Human Stool OSMR And TFPI2 Gene Methylation Analysis In The Diagnosis Of Colorectal Cancer

Colorectal cancer is a common digestive system cancer. CRC largely can be prevented by the detection and removal of adenomatous polyps, and survival is significantly better when CRC is diagnosed while still localized. The goal of cancer screening is to reduce mortality through a reduction in incidence of advanced disease. Today there is a range of options for CRC screening in the average-risk population, with current technology falling into 2 general categories:stool tests, which include tests for occult blood or exfoliated DNA; and structural exams.Stool tests are best suited for the detection of cancer, although they also will deliver positive findings for some advanced adenomas, It is not dependent on the detection of occult bleeding, which is intermittent and nonspecific, and it requires only a single stool collection. Further, newer versions may have better sensitivity as more is learned about markers that are common across all prevalent CRC, as well as advanced adenomas.OSMR is a receptor of Oncostatin M (OSM), an interleukin-6(IL-6)-type cytokine identified as a potent suppressor of tumor cells.Human OSM was originally described by its capacity to inhibit melanoma proliferation in vitro [33,34], and its targets for growth inhibition include lung carcinomas, ovarian carcinomas,and breast tumors. The biological effects of OSM are mediated through a heterodimeric complex of gp130 and OSM receptor (OSMR) The escape of cell growth control by OSM in metastatic melanoma cells has recently been reported to be caused by the epigenetic silencing of OSMR due to aberrant histone modification.Tissue factor pathway inhibitor (TFPI2), a Kunitz-type serine proteinase inhibitor that protects the extracellular matrix of cancer cells from degradation and inhibits in vitro colony formation and proliferation. It is thought that loss of TFPI2 function could predispose cells toward a proinvasive program, consistent with an important role for this protein in later stages of carcinogenesis.Epigenetic is a modification of the genome, which dose not changes the genomic DNA sequence but can be transmitted to progeny cells in the process of cell division. DNA methylation and histone acetylating modification are the main forms of epigenetic modification. DNA methylation is often the early phase in the development of tumor.Epigenetic events which are seen more frequently than genetic events may have a potential to achieve high sensitivity and specificity using a single gene.ObjectiveTo study the feasibility and clinical significance of OMSR and TFPI2 gene methylation in stool of colorectal cancer patients with methylation-specific PCR.Materials and methodsFrom May,2009 to September,2010, We selected 107 patients who admitted to the first affiliated hospital of Zhengzhou University, including 60 cases of colorectal cancer and 30 normal controls and 17 colorectal polyps, and All specimens were confirmed histologically, as for 30 cases of normal controls, all colonoscopy were negative. Stool specimens were collected before colonoscopy or surgery from the 77 patients, then sent to the laboratory immediately and preserved in -70℃refrigerator. DNA was extracted with QIAamp DNA Stool mini kit(QIAGEN,Germany) and saved in -20℃refrigerator. Then DNA was modified by Bisulfite and purified according to EpiTect Bisulfite Kit(Qiagen) operation instructions and saved in -20℃refrigerator. The methylation status of genes TFPI2 and osmr was detected with the DNA with bisulfate modification as the template of MSP. The results of the experiment were processed by statistical software SPSS15.0, with gene frequencies obtained through direct counting method and each group rate compared by the X2 test. The test results signify statistically with P<0.05.Results1. We observed methylation of OSMR in 21 of 60 (35%) colorectal cancers,2 of 17 (12%) colorectal polyps, and methylation of TFPI2 in 42 of 60 (70%) colorectal cancers,3 of 17 (18%) colorectal polyps. The total positive rate was 85% (51/60).The methylation of OSMR and TFPI2 was 7% (2/30) and 3% (1/30) in normal controls.2. TFPI2 gene methylation and patients sex and tumor location have no signification correlation (P>0.05).3. Osmr gene methylation and patients sex and tumor location and Dukes stages have no signification correlation (P>0.05).4. The positive rates of TFPI2 gene methylation in Dukes stages A,B (57%) and C,D (81%) were significantly different(P<0.05). poorly differentiated tumor tissue TFP12 gene methylation positive rate was 86%. In well-differentiated tumor tissue TFPI2 gene methylation positive rate was 61%. The difference was significant (p<0.05).5. Poorly differentiated tumor tissue osmr gene methylation positive rate was 55%. In well-differentiated tumor tissue osmr gene methylation positive rate was 24%. The difference was significant (p<0.05).Conclusion1. TFPI2 and OSMR gene methylation may be one of molecular markers which are used non-invasive diagnosis for colorectal cancer.2. The correlation between TFPI2 gene methylation status and colorectal cancer patient's Dukes staging and tumor differentiation degree is obvious.TFPI2 gene methylation status and colorectal cancer patient's sex and tumor location have no signification correlation.3. The correlation between OSMR gene methylation status and colorectal cancer patient's tumor differentiation degree is obvious.OSMR gene methylation status and colorectal cancer patient's sex and tumor location and Dukes staging have no signification correlation.