Mismatch Repair Research Articles

Clinical and Pathologic Response to Neoadjuvant Immunotherapy in DNA Mismatch Repair Protein-Deficient Gastroesophageal Cancers.

Mismatch repair deficient (dMMR) gastroesophageal cancers (GEC) are a distinct subgroup. Among patients with locally advanced disease, previous trial data suggest a good response to neoadjuvant immune checkpoint inhibitors (nICI). Since 2019, our institution has routinely performed MMR testing for new GEC cases. Patients diagnosed with GEC (2019-2024) were included in the study. Quantitative data are described as the median and interquartile range (IQR); qualitative data are described as quantities and percentages. A total of 24 patients with dMMR GEC were identified following implementation of routine immunohistochemical testing; 14 were potentially resectable with a median follow-up of 14 months (IQR 8-27). All patients underwent pre-treatment positron emission tomography (PET; median SUV 20.9). Among the 14 potentially resectable patients, 4 underwent immediate surgery, 10 were treated with nICI, and 5 underwent surgical resection to date. All regimens included PD-1 inhibitors, with 70% receiving pembrolizumab. Re-staging PET was performed in five patients; the median post-nICI SUV was 5.1 (range 4.7-6.3). All resected specimens had gross ulceration after nICI, but 60% (N = 3) had a pathologic complete response (pCR) following nICI; one patient had a near-complete response (nCR) and one patient had a partial response (pPR). Reduction in SUV was 75% and 82% in the pCR patients, 25% in the nCR patient, and 43% in the pPR patient. dMMR GECs are responsive to nICI in this limited experience, mirroring early clinical trial data. Given persistent metabolic activity and visible ulceration despite pCR, studies should continue to optimize tools for estimating post-nICI pCR in these patients.

SENECA study: staging endometrial cancer based on molecular classification

ObjectiveManagement of endometrial cancer is advancing, with accurate staging crucial for guiding treatment decisions. Understanding sentinel lymph node (SLN) involvement rates across molecular subgroups is essential. To evaluate SLN involvement...

A comprehensive molecular characterization of a claudin-low luminal B breast tumor

Breast cancer is the most common cause of death from cancer in women. Here, we present the case of a 43-year-old woman, who received a diagnosis of claudin-low luminal B breast cancer. The lesion revealed to be a poorly differentiated high-grade infiltrating ductal carcinoma, which was strongly estrogen receptor (ER)/progesterone receptor (PR) positive and human epidermal growth factor receptor (HER2) negative. Her tumor underwent in-depth chromosomal, mutational and gene expression analyses. We found a pathogenic protein truncating mutation in the TP53 gene, which is predicted to disrupt its transcriptional activity. The patient also harbors germline mutations in some mismatch repair (MMR) genes, and her tumor displays the presence of immune infiltrates, high tumor mutational burden (TMB) status and the apolipoprotein B mRNA editing enzyme catalytic polypeptide 3 (APOBEC3) associated signatures, which, overall, are predictive for the use of immunotherapy. Here, we propose promising prognostic indicators as well as potential therapeutic strategies based on the molecular characterization of the tumor.

NTRK gene alterations were enriched in hepatoid or enteroblastic differentiation type of gastric cancer

AimsCurrently, the clinicopathological characteristics of gastric cancer (GC) with oncogenic NTRK alterations are not well known. Although NTRK fusion has been identified as prevalent in DNA mismatch repair protein deficient...

Integrated genomic/epigenomic analysis stratifies subtypes of clear cell ovarian carcinoma, highlighting their cellular origin

