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Department of Urology, University of California San Francisco and VA Medical Center, San Francisco, California 94121 [M. S., L. K., A. D., R. D.], and Department of Obstetrics and Gynecology, School of Medicine, Hokkaido University, Kita-ku, Sapporo, Japan [M. S., S. F.]
We hypothesize that estrogen receptors (ERs) are differentially expressed in endometrial cancer. To test this hypothesis, we investigated the expression profile of ER
(ER-A, ER-B, ER-C) and ERß genes and CpG methylation status in endometrial cancer cell lines and tissues using reverse transcription-PCR and methylation-specific PCR and direct DNA sequencing. The results demonstrated that ER-A, ER-B, and ERß were normally expressed whereas ER-C gene was inactivated in all endometrial cancer cell lines. We further investigated the mechanisms of ER-C gene inactivation through CpG methylation pathways. The treatment with demethylating agent (5'-aza-2'-deoxycytidine) restored ER-C gene expression in all endometrial cancer cell lines. We further confirmed these findings with methylation-specific PCR and direct DNA sequencing and found that only ER-C was methylated on all five different CpG sites in all cell lines. We further analyzed 88 cancerous and 46 normal endometrial tissues. The results demonstrated that only ER-C was inactivated and methylated in 94% of cancer tissues. In 32 pairs of cancerous and normal endometrial tissues from the same patient, ER-C was methylated in 29 of 32 cancer tissues but unmethylated in all normal endometrial tissues. This is the first report that demonstrates selective ER-C gene inactivation through CpG methylation pathway in uterine endometrial cancer.
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