CA19-9 in Combination with Methylated HOXA1 and SST Is Useful to Diagnose Stage I Pancreatic Cancer.

INTRODUCTION: We previously developed a novel methylation assay, the combined restriction digital PCR (CORD) assay, consisting of treatment of DNA with methylation-sensitive restriction enzymes and droplet digital PCR. METHODS: In this study, we assessed the diagnostic performance of serum methylated Homeobox A1 (mHOXA1) and methylated somatostatin (mSST) using the CORD assay in combination with CA19-9 for pancreatic cancer using serum samples from 82 healthy individuals, 13 patients with benign pancreatic disease, 3 patients with branched-duct intraductal papillary mucinous neoplasm, and 91 patients with pancreatic cancer. RESULTS: For the single marker tests, sensitivity for all stages of pancreatic cancer, stage I cancer, and specificity were, respectively, 71.4%, 50.0%, and 94.9% for CA19-9; 51.6%, 68.8%, and 90.8% for mHOXA1; and 50.1%, 68.8%, and 94.9% for mSST. Those for the combined marker tests were, respectively, 86.8%, 81.3%, and 85.7% for combined mHOXA1 and CA19-9; 86.8%, 87.5%, and 89.8% for combined mSST and CA19-9; and 89.0%, 87.5%, and 85.7% for all three markers combined. CONCLUSION: The combination of mHOXA1 and mSST with CA19-9 appears to be useful to detect pancreatic cancer even at an early stage.

Comparison of two primer-probe sets of Fusobacterium nucleatum using droplet digital polymerase chain reaction for the detection of colorectal neoplasia from faecal samples.

BACKGROUND: Although faecal DNA testing of Fusobacterium nucleatum (Fn) is expected to be useful for colorectal neoplasia detection, there is no standardized quantification method of Fn. We performed this study to establish a possible standardized method. METHODS: In this study, 322 participants including 71 subjects without colorectal neoplasia (control group), 31 patients with non-advanced colorectal adenoma, 93 patients with advanced colorectal adenoma, and 127 patients with colorectal cancer were enrolled. Faecal Fn were quantified by droplet digital PCR (ddPCR) using two PCR primer-probe sets reported previously that are tentatively named Fn1 and Fn2. Fn1 has been used in ddPCR by us and Fn2 has been widely used in quantitative real-time PCR. RESULTS: The Fn copy number using Fn1 was five times higher than that using Fn2, with a linear relationship shown between them. Receiver operating characteristic curve analysis showed the area under the curve (AUC) to be almost the same between Fn1 and Fn2 in discriminating between the control group and the colorectal cancer group (AUC = 0.81 and 0.81, respectively), and between the control/non-advanced colorectal adenoma group and the advanced colorectal adenoma/colorectal cancer group (AUC = 0.74 and 0.74, respectively). CONCLUSIONS: As the diagnostic performance was quite similar between Fn1 and Fn2, ddPCR-based Fn testing using Fn1 and Fn2 could be a possible standardized method for a colorectal neoplasia screening test, considering that Fn levels quantified by Fn1 are about five times higher than those by Fn2.

Combination of CA19-9 and Blood Free-Circulating Methylated RUNX3 May Be Useful to Diagnose Stage I Pancreatic Cancer.

BACKGROUND: Although serum carbohydrate antigen 19-9 (CA19-9) is widely used as a useful biomarker of pancreatic cancer for monitoring the response to therapy, it is not recommended for screening of early pancreatic cancer because of its limited sensitivity for small tumors. Thus, it is critical to discover novel serum biomarkers to complement CA19-9 in order to improve sensitivity. Although methylated runt-related transcription factor 3 (RUNX3) is a biomarker of pancreatic cancer, its detection by conventional bisulfite-based methylation assays from a small serum sample amount is very difficult. Therefore, we developed a new methylation assay, the combined restriction digital PCR (CORD) assay, that enables counting of even one copy of a methylated gene in a small DNA sample amount without DNA bisulfite treatment. OBJECTIVES: We evaluated the sensitivity and specificity of serum DNA testing of methylated RUNX3 by the CORD assay in combination with and without CA19-9 for the detection of pancreatic cancer in 55 patients with pancreatic cancer, 12 patients with benign pancreatic disease, and 80 healthy individuals. RESULTS: The CORD assay of methylated RUNX3 had a sensitivity of 50.9% (28/55) and specificity of 93.5% (86/92). Combination of the CORD assay of methylated RUNX3 and CA19-9 resulted in a sensitivity of 85.5% (47/55) and specificity of 93.5% (86/92) for all stages of pancreatic cancer and a sensitivity of 77.8% (7/9) for stage I pancreatic cancer. CONCLUSIONS: ombination of the CORD assay and CA19-9 may provide an alternative screening strategy for detecting early-stage pancreatic cancer.

