rs12904 polymorphism in the 3'-untranslated region of ephrin A1 ligand and the risk of sporadic colorectal cancer in the Iranian population

Background: Colorectal cancer (CRC) is rated as the second cause of cancer death. Genetic determinants are considered as driving forces in the development of sporadic CRC. Single-nucleotide polymorphisms (SNPs), due to their abundance in the human genome with collectively huge effect on cellular signaling pathways, are attributed as the main genetic factor in disease susceptibility including cancers. MicroRNAs are contributing to posttranslational gene regulation. They exert their regulatory function by binding to their specific recognition sequences located at 3'-untranslated region (UTR) of mRNAs. In the present study, we have elucidated the role of rs12904, a naturally occurring SNP, in the recognition site of miR200c in the 3'UTR of ephrin A1 ligand gene, in the development of sporadic CRC in the Iranian population. Materials and Methods: A case–control study using 152 CRC patients and 160 noncancerous counterparts was conducted to determine the rs12904 genotypes using polymerase chain reaction–restriction fragment length polymorphism method. Results: The results revealed no significant association between the rs12904 and sporadic CRC (odds ratio = 0.97, 95% confidence interval = 0.70–1.34). The frequency of genotypes and also alleles of the mentioned polymorphism were not significantly different between case and control groups (P = 0.765 and P = 0.847, respectively). Conclusion: The results suggest that this polymorphism probably has not a crucial role in the Iranian CRC risk and is not an important potential risk factor in molecular diagnostics of mentioned disease among the Iranian population.

A Case of Colonic Mucinous Adenocarcinoma in 19-year-old Male Patient

Colorectal cancer is rare in teenagers, especially without known risk factors. Colon cancer in young age is more likely to be diagnosed at advanced-stage, to present unfavorable tumor histology such as mucinous carcinoma, and poor outcome. We report a case of sporadic mucinous adenocarcinoma of the colon in a 19-year-old male patient without any risk factors. He complained of severe left abdominal pain that developed 1 month ago. He had a distended abdomen with severe tenderness on the left lower quadrant. A distal descending colon mass causing mechanical obstruction was observed on abdominal computed tomography. Emergency colonoscopy showed a large, fungating mass obstructing the lumen at 40 cm from the anal verge. Biopsy of the colonic mass suggested a mucinous adenocarcinoma. After decompression by colonic stent, the patient was transferred to the general surgery department for left hemicolectomy. The lesion was confirmed to be a mucinous adenocarcinoma (7.0x4.5 cm). For hereditary nonpolyposis colorectal cancer evaluation, immunohistochemical staining for MLH1 and MSH2 was normal. Reverse transcription polymerase chain reaction analysis did not detect microinstability in any of the markers tested. The patient had no familial history of cancer. Mucinous adenocarcinoma has high frequencies of poor differentiation, advanced tumor stage, loss of mismatch repair gene expression, and increased MUC2 expression. A mucinous histology is considerably more frequent in children and adolescent than in adults. Adequate invasive study is also necessary for young age patients.

Clinicopathologic Features of Sporadic Colorectal Cancer with MLH1/MSH2 Loss of Expression - Reduced Likelihood of Metastases

PURPOSE: This study was designed to determine the frequency of MMR defective sporadic colorectal cancer (CRC) by using immunohistochemistry and to investigate the correlation between the MMR status and the metastatic potential. METHODS: The study included 249 patients with sporadic colorectal cancer who underwent surgical resection. The MMR status was determined by using an immunohistochemical analysis of MLH1 and MSH2 expression. RESULTS: Twenty seven (10.8%) carcinomas showed abnormal MMR protein expression (18 MLH1 negative and 9 MSH2 negative) and were classified as MMR defective tumors whereas 222 tumors demonstrated normal MLH1/MSH2 immunoreactivity (MMR intact tumor). MMR defective tumors developed at significantly higher frequencies in a proximal site (59.3% vs. 27.5%, P=0.001) and tended to be larger in size (6.3+/-2.4 cm vs. 5.1+/-2.1 cm, P=0.026). They showed significantly lower overall stage, N stage, and M stage at the time of diagnosis (P=0.002, P=0.014, P=0.010, respectively). In patients who had MMR defective tumors, lymphocytic infiltration (40.7% vs. 8.7%, P<0.001) and poor differentiation (22.2% vs. 11.7%, P=0.012) were more frequently observed. Less frequently MMR defective tumors displayed lymphatic invasion (40.7% vs. 67.1%, P=0.007) and infiltrative borders (22.2% vs. 51.8%, P=0.004). The MMR defect was strongly associated with a decreased likelihood of lymph node (odds ratio: 0.34, 95% CI: 0.13~0.95) and distant organ metastases at diagnosis (odds ratio: 0.09, 95% CI: 0.01~0.94), independent of the clinicopathologic features. CONCLUSIONS: mmunohistochemical analysis revealed that 10.8% of sporadic CRC cases showed no staining for MLH1 or MSH2. Lymphatic invasion and distant metastases were found at lower rates in these MMR defective tumors.

