Noriyuki Horiguchi

Mutation spectrum of TP53 gene predicts clinicopathological features and survival of gastric cancer

Background and aim TP53 gene is frequently mutated in gastric cancer (GC), but the relationship with clinicopathological features and prognosis is conflicting. Here, we screened TP53 mutation spectrum of 214 GC patients in relation to their clinicopathological features and prognosis. Results TP53 nonsilent mutations were detected in 80 cases (37.4%), being frequently occurred as C:G to T:A single nucleotide transitions at 5′-CpG-3′ sites. TP53 mutations occurred more frequently in differentiated histologic type than in undifferentiated type in the early stage (48.6% vs. 7%, P=0.0006), while the mutations correlated with venous invasion among advanced stage (47.7% vs. 20.7%, P=0.04). Subset of GC with TP53 hot spot mutations (R175, G245, R248, R273, R282) presented significantly worse overall survival and recurrence free survival compared to others (both P=0.001). Methods Matched biopsies from GC and adjacent tissues from 214 patients were used for the experiment. All coding regions of TP53 gene (exon2 to exon11) were examined using Sanger sequencing. Conclusion Our data suggest that GC with TP53 mutations seems to develop as differentiated histologic type and show aggressive biological behavior such as venous invasion. Moreover, our data emphasizes the importance of discriminating TP53 hot spot mutations (R175, G245, R248, R273, R282) to predict worse overall survival and recurrence free survival of GC patients.

Potential link between Fusobacterium enrichment and DNA methylation accumulation in the inflammatory colonic mucosa in ulcerative colitis

BACKGROUND AND AIM Fusobacterium enrichment has been associated with colorectal cancer development. Ulcerative colitis (UC) associated tumorigenesis is characterized as high degree of methylation accumulation through continuous colonic inflammation. The aim of this study was to investigate a potential link between Fusobacterium enrichment and DNA methylation accumulation in the inflammatory colonic mucosa in UC. METHODS In the candidate analysis, inflamed colonic mucosa from 86 UC patients were characterized the methylation status of colorectal a panel of cancer related 24 genes. In the genome-wide analysis, an Infinium HumanMethylation450 BeadChip array was utilized to characterize the methylation status of >450,000 CpG sites for fourteen UC patients. Results were correlated with Fusobacterium status. RESULTS UC with Fusobacterium enrichment (FB-high) was characterized as high degree of type C (for cancer-specific) methylation compared to other (FB-low/neg) samples (P<0.01). Genes hypermethylated in FB-high samples included well-known type C genes in colorectal cancer, such as MINT2 and 31, P16 and NEUROG1. Multivariate analysis demonstrated that the FB high status held an increased likelihood for methylation high as an independent factor (odds ratio: 16.18, 95% confidence interval: 1.94-135.2, P=0.01). Genome-wide methylation analysis demonstrated a unique methylome signature of FB-high cases irrespective of promoter, outside promoter, CpG and non-CpG sites. Group of promoter CpG sites that were exclusively hypermethylated in FB-high cases significantly codified the genes related to the catalytic activity (P=0.039). CONCLUSION Our findings suggest that Fusobacterium accelerates DNA methylation in specific groups of genes in the inflammatory colonic mucosa in UC.BACKGROUND AND AIM Fusobacterium enrichment has been associated with colorectal cancer development. Ulcerative colitis (UC) associated tumorigenesis is characterized as high degree of methylation accumulation through continuous colonic inflammation. The aim of this study was to investigate a potential link between Fusobacterium enrichment and DNA methylation accumulation in the inflammatory colonic mucosa in UC. METHODS In the candidate analysis, inflamed colonic mucosa from 86 UC patients were characterized the methylation status of colorectal a panel of cancer related 24 genes. In the genome-wide analysis, an Infinium HumanMethylation450 BeadChip array was utilized to characterize the methylation status of >450,000 CpG sites for fourteen UC patients. Results were correlated with Fusobacterium status. RESULTS UC with Fusobacterium enrichment (FB-high) was characterized as high degree of type C (for cancer-specific) methylation compared to other (FB-low/neg) samples (P<0.01). Genes hypermethylated in FB-high samples included well-known type C genes in colorectal cancer, such as MINT2 and 31, P16 and NEUROG1. Multivariate analysis demonstrated that the FB high status held an increased likelihood for methylation high as an independent factor (odds ratio: 16.18, 95% confidence interval: 1.94-135.2, P=0.01). Genome-wide methylation analysis demonstrated a unique methylome signature of FB-high cases irrespective of promoter, outside promoter, CpG and non-CpG sites. Group of promoter CpG sites that were exclusively hypermethylated in FB-high cases significantly codified the genes related to the catalytic activity (P=0.039). CONCLUSION Our findings suggest that Fusobacterium accelerates DNA methylation in specific groups of genes in the inflammatory colonic mucosa in UC.

