A global functional analysis of missense mutations reveals two major hotspots in the PALB2 tumor suppressor

Rodrigue, Amélie; Margaillan, Guillaume; Gomes, Thiago T.; Coulombe, Yan; Montalban, Gemma; Carvalho, Simone da Costa e Silva; Milano, Larissa; Ducy, Mandy; De-Gregoriis, Giuliana; Dellaire, Graham; Silva, Wilson A.; Monteiro, Alvaro N.A.; Carvalho, Marcelo A.; Simard, Jacques; Masson, Jean‐Yves

doi:10.1093/nar/gkz780

Cited by 39 publications

(77 citation statements)

References 58 publications

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“…3d). This region is important for MutLα heterodimers formation, necessary for the correct mismatched DNA fragment excision [97]. The p.Val717Met (c.2149G > A) is a VUS that presents conflicting information of pathogenicity by ClinVar database and only AlignGVGD does not predict it as pathogenic.…”

Section: Discussionmentioning

confidence: 99%

Germline variants in DNA repair genes associated with hereditary breast and ovarian cancer syndrome: analysis of a 21 gene panel in the Brazilian population

et al. 2020

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Background: The Hereditary Breast and Ovarian Cancer Syndrome (HBOC) occurs in families with a history of breast/ ovarian cancer, presenting an autosomal dominant inheritance pattern. BRCA1 and BRCA2 are high penetrance genes associated with an increased risk of up to 20-fold for breast and ovarian cancer. However, only 20-30% of HBOC cases present pathogenic variants in those genes, and other DNA repair genes have emerged as increasing the risk for HBOC. In Brazil, variants in ATM, ATR, CHEK2, MLH1, MSH2, MSH6, POLQ, PTEN, and TP53 genes have been reported in up to 7.35% of the studied cases. Here we screened and characterized variants in 21 DNA repair genes in HBOC patients. Methods:We systematically analyzed 708 amplicons encompassing the coding and flanking regions of 21 genes related to DNA repair pathways (ABRAXAS1, ATM, ATR, BARD1, BRCA1, BRCA2, BRIP1, CDH1, CHEK2, MLH1, MRE11, MSH2, MSH6, NBN, PALB2, PMS2, PTEN, RAD50, RAD51, TP53 and UIMC1). A total of 95 individuals with HBOC syndrome clinical suspicion in Southeast Brazil were sequenced, and 25 samples were evaluated for insertions/deletions in BRCA1/BRCA2 genes. Identified variants were assessed in terms of population allele frequency and their functional effects were predicted through in silico algorithms. Results: We identified 80 variants in 19 genes. About 23.4% of the patients presented pathogenic variants in BRCA1, BRCA2 and TP53, a frequency higher than that identified among previous studies in Brazil. We identified a novel variant in ATR, which was predicted as pathogenic by in silico tools. The association analysis revealed 13 missense variants in ABRAXAS1, BARD1, BRCA2, CHEK2, CDH1, MLH1, PALB2, and PMS2 genes, as significantly associated with increased risk to HBOC, and the patients carrying those variants did not present large insertions or deletions in BRCA1/BRCA2 genes.

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Section: Discussionmentioning

confidence: 99%

Germline variants in DNA repair genes associated with hereditary breast and ovarian cancer syndrome: analysis of a 21 gene panel in the Brazilian population

et al. 2020

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“…Assays using HR as a read-out have emerged as the standard for the functional characterisation of VUS in BRCA1 and BRCA2 (Bouwman et al, 2013;Woods et al, 2016;Shimelis et al, 2017;Starita et al, 2018;Mesman et al, 2019). More recently, VUS in PALB2 have also been characterized in a similar manner (Figures 1, 2) (Park et al, 2014;Foo et al, 2017;Boonen et al, 2019;Rodrigue et al, 2019;Wiltshire et al, 2019). To identify variants that impact HR, the well described DR-GFP reporter, as well as the more recently introduced CRISPR-LMNA HR assay were used (Kass et al, 2013;Ducy et al, 2019).…”

Section: A Comprehensive Functional Analysis Of Vus In Palb2mentioning

confidence: 99%

“…PALB2-interacting proteins are depicted underneath their respective PALB2 interacting domain/regions. All PALB2 genetic variants from five functional studies (Park et al, 2014;Foo et al, 2017;Boonen et al, 2019;Rodrigue et al, 2019;Wiltshire et al, 2019) are shown and categorized per (functional) domain as benign (green framed sections), truncating (red framed sections), or VUS and synthetic missense variants (MVs) (blue framed sections) based on ClinVar. All functionally damaging PALB2 VUS with an HR efficiency < 50% compared to wild type PALB2 in at least one functional assay are highlighted in red.…”