Fecal DNA Testing of TWIST1 Methylation Identifies Patients With Advanced Colorectal Adenoma Missed by Fecal Immunochemical Test for Hemoglobin.

INTRODUCTION: We have reported previously that fecal DNA testing of TWIST1 methylation in combination with the fecal immunochemical test for hemoglobin (FIT) (combination test) is useful for colorectal neoplasia screening. In this study, using larger sample sizes, we studied the clinical performance of the combination test for the detection of colorectal neoplasia and, especially, advanced colorectal adenoma. METHODS: We performed a prospective study in which FIT, fecal DNA testing of TWIST1 methylation, and colonoscopy were performed on 372 patients with colorectal neoplasia and 71 subjects without colorectal neoplasia. We assessed the individual clinical performance of each of FIT and fecal DNA testing of TWIST1 methylation and of the combination test for the detection of colorectal neoplasia including advanced adenoma based on morphologic subtypes. RESULTS: The FIT alone had a sensitivity of 7.5% (3/40) for nonadvanced adenoma, 32.3% (41/127) for advanced adenoma, and 93.7% (192/205) for colorectal cancer and a specificity of 87.3% (62/71). The combination test had a sensitivity of 35.0% (14/40) for nonadvanced adenoma, 68.5% (87/127) for advanced adenoma, and 95.6% (196/205) for colorectal cancer and a specificity of 80.3% (57/71). For morphological subtypes of advanced adenoma, the sensitivity of FIT was only 28.2% (20/71) for polypoid type and 16.1% (5/31) for nonpolypoid type, whereas the combination test increased the sensitivities to 64.8% (46/71) and 71.0% (22/31), respectively. DISCUSSION: The combination of the fecal DNA test with FIT seemed to be useful to detect colorectal neoplasia and, especially, advanced adenoma of the nonpolypoid type.

Blood Free-Circulating DNA Testing of Methylated RUNX3 Is Useful for Diagnosing Early Gastric Cancer.

The main modalities for gastric cancer screening are limited to upper gastrointestinal endoscopy and contrast radiography. The former is invasive, and the latter has high false-negative rates. Thus, alternative diagnostic strategies are required. One solution may be a liquid biopsy. Methylated RUNX3 is a well-known biomarker of gastric cancer but it is very difficult to detect with conventional bisulfite-based methylation assays when only a small amount of serum is available. We developed the combined restriction digital PCR (CORD) assay, a new methylation assay allowing for the counting of as little as one copy of a methylated gene in a small sample of DNA without necessitating DNA bisulfite treatment. We evaluated the sensitivity and specificity of the serum DNA testing of methylated RUNX3 by the CORD assay for the detection of early gastric cancer using 50 patients with early gastric cancer and 61 control individuals. The CORD assay had a sensitivity of 50.0% and a specificity of 80.3% for early gastric cancer. Methylated RUNX3 copies were significantly associated with tumor size, massive submucosal invasion, and lymph-vascular invasion. After the treatment, the median number of methylated RUNX3 copies was significantly decreased. The CORD assay may provide an alternative screening strategy to detect even early-stage gastric cancer.

Novel Liquid Biopsy Test Based on a Sensitive Methylated SEPT9 Assay for Diagnosing Hepatocellular Carcinoma.