Demonstration of potential link between helicobacter pylori related promoter CpG island methylation and telomere shortening in human gastric mucosa

Background Telomere length shortening in Helicobacter pylori (H. pylori) infected gastric mucosa constitutes the earliest steps toward neoplastic transformation. In addition to this genotoxic changes, epigenetic changes such as promoter CpG island (PCGI) methylation are frequently occurred in H. pylori infected gastric mucosa. The aim of this study was to investigate a potential link between H. pylori related PCGI methylation and telomere length shortening in the human gastric mucosa. Methods Telomere length was measured in non-neoplastic gastric mucosa from 106 cancer-free subjects. To identify H. pylori related PCGI methylation, bisulfite pyrosequencing was used to quantify the methylation of 49 PCGIs from 47 genes and LINE1 repetitive element Results We identified five PCGIs (IGF2, SLC16A12, SOX11, P2RX7 and MYOD1), which the methylation is closely associated with H. pylori infection. Hypermethylation of all these PCGIs was associated with development of pathological state from normal to mild, active, and atrophic gastritis (P<0.001) and lower pepsinogen I/II ratio (P<0.05), an indicator for gastric mucosal atrophy. Telomere shortening was significantly associated with mean Z score methylation of five PCGIs (R=−0.39, P<0.0001) and four of these locus (IGF2: R=−0.35, P=0.0003, SLC16A12: R=−0.35, P=0.0002, P2RX7: R=−0.29, P=0.003, and MYOD1: R=−0.33, P=0.0005). Multivariate analysis revealed that telomere shortening held an increased risk for hypermethylation (odds ratio: 1.71, 95% confidence interval: 1.11-2.63, P=0.016). Conclusion Potential link between H. pylori related PCGI methylation and telomere shortening emphasize the importance of genotoxic-epigenetic interaction in the pathological state of H. pylori infected gastric mucosa.

Evaluations of primary lesions by endoscopy clearly distinguishes prognosis in patients with gastric cancer who receive chemotherapy

Background Chemotherapy may improve outcomes in gastric cancer (GC), especially for the patients with advanced stage. To explore useful predictive factor for GC performing chemotherapy, we compared the tumor responses assessed using computed tomography (CT) with endoscopy based criteria. Methods 192 GC patients performing chemotherapy were retrospectively studied. CT based response assessment was performed after 2 courses of treatment. Endoscopic evaluation according to The Japanese classification of gastric carcinoma was also performed at same period. Data were correlated with overall survival (OS) and progression-free survival (PFS). Results Majority of the cases (n = 178, 93%) received S-1 based chemotherapy as the first line treatment. 55 (29%) and 91 (47%) cases were considered to be CT and endoscopic responders. Endoscopic responder was more clearly associated with better OS and PFS compared to CT based responder by the log-rank test (P<0.0001 vs. 0.01 and P<0.0001 vs. 0.008, respectively). The association was more striking among patients performing neoadjuvant chemotherapy (P<0.0001 vs. 0.15 and P<0.0001 vs. 0.1, respectively). Multivariate survival analysis using Coxs regression model revealed that endoscopic non-responder was the independent predictive factor, being more strongly associated with worse OS when compared to CT non-responder (hazard ratio: 4.60 vs. 1.77, 95% confidence interval: 2.83–7.49 vs.1.08–2.89, P<0.0001 vs. 0.02). More advanced T, N stage and cases who had peritoneal dissemination were significantly associated with endoscopic non-responder (all P values <0.01). Conclusion Endoscopy based evaluation of primary lesions are clearly associated with prognosis in patients with GC who perform chemotherapy.