Efficacy of hMLH1/hMSH2 Immunohistochemical Staining as Representative Index for Microsatellite Instability Status in Sporadic Colorectal Cancer

PURPOSE: Sporadic colorectal cancer with micosatellite instability (MSI) is supposed to have a distinct molecular profile, distinct clinocopathologic feature, and a distinct prognosis. However, the test for MSI is still expensive, and a big machine is needed for routine screening. This study was performed to examine the clinicopathologic of characteristics of MSI sporadic colorectal cancer and the efficacy of immunohistochemical staining for hMLH1 and hMSH2. METHODS: Five hundred sixty nine colorectal adenocarinomas resected from September 2003 to August 2004 at Asan Medical Center were prospectively collected. FAP (familial adenomatous polyposis), HNPCC (hereditary non-polyposis colo-rectal cancer), and incomplete tests of immunohistochemical staining or MSI were excluded. The MSI status was determined by using PCR (polymerase chain reaction). A first round of immunohistochemical staining for hMLH1/hMSH2 was performed, and a second round was performed for cases showing a disparity between the two exams. The clinicopathologic variables regarding the MSI status were analyzed, and the sensitivity and the specificity of immunohistochemical staining were evaluated. RESULTS: Sporadic colorectal cancers with MSI-H were 8.4% (n=48) and were associated with age (< or = 60 years), colorectal cancer familial history, synchronous colorectal cancer, right side tumor location, and poorly differentiated or mucinous cell type. However, age, synchronous colorectal cancer, and right side tumor location were associated an the multivariate analysis. In the first round of immunohistochemical staining, no expression of hMLH1 and/or hMSH2 was obserred in 71 cases (12.5%), and the sensitivity and the specificity were 50.0% and 91.9%, respectively. After repetitive immunohistochemical staining for the 71 cases showing disagreement with the to MSI status, the sensitivity and the specificity of the second round of immunohistochemical staining were 53.3% and 97.6%, respectively. CONCLUSIONS: Sporadic colorectal cancer with MSI appears to have distinct characteristics. However, immunohistochemical staining for hMLH1 and hMSH2 is not accurate enough to be used instead of MSI.

Relation between the Low Molecular Weight Cyclin E and Mutator Phenotype in Sporadic Colorectal Cancer

PURPOSE: Cyclin E/CDK2 complexes are thought to play critical roles in multiple cell cycle events, including DNA replication, centrosome duplication, and activation of the E2F transcriptional program. Deregulation of the cell cycle control mechanism is an obligatory step in tumorigenesis. Cyclin E gene amplification and the high expression of low molecular weight (LMW) cyclin E proteins are reported to be important events in breast, and other cancers. According to recent studies, the overexpression of cyclin E protein has the role of developing chromosomal and microsatellite instability (MSI). MSI is known as one of the pathways by which colorectal cancer develops. LMW cyclin E variants are also expressed exclusively in cancer tissues. Therefore, we hypothesize that the LMW cyclin E maybe related to the MSI in sporadic colorectal cancers. METHODS: The expressions of the LMW cyclin E, CDK2 proteins and MSI stati were detected by western blot and PCR-SSCP analysis, respectively, using five Bethesda microsatellite markers in 49 sporadic colorectal cancers, which were compared with matched normal colonic mucosal tissues. RESULTS: There were 5, 10 and 34 cases of MSI-H (10.2%), MSI-L (20.4%) and MSS (69.4%), respectively. LMW cyclin E was over-expressed in 4 of the 5 MSI-H (80%) and 31 of the 44 MSI-L and MSS cases (70.5%). No correlation was found between LMW cyclin E (Fisher exact one-tailed P= 0.554), CDK2 expression (Fisher exact one-tailed P=0.569) and microsatellite instability in sporadic colorectal cancers. CONCLUSION: The expression of the LMW cyclin E variant was not associated with the MSI status in sporadic colorectal cancers.

