Clinicopathological Characteristics and Mutational Landscape of APC, HOXB13, and KRAS among Rwandan Patients with Colorectal Cancer.

Cancer research in Rwanda is estimated to be less than 1% of the total African cancer research output with limited research on colorectal cancer (CRC). Rwandan patients with CRC are young, with more females being affected than males, and most patients present with advanced disease. Considering the paucity of oncological genetic studies in this population, we investigated the mutational status of CRC tissues, focusing on the Adenomatous polyposis coli (APC), Kirsten rat sarcoma (KRAS), and Homeobox B13 (HOXB13) genes. Our aim was to determine whether there were any differences between Rwandan patients and other populations. To do so, we performed Sanger sequencing of the DNA extracted from formalin-fixed paraffin-embedded adenocarcinoma samples from 54 patients (mean age: 60 years). Most tumors were located in the rectum (83.3%), and 92.6% of the tumors were low-grade. Most patients (70.4%) reported never smoking, and 61.1% of patients had consumed alcohol. We identified 27 variants of APC, including 3 novel mutations (c.4310_4319delAAACACCTCC, c.4463_4470delinsA, and c.4506_4507delT). All three novel mutations are classified as deleterious by MutationTaster2021. We found four synonymous variants (c.330C>A, c.366C>T, c.513T>C, and c.735G>A) of HOXB13. For KRAS, we found six variants (Asp173, Gly13Asp, Gly12Ala, Gly12Asp, Gly12Val, and Gln61His), the last four of which are pathogenic. In conclusion, here we contribute new genetic variation data and provide clinicopathological information pertinent to CRC in Rwanda.

Suprapapillary placement of plastic versus metal stents for malignant biliary hilar obstructions: a multicenter, randomized trial.

BACKGROUND AND AIMS: The efficacy of the suprapapillary placement of inside plastic stents (iPSs) for unresectable malignant hilar biliary obstructions (MHOs) is unknown compared with that of uncovered inside metal stents (iMSs). This randomized controlled trial was designed to evaluate the outcomes of endoscopic placement of these stents for unresectable MHOs. METHODS: This open-label, randomized study was conducted at 12 Japanese institutions. The enrolled patients with unresectable MHOs were allocated to iPS and iMS groups. The primary outcome was defined as the time to recurrent biliary obstruction in patients for whom the intervention was both technically and clinically successful. RESULTS: Among 87 enrollments, 38 patients in the iPS group and 46 patients in the iMS group were analyzed. Technical success rates were 100% (38 of 38) and 96.6% (44 of 46), respectively (P = 1.00). After transferring 1 unsuccessful iMS-group patient to the iPS group (since iPSs were deployed), the clinical success rates were 90.0% (35 of 39) for the iPS group and 88.9% (40 of 45) for the iMS group from a per-protocol analysis (P = 1.00). Among the patients with clinical success, the median times to recurrent biliary obstruction were 250 (95% confidence interval, 85-415) and 361 (95% confidence interval, 107-615) days (log-rank test, P = .34). No differences were detected in rates of adverse events. CONCLUSIONS: This Phase II randomized trial did not show any statistically significant difference in stent patency between suprapapillary plastic versus metal stents. Considering the potential advantages of plastic stents for malignant hilar obstruction, these findings suggest that suprapapillary plastic stents could be a viable alternative to metal stents for this condition.

Isolation of New Colibactin Metabolites from Wild-Type Escherichia coli and In Situ Trapping of a Mature Colibactin Derivative.

Colibactin is a polyketide-nonribosomal peptide hybrid secondary metabolite that can form interstrand cross-links in double-stranded DNA. Colibactin-producing Escherichia coli has also been linked to colorectal oncogenesis. Thus, there is a strong interest in understanding the role colibactin may play in oncogenesis. Here, using the high-colibactin-producing wild-type E. coli strain we isolated from a clinical sample with the activity-based fluorescent probe we developed earlier, we were able to identify colibactin 770, which was recently identified and proposed as the complete form of colibactin, along with colibactin 788, 406, 416, 420, and 430 derived from colibactin 770 through structural rearrangements and solvolysis. Furthermore, we were able to trap the degrading mature colibactin species by converting the diketone moiety into quinoxaline in situ in the crude culture extract to form colibactin 860 at milligram scale. This allowed us to determine the stereochemically complex structure of the rearranged form of an intact colibactin, colibactin 788, in detail. Furthermore, our study suggested that we were capturing only a few percent of the actual colibactin produced by the microbe, providing a crude quantitative insight into the inherent instability of this compound. Through the structural assignment of colibactins and their degradative products by the combination of LC-HRMS and NMR spectroscopies, we were able to elucidate further the fate of inherently unstable colibactin, which could help acquire a more complete picture of colibactin metabolism and identify key DNA adducts and biomarkers for diagnosing colorectal cancer.

