a significant part of the mutation spectrum of these genes. Since 2004, the National Institute of Oncology, Hungary, has been involved in screening for LGRs of breast or ovarian cancer families enrolled for genetic testing. LGRs were detected by multiplex ligation probe amplification method, or next-generation sequencing. Where it was possible, transcript-level characterization of LGRs was performed. Phenotype data were collected and analyzed too. Altogether 28 different types of LGRs in 51 probands were detected. Sixteen LGRs were novel. Forty-nine cases were deletions or duplications in BRCA1 and two affected BRCA2. Rearrangements accounted for 10% of the BRCA1 mutations. Three exon copy gains, two complex rearrangements, and 23 exon losses were characterized by exact breakpoint determinations. The inferred mechanisms for LGR formation were mainly end-joining repairs utilizing short direct homologies. Comparing phenotype features of the LGR-carriers to that of the non-LGR BRCA1 mutation carriers, revealed no significant differences. Our study is the largest comprehensive report of LGRs of BRCA1/2 in familial breast and ovarian cancer patients in the Middle and Eastern European region. Our data add novel insights to genetic interpretation associated to the LGRs.

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Section: Discussionsupporting

confidence: 84%

Complex Characterization of Germline Large Genomic Rearrangements of the BRCA1 and BRCA2 Genes in High-Risk Breast Cancer Patients—Novel Variants from a Large National Center

Bozsik

Pócza

Papp

et al. 2020

IJMS

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“…However the possibility of analyzing all types of mutations through a single workflow offered by the NGS is appealing. Therefore several research groups have tested different NGS platforms for CNVs detection by implementing and validating customized workflow [ 21 , 28 – 30 ].…”

Section: Discussionmentioning

confidence: 99%

“…Pathogenic CNVs in BRCA1 and BRCA2 genes range respectively from 0 to 27% and 0% to 8% [ 12 – 14 ] The higher frequency of LGRs in BRCA1 compared to BRCA2 gene is caused by higher density of ALU sequences [ 15 ], which mediate the formation of LGRs [ 16 , 17 ]. LGRs in BRCA1 and BRCA2 vary across populations and ethnicities [ 18 – 20 ], with a frequency up to 12% in Italy [ 21 ]. Many approaches have been used for detecting BRCA LGRs, such as comparative genomic hybridization (CGH), array comparative genomic hybridization (aCGH), real Time PCR (qPCR) and fluorescent in situ Hybridization (FISH) [ 22 ].…”

Section: Introductionmentioning

confidence: 99%

“…Nowadays, the Multiplex ligation-dependent probe amplification (MLPA, MRC-Holland, Amsterdam, the Netherlands) is the primary method for detecting LGRs in BRCA1/2 with the Multiplex Amplicon Quantification (MAQ) (Multiplicon, Niel, Belgium), as an alternative way [ 23 – 25 ]. Both approaches are consistent, but they always need the confirmation with other techniques, because of the possibility of false positive results [ 21 ]. For many years, BRCA gene analysis on peripheral blood has been using Sanger sequencing and MLPA.…”

Section: Introductionmentioning

confidence: 99%

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Rapid detection of copy number variations and point mutations in BRCA1/2 genes using a single workflow by ion semiconductor sequencing pipeline

Germani

Libi²,

Maggi³

et al. 2018

Oncotarget

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Molecular analysis of BRCA1 (MIM# 604370) and BRCA2 (MIM #600185) genes is essential for familial breast and ovarian cancer prevention and treatment. An efficient, rapid, cost-effective accurate strategy for the detection of pathogenic variants is crucial. Mutations detection of BRCA1/2 genes includes screening for single nucleotide variants (SNVs), small insertions or deletions (indels), and Copy Number Variations (CNVs). Sanger sequencing is unable to identify CNVs and therefore Multiplex Ligation Probe amplification (MLPA) or Multiplex Amplicon Quantification (MAQ) is used to complete the BRCA1/2 genes analysis. The rapid evolution of Next Generation Sequencing (NGS) technologies allows the search for point mutations and CNVs with a single platform and workflow. In this study we test the possibilities of NGS technology to simultaneously detect point mutations and CNVs in BRCA1/2 genes, using the OncomineTM BRCA Research Assay on Personal Genome Machine (PGM) Platform with Ion Reporter Software for sequencing data analysis (Thermo Fisher Scientific). Comparison between the NGS-CNVs, MLPA and MAQ results shows how the NGS approach is the most complete and fast method for the simultaneous detection of all BRCA mutations, avoiding the usual time consuming multistep approach in the routine diagnostic testing of hereditary breast and ovarian cancers.

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“…In particular, our laboratory (Molecular and Genomics Diagnostics Unit, Fondazione Policlinico Universitario Agostino Gemelli, IRCCS) remodulated its priorities by temporarily interrupting most of the molecular tests and guaranteeing only those considered "urgent" and not postponable. Herein, we report changes regarding the execution of the germline BRCA (gBRCA ) testing, during the COVID-19 outbreak in March 2020, in a referral center for the gBRCA testing [2][3][4][5].…”

Section: Introductionmentioning

confidence: 99%

BRCA testing in a genomic diagnostics referral center during the COVID-19 pandemic

et al. 2020

Self Cite

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The first person-to-person transmission of the 2019-novel coronavirus in Italy on 21 February 2020 led to an infection chain that represents one of the largest known COVID-19 outbreaks outside Asia. Hospitals have been forced to reorganized their units in response to prepare for an unforeseen healthcare emergency. In this context, our laboratory (Molecular and Genomic Diagnostics Unit, Fondazione Policlinico Universitario Agostino Gemelli IRCCS) re-modulated its priorities by temporarily interrupting most of the molecular tests guaranteeing only those considered "urgent" and not postponable. In particular, this paper details changes regarding the execution of germline BRCA (gBRCA ) testing in our laboratory. A substantial reduction in gBRCA testing (about 60%) compared to the first 2 months of the current year was registered, but the requests have not been reset. The requesting physicians were mainly gynaecologists and oncologists. These evidences further emphasize the new era of gBRCA testing in the management of cancer patients and confirms definitively the integration of gBRCA testing/ Next Generation Sequencing (NGS) into clinical oncology. Finally, a re-organization of gBRCA testing in our Unit, mainly related to delayed and reduced arrival of tests was necessary, ensuring, however, a high-quality standard and reliability, mandatory for gBRCA testing in a clinical setting.

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A comprehensive BRCA1/2 NGS pipeline for an immediate Copy Number Variation (CNV) detection in breast and ovarian cancer molecular diagnosis

Cited by 32 publications

References 15 publications

Complex Characterization of Germline Large Genomic Rearrangements of the BRCA1 and BRCA2 Genes in High-Risk Breast Cancer Patients—Novel Variants from a Large National Center

Complex Characterization of Germline Large Genomic Rearrangements of the BRCA1 and BRCA2 Genes in High-Risk Breast Cancer Patients—Novel Variants from a Large National Center

Rapid detection of copy number variations and point mutations in BRCA1/2 genes using a single workflow by ion semiconductor sequencing pipeline

BRCA testing in a genomic diagnostics referral center during the COVID-19 pandemic

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