Clinicopathological Characteristics of Sporadic Colorectal Cancer with DNA Microsatellite Instability

PURPOSE: Sporadic colorectal cancers, with DNA microsatellite instability (MSI), have been characterized by a predilection area of proximal colon, younger age onset, exophytic growth and larger tumor size. MSI colorectal cancers have recently been had a good survival rate. The aim of this study is to determine the MSI status in sporadic colorectal cancers, and compare their clinical and pathological characteristics with those of MSS (Microsatellite Stable) cancers. METHODS: Between March 1995 and December 1997, deep frozen fresh tissue of 107 eligible colorectal cancer patients, who underwent surgical resections, were used for analysis. Hereditary nonpolyposis colorectal cancer, and familial adenomatous polyposis, patients were excluded. All the patients were registered on a colorectal cancer database, and followed up completely with regular visits for a potential recurrence. Genomic DNA was prepared by the SDS-proteinase K and phenol chloroform extraction methods. The DNA was amplified by PCR at five microsatellite loci (BAT26, BAT25, D2S123, D5S346, and D17S250) to evaluate the MSI. The PCR products were separated in 6% polyacrylamide gels, containing 5.6 M urea, followed by autoradiography. The MSI was defined as being over 2 marker positive, and the MSS as 1 marker positive, all marker negatives were classed as MSS. The survival rates were calculated by the Kaplan- Meier methods. RESULTS: MSI was noted at 16/107 (15%), with mean ages for the patients of 51.8 vs. 58.6 years old for MSI and MSS, respectively. For the patients under 40 years old 5 (31.3%) vs. 6 (6.6%) had MSI and MSS, respectively (P<0.01). The cancer was located in the right colon in 12 of each of the MSI and MSS (P<0.01). There were no MSI rectal cancer tumors. The average tumor sizes were 7.6 3.6 cm vs. 5.3 2.2 cm (P<0.01) for MSI and MSS, respectively, but there were no correlations with the frequency of associated polyps, recurrence and distant metastasis between MSI and MSS. The cells were well differentiated (12.5% vs. 17.6%), moderately differentiated (68.8% vs. 76.9%), poorly differentiated (6.2% vs. 3.3%), and mucinous type (12.5% vs. 2.2%), with MSI and MSS, respectively. The overall survival rates were 93.8% vs. 73.8% for MSI and MSS (P=0.07), respectively. CONCLUSIONS: Sporadic colorectal cancer, with DNA microsatellite instability (MSI), was located predominantly in the proximal colon, in the younger age onset, and larger size of tumor. The survival rate of the patients with MSI tumors were good, but with no statistical significance.

Genetic Instability and Mutations of Mismatch Repair (MMR) and p53 Gene in Colorectal Cancers with Multiple Polyps and Sporadic Colorectal Cancers

PURPOSE: General conceptions of carcinogenic mechanisms by recent reports are ras-p53 gene pathway in sporadic colorectal cancers (SCC), MMR gene pathway in hereditary nonpolyposis colorectal cancer (HNPCC) and APC gene in familial adenomatous polyposis (FAP). But in the colorectal cancer with multiple polyps (CCMP), the carcinogenic pathway is not still defined exactly. In order to find out the which carcinogenic pathway control the CCMP and SCC, genetic instability were studied in CCMP and SCC. METHODS: In this study, genetic instability on D2S123, D3S1029, D5S107, D6S87 and AP delta3 foci and gene mutations of hMLH1 (exon 2, 16, 19), hMSH2 (exon 11, 12, 13, 14) gene of MMR gene, p53 gene (exon 5, 6, 7, 8, 9) were studied on the 60 DNA samples of CCMP (30 cases) and SCC (30 cases). RESULTS: 1. Observed positive genetic instability was higher in CCMP (30%) than SCC (20%), and was higher in right colon cancers (33%) than left colon cancers (23%) or rectal cancers (17%), but not significant statisitically. And observed positive genetic instability was lower in moderate differentiated cancers (16%) than well (67%) or poorly (60%) differentiated cancers (P=0.005). 2. Any mutations of hMLH1 and hMSH2 gene of MMR gene were not observed in both of CCMP and SCC, but 3 cases (2 CCMPs and 1 SCC) point mutations of intron of hMSH2 gene, which were higher in positive genetic instability than negative (P=0.002). 3. This 3 cases point mutations were C for T which were on 6th bases upstream from codon 669. 4. From the results of SSCP for nucleotide sequencing of p53 gene, the abnormal bands were observed in 30% (9 of 30) of CCMP and SCC. Also the abnormal bands were observed in both of positive or negative genetic instability without differences. CONCLUSIONS: With above results the authors suggested that the mechanism of genetic instability and mutations of p53 gene strongly affect the mechanism of carcinogenesis in SCC and CCMP. And there are relationship between genetic instability and point mutation at intron region of hMSH2 gene. However further evaluation and research is needed to establish relation between APC gene and other different kind of MMR gene.

