Endometrial carcinoma, the most common malignancy of the female genital tract, remains challenging to treat despite early-stage dominance. Surgical interventions and irradiation are insufficient for advanced endometrial cancer. Our aim was to investigate to explore the in vitro cytotoxicity and apoptotic effects of boric acid (BA) on endometrial adenocarcinoma cell lines (Ishikawa and HEC-1B cell lines), providing experimental evidence for the potential application of boric acid as an anticancer drug. Time- and dose-dependent cell viability was determined with the XTT cell proliferation test. Differences in mRNA levels were determined by RT-PCR using cDNAs and SYBR green assay. Colony formation and the effect of BA on wound healing were evaluated. Immunocytochemistry and TUNEL tests were performed to evaluate apoptosis. BA increased the expression of Caspase 3 and Bax in HEC-1B and Ischikawa cell lines. It was determined that BA significantly decreased the number of colonies in both cell lines (p < 0.05). In HEC-1B and Ishikawa cell lines, there was an increase in cell migration in the control group at 16 and 24 h. The apoptotic index was higher in the BA group, although it was not statistically significant. According to immunohistochemistry results, Caspase 3 and Bax expression in HEC-1B and Ishikawa cell lines were statistically increased in BA group. The expression of Bcl-2 was decreased statistically with BA treatment in both cell lines (p = 0.0001). BA treatment inhibited cell migration and colony formation, which are important for carcinogenesis, in endometrial adenocarcinoma cell lines. This inhibition was shown to occur through the apoptotic pathway.