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Hum Mol Genet. 2016 Jun 1;25(11):2256-2268. Epub 2016 Mar 23.

Combined genetic and splicing analysis of BRCA1 c.[594-2A>C; 641A>G] highlights the relevance of naturally occurring in-frame transcripts for developing disease gene variant classification algorithms.

Author information

1
Molecular Oncology Laboratory, Instituto de Investigacion Sanitaria San Carlos (IdISSC), Hospital Clinico San Carlos, Madrid, Spain amanda.spurdle@qimr.edu.au mhoya@hotmail.com.
2
Inserm U1079-IRIB, University of Rouen, Normandy Centre for Genomic and Personalized Medicine, Rouen, France.
3
Molecular Oncology Laboratory, Instituto de Investigacion Sanitaria San Carlos (IdISSC), Hospital Clinico San Carlos, Madrid, Spain.
4
Fundacion Publica Galega de Medicina Xenómica-SERGAS Grupo de Medicina Xenómica-USC, IDIS, CIBERER, Santiago de Compostela 15706, Spain.
5
Department of Pathology, University of Otago, Christchurch 8140, New Zealand.
6
Department of Clinical Genetics, Leiden University Medical Centre, Leiden 2300, The Netherlands.
7
Human Development and Health, Faculty of Medicine, University of Southampton, Southampton S016 5YA, UK.
8
CIBERER, Grupo de Medicina Xenómica-USC, Universidade de Santiago de Compostela, Fundacion Galega de Medicina Xenómica (SERGAS), Santiago de Compostela 15706, Spain.
9
Department of Clinical Genetics, Leiden University Medical Centre, Leiden 2300, The Netherlands Department of Clinical Genetics, Leiden University Medical Centre, Leiden 2300, The Netherlands.
10
Department of Genetics and Computational Biology, QIMR Berghofer Medical Research Institute, Brisbane, QLD 4006, Australia.
11
Center for Hereditary Breast and Ovarian Cancer, Center for Integrated Oncology (CIO), Medical Faculty, University Hosptial Cologne, Cologne 50931, Germany.
12
Center for Genomic Medicine, Rigshospitalet, Copenhagen University Hospital, Copenhagen DK-2100, Denmark.
13
Peter MacCallum Cancer Center, University of Melbourne, Melbourne, VIC 3002, Australia.
14
Department of Gynaecology and Obstetrics, University Hospital Düsseldorf, Heinrich-Heine University Düsseldorf, Düsseldorf 40225, Germany.
15
Department of Gynaecology and Obstetrics, University Hospital of Schleswig-Holstein, Campus Kiel, Christian-Albrechts University Kiel, Kiel 24105, Germany.
16
Institute of Human Genetics, University of Münster, Münster 48149, Germany.
17
Institute of Human Genetics, Hannover Medical School, Hannover 30625, Germany.
18
Institute of Medical Genetics and Applied Genomics, University Hospital Tuebingen, Tuebingen 72076, Germany.
19
National Institutes of Health, Bethesda, MD 20892-2152, USA.
20
Center for Medical Genetics, NorthShore University Health System, University of Chicago Pritzker School of Medicine, Evanston, IL 60201, USA.
21
Service de Génétique, Department de Biologie des Tumeurs, Institut Curie and INSERM U830, Centre de Recherche de l'Institut Curie, Paris, and Universite Paris Descartes, Sorbonne Paris Cite, Paris 75248, France.
22
Service de Génétique, Department de Biologie des Tumeurs, Institut Curie, Paris 75248, France.
23
Centre Francois Baclesse, Laboratoire de Biologie et de Genetique du Cancer, 14076 Caen, Paris 75248, France.
24
Ambry Genetics, Aliso Viejo, CA 92656, USA.
25
Department of Clinical Genetics, Royal Devon and Exeter Hospital, Exeter, UK.
26
Genetic Epidemiology Laboratory, Department of Pathology, The University of Melbourne, Melbourne, VIC 3010, Australia.
27
Department of Obstetrics and Gynaecology, Medical University of Vienna, Vienna, Austria, Waehringer Guertel 18-20, A 1090 Vienna, Austria.
28
Genetic Health Service NZ, South Island Hub, Christchurch Hospital, Christchurch 8140, New Zealand.
29
Adult Genetics Unit, South Australian Clinical Genetics Service, SA Pathology at the Women's and Children's Hospital, North Adelaide, SA 5067, Australia University Department of Paediatrics, University of Adelaide, North Terrace, Adelaide, SA 5000, Australia.
30
Clinical Genetics Branch, DCEG, NCI, NIH, Bethesda, MD, USA.
31
London School of Hygiene and Tropical Medicine, London WC1E 7HT, UK.
32
Breakthrough Breast Cancer Research Centre, Institute of Cancer Research, London SW3 6JB, UK Division of Breast Cancer Research, Institute of Cancer Research, London SW3 6JB, UK.
33
Molecular Biology of Breast Cancer, Department of Gynecology and Obstetrics, University of Heidelberg, Heidelberg 69120, Germany Molecular Epidemiology Group, German Cancer Research Center, DKFZ, Heidelberg 69120, Germany.
34
Copenhagen General Population Study, Herlev and Gentofte Hospital, Copenhagen 2730, Denmark Department of Clinical Biochemistry, Herlev Hospital, Copenhagen University Hospital, Herlev 2730, Denmark Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen 1165, Denmark.
35
Department of Breast Surgery, Herlev and Gentofte Hospital, Copenhagen University Hospital, 2730 Denmark.
36
Department of Molecular Medicine and Surgery, Karolinska Institutet, Stockholm SE-171 77, Sweden.
37
Department of Oncology Pathology, Karolinska Institutet, Stockholm SE-171 77, Sweden.
38
Division of Cancer Epidemiology, German Cancer Research Center (DKFZ), Heidelberg 69120, Germany University Cancer Center Hamburg (UCCH), University Medical Center Hamburg-Eppendorf, Hamburg 20246, Germany.
39
Division of Cancer Epidemiology, German Cancer Research Center (DKFZ), Heidelberg 69120, Germany.
40
Unit of "Molecular bases of genetic risk and genetic testing", Department of Preventive and Predictive Medicine, Fondazione IRCCS Istituto Nazionale dei Tumori, Milano 20139, Italy.
41
Associazione Volontari Italiani Sangue (AVIS) comunale di Milano, Milano 20139, Italy.
42
Department of Health Sciences Research, Mayo Clinic, Rochester, MN 55905, USA.
43
Centre for Epidemiology and Biostatistics, Melbourne School of Population and Global Health, University of Melbourne, VIC 3010, Australia Cancer Epidemiology Centre, Cancer Council Victoria, Melbourne, VIC 3004, Australia.
44
Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada Department of Molecular Genetics, University of Toronto, M5B 1W8, Canada.
45
Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada.
46
Department of Medical Epidemiology and Biostatistics, Karolinska Institutet, Stockholm SE-171 77, Sweden.
47
Centre for Cancer Genetic Epidemiology, Department of Oncology, University of Cambridge, Cambridge CB1 8RN, UK.
48
Centre for Cancer Genetic Epidemiology, Department of Public Health and Primary Care, University of Cambridge, Cambridge CB1 8RN, UK.
49
Centre for Cancer Genetic Epidemiology, Department of Public Health and Primary Care, University of Cambridge, Cambridge CB1 8RN, UK Department of Electron Microscopy/Molecular Pathology, The Cyprus Institute of Neurology and Genetics, 1683, Nicosia, Cyprus.
50
Department of Health Sciences Research, Mayo Clinic, Rochester, MN 55905, USA Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN 55905, USA.
51
Department of Oncological Sciences.
52
Department of Dermatology, Huntsman Cancer Institute, University of Utah School of Medicine, Salt Lake City, UT 84132, USA.
53
Department of Genetics and Computational Biology, QIMR Berghofer Medical Research Institute, Brisbane, QLD 4006, Australia amanda.spurdle@qimr.edu.au mhoya@hotmail.com.

