Clinical, splicing, and functional analysis to classify BRCA2 exon 3 variants: Application of a points-based ACMG/AMP approach

Thomassen, M; Mesman, RLS; Hansen, TVO; Menendez, M; Rossing, M; Esteban-Sanchez, A; Tudini, E; Torngren, T; Parsons, MT; Pedersen, IS; Teo, SH; Kruse, TA; Moller, P; Borg, A; Jensen, UB; Christensen, LL; Singer, CF; Muhr, D; Santamarina, M; Brandao, R; Andresen, BS; Feng, BJ; Canson, D; Richardson, ME; Karam, R; Pesaran, T; LaDuca, H; Conner, BR; Abualkheir, N; Hoang, L; Calleja, FMGR; Andrews, L; James, PA; Bunyan, D; Hamblett, A; Radice, P; Goldgar, DE; Walker, LC; Engel, C; Claes, KBM; Machackova, E; Baralle, D; Viel, A; Wappenschmidt, B; Lazaro, C; Vega, A; ENIGMA Consortium; Vreeswijk, MPG; de la Hoya, M; Spurdle, AB

Author(s): Thomassen, M; Mesman, RLS; Hansen, TVO; Menendez, M; Rossing, M; Esteban-Sanchez, A; Tudini, E; Torngren, T; Parsons, MT; Pedersen, IS; Teo, SH; Kruse, TA; Moller, P; Borg, A; Jensen, UB; Christensen, LL; Singer, CF; Muhr, D; Santamarina, M; Brandao, R; Andresen, BS; Feng, BJ; Canson, D; Richardson, ME; Karam, R; Pesaran, T; LaDuca, H; Conner, BR; Abualkheir, N; Hoang, L; Calleja, FMGR; Andrews, L; James, PA; Bunyan, D; Hamblett, A; Radice, P; Goldgar, DE; Walker, LC; Engel, C; Claes, KBM; Machackova, E; Baralle, D; Viel, A; Wappenschmidt, B; Lazaro, C; Vega, A; ENIGMA Consortium; Vreeswijk, MPG; de la Hoya, M; Spurdle, AB;
Details: Publication Year 2022-12,Volume 43,Issue #12,Page 1921-1944
Journal Title: Human Mutation
Publication Type: Research article
Abstract: Skipping of BRCA2 exon 3 (∆E3) is a naturally occurring splicing event, complicating clinical classification of variants that may alter ∆E3 expression. This study used multiple evidence types to assess pathogenicity of 85 variants in/near BRCA2 exon 3. Bioinformatically predicted spliceogenic variants underwent mRNA splicing analysis using minigenes and/or patient samples. ∆E3 was measured using quantitative analysis. A mouse embryonic stem cell (mESC) based assay was used to determine the impact of 18 variants on mRNA splicing and protein function. For each variant, population frequency, bioinformatic predictions, clinical data, and existing mRNA splicing and functional results were collated. Variant class was assigned using a gene-specific adaptation of ACMG/AMP guidelines, following a recently proposed points-based system. mRNA and mESC analysis combined identified six variants with transcript and/or functional profiles interpreted as loss of function. Cryptic splice site use for acceptor site variants generated a transcript encoding a shorter protein that retains activity. Overall, 69/85 (81%) variants were classified using the points-based approach. Our analysis shows the value of applying gene-specific ACMG/AMP guidelines using a points-based approach and highlights the consideration of cryptic splice site usage to appropriately assign PVS1 code strength.
Publisher: Wiley
Keywords: Animals; Humans; Mice; Alternative Splicing; BRCA2 Protein/genetics/metabolism; *Genes, BRCA2; *RNA Splice Sites; RNA Splicing; RNA, Messenger/genetics/metabolism; ACMG/AMP classification; Brca2; dPCR; functional analysis; quantitation; splicing
Department(s): Familial Cancer Centre
PubMed ID: 35979650
Publisher's Version: https://doi.org/10.1002/humu.24449
Open Access at Publisher's Site: https://doi.org/10.1002/humu.24449
Terms of Use/Rights Notice: Refer to copyright notice on published article.

Creation Date: 2025-04-17 07:51:27

Last Modified: 2025-04-17 07:51:48