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2.
Gastric Cancer ; 26(5): 653-666, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37249750

RESUMO

BACKGROUND: Germline CDH1 pathogenic or likely pathogenic variants cause hereditary diffuse gastric cancer (HDGC). Once a genetic cause is identified, stomachs' and breasts' surveillance and/or prophylactic surgery is offered to asymptomatic CDH1 carriers, which is life-saving. Herein, we characterized an inherited mechanism responsible for extremely early-onset gastric cancer and atypical HDGC high penetrance. METHODS: Whole-exome sequencing (WES) re-analysis was performed in an unsolved HDGC family. Accessible chromatin and CDH1 promoter interactors were evaluated in normal stomach by ATAC-seq and 4C-seq, and functional analysis was performed using CRISPR-Cas9, RNA-seq and pathway analysis. RESULTS: We identified a germline heterozygous 23 Kb CDH1-TANGO6 deletion in a family with eight diffuse gastric cancers, six before age 30. Atypical HDGC high penetrance and young cancer-onset argued towards a role for the deleted region downstream of CDH1, which we proved to present accessible chromatin, and CDH1 promoter interactors in normal stomach. CRISPR-Cas9 edited cells mimicking the CDH1-TANGO6 deletion display the strongest CDH1 mRNA downregulation, more impacted adhesion-associated, type-I interferon immune-associated and oncogenic signalling pathways, compared to wild-type or CDH1-deleted cells. This finding solved an 18-year family odyssey and engaged carrier family members in a cancer prevention pathway of care. CONCLUSION: In this work, we demonstrated that regulatory elements lying down-stream of CDH1 are part of a chromatin network that control CDH1 expression and influence cell transcriptome and associated signalling pathways, likely explaining high disease penetrance and very young cancer-onset. This study highlights the importance of incorporating scientific-technological updates and clinical guidelines in routine diagnosis, given their impact in timely genetic diagnosis and disease prevention.


Assuntos
Adenocarcinoma , Neoplasias Gástricas , Humanos , Adulto , Neoplasias Gástricas/patologia , Penetrância , Predisposição Genética para Doença , Caderinas/genética , Cromatina , Mutação em Linhagem Germinativa , Antígenos CD/genética
3.
J Med Genet ; 60(6): 557-567, 2023 06.
Artigo em Inglês | MEDLINE | ID: mdl-36270769

RESUMO

BACKGROUND: Patients with serrated polyposis syndrome (SPS) have multiple and/or large serrated colonic polyps and higher risk for colorectal cancer. SPS inherited genetic basis is mostly unknown. We aimed to identify new germline predisposition factors for SPS by functionally evaluating a candidate gene and replicating it in additional SPS cohorts. METHODS: After a previous whole-exome sequencing in 39 SPS patients from 16 families (discovery cohort), we sequenced specific genes in an independent validation cohort of 211 unrelated SPS cases. Additional external replication was also available in 297 SPS cases. The WNK2 gene was disrupted in HT-29 cells by gene editing, and WNK2 variants were transfected using a lentiviral delivery system. Cells were analysed by immunoblots, real-time PCR and functional assays monitoring the mitogen-activated protein kinase (MAPK) pathway, cell cycle progression, survival and adhesion. RESULTS: We identified 2 rare germline variants in the WNK2 gene in the discovery cohort, 3 additional variants in the validation cohort and 10 other variants in the external cohorts. Variants c.2105C>T (p.Pro702Leu), c.4820C>T (p.Ala1607Val) and c.6157G>A (p.Val2053Ile) were functionally characterised, displaying higher levels of phospho-PAK1/2, phospho-ERK1/2, CCND1, clonogenic capacity and MMP2. CONCLUSION: After whole-exome sequencing in SPS cases with familial aggregation and replication of results in additional cohorts, we identified rare germline variants in the WNK2 gene. Functional studies suggested germline WNK2 variants affect protein function in the context of the MAPK pathway, a molecular hallmark in this disease.


Assuntos
Polipose Adenomatosa do Colo , Pólipos do Colo , Neoplasias Colorretais , Humanos , Mutação em Linhagem Germinativa/genética , Polipose Adenomatosa do Colo/genética , Pólipos do Colo/genética , Genótipo , Neoplasias Colorretais/genética , Proteínas Serina-Treonina Quinases/genética
5.
Eur J Med Genet ; 65(5): 104475, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35283344

RESUMO

BACKGROUND: Patients and families with suspected, but genetically unexplained (unsolved) genetic tumour risk syndromes lack appropriate treatment and prevention, leading to preventable morbidity and mortality. To tackle this problem, patients from the European Reference Network on Genetic Tumour Risk Syndromes (ERN GENTURIS) are analysed in the European Commission's research project "Solving the unsolved rare diseases" (Solve-RD). The aim is to uncover known and novel cancer predisposing genes by reanalysing available whole-exome sequencing (WES) data of large cohorts in a combined manner, and applying a multidimensional omics approach. APPROACH: Around 500 genetically unsolved cases with suspected hereditary gastrointestinal tumour syndromes (polyposis, early-onset/familial colorectal cancer and gastric cancer) from multiple European centres are aimed to be included. Currently, clinical and germline WES data from 294 cases have been analysed. In addition, an extensive molecular profiling of gastrointestinal tumours from these patients is planned and deep learning techniques will be applied. The ambitious, multidisciplinary project is accompanied by a number of methodical, technical, and logistic challenges, which require the development and implementation of new analysis tools, the standardisation of bioinformatics pipelines, and strategies to exchange data and knowledge. RESULTS: and Outlook. The first re-analysis of 229 known and proposed cancer predisposition genes allowed solving 2-3% of previously unsolved GENTURIS cases. The integration of expert knowledge and new technologies will help to identify the genetic basis of additional unsolved cases within the ongoing project. The ERN GENTURIS approach might serve as a model for other genomic initiatives.


