RNA Analysis Identifies Pathogenic Duplications in MSH2 in Patients With Lynch Syndrome.
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ABSTRACT:
SUBMITTER: Conner BR
PROVIDER: S-EPMC10308853 | biostudies-literature | 2019 May
REPOSITORIES: biostudies-literature
Publications
RNA Analysis Identifies Pathogenic Duplications in MSH2 in Patients With Lynch Syndrome.
Conner Blair R BR Hernandez Felicia F Souders Beth B Landrith Tyler T Boland C Richard CR Karam Rachid R
Gastroenterology 20190130 6
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Project description:AimTo investigate the protein expression profile of mismatch repair (MMR) genes in suspected cases of Lynch syndrome and to characterize the associated germline mutations.MethodsImmunohistochemical analysis of tumor samples was performed to determine the protein expression profile of MMR protein. Germline mutation screening was carried out on peripheral blood samples. The entire exon regions of MLH1 and MSH2 genes were amplified by polymerase chain reaction, screened by denaturing high performance liquid chromatography (dHPLC) and analyzed by DNA sequencing to characterize the germline mutations.ResultsThree out of 34 tissue samples (8.8%) and four out of 34 tissue samples (11.8%) showed loss of nuclear staining by immunohistochemistry, indicating the absence of MLH1 and MSH2 protein expression in carcinoma cells, respectively. dHPLC analysis followed by DNA sequencing showed these samples to have germline mutations of MSH2 gene. However, no deleterious mutations were identified in any of the 19 exons or coding regions of MLH1 gene, but we were able to identify MLH1 promoter polymorphism, -93G > A (rs1800734), in 21 out of 34 patients (61.8%). We identified one novel mutation, transversion mutation c.2005G > C, which resulted in a missense mutation (Gly669Arg), a transversion mutation in exon 1, c.142G > T, which resulted in a nonsense mutation (Glu48Stop) and splice-site mutation, c.2006-6T > C, which was adjacent to exon 13 of MSH2 gene.ConclusionGermline mutations were identified in four Malaysian Lynch syndrome patients. Immunohistochemical analysis of tumor tissue proved to be a good pre-screening test before proceeding to germline mutation analysis of DNA MMR genes.
| S-EPMC3286145 | biostudies-literature
Project description:Lynch syndrome (LS) is the most common hereditary colorectal cancer (CRC) syndrome, characterized by germline pathogenic variants in mismatch repair (MMR)-related genes that lead to microsatellite instability. Patients who meet the clinical criteria for LS and MMR deficiency and without any identified germline pathogenic variants are frequently considered to have Lynch-like syndrome (LLS). These patients have a higher risk of CRC and extracolonic tumors, and little is known about their underlying genetic causes. We investigated the germline spectrum of LLS patients through whole-exome sequencing (WES). A total of 20 unrelated patients with MMR deficiency who met the clinical criteria for LS and had no germline variant were subjected to germline WES. Variant classification was performed according to the American College of Medical Genetics and Genomics (ACMG) criteria. Pathogenic/likely pathogenic variants were identified in 35% of patients in known cancer genes such as MUTYH and ATM. Besides this, rare and potentially pathogenic variants were identified in the DNA repair gene POLN and other cancer-related genes such as PPARG, CTC1, DCC and ALPK1. Our study demonstrates the germline mutational status of LLS patients, a population at high risk of colorectal cancer.
| S-EPMC9454535 | biostudies-literature
Project description:Lynch syndrome (LS) is a hereditary cancer-predisposing syndrome associated most frequently with epithelial tumors, particularly colorectal (CRC) and endometrial carcinomas (EC). The aim of this study was to investigate the relationship between sarcomas and LS by performing clinical and molecular characterization of patients presenting co-occurrence of sarcomas and tumors from the LS spectrum. We identified 27 patients diagnosed with CRC, EC, and other LS-associated tumors who had sarcomas in the same individuals or families. Germline genetic testing, mismatch repair (MMR) protein immunohistochemistry, microsatellite instability (MSI), and other molecular analyses were performed. Five LS patients presenting personal or family history of sarcomas were identified (3 MSH2 carriers and 2 MLH1), with 2 having Muir-Torre phenotypes. For two MSH2 carriers we confirmed the etiology of the sarcomas (one liposarcoma and two osteosarcomas) as LS-related, since the tumors were MSH2/MSH6-deficient, MSI-high, or presented a truncated MSH2 transcript. Additionally, we reviewed 43 previous reports of sarcomas in patients with LS, which revealed a high frequency (58%) of MSH2 alterations. In summary, sarcomas represent a rare clinical manifestation in patients with LS, especially in MSH2 carriers, and the analysis of tumor biological characteristics can be useful for definition of tumor etiology and novel therapeutic options.
