MSH2 Research Articles

Frequency of Pathogenic Germline Mutations in Early and Late Onset Familial Breast Cancer Patients Using Multi-Gene Panel Sequencing: An Egyptian Study.

Precision oncology has been increasingly used in clinical practice and rapidly evolving in the oncology field. Thus, this study was performed to assess the frequency of germline mutations in early and late onset familial breast cancer (BC) Egyptian patients using multi-gene panel sequencing to better understand the contribution of the inherited germline mutations in BC predisposition. Moreover, to determine the actionable deleterious mutations associated with familial BC that might be used as biomarker for early cancer detection. Whole blood samples were collected from 101 Egyptian patients selected for BC family history, in addition to 50 age-matched healthy controls. A QIAseq targeted DNA panel (human BC panel) was used to assess the frequency of germline mutations. A total of 58 patients (57.4%) out of 101 were found to have 27 deleterious germline mutations in 11 cancer susceptibility genes. Of them, 32 (31.6%) patients carried more than one pathogenic mutation and each one carried at least one pathogenic mutation. The major genes harboring the pathogenic mutations were: ATM, BRCA2, BRCA1, VHL, MSH6, APC, CHEK2, MSH2, MEN1, PALB2, and MUTYH. Thirty-one patients (30.6%) had BRCA2 mutations and twenty (19.8%) had BRCA1 mutations. Our results showed that exon 10 and exon 11 harbored 3 and 5 mutations, respectively, in BRCA1 and BRCA2 genes. Our analysis also revealed that the VHL gene significantly co-occurred with each of the BRCA2 gene (p = 0.003, event ratio 11/21), the MSH2 gene (p = 0.01, 4/10), the CHEK2 gene (p = 0.02, 4/11), and the MSH6 gene (p = 0.04, 4/12). In addition, the APC gene significantly co-occurred with the MSH2 gene (p = 0.01, 3/7). Furthermore, there was a significant mutually exclusive event between the APC gene and the ATM gene (p = 0.04, 1/36). Interestingly, we identified population specific germline mutations in genes showing potentials for targeted therapy to meet the need for incorporating precision oncology into clinical practice. For example, the mutations identified in the ATM, APC, and MSH2 genes. Multi-gene panel sequencing was used to detect the deleterious mutations associated with familial BC, which in turns mitigate the essential need for implementing next generation sequencing technologies in precision oncology to identify cancer predisposing genes. Moreover, identifying DNA repair gene mutations, with focus on non-BRCA genes, might serve as candidates for targeted therapy and will be increasingly used in precision oncology.

Logistic Regression Modeling with A Bayesian Approach to The Risk Factors of Colorectal Cancer Patients

This research aims to create a model, determine the factors that affect the probability of someone suffering from colorectal cancer also design a web application to do prediction using the probability logit function which has been made. There are 4 independent variables and 1 dependent variable which are used in this research. The independent variables used in this research are heredity-pedigree value, age, Adenomatous Polyposis Coli, and MutS Homolog 2. And the dependent variable used in this research is someone’s status of suffering from colorectal cancer symbolized with Y. The stages of designing a web application include making use case diagrams, use case descriptions, activity diagrams, sequence diagrams, class diagrams, entity relationship diagrams, and designing the application interface. The results of this research will be a logit probability model which can be used to calculate the probability value of someone suffering from colorectal cancer. Model constructed using significant 3 independent variables which are heredity-pedigree, age, and Adenomatous Polyposis Coli.

Cancer prevention in cancer predisposition syndromes: A protocol for testing the feasibility of building a hereditary cancer research registry and nurse navigator follow up model.