The cellular origin of clear cell ovarian carcinoma (CCOC), a major histological subtype of ovarian carcinoma remains elusive. Here, we explored the candidate cellular origin and identify molecular subtypes using integrated genomic/epigenomic analysis. We performed whole exome-sequencing, microarray, and DNA methylation array in 78 CCOC samples according to the original diagnosis. The findings revealed that ARID1A and/or PIK3CA mutations were mutually exclusive with DNA repair related genes, including TP53, BRCA1, and ATM. Clustering of CCOC and other ovarian carcinomas (n = 270) with normal tissues from the fallopian tube, ovarian surface epithelium, endometrial epithelium, and pelvic peritoneum mesothelium (PPM) in a methylation array showed that major CCOC subtypes (with ARID1A and/or PIK3CA mutations) were associated with the PPM-lile cluster (n = 64). This cluster was sub-divided into three clusters: (1) mismatch repair (MMR) deficient with tumor mutational burden-high (n = 2), (2) alteration of ARID1A (n = 51), and (3) ARID1A wild-type (n = 11). The remaining samples (n = 14) were subdivided into (4) ovarian surface epithelium-like (n = 11) and (5) fallopian tube-like (considered as high-grade serous histotype; n = 3). Among these, subtypes (1–3) and others (4 and 5) were found to be associated with immunoreactive signatures and epithelial-mesenchymal transition, respectively. These results contribute to the stratification of CCOC into biological subtypes.

PMS2 has both pro-mutagenic and anti-mutagenic effects on repeat instability in the Repeat Expansion Diseases.

Genome Wide Association studies (GWAS) have implicated PMS2 as a modifier of somatic expansion in Huntington's disease (HD), one of >45 known Repeat Expansion Diseases (REDs). PMS2 is a subunit of the MutLα complex, a major component of the mismatch repair (MMR) system, a repair pathway that is involved in the generation of expansions in many different REDs. However, while MLH3, a subunit of a second MutL complex, MutLγ, is required for all expansions, PMS2 has been shown to protect against expansion in some model systems but to drive expansion in others. To better understand PMS2's behavior, we have compared the effect of the loss of PMS2 in different tissues of an HD mouse model (CAG/CTG repeats) and a mouse model for the Fragile X-related disorders (FXDs), disorders that result from a CGG/CCG repeat expansion. Mice heterozygous for Pms2 show increased expansions in most expansion-prone tissues in both disease models. However, in Pms2 null mice expansions of both repeats increased in some tissues but decreased in others. Thus, the previously reported differences in the effects of PMS2 in different model systems do not reflect fundamentally different roles played by PMS2 in different REDs, but rather the paradoxical effects of PMS2 in different cellular contexts. These findings have important implications not only for the mechanism of expansion and the development of therapeutic approaches to reduce the pathology generated by repeat expansion, but also for our understanding of normal MMR.

Enhanced Commendable Sensitivity and Specificity for MSI in Colorectal Cancer by a New PCR-HRM Based 8-Loci MSI Assay.

This study evaluated the performance of the PCR-HRM assay by comparing it with immunohistochemistry (IHC) for mismatch repair (MMR) proteins and the PCR capillary electrophoresis (PCR-CE) methods. A total of 224 patients with colorectal cancer participated in the study, with nearly half having mismatch repair deficiency (dMMR) tissues and the remainder possessing pMMR tissues. There was a 97.77% concordance between the PCR-HRM assay and IHC, and a 97.56% concordance between PCR-HRM and the PCR-CE assay. In comparison with IHC for dMMR proteins, the PCR-HRM demonstrated a sensitivity of 96.36% and a specificity of 99.12%. When juxtaposed with the PCR-CE assay, its sensitivity was 98.96% and specificity stood at 96.33%. The mutations observed in the microsatellite loci were uniformly distributed across all eight loci. Discrepant outcomes were more frequent in instances of MLH1 and PMS2 deficiency. Furthermore, the germline mutation status of MLH1, MSH2, PMS2, and MSH6 in 62 patients was ascertained using next-generation sequencing. All patients displaying MMR gene pathogenic mutations (N = 14) were identified as MSI-H by PCR-HRM, whereas those with MSS tissues (N = 43) did not exhibit MMR gene pathogenic mutations. Thus, the PCR-HRM method proficiently pinpoints tumors with verified germline MMR mutations, indicative of Lynch syndrome. Conclusively, the PCR-HRM assay emerges as a swift and congruent diagnostic tool for microsatellite instability, boasting commendable sensitivity and specificity in colorectal cancer.

Comparing Clinicopathological and Immunohistochemical Features of Colorectal Carcinoma between Young and Old Age Groups.