Assuntos
Neoplasias Colorretais , Síndromes Neoplásicas Hereditárias , Neoplasias Colorretais/genética , Predisposição Genética para Doença , Genômica , Humanos , Síndromes Neoplásicas Hereditárias/genética , Sequenciamento do Exoma
6.
Eur J Hum Genet ; 29(9): 1354-1358, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34075207

RESUMO

Hereditary diffuse gastric cancer (HDGC) is associated with germline deleterious variants in CDH1 and CTNNA1. The majority of HDGC-suspected patients are still genetically unresolved, raising the need for identification of novel HDGC predisposing genes. Under the collaborative environment of the SOLVE-RD consortium, re-analysis of whole-exome sequencing data from unresolved gastric cancer cases (n = 83) identified a mosaic missense variant in PIK3CA in a 25-year-old female with diffuse gastric cancer (DGC) without familial history for cancer. The variant, c.3140A>G p.(His1047Arg), a known cancer-related somatic hotspot, was present at a low variant allele frequency (18%) in leukocyte-derived DNA. Somatic variants in PIK3CA are usually associated with overgrowth, a phenotype that was not observed in this patient. This report highlights mosaicism as a potential, and understudied, mechanism in the etiology of DGC.


Assuntos
Classe I de Fosfatidilinositol 3-Quinases/genética , Mosaicismo , Neoplasias Gástricas/genética , Adulto , Feminino , Humanos , Mutação de Sentido Incorreto , Neoplasias Gástricas/patologia , Sequenciamento do Exoma
7.
Int J Mol Sci ; 21(22)2020 Nov 19.
Artigo em Inglês | MEDLINE | ID: mdl-33228212

RESUMO

To discover novel high-penetrant risk loci for hereditary colorectal cancer (hCRC) and polyposis syndromes many whole-exome and whole-genome sequencing (WES/WGS) studies have been performed. Remarkably, these studies resulted in only a few novel high-penetrant risk genes. Given this observation, the possibility and strategy to identify high-penetrant risk genes for hCRC and polyposis needs reconsideration. Therefore, we reviewed the study design of WES/WGS-based hCRC and polyposis gene discovery studies (n = 37) and provide recommendations to optimize discovery and validation strategies. The group of genetically unresolved patients is phenotypically heterogeneous, and likely composed of distinct molecular subtypes. This knowledge advocates for the screening of a homogeneous, stringently preselected discovery cohort and obtaining multi-level evidence for variant pathogenicity. This evidence can be collected by characterizing the molecular landscape of tumors from individuals with the same affected gene or by functional validation in cell-based models. Together, the combined approach of a phenotype-driven, tumor-based candidate gene search might elucidate the potential contribution of novel genetic predispositions in genetically unresolved hCRC and polyposis.


Assuntos
Proteína da Polipose Adenomatosa do Colo/genética , Polipose Adenomatosa do Colo/genética , Neoplasias Colorretais Hereditárias sem Polipose/genética , Loci Gênicos , PTEN Fosfo-Hidrolase/genética , Polipose Adenomatosa do Colo/diagnóstico , Polipose Adenomatosa do Colo/metabolismo , Polipose Adenomatosa do Colo/patologia , Proteína da Polipose Adenomatosa do Colo/metabolismo , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais Hereditárias sem Polipose/diagnóstico , Neoplasias Colorretais Hereditárias sem Polipose/metabolismo , Neoplasias Colorretais Hereditárias sem Polipose/patologia , Exoma , Estudos de Associação Genética , Predisposição Genética para Doença , Genótipo , Humanos , Proteína 1 Homóloga a MutL/genética , Proteína 1 Homóloga a MutL/metabolismo , Proteína 2 Homóloga a MutS/genética , Proteína 2 Homóloga a MutS/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Penetrância , Fenótipo , Sequenciamento do Exoma
8.
Kidney Int ; 97(2): 370-382, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31874800

RESUMO

Autosomal dominant polycystic kidney disease (ADPKD) is an inherited, progressive nephropathy accounting for 4-10% of end stage renal disease worldwide. PKD1 and PKD2 are the most common disease loci, but even accounting for other genetic causes, about 7% of families remain unresolved. Typically, these unsolved cases have relatively mild kidney disease and often have a negative family history. Mosaicism, due to de novo mutation in the early embryo, has rarely been identified by conventional genetic analysis of ADPKD families. Here we screened for mosaicism by employing two next generation sequencing screens, specific analysis of PKD1 and PKD2 employing long-range polymerase chain reaction, or targeted capture of cystogenes. We characterized mosaicism in 20 ADPKD families; the pathogenic variant was transmitted to the next generation in five families and sporadic in 15. The mosaic pathogenic variant was newly discovered by next generation sequencing in 13 families, and these methods precisely quantified the level of mosaicism in all. All of the mosaic cases had PKD1 mutations, 14 were deletions or insertions, and 16 occurred in females. Analysis of kidney size and function showed the mosaic cases had milder disease than a control PKD1 population, but only a few had clearly asymmetric disease. Thus, in a typical ADPKD population, readily detectable mosaicism by next generation sequencing accounts for about 1% of cases, and about 10% of genetically unresolved cases with an uncertain family history. Hence, identification of mosaicism is important to fully characterize ADPKD populations and provides informed prognostic information.


Assuntos
Rim Policístico Autossômico Dominante , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Mosaicismo , Mutação , Rim Policístico Autossômico Dominante/diagnóstico , Rim Policístico Autossômico Dominante/genética , Canais de Cátion TRPP/genética
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