Mother-to-infant transmission of the carcinogenic colibactin-producing bacteria.

BACKGROUND: The Escherichia coli strain that is known to produce the genotoxic secondary metabolite colibactin is linked to colorectal oncogenesis. Therefore, understanding the properties of such colibactin-positive E. coli and the molecular mechanism of oncogenesis by colibactin may provide us with opportunities for early diagnosis or prevention of colorectal oncogenesis. While there have been major advances in the characterization of colibactin-positive E. coli and the toxin it produces, the infection route of the clb + strain remains poorly characterized. RESULTS: We examined infants and their treatments during and post-birth periods to examine potential transmission of colibactin-positive E. coli to infants. Here, analysis of fecal samples of infants over the first month of birth for the presence of a colibactin biosynthetic gene revealed that the bacterium may be transmitted from mother to infant through intimate contacts, such as natural childbirth and breastfeeding, but not through food intake. CONCLUSIONS: Our finding suggests that transmission of colibactin-positive E. coli appears to be occurring at the very early stage of life of the newborn and hints at the possibility of developing early preventive measures against colorectal cancer.

[Pulse steroid therapy to treat liver injury with risk of liver failure due to immune-related adverse events:a case report].

A 75-year-old woman was diagnosed with clinical stage III lung cancer. The patient was treated with chemoradiotherapy and subsequent durvalumab, an anti-PD-L1 antibody immune checkpoint inhibitor (ICI). Liver dysfunction was observed 14 days after the start of durvalumab therapy (aspartate transaminase, 218U/l;alanine aminotransferase, 169U/l). This corresponded to a grade 3 adverse event according to the Common Terminology Criteria for Adverse Events. The second course of durvalumab was withheld. The patient was hospitalized 31 days after durvalumab therapy because of worsening liver dysfunction. Laboratory findings and imaging examinations suggested liver injury due to an immune-related adverse event (irAE). Liver biopsy performed 38 days after durvalumab therapy showed severe lymphocyte and plasma cell infiltration into the portal tract, focal necrosis in the hepatic lobules, and necrotic changes around the hepatic lobules. These findings were similar to those of autoimmune hepatitis (AIH). Immunohistochemical results revealed infiltration of CD3- and CD8-positive lymphocytes and mild infiltration of CD4-positive lymphocytes. Pathological findings in the liver tissue were consistent with an irAE. Jaundice worsened and the prothrombin time was prolonged, leading to a risk of progression to liver failure. Thus, pulse steroid therapy was performed with methylprednisolone (mPSL) starting at 0.8mg/kg. Liver dysfunction lessened and the mPSL dose was gradually reduced. Moreover, ICIs exert antitumor effects by inhibiting the immune checkpoint system but can cause irAEs in various organs. Liver injury is also relatively common. Liver tissue findings are similar to those in AIH, but immunostaining reveals the presence of numerous CD8-positive lymphocytes. Fewer CD4-positive lymphocytes exist in irAE-associated liver injury than in AIH. Medical departments must cooperate and effectively manage irAEs because ICIs are increasingly being used and can occur in organs throughout the body. In principle, irAEs are treated with steroids. Thus, high-dose steroids diminishing the therapeutic effect of ICIs is a concern, and it is important to control irAEs with low-dose steroids that are started earlier.

Distinct prognostic roles and heterogeneity of TTF1 copy number and TTF1 protein expression in non-small cell lung cancer.

