DNA Mismatch Repair Genes Research Articles

Prognostic Values of METTL3 and Its Roles in Tumor Immune Microenvironment in Pan-Cancer

Background: N6-methyladenosine (m6A) is among the most prevalent RNA modifications regulating RNA metabolism. The roles of methyltransferase-like 3 (METTL3), a core catalytic subunit, in various cancers remain unclear. Methods: The expression levels of METTL3 in pan-cancer were profiled and their prognostic values were examined. We assessed the relationships between METTL3 expression levels and tumor immune infiltration levels, immune checkpoint gene expression, immune neoantigens, tumor mutation burden, microsatellite instability, and DNA mismatch repair gene expression. Furthermore, a protein-protein interaction network was drawn, and gene set enrichment analysis was conducted to explore the functions of METTL3. Results: METTL3 expression levels were elevated in most cancers, with high expression associated with poorer overall and disease-free survival. METTL3 levels were significantly related to immune cell infiltration, tumor mutation burden, microsatellite instability, mismatch repair genes, and immune checkpoint gene levels. METTL3 was enriched in pathways related to RNA modification and metabolism and correlated with epithelial-mesenchymal transition. Conclusions: METTL3 serves as an oncogene in most cancer types and shows potential as a prognostic biomarker. Additionally, our comprehensive pan-cancer analysis suggested that METTL3 is involved in regulating the tumor immune microenvironments and epithelial-mesenchymal transition via modulating RNA modification and metabolism, making it a potential therapeutic target.

A Novel Immune Gene-Related Prognostic Score Predicts Survival and Immunotherapy Response in Glioma.

Background and Objectives: The clinical prognosis and survival prediction of glioma based on gene signatures derived from heterogeneous tumor cells are unsatisfactory. This study aimed to construct an immune gene-related prognostic score model to predict the prognosis of glioma and identify patients who may benefit from immunotherapy. Methods: 23 immune-related genes (IRGs) associated with glioma prognosis were identified through weighted gene co-expression network analysis (WGCNA) and Univariate Cox regression analysis based on large-scale RNA-seq data. Eight IRGs were retained as candidate predictors and formed an immune gene-related prognostic score (IGRPS) by multifactorial Cox regression analysis. The potential efficacy of immune checkpoint blockade (ICB) therapy of different subgroups was compared by The Tumor Immune Dysfunction and Exclusion (TIDE) algorithm. We further adopted a series of bioinformatic methods to characterize the differences in clinicopathological features and the immune microenvironment between the different risk groups. Finally, a nomogram integrating IGRPS and clinicopathological characteristics was built to accurately predict the prognosis of glioma. Results: Patients in the low-risk group had a better prognosis than those in the high-risk group. Patients in the high-risk group showed higher TIDE scores and poorer responses to ICB therapy, while patients in the low-risk group may benefit more from ICB therapy. The distribution of age and tumor grade between the two subgroups was significantly different. Patients with low IGRPS harbor a high proportion of natural killer cells and are sensitive to ICB treatment. While patients with high IGRPS display relatively poor prognosis, a higher expression level of DNA mismatch repair genes, high infiltrating of immunosuppressive cells, and poor ICB therapeutic outcomes. Conclusions: We demonstrated that the IGRPS model can independently predict the clinical prognosis as well as the ICB therapy responses of glioma patients, thus having important implications on the design of immune-based therapeutic strategies.

Exposure of Pseudomonas aeruginosa to Cinnamaldehyde Selects Multidrug Resistant Mutants.

Cinnamaldehyde (CNA), the main component of cinnamon essential oil, is one of the most active plant compounds against nosocomial pathogen Pseudomonas aeruginosa. Exposure of wild-type strain PA14 (MIC 700 µg/mL) for 5 to 10 days to fixed (900 µg/mL) or increasing (from 900 to 1400 µg/mL) concentrations of this natural antibacterial resulted in emergence of resistant mutants CNA-A1 to A3, and CNA-B1 to B7, respectively. Genome sequencing experiments showed that each of CNA-A1 to A3 mutants differed from PA14 by one SNP, and a slight increase in CNA resistance level (from 700 to 900 µg/mL). By comparison, mutants B1 to B7 were more resistant (up to 1100 µg/mL); each of them harbored multiple SNPs (from 24 to 39) likely as a consequence of alteration of DNA mismatch repair gene mutS. Of the ten mutants selected, eight contained mutations in gene nalC, which indirectly downregulates expression of the operon that codes for multidrug efflux system MexAB-OprM, and showed increased resistance (up to 16-fold versus PA14) to antibiotic molecules exported by the pump, including ß-lactams and fluoroquinolones. Of the six mutants with the highest CNA resistance, five were no longer motile because of alteration of genes flgJ, fliE and/or pilJ genes. Altogether, our data show that P. aeruginosa is able to adapt to strong electrophilic molecules such as CNA by upregulating its intrinsic efflux pump MexAB-OprM, and through less well-characterized pleiotropic changes. Whether multidrug-resistant mutants can emerge in patients using cinnamon essential oil as self-medication needs to be assessed further.