Monogenic, high penetrance syndromes, conferring an increased risk of malignancies in multiple organs, are important contributors to the hereditary burden of cancer. Early detection and risk reduction strategies in patients with a cancer predisposition syndrome can save their lives. However, despite evidence supporting the benefits of early detection and risk reduction strategies, most Canadian jurisdictions have not implemented programmatic follow up of these patients. In our study site in the province of Newfoundland and Labrador (NL), Canada, there is no centralized, provincial registry of high-risk individuals. There is no continuity or coordination of care providing cancer genetics expertise and no process to ensure that patients are referred to the appropriate specialists or risk management interventions. This paper describes a study protocol to test the feasibility of obtaining and analyzing patient risk management data, specifically patients affected by hereditary breast ovarian cancer syndrome (HBOC; BRCA 1 and BRCA 2 genes) and Lynch syndrome (LS; MLH1, MSH2, MSH6, and PMS2 genes). Through a retrospective cohort study, we will describe these patients' adherence to risk management guidelines and test its relationship to health outcomes, including cancer incidence and stage. Through a qualitative interviews, we will determine the priorities and preferences of patients with any inherited cancer mutation for a follow up navigation model of risk management. Study data will inform a subsequent funding application focused on creating and evaluating a research registry and follow up nurse navigation model. It is not currently known what proportion of cancer mutation carriers are receiving care according to guidelines. Data collected in this study will provide clinical uptake and health outcome information so gaps in care can be identified. Data will also provide patient preference information to inform ongoing and planned research with cancer mutation carriers.

Saturation-scale functional evidence supports clinical variant interpretation in Lynch syndrome

BackgroundLynch syndrome (LS) is a cancer predisposition syndrome affecting more than 1 in every 300 individuals worldwide. Clinical genetic testing for LS can be life-saving but is complicated by the heavy burden of variants of uncertain significance (VUS), especially missense changes.ResultTo address this challenge, we leverage a multiplexed analysis of variant effect (MAVE) map covering >94% of the 17,746 possible missense variants in the key LS gene MSH2. To establish this map’s utility in large-scale variant reclassification, we overlay it on clinical databases of >15,000 individuals with LS gene variants uncovered during clinical genetic testing. We validate these functional measurements in a cohort of individuals with paired tumor-normal test results and find that MAVE-based function scores agree with the clinical interpretation for every one of the MSH2 missense variants with an available classification. We use these scores to attempt reclassification for 682 unique missense VUS, among which 34 scored as deleterious by our function map, in line with previously published rates for other cancer predisposition genes. Combining functional data and other evidence, ten missense VUS are reclassified as pathogenic/likely pathogenic, and another 497 could be moved to benign/likely benign. Finally, we apply these functional scores to paired tumor-normal genetic tests and identify a subset of patients with biallelic somatic loss of function, reflecting a sporadic Lynch-like Syndrome with distinct implications for treatment and relatives’ risk.ConclusionThis study demonstrates how high-throughput functional assays can empower scalable VUS resolution and prospectively generate strong evidence for variant classification.

Loss of mismatch repair promotes a direct selective advantage in human stem cells.

Lynch syndrome (LS) is the most common hereditary form of colon cancer, resulting from a germline mutation in a DNA mismatch repair (MMR) gene. Loss of MMR in cells establishes a mutator phenotype, which may underlie its link to cancer. Acquired downstream mutations that provide the cell a selective advantage would contribute to tumorigenesis. It is unclear, however, whether loss of MMR has other consequences that would directly result in a selective advantage. We found that knockout of the MMR gene MSH2 results in an immediate survival advantage in human stem cells grown under standard cell culture conditions. This advantage results, in part, from an MMR-dependent response to oxidative stress. We also found that loss of MMR gives rise to enhanced formation and growth of human colonic organoids. These results suggest that loss of MMR may affect cells in ways beyond just increasing mutation frequency that could influence tumorigenesis.

Differential operation of MLH1/MSH2 and FANCD2 crosstalk in chemotolerant bladder carcinoma: a clinical and therapeutic intervening study.