| S-EPMC7408879 | biostudies-literature
Project description:Individuals with Lynch syndrome (LS), have an increased risk of developing cancer. Common genetic variants of telomerase reverse transcriptase (TERT) have been associated with a wide range of cancers, including colorectal cancer (CRC) in LS. We combined genotype data from 1881 LS patients, carrying pathogenic variants in MLH1, MSH2 or MSH6, for rs2075786 (G>A, intronic variant), 1207 LS patients for rs2736108 (C>T, upstream variant) and 1201 LS patients for rs7705526 (C>A, intronic variant). The risk of cancer was estimated by heterozygous/homozygous odds ratio (OR) with mixed-effects logistic regression to adjust for gene/gender/country of sample origin considering family identity. The AA genotype of SNP rs2075786 is associated with 85% higher odds at developing cancer compared to GG genotype in MSH2 pathogenic variant carriers (p = 0.0160). Kaplan-Meier analysis also shows an association for rs2075786; the AA allele for MSH2 variant carriers confers risk for earlier diagnosis of LS cancer (log-rank p = 0.0011). We report a polymorphism in TERT to be a possible modifier of disease risk in MSH2 pathogenic variant carriers. The rs2075786 SNP in TERT is associated with a differential risk of developing cancer for MSH2 pathogenic variant carriers. Use of this information has the potential to personalise screening protocols for LS patients.
| S-EPMC8166931 | biostudies-literature
Project description:Background. Lynch Syndrome (LS) is characterized by germline mutations in the DNA mismatch repair (MMR) genes MLH1, MSH2, MSH6, and PMS2. This syndrome is inherited in an autosomal dominant pattern and is characterized by early onset colorectal cancer (CRC) and extracolonic tumors. The aim of this study was to identify mutations in MMR genes in three Mexican patients with LS. Methods. Immunohistochemical analysis was performed as a prescreening method to identify absent protein expression. PCR, Denaturing High Performance Liquid Chromatography (dHPLC), and Sanger sequencing complemented the analysis. Results. Two samples showed the absence of nuclear staining for MLH1 and one sample showed loss of nuclear staining for MSH2. The mutations found in MLH1 gene were c.2103+1G>C in intron 18 and compound heterozygous mutants c.1852_1854delAAG (p.K618del) and c.1852_1853delinsGC (p.K618A) in exon 16. In the MSH2 gene, we identified mutation c.638dupT (p.L213fs) in exon 3. Conclusions. This is the first report of mutations in MMR genes in Mexican patients with LS and these appear to be novel.
| S-EPMC4877485 | biostudies-literature
Project description:BackgroundApproximately 5% of colorectal cancers (CRCs) are hereditary. Lynch syndrome (LS), also known as hereditary nonpolyposis colorectal cancer (HNPCC), is the most common form of recognized hereditary CRC. Although Iran, as a developing country, has a high incidence of CRC, the spectrum of variants has yet to be thoroughly investigated.AimsThis study aimed to investigate pathogenic and non-pathogenic variants in MLH1 and MSH2 genes in Iranian patients with suspected Lynch syndrome (sLS).Methods and resultsIn the present study, 25 peripheral blood samples were collected from patients with sLS and high microsatellite instability (MSI-H). After DNA extraction, all samples underwent polymerase chain reaction and Sanger sequencing to identify the variants in the exons of MLH1 and MSH2 genes. The identified variants were interpreted using prediction tools, and were finally reported under ACMG guidelines. In our study population, 13 variants were found in the MLH1 gene and 8 in the MSH2 gene. Interestingly, 7 of the 13 MLH1 variants and 3 of the 8 MSH2 variants were novel, whereas the remaining variants were previously reported or available in databases. In addition, some patients with sLS did not have variants in the exons of the MLH1 and MSH2 genes. The variants detected in the MLH1 and MSH2 genes had specific characteristics regarding the number, area of occurrence, and their relationship with demographic and clinicopathologic features.ConclusionOverall, our results suggest that analysis of MLH1 and MSH2 genes alone is insufficient in the Iranian population, and more comprehensive tests are recommended for detecting LS.