Usefulness of confocal laser endomicroscopy to diagnose ulcerative colitis‐associated neoplasia

Chromoendoscopy, narrow‐band imaging (NBI), and confocal laser endomicroscopy (CLE) have been introduced in ulcerative colitis (UC)‐associated neoplasia surveillance. We aimed to determine the ability of CLE to differentiate among UC‐associated neoplasia (differentiated type or undifferentiated type), sporadic adenoma, and circumscribed regenerative lesions. Of 665 patients with UC, we carried out probe‐based CLE (pCLE) on 12 patients with suspected UC‐associated neoplasia in addition to magnifying chromoendoscopy with crystal violet and NBI. We compared pCLE findings with pathological diagnoses. pCLE could differentiate UC‐associated differentiated cancer from other pathologies such as solitary adenoma and non‐neoplastic circumscribed regenerative lesions on the basis of back‐to‐back orientation of crypts (P = 0.048), and UC‐associated undifferentiated cancer from other pathologies on the basis of dark trabecular architecture (P = 0.015). Sensitivity, specificity, and accuracy of combination of back‐to‐back orientation of crypts and dark trabecular architecture for carcinoma or dysplasia were 100%, 83%, and 92%, respectively. In vivo microscopic observation with pCLE was helpful to evaluate the suspected UC‐associated neoplasia.

Distinct Clinic-Pathological Features of Early Differentiated-Type Gastric Cancers after Helicobacter pylori Eradication.

Background. Gastric cancer is discovered even after successful eradication of H. pylori. We investigated clinic pathological features of early gastric cancers after H. pylori eradication. Methods. 51 early gastric cancers (EGCs) from 44 patients diagnosed after successful H. pylori eradication were included as eradication group. The clinic-pathological features were compared with that of 131 EGCs from 120 patients who did not have a history of H. pylori eradication (control group). Results. Compared with control group, clinic-pathological features of eradication group were characterized as depressed (p < 0.0001), reddish (p = 0.0001), and smaller (p = 0.0095) lesions, which was also confirmed in the comparison of six metachronous lesions diagnosed after initial ESD and subsequent successful H. pylori eradication. Prevalence of both SM2 (submucosal invasion greater than 500 μm) and unexpected SM2 cases tended to be higher in eradication group (p = 0.077, 0.0867, resp.). Prevalence of inconclusive diagnosis of gastric cancer during pretreatment biopsy was also higher in the same group (26.0% versus 1.6%, p < 0.0001). Conclusions. Informative clinic pathological features of EGC after H. pylori eradication are depressed, reddish appearances, which should be treated as a caution because histological diagnosis of cancerous tissue is sometimes difficult by endoscopic biopsy.

In vivo diagnosis of early‐stage gastric cancer found after Helicobacter pylori eradication using probe‐based confocal laser endomicroscopy

Early‐stage gastric cancer (EGC) found after Helicobacter pylori (Hp) eradication often displays non‐tumorous regenerative epithelium and/or maturated tumorous epithelium overlying the cancerous tissue, which may confuse endoscopic and histological diagnosis. Probe‐based confocal laser endomicroscopy (pCLE) enables in vivo real‐time optical biopsy. We compared the diagnostic yields for these EGC cases using conventional white light endoscopy (WL), magnifying endoscopy with narrow‐band imaging (ME‐NBI), pCLE, and endoscopic biopsy; we also compared the accuracy of the horizontal extent diagnosis between ME‐NBI and pCLE.

In vivo characterization of abnormalities in small-bowel diseases using probe-based confocal laser endomicroscopy

