Open, multicentre study for the diagnosis of Lynch syndrome using foss sequencing platform in colorectal cancer patients with immunohistochemical indications of MMR protein deficiency
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ABSTRACT:
DISEASE(S): Colorectal Cancer
PROVIDER: 2688576 | ecrin-mdr-crc |
REPOSITORIES: ECRIN MDR
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Project description:The current algorithm for Lynch syndrome diagnosis is highly complex with multiple steps which can result in an extended time to diagnosis while depleting precious tumor specimens. Here we describe the analytical validation of a custom probe-based NGS tumor panel, TumorNext-Lynch-MMR, which generates a comprehensive genetic profile of both germline and somatic mutations that can accelerate and streamline the time to diagnosis and preserve specimen. TumorNext-Lynch-MMR can detect single nucleotide variants, small insertions and deletions in 39 genes that are frequently mutated in Lynch syndrome and colorectal cancer. Moreover, the panel provides microsatellite instability status and detects loss of heterozygosity in the five Lynch genes; MSH2, MSH6, MLH1, PMS2 and EPCAM. Clinical cases are described that highlight the assays ability to differentiate between somatic and germline mutations, precisely classify variants and resolve discordant cases.
| S-EPMC5945525 | biostudies-literature
Project description:Universal screening for Lynch syndrome in colorectal cancer is recommended, and immunohistochemistry for the mismatch repair proteins is commonly used. To reduce cost, some screen using only MSH6 and PMS2, with reflex to the partner stain if either are absent (two-stain method). An expression pattern revealing absent MSH2 and intact MSH6 is not expected, but could result in failed Lynch syndrome detection. We analyzed tumors with absent MSH2 but any degree of MSH6 expression to determine if the two-stain method could miss MSH2 mutations. One-thousand seven-hundred thirty colorectal cancer patients from the Ohio Colorectal Cancer Prevention Initiative underwent tumor screening using microsatellite instability and immunohistochemistry. The two-stain method was used for 1235 cases; staining for all four proteins was completed for 495 cases. The proportion of positive cells and staining intensity were reviewed for MSH6, as well as MSH2 when available. Patients with mismatch repair deficiency underwent next-generation sequencing of germline DNA for mismatch repair genes. If negative, tumor next-generation sequencing was performed to assess for somatic mutations. Overall, thirty-three (1.9%, 33/1730) MSH2-absent cases were identified. Of those, fourteen had no MSH6 expression but eight (0.5%, 8/1730) had ambiguous and eleven (0.6%, 11/1730) had convincing MSH6 expression that could have been interpreted as intact. Germline next-generation sequencing identified MSH2 mutations in 11/14 cases with absence of both stains, 7/8 cases with ambiguous MSH6 expression, and 9/11 cases with convincing MSH6 expression. All remaining cases, except one, had double somatic mutations. The two-stain method fails to detect some patients with Lynch syndrome: (1) significant staining weaker than the control may be incorrectly interpreted as intact MSH6, or (2) Weak or focal/patchy MSH6 can be retained with the absence of MSH2. Accordingly, we recommend the four-stain method be used for optimal Lynch syndrome screening detection.
| S-EPMC6800091 | biostudies-literature
Project description:Several studies have reported that, in Lynch syndrome resulting from mutations of the mismatch repair (MMR) genes, a CA repeat ?17 within the IGF1 promoter, SNPs within the xenobiotic metabolizing enzyme gene CYP1A1 and SNPs on 8q23.3 and 11q23.1 modify colorectal cancer (CRC) risk in MMR mutation carriers. We analysed the impact of these polymorphisms on CRC risk in 748 French MMR mutation carriers derived from 359 families. We also analysed the effect of the Novel 1 SNP (18q21), which has recently been shown to increase CRC risk in the general population. We observed a significant difference in the CRC-free survival time between males and females, between MSH2 and MSH6 mutation carriers and between MLH1 and MSH6, indicating that this series is representative of Lynch syndrome. In contrast, the univariate log-rank test, as well as multivariate Cox model analysis controlling for familial aggregation and mutated MMR gene, year of birth and gender showed that the polymorphic alleles tested were not associated with a significant CRC risk increase, neither on the entire sample nor among males and females. This discrepancy with previous reports might be explained both by the genetic heterogeneity between the different populations analysed and the allelic heterogeneity of the MMR mutations. We conclude that genotyping of these polymorphisms is not useful to evaluate CRC risk in MMR mutation carriers and to optimize their clinical follow-up.