Section: A Comprehensive Functional Analysis Of Vus In Palb2mentioning

confidence: 99%

“…This phenotype was used as a readout for the functional characterization of several PALB2 variants, revealing that two variants, p.L939W and p.L1143P, impaired PALB2 function (Park et al, 2014). Lastly, since PALB2 interacts with BRCA1 and BRCA2 to load RAD51 at sites of DSBs, co-immunoprecipitation, recruitment to laser micro-irradiation induced DNA damage, and DNA-damage-induced RAD51 foci formation were among the additional functional readouts to study the impact of PALB2 variants on HR ( Figure 2) (Park et al, 2014;Foo et al, 2017;Boonen et al, 2019;Rodrigue et al, 2019;Wiltshire et al, 2019). A complete overview of all functional assays that were performed by three recent studies is provided in Table 1 (Boonen et al, 2019;Rodrigue et al, 2019;Wiltshire et al, 2019).…”

Section: A Comprehensive Functional Analysis Of Vus In Palb2mentioning

confidence: 99%

“…Rodrigue et al (2019) first tested their set of 41 PALB2 VUS using PARPi sensitivity assays ( Table 2). Their assay was set up in HeLa cells in which endogenous PALB2 was depleted Park et al, 2014Foo et al, 2017Boonen et al, 2019Wiltshire et al, 2019Rodrigue et al, 2019 Nucleotide change Protein change Park et al, 2014Foo et al, 2017Boonen et al, 2019Wiltshire et al, 2019Rodrigue et al, 2019 Nucleotide change Protein change Park et al, 2014Foo et al, 2017Boonen et al, 2019Wiltshire et al, 2019Rodrigue et al, 2019 Nucleotide change Protein change by siRNA treatment. Following transient expression of siRNAresistant YFP-PALB2 cDNA, with or without a variant, cells were assayed for PARPi sensitivity.…”

Section: A Comprehensive Functional Analysis Of Vus In Palb2mentioning

confidence: 99%

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Functional Characterization of PALB2 Variants of Uncertain Significance: Toward Cancer Risk and Therapy Response Prediction

Boonen

Vreeswijk

Attikum

2020

Front. Mol. Biosci.

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In recent years it has become clear that pathogenic variants in PALB2 are associated with a high risk for breast, ovarian and pancreatic cancer. However, the clinical relevance of variants of uncertain significance (VUS) in PALB2, which are increasingly identified through clinical genetic testing, is unclear. Here we review recent advances in the functional characterization of VUS in PALB2. A combination of assays has been used to assess the impact of PALB2 VUS on its function in DNA repair by homologous recombination, cell cycle regulation and the control of cellular levels of reactive oxygen species (ROS). We discuss the outcome of this comprehensive analysis of PALB2 VUS, which showed that VUS in PALB2's Coiled-Coil (CC) domain can impair the interaction with BRCA1, whereas VUS in its WD40 domain affect PALB2 protein stability. Accordingly, the CC and WD40 domains of PALB2 represent hotspots for variants that impair PALB2 protein function. We also provide a future perspective on the high-throughput analysis of VUS in PALB2, as well as the functional characterization of variants that affect PALB2 RNA splicing. Finally, we discuss how results from these functional assays can be valuable for predicting cancer risk and responsiveness to cancer therapy, such as treatment with PARP inhibitor-or platinumbased chemotherapy.

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Germline testing for homologous recombination repair genes—opportunities and challenges

Hirsch

Gieldon

Sutter

et al. 2020

Genes Chromosomes & Cancer

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Pathogenic variants in the BRCA1 and BRCA2 genes are well known causes of hereditary breast and ovarian cancer. Other genes involved in the homologous recombination pathway can also be associated with increased probability of cancer development, for example, breast and ovarian cancer, prostate and pancreatic cancer, colorectal cancer, and even childhood tumors like medulloblastoma. Traditionally, patients and families likely to harbor a genetic predisposition have been identified using personal and family history. Several checklists and risk prediction tools have proven to be useful in the clinic. Through the widespread application of next generation sequencing of tumor tissue, a growing number of individuals with genetic cancer predisposition is now identified molecularly, even in the absence of a suggestive family history. Any constitutional variant identified during molecular genetic testing has to be assessed for its relevance, both functionally and in the context of patient phenotype. Variant curation is time consuming, but has been increasingly standardized by introduction of several guidelines to allow reliable and reproducible classification of constitutional variants. Variant classification by expert panels using data mining tools, evidence‐based decision trees and gene specific criteria represents the gold standard. Participation of geneticists in molecular tumor boards facilitates the curation of potential constitutional variants, germline validation and thus directing the patient to appropriate counselling and care pathways. Due to the high relevance of germline variants for treatment and surveillance of the index patient and predictive testing and surveillance of relatives, only pathogenic or likely pathogenic variants must be used for clinical decision‐making.

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A global functional analysis of missense mutations reveals two major hotspots in the PALB2 tumor suppressor

Cited by 39 publications

References 58 publications

Germline variants in DNA repair genes associated with hereditary breast and ovarian cancer syndrome: analysis of a 21 gene panel in the Brazilian population

Germline variants in DNA repair genes associated with hereditary breast and ovarian cancer syndrome: analysis of a 21 gene panel in the Brazilian population

Functional Characterization of PALB2 Variants of Uncertain Significance: Toward Cancer Risk and Therapy Response Prediction

Germline testing for homologous recombination repair genes—opportunities and challenges

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