title: Post-mortem testing; germline BRCA1/2 variant detection using archival FFPE non-tumor tissue. A new paradigm in genetic counseling.
abstract: Accurate estimation of cancer risk in HBOC families often requires BRCA1/2 testing, but this may be impossible in deceased family members. Previous, testing archival formalin-fixed, paraffin-embedded (FFPE) tissue for germline BRCA1/2 variants was unsuccessful, except for the Jewish founder mutations. A high-throughput method to systematically test for variants in all coding regions of BRCA1/2 in archival FFPE samples of non-tumor tissue is described, using HaloPlex target enrichment and next-generation sequencing. In a validation study, correct identification of variants or wild-type was possible in 25 out of 30 (83%) FFPE samples (age range 1-14 years), with a known variant status in BRCA1/2. No false positive was found. Unsuccessful identification was due to highly degraded DNA or presence of large intragenic deletions. In clinical use, a total of 201 FFPE samples (aged 0-43 years) were processed. Thirty-six samples were rejected because of highly degraded DNA or failed library preparation. Fifteen samples were investigated to search for a known variant. In the remaining 150 samples (aged 0-38 years), three variants known to affect function and one variant likely to affect function in BRCA1, six variants known to affect function and one variant likely to affect function in BRCA2, as well as four variants of unknown significance (VUS) in BRCA1 and three VUS in BRCA2 were discovered. It is now possible to test for germline BRCA1/2 variants in deceased persons, using archival FFPE samples from non-tumor tissue. Accurate genetic counseling is achievable in families where variant testing would otherwise be impossible.
authors.affiliation: Department of Clinical Genetics, Vejle Hospital, Lillebaelt Hospital, Vejle, Denmark.
authors.forename: Annabeth Høgh
authors.initials: AH
authors.lastname: Petersen
authors.affiliation: Department of Clinical Genetics, Vejle Hospital, Lillebaelt Hospital, Vejle, Denmark.
authors.forename: Mads Malik
authors.initials: MM
authors.lastname: Aagaard
authors.affiliation: Department of Clinical Genetics, Vejle Hospital, Lillebaelt Hospital, Vejle, Denmark.
authors.forename: Henriette Roed
authors.initials: HR
authors.lastname: Nielsen
authors.affiliation: Department of Clinical Oncology, Vejle Hospital, Lillebaelt Hospital, Vejle, Denmark.|Faculty of Health Sciences, Institute of Regional Health Services Research, University of Southern Denmark, Odense, Denmark.
authors.forename: Karina Dahl
authors.initials: KD
authors.lastname: Steffensen
authors.affiliation: Faculty of Health Sciences, Institute of Regional Health Services Research, University of Southern Denmark, Odense, Denmark.|Department of Pathology, Vejle Hospital, Lillebaelt Hospital, Vejle, Denmark.
authors.forename: Marianne
authors.initials: M
authors.lastname: Waldstrøm
authors.affiliation: Department of Clinical Genetics, Vejle Hospital, Lillebaelt Hospital, Vejle, Denmark.|Faculty of Health Sciences, Institute of Regional Health Services Research, University of Southern Denmark, Odense, Denmark.
authors.forename: Anders
authors.initials: A
authors.lastname: Bojesen
mesh_terms: D001344:Autopsy; D019313:BRCA1 Protein / Q000235:genetics*; D024682:BRCA2 Protein / Q000235:genetics*; D001943:Breast Neoplasms / Q000175:diagnosis / Q000235:genetics*; D005189:False Positive Reactions; D005260:Female; D005817:Genetic Counseling / Q000379:methods* / Q000592:standards; D005820:Genetic Testing / Q000379:methods* / Q000592:standards; D018095:Germ-Line Mutation; D006801:Humans; D008297:Male; D010375:Pedigree; D017422:Sequence Analysis, DNA / Q000379:methods / Q000592:standards; D016707:Tissue Fixation / Q000379:methods
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D024682:BRCA2 Protein; C551750:BRCA2 protein, human