We aimed to understand the crosstalk between mismatch repair (MMR) and FA-BRCA pathway in primary bladder carcinoma (BlCa) samples as well as in chemotolerant cell line. We analysed the genetic alterations of MLH1 and MSH2 (MMR-related genes) and after that we correlated it with the nuclear translocation of FANCD2 protein. Next, we evaluated this crosstalk in T24 BlCa cell line in response to doxorubicin treatment. In primary BlCa tumors, infrequent genetic deletion (17-20%) but frequent promoter methylation (28-55%) of MLH1 and MSH2 was observed, where MLH1 was significantly (p < 0.05) more methylated among the early staged samples (NMIBC). However, MSH2 was significantly more altered among the NMIBC samples, signifying the importance of MMR pathway during the early pathogenesis of the disease. Furthermore, BlCa samples with underexpressed MLH1/MSH2 protein possessed cytoplasmic FANCD2 protein; encouraging that inefficiency of MMR proteins might restrict FANCD2 nuclear translocation. Next, we analysed publicly available data in GEO2R tool where we observed that in response to chemotherapeutic drugs, expression of MLH1, MSH2 and FANCD2 were diminishing. Validating this result in doxorubicin tolerant T24 cells, we found that expression of MLH1 and MSH2 was gradually decreased with increasing dose of doxorubicin. Interestingly, FANCD2 mono-ubiquitination (L-form) was also reduced in chemotolerant T24 cells. The crosstalk between MMR and FA-BRCA pathway was substantiated in the primary BlCa tumors. Further, in response to doxorubicin, this crosstalk was found to be hampered due to under-expression of MLH1 and MSH2 gene, thereby rendering chemotolerance.

Lynch syndrome, molecular mechanisms and variant classification.

Patients with the heritable cancer disease, Lynch syndrome, carry germline variants in the MLH1, MSH2, MSH6 and PMS2 genes, encoding the central components of the DNA mismatch repair system. Loss-of-function variants disrupt the DNA mismatch repair system and give rise to a detrimental increase in the cellular mutational burden and cancer development. The treatment prospects for Lynch syndrome rely heavily on early diagnosis; however, accurate diagnosis is inextricably linked to correct clinical interpretation of individual variants. Protein variant classification traditionally relies on cumulative information from occurrence in patients, as well as experimental testing of the individual variants. The complexity of variant classification is due to (1) that variants of unknown significance are rare in the population and phenotypic information on the specific variants is missing, and (2) that individual variant testing is challenging, costly and slow. Here, we summarise recent developments in high-throughput technologies and computational prediction tools for the assessment of variants of unknown significance in Lynch syndrome. These approaches may vastly increase the number of interpretable variants and could also provide important mechanistic insights into the disease. These insights may in turn pave the road towards developing personalised treatment approaches for Lynch syndrome.

Prevalence and Genotype-Phenotype Correlation of Lynch Syndrome in a Selected High-Risk Cohort from Qatar's Population.

Lynch syndrome (LS) is the most common cause of hereditary colorectal cancers (CRC) and is associated with an increased risk for ovarian and endometrial cancers. There is lack of knowledge on the epidemiology of LS in the non-Caucasian populations especially in Qatar. The aim of this retrospective study is to explore the prevalence of LS in a selected high-risk cohort in the State of Qatar in addition to investigating the frequency and genotype-phenotype correlation associated with mismatch repair genes pathogenic variants. Retrospective review of medical records of 31 individuals with LS, 20 affected with colorectal cancer and 11 unaffected with family history of cancers, referred from January 2017 until August 2020. The prevalence of LS among affected and unaffected patients is 22% (20/92) and 2.2% respectively. Among affected individuals, MLH1 and MSH2 genes were highly frequent while for unaffected individuals, a recurrent PMS2 pathogenic variant was reported in several related individuals suggesting a tribal effect. This study highlights the epidemiology of LS in high-risk cohort in Qatar which helps to provide recommendations on genetic testing, and personalize surveillance and management programs.

Recurrent posterior fossa group A (PFA) ependymoma in a young child with constitutional mismatch repair deficiency (CMMRD).

Recurrent posterior fossa group A (PFA) ependymoma in a young child with constitutional mismatch repair deficiency (CMMRD).

