Inhibition of thioredoxin reductase by auranofin induces apoptosis in adriamycin-resistant human K562 chronic myeloid leukemia cells.

Mammalian thioredoxin reductase (TrxR) catalyzes the NADPH-dependent reduction of oxidized thioredoxin (Trx) and plays a central role in regulating cellular redox homeostasis, cell growth and apoptosis. Increasing evidence shows that TrxR is over-expressed or constitutively active in many tumor cells. Moreover, TrxR appears to contribute to increased tumor cell growth and a resistance to chemotherapy. In this study, we evaluated the activity of TrxR in adriamycin-resistant leukemic cells (K562/ADM) and adriamycin-sensitive parental lines (K562), and found that TrxR activity was higher in the drug resistant cell sublines K562/ADM than in K562 drug sensitive parental cells. Auranofin, a gold(I) compound clinically used as an antirheumatic agent, reduced TrxR activity and was more effective than adriamycin in decreasing cell viability in K562/ADM cells. In addition, auranofin induced apoptosis in dose-dependent manners, accompanied by caspase-3 activation in K562/ADM cells. Our results demonstrate that inhibition of TrxR and induction of apoptosis by auranofin provides its ability in overcoming adriamycin resistance in K562/ADM cells.

[siRNA silences mdr1 gene expression and reverses apoptosis resistance of K562/ADM cells line].

OBJECTIVE: To explore the effect of small interfering RNA(siRNA) on silence of mdr1 gene and reversal of apoptosis resistance in multidrug-resistant (MDR) human leukemia K562/ADM cell. METHODS: Human MDR leukemia cell line K562/ADM was used as the target cells. Two siRNAs (mdr1 siRNA-1 and mdr1 siRNA-2) targeted mdr1 gene were chemically synthesized and transfected into K562/ADM cells with liposome. Expression of mdr1 mRNA was determined by real-time PCR, P-glycoprotein (P-gp) expression and caspase-3 activity were measured with flow cytometry (FCM), and the cell apoptosis was observed by optical and electronic microscopy for morphology and Annexin V/PI staining. RESULTS: The mdr1 siRNA-1 and mdr1 siRNA-2 could markedly down-regulate the expression of mdr1 gene in K562/ADM cells, the expression of mdr1 mRNA decreased by 91.2% and 82.0% , and the P-gp by 74.1% and 84.4%, respectively. The caspase-3 activity was markedly enhanced, and the active caspase-3 in K562/ADM cells increased by about 40% compared to liposome alone and non-silencing controls. the sensitivity of K562/ADM cells to adriamycin-induced apoptosis was significantly augmented, the apoptotic rate of the cells treated with siRNA plus adriamycin increased by about 60% compared to adriamycin alone. CONCLUSION: siRNAs silence the expression of mdr1/P-gp to overcome the P-gp-mediated apoptosis resistance in drug-resistant K562/ADM cells.

Arsenic trioxide overcomes apoptosis inhibition in K562/ADM cells by regulating vital components in apoptotic pathway.

Extensive researches have revealed that arsenical can exert anti-tumor efficacy against several kinds of cancers including leukemia. Though, little is known about the effects of arsenical on leukemia resistant to chemotherapy, emerging as a serious clinical problem. In this study, we tested arsenic trioxide (As(2)O(3))-induced apoptosis in K562/ADM multidrug-resistant leukemic cells and investigated its possible mechanisms. Using microscopy, flow cytometry (FCM) and DNA electrophoresis, we found that As(2)O(3) could induce the cells to undergo G2/M phase arrest and apoptosis. Further, it was shown that the levels of FAS and P53 proteins increased and P-glycoprotein (P-gp) decreased upon drug action by employing FCM. Reverse transcription polymerase chain reaction (RT-PCR) detected increased mRNA product of FAS and caspase-3 genes and reduced MDR1 mRNA. CASPASE-3 activity was also enhanced after As(2)O(3) treatment. However, the expression of BCL-2 protein was not affected by the drug. Taken together, As(2)O(3) is able to reverse the apoptosis resistance in drug-resistant K562/ADM cells by modulating expression or activity of key factors associated with apoptosis induction.

[Arsenic trioxide inhibits P-glycoprotein expression in multidrug-resistant human leukemia K562/ADM cell line that overexpresses mdr-1 gene and enhances their chemotherapeutic sensitivity].

OBJECTIVE: To investigate the effects of arsenic trioxide (As(2)O(3)) on the apoptosis and P-glyco-protein (P-gp) expression of multidrug-resistant human leukemia K562/ADM cells, and the combined effects of As(2)O(3) with conventional chemotherapeutic agents. METHODS: Multidrug-resistant human leukemia cell line K562/ADM that overexpresses mdr-1 gene was used as the target cells. The cell proliferating activity was assessed with a MTT assay. Cell morphology was examined by light microscopy, confocal microscopy and electron-microscopy. P-gp expression, cell-cycle status were determined by flow cytometry. RESULTS: K562/ADM cells were highly resistant to adriamycin, and cross-resistant to daunorubicin and etoposide. As(2)O(3) at concentrations of 0.5 to 20 micromol/L inhibited the proliferation of K562/ADM cells, and K562/ADM cells were more sensitive to As(2)O(3) than their parent K562 cells did. As(2)O(3) induced marked apoptosis of K562/ADM cells showed by typical apoptotic morphological changes and the appearance of high sub-G(1) cell population. As(2)O(3) significantly inhibited the P-gp expression in K562/ADM cells, and exerted a synergistic effect on the enhancement of the cell sensitivity to adriamycin, daunorubicin and etoposide. CONCLUSION: As(2)O(3) induces growth-inhibition and apoptosis of multidrug-resistant K562/ADM cells, and augments synergistically the sensitivity of the cells to conventional chemotherapeutic agents via down-regulation of P-gp expression.