Association Studies Of Promoter Hypermethylation Of Multiple Candidate Genes With Colorectal Cancer

Objective:The development of colorectal cancer(CRC) involves both genetic and epigenetic modifications and aberrant DNA methylation within gene promoters is primary mediator of epigenetic inheritance in CRC. Promoter methylation of the PCDH-γ-A12, SLC19A1, CREB and CYLD genes was shown to regulate their gene expression levels, while the hypermethylation of the PCDH-γ-A12, SLC19A1, CREB and CYLD promoters has not been investigated in CRC. In light of the previous findings, we investigated whether PCDH-γ-A12, SLC19A1, CREB and CYLD gene promoter methylation contributed to the occurrence and metastasis of colorectal cancer.Methods:1. 42 CRC samples and 42 adjacent normal tissue samples were collected, and then the clinical and pathological data were collated. 2. Genomic DNA was isolated and its concentration and quality were detected by spectrophotometer. 3. DNA was bisulfite-modified with the DNA Methylation-Gold Kit. 4. Methylation-specific PCR(MSP) was performed and PCR products were then electrophoresed on the agarose gels. 5. Three PCR products randomly obtained from the methylation PCR experiments were sequenced bidirectionally. 6. Statistical analysis was performed by the SPSS 18.0 and Chi-square test, and p < 0.05 was considered significant statistically.Results:PCDH-γ-A12 and SLC19A1 gene promoters were more frequently methylated in CRC tissues than normal tissues(83.33% vs 57.14%, p = 0.009 and 78.57% vs 42.54%, p = 0.002), while the methylation of CREB and CYLD gene promoters between CRC tissues and normal tissues had no significant difference(26.19% vs 11.90%, p=0.095 and 14.29% vs 11.90%, p=0.746). PCDH-γ-A12 and SLC19A1 gene promoters were more frequently methylated in lymph metastasis CRC tissues than non-lymph metastasis CRC tissues(100.00% vs 66.67%, p=0.013 and 95.24% vs 61.90%, p=0.024), while the methylation of CREB and CYLD gene promoters between lymph metastasis CRC tissues and non-lymph metastasis CRC tissues had no significant difference(33.33% vs 19.05%, p=0.292 and 19.05% vs 9.52%, p=0.659). Cp Gdinucleotides of the PCDH-γ-A12 and SLC19A1 promoters in the samples were extensive hypermethylation, whereas the CREB and CYLD promoters were unmethylated at these Cp G dinucleotides. In addition, the methylation status of the PCDH-γ-A12, SLC19A1, CREB and CYLD gene promoters had no significant correlation with the clinicopathological characteristics of CRC.Conclusions : the PCDH-γ-A12 and SLC19A1 promoters were hypermethylated and contributed to the occurrence and metastasis of colorectal cancer, while the methylation status of the CREB and CYLD gene promoters had no significant correlation with CRC. PCDH-γ-A12 and SLC19A1 promoters methylation can be a marker of early diagnosis of CRC, and may offer a new strategy for the detection and treatment of CRC.