Case report: Undifferentiated sarcoma with multiple tumors involved in Lynch syndrome: Unexpected favorable outcome to sintilimab combined with chemotherapy

Patients with Lynch syndrome are at an increased risk of developing simultaneous or metachronous tumors, while sarcomas have been occasionally reported. Sarcomas are generally not considered part of the common Lynch syndrome tumor spectrum. However, more and more studies and case reports suggested that sarcoma could be a rare clinical manifestation of Lynch syndrome, leading to new treatment strategies for sarcoma. We report the case of a 74-year-old male patient with Lynch syndrome who had rectal mucinous adenocarcinoma and prostate adenocarcinoma and then developed undifferentiated sarcoma of the left neck two years later. Mismatch repair deficiency (dMMR) was confirmed by immunohistochemical staining for the mismatch repair proteins MSH2, MSH6, MLH1 and PMS2. The result of polymerase chain reaction (PCR) microsatellite instability (MSI) testing of sarcoma showed high-level microsatellite instability (MSI-H). Additionally, a pathogenic germline mutation in MSH2 (c.2459-12A>G) was detected by next-generation sequencing (NGS). Taking into account HE morphology, immunohistochemical phenotype, MSI status, NGS result, medical history and germline MSH2 gene mutation, the pathological diagnosis of left neck biopsy tissue was Lynch syndrome related undifferentiated sarcoma with epithelioid morphology. The patient has been receiving immunotherapy (sintilimab) combined with chemotherapy (tegafur, gimeracil and oteracil potassium capsules) and currently has stable disease. We also reviewed the literature to understand the association between sarcoma and Lynch syndrome. Sarcoma may now be considered a rare clinical manifestation of Lynch syndrome. Attention and awareness about the association between Lynch syndrome and sarcoma need to be increased. Therefore, timely detection of MMR proteins and validation at the gene level for suspicious patients are the keys to avoiding missed or delayed diagnosis and to identifying patients suited for immunotherapy, which may also help to provide appropriate genetic counseling and follow-up management for patients.

EXTH-95. UNCOVERING THERAPEUTIC VULNERABILITIES IN MISMATCH REPAIR-DEFICIENT GLIOMAS

Abstract Approximately 25% of recurrent gliomas exhibit hypermutation, which is most often acquired in chemotherapy sensitive gliomas post-treatment with standard of care alkylating agent temozolomide, Intriguingly, nearly all of these recurrent hypermutant tumors harbor deficiencies in mismatch repair (MMR) genes. Unlike other MMR-deficient cancers, hypermutant gliomas do not exhibit detectable microsatellite instability (MSI) at the population level, and do not share similar dependencies. Thus, strategies to therapeutically target MMR-deficient and hypermutant gliomas are urgently needed, and likely to impact many patients. To evaluate vulnerabilities associated with MMR-deficient gliomas, we have generated isogenic MMR-deficient models by systematic ablation of core MMR genes MSH2, MSH6, MLH1, and PMS2, in multiple patient-derived glioma cell lines using a robust all-in-one CRISPR-Cas9 system. We characterized these isogenic MMR-deficient glioma lines in comparison to respective MMR-proficient control line by performing differential gene expression analysis and subsequent gene set enrichment analyses. Our analyses reveal in an unbiased fashion an enrichment of several hallmark gene sets including cell cycle control and DNA repair in the MMR-deficient cell lines. We performed a CRISPR-Cas9 knockout screen in these MMR deficient and hypermutant models and identified candidate genes involved in DNA repair, cell cycle, RNA metabolism and other pathways as preferential dependencies in the MMR-deficient glioma cells. We also screened multiple MMR-deficient isogenic models against the high-throughput drug repurposing (REPO) chemical library and our preliminary results reveal a number of compounds from a variety of drug classes that selectively kill the MMR-deficient glioma cells, indicating that the loss of MMR confers differential dependencies to these small molecule inhibitors. The candidate drugs and genes identified from these high-throughput assays are currently being subjected to further validation, and they possibly will help us identify novel therapeutic strategies or improve existing therapeutic strategies to target recurrent MMR-deficient gliomas.