Liquid biopsies are not used in practice for hepatocellular carcinoma (HCC). Epi proColon is the first commercial blood-based test for colorectal cancer screening based on methylated DNA testing of the septin 9 gene (SEPT9). However, Epi proColon has some disadvantages, including the requirement of a large amount of blood and lack of quantitative performance. Therefore, we previously developed a novel liquid biopsy test that can quantitatively detect even a single copy of methylated SEPT9 in a small amount of DNA. In the current study, we evaluated the application potential of this assay for diagnosing HCC. Study subjects included 80 healthy volunteers, 45 patients with chronic liver disease (CLD) without HCC, and 136 patients with HCC (stage 0, 12; stage A, 50; stage B, 31; stage C, 41; and stage D, 2), according to the Barcelona Clinic Liver Cancer staging system. For the assay, DNA was treated with methylation-sensitive restriction enzymes in two steps, followed by multiplex droplet digital polymerase chain reaction. The median copy number of methylated SEPT9 was 0.0, 2.0, and 6.4 in the healthy control, CLD, and HCC groups, respectively, with significant differences among the groups (HCC vs. healthy control, P < 0.001; HCC vs. CLD, P = 0.002; CLD vs. healthy control, P = 0.008). Assay sensitivity and specificity were 63.2% and 90.0%, respectively (cutoff value, 4.6 copies), in detecting HCC when compared with healthy subjects. The positive rate of methylated SEPT9 increased with HCC progression (stage 0, 41.7%; stage A, 58.0%; stage B, 61.3%; stage C, 75.6%; and stage D, 100%). Conclusion: We developed a sensitive methylated SEPT9 assay that might serve as a liquid biopsy test for diagnosing HCC.

Blood free-circulating DNA testing by highly sensitive methylation assay to diagnose colorectal neoplasias.

Although methylated TWIST1 is a biomarker of colorectal neoplasia, its detection from serum samples is very difficult by conventional bisulfite-based methylation assays. Therefore, we have developed a new methylation assay that enables counting of even one copy of a methylated gene in a small DNA sample amount without DNA bisulfite treatment. We performed this study to evaluate the sensitivity and specificity of serum DNA testing by the new methylation assay in combination with and without the fecal immunochemical test for hemoglobin for the detection of colorectal neoplasia. This study comprised 113 patients with colorectal neoplasia and 25 control individuals. For the new methylation assay, DNA was treated in two stages with methylation-sensitive restriction enzymes, followed by measurement of copy numbers of hTERT and methylated TWIST1 by multiplex droplet digital PCR. The fecal immunochemical test had a sensitivity of 8.0% for non-advanced adenoma, 24.3% for advanced adenoma, and 44.4% for colorectal cancer, and a specificity of 88.0%. The new assay had a sensitivity of 36.0% for non-advanced adenoma, 30.0% for advanced adenoma, and 44.4% for colorectal cancer, and a specificity of 92.0%. Combination of the both tests increased the sensitivity to 40.0%, 45.7%, and 72.2% for the detection of non-advanced adenoma, advanced adenoma, and colorectal cancer, respectively, and resulted in a specificity of 84.0%. Combination of both tests may provide an alternative screening strategy for colorectal neoplasia including potentially precancerous lesions and colorectal cancer.

Highly sensitive faecal DNA testing of TWIST1 methylation in combination with faecal immunochemical test for haemoglobin is a promising marker for detection of colorectal neoplasia.

Background As TWIST1 methylation is specific to colorectal neoplasia, detection of TWIST1 methylation from faeces samples might be useful for colorectal neoplasia screening. However, because the content of human DNA in faeces is very small, it is very difficult to detect TWIST1 methylation by conventional bisulphite-based methylation assays. Therefore, we developed a new methylation assay without bisulphite treatment, the combined restriction digital PCR assay, and evaluated its sensitivity and specificity in combination with and without the faecal immunochemical test for haemoglobin for colorectal neoplasia detection from faeces samples. Methods For the combined restriction digital PCR assay, DNA was treated with three methylation-sensitive restriction enzymes and an exonuclease, followed by measurement of TWIST1 methylation level by droplet digital PCR. Faecal DNA testing and faecal immunochemical test for haemoglobin were performed on 109 patients with colorectal neoplasia and 10 control individuals. Results Basic performance testing showed that the combined restriction digital PCR assay enabled detection of 0.14% of the TWIST1 methylation level for the lymphocyte DNA. The combined restriction digital PCR assay from faeces samples had a sensitivity of 22.2% (95% confidence interval, 2.8-60.0%) for non-advanced adenoma, 47.1% (95% confidence interval, 23.0-72.2%) for advanced adenoma, and 33.7% (95% confidence interval, 23.7-45.0%) for colorectal cancer, and a specificity of 100.0%. Combination of faecal immunochemical test for haemoglobin and faecal combined restriction digital PCR assay increased sensitivity to 82.4% (95% confidence interval, 56.6-96.2%) for the detection of advanced adenoma. Conclusions We developed the combined restriction digital PCR assay, a possible highly sensitive methylation assay. Combination of faecal combined restriction digital PCR assay with faecal immunochemical test for haemoglobin may provide an alternative screening strategy for colorectal neoplasia, especially for potentially precancerous lesions.