Thyroid transcription factor 1 (TTF1) located on chromosome band 14q13.3 is an oncogene and a suppressor gene in non-small cell lung cancer (NSCLC). The prognostic relevance of TTF1 copy number alterations (CNAs) and their association with TTF1 protein expression are poorly understood. Here, we assessed TTF1 CNAs and protein expression using microarrays in a cohort of 636 NSCLC, including 423 adenocarcinoma (ADC) and 171 squamous cell carcinoma (SCC). In addition, fluorescent in situ hybridization and immunohistochemistry were performed. TTF1 CNAs were detected in 23% of NSCLC (23% of ADC and 20% of SCC). Specifically, TTF1 amplification and polysomy were observed in 5% and 18% of NSCLC, and in 7% and 16% of ADC, respectively. TTF1 expression was detected in 85% of ADC. TTF1 CNAs were significantly associated with advanced tumor stage, EGFR mutations, and TTF1 expression. A multivariate Cox hazards model analysis of overall survival and recurrence-free survival demonstrated that both TTF1 amplification and polysomy were independent indicators of an unfavorable prognosis in patients with NSCLC. Survival was inversely correlated with TTF1 copy number. In contrast, TTF1 protein expression was an independent favorable prognostic factor. Intratumoral and intertumoral heterogeneities of TTF1 CNAs and TTF1 protein expression were assessed using primary cores from 138 pairs of primary tumors and corresponding nodal metastases. The concordance rate for TTF1 CNAs and TTF1 protein expression was high within tumors and between primary and metastatic tumors. Altogether, these results suggest that TTF1 CNAs are correlated with TTF1 protein expression, but have opposing effects on survival.

Characterization of V-set and immunoglobulin domain containing 1 exerting a tumor suppressor function in gastric, lung, and esophageal cancer cells.

V-set and immunoglobulin domain containing 1 (VSIG1) is a newly discovered member of the immunoglobulin superfamily of proteins, expressed in normal stomach and testis. In cancers, however, the clinical and biological roles of VSIG1 remain unknown. Here we investigated VSIG1 expression in 11 cancers and assessed the prognostic roles of VSIG1 in patients with gastric cancer (GC) (n = 362) and non-small-cell lung cancer (n = 650). V-set and immunoglobulin domain containing 1 was downregulated in 60.5% of GC specimens, and high VSIG1 expression was identified as an independent favorable prognostic factor for overall survival in GC patients (hazard ratio, 0.58; 95% confidence interval, 0.35-0.96). Among lung adenocarcinomas (n = 428), VSIG1 was significantly and inversely associated with thyroid transcription factor 1 expression and was frequently expressed in the invasive mucinous subtype (17 of 19, 89.5%). In addition, VSIG1 was expressed in a subset of pancreatic, ovarian, and prostate cancers. The variant 2 VSIG1 transcript was the dominant form in these tissues and cancer cells. Introduction of VSIG1 significantly reduced the proliferative ability of MKN1 and MKN28 GC cells and H1299 lung cancer cells and downregulated cell migration of these cells, as well as of KYSE150, an esophageal cancer cell line. Cell invasion of MKN1, MKN28, and KYSE150 cells was also reduced by VSIG1 introduction. In vitro characterization revealed that VSIG1 forms homodimers through homophilic cis-interactions but not through homophilic trans-interactions. These results suggest that VSIG1 possesses tumor suppressive functions that are translated into favorable prognosis of VSIG1-expressing GC patients.

Diagnostic efficacy of liquid-based cytology for solid pancreatic lesion samples obtained with endoscopic ultrasound-guided fine-needle aspiration: Propensity score-matched analysis.