EPCO-11. GATEKEEPER INACTIVATION DRIVES TUMOR PROGRESSION TO GRADE IV ASTROCYTOMA

Abstract IDH-mutant low-grade diffuse astrocytomas frequently progress to grade IV astrocytomas with implications for patient prognosis. To better understand this process, we applied whole-genome and transcriptome sequencing to matched tumor samples collected before and after progression to grade IV astrocytomas from five patients. All tumors carried an IDH1 mutation. The number of chromosomal rearrangements was increased between 1.3 and 3.5-fold in the tumors upon progression, with the exception of one case, in which the increase was only 1.03-fold. This case exhibited a hypermutation signature caused by homozygous deletion of the MSH2 gene, which encodes a member of the DNA mismatch repair complex. The most common genomic alterations acquired at progression were homozygous deletions in the CDKN2A/ RB1 -pathway or hemizygous deletion of PTEN. Additionally, PDGFRA was amplified in two grade IV tumors, with concordantly increased expression. For one of these cases, a PDGFRA-amplified subclone is likely to be present already in the low-grade astrocytoma. We further detected intrachromosomal rearrangements closeby the genes NRG3 in the progressed tumors as well as in the The Cancer Genome Atlas (TCGA) cohort. The expression of NRG3 decreased with increasing grade in the TCGA cohort and the gene was frequently deleted. Lower NRG3 expression was associated with shorter survival in the TCGA cohort. Several miRNAs showed differential expression upon progression. For two miRNAs the predicted targets were associated with cell cycle regulation and we detected inverse correlation between miRNA and target mRNA expression. While progression seems to occur via different pathways, the predicted outcome for many of the alterations was the inactivation of tumor suppressor genes and further dysregulation of cell proliferation.

Inherited Cancer Susceptibility Gene Sequence Variations Among Patients With Appendix Cancer

Germline sequence variations in APC, BMPR1A, CDH1, CHEK2, EPCAM, MLH1, MSH2, MSH6, MUTYH, PMS2, PTEN, SMAD4, STK11, and TP53 genes are associated with susceptibility to gastrointestinal cancers. As a rare cancer, the evaluation of appendiceal cancer (AC) predisposition has been limited. To assess the prevalence and spectrum of inherited cancer susceptibility gene sequence variations in patients with AC and the utility of germline genetic testing for this population. This cohort study included patients with AC who underwent germline genetic testing of 14 cancer susceptibility genes performed by a clinical testing laboratory between March 1, 2012, and December 31, 2016. Data were analyzed from March to August 2022. Clinical, individual, and family histories were obtained from clinician-completed test requisition forms. Multigene panel testing was performed by targeted custom capture and sequencing and chromosome rearrangement analysis. The main outcomes were germline sequence variation prevalence and spectrum in patients with AC. Among the 131 patients with AC in the cohort (90 female [68.7%]), a total of 16 deleterious sequence variations were identified in 15 patients (11.5%). Similarly, when limited to the 74 patients with AC as the first and only primary tumor, a total of 8 patients (10.8%) had at least 1 deleterious sequence variation in a cancer susceptibility gene. Overall, 6 patients (4.6%) had a deleterious sequence variation observed in MUTYH (5 with monoallelic MUTYH and 1 with biallelic MUTYH). All 4 patients with Lynch syndrome (3.1%) had a sequence variation in the MLH1 gene, of whom 3 were aged 50 years or older at AC diagnosis. Five patients (3.8%) had deleterious sequence variations in other cancer predisposition genes (1 with APC [c.3920T>A, p.I1307K], 2 with CHEK2 [c.470T>C, p.I157T], 1 with SMAD4 [c.263 287dup, p.L98IFS*14], and 1 with TP53 [c.524G>A, p.R175H]). In this cohort study, 1 in every 10 patients with AC who underwent testing for hereditary cancer predisposition carried an inherited gene sequence variation associated with cancer susceptibility. Given the high frequency and broad spectrum of germline gene sequence variations, these data suggest that genetic evaluation might be warranted for all patients diagnosed with this rare malignant tumor. A systemic sequencing effort for all patients with AC may also identify cancer vulnerabilities to exploit for therapeutic development in a cancer type for which clinical trials are limited.