| S-EPMC10809271 | biostudies-literature
Project description:Lynch syndrome, a heritable form of cancer predisposition, is caused by germline mutations within genes of the DNA mismatch repair family, and can be rapidly identified in young onset cancer patients through the detection of loss of expression of at least one of these genes in tumour samples. To date, such causative mutations have only been identified within exonic and splice site regions. Though this approach has been successful in the majority of families, a considerable number remain in which no mutation has been found. To address this situation, we used an alternative mutation discovery procedure which involved haplotype analysis of the locus containing the gene lost in the tumour and delineation of segregating haplotypes, followed by an investigation of splicing aberrations to uncover cryptic splice sites which lay outside the genomic regions routinely examined for mutations. In this report, we show that an intronic mutation 478 bp upstream of exon 2 in the MSH2 gene causes Lynch syndrome through creation of a novel splice donor site with subsequent pseudoexon activation, thus highlighting the need for more extensive sequencing approaches in families where routine procedures fail to find a mutation.
| S-EPMC4580736 | biostudies-literature
Project description:A subset of genetic variants found through screening of patients with hereditary breast and ovarian cancer syndrome (HBOC) and Lynch syndrome impact RNA splicing. Through target enrichment of the transcriptome, it is possible to perform deep-sequencing and to identify the different and even rare mRNA isoforms. A targeted RNA-seq approach was used to analyse the naturally-occurring splicing events for a panel of 8 breast and/or ovarian cancer susceptibility genes (BRCA1, BRCA2, RAD51C, RAD51D, PTEN, STK11, CDH1, TP53), 3 Lynch syndrome genes (MLH1, MSH2, MSH6) and the fanconi anaemia SLX4 gene, in which monoallelic mutations were found in non-BRCA families. For BRCA1, BRCA2, RAD51C and RAD51D the results were validated by capillary electrophoresis and were compared to a non-targeted RNA-seq approach. We also compared splicing events from lymphoblastoid cell-lines with those from breast and ovarian fimbriae tissues. The potential of targeted RNA-seq to detect pathogenic changes in RNA-splicing was validated by the inclusion of samples with previously well characterized BRCA1/2 genetic variants. In our study, we update the catalogue of normal splicing events for BRCA1/2, provide an extensive catalogue of normal RAD51C and RAD51D alternative splicing, and list splicing events found for eight other genes. Additionally, we show that our approach allowed the identification of aberrant splicing events due to the presence of BRCA1/2 genetic variants and distinguished between complete and partial splicing events. In conclusion, targeted-RNA-seq can be very useful to classify variants based on their putative pathogenic impact on splicing.
| S-EPMC6635756 | biostudies-literature
Project description:BackgroundA large proportion of patients with Lynch syndrome (LS) have MSH2 abnormalities, but genotype-phenotype studies of MSH2 mutations in LS are still lacking. The aim of this study was to comprehensively analyze the clinicopathological characteristics and molecular basis of colorectal cancer (CRC) in patients with uncommon MSH2 cytoplasmic expression.MethodsWe retrospectively reviewed 4195 consecutive cases of CRC patients diagnosed between January 2015 and December 2017 at the Cancer Hospital Chinese Academy of Medical Sciences. Of the 4195 patients with CRC, 69 were indicated to have abnormal MSH2 expression through tumor immunohistochemical staining. Genetic tests, such as next-generation sequencing, large genomic rearrangement (LGR) analysis, microsatellite instability status analysis and genomic breakpoint analysis, were performed. Clinicopathological and molecular characteristics and clinical immunotherapy response were analyzed.ResultsForty-five of 69 patients were identified to have LS with pathogenic germline mutations in MSH2 and/or EPCAM. Of these LS patients, 26.7% were confirmed to harbor large genomic rearrangements (LGRs). Of note, three tumors from two unrelated family pedigrees exhibited a rare cytoplasmic MSH2 staining pattern that was found in LS patients with EPCAM/MSH2 deletions. RNA analysis showed that two novel mRNA fusions of EPCAM and MSH2 resulted in the predicted protein fusion with MSH2 cytoplasmic localization. Analyses of genomic breakpoints indicated that two novel deletions of EPCAM and MSH2 originated from Alu repeat-mediated recombination events. Our study also provides clinical evidence for the beneficial effect of the PD-1 inhibitor pembrolizumab for CRC patients that exhibit cytoplasmic MSH2 staining.ConclusionOur study demonstrates that the rare cytoplasmic MSH2 staining pattern should be fully recognized by pathologists and geneticists. Given the specific genotype-phenotype correlation in LS screening, we advocate that all CRC patients with cytoplasmic MSH2 staining in histology should be screened for LGRs of EPCAM and MSH2.
| S-EPMC8162378 | biostudies-literature