Prx II(Peroxiredoxin II) Activity Has Important Roles As a New Mechanism to Control ABL Tyrosine Kinase Activity in STIs(Signal Transduction Inhibitor) Treated CML Patients

Purpose: In this study, we investigated the roles of PRX II, one of 3 critical peroxidases besides catalase and GPXs, in CML primary cells at diagnosis and remission while patients were treated with STI(Signal Transduction Inhibitor) and tested the same roles in Imatinib(IM) sensitive Ph+ cell lines and resistant cell lines as well.

Methods: Newly diagnosed CML cells, IM resistant K562 cells and parental K562 cells were treated STIs and analyzed western blot assay to detect BCR-ABL, phosphorylated BCR-ABL and PRX2 protein expression level. We added N-Acetylcysteine (0-5mM, 6hr) to K562 cells to show antioxidant effect of imatinib and analyzed DCF-DA detection for intracellular ROS level and western blot for PRX2 protein level. MTT assay was performed to detect cell death by NAC time-dependent treatment of 5mM NAC(0, 24, 40, 48hr). Imatinib resistant K562 cells were established by treatment of gradual increment of imatinib. We also repeatedly investigated the effects of IM therapy using PRXII overexpressed K562 cells by transfection.

Results: At diagnosis of CML, ROS level was elevated and PRX II was either absent or significantly suppressed. As Ph chromosomes were decreased with STIs, suppressed or absent PRXs levels were restored to the level of normal individuals. These findings were also inversely correlated with the level of Ph chromosomes in the cases of disease progression and re-remission with further treatment. When STI were treated in Ph positive cell line, we found deceased cell survival and ROS level by MTT assay and DCF-DA methods respectively, but elevated PRX II by western blot. By the treatment of NAC into Ph+ cell lines, the level of DCF-DA was decreased and MTT level was down, but PRX II level was elevated. Interestingly, the level of BCR-ABL oncogene were decreased in PRX II tranfected cells. Meanwhile, we observed that PRX II restoration was mild or weak in Imatinib resistant K-562, which we established in our lab.

Conclusions: The importance of the roles of ROS and its PRX II, antioxydant enzymes in CML is further extablished by our work. Our finding may contribute to find a new pathway on which TKIs are working besides the mechanisms of ATP binding competitively, blocking the binding of ABL-BCR kinase and substract resulting apoptosis of Ph+ cells. Furthermore, our finding may be useful to overcome the STIs resistant CML in the clinics.