Abstract IA018: Cancer immune-interception in Lynch Syndrome: Neoantigen-based vaccine development

Abstract Lynch Syndrome (LS) is the most common cause of hereditary colorectal cancer and also conveys significantly increased risks for several other malignancies, including endometrial, small bowel, gastric, ovarian, adrenocortical tumors, and others. LS results from a heterozygous germline mutation in one of four DNA mismatch repair (MMR) genes. The acquisition of a somatic “second hit” causes affected cells to lose the ability to repair DNA mismatch replication errors that frequently occur in microsatellite regions of the genome. Consequently, hundreds to thousands of small insertion/deletions (indels) accumulate in these regions, and when expressed, result in frameshift peptides that are recognized as neoantigens that are then presented on the cell surface via the major histocompatibility complexes (MHC-I/II) for immune cell recognition. Vaccine development efforts in this high-risk population have been challenged by the substantial inter-individual variability in both the set of expressed neoantigens and MHC I/II responses. Yet, recent advances in next-generation sequencing and associated bioinformatic approaches are now allowing for more accurate profiling of the most frequently recurring and shared mutated neoantigens in LS-associated tumors. This allows for identification of the most immunogenic neoantigens that can be incorporated into different vaccine platforms to test the development of a population-based vaccine. The Vilar Lab has collaborated with industry and academic partners such as Nouscom, s.r.l. and the National Cancer Institute to develop two different clinical trials to bring new vaccines to the LS population based on mutated neoantigens and tumor associated antigens. In this presentation, we will focus on mutated neoantigen-based strategies and present the Phase I clinical trial (NCT05078866) using a viral-based vaccine including 209 distinct neoantigens. The primary endpoint of this trial is safety and immunogenicity. A total of 45 participants will be enrolled to receive a prime and boost vaccination. Different secondary biomarkers will explore immunological aspects of this novel immune-interception strategy. Citation Format: Eduardo Vilar-Sanchez. Cancer immune-interception in Lynch Syndrome: Neoantigen-based vaccine development [abstract]. In: Proceedings of the Second Biennial NCI Meeting: Translational Advances in Cancer Prevention Agent Development (TACPAD); 2022 Sep 7-9. Philadelphia (PA): AACR; Can Prev Res 2022;15(12 Suppl_2): Abstract nr IA018.

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.

Functional characterization of MLH1 missense variants unveils mechanisms of pathogenicity and clarifies role in cancer.

Lynch syndrome is a heritable condition caused by a heterozygous germline inactivating mutation of the DNA mismatch repair (MMR) genes, most commonly the MLH1 gene. However, one third of the identified alterations are missense variants, for which the clinical significance is unclear in many cases. We have identified three MLH1 missense alterations (p.(Glu736Lys), p.(Pro640Thr) and p.(Leu73Pro)) in six individuals from large Tunisian families. For none of these alterations, a classification of pathogenicity was available, consequently diagnosis, predictive testing and targeted surveillance in affected families was impossible. We therefore performed functional laboratory testing using a system testing stability as well as catalytic activity that includes clinically validated reference variants. Both p.(Leu73Pro) and p.(Pro640Thr) were found to be non-functional due to severe defects in protein stability and catalytic activity. In contrast, p.(Glu736Lys) was comparable to the wildtype protein and therefore considered a neutral substitution. Analysis of residue conservation and of the structural roles of the substituted residues corroborated these findings. In conjunction with the available clinical data, two variants fulfil classification criteria for class 4 "likely pathogenic". The findings of this work clarify the mechanism of pathogenicity of two unclear MLH1 variants and enables predictive testing and targeted surveillance in members of carrier families worldwide.

Comprehensive Pan-Cancer Analysis of GINS2 for Human Tumour Prognosis and as an Immunological Biomarker.