Abstract

A recent analysis using family history weighting and co-observation classification modeling indicated that BRCA1 c.594-2A > C (IVS9-2A > C), previously described to cause exon 10 skipping (a truncating alteration), displays characteristics inconsistent with those of a high risk pathogenic BRCA1 variant. We used large-scale genetic and clinical resources from the ENIGMA, CIMBA and BCAC consortia to assess pathogenicity of c.594-2A > C. The combined odds for causality considering case-control, segregation and breast tumor pathology information was 3.23 × 10-8 Our data indicate that c.594-2A > C is always in cis with c.641A > G. The spliceogenic effect of c.[594-2A > C;641A > G] was characterized using RNA analysis of human samples and splicing minigenes. As expected, c.[594-2A > C; 641A > G] caused exon 10 skipping, albeit not due to c.594-2A > C impairing the acceptor site but rather by c.641A > G modifying exon 10 splicing regulatory element(s). Multiple blood-based RNA assays indicated that the variant allele did not produce detectable levels of full-length transcripts, with a per allele BRCA1 expression profile composed of ≈70-80% truncating transcripts, and ≈20-30% of in-frame Δ9,10 transcripts predicted to encode a BRCA1 protein with tumor suppression function.We confirm that BRCA1c.[594-2A > C;641A > G] should not be considered a high-risk pathogenic variant. Importantly, results from our detailed mRNA analysis suggest that BRCA-associated cancer risk is likely not markedly increased for individuals who carry a truncating variant in BRCA1 exons 9 or 10, or any other BRCA1 allele that permits 20-30% of tumor suppressor function. More generally, our findings highlight the importance of assessing naturally occurring alternative splicing for clinical evaluation of variants in disease-causing genes.

PMID:
27008870
PMCID:
PMC5081057
DOI:
10.1093/hmg/ddw094
[Indexed for MEDLINE]
Free PMC Article

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