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Alternative splicing and ACMG-AMP-2015-based classification of PALB2 genetic variants : An ENIGMA report

Lopez-Perolio, Irene; Leman, Raphaël; Behar, Raquel; Lattimore, Vanessa; Pearson, John F.; Castéra, Laurent; Martins, Alexandra; Vaur, Dominique; Goardon, Nicolas and Davy, Grégoire, et al. (2019) In Journal of Medical Genetics
Abstract

Background: PALB2 monoallelic loss-of-function germ-line variants confer a breast cancer risk comparable to the average BRCA2 pathogenic variant. Recommendations for risk reduction strategies in carriers are similar. Elaborating robust criteria to identify loss-of-function variants in PALB2 - without incurring overprediction - is thus of paramount clinical relevance. Towards this aim, we have performed a comprehensive characterisation of alternative splicing in PALB2, analysing its relevance for the classification of truncating and splice site variants according to the 2015 American College of Medical Genetics and Genomics-Association for Molecular Pathology guidelines. Methods: Alternative splicing was characterised in RNAs extracted... (More)

Background: PALB2 monoallelic loss-of-function germ-line variants confer a breast cancer risk comparable to the average BRCA2 pathogenic variant. Recommendations for risk reduction strategies in carriers are similar. Elaborating robust criteria to identify loss-of-function variants in PALB2 - without incurring overprediction - is thus of paramount clinical relevance. Towards this aim, we have performed a comprehensive characterisation of alternative splicing in PALB2, analysing its relevance for the classification of truncating and splice site variants according to the 2015 American College of Medical Genetics and Genomics-Association for Molecular Pathology guidelines. Methods: Alternative splicing was characterised in RNAs extracted from blood, breast and fimbriae/ovary-related human specimens (n=112). RNAseq, RT-PCR/CE and CloneSeq experiments were performed by five contributing laboratories. Centralised revision/curation was performed to assure high-quality annotations. Additional splicing analyses were performed in PALB2 c.212-1G>A, c.1684+1G>A, c.2748+2T>G, c.3113+5G>A, c.3350+1G>A, c.3350+4A>C and c.3350+5G>A carriers. The impact of the findings on PVS1 status was evaluated for truncating and splice site variant. Results: We identified 88 naturally occurring alternative splicing events (81 newly described), including 4 in-frame events predicted relevant to evaluate PVS1 status of splice site variants. We did not identify tissue-specific alternate gene transcripts in breast or ovarian-related samples, supporting the clinical relevance of blood-based splicing studies. Conclusions: PVS1 is not necessarily warranted for splice site variants targeting four PALB2 acceptor sites (exons 2, 5, 7 and 10). As a result, rare variants at these splice sites cannot be assumed pathogenic/likely pathogenic without further evidences. Our study puts a warning in up to five PALB2 genetic variants that are currently reported as pathogenic/likely pathogenic in ClinVar.

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@article{1528ef2d-baeb-49f9-9abb-017ef2455760,
  abstract     = {<p>Background: PALB2 monoallelic loss-of-function germ-line variants confer a breast cancer risk comparable to the average BRCA2 pathogenic variant. Recommendations for risk reduction strategies in carriers are similar. Elaborating robust criteria to identify loss-of-function variants in PALB2 - without incurring overprediction - is thus of paramount clinical relevance. Towards this aim, we have performed a comprehensive characterisation of alternative splicing in PALB2, analysing its relevance for the classification of truncating and splice site variants according to the 2015 American College of Medical Genetics and Genomics-Association for Molecular Pathology guidelines. Methods: Alternative splicing was characterised in RNAs extracted from blood, breast and fimbriae/ovary-related human specimens (n=112). RNAseq, RT-PCR/CE and CloneSeq experiments were performed by five contributing laboratories. Centralised revision/curation was performed to assure high-quality annotations. Additional splicing analyses were performed in PALB2 c.212-1G&gt;A, c.1684+1G&gt;A, c.2748+2T&gt;G, c.3113+5G&gt;A, c.3350+1G&gt;A, c.3350+4A&gt;C and c.3350+5G&gt;A carriers. The impact of the findings on PVS1 status was evaluated for truncating and splice site variant. Results: We identified 88 naturally occurring alternative splicing events (81 newly described), including 4 in-frame events predicted relevant to evaluate PVS1 status of splice site variants. We did not identify tissue-specific alternate gene transcripts in breast or ovarian-related samples, supporting the clinical relevance of blood-based splicing studies. Conclusions: PVS1 is not necessarily warranted for splice site variants targeting four PALB2 acceptor sites (exons 2, 5, 7 and 10). As a result, rare variants at these splice sites cannot be assumed pathogenic/likely pathogenic without further evidences. Our study puts a warning in up to five PALB2 genetic variants that are currently reported as pathogenic/likely pathogenic in ClinVar.</p>},
  author       = {Lopez-Perolio, Irene and Leman, Raphaël and Behar, Raquel and Lattimore, Vanessa and Pearson, John F. and Castéra, Laurent and Martins, Alexandra and Vaur, Dominique and Goardon, Nicolas and Davy, Grégoire and Garre, Pilar and García-Barberán, Vanesa and Llovet, Patricia and Pérez-Segura, Pedro and Díaz-Rubio, Eduardo and Caldés, Trinidad and Hruska, Kathleen S. and Hsuan, Vickie and Wu, Sitao and Pesaran, Tina and Karam, Rachid and Vallon-Christersson, Johan and Borg, Ake and Investigators, Kconfab and Valenzuela-Palomo, Alberto and Velasco, Eladio Andrés and Southey, Melissa and Vreeswijk, Maaike P.G. and Devilee, Peter and Kvist, Anders and Spurdle, Amanda B. and Walker, Logan C. and Krieger, Sophie and De La Hoya, Miguel},
  issn         = {0022-2593},
  keyword      = {acmg-amp guidelines,palb2,pvs1,splicing,variant classification},
  language     = {eng},
  publisher    = {BMJ Publishing Group},
  series       = {Journal of Medical Genetics},
  title        = {Alternative splicing and ACMG-AMP-2015-based classification of PALB2 genetic variants : An ENIGMA report},
  url          = {http://dx.doi.org/10.1136/jmedgenet-2018-105834},
  year         = {2019},
}