Background and study aims  Probe-based confocal laser endomicroscopy (pCLE) enables real-time optical biopsy. Little is known about pCLE imaging deep inside the small bowel, therefore the aim of this study was to determine its usefulness. Patients and methods  Between April 2014 and January 2016, we performed 38 pCLE examinations during double-balloon enteroscopy with intravenous fluorescein in 37 patients with: tumors (n = 10), vascular disorders (n = 6), inflammatory diseases and drug injuries (n = 13), other disorders (n = 4), and normal findings (n = 4). We measured the calibers of capillary and lymphatic vessels at 15 different sites and compared the calibers between pCLE images and histopathology. We also compared pCLE findings with pathologic diagnosis. Results  The inner diameters of capillary vessels beneath the epithelium and in the middle of villi were 16.2 ± 3.0 µm and 14.5 ± 3.1 µm, respectively, in the pCLE images, but these were not consistent with formalin-fixed paraffin-embedded histology. In tumors, larger capillary vessels were observed in aggressive malignant lymphoma and metastasis, and a “soccer ball-like pattern” constituting homogenous dark cells packed with polygonal, narrower capillary vessels was characteristic in pCLE images of a malignant lymphoma follicle. Hereditary hemorrhagic telangiectasia and angiodysplasia contained anastomosis of capillary vessels of different calibers. In IgA vasculitis, segmental capillary strictures were observed. Intestinal lymphangiectasia with protein-losing enteropathy contained a reticular network of lymphatic vessels and dilated lymphatic ducts accompanied by numerous cell gaps. pCLE findings corresponded to pathologic diagnosis in 32 examinations (91 %). Conclusions  pCLE is useful for in vivo analysis of abnormalities of the capillary and lymphatic vessels and epithelium, and for diagnosis in various small-bowel diseases.

A Possible Link between Gastric Mucosal Atrophy and Gastric Cancer after Helicobacter pylori Eradication

Background The effect of H. pylori eradication in gastric cancer prevention can be attributed to the improvement of atrophic gastritis, which is a known risk of gastric cancer. However, gastric cancer has also been diagnosed after long-term H. pylori eradication. This study aimed to clarify the association between gastric atrophy and gastric cancer after H. pylori eradication, including its clinicopathological features. Methods A total of 55 consecutive patients with 64 early gastric cancers (EGCs) diagnosed after H. pylori eradication were enrolled. The degree of endoscopic atrophy and the histological degrees of mononuclear cell infiltration, atrophy, and metaplasia in the corpus and adjacent mucosa of the EGCs were determined and scored. Results The majority of EGCs (63/64) were located within the endoscopically assessed atrophic mucosa or along the atrophic border. The adjacent mucosa of the EGCs presented significantly higher degrees of all histological parameters than in the corpus (mononuclear cell infiltration, 0.86+/-0.09 vs. 0.51+/-0.11, P = 0.016; atrophy, 1.77+/-0.13 vs. 0.65+/-0.14, P<0.0001; metaplasia, 1.68+/-0.13 vs. 0.48+/-0.1, P<0.0001). The degree of endoscopic atrophy improved in the patients with longer post-H. pylori eradication periods; however, this trend was not observed for the histological parameters, and high degrees of atrophy and metaplasia were observed in the adjacent mucosa of the EGCs compared with the corpus during all periods (all P<0.05). The histological degrees of atrophy and metaplasia in the adjacent mucosa were particularly higher in the patients who underwent eradication due to gastric ulcers. Conclusions Severe gastric atrophy remained in the adjacent mucosa of the EGCs after H. pylori eradication, which may be linked to gastric carcinogenesis.

Unusual growth of an Epstein-Barr virus-associated differentiated early-stage gastric carcinoma: A case report

Epstein-Barr virus (EBV) infection is involved in a subset of gastric carcinoma (GC) cases, and is associated with distinct clinicopathological features. The present study reports a unique case of EBV-associated early-stage GC compared with the other cases experienced in our hospital. A 72-year-old male receiving treatment for cerebral infarction underwent an esophagogastroduodenoscopy (EGD) for suspicion of gastrointestinal bleeding. EGD revealed a type 0-I protruding lesion in the lesser curvature of the upper gastric corpus. Biopsy indicated well-differentiated adenocarcinoma. As the tumor diameter was >3 cm and the thickness of the tumor suggested submucosal invasion, laparoscopic gastrectomy was performed. Histological assessment revealed polypoid growth of an intramucosal, differentiated, tubular or papillary adenocarcinoma, with dense infiltration of lymphocytes. The carcinoma crypts were found to be EBV-positive on in situ hybridization. A review of the clinicopathological features of 25 EBV-associated GCs from 20 patients treated in our hospital between 2005 and 2014 was performed. All of these tumors, except that in the current case, appeared as shallow, depressed or ulcerative lesions. Thus, the current case appears to represent an unusual growth of EBV-positive GC.