Induced Apoptosis Of Multidrug Resistance Leukemia K562/A02 Cells By Magnetic Nanoparticle Of Fe₃O₄ Copolymerizating JNK1 shRNA Expression Vector and Daunorubicin

Currently, serious toxicity and poor antitumor efficacy resulting from the induction of multidrug resistance (MDR) in hematological malignancies and solid tumors are common hindrances to successful systemic chemotherapy. One of the important mechanisms of tumorigenesis and resistance to anticancer drugs was blockade of the apoptosis-inducing pathway. In order to overcome tumor MDR while maintaining good antitumor activity, biomaterial combine with gene therapy provides a novel strategy for the treatment of MDR.The aim of this study was to explore the induction of apoptosis by magnetic nanoparticle of Fe₃O₄ (MNPs-Fe₃O₄), a novel delivery system for chemotherapeutic regimen that is chemically stable, environmentally friendly, and noncytotoxic, copolymerizating JNK1shRNA expression vector with DNR, which could overcome siRNA technique instability and enhance DNR chemotherapeutic efficiency and provide a new field to reverse tumor MDR. JNK1shRNA expression vector was constructed and screened the most effective plasmid and transfected the cells. Typical apoptotic characteristics and apoptosis effect of MNPs-Fe₃O₄ and PGM-3 with DNR were investigated by DAPI staining and FCM assay, respectively. Additionally, drug accumulation and apoptosis relative-genes were evaluated in K562/AO2 leukemia cells.The results showed that PGCsilencer-U6-neo-GFP-GV102 shRNA/JNK1 was successfully constructed, which was confirmed by sequencing that expression of JNK1 gene in mRNA and protein level that had the less in PGM-3 than that of PGM-1 or PGM-2. Meanwhile, MNPs-Fe₃O₄ and PGM-3 with DNR (DNR/PGM-3/MNPs-Fe₃O₄) could synergistically induced typical apoptotic characteristics of chromatin condensation and apoptotic body with DAPI staining, and a higher rate of apoptosis were detected in K562/A02 cells treated with DNR/PGM-3/MNPs-Fe₃O₄. Intracellular DNR was higher in K562/A02 cells treated with DNR/PGM-3/MNPs-Fe₃O₄ than that of DNR/MNPs-Fe₃O₄ by FCM analysis.  Further study demonstrated that both DNR/PGM-3/MNPs-Fe₃O₄ and DNR/PGM-3 reduced the gene transcriptions and protein expressions of bcl-2, enhanced that of bax and caspase-3, decreased survivin in protein level, and has no different change in mRNA level by qRT-PCR and Western blotting analyses, respectively. These findings showed that the combination of MNPs-Fe₃O₄ with JNK1 shRNA expression vector and DNR could induce apoptosis of K562/A02 cells through elevating the ratio of bax/bcl-2, activating caspase-3, and decreasing survivin in protein level.