The purpose of this study was to analyze the effects of bufalin on the gene expression of K562 cells and on the expression of BMI‑1 pathway constituents in K562 cell apoptosis. K562 cells were treated with bufalin, and the inhibition rate and apoptosis were detected by an MTT assay, flow cytometry and a microarray assay. BMI‑1, p16INK4a and p14ARF were examined by quantitative polymerase chain reaction (qPCR). Bufalin induced significant changes in the gene expression of the K562 cells; 4296 genes were differentially expressed, 2185 were upregulated and 2111 were downregulated. The most upregulated genes were associated with transcription regulation, while the most downregulated genes were associated with the non-coding RNA metabolic processes and DNA repair. qPCR analysis demonstrated that BMI‑1 was overexpressed in the K562 cells. Bufalin is able to downregulate BMI‑1 expression levels in K562 cells prematurely and cause an increase in the expression levels of p16INK4a and p14ARF. Moreover, bufalin downregulated BCR/ABL expression levels in a time‑dependent manner, and the expression of BCR/ABL was not associated with the upregulation or downregulation of BMI‑1 expression. Bufalin may induce K562 cell apoptosis by downregulating BMI‑1 expression levels and accordingly upregulating the expression levels of p16INK4a and p14ARF. Bufalin may also induce K562 cell apoptosis via downregulating BCR/ABL expression levels, and this pathway may be independent of the BMI‑1 pathway.