The incidence of colorectal carcinoma (CRC) is increasing among individuals younger than 50, and some studies suggest the presence of differences in CRC among old and young individuals regarding clinical and histopathological features. The aim of this study was to compare clinicopathological features, mismatch repair protein status, and expression of certain immunohistochemical stains between young and old groups. The study included 180 cases and found significant histological and immunohistochemical differences between the two groups. CRC in the young tends to be more right-sided and has a higher percentage of dMMR proteins, but less expression of p53 mutations. These features are commoner in Lynch syndrome, and more investigations to study the relationship between young-onset CRC and hereditary syndromes are needed. Young-onset CRC also tends to show higher expression of tumor cell PD-L1, which is an expected finding, as dMMR cases are more likely to be immunogenic. Two other significant differences are the higher percentage of mucinous carcinoma and the higher tumor grade in young-onset CRC. These two features suggest a more advanced disease with possibly worse outcomes; however, there is no difference in disease stage between the two age groups.

First-line (1L) anti-PD-1 plus chemotherapy in HER2-negative advanced gastric cancer: Real-world evidence from a single Korean center.

83 Background: The implementation of immune checkpoint inhibitor combinations as first-line (1L) treatment has shown promising outcomes and is now the standard of care for HER2-negative advanced gastric cancer. However, real-world evidence supporting this approach as 1L treatment remains limited. Here, we present an analysis of real-world outcomes, comparing 1L anti-PD-1 combined with doublet chemotherapy to the previous standard chemotherapy. Methods: Patient from January 2005 to August 2023 were retrospectively reviewed. The study defined the molecular epidemiology cohort (M cohort) as patients with results for HER2, Epstein-Barr virus (EBV), mismatch repair (MMR), or PD-L1. HER2-negative patients who received palliative first-line chemotherapy doublet regimens with or without PD-1 inhibitor were included. Survival analysis included progression-free survival (PFS) and overall survival (OS), as well as subgroup analyses for molecular subtypes. Results: Total 3,028 were enrolled in the M cohort. The incidence rates of HER2-positive, EBV-positive, and deficient Mismatch Repair (dMMR) were 20.0% (606/3028), 3.8% (93/2432), and 4.7% (126/2660), respectively. PD-L1 testing was conducted on 1,394 patients, with 54.3% (1634/3028) having a CPS (Combined Positive Score) of ≥10, and 47.6% (664/1394) having a CPS of ≥1. Among these patients 1,527 had received chemotherapy doublet, and 153 had received chemotherapy doublet with PD-1 inhibitor as their 1L treatment. During a median follow-up of 56.5 months (95% CI,49.9-66.2), patients who had received PD-1 inhibitor combination therapy showed improved PFS (7.1 vs. 6.1 months; Hazard Ratio [HR], 0.77; 95% Confidence Interval [CI], 0.64–0.93) and OS (18.1 vs. 14.6 months; HR 0.80; 95% CI, 0.63–1.01) compared to those without it. The benefits in PFS from PD-1 inhibitor combination therapy were particularly notable in patients with deficient MMR status (HR, 0.22 vs. 0.80 in those without PD-1 inhibitor combination; p for interaction <0.05) and in patients with signet ring cell carcinoma histology (HR, 0.58 vs. 0.92 in those without PD-1 inhibitor combination; p for interaction <0.05). Conclusions: We observed real-world efficacy of the combination of PD-1 inhibitor and chemotherapy doublet as 1L treatment for HER2-negative advanced gastric cancer, consistent with previous clinical trials. [Table: see text]

