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Detecting Large Germline Rearrangements of BRCA1 by Next Generation Tumor Sequencing

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Abstract

A majority of BRCA1/2 (BRCA) pathogenic variants (PVs) are single nucleotide substitutions or small insertions/deletions. Copy number variations (CNVs), also known as large genomic rearrangements (LGRs), have been identified in BRCA genes. LGRs detection is a mandatory analysis in hereditary breast and ovarian cancer families, if no predisposing PVs are found by sequencing. Next generation sequencing (NGS) may be used to detect structural variation, since quantitative analysis of sequencing reads, when coupled with appropriate bioinformatics tools, is capable of estimating and predicting germline LGRs (gLGRs). However, applying this approach to tumor tissue is challenging, and the pipelines for determination of CNV are yet to be optimized. The aim of this study was to validate the Next Generation Tumor Sequencing (NGTS) technology to detect various gLGRs of BRCA1 locus in surgical tumor tissue samples. In this study, seven different BRCA1 gLGRs, previously found in high-grade serous ovarian cancers (HGSOC) patients, were detected in tumor samples collected from the patients at a time of HGSOC surgery. This study demonstrated that NGS can accurately detect BRCA1 gLGRs in primary tumors, suggesting that gLGR evaluation in BRCA1 locus should be performed in cases when the screening for BRCA alterations starts from tumor instead of blood. NGS sequencing of tumor samples may become the preferred method to detect both somatic and germline gLGRs in BRCA-encoding loci.

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ACKNOWLEDGMENTS

We would like to thank Franziska M. Lohmeyer for critically reviewing and editing our manuscript.

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Authors and Affiliations

  1. Molecular Diagnostic and Genomics Laboratory, Fondazione Policlinico Universitario A. Gemelli IRCCS, 00168, Rome, Italy

    A. Minucci, G. Mazzuccato & A. Urbani

  2. Division of Oncological Gynecology, Department of Women’s and Children’s Health, Fondazione Policlinico Universitario A. Gemelli IRCCS, 00168, Rome, Italy

    C. Marchetti, A. Pietragalla, G. Scambia & A. Fagotti

  3. Catholic University of the Sacred Heart, 00168, Rome, Italy

    C. Marchetti, G. Scambia, A. Fagotti & A. Urbani

Authors
  1. A. Minucci
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  2. G. Mazzuccato
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  3. C. Marchetti
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  4. A. Pietragalla
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  5. G. Scambia
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  6. A. Fagotti
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  7. A. Urbani
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Contributions

Conception and design: Angelo Minucci, Claudia Marchetti, Giovanni Scambia, Anna Fagotti. Collection and assembly of data: Angelo Minucci and Giorgia Mazzuccato. Data analysis and interpretation: Angelo Minucci. Manuscript writing: Angelo Minucci. Final approval of manuscript: All authors. Accountable for all aspects of the work: All authors.

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Correspondence to A. Minucci or A. Urbani.

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COMPLIANCE WITH ETHICAL STANDARDS

Conflict of interest. The authors declare that they have no conflict of interest.

Statement of compliance with standards of research involving humans as subjects. All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards. Informed consent was obtained from all patients.

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Abbreviations: gBRCA, germline BRCA1/2; HGSOC, High Grade Serous Ovarian Cancer; PVs, Pathogenic variants; PARP-1, Poly-(ADP-ribose) polymerase; tBRCA, tumor BRCA; FFPE, formalin-fixed paraffin-embedded; FFT, fresh frozen tissue; NGS, Next generation sequencing; CNVs, Copy number variations; LGRs, large genomic rearrangements; MLPA, multiplex ligation-dependent probe amplification; DoC, depth of coverage; gLGRs, germline large genomic rearrangements; LOH, loss of heterozygosity; NGTS, next generation tumor sequencing.

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Minucci, A., Mazzuccato, G., Marchetti, C. et al. Detecting Large Germline Rearrangements of BRCA1 by Next Generation Tumor Sequencing. Mol Biol 54, 464–473 (2020). https://doi.org/10.1134/S0026893320030127

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  • DOI: https://doi.org/10.1134/S0026893320030127

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