| BackgroundThe morbidity and mortality of colorectal cancer (CRC) has increased year by year, it's the third-fourth leading causes of cancer-related death in China,and age at onset was significantly ahead of schedule,the cost of treating CRC is raised at the same time. As a curable disease, the outcome of the patient depends on the early diagnosis and early detection: earlier stage means better outcome. So the CRC screening of the average risk people is of great social and economic significance. Among the frequently used screening methods, such as colonoscopy, fecal occult blood testing (FOBT), lectronic colonoscopy and iconography can not satisfy us completely, because of poor sensitivity and specificity, or lack of patients compliance, or the high cost,even serious complications such as bleeding and perforation et al.so these methods is not suitable for CRCscreening. Along with the development of molecular Biology and gene technology, detecting DNA alterations in stool DNA, as a promising noninvasive screening method for CRC, is recommended. With the theory of epigenetics and technology development, a lot of gene aberrant methylation status was closely confirmed with CRC occurrence and development .And some pilot studies have found that detection of epigenetic alterations in stool DNA, such as DNA methylation, can get better outcome than that of detection of genetic alteration, such as mutation. Studies shows that MAL ,CDKNA2A, MGMT gene promoter region hypermethylation status and are closely related to colorectal cancer progress.AS we know , a number of genetic alterations have been associated with CRC's occurrence and development ,and detection of a single gene often exist the problem of low sensitivity, which suggested that a panel of methylated genes might be required to improve the detection sensitivity for CRC.ObjectiveWe analyzed methylation of three genes: MAL ,CDKNA2A and MGMT in stools from patients with CRC and colorectal polyps as well as from normal controls by MSP, and evaluated the feasibility of detecting hypermethylation in stools as a non-invasive screening tool for CRC and precancerouslesions and it's relationship with the clinicopathological characteristics of CRC patients. We also evaluated the stool DNA isolation kit in isolating and purifying stool DNA at the same time.MethodsStool from patients suffering from colorectal cancer, colorectal polyps or normal people , who were admitted to Chang Hai Hospital in the second department of general surgery during September 2008 to January 2009, were collected. Clinical characteristics of these patients and medical record reports were reviewed. After stool DNA isolated by the stool DNA isolation kit, MS-PCR (methylation-specific PCR) was applied to analyze the promoter hypermethylation of MAL,CDKNA2A and MGMT gene in the stool DNA.ResultsStool DNA were isolated from stool samples of 144patients and at last 144 samples passed all the testing procedures, including 69 patients with CRC and 49 patients with colorectal polyps and 26 patients without colorectal neoplasia. The patients without colorectal neoplasia were included in control group. Methylated MAL,CDKN2A and MGMT were detected in 78.3%, 52.3%,55.1% of CRC patients,respectively. And the overall prevalence of fecal DNA with at least one methylated gene was 92.7% in patients with CRC; Methylated MAL,CDKN2A and MGMT were detected in 72.7%, 54.5%, 45.51% of advanced adenomas(size >1 cm,intraepithelial pecilotumor,Villous Adenoma or Tubulovillous adenoma )patients,respectively. And the overall prevalence of fecal DNA with at least one methylated gene was 81.8% in patients with advanced adenomas; Methylated MAL,CDKN2A and MGMT were detected in 46.2%, 30.8%,30.8% of non-advanced adenomas patients,respectively. And the overall prevalence of fecal DNA with at least one methylated gene was 61.5% in patients with non-advanced adenomas; Methylated MAL,CDKN2A and MGMT were detected in 50.0%, 16.7%,33.3% of serrated adenoma(named mixed hyperp lastic adenomatous polyps in department of pathology of Changhai Hosptial)patients,respectively. And the overall prevalence of fecal DNA with at least one methylated gene was 66.7% in patients with serrated adenomas.In contrast, Methylated MAL,CDKN2A and MGMT were detected in 26.3%, 15.8%,10.5% of hyperplastic polyp patients,respectively. And the overall prevalence of fecal DNA with at least one methylated gene was 42.1% in patients with hyperplastic polyps; Methylated MAL,CDKN2A and MGMT were detected in 3.8%, 0%,3.8% of patients without colorectal neoplasia,respectively. Methylated MAL,CDKN2A and MGMT were detected in 73.8%, 54.8%,52.4% ofâ… -â…¡stage CRC and precancerous lesion(advanced adenoma),respectively. And the overall prevalence of fecal DNA with at least one methylated gene was 42.1% in patients withâ… -â…¡stage CRC and precancerous lesion. The promoter hypermethylation of MAL,CDKN2A and MGMT gene didn't correlate with age, gender, tumor site,TNM stage,lymphatic metastasis and beyond metastasis. ConclusionsThe stool DNA isolation kit can provide purified stool DNA. The MAL ,CDKN2A and MGMT gene promoter hypermethylation was not associated with the clinicopathological characteristics of the CRC patients, including age, gender, tumor site,TNM stage ,lymphatic metastasis and beyond metastasis. The stool DNA MAL , CDKN2A and MGMT gene promoter hypermethylation test carried high potential for the remote detection of CRC of early stage and precancerous lesions as non-invasive screening method, and combined analysis of hypermethylated MAL ,CDKN2A and MGMT gene in fecal could further increase the sensibility,although there would be a little descend of specificity.The combined detection of genes promoter hypermethylation in stool DNA is a potential noninvasive screening method for CRC and deserves further research.
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