Mismatch repair deficiency in bilateral breast cancer

BackgroundSince the FDA approved immune-enhancing therapies for patients with high microsatellite instability (MSI-H) and/or mismatch repair deficiency (dMMR), recognizing these biomarkers in solid tumors has gained clinical importance. Although MSI-H and dMMR are considered uncommon in breast cancer, previous studies on bilateral breast cancer (biBC) identified a surprisingly high frequency of MSI.MethodsIn this study, we aimed to describe the prevalence of dMMR and its association with clinicopathologic parameters in biBC. We performed immunohistochemistry with anti-MMR proteins on tissue microarrays (TMAs) with 58 bilateral breast cancer cases. The biomarkers used were MLH1, PMS2, MSH2, MSH6, ER, PR, HER2 and Ki67. SPSS was used for data analysis.ResultsFour (6.9%) cases showed dMMR on TMAs. Three (75%) of the dMMR cases were luminal and one (25%) was triple negative. Two biBC cases presented unilateral dMMR. No association between dMMR status and clinicopathologic parameters was found.ConclusionsThis work highlights a noticeable frequency of dMMR in bilateral breast cancer and builds upon previous research in this area, suggesting routine MMR protein testing as part of the immunohistochemical panel for biBC to identify candidates for immune-enhancing therapies.

Systemic treatment post curative-intent intervention based on tumor-informed circulating tumor DNA result for oligometastatic colorectal cancer.

84 Background: Local therapies potentially offer a curative approach for patients with oligometastatic colorectal cancer (CRC). The effectiveness of systemic chemotherapy following definitive local treatment in this setting is not well-defined. Tumor-informed circulating tumor DNA (ctDNA) might guide management strategies after curative-intent local treatment. Methods: This is a single institution retrospective study of patients with CRC who underwent curative-intent local therapy to an isolated site of metastatic disease and had post-intervention tumor-informed ctDNA (Signatera) testing. The study was approved by the institutional review board. Clinical characteristics, including ctDNA results, were collected and evaluated. Kaplan-Meier method and log-rank test were used to calculate and compare disease-free survival (DFS). Factors associated with DFS were analyzed using multivariate Cox proportional hazards model. Results: 45 patients were included with median age of 59 years, 62% being male, 91% white, and 53% with stage IV disease at diagnosis. 37% of the patients had RAS mutations, no patient had BRAF mutation, and 2.2% patient had mismatch repair deficient disease. Isolated liver metastases were present in 80% of patients, lung metastases in 11% and other sites in 9%. 44 patients received chemotherapy prior to definitive treatment, with a median treatment duration of 16 weeks. ctDNA tests were done prior to local therapy in 23 patients, with 14 cases (61%) testing positive. Definitive treatment included resection (58%), ablation (13%), radiotherapy (4%), and multimodality (25%). Post-definitive treatment chemotherapy was administered to 24 (53%) patients. ctDNA positivity following definitive therapy was observed in 10 patients (22%), which was associated with a poorer prognosis, with a median DFS of 5.3 months compared to 21.3 months for the ctDNA-negative group (p<0.001). These findings were confirmed on multivariate analysis. In patients who had negative ctDNA after definitive local treatment, median DFS was 14.9 months for those who received post-intervention chemotherapy versus 21.3 months for those without post-intervention chemotherapy (p=0.835). Conclusions: Patients with negative ctDNA results following definitive local therapy for isolated metastatic CRC have better prognosis. Post-intervention chemotherapy in this group of patients was not associated with improved DFS. ctDNA guided omission of post-intervention chemotherapy following curative-intent local treatment of oligometastatic CRC warrants further study. [Table: see text]

Identification of Key Toxic Substances Considering Metabolic Activation: A Combination of Transcriptome and Nontarget Analysis.

There have been numerous studies using effect-directed analysis (EDA) to identify key toxic substances present in source and drinking water, but none of these studies have considered the effects of metabolic activation. This study developed a comprehensive method including a pretreatment process based on an in vitro metabolic activation system, a comprehensive biological effect evaluation based on concentration-dependent transcriptome (CDT), and a chemical feature identification based on nontarget chemical analysis (NTA), to evaluate the changes in the toxic effects and differences in the chemical composition after metabolism. Models for matching metabolites and precursors as well as data-driven identification methods were further constructed to identify toxic metabolites and key toxic precursor substances in drinking water samples from the Yangtze River. After metabolism, the metabolic samples showed a general trend of reduced toxicity in terms of overall biological potency (mean: 3.2-fold). However, metabolic activation led to an increase in some types of toxic effects, including pathways such as excision repair, mismatch repair, protein processing in endoplasmic reticulum, nucleotide excision repair, and DNA replication. Meanwhile, metabolic samples showed a decrease (17.8%) in the number of peaks and average peak area after metabolism, while overall polarity, hydrophilicity, and average molecular weight increased slightly (10.3%). Based on the models for matching of metabolites and precursors and the data-driven identification methods, 32 chemicals were efficiently identified as key toxic substances as main contributors to explain the different transcriptome biological effects such as cellular component, development, and DNA damage related, including 15 industrial compounds, 7 PPCPs, 6 pesticides, and 4 natural products. This study avoids the process of structure elucidation of toxic metabolites and can trace them directly to the precursors based on MS spectra, providing a new idea for the identification of key toxic pollutants of metabolites.

