The Effect of HO-1 on AMN107 In Promoting K562/A02 Apoptosis

• Published in: Blood 2010-11, Vol.116 (21), p.1231-1231
• Main Authors: Wang, Jishi, Chai, Baisheng, Chen, Cheng, Fang, Qin
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V116.21.1231.1231

Abstract 1231 Heme oxygenase-1(HO-1), also known as heat shock protein 32(Hsp32), has recently been identified as a stress-related survival molecule that acts anti-apoptotic and cytoprotective in inflammatory reactions. In the present study, we provide evidence that HO-1 is effective on AMN107(Nilotinib) in promoting K562/A02 apoptosis which is induced by STI571(Imatinib),and investigate AMN107 on the mechanism of resistance to STI571. HO-1 gene was cloned from rat liver by RT-PCR. And the retrovirus vector pQCXIP-EGFP-C1 was constructed. K562/A02 cell which was expressed HO-1 highly was seemed as gene-transfected group. At the same time, we set the empty vector transfected group and untransfected group. K562/A02 cell was cultivated with AMN107 in gene-transfected, empty vector transfected and untransfected groups. Expression of HO-1 mRNA was demonstrable by RT-PCR, Real-time PCR, and the HO-1 protein by Western blotting. Constant MTT assay and cell number count were used to detect the proliferation of leukemia cells after treatment with AMN107. Apoptosis was determined by morphological observation and flow cytomertry analysis after AnnexinV/PI double labeling. Also we detected the intracellular drug concentration by HPLC after treatment with the same concentration of AMN107. HO-1 gene was cloned from rat liver successfully. The sequences were confirmed by restriction enzyme digestion analysis and sequencing. The virus was packaged in 293T cells and titer of virus was tested by Real-Time PCR, 8.09×1010v.p./mL. Following transfer the retrovirus vector into K562/A02, 72 hours after transfection, it showed that HO-1 was expressed highest by fluorescence micrope. High expression of HO-1 was detected by RT-PCR, Real-Time PCR and Western blotting. After 10umol/L AMN107 treated, the expression of HO-1 was clearly lower in empty vector transfected group and untransfected group (p