Highly sensitive stool DNA testing of Fusobacterium nucleatum as a marker for detection of colorectal tumours in a Japanese population.

Background Accumulating evidence shows an over-abundance of Fusobacterium nucleatum in colorectal tumour tissues. Although stool DNA testing of Fusobacterium nucleatum might be a potential marker for the detection of colorectal tumours, the difficulty in detecting Fusobacterium nucleatum in stool by conventional methods prevented further explorations. Therefore, we developed a droplet digital polymerase chain reaction (PCR) assay for detecting Fusobacterium nucleatum in stool and investigated its clinical utility in the management of colorectal tumours in a Japanese population. Methods Feces were collected from 60 healthy subjects (control group) and from 11 patients with colorectal non-advanced adenomas (non-advanced adenoma group), 19 patients with colorectal advanced adenoma/carcinoma in situ (advanced adenoma/carcinoma in situ (CIS) group) and 158 patients with colorectal cancer of stages I to IV (colorectal cancer group). Absolute copy numbers of Fusobacterium nucleatum were measured by droplet digital PCR. Results The median copy number of Fusobacterium nucleatum was 17.5 in the control group, 311 in the non-advanced adenoma group, 122 in the advanced adenoma/CIS group, and 317 in the colorectal cancer group. In comparison with that in the control group, the Fusobacterium nucleatum level was significantly higher in the non-advanced adenoma group, the advanced adenoma/CIS group and the colorectal cancer group. Conclusions This study illustrates the potential of stool DNA testing of Fusobacterium nucleatum by droplet digital PCR to detect individuals with colorectal tumours in a Japanese population.

Image quality of a novel light-emitting diode (LED)-illuminated colonoscope.

BACKGROUND AND STUDY AIMS: Light-emitting diodes (LEDs) are used widely for their high luminous efficiency and durability. We developed a novel prototype high definition endoscope with white LEDs and evaluated the image quality it produced against a commercial endoscope with conventional light source. PATIENTS AND METHODS: The specifications of both colonoscopes were identical, except for the LED light source at the tip of the prototype. We examined 20 patients with rectal or sigmoid colon lesions and the image quality was evaluated in 40 images (one image from the LED colonoscope and one from the conventional colonoscope for each lesion) by three endoscopists. We additionally evaluated the 17 videos recorded with the LED colonoscope that were available. Image quality, mucosal and vascular color, and luminous distribution and intensity were scored on a 5-point scale. RESULTS: The mean score for vascular color given by one evaluator was significantly higher using the LED colonoscope than using the conventional colonoscope. The mean scores for mucosal color and luminous intensity from another evaluator were significantly lower with the LED colonoscope than with the conventional colonoscope. There were no significant differences in the luminous distribution scores for any of the evaluators. The image quality of the videos was evaluated as being similar with both colonoscopes. CONCLUSIONS: Image quality from the LED and conventional colonoscopes were similar, although the luminous intensity of the LEDs is inferior to that of the conventional light source at the present time.

Long-fragment DNA as a potential marker for stool-based detection of colorectal cancer.

Neoplastic cells that are exfoliated from the colorectal epithelium exhibit dysfunctional apoptotic mechanisms, and thus it is possible to identify high-molecular-weight DNA fragments (long DNA) in feces. In the present study, the sensitivity and specificity of fecal-based long DNA assays were evaluated for the detection of colorectal cancer (CRC). Feces were collected from 54 healthy volunteers and 130 patients with CRC prior to surgical treatment. The presence of long DNA of the adenomatosis polyposis coli, Kirsten rat sarcoma viral oncogene homolog (KRAS), B-raf proto-oncogene, serine/threonine kinase and p53 genes was assessed by polymerase chain reaction followed by electrophoresis. The identification of long DNA in feces was found to exhibit a sensitivity of 56.2% and specificity of 96.3% for CRC detection. In addition, long DNA was identified in the feces of 58/90 (64.4%) patients with distal CRC and 15/40 (37.5%) patients with proximal CRC. This study indicates the potential of the fecal long DNA assay as a non-invasive and easily performed method for the detection of individuals with CRC.