ASO Visual Abstract: Mitochondrial DNA Copy Number as a Biomarker for Guiding Adjuvant Chemotherapy in Stages II and III Colorectal Cancer Patients with Mismatch Repair Deficiency: Seeking Benefits and Avoiding Harms.

ASO Visual Abstract: Mitochondrial DNA Copy Number as a Biomarker for Guiding Adjuvant Chemotherapy in Stages II and III Colorectal Cancer Patients with Mismatch Repair Deficiency: Seeking Benefits and Avoiding Harms.

Clinicopathological and immune characterization of mismatch repair deficient endocervical adenocarcinoma.

Endocervical adenocarcinoma (ECA) is reported increasingly often in young women, and this aggressive disease lacks effective methods of targeted therapy. Since mismatch repair deficiency (dMMR) is an important biomarker for predicting response to immune checkpoint inhibitors, it is important to investigate the clinicopathological features and immune microenvironment of dMMR ECAs. We assessed 617 ECAs from representative tissue microarray sections, gathered clinicopathologic information, reviewed histological characteristics, and performed immunohistochemical staining for MMR, programmed cell death 1 (PD-L1), and other immune markers. Of 617 ECA samples, 20 (3.2%) cases had dMMR. Among them, loss of MMR-related proteins expression was observed in 17/562 (3.0%) human papilloma virus-associated (HPVA) adenocarcinoma and 3/55 (5.5%) non-HPV-associated (NHPVA) adenocarcinoma. In NHPVA cohort, dMMR status was observed in 3 (3/14, 15.0%) patients with clear cells. dMMR ECAs had a higher tendency to have a family history of cancer, larger tumor size, p16 negative, HPV E6/E7 mRNA in situ hybridization (HPV E6/E7 RNAscope) negative, and lower ki-67 index. Among the morphological variables evaluated, poor differentiation, necrosis, stromal tumor-infiltrating lymphocytes, peritumoral lymphocytes, and lymphoid follicles were easily recognized in the dMMR ECAs. In addition, dMMR ECAs had higher CD3+, CD8+, CD38+, CD68+ and PD-1+ immune cells. A relatively high prevalence of PD-L1 expression was observed in dMMR ECAs. dMMR ECAs were significantly more likely to present with a tumor-infiltrating lymphocytes -high/PD-L1-positive status. In conclusion, dMMR ECAs have some specific morphological features and a critical impact on the immune microenvironment, which may provide insights into improving responses to immunotherapy-included comprehensive treatment for ECAs in the future.

Approaches for Lynch syndrome screening and characteristics of subtypes with mismatch repair deficiency in patients with colorectal carcinoma.

To evaluate different Lynch syndrome (LS) screening approaches and establish an efficient and sensitive strategy are critical for clinical practice. In total, 583 patients with colorectal carcinoma (CRC) at Fudan University Shanghai Cancer Center were enrolled. Patient samples were examined by immunohistochemistry (IHC) and next-generation sequencing (NGS), and MLH1 promoter hypermethylation (MPH) was detected in MLH1-deficient cases. Germline genetic testing was performed in cases with deleterious variants and large genomic rearrangements (LGRs) of tumor MMR genes were detected in cases with dMMR or MSI-H cases with no MMR germline variants. Our results showed that triage with IHC and followed by BRAF/MLH1 methylation testing (Strategy 1) identified 93.3% (70/75) of LS cases. IHC followed by germline NGS (Strategy 2) or direct tumor NGS (Strategy 3) both identified 98.7% (74/75) of LS cases. The proportion of LGRs in LS cases was 16.0% (12/75), while 84.0% (63/75) showed SNV/Indel. The average cost per patient was ¥6010.81, ¥6058.48, and ¥8029.98 for Strategy 1, Strategy 2 and Strategy 3, respectively. The average time spent on different strategies was 4.74 days (Strategy 1), 4.89 days (Strategy 2), and 14.50 days (Strategy 3) per patient, respectively. LS and Lynch-like syndrome (LLS) were associated with an earlier onset age than MPH. In conclusion, we compared different workflows for LS screening and IHC plus germline NGS is recommended for LS screening when taking sensitivity, time, and cost into account. Moreover, multiplex ligation-dependent probe amplification made up for the shortcoming of NGS and should be incorporated into routine screening.