RF21 | PSUN07 Germline Genetic Testing in a Cohort of Adults with Adrenocortical Carcinoma : Insights From a Clinical Care Setting

Abstract Context About 5-10% of adrenocortical cancer (ACC) arise in patients with genetic predisposition syndromes, including Li-Fraumeni, Lynch, MEN1, or Familial Adenomatous Polyposis (APC). Recent European (2018) and American guidelines (2020) recommend that all adults with ACC should be offered clinical genetic counseling. There is a paucity of data regarding systematic germline testing in adults with ACC and the extent of the genetic evaluation is unclear. Objective To describe the germline genetic characteristics in a cohort of adult patients with ACC evaluated in a tertiary clinical care center. Methods Data including demographics, personal and family history of neoplasia, pathology reports, clinical features and genetic testing were retrospectively collected from charts of patients treated at Centre hospitalier de l'Université de Montréal (CHUM). After genetic counseling, genetic testing was proposed to patients with a pathologic diagnosis of ACC. From 2005 to 2015, TP53 gene analysis was performed using direct sequencing and MLPA. Since 2016, multigene testing was performed for a progressively increasing number of oncogenic genes using a custom next-generation panel for germline leucocyte DNA including at least the TP53, MSH2, MSH6, MLH1,PMS2, EPCAM, MEN1, BRCA1 and BRCA2 genes (Invitae, CA). An extended panel, based on past medical and familial history, was performed at the discretion of the geneticist. Patient data was retrospectively investigated. Results We analyzed data from 44 patients with available germline genetic results. Median age of the patients was 46 years (ranging from 22 to 79 years), including 11 males (25.0%) and 33 females (75.0%). Ten patients (22.7%) underwent only TP53 gene analysis and 32 patients (72.7%) were studied using the larger oncogenic genetic panel. Germline pathogenic or likely pathogenic variants were identified in 5 of the 44 patients (11.4%) while genetic variants of unknown significance (VUS) were found in 7 of the 44 patients (15.9%). Among patients with pathogenic variants, one had a family history and known germline mutation in the BRCA2 gene (8765delAG, p.Glu2846GlyfsX23). One patient had a personal and familial medical history suggesting a MEN1 syndrome that was confirmed with the finding of a germline MEN1 mutation (c.1556delC, p.Pro519Leu fs40). Unsuspected germline pathogenic variants were found in three patients: 1) TP53 (c.425delC, p.Pro142fs), 2) MUTYH (c.536A&amp;gt;G, p.Tyr179Cys) and 3) MSH6 (c.3649-3655, p.Arg1217Leu fs9) genes. Variants of unknown significance (VUS) were found in the following genes: POT1, MSH3, PALB2, RAD51d, APC, ATM and BRCA2. Conclusions Germline pathogenic variants were found in 11.4% of our cohort of patients with ACC. VUS were found in 15.9% of patients but their significance remains to be determined. Genetic counseling and germline genetic testing should be offered to all patients with ACC; however the optimal use and extent of oncogenic gene panels need to be better defined. Presentation: Sunday, June 12, 2022 12:30 p.m. - 2:30 p.m., Sunday, June 12, 2022 12:48 p.m. - 12:53 p.m.

Intellectual enrichment and genetic modifiers of cognition and brain volume in Huntington's disease.