BACKGROUND AND AIM: There is a paucity of data on the diagnostic efficacy of liquid-based cytology (LBC) for pancreatic samples obtained by endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA). Using propensity score matching, we retrospectively analyzed the additional diagnostic value of LBC compared to a conventional Papanicolaou smear (CPS) for samples of solid pancreatic lesions obtained by EUS-FNA. METHODS: This cohort study included 126 matched patients who underwent initial EUS-FNA for solid pancreatic lesions between January 2009 and August 2014. CPS was used for cytology of EUS-FNA samples obtained until May 2012 (63 patients). Subsequently, LBC was used for cytological analysis (63 patients). Diagnostic yields of CPS and LBC for malignancy were compared. Risk factors for cytological misdiagnosis with LBC were investigated. RESULTS: Overall rate of malignancy was 86% after matching. LBC had higher diagnostic sensitivity and accuracy than CPS (96.6% vs 84.0%, P = 0.03; and 96.8% vs 87.3%, P = 0.05). LBC was significantly more sensitive for diagnosing pancreatic head lesions (96.4% vs 78.1%, P = 0.04). The sensitivity for pancreatic ductal adenocarcinoma (PDAC) with LBC was higher (98.1% vs 83.0%, P = 0.009). Multivariate analysis revealed that malignant tumors other than PDAC (P = 0.004) and lesion size ≤20 mm (P = 0.046) were risk factors for LBC misdiagnosis in all participants. CONCLUSIONS: For solid pancreatic lesions, LBC of EUS-FNA samples contributes to the diagnosis of malignancy. Malignant tumors other than PDAC and small tumors are difficult to diagnose using EUS-FNA and LBC.

[Jejunal duplication in an adult presenting with hematochezia].

A 56-year-old man was admitted to our hospital with appetite loss, palpitations, orthostatic syncope, and hematochezia. Contrast-enhanced abdominal computed tomography (CT) revealed a proximal jejunal diverticulum with contrast extravasation. We immediately performed transoral double balloon enteroscopy (DBE) to treat the bleed in the jejunum, and this revealed a small ulcer with an exposed vessel at the opening of the jejunal diverticulum. Hemostasis was achieved endoscopically with argon plasma coagulation (APC) and hemoclips. During subsequent surgery, the diverticulum was found on the mesenteric side of the jejunum. We performed laparoscopy-assisted partial resection of the jejunum, and pathological examination showed that the diverticulum shared a common proper muscle layer with the jejunum and was covered by jejunal mucosa with no ectopic mucosa. Therefore, we diagnosed jejunal duplication. After hospital discharge, the patient had no recurrence of hematochezia or anemia. We report a rare case of jejunal duplication presenting with hematochezia, which was diagnosed as jejunal diverticular bleeding by CT and DBE before surgery. Pathological analysis confirmed jejunal duplication after surgery. We suggest that intestinal diverticular bleeding, as well as duplication of the gastrointestinal tract, should be considered as part of the differential diagnosis of obscure gastrointestinal bleeding.

The spectrum of TP53 mutations in Rwandan patients with gastric cancer.

BACKGROUND: Gastric cancer is the sixth most frequently diagnosed cancer and third in causing cancer-related death globally. The most frequently mutated gene in human cancers is TP53, which plays a pivotal role in cancer initiation and progression. In Africa, particularly in Rwanda, data on TP53 mutations are lacking. Therefore, this study intended to obtain TP53 mutation status in Rwandan patients with gastric cancer. RESULTS: Formalin-fixed paraffin-embedded tissue blocks of 95 Rwandan patients with histopathologically proven gastric carcinoma were obtained from the University Teaching Hospital of Kigali. After DNA extraction, all coding regions of the TP53 gene and the exon-intron boundary region of TP53 were sequenced using the Sanger sequencing. Mutated TP53 were observed in 24 (25.3%) of the 95 cases, and a total of 29 mutations were identified. These TP53 mutations were distributed between exon 4 and 8 and most of them were missense mutations (19/29; 65.5%). Immunohistochemical analysis for TP53 revealed that most of the TP53 missense mutations were associated with TP53 protein accumulation. Among the 29 mutations, one was novel (c.459_477delCGGCACCCGCGTCCGCGCC). This 19-bp deletion mutation in exon 5 caused the production of truncated TP53 protein (p.G154Wfs*10). Regarding the spectrum of TP53 mutations, G:C > A:T at CpG sites was the most prevalent (10/29; 34.5%) and G:C > T:A was the second most prevalent (7/29; 24.1%). Interestingly, when the mutation spectrum of TP53 was compared to three previous TP53 mutational studies on non-Rwandan patients with gastric cancer, G:C > T:A mutations were significantly more frequent in this study than in our previous study (p = 0.013), the TCGA database (p = 0.017), and a previous study on patients from Hong Kong (p = 0.006). Even after correcting for false discovery, statistical significance was observed. CONCLUSIONS: Our results suggested that TP53 G:C > T:A transversion mutation in Rwandan patients with gastric cancer is more frequent than in non-Rwandan patients with gastric cancer, indicating at an alternative etiological and carcinogenic progression of gastric cancer in Rwanda.