Comparison of antigenicity between frozen section vs non-frozen section tissue blocks: An immunohistochemical study of antibodies commonly used in gynecologic pathology.

Frozen section (FS) is a technique widely used intraoperatively to render a preliminary histopathologic diagnosis, allowing for immediate decisions at the time of surgery. We aimed to investigate potential variations in tissue antigenicity induced by rapid freezing in a variety of gynecologic tumor samples. A total of 177 FS and 177 non-frozen section (NFS) tissue slides were tested using a panel of immunostains commonly used in gynecologic pathology, including hormone receptors (estrogen receptor, progesterone receptor), HER2, mismatch repair proteins (MSH6, PMS2), programmed cell death 1 ligand 1 (PD-L1), p53, napsin A, and ɑ-methylacyl coenzyme-A racemase. Immunohistochemistry results were categorized as positive or negative, and positive cases were subsequently scored based on the distribution and intensity of the staining. Certain immunostains, such as HER2, PD-L1, and p53, were scored according to the established guidelines. The overall concordance between FS and NFS blocks was 87%; among the 13% of discrepant cases, most (10.7%) were classified as minor, with only quantitative differences without foreseeable clinical significance. In 2.3% of cases, there were major qualitative changes with potential impact on disease management. We concluded that FS tissue blocks may, in most cases, safely be used for immunohistochemical studies because most discrepant cases showed only minor differences in staining, with no anticipated clinical significance. Nevertheless, for certain markers, including HER2, p53, and PMS2, a NFS block is preferred when that option is available.

Paneth cell differentiation associated with neoadjuvant therapy in esophageal adenocarcinoma.

Paneth cells and Paneth cell metaplasia are well-known in pathology as foundational components in the gastrointestinal system. When within malignant cells (Paneth cell differentiation [PCD]), however, the function and significance of these cells is less well understood. Here, we present findings from the first study focused on PCD in postneoadjuvant esophageal adenocarcinoma (EAC) resection specimens. Patients with EAC treated with neoadjuvant chemoradioation and followed by esophagectomy between 2012 and 2018 in our institution were retrospectively evaluated. A tissue microarray was constructed, and special and immunohistochemical stains were performed. A total of 64 cases were collected, of which 8 had PCD, as highlighted by periodic acid-Schiff with diastase staining. Adenocarcinomas with PCD were more commonly seen in patients 60 to 70 years of age and typically had a poorly differentiated morphology, observationally fewer stromal mucinous changes, and less lymph node metastasis. β-catenin activation induced by neoadjuvant therapy was more frequent in the PCD-positive cases. Patients with PCD-positive disease had low programmed cell death 1 ligand 1 levels, no positive or equivocal ERBB2 (HER2) expression, and low CD8-positive T-cell infiltration; they were also mismatch repair proficient. Patients with PCD-positive disease showed a survival pattern inferior to that of patients with PCD-negative disease. When induced by neoadjuvant therapy in EAC, PCD is associated with high β-catenin activation, less expression of targetable biomarkers, and a potentially worse clinical prognosis.

Right vs. Left colorectal cancer – Where do we draw the line?