In recent years, more and more reports have shown that GINS complex subunit 2 (GINS2) plays an important role in the occurrence and progression of tumours. However, there is a lack of comprehensive and systematic research on its prognostic and immune effects in pan-cancer. Therefore, this study is aimed at investigating the prognostic value and immune-related role of GINS2 in human tumours and providing a comprehensive understanding of its carcinogenic mechanism in pan-cancer. We investigated different databases, including TIMER, TCGA, GTEX, CPTAC, GEPIA, and SangerBox. The study was carried out on the expression and prognosis of GINS2 in human tumours, immune infiltration and microenvironment, immune checkpoints, neoantigens, tumour mutational burden, microsatellite instability, mismatch repair (MMR) genes, methylation, cancer-associated fibroblasts (CAFs), and enrichment analysis of gene set. GINS2 plays a potential carcinogenic role in various human tumours through mRNA and protein levels. It is highly expressed in most cancers, and its expression is significantly correlated with tumour prognosis. In addition, the expression of GINS2 is associated with immune microenvironment and immune infiltration, especially in brain lower-grade glioma, lung squamous cell carcinoma, TGCT, breast invasive carcinoma, and glioblastoma multiforme. At the same time, GINS2 is related to immune neoantigens and the expression profiles of immune checkpoint genes in pan-cancer. It also affects the expression of DNA MMR genes and methyltransferase in pan-cancer. Finally, the correlation between GINS2 and CAF abundance in most tumours was studied, and an enrichment analysis of GINS2 and its related proteins was also carried out. This is the first study on GINS2 as a prognostic and immune mechanism in pan-cancer. GINS2 may be a valuable prognostic immunological biomarker of pan-cancer. This paper provides a relatively comprehensive understanding on the correlation of GINS2 with pan-cancer.

Patterns of DNA mismatch repair protein expression for primary and recurrent colorectal cancer at an advanced surgical unit: A retrospective audit.

Lynch syndrome is an inherited cancer syndrome associated with an increased lifetime risk of colorectal cancer (CRC) and characterized by germline mutations to one of four DNA mismatch repair (MMR) genes. Immunohistochemical (IHC) testing is used to screen for Lynch syndrome; however, despite routine completion following resection of primary CRC, it is only variably completed following resection of recurrent disease. This may be significant, as MMR protein expression can change from primary to recurrent CRC. The primary aim of this study is to investigate how MMR profiles change from primary to recurrent CRC; the secondary aim is to assess rates of MMR testing of primary and recurrent disease. We conducted a retrospective analysis of patients undergoing surgery for recurrent CRC from 2018-19 at a high-volume institution. MMR profiles were obtained following both primary and recurrent resection of CRC, and MMR protein expression was evaluated from both time points. A total of 107 patients met the inclusion criteria and IHC results were obtained for both primary and recurrent resections in 85 cases. MMR profiles changed in nine patients (10.6%), with a loss of staining from primary to recurrent disease in six (7.1%) and a gain of staining in three (3.5%). IHC testing was completed following 88.7% of primary and 39.3% of recurrent resections. MMR profiles can change from primary to recurrent CRC and repeat MMR testing for recurrent CRC is completed in only a minority of cases.

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).

Comprehensive analysis of HHV-6 and HHV-7-related gene signature in prognosis and response to temozolomide of glioma.

Human herpesvirus (HHV)-6 and HHV-7 have been detected in central nervous system and glioma tissue, while their exact role in glioma remains uncertain. Omics profiles and clinical information were downloaded from public databases, including The Cancer Genome Atlas cohort for training set and the Chinese Glioma Genome Atlas cohorts for validation sets. Differentially expressed genes between HHV-6 and HHV-7 infected or noninfected glioma patients were screened for establishing the HHV-6 and HHV-7 infection (HI) model through Lasso regression analysis. Bioinformatics methods were used to analyze the correlation between HI scores and prognosis, metastasis in glioma patients. Predictable efficacy of HI in temozolomide-resistance and HI-related genetic signatures were also explored. The HI model was constructed as: Risk score = (0.014709*DIRAS3) + (0.029787*TEX26) + (0.223492*FBXO39) + (0.074951*MYBL1) + (0.060202*HILS1). The five gene signature showed good performance in predicting survival time for glioma patients, while higher HI score is correlated with malignant features. Moreover, DNA mismatch repair genes were augmented in glioma patients with higher HI score as well as nonresponse to temozolomide treatment, which was in parallel with the transcriptomic result of temozolomide-resistant glioma cell. Targeting the five gene signature is beneficial for prognosis of glioma patients, especially in glioma patients underwent temozolomide treatment.