Genome-wide repression of NF-κB target genes by transcription factor MIBP1 and its modulation by O-linked ß-N-acetylglucosamine (O-GlcNAc) transferase.

The transcription factor c-MYC intron binding protein 1 (MIBP1) binds to various genomic regulatory regions, including intron 1 of c-MYC. This factor is highly expressed in postmitotic neurons in the fetal brain and may be involved in various biological steps, such as neurological and immunological processes. In this study, we globally characterized the transcriptional targets of MIBP1 and proteins that interact with MIBP1. Microarray hybridization followed by gene set enrichment analysis revealed that genes involved in the pathways downstream of MYC, NF-κB, and TGF-ß were down-regulated when HEK293 cells stably overexpressed MIBP1. In silico transcription factor binding site analysis of the promoter regions of these down-regulated genes showed that the NF-κB binding site was the most overrepresented. The up-regulation of genes known to be in the NF-κB pathway after the knockdown of endogenous MIBP1 in HT1080 cells supports the view that MIBP1 is a down-regulator of the NF-κB pathway. We also confirmed the binding of the MIBP1 to the NF-κB site. By immunoprecipitation and mass spectrometry, we detected O-linked ß-N-acetylglucosamine (O-GlcNAc) transferase as a prominent binding partner of MIBP1. Analyses using deletion mutants revealed that a 154-amino acid region of MIBP1 was necessary for its O-GlcNAc transferase binding and O-GlcNAcylation. A luciferase reporter assay showed that NF-κB-responsive expression was repressed by MIBP1, and stronger repression by MIBP1 lacking the 154-amino acid region was observed. Our results indicate that the primary effect of MIBP1 expression is the down-regulation of the NF-κB pathway and that this effect is attenuated by O-GlcNAc signaling.

Transgenic expression of human neutrophil peptide-1 enhances hepatic fibrosis in mice fed a choline-deficient, L-amino acid-defined diet.

BACKGROUND: Neutrophils infiltrate the livers of patients with nonalcoholic steatohepatitis (NASH). Human neutrophil peptides (HNPs) induce cytokine and chemokine production under inflammatory conditions, which may contribute to the progression of NASH. In this study, we focused on the effects of HNP-1 on hepatic steatosis and fibrosis in a mouse model of NASH induced by a choline-deficient, L-amino acid-defined (CDAA) diet. MATERIALS & METHODS: We generated transgenic mice expressing HNP-1 under the control of a ß-actin-based promoter. HNP-1 transgenic and wild-type C57BL/6N mice were fed a CDAA diet for 16 weeks to induce hepatic steatosis and fibrosis. Serological and histological features were examined, and the effects of HNP-1 on hepatic stellate cell lines were assessed. RESULTS: HNP-1 transgenic and wild-type mice fed the CDAA diet showed no significant differences in serum alanine aminotransferase levels or the degree of hepatic steatosis based on Oil red O staining and hepatic triglyceride content. In contrast, Sirius Red and Azan staining showed significantly more severe hepatic fibrosis in HNP-1 transgenic mice compared with wild-type mice. In addition, significantly more α-smooth muscle actin-positive hepatic stellate cells were observed in the transgenic mice than in the wild-type mice. Finally, the proliferation of the LI90 hepatic stellate cell line increased in response to HNP-1. CONCLUSION: Our data indicate that HNP-1 enhances hepatic fibrosis in fatty liver by inducing hepatic stellate cell proliferation. Thus, neutrophil-derived HNP-1 may contribute to the progression of NASH.

Efficacy and safety of a novel anti-reflux metal stent during neoadjuvant chemotherapy for pancreatic cancer: A prospective multicenter exploratory study.