An important step towards the development of treatments for cognitive impairment in ageing and neurodegenerative diseases is to identify genetic and environmental modifiers of cognitive function and understand the mechanism by which they exert an effect. In Huntington's disease, the most common autosomal dominant dementia, a small number of studies have identified intellectual enrichment, i.e. a cognitively stimulating lifestyle and genetic polymorphisms as potential modifiers of cognitive function. The aim of our study was to further investigate the relationship and interaction between genetic factors and intellectual enrichment on cognitive function and brain atrophy in Huntington's disease. For this purpose, we analysed data from Track-HD, a multi-centre longitudinal study in Huntington's disease gene carriers and focused on the role of intellectual enrichment (estimated at baseline) and the genes FAN1, MSH3, BDNF, COMT and MAPT in predicting cognitive decline and brain atrophy. We found that carrying the 3a allele in the MSH3 gene had a positive effect on global cognitive function and brain atrophy in multiple cortical regions, such that 3a allele carriers had a slower rate of cognitive decline and atrophy compared with non-carriers, in agreement with its role in somatic instability. No other genetic predictor had a significant effect on cognitive function and the effect of MSH3 was independent of intellectual enrichment. Intellectual enrichment also had a positive effect on cognitive function; participants with higher intellectual enrichment, i.e. those who were better educated, had higher verbal intelligence and performed an occupation that was intellectually engaging, had better cognitive function overall, in agreement with previous studies in Huntington's disease and other dementias. We also found that intellectual enrichment interacted with the BDNF gene, such that the positive effect of intellectual enrichment was greater in Met66 allele carriers than non-carriers. A similar relationship was also identified for changes in whole brain and caudate volume; the positive effect of intellectual enrichment was greater for Met66 allele carriers, rather than for non-carriers. In summary, our study provides additional evidence for the beneficial role of intellectual enrichment and carrying the 3a allele in MSH3 in cognitive function in Huntington's disease and their effect on brain structure.

An MSH6 germline pathogenic variant p.Gly162Ter associated with Lynch syndrome

We identified a three-generation Russian family with Lynch syndrome with a novel germline variant of the MSH6 gene. An 84-year-old female was diagnosed with endometrial adenocarcinoma at the age of 49 years. Her son was diagnosed with colorectal tubular adenoma at the age of 32 years. A germline nonsense variant (c.484 G > T:p.Gly162Ter) in exon 3 of the MSH6 gene was revealed by whole-exome sequencing. Sanger sequencing confirmed the cosegregation of the MSH6 nonsense variant in family members.

Lynch Syndrome Identification in Saudi Cohort of Endometrial Cancer Patients Screened by Universal Approach.

Lynch syndrome (LS) is the most common cause of inherited endometrial cancer (EC). The prevalence and molecular characteristic of LS in Middle Eastern women with EC have been underexplored. To evaluate the frequency of LS in a cohort of EC patients from Saudi Arabia, a total of 436 EC cases were screened utilizing immunohistochemistry (IHC), MLH1 promoter methylation analysis and next-generation sequencing technology. A total of 53 of 436 (12.2%) ECs were classified as DNA mismatch repair-deficient (dMMR). MLH1 promoter hypermethylation was detected in 30 ECs (6.9%). Three ECs (0.7%) were found to be LS harboring germline pathogenic variants (PVs)/likely pathogenic variants (LPVs): two in the MSH2 gene and one in the MSH6 gene. Three ECs (0.7%) were Lynch-like syndrome (LLS) carrying double somatic MSH2 PVs/LPVs. Seven cases were found to have variants of uncertain significance in cancer-related genes other than MMR genes. Our results indicate that LS prevalence is low among Saudi EC patients and LLS is as common as LS in this ethnicity. Our findings could help in better understanding of the prevalence and mutational spectrum of this syndrome in Saudi Arabia, which may help in defining best strategies for LS identification, prevention and genetic counseling for EC patients.

High MutS homolog 2 expression predicts poor prognosis and is related to immune infiltration in endometrial carcinoma.