PurposeNo consensus on the definition of right and left colorectal cancer (CRC) exists, nor studies offering histological or molecular basis for such categorisation. This study investigated the regional variations in the histological and molecular characteristics of CRCs, with the objective of determining an optimal division point between right and left CRCs. Materials and methodsAn observational study of consecutive patients who underwent CRC resection (1995–2022) at Concord Hospital, Sydney was performed. Clinicopathological data were extracted from a prospective database and seven permutations of right-left divisions considered. Logistic regression tested association between the right-left divisions and pathological characteristics. Receiver operating characteristic and area under the curve (AUC) analyses determined the discriminative ability of each division to predict 18 pathology characteristics. Results3753 patients underwent a CRC resection (2120 male; mean 69.5yrs [SD12.6]). There was regional variation in tumours with respect to tumour infiltrating lymphocytes (TILs), mismatch repair deficiency (dMMR), and mutant BRAF (mBRAF). Left-sided tumours were less likely to demonstrate TILs (P < 0.001), be dMMR (P < 0.001), and express mBRAF (P < 0.001). Division at the descending-sigmoid junction yielded highest discriminative abilities: TILs – AUC 0.66, dMMR – AUC 0.76, and mBRAF – AUC 0.73. ConclusionThis is the first study to provide a pathological basis on which right- and left-sided cancers may be defined, and found the optimal division point between the right and left colorectum to be at the descending-sigmoid junction. Further research is needed to determine whether this can facilitate individualised patient management.

Microsatellite instability at U2AF-binding polypyrimidic tract sites perturbs alternative splicing during colorectal cancer initiation

BackgroundMicrosatellite instability (MSI) due to mismatch repair deficiency (dMMR) is common in colorectal cancer (CRC). These cancers are associated with somatic coding events, but the noncoding pathophysiological impact of this genomic instability is yet poorly understood. Here, we perform an analysis of coding and noncoding MSI events at the different steps of colorectal tumorigenesis using whole exome sequencing and search for associated splicing events via RNA sequencing at the bulk-tumor and single-cell levels.ResultsOur results demonstrate that MSI leads to hundreds of noncoding DNA mutations, notably at polypyrimidine U2AF RNA-binding sites which are endowed with cis-activity in splicing, while higher frequency of exon skipping events are observed in the mRNAs of MSI compared to non-MSI CRC. At the DNA level, these noncoding MSI mutations occur very early prior to cell transformation in the dMMR colonic crypt, accounting for only a fraction of the exon skipping in MSI CRC. At the RNA level, the aberrant exon skipping signature is likely to impair colonic cell differentiation in MSI CRC affecting the expression of alternative exons encoding protein isoforms governing cell fate, while also targeting constitutive exons, making dMMR cells immunogenic in early stage before the onset of coding mutations. This signature is characterized by its similarity to the oncogenic U2AF1-S34F splicing mutation observed in several other non-MSI cancer.ConclusionsOverall, these findings provide evidence that a very early RNA splicing signature partly driven by MSI impairs cell differentiation and promotes MSI CRC initiation, far before coding mutations which accumulate later during MSI tumorigenesis.

Functions of PMS2 and MLH1 important for regulation of divergent repeat-mediated deletions.

Repeat-mediated deletions (RMDs) are a type of deletion rearrangement that utilizes two repetitive elements to bridge a DNA double-strand break (DSB) that leads to loss of the intervening sequence and one of the repeats. Sequence divergence between repeats causes RMD suppression and indeed this divergence must be resolved in the RMD products. The mismatch repair factor, MLH1, was shown to be critical for both RMD suppression and a polarity of sequence divergence resolution in RMDs. Here, we sought to study the interrelationship between these two aspects of RMD regulation (i.e., RMD suppression and polar divergence resolution), by examining several mutants of MLH1 and its binding partner PMS2. To begin with, we show that PMS2 is also critical for both RMD suppression and polar resolution of sequence divergence in RMD products. Then, with six mutants of the MLH1-PMS2 heterodimer, we found several different patterns: three mutants showed defects in both functions, one mutant showed loss of RMD suppression but not polar divergence resolution, whereas another mutant showed the opposite, and finally one mutant showed loss of RMD suppression but had a complex effect on polar divergence resolution. These findings indicate that RMD suppression vs. polar resolution of sequence divergence are distinct functions of MLH1-PMS2.