The BCR/ABL tyrosine kinase inhibitor, imatinib, has shown substantial effects in blast crises of chronic myelogenous leukemia. However, most patients relapse after an initial clinical response, indicating that drug resistance is a major problem for patients being treated with imatinib. In this study, we generated a new imatinib-resistant BCR/ABL-positive cell line, KCL22/SR. The 50% inhibitory concentration of imatinib was 11-fold higher in KCL22/SR than in the imatinib-sensitive parental cell line, KCL22. However, KCL22/SR showed no mutations in the BCR/ABL gene and no increase in the levels of BCR/ABL protein and P-glycoprotein. Furthermore, the level of phosphorylated BCR/ABL protein was suppressed by imatinib treatment, suggesting that mechanisms independent of BCR/ABL signaling are involved in the imatinib resistance in KCL22/SR cells. DNA microarray analyses demonstrated that the signal transduction-related molecules, RAS p21 protein activator and RhoA, which could affect Ras signaling, and a surface tumor antigen, L6, were upregulated, while c-Myb and activin A receptor were downregulated in KCL22/SR cells. Furthermore, imatinib treatment significantly suppressed the level of phosphorylated p44/42 in KCL22 cells but not in KCL22/SR cells, even when BCR/ABL was inhibited by imatinib. These results suggest that various mechanisms, including disturbance of Ras-mitogen-activated protein kinase signaling, are involved in imatinib resistance.