Several studies have shown that MutS homolog 2 (MSH2) is highly expressed in many cancer tissues. Transcriptome expression data were collected from the Cancer Genome Atlas (TCGA) database. We analyzed the expression of MSH2 in normal and tumor tissues, the relationship between MSH2 expression and various prognostic factors, and the relationship between MSH2 expression and overall survival, disease specific survival, and progression free interval. We also examined MSH2 promoter methylation between endometrial cancer and normal endometrial tissues, and identified the prognostic value of MSH2 methylation in endometrial cancer. MSH2 was highly expressed in endometrial cancer tumor tissues compared with normal tissues. High MSH2 expression might be an independent prognostic factor for OS, DSS, and PFI. Further, high MSH2 expression was correlated with age and histological type, but not with BMI, clinical stage, tumor invasion, or other clinical features. MSH2 promoter methylation in endometrial cancer was significantly lower than in normal tissues. Additionally, MSH2 levels, OS, DSS, and PFI were associated with BMI, age, tumor invasion, and histological type. ssGSEA showed that MSH2 expression was positively correlated with the infiltration of Th2 cells, Tcm cells, T helper cells, and Tgd cells, whereas it was negatively correlated with NK CD56 bright cells, pDC cells, iDC cells, cytotoxic cells, and neutrophils. Increased MSH2 expression and reduced MSH2 methylation in endometrial cancer predicts poor prognosis. MSH2 may be used as a biomarker for the diagnosis and prognosis of endometrial cancer and as an immunotherapy target.

MutS functions as a clamp loader by positioning MutL on the DNA during mismatch repair

Highly conserved MutS and MutL homologs operate as protein dimers in mismatch repair (MMR). MutS recognizes mismatched nucleotides forming ATP-bound sliding clamps, which subsequently load MutL sliding clamps that coordinate MMR excision. Several MMR models envision static MutS-MutL complexes bound to mismatched DNA via a positively charged cleft (PCC) located on the MutL N-terminal domains (NTD). We show MutL-DNA binding is undetectable in physiological conditions. Instead, MutS sliding clamps exploit the PCC to position a MutL NTD on the DNA backbone, likely enabling diffusion-mediated wrapping of the remaining MutL domains around the DNA. The resulting MutL sliding clamp enhances MutH endonuclease and UvrD helicase activities on the DNA, which also engage the PCC during strand-specific incision/excision. These MutS clamp-loader progressions are significantly different from the replication clamp-loaders that attach the polymerase processivity factors β-clamp/PCNA to DNA, highlighting the breadth of mechanisms for stably linking crucial genome maintenance proteins onto DNA.

Individualized therapy based on the combination of mini-PDX and NGS for a patient with metastatic AFP-producing and HER-2 amplified gastric cancer.

Mini-patient-derived xenograft (mini-PDX) is a novel, rapid and accurate method used to assess in vivo drug susceptibility. In the present study, a mini-PDX combined with next-generation sequencing (NGS) was used to guide the individualized treatment of a patient with metastatic a-fetoprotein-producing and human epidermal growth factor receptor 2 (HER-2) amplified gastric cancer (GC). Tumor cells were isolated from the tumor tissue obtained from gastroscopic biopsy, transferred into capsules and implanted into severe combined immunodeficiency mice to determine their sensitivity to various drug regimens. NGS was also performed to assess the mutation spectrum of the cells. The results were analyzed to select the most appropriate treatment regimen for the patient. The mini-PDX model confirmed that the patient's tumor was sensitive to a combination regimen of irinotecan and tegafur-gimeracil-oteracil (S-1). Fluorescence in situ hybridization assay of the tumor tissue confirmed HER-2 amplification. The NGS results indicated ERBB2 amplification, and tumor protein P53 [c.659A>G (p.Y220C)], ataxia-telengiectasia mutated [c.125A>G (p.H42R)] and MutS homolog 6 [c.3254C(8>7) (p.F1088Sfs*2)] mutation, in which UGT1A1*28, TA6/7 (rs8175347) was a mutant heterozygote. After six courses of treatment with a regimen comprising 300 mg irinotecan on day 1 + 40 mg S-1 twice daily on days 2–15 + 350 mg trastuzumab once-every 3 weeks, the patient continued with S-1 treatment for 4 courses and trastuzumab for 1 year. The patient retained progression-free survival status at the 32-month follow-up. Thus, the mini-PDX model combined with the NGS rapidly assessed drug sensitivity in a patient with GC and revealed key genetic mutations. However, the proposed technique requires further research to confirm its potential in the individualized treatment of patients with refractory malignancies.