| S-EPMC3172927 | biostudies-literature
Project description:MMR defects study in mCRPC using targeted DNA repair panel
| PRJEB27287 | ENA
Project description:Background: In ~50% of Lynch syndrome (LS)-suspected patients (also called Lynch-like syndrome, LLS), the causal mechanism for cancer predisposition remains unknown. Our aim was to elucidate the constitutional basis of mismatch repair (MMR) deficiency in LLS patients throughout a comprehensive genetic and epigenetic analysis. Methods: One hundred and fifteen LLS patients harboring MMR deficient tumors and no pathogenic germline MMR gene mutations were included in this study. Mutational analysis of 26 colorectal cancer associated genes was performed by using a customized multigene panel and massively parallel sequencing. Pathogenicity of MMR variants was assessed by mRNA analysis and multifactorial likelihood calculations. Genome-wide methylome analysis was perfomed by using the Infinium HumanMethylation450K BeadChip. Results: The multigene panel analysis revealed the presence of two MMR gene truncating mutations not found in previous analysis. Of a total of 15 MMR variants of unknown significance identified, five - present in 6 unrelated individuals- were reclassified as pathogenic. In addition, 13 predicted deleterious variants in other CRC-predisposing genes (MSH3, MUTYH, POLD1, APC, EPCAM, BUB1, FAN1, EXO1 or PSM1) were found in 12 probands. Methylome analysis detected one constitutional MLH1 epimutation in an individual diagnosed with CRC at age 42, but no additional differentially methylated regions were identified in LLS compared to LS patients or healthy individuals. Conclusions: In conclusion, the use of an ad-hoc designed gene panel combined with pathogenicity assessment of variants allows the identification of deleterious MMR mutations as well as new LLS candidate genes. Moreover, constitutional epimutations in non-LS-associated genes are not responsible for the MMR-deficient phenotype observed in LLS patients.
2019-03-09 | GSE128067 | GEO
Project description:Background: In ~50% of Lynch syndrome (LS)-suspected patients (also called Lynch-like syndrome, LLS), the causal mechanism for cancer predisposition remains unknown. Our aim was to elucidate the constitutional basis of mismatch repair (MMR) deficiency in LLS patients throughout a comprehensive genetic and epigenetic analysis. Methods: One hundred and fifteen LLS patients harboring MMR deficient tumors and no pathogenic germline MMR gene mutations were included in this study. Mutational analysis of 26 colorectal cancer associated genes was performed by using a customized multigene panel and massively parallel sequencing. Pathogenicity of MMR variants was assessed by mRNA analysis and multifactorial likelihood calculations. Genome-wide methylome analysis was perfomed by using the Infinium HumanMethylation450K BeadChip. Results: The multigene panel analysis revealed the presence of two MMR gene truncating mutations not found in previous analysis. Of a total of 15 MMR variants of unknown significance identified, five - present in 6 unrelated individuals- were reclassified as pathogenic. In addition, 13 predicted deleterious variants in other CRC-predisposing genes (MSH3, MUTYH, POLD1, APC, EPCAM, BUB1, FAN1, EXO1 or PSM1) were found in 12 probands. Methylome analysis detected one constitutional MLH1 epimutation in an individual diagnosed with CRC at age 42, but no additional differentially methylated regions were identified in LLS compared to LS patients or healthy individuals. Conclusions: In conclusion, the use of an ad-hoc designed gene panel combined with pathogenicity assessment of variants allows the identification of deleterious MMR mutations as well as new LLS candidate genes. Moreover, constitutional epimutations in non-LS-associated genes are not responsible for the MMR-deficient phenotype observed in LLS patients.
