Low concentrations of 5-aza-2′-deoxycytidine induce breast cancer stem cell differentiation by triggering tumor suppressor gene expression

BACKGROUND Breast cancer stem cells (BCSCs) are considered the cause of tumor growth, multidrug resistance, metastasis, and recurrence. Therefore, differentiation therapy to reduce self-renewal of BCSCs is a promising approach. We have examined the effects of 5-aza-2'-deoxycytidine (DAC) on BCSC differentiation. MATERIALS AND METHODS BCSCs were treated with a range of DAC concentrations from 0.625 to 100 µM. The differentiation status of DAC-treated BCSCs was graded by changes in cell proliferation, CD44(+)CD24(-) phenotype, expression of tumor suppressor genes, including BRCA1, BRCA2, p15, p16, p53, and PTEN, and antitumor drug resistance. RESULTS DAC treatment caused significant BCSC differentiation. BCSCs showed a 15%-23% reduction in proliferation capacity, 3.0%-21.3% decrease in the expression of BCSC marker CD44(+)/CD24(-), activation of p53 expression, and increased p15, p16, BRCA1, and BRCA2 expression. Concentrations of DAC ranging from 0.625 to 40 µM efficiently induce cell cycle arrest in S-phase. ABCG2, highly expressed in BCSCs, also decreased with DAC exposure. Of particular note, drug-sensitivity of BCSCs to doxorubicin, verapamil, and tamoxifen also increased 1.5-, 2.0-, and 3.7-fold, respectively, after pretreatment with DAC. CONCLUSION DAC reduced breast cancer cell survival and induced differentiation through reexpression of tumor suppressor genes. These results indicate the potential of DAC in targeting specific chemotherapy-resistant cells within a tumor.