BACKGROUND/PURPOSE: The benefits of anti-reflux metal stents, used for treating biliary obstruction in patients receiving neoadjuvant chemotherapy (NAC) for pancreatic cancer, are yet unknown. Herein, the safety and efficacy of the novel duckbill-type anti-reflux metal stent (D-ARMS) were prospectively evaluated for biliary drainage. Additionally, the incidence of recurrent biliary obstruction (RBO) after placement of D-ARMS vs conventional covered self-expandable metal stents (CCSEMSs) was retrospectively compared. METHODS: Patients who received D-ARMS (n = 33) for treatment of distal biliary obstruction before NAC between September 2019 and January 2021 and those that received CCSEMSs (n = 38) between January 2013 and August 2019 were included in the historical control group. Technical and clinical successes, rate of RBO, and cumulative incidence of RBO were compared between the two groups. RESULTS: The technical success rate was 100% for both the D-ARMS and CCSEMS groups, and the clinical success rate were not significantly different (93.9% and 89.5%, respectively; P = .68). In the multivariate analysis, D-ARMS was identified as the independent factor for cumulative incidence of RBO (P = .03). The cumulative incidence of RBO was significantly lower in the D-ARMS group than that in the CCSEMS group (P = .04). CONCLUSIONS: D-ARMS is safe and effective for patients receiving NAC.

TP53 mutations in Romanian patients with colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) has been ranked as the second most deadly cancer and the third most diagnosed cancer cases for the year 2020. Specifically for Romania, the number of CRC-related deaths in 2019 was estimated at 6307 people, with a standardized mortality rate of 33.8 per 100,000 inhabitants. Although the tumor protein 53 (TP53) gene is intensively studied, there are few data on TP53 mutations in Romanian CRC. Furthermore, since genetic alterations may show geographical differences, our study aimed to analyze the clinical status and TP53 somatic variation in Romanian CRC patients. SUBJECTS AND METHODS: DNA from 40 randomly selected cases of CRC was extracted from formalin-fixed paraffin-embedded tissues and sequenced using direct Sanger sequencing techniques, and variants were annotated according to the recommendations of the Human Genome Variation Society. Novel variants were analyzed using MutationTaster2021 to predict their effects. RESULTS: The mean age was 63.6 years (range 33-85 years) with a male to female ratio of 2.3. More than 45% (18/40) had an advanced cancer stage (≥ stage III). Mutations were found in 21/40 cases (52.5%), with one case having two mutations, giving a total of twenty-two mutations in the TP53 coding DNA. These mutations include 3 (13.6%) insertion-deletion mutations, two of which are novel frameshift mutations: c.165delT (in exon 4) and c.928_935dup (in exon 9), both of which are predicted to lead to nonsense-mediated mRNA decay and are classified as deleterious. The remaining 19 (86.36%) were substitution mutations: 1 nonsense and 18 (81.8%) missense mutations, with G > A (n = 7/19; 36.8%) and C > T (n = 6/19; 31.5%) transitions being the most common. The G > T transversion was found in 21.05% (4/19) of the substitution mutations. CONCLUSION: We have described two novel frameshift mutations in TP53. The discovery of novel mutations following the efforts of The Cancer Genome Atlas and other large-scale cancer genome sequencing projects may be further evidence of the heterogeneous nature of mutations in cancer and may indicate that the identification of carcinogenic mutations is not yet saturated. Further sequencing is therefore needed, especially in less studied populations. Importantly, consideration of their geographical environment will shed light on population-specific carcinogenesis.

High Expression of Fas-Associated Factor 1 Indicates a Poor Prognosis in Non-Small-Cell Lung Cancer.

Fas-associated factor 1 (FAF1) is a death-promoting protein identified as an interaction partner of the death receptor Fas. The downregulation and mutation of FAF1 have been reported in a variety of human tumors, but there have been few studies on lung cancer. Here, we investigated the prognostic significance of FAF1 expression in non-small-cell lung cancer (NSCLC), and whether aberrant FAF1 expression may be involved in the pathogenesis and prognosis of NSCLC. FAF1 expression was examined in NSCLC specimens as well as human lung cancer cell lines. In addition, changes in cell viability and apoptosis upon regulating FAF1 expression were investigated in lung cancer cell lines. As a result, high FAF1 expression was significantly associated with a poor prognosis in NSCLC. In lung cancer cell lines, FAF1 downregulation hindered cell viability and tended to promote early apoptosis. In conclusion, this is the first study of the clinical significance of FAF1 in NSCLC, showing that FAF1 overexpression is associated with a poor prognosis in NSCLC and that FAF1 acts as a dangerous factor rather than an apoptosis promoter in NSCLC.

