Study On The Mechanisms Of Leukemia Cells' Resistance To Chemotherapeutic And Biotherapeutic Agents Mediated By Glutathione S-transferase

Chronic myelogenous leukemia (CML) is one of malignant diseases in blood system. Hydroxyurea (HU) is one of the chemotherapeutics for CML. Interferon-a (IFN-a) is an effective biotherapeutic agent for hematopoietic malignancies and lymphoma. However, CML cells showed resistance to both of HU and IFN-a. GSTPi, as a member of glutathione S-transferases (GSTs) which belong to drug metabolism enzymes, may be involved in the process of CML cells'resistance to chemotherapeutics and biotherapeutic agents.Objectives:This study uses CML cell lines such as K562,KT-1/A3 and KT-1/A3R cells as the experimental models to explore the influences of CML cells'proliferation and apoptosis induced by HU and/or IFN-a. We'll analyze the difference of GSTPi expression in the conditions with or without the inducement by HU and/or IFN-a. This study also hope to elucidate the influences and the related mechanisms of GSTPi in the process of CML cells'resistance to chemotherapeutics and biotherapeutic agents.Methods:1. Analyzing proliferation and growth states of K562,KT-1/A3 and KT-1/A3R cells by using cell counting assay, and drawing the cell growth curve diagrams.2. Observing the cell growth inhibition rates of K562, KT-1/A3 and KT-1/A3R cells by employing MTT assay at 48h after HU and/or IFN-a inducement.3. The cell cycle progression and cell apoptosis rates were examined by flow cytometry (FCM), at 48h after IFN-a, HU, and HU plus IFN-a induction of K562, KT-1/A3 and KT-1/A3R cells.4. The expression level of GSTPi was analyzed by the relatively quantitative RT-PCR and Western-blotting at 48h after cultivation of K562, KT-1/A3 and KT-1/A3R cell lines with HU and/or IFN-α.Results:1. HU can inhibit cell proliferation and induce apoptosis of K562, KT-1/A3 and KT-1/A3R cells, and the differences were significant comparing with that of the control group (P<0.01 and P<0.05 respectively).2. HU combined with IFN-a can inhibit the proliferation and induce apoptosis of CML cells more effective than alone use of HU or IFN-a (P<0.05).3. RT-PCR results showed that GSTPi gene expression in K562, KT-1/A3R cells was down-regulated by HU inducement (P<0.05).4. Western blotting showed that GSTPi protein expression in K562, KT-1/A3R cells was also down-regulated by HU inducement (P＜0.05).Conclusions:1.HU and HU plus IFN-a can inhibit the proliferation of CML cells and induce apoptosis of them. HU plus IFN-a is more effective than the alone use of HU or IFN-a.2. The results suggest that GSTPi expression is related to K562 and KT-1/A3R cells'resistance to IFN-a, and HU can reduce GSTPi expression level.3. K562, KT-1/A3R cells display the resistance to IFN-a, and HU or HU combined with IFN-a can reduce K562, KT-1/A3R cells' resistance to IFN-a by inhibiting proliferation and inducing apoptosis of the cells.