Preoperative evaluation of early colorectal cancer using an ultrasound mini probe.

BACKGROUND/AIMS: We investigated whether endoscopic ultrasonography (EUS) can assess the depth of invasion of early colorectal cancer exhibiting the V pit pattern on magnifying endoscopy with submucosal invasion of 1,000 µm or deeper. METHODOLOGY: Among 38 colorectal tumors exhibiting the V pit pattern on magnifying endoscopy, the findings on EUS with a mini-probe (15 MHz) were compared with histopathological findings. The diagnostic accuracy, sensitivity and specificity of EUS were examined separately in tumors exhibiting the Vi or V5 pit pattern. RESULTS: Diagnostic accuracy of EUS for cancers exhibiting the Vi pit pattern on magnifying endoscopy with submucosal invasion of 1,000 µm or deeper was 9/15 (60%). Sensitivity was 90%, specificity 14.3%, positive predictive value 31.7% and negative predictive value 76.3%. Diagnostic accuracy of EUS for cancers exhibiting the VN pit pattern on magnify-ing endoscopy with submucosal invasion of 1,000 µm or deeper was 13/18 (72%). The sensitivity and specificity of EUS were 100% and 37.5%, respectively. CONCLUSIONS: EUS tended to diagnose the invasion depth of cancer with submucosal invasion exhibiting the V pit pattern as deeper than it actually was. EUS accurately diagnosed early colorectal cancer with shallow invasion exhibiting the VN pit pattern and surgery was avoided.

A novel colonoscope with high color-rendering white light-emitting diodes.

BACKGROUND: Light-emitting diodes (LEDs) are being used for a variety of new uses because of their cost-effectiveness and durability. We therefore considered using white LEDs as a light source for GI endoscopes to simplify the endoscopic system. OBJECTIVE: To assess the feasibility and safety of an LED-illuminated colonoscope. DESIGN: Pilot study of the LED colonoscope in healthy human volunteers and patients with colorectal lesions. SETTING: Yamaguchi University Hospital, Ube, Japan. INTERVENTIONS: We performed a total colonoscopy for 2 healthy volunteers and a sigmoidoscopy for 15 patients with colorectal lesions by using both LED and conventional colonoscopes. We assessed the feasibility and safety of the LED colonoscope by using a 5-grade scale. The 30 images of 15 colorectal lesions obtained by using each endoscope were evaluated in a blind, randomized order by 2 endoscopists. MAIN OUTCOME MEASUREMENTS: The mean scores of the LED colonoscope for the operability, ease of manipulation, image quality, and safety. RESULTS: We manufactured a prototype LED colonoscope with white LEDs on its tip. The LED colonoscope did not require an external light source or light-guide fiber bundle. The operability and ease of manipulation of the LED colonoscope were evaluated as similar to those of the conventional colonoscope. The color of the colonic mucosa and the vascular pattern were clearly visualized in the volunteers. For the 15 colorectal lesions, the mean score for image quality was not significantly different between the colonoscopes. The study was performed safely without any incident. LIMITATIONS: Single-center, small number of patients. CONCLUSIONS: The use of an LED colonoscope is feasible, and LED illumination may simplify the endoscope system.

Usefulness of computed tomography enteroclysis in the intestinal tract.

PURPOSE: The aim of this study was to evaluate computed tomography enteroclysis (CTE) of the small intestinal tract. MATERIALS AND METHODS: A total of 36 patients underwent CTE for further examination of small intestinal disease. RESULTS: The indications were obscure gastrointestinal bleeding (OGIB) (n = 16), suspected Crohn's disease (n = 7), suspected and diagnosed by exclusion a small intestinal tumor (n = 5), and others (n = 8). Regarding OGIB, positive findings were observed in nine patients: angiodysplasia (n = 2), suspected ileac tumor (n = 2), colon cancer (n = 1), colon diverticulosis and diverticulitis (n = 2), Crohn's disease (n = 1), and enteritis (n = 1). As for Crohn's disease, hyperplasia of the small intestinal wall was shown in six patents. Positive findings of a small intestinal tumor were observed in two patients. In the "others" category, colon diverticulitis was found in three patients and isolated dissection of the superior mesenteric artery in one of four patients with abdominal pain. Primary carcinoid was identified in the pancreas in one patient, with liver carcinoid metastasis in the remaining four patients. CONCLUSION: CT enteroclysis is a noninvasive method and useful approach in the diagnosis of small intestinal diseases.