Both MLH1 deficiency and BRAFV600E mutation are a unique characteristic of colorectal medullary carcinoma: An observational study.

Although immunohistochemistry (IHC) for mismatch repair (MMR) proteins (MMR IHC) is used to identify DNA MMR status, universal screening of all patients with colorectal cancer (CRC) using a combination of both MMR IHC and genetic testing for the BRAFV600E mutation is limited in Japan. This study aimed to better understand the histopathological characteristics of CRCs, which exhibit both deficient mismatch repair (dMMR) and BRAFV600E mutation. MMR IHC of formalin-fixed paraffin-embedded tissues from tumor areas obtained from 651 patients with CRC who underwent surgical resection at Hamamatsu University Hospital (Hamamatsu, Japan) between August 2016 and March 2022 were used to evaluate MMR status, which was determined by staining for the expression of 4 MMR proteins (MLH1, MSH2, PMS2, and MSH6). All dMMR tumors were additionally evaluated for BRAFV600 mutation status via Sanger sequencing. Patient clinical characteristics (age, sex, tumor location, size, and tumor pathology) were then classified using their dMMR and BRAFV600 mutation statuses. Among the 651 patients with CRC, 58 carried tumors with dMMR, of which 52 were deficiency in MLH1 (dMLH1). Interestingly, all 16 medullary carcinomas that were analyzed showed characteristics corresponding to the presence of both dMLH1 and BRAFV600E mutation (P = .01). These results suggest that colorectal medullary carcinomas can be diagnosed based on their unique characteristics of harboring the BRAFV600E mutation and exhibiting dMLH1 expression.

Comparison of recurrent biliary obstruction with the use of metal and plastic stents in EUS-guided biliary drainage: A propensity score-matched analysis.

Background and Objectives: In transpapillary biliary drainage, metal stents (MSs) exhibit a lower incidence of a biliary obstruction than plastic stents (PSs). However, few studies have compared recurrent biliary obstruction (RBO) when MSs and PSs are used in EUS-guided hepaticogastrostomy (EUS-HGS) and choledochoduodenostomy (EUS-CDS). We retrospectively evaluated the RBO for both stents in each procedure. Patients and Methods: : Between November 2012 and December 2020, 85 and 53 patients who underwent EUS-HGS and EUS-CDS for unresectable malignant biliary obstruction, respectively, were enrolled. Factors associated with RBO were assessed. Clinical outcomes were compared between the MS and PS groups using propensity score matching. Results: : The clinical success rate and procedure-related adverse events were similar in the MS and PS groups. Multivariate analysis identified the use of PS as a factor associated with RBO (EUS-HGS, P = 0.03; EUS-CDS, P = 0.02). After matching, the median time to RBO in EUS-HGS (MS: 313; PS: 125 days; P = 0.01) in the MS group was longer than that in the PS group. The cumulative incidence of RBO at 1, 3, and 6 months in the MS group was significantly lower than that in the PS group for EUS-HGS (MS: 4.0%, 8.2%, and 8.2%; PS: 12.4%, 24.9%, and 39.5%, respectively, P = 0.01). Conclusions: : MS exhibited a lower rate of RBO than PS for EUS-HGS and EUS-CDS.

Non-CpG sites preference in G:C > A:T transition of TP53 in gastric cancer of Eastern Europe (Poland, Romania and Hungary) compared to East Asian countries (China and Japan).