MODL-20. REDUCTION OR LOSS OF MSH6 CONFERS RESISTANCE TO TEMOZOLOMIDE IN GLIOBLASTOMA

Abstract Glioblastoma (GBM) is the most common and aggressive primary brain tumor. The only intervention that has improved the survival rate of GBM patients over the past several decades has been combining temozolomide (TMZ) with radiotherapy (RT), which increased median survival by only ~2.5 months to where it currently stands at ~15 months. Unfortunately, all GBM patients eventually die due to tumor recurrence. Intrinsic or acquired resistance to TMZ is a significant contributing factor to tumor progression, and various mechanisms have been suggested, including deficiencies in DNA mismatch repair (MMR) genes such as MSH6. To test the biological significance of Msh6 in GBM growth and resistance to TMZ and immunotherapy, we have generated immunocompetent genetically engineered mouse models (GEMMs) of GBM with germline loss of Msh6. To generate tumors with reduced or abrogated Msh6, we utilized GEMMs based on the RCAS/tv-a gene transfer system in combination with mice that exhibited heterozygous and homozygous loss of Msh6. When PDGFB was overexpressed in combination with the silencing of Tp53 to induce tumors, tumor-bearing mice with reduced or deficient Msh6 demonstrated increased tumor growth and shorter survival time compared to wild-type tumor-bearing mice. Our data demonstrate that two weeks of TMZ treatment at a clinically relevant dose of 25 mg/kg provides a significant survival advantage in WT-tumor-bearing mice, while no efficacy was observed in tumor-bearing mice with either heterozygous or homozygous loss of Msh6. We are currently evaluating the mechanism by which of Msh6 confers TMZ-resistance.

Modeling Therapy-Driven Evolution of Glioblastoma with Patient-Derived Xenografts.

Adult-type diffusely infiltrating gliomas, of which glioblastoma is the most common and aggressive, almost always recur after treatment and are fatal. Improved understanding of therapy-driven tumor evolution and acquired therapy resistance in gliomas is essential for improving patient outcomes, yet the majority of the models currently used in preclinical research are of therapy-naïve tumors. Here, we describe the development of therapy-resistant IDH-wildtype glioblastoma patient-derived xenografts (PDX) through orthotopic engraftment of therapy naïve PDX in athymic nude mice, and repeated in vivo exposure to the therapeutic modalities most often used in treating glioblastoma patients: radiotherapy and temozolomide chemotherapy. Post-temozolomide PDX became enriched for C>T transition mutations, acquired inactivating mutations in DNA mismatch repair genes (especially MSH6), and developed hypermutation. Such post-temozolomide PDX were resistant to additional temozolomide (median survival decrease from 80 days in parental PDX to 42 days in a temozolomide-resistant derivative). However, temozolomide-resistant PDX were sensitive to lomustine (also known as CCNU), a nitrosourea which induces tumor cell apoptosis by a different mechanism than temozolomide. These PDX models mimic changes observed in recurrent GBM in patients, including critical features of therapy-driven tumor evolution. These models can therefore serve as valuable tools for improving our understanding and treatment of recurrent glioma.

Pediatric manifestations of Lynch Syndrome: A single center experience

Lynch syndrome is an autosomal dominant condition caused by a heterozygous variation in one of the DNA mismatch repair (MMR) genes that pre-disposes individuals to early onset colorectal cancers and other malignancies. Lynch syndrome is generally considered an adult-onset disorder, with malignancy rarely manifesting in childhood. Colorectal cancer is extremely rare in children, but hereditary syndromes including Lynch syndrome are an important cause. We aimed to assess the frequency and clinical course of children with Lynch syndrome associated pediatric colorectal cancers at our institution over the last 20 years. In this retrospective study, we describe four cases of children with Lynch syndrome-associated colorectal cancers age 14-17 years at diagnosis. All patients were diagnosed with Lynch syndrome after diagnosis, despite three of them having family histories consistent with Lynch syndrome. This series highlights a rare but important cause of pediatric malignancy and points to the need for early education on colorectal cancer warning symptoms and open discussion about this condition in affected families. It also illustrates the need for a thorough family history and a high level of suspicion for Lynch syndrome in children based on family background, as early detection may be key to improving cancer outcomes.