2019-03-09 | GSE128064 | GEO
Project description:We have developed a multi-analyte fluid-phase protein array technology termed high-throughput immunophenotyping using transcription (HIT). This method employs a panel of monoclonal antibodies, each tagged with a unique oligonucleotide sequence that serves as a molecular barcode. After staining a sample, T7 polymerase amplifies the tags which are then hybridized to a DNA microarray for indirect measurement of each analyte. Here we screened 90 antibodies directed against a panel of cell surface markers as well as 4 isotype controls to compare resting human naive CD4+ T cells versus CD4+ T cells activated for 48 h with anti-CD3/anti-CD28 coated beads. Keywords: protein profiling, response to stimulus
2008-03-03 | GSE10665 | GEO
Project description:Lynch syndrome (LS), also known as hereditary nonpolyposis colorectal cancer (HNPCC), is a disorder caused by an autosomal dominant heterozygous germline mutation in one of the DNA mismatch repair (MMR) genes. Individuals with LS are at an increased risk of developing colorectal and extracolonic cancers, such as endometrial, small bowel, or ovarian. In this review, the mutations involved with LS and their diagnostic methods are described and compared, as are their current uses in clinical decision making. Nowadays, LS diagnosis is based on a review of family medical history, and when necessary, microsatellite instability (MSI) or/and immunohistochemistry (IHC) analyses should be performed. In the case of a lack of MMR protein expression (dMMR) or MSI-H (MSI-High) detection in tumor tissue, molecular genetic testing can be undertaken. More and more genetic testing for LS is based mainly on next-generation sequencing (NGS) and multiplex ligation-dependent probe amplification (MLPA), which provide better and quicker information about the molecular profile of patients as well as individuals at risk. Testing based on these two methods should be the standard and commonly used. The identification of individuals with mutations provides opportunities for the detection of cancer at an early stage as well as the introduction of proper, more effective treatment, which will result in increased patient survival and reduced costs of medical care.
| S-EPMC7600989 | biostudies-literature
Project description:We have developed a multi-analyte fluid-phase protein array technology termed high-throughput immunophenotyping using transcription (HIT). This method employs a panel of monoclonal antibodies, each tagged with a unique oligonucleotide sequence that serves as a molecular barcode. After staining a sample, T7 polymerase amplifies the tags which are then hybridized to a DNA microarray for indirect measurement of each analyte. Here we screened 90 antibodies directed against a panel of cell surface markers as well as 4 isotype controls to compare resting human naive CD4+ T cells versus CD4+ T cells activated for 48 h with anti-CD3/anti-CD28 coated beads. Keywords: protein profiling, response to stimulus Two-condition experiment, activated versus resting cells. Biological replicates: 3 normal human donors, each donor served as an internal control. One replicate per array. One of the biological replicates was performed as a dye-swap to monitor dye-bias. We also performed a self-self comparison between activated cells from two different donors.
2010-05-26 | E-GEOD-10665 | biostudies-arrayexpress
Project description:Today most patients with Lynch syndrome (LS) survive their first cancer. There is limited information on the incidences and outcome of subsequent cancers. The present study addresses three questions: (i) what is the cumulative incidence of a subsequent cancer; (ii) in which organs do subsequent cancers occur; and (iii) what is the survival following these cancers?Information was collated on prospectively organised surveillance and prospectively observed outcomes in patients with LS who had cancer prior to inclusion and analysed by age, gender and genetic variants.1273 patients with LS from 10 countries were followed up for 7753 observation years. 318 patients (25.7%) developed 341 first subsequent cancers, including colorectal (n=147, 43%), upper GI, pancreas or bile duct (n=37, 11%) and urinary tract (n=32, 10%). The cumulative incidences for any subsequent cancer from age 40 to age 70 years were 73% for pathogenic MLH1 (path_MLH1), 76% for path_MSH2 carriers and 52% for path_MSH6 carriers, and for colorectal cancer (CRC) the cumulative incidences were 46%, 48% and 23%, respectively. Crude survival after any subsequent cancer was 82% (95% CI 76% to 87%) and 10-year crude survival after CRC was 91% (95% CI 83% to 95%).Relative incidence of subsequent cancer compared with incidence of first cancer was slightly but insignificantly higher than cancer incidence in patients with LS without previous cancer (range 0.94-1.49). The favourable survival after subsequent cancers validated continued follow-up to prevent death from cancer. The interactive website http://lscarisk.org was expanded to calculate the risks by gender, genetic variant and age for subsequent cancer for any patient with LS with previous cancer.
| S-EPMC5561364 | biostudies-other