Characterization of cancer stem-like cells in a imatinib mesylate-resistant chronic myeloid leukemia cell line / 中国组织工程研究

ABSTRACT

BACKGROUND: The treatment of chronic myelogenous leukemia (CML) is revolutionized by the tyrosine kinase inhibitor imatinib mesylate (imatinib). However, resistance to imatinib is increasingly recognized as a clinical problem, the prognosis of patients who develop imatinib resistance is poor, particularly in acute transformation phase of leukemia.OBJECTIVE: To characterize a novel CML cell line and to further elucidate the mechanisms of resistance to STI571.DESIGN: An observational comparative experiment.SETTING: Henan Institute of Haematology, Henan Tumor Hospital.MATERIALS Thirty female BALB/c nu/nu mice with 5 weeks old were purchased from Animal Center, Chinese Academy of Medical Sciences. STI571 was kindly provided by Novartis (Nuremberg, Germany). VP-16 was purchased from Bristol-Myers Squibb (Munich, Germany); anti-P-gp from Santa Cruz Company, USA; anti-ab1 from BD Biosciences Company, USA. The disposal of experimental animal was coincidence with the ethical standard.METHODS: The experiment was performed in the Henan Institute of Haematology from September 2003 to November 2005. A novel K562 cell line (K562/VP16) was achieved after exposure of the K562 cells to VP16. A small subpopulation (SP K562/VP16) that was capable of excluding Hoechst 33342 in the K562/VP16 cell line was isolated by flowcytometry sorting. The rest of the K562/VP16 cells were classified as non-SP K562/VP16. In order to elucidate the mechanisms involved in K562/VP16 SP cells which became resistant to STI571, the expression of multidrug-resistant gene 1 (MDR1), Bcr-Abl and P-gp was detected in K562, non-SP K562/VP16, or K562/VP16 SP calls, respectively. Furthermore, one thousand cells of K562, K562/VP16 SP and non-SP cells were injected,respectively, intraperitoneally into the right flanks of ten 5-week-old female BALB/c nu/nu mice. The same experiment was repeated twice.MAIN OUTCOME MEASURES: Comparison of STI571 resistance and oncogenicity of non-SP K562/VP16 and K562/VP16 SP cells.RESULTS: The MDR-1 gene expression of the Mr 170 000 P-gp was detected in K562/VP16 non-SP and K562/VP16 SP cells but not in K562 cells. The expression levels of P-gp in the two K562/VP16 cell lines were similar (P ＞ 0.05).The levels of Bcr-Abl and Abl proteins were similar in the K562 cell line and in non-SP K562/VP16 and K562/VP16 SP cells (P ＞ 0.05). Compared with non-SP K562/VP16, the K562/VP16 SP cells were more resistant to STI571, and this resistance could hardly be reversed by many multidrug resistance inhibitors. In addition, in vivo study showed that the K562/VP16 SP cells induced oncogenicity in mice, while the K562/VP16 non-SP cells failed to do so.CONCLUSION: Bcr/abl gene amplification and MDR1 overexpression may not be an important clinical mechanism in the diversity of resistance development to imatinib treatment, and the development of drug resistance by leukemia cells may be at least partly due to a rare SP cells which drives leukemia occurrence and maintenance. So, these SP cells need to be targeted for effective cancer therapy.