"Lynch Syndrome: Current Status of Crc in Biotechnological and Biomedical Fields"

Lynch syndrome (LS) is an autosomal dominant genetic condition caused by mutations in the DNA mismatch repair (MMR) genes in the germline. Colorectal cancer and/or LS-associated cancer are more likely in people who carry pathogenic mutations in these genes. Cancers of the endometrium, small intestine, stomach, pancreas, and biliary tract, ovarian, brain, upper urinary tract, and skin are among the cancer types linked to LS. The criteria for a clinical diagnosis of LS, as well as the processes for genetic testing to identify carriers of pathogenetic mutations in MMR genes, have been known for a long time.

Is HLA type a possible cancer risk modifier in Lynch syndrome?

Lynch syndrome (LS) is the most common inherited cancer syndrome. It is inherited via a monoallelic germline variant in one of the DNA mismatch repair (MMR) genes. LS carriers have a broad 30% to 80% risk of developing various malignancies, and more precise, individual risk estimations would be of high clinical value, allowing tailored cancer prevention and surveillance. Due to MMR deficiency, LS cancers are characterized by the accumulation of frameshift mutations leading to highly immunogenic frameshift peptides (FSPs). Thus, immune surveillance is proposed to inhibit the outgrowth of MMR-deficient cell clones. Recent studies have shown that immunoediting during the evolution of MMR-deficient cancers leads to a counter-selection of highly immunogenic antigens. The immunogenicity of FSPs is dependent on the antigen presentation. One crucial factor determining antigen presentation is the HLA genotype. Hence, a LS carrier's HLA genotype plays an important role in the presentation of FSP antigens to the immune system, and may influence the likelihood of progression from precancerous lesions to cancer. To address the challenge of clarifying this possibility including diverse populations with different HLA types, we have established the INDICATE initiative (Individual cancer risk by HLA type, http://indicate-lynch.org/), an international network aiming at a systematic evaluation of the HLA genotype as a possible cancer risk modifier in LS. Here we summarize the current knowledge on the role of HLA type in cancer risk and outline future research directions to delineate possible association in the scenario of LS with genetically defined risk population and highly immunogenic tumors.

Characterization of hotspot exonuclease domain mutations in the DNA polymerase ϵ gene in endometrial cancer.

ObjectiveThis study was aimed to profile hotspot exonuclease domain mutations (EDMs) of the DNA polymerase ϵ gene (POLE) in endometrial cancer (EC) and to investigate the effects of EDMs on tumor cell behavior and catalytic activities of Polϵ.Methods POLE sequencing was performed in tumor tissue samples from patients with EC to identify hotspot EDMs. Bioinformatics tools were used to select the potential pathogenic EDMs. The association of EDMs with the clinical outcomes of patients was assessed. EC cells were transfected with wildtype POLE or POLE variants to examine the effects of the EDMs on EC cell behavior, including cell cycle, migration, and invasion. Co-immunoprecipitation was employed to obtain FLAG-tagged wildtype and mutant catalytic subunits of Polϵ, followed by the assessment of polymerase and exonuclease activities.ResultsIn addition to previously reported P286R and V411L, R375Q and P452L were identified as novel, and deleterious POLE hotspot EDMs of EC. Patients in EDM group had significantly better clinical outcomes than the rest of the cohort. Compared with wildtype POLE, overexpression of POLE variants promoted cisplatin resistance, G0/G1 cell cycle arrest, and cell migration and invasion in EC cells. Overexpression of POLE variants significantly increased the abundance of 3’-OH and upregulated the expression of DNA mismatch repair genes in HEK293T cells. Compared with wildtype Polϵ, Pol ϵ mutants exhibited undermined polymerase and exonuclease abilities in the presence of mismatched nucleotides in HEK293 cells.ConclusionWe characterized the of hotspot exonuclease domain mutations in the DNA polymerase ϵ gene and identified P286R, V411L, R375Q, and P452L as pathogenic POLE hotspot EDMs in endometrial cancer. These hotspot EDMs are associated with the malignant behavior of endometrial cancer cells in vitro and favorable prognosis in patients, suggesting that POLE affects a wide range of cellular processes beyond DNA replication and proofreading.