TWIST1 hypermethylation is observed frequently in colorectal tumors and its overexpression is associated with unfavorable outcomes in patients with colorectal cancer.

Although growing evidence demonstrates that TWIST1 is an interesting tumor biomarker, little is known about the clinical significance of TWIST1 expression and TWIST1 methylation in human primary colorectal cancer. In this study, we examined the association of TWIST1 expression and TWIST1 methylation with clinicopathologic features in human primary colorectal tumors. Primary colorectal cancer (CRC) specimens from 319 patients, corresponding normal colorectal nontumorous mucosa from 251 patients with cancer, and colorectal adenomas from 189 patients were used. Methylation and expression levels of TWIST1 were compared with clinicopathologic features. The TWIST1 methylation level was higher in colorectal adenoma and cancer than in normal colorectal mucosa. Elevated TWIST1 mRNA expression in normal colorectal mucosa in patients with CRC as well as in primary CRC specimens was associated with unfavorable outcomes. There was no correlation between TWIST1 methylation and TWIST1 expression. Our results suggest that TWIST1 methylation may be a useful biomarker for screening colorectal tumors. In addition, TWIST1 mRNA expression is a possible molecular marker for predicting the outcome in patients with CRC. Confirmatory studies using independent data sets are needed to confirm our findings.

Computed tomography enteroclysis for recurrent severe gastrointestinal bleeding in a patient with vascular malformation of the small bowel.

This is a case study of a 66-year-old woman who had a vascular malformation of the small bowel that was visualized on computed tomography enteroclysis (CTE). She presented with repeated tarry stool and severe anemia. Although the source of bleeding was not identified on upper and lower gastrointestinal endoscopy, active bleeding was revealed by capsule endoscopy in the deep jejunum. The cause of bleeding was not found on capsule endoscopy. CTE was requested as double-balloon endoscopy would have been difficult because of strong adhesion of the small intestine. A continual subtle vascular malformation of the jejunum, starting from the third jejunal branch end, was demonstrated on CTE with dynamic contrast enhancement. Because this vascular malformation was considered the cause of small intestinal bleeding, selective arterial coil embolization was performed. After embolization, the repeated tarry stool disappeared and the severe anemia dramatically improved. CTE may be a safe and useful method for determining the cause of small intestinal bleeding and for subsequent therapy.

Does chromoendoscopy improve the colonoscopic adenoma detection rate?

BACKGROUND/AIMS: It is not clear whether chromoendoscopy with indigo carmine improves the rate of detection of colorectal polyps compared to: detection via standard colonoscopy. The aim of our study was to determine whether chromoendoscopy with indigo carmine significantly improves the detection of adenomas in the distal colon and rectum. METHODOLOGY: Using back-to-back sigmoidoscopies in each study patient, we prospectively evaluated whether chromoendoscopy with indigo carmine picked up more adenomatous polyps than standard colonoscopy. In all patients, standard high-resolution complete colonoscopy without indigo carmine was performed at the first examination. The second examination was restricted to colonoscopy distal to the splenic flexure of the colon. For the second examination, patients were randomized to chromoendoscopy with indigo carmine or colonoscopy without indigo carmine application. The second examination's detection rate was compared between the two groups. RESULTS: In the 60 patients in the chromoendoscopy with indigo carmine group, 38 adenomas were found in the first examination and 14 adenomas in the second examination. In the 70 patients in the standard colonoscopy group, 66 adenomas were found in the first examination and 32 adenomas in the second examination. The detection rates in the two groups were 26.9% and 32.7%, re spectively, which were not significantly different (p = 0.47). CONCLUSION: Chromoendoscopy did not detect more adenomatous polyps in comparison to standard colonoscopy.