AIM: Mutation spectrum of TP53 in gastric cancer (GC) has been investigated world-widely, but a comparison of mutation spectrum among GCs from various regions in the world are still sparsely documented. In order to identify the difference of TP53 mutation spectrum in GCs in Eastern Europe and in East Asia, we sequenced TP53 in GCs from Eastern Europe, Lujiang (China), and Yokohama, Kanagawa (Japan) and identified the feature of TP53 mutations of GC in these regions. SUBJECTS AND METHOD: In total, 689 tissue samples of GC were analyzed: 288 samples from East European populations (25 from Hungary, 71 from Poland and 192 from Romania), 268 from Yokohama, Kanagawa, Japan and 133 from Lujiang, Anhui province, China. DNA was extracted from FFPE tissue of Chinese, East European cases; and from frozen tissue of Japanese GCs. PCR products were direct-sequenced by Sanger method, and in ambiguous cases, PCR product was cloned and up to 8 clones were sequenced. We used No. NC_000017.11(hg38) as the reference sequence of TP53. Mutation patterns were categorized into nine groups: six base substitutions, insertion, deletion and deletion-insertion. Within G:C > A:T mutations the mutations in CpG and non-CpG sites were divided. The Cancer Genome Atlas data (TCGA, ver.R20, July, 2019) having somatic mutation list of GCs from Whites, Asians, and other ethnicities were used as a reference for our data. RESULTS: The most frequent base substitutions were G:C > A:T transition in all the areas investigated. The G:C > A:T transition in non-CpG sites were prominent in East European GCs, compared with Asian ones. Mutation pattern from TCGA data revealed the same trend between GCs from White (TCGA category) vs Asian countries. Chinese and Japanese GCs showed higher ratio of G:C > A:T transition in CpG sites and A:T > G:C mutation was more prevalent in Asian countries. CONCLUSION: The divergence in mutation spectrum of GC in different areas in the world may reflect various pathogeneses and etiologies of GC, region to region. Diversified mutation spectrum in GC in Eastern Europe may suggest GC in Europe has different carcinogenic pathway of those from Asia.

Induction of DNA Damage in Mouse Colorectum by Administration of Colibactin-producing Escherichia coli, Isolated from a Patient With Colorectal Cancer.

BACKGROUND/AIM: Among colorectal cancer-associated intestinal microbiota, colibactin-producing (clb+) bacteria are attracting attention. We aimed to clarify the interaction between clb+ Escherichia coli and normal colorectal epithelial cells in vivo and in vitro. MATERIALS AND METHODS: Five-week-old female Balb/c mice were divided in an untreated group, a group treated with clb+ E. coli isolated from a Japanese patient with colorectal cancer (E. coli-50), and a group treated with non colibactin-producing E. coli (E. coli-50/ΔclbP). Mice were sacrificed at 18 weeks of treatment. RESULTS: Treatment with clb+ E. coli increased positivity for H2A histone family member X phosphorylated at Ser-139 (γH2AX) in epithelial cells of the luminal surface of the mouse rectum but this did not occur in the E. coli-50/ΔclbP and untreated groups. In an in vitro setting, the ratio of apoptotic cells was increased and cell counts were reduced by treatment with clb+ E. coli more than in untreated cells and normal rat colorectal epithelial cells. CONCLUSION: E. coli-50 induced DNA damage in the mouse rectum, possibly by direct interaction between clb+ E. coli and normal colorectal epithelial cells. Our findings imply that regulation of clb+ E. coli infection may be a useful strategy for colorectal cancer control.

m6A demethylase ALKBH5 promotes tumor cell proliferation by destabilizing IGF2BPs target genes and worsens the prognosis of patients with non-small-cell lung cancer.

The modification of N6-methyladenosine (m6A) in RNA and its eraser ALKBH5, an m6A demethylase, play an important role across various steps of human carcinogenesis. However, the involvement of ALKBH5 in non-small-cell lung cancer (NSCLC) development remains to be completely elucidated. The current study revealed that the expression of ALKBH5 was increased in NSCLC and increased expression of ALKBH5 worsened the prognosis of patients with NSCLC. In vitro study revealed that ALKBH5 knockdown suppressed cell proliferation ability of PC9 and A549 cells and promoted G1 arrest and increased the number of apoptotic cells. Furthermore, ALKBH5 overexpression increased the cell proliferation ability of the immortalized cell lines. Microarray analysis and western blotting revealed that the expression of CDKN1A (p21) or TIMP3 was increased by ALKBH5 knockdown. These alterations were offset by a double knockdown of both ALKBH5 and one of the IGF2BPs. The decline of mRNAs was, at least partly, owing to the destabilization of these mRNAs by one of the IGF2BPs. In conclusions, the ALKBH5-IGF2BPs axis promotes cell proliferation and tumorigenicity, which in turn causes the unfavorable prognosis of NSCLC.