Study Of Hypomethylation Of CAGE, MAGE-A1 And MAGE-A3 Promoter In Gastric Carcinoma

Objective:DNA methylation,as an important epigenetic modification,plays an important role in tumorigenesis of gastric cancer.Aberrant DNA methylation,such as the hypermethylation of tumor suppressor genes and the hypomethylation of oncogene,is recognized as the main mechanisms of tumor suppressor gene inactivation and oncogene activation.Therefore,it is important to study aberrant DNA methylation for tumor prevention and treatment.CAGE,MAGE-A1 and MAGE-A3 are included in cancer/testis antigen genes.The hypomethylation of the genes promoter has been reported in some tumor tissues and is related to the occurrence and development of tumor.The objectives of the study were to reveal the relationship between the hypomethylation of CAGE,MAGE-A1,MAGE-A3 promoter CpG island and clinicopathological features and to explore the role of the hypomethylation of CAGE, MAGE-A1,MAGE-A3 promoter CpG island in earlier and development of gastric carcinoma,in order to search new molecular markers for cancer detection.Materials and Methods:1.The endoscopic biopsy specimens were collected randomly from 25 normal control,32 premalignant lesions and 30 gastric carcinoma in Department of Gastroenterology.The specimens were divided into three groups by pathological results including normal control,premalignant lesions and gastric carcinoma.25 cases of normal control included 14 males and 11 females,average age 52 years old. 32 cases of premalignant lesions included 20 males and 12 females,average age of 56 years old.30 cases of gastric carcinoma included 18 males and 12 females,average age of 58 years old.Gastric carcinoma group,according to different histological types, were divided into 3 groups including adenocarcinoma,signet-ring cell carcinoma and undifferentiated carcinoma;according to different differentiation degree,were divided into 3 groups including well-differentiated,moderately-differentiated and poorly-differentiated;according to different lymphatic metastasis,were divided into 2 groups including lymphatic metastasis positive and lymphatic metastasis negative; according to clinical stages,was divided into 2 groups includingⅠ+ⅡandⅢ+Ⅳ.2.DNA was extracted from all tissues and then treated by Sulfite.The hypomethylation status of CAGE,MAGE-A1 and MAGE-A3 promoter was detected by MSP. Results:1.The hypomethylation of the CAGE,MAGE-A1 and MAGE-A3 promoter 1.1 The positive rate of hypomethylation of CAGE promoter was 4.0%(1/25), 37.5%(12/32) and 80.0%(24/30) in normal control,premalignant lesions and gastric carcinoma respectively.There was significant difference among three groups (P＜0.05).1.2 The positive rate of hypomethylation of MAGE-A1 promoter was 0.0%(0/25), 21.9%(7/32) and 60.0%(18/30) in normal control,premalignant lesions and gastric carcinoma respectively.There was significant difference among three groups (P＜0.05).1.3 The positive rate of hypomethylation of MAGE-A3 promoter was 8%(2/25), 12.5%(4/32) and 46.7%(14/30) in normal control,premalignant lesions and gastric carcinoma respectively.The positive rate of hypomethylation in normal mucosa and premalignant lesions was significantly lower than that of the gastric carcinoma tissues (P＜0.05),but the difference between the normal mucosa and the premalignant lesions was not statistically significant(P＞0.05).1.4 Among the 30 gastric carcinoma tissues,more than one gene promoter hypomethylation were found in 27 eases;more than two genes promoter hypomethylation were found in 23 cases;three genes promoter hypomethylation were found in 7 cases.Therefore,the combined hypomethylation analysis of the genes promoter may enhance the efficiency of detecting the hypomethylation.2.The relationship between the hypomethylation of CAGE,MAGE-A1,MAGE-A3 promoter and the clinicopathologic features.2.1 The relationship between the hypomethylation of the genes promoter and the differentiation:The positive rate of hypomethylation of CAGE promoter in well-differentiated, moderately-differentiated and poorly-differentiated groups was 62.5%, 81.8%and 90.9%respectively.No statistical significance was observed among three groups(P＞0.05).The positive rate of hypomethylation of MAGE-A1 promoter was 25.0%,63.6%and 81.8%respectively.The positive rate of hypomethylation of MAGE-A3 promoter was 12.5%,45.5%and 72.7%respectively.There was significant difference among three groups.(P＜0.05)2.2 The relationship between the hypomethylation of the genes promoter and the lymphatic metastasis:The positive rate of hypomethylation of CAGE promoter in the lymphatic metastasis positive and negative groups was 94.7%and 54.5%respectively. There was significant difference between two groups.The positive rate of hypomethylation of MAGE-A1 promoter was 68.4%and 45.5%respectively.No significant difference was observed(P＞0.05).The positive rate of hypomethylation of MAGE-A3 promoter was 63.2%and 18.8%respectively.There was significant difference between two groups.(P＜0.05).2.3 The relationship between the hypomethylation of the genes promoter and the TNM stage:The positive rate of hypomethylation of CAGE promoter inⅠ+ⅡandⅢ+Ⅳgroups was 58.3%and 94.4%respectively.The positive rate of hypomethylation of MAGE-A1 promoter inⅠ+Ⅱgroup andⅢ+Ⅳgroup was 25.0% and 83.3%respectively.There was significant difference betweenⅠ+ⅡandⅢ+Ⅳgroups.The positive rate of hypomethylation of MAGE-A3 was 33.3%and 55.6% respectively.No significant difference was observed(P＞0.05).2.4 No statistical significance was observed between the positive rate of hypomethylation of CAGE,MAGE-A1,MAGE-A3 promoter and age,sex and tumor histological type(P＞0.05).Conclusions:1.The hypomethylation of CAGE,MAGE-A1 and MAGE-A3 promoter was significantly increased from normal mucosa,premalignant lesion to gastric carcinoma. The hypomethylation of CAGE,MAGE-A1 and MAGE-A3 promoter may occur in the early stage of carcinogenesis and plays a crucial role in the occurrence and development of gastric cancer.2.Hypomethylation of CAGE promoter was significantly correlated with lymphatic metastasis and TNM stage;Hypomethylation of MAGE-A1 promoter was significantly correlated with the differentiation and TNM stage;Hypomethylation of MAGE-A3 promoter was significantly correlated with the differentiation and lymphatic metastasis.3.The combined hypomethylation analysis of CAGE,MAGE-A1 and MAGE-A3 promoter may enhance the efficiency of detecting the hypomethylation.