Hypermethylation status of APC inversely correlates with the presence of submucosal invasion in laterally spreading colorectal tumors.

Little is known about epigenetic alterations in laterally spreading colorectal tumors (LSTs). The goal of the present study was to elucidate the epigenetic background of LSTs and compare the methylation status of DNA CpG islands (CGIs) with clinicopathologic features. Methylation of MINT1, MINT2, MINT31, p16, O(6)-methylguanine-DNA methyltransferase (MGMT), adenomatous polyposis coli (APC), and human MutL homologue 1 (hMLH1) in 42 LSTs was assessed by methylation-specific polymerase chain reaction (MSP) and compared with clinicopathologic parameters. The frequency of hypermethylation was 12.5% (4/32) for MINT1, 40.0% (16/40) for MINT2, 25.0% (10/40) for MINT31, 25.7% (9/35) for p16, 7.7% (3/39) for hMLH1, 26.5% (9/34) for MGMT, and 35.9% (14/39) for APC. APC methylation was inversely associated with submucosal invasion (P = 0.034), which was not found in any of 14 LST cases with APC methylation, whereas submucosal invasion was present in 8 of 25 (32.0%) cases without APC methylation. These data suggest that hypermethylation of APC could be a predictive marker for the absence of submucosal invasion of LSTs.

A reduced COX-2 expression and a reduced number of pericryptal myofibroblasts are associated with depressed adenoma of the colon.

The histogenesis of depressed adenoma of the colon has not been sufficiently investigated. Pericryptal myofibroblasts are stromal cells expressing smooth muscle actin, and are involved in the differentiation and multiplication of epithelial cells in the colonic epithelium. COX-2 has been reported to be involved in the development of colon adenoma. We studied the histogenesis of depressed adenoma of the colon by examining the relationship between the presence of pericryptal myofibroblasts and COX-2 expression. Twenty-one depressed adenomas of the colon that had been resected endoscopically between June 1998 and May 2003 (mild-moderate atypia; mean diameter, 6.7 mm) and 23 elevated adenomas that had been resected endoscopically in 2003 (mild-moderate atypia; mean diameter, 11.7 mm), were studied. We performed immunohistochemical staining using alpha-smooth muscle actin antibody to detect pericryptal myofibroblasts. We also performed immunohistochemical staining for Cox-2. Eighteen (78.3%) of the 23 elevated adenomas and six (28.6%) of the 21 depressed adenomas were positive for pericryptal myofibroblasts immunohistochemically, showing a significant difference (P<0.001). Seventeen elevated adenomas (73.9%) and eight depressed adenomas (38.1%) were positive for COX-2 expression (P=0.016). COX-2 expression was detected in the stroma, and the sites of COX-2 expression coincided with the sites of pericryptal myofibroblasts. The histogenesis of depressed adenomas differs from that of elevated adenomas. Our results suggest that a low number of pericryptal myofibroblasts and a low COX-2 expression are associated with depressed adenomas.

Relationship between molecular markers and endoscopic findings in laterally spreading tumors.

BACKGROUND AND AIM: Characteristic clinicopathological features of laterally spreading tumors (LSTs) have been reported by endoscopists; however, only a few studies have been conducted on the biological features. These studies were not fully associated with the endoscopic findings of LSTs. The aim of this study was to estimate the biological features of each type of endoscopic finding of LST using two molecular markers, matrix metalloproteinase-7 (MMP-7) and beta-catenin. METHODS: Expression of the molecular markers and the endoscopic findings were compared in 22 LSTs and 14 subpedunculated polyps. MMP-7 and beta-catenin were immunostained. Three types of representative endoscopic findings of LST were defined as segments in LSTs. They were 15 granular segments, seven large nodular segments, and seven flat segments that corresponded to the area composed of aggregates of similar size granules, large nodules of diameter more than 10 mm, and a flat surface with no granule, respectively. RESULTS: Expression of MMP-7 and coexpression of MMP-7 and beta-catenin were higher in large nodular segments than in granular segments (P < 0.0167). Among the three types of segments, flat segments showed the highest expression densities of beta-catenin accumulated in the nucleus. Large nodular segments and subpedunculated polyps showed similar expression patterns for the molecular markers. CONCLUSIONS: This study provides new and important information on the relationship between the molecular markers and endoscopic findings of LSTs.