Abstract: Objective To explore the effect of demethylating drug 5-Aza-2′-deoxycytidine (5-Aza-CdR) on methtylation status of the Ras-association domain family1A gene (RASSF1A) in human endometrial carcinoma. Methods Randomly assign the human endometrial carcinoma cell line HEC-1-B into groups and use demethylating drug 5-Aza-CdR of different concentration to treat them. Then Methylation-specific polymerase chain reaction (MSP), real-time PCR, Western blot, TUNEL technology were used to analyze methylation status of RASSF1A promoter CpG islands,RASSF1A mRNA expression, RASSF1A protein expression and apoptosis of HEC-1-B cell. Results High DNA methylation in RASSF1A gene promoter region, low RASSF1A mRNA level and protein expression and out of control of human endometrial carcinoma cell HEC-1-B apoptosis were observed. 5-Aza-CdR of different concentration could reverse RASSF1A gene’s methylation status, recover the expression of mRNA and protein, and control the growth of HEC-1-B by inducing apoptosis. Conclusion Aberrant methylation of RASSF1A in endometrial cancer as a therapeutic target, demethylating agent 5-Aza-CdR could be an effective way of gene therapy.