Article open access publication

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

Human Molecular Genetics, Oxford University Press (OUP), ISSN 1460-2083

Volume 25, 11, 2016

DOI:10.1093/hmg/ddw094, Dimensions: pub.1059645866, PMC: PMC5081057, PMID: 27008870,


Vega, Ana (3)
Poplawski, Nicola (26) (27)
Burwinkel, Barbara (31) (32)
Surowy, Harald (31) (32)
Bojesen, Stig E. (33) (34) (35)
Giles, Graham G (23) (42)
Milne, Roger L. (23) (42)
Andrulis, Irene L. (43) (44)
Hall, Per (37)
Wang, Qin (45)



  1. (1) Hospital Clínico San Carlos, grid.411068.a
  2. (2) University of Rouen, grid.10400.35
  3. (3) Instituto de Investigación Sanitaria de Santiago, grid.488911.d
  4. (4) University of Otago, grid.29980.3a
  5. (5) Leiden University Medical Center, grid.10419.3d
  6. (6) University of Southampton, grid.5491.9
  7. (7) University of Santiago de Compostela, grid.11794.3a
  8. (8) QIMR Berghofer Medical Research Institute, grid.1049.c
  9. (9) Philipp University of Marburg, grid.10253.35
  10. (10) Center for Genomic Medicine, Rigshospitalet, Copenhagen University Hospital, Copenhagen DK-2100, Denmark
  11. (11) Peter MacCallum Cancer Center, University of Melbourne, Melbourne, VIC 3002, Australia
  12. (12) Department of Gynaecology and Obstetrics, University Hospital Düsseldorf, Heinrich-Heine University Düsseldorf, Düsseldorf 40225, Germany
  13. (13) Department of Gynaecology and Obstetrics, University Hospital of Schleswig-Holstein, Campus Kiel, Christian-Albrechts University Kiel, Kiel 24105, Germany
  14. (14) University of Münster, grid.5949.1
  15. (15) Hannover Medical School, grid.10423.34
  16. (16) Universitätsklinikum Tübingen, grid.411544.1
  17. (17) National Institutes of Health, grid.94365.3d
  18. (18) NorthShore University HealthSystem, grid.240372.0
  19. (19) Institute Curie, grid.418596.7
  20. (20) Centre Franois Baclesse, grid.476192.f
  21. (21) Ambry Genetics (United States), grid.465138.d
  22. (22) Royal Devon and Exeter Hospital, grid.416118.b
  23. (23) University of Melbourne, grid.1008.9
  24. (24) Medical University of Vienna, grid.22937.3d
  25. (25) Christchurch Hospital, grid.414299.3
  26. (26) South Australia Pathology, grid.414733.6
  27. (27) University of Adelaide, grid.1010.0
  28. (28) National Institute of Arthritis and Musculoskeletal and Skin Diseases, grid.420086.8
  29. (29) London School of Hygiene & Tropical Medicine, grid.8991.9
  30. (30) Institute of Cancer Research, grid.18886.3f
  31. (31) Heidelberg University, grid.7700.0
  32. (32) German Cancer Research Center, grid.7497.d
  33. (33) Gentofte Hospital, grid.411646.0, Capital Region
  34. (34) Herlev Hospital, grid.411900.d, Capital Region
  35. (35) University of Copenhagen, grid.5254.6, KU
  36. (36) Department of Breast Surgery, Herlev and Gentofte Hospital, Copenhagen University Hospital, 2730 Denmark
  37. (37) Karolinska Institute, grid.4714.6
  38. (38) University Cancer Center Hamburg, grid.412315.0
  39. (39) Fondazione IRCCS Istituto Nazionale dei Tumori, grid.417893.0
  40. (40) Associazione Volontari Italiani Sangue (AVIS) comunale di Milano, Milano 20139, Italy
  41. (41) Mayo Clinic, grid.66875.3a
  42. (42) Cancer Council Victoria, grid.3263.4
  43. (43) University of Toronto, grid.17063.33
  44. (44) Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON M5G 1X5, Canada
  45. (45) University of Cambridge, grid.5335.0
  46. (46) Cyprus Institute of Neurology and Genetics, grid.417705.0
  47. (47) Department of Oncological Sciences
  48. (48) University of Utah, grid.223827.e


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.


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