Selenoprotein GPX1 is a prognostic and chemotherapy-related biomarker for brain lower grade glioma

ObjectiveGlutathione peroxidase 1 (GPX1) is a major selenoprotein in most animal tissues, primarily expressed in the cytoplasm and mitochondria of cells and peroxidase structures of certain cells. GPX1 expression is highly correlated with carcinogenesis and disease progression. The goal of the study was to determine the association between GPX1 expression and tumor therapy, and to identify GPX1 prognostic value in various malignancies. MethodsThe Cancer Genome Atlas (TCGA), Genotype-Tissue Expression (GTEx), and Human Protein Atlas (HPA) databases were used to detect the levels of GPX1 expression in human tumor tissues and normal tissues. Indeed, correlations between GPX1 and tumor purity, tumor mutation burden (TMB), microsatellite instability (MSI), and DNA mismatch repair genes (MMRs) were explored using the TCGA cohort. Functional and enrichment analyses were performed by the GeneMANIA database and Gene Set Enrichment Analysis (GSEA), respectively. Cox regression models and Kaplan – Meier curves were used to screen for independent risk factors and estimate brain lower-grade glioma (LGG) survival probability. The Chinese Glioma Genome Atlas (CGGA) database was used to determine whether GPX1 had a race-specific effect on overall survival (OS) in LGG. The cross-interaction between GPX1 and chemoradiotherapy on LGG OS was determined by Kaplan – Meier curves. Logistic regression models of multiplicative interactions were constructed. Furthermore, the relationship between GPX1 and LGG treatment regimens was also explored through the Genomics of Drug Sensitivity in Cancer (GDSC) database. ResultsGPX1 was highly expressed in various tumors, GPX1 overexpression was significantly correlated with the poor prognosis of LGG. GPX1 was found to be an independent predictive factor for LGG in both univariate and multivariate Cox models. The nomogram showed a high predictive accuracy (C-index: 0.804, 95% CI: 0.74–0.86). In addition, GPX1 was significantly associated with TMB, MSI, and MMRs in diverse cancers. GPX1 was involved in IL6/JAK/STAT3, inflammatory response, and apoptosis signaling pathways. Besides, non-radiotherapy, chemotherapy, and low GPX1 expression were important factors affecting the better prognosis of LGG. GPX1 acted as a tumor promoter, which has taken the worst effect on LGG survival, but a multiplicative interaction of GPX1*chemoradiotherapy may improve the poor clinical outcome. GPX1 was negatively correlated with the half inhibition concentration (IC50) of temozolomide (TMZ) (Spearman = −0.44, P = 4.52 ×10−26). ConclusionIn LGG patients, high GPX1 expression was linked to a shorter OS. The interaction between GPX1 and chemoradiotherapy exhibits a beneficial clinical effect and chemotherapy was recommended for LGG patients, especially for those with high GPX1 expression. Besides, high GPX1 expression can predict TMZ sensitivity in LGG, providing potential evidence for chemotherapy. On the whole, this study presents a wealth of biological as well as clinical significance for the roles of GPX1 in human tumors, particularly in LGG.

Comprehensive RNA and protein functional assessments contribute to the clinical interpretation of MSH2 variants causing in-frame splicing alterations

BackgroundSpliceogenic variants in disease-causing genes are often presumed pathogenic since most induce frameshifts resulting in loss of function. However, it was recently shown in cancer predisposition genes that some may...

A calibrated cell-based functional assay to aid classification of MLH1 DNA mismatch repair gene variants.

Functional assays provide important evidence for classifying the disease significance of germline variants in DNA mismatch repair genes. Numerous laboratories, including our own, have developed functional assays to study mismatch repair gene variants. However, previous assays are limited due to the model system employed, the manner of gene expression, or the environment in which function is assessed. Here, we developed a human cell-based approach for testing the function of variants of uncertain significance (VUS) in the MLH1 gene. Using clustered regularly interspaced short palindromic repeats gene editing, we knocked inMLH1 VUS into the endogenous MLH1 loci in human embryonic stem cells. We examined their impact on RNA and protein, including their ability to prevent microsatellite instability and instigate a DNA damage response. A statistical clustering analysis determined the range of functions associated with known pathogenic or benign variants, and linear regression was performed using existing odds in favor of pathogenicity scores for these control variants to calibrate our functional assay results. By converting the functional outputs into a single odds in favor of pathogenicity score, variant classification expert panels can use these results to readily reassess these VUS. Ultimately, this information will guide proper diagnosis and disease management for suspected Lynch syndrome patients.

372P Microsatellite instability testing practices in colorectal cancer patients, in Europe and Asia

Microsatellite instability (MSI) is caused by mutations in the DNA mismatch repair genes and has been shown to predict response to checkpoint inhibitors (CPIs). Both FDA & EMA have approved CPIs in MSI-High (MSI-H) colorectal cancer (CRC). The aim of this study was to understand MSI testing practices for CRC patients, among physicians in Europe and Asia.