hMLH1/hMSH2 Protein Expression in Sporadic Colorectal Carcinoma and Its Clinicopathological Significance

PURPOSE: DNA replication errors (RERs) in repeated nucleotide sequences (microsatellite instability) is caused by defective mismatch repair (MMR) genes. Ninety percent of colorectal carcinomas in hereditary nonpolyposis colorectal cancer (HNPCC) patients and 10-15% of sporadic colorectal cancers show microsatellite instability. In the majority of colorectal cancers with microsatellite instability, the defective MMR gene is hMLH1 or hMSH2. The author examined immunohistochemical expression of hMLH1 and hMSH2 in 75 cases of colorectal carcinomas excluding HNPCC, based on Amsterdam criteria for investigating clinicopathological characteristics and prognosis in hMLH1/hMSH2 negative cases. METHODS: Formalin fixed, paraffin blocks obtained from tumors of 75 cases of colorectal cancers were stained with two monoclonal antibodies (hMLH1 and hMSH2). The correlation between hMLH1/hMSH2 negativity, and clinicopathological feature and prognosis were statistically analysed. RESULTS: Twelve cases (16.0%) showed hMLH1/hMSH2 negativity. Negative expression of hMLH1/hMSH2 was associated with early onset (under age 50), proximal location, multiplicity, mucinous histologic type and poor differentiation. There was a significant survival advantage in patients with hMLH1/hMSH2 negative colorectal carcinoma. CONCLUSIONS: This study shows that hMLH1/hMSH2 negative colorectal carcinomas have the same clinicopathological characteristics of colorectal carcinomas with microsatellite instability. The immunohistochemical test for hMLH1/hMSH2 protein can be a simple screening method routinely applicable. The result of this test is available for establishing guidelines for management, and an independent prognostic factor for sporadic colorectal cancers.

Correlation between patient′s age at onset of sporadic colorectal cancer and microsatellite instability in cancer tissues / 中国癌症杂志

Purpose:To investigate the relationship between inactivity of mismatch repair system in colorectal cancer (CRC) in Chinese patients and their age at onset of CRC.Methods:Genomic DNA extracted from colorectal cancer tissues and their normal colon tissues were subjected to analysis for microstallite instability (MSI) in six of DNA markers in 100 colorectal cancer patients.Results:Microsatellite instability positive (MSI + ) was detected in 46 out of 100 (46%) of CRC tissues, MSI + H in 18/100 (18%) and MSI + L in 28/100 (28%). The rate of MSI + in the CRC patients who are younger than 45 years old is higher than that in older patients (≥65 years ) ( P

Microsatellite Instability and hMSH2 Gene Mutations in Sporadic Colorectal Cancers

Microsatellites are short nucleotide repeat sequences present throughout the human genome. Alterations of microsatellites, comprising extra or missing copies of these se quences, have been termed microsatellite instability(MSI, genetic instability, replication errors, RER(+) phenotype). To date, at least four genes involved in DNA mismatch repair, hMSH2, hMLH1, hPMS1 and hPMS2, are thought to account for the observation of microsatellite instability in tumor from Hereditary nonpolyposis colorectal cancer (HNPCC) patients. The genetic defect responsible for the MIN+ phenotype in sporadic colorectal cancer, however, has yet to be clearly delineated. The purpose of this study was to determine the presence of MSI in sporadic cancer and to correlate its occurrence with clinicopathological parameters, we have studied six microsatellite loci by use of polymerase chain reaction amplification and denaturing polyacrylamide gel electrophoresis. We found that 20%(9 of 46 cases) sporadic colorectal cancers showed RER at two or several loci(RER+). Microsatellite instability was associated with location of the tumor in the proximal colon 66%(6 of 9 cases) and with poorly differentiated tumor phenotype 56%(5 of 9 cases). In order to better understand the role of somatic alterations within hMSH2 in the process of colorectal tumorigenesis, we examined the most conserved regions(codon 598~789) of this gene in nine patients with MIN spotadic colorectal cancer. 6 patient of RER(+) colorectal ca. patients had a polymorphism which was a T to C base change in the intron sequence at -6 position of the splice acceptor site at the 5'end of exon 13. This particular sequence variation is a polymorphism rather than a mutation which increase cancer susceptability. These data suggest that the genetic instability is detect ed in some colorectal cancers and play an important role in the pathogenesis of sporadic colorectal cancer.