Promoter Methylation Regulates Cyclooxygenase Expression in
Promoter Methylation Regulates Cyclooxygenase Expression in
Introduction: Overexpression of cyclooxygenase (COX-2) is commonly observed in human cancers. In a murine model of metastatic breast cancer, we observed that COX-2 expression and enzyme activity were associated with enhanced tumorigenic and metastatic potential. In contrast to the high COX-2 expression in metastatic tumors, transplantation of poorly tumorigenic tumor cell lines to syngeneic mice results in less COX-2 expression and less COX-2 activity in vivo. Aberrant CpG island methylation, and subsequent silencing of the COX-2 promoter, has been observed in human cancer cell lines and in some human tumors of the gastrointestinal tract.
Methods: Using bisulfite modification and a methylation-specific PCR, we examined the methylation status of the COX-2 promoter in a series of four closely-related murine mammary tumors differing in COX-2 expression and metastatic potential.
Results: We showed that line 410, which does not express COX-2 in vivo, exhibited evidence of promoter methylation. Interestingly, the metastatic counterpart of this cell (line 410.4) displayed only the unmethylated COX-2 promoter, as did two additional cell lines (lines 66.1 and 67). The methylation patterns observed in vitro were maintained when these murine mammary tumor lines were transplanted to syngeneic mice. Treatment with the DNA demethylating agent 5-aza-deoxycytidine increased COX-2 mRNA, increased protein and increased enzyme activity (prostaglandin synthesis).
Conclusions: These results indicate that COX-2 promoter methylation may be one mechanism by which tumor cells regulate COX-2 expression. Upregulation of COX-2 expression in closely related metastatic lesions versus nonmetastatic lesions may represent a shift towards the unmethylated phenotype.
It is now well established that the inducible isoform of cyclooxygenase, COX-2, is commonly overexpressed in many solid tumors. Epidemiological studies as well as clinical trials employing selective and nonselective COX-2 inhibitors indicate that COX-2 is mechanistically involved in colorectal carcinogenesis, and possibly in other sites of carcinogenesis. In addition to early cancer development, evidence is beginning to accumulate that COX-2 may also contribute to late-stage progression (i.e. tumor metastasis). Early reports in breast cancer suggested a linkage between prostaglandin production and aggressive disease, but less is known regarding the specific contribution of the COX-2 isoform to behavior. Studies are beginning to emerge that suggest heightened COX-2 expression is associated with more aggressive breast cancer.
We have examined COX-2 expression in a murine model of metastatic breast cancer and have observed that COX-2 protein expression in vivo, as well as COX-2 enzyme activity (i.e. prostaglandin E2 [PGE2] synthesis), is positively correlated with more aggressive disease. Furthermore, selective COX-2 or COX-1 inhibitors control metastatic disease in this model system.
COX-2 expression and PGE2 production were very low or absent in tumors derived from transplantation of nonmetastatic cell lines to syngeneic mice, but metastatic tumors expressed high levels of COX-2 protein, mRNA and PGE2. COX-2 expression is controlled in normal tissues at the transcriptional level. In cancer, expression of many genes is regulated by aberrant promoter methylation. Several recent studies have likewise suggested that methylation of COX-2 promoter DNA at areas of CpG islands may result in aberrant COX-2 methylation in human tumors of colorectal or gastric origin. Based on these studies, we asked whether COX-2 promoter methylation regulates gene expression in mammary epithelial tumors.
Introduction: Overexpression of cyclooxygenase (COX-2) is commonly observed in human cancers. In a murine model of metastatic breast cancer, we observed that COX-2 expression and enzyme activity were associated with enhanced tumorigenic and metastatic potential. In contrast to the high COX-2 expression in metastatic tumors, transplantation of poorly tumorigenic tumor cell lines to syngeneic mice results in less COX-2 expression and less COX-2 activity in vivo. Aberrant CpG island methylation, and subsequent silencing of the COX-2 promoter, has been observed in human cancer cell lines and in some human tumors of the gastrointestinal tract.
Methods: Using bisulfite modification and a methylation-specific PCR, we examined the methylation status of the COX-2 promoter in a series of four closely-related murine mammary tumors differing in COX-2 expression and metastatic potential.
Results: We showed that line 410, which does not express COX-2 in vivo, exhibited evidence of promoter methylation. Interestingly, the metastatic counterpart of this cell (line 410.4) displayed only the unmethylated COX-2 promoter, as did two additional cell lines (lines 66.1 and 67). The methylation patterns observed in vitro were maintained when these murine mammary tumor lines were transplanted to syngeneic mice. Treatment with the DNA demethylating agent 5-aza-deoxycytidine increased COX-2 mRNA, increased protein and increased enzyme activity (prostaglandin synthesis).
Conclusions: These results indicate that COX-2 promoter methylation may be one mechanism by which tumor cells regulate COX-2 expression. Upregulation of COX-2 expression in closely related metastatic lesions versus nonmetastatic lesions may represent a shift towards the unmethylated phenotype.
It is now well established that the inducible isoform of cyclooxygenase, COX-2, is commonly overexpressed in many solid tumors. Epidemiological studies as well as clinical trials employing selective and nonselective COX-2 inhibitors indicate that COX-2 is mechanistically involved in colorectal carcinogenesis, and possibly in other sites of carcinogenesis. In addition to early cancer development, evidence is beginning to accumulate that COX-2 may also contribute to late-stage progression (i.e. tumor metastasis). Early reports in breast cancer suggested a linkage between prostaglandin production and aggressive disease, but less is known regarding the specific contribution of the COX-2 isoform to behavior. Studies are beginning to emerge that suggest heightened COX-2 expression is associated with more aggressive breast cancer.
We have examined COX-2 expression in a murine model of metastatic breast cancer and have observed that COX-2 protein expression in vivo, as well as COX-2 enzyme activity (i.e. prostaglandin E2 [PGE2] synthesis), is positively correlated with more aggressive disease. Furthermore, selective COX-2 or COX-1 inhibitors control metastatic disease in this model system.
COX-2 expression and PGE2 production were very low or absent in tumors derived from transplantation of nonmetastatic cell lines to syngeneic mice, but metastatic tumors expressed high levels of COX-2 protein, mRNA and PGE2. COX-2 expression is controlled in normal tissues at the transcriptional level. In cancer, expression of many genes is regulated by aberrant promoter methylation. Several recent studies have likewise suggested that methylation of COX-2 promoter DNA at areas of CpG islands may result in aberrant COX-2 methylation in human tumors of colorectal or gastric origin. Based on these studies, we asked whether COX-2 promoter methylation regulates gene expression in mammary epithelial tumors.
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