| Multidrug resistance(MDR)in cancer cells is a terminology used in reference to the phenomenon of tumor cells exhibiting simultaneous resistance to a number of structurally and functionally unrelated chemotherapeutic agents and is a major impediment to effective chemotherapy in human cancer.Mitoxantrone(MXT)has major clinical activity in the treatment of several leukaemias and breast cancer with a biological activity of interaction with DNA and the inhibition of topoisomeraseâ…¡,which induce the inhibition of S phase ana G2/M phase of cell cycle.PI3K/Akt pathway plays an important role in cell survival, growth,proliferation and chemotherapy resistance in cancer cells.Inhibition of PI3K/Akt pathway could sensitize cancer cells to chemotherapy.LY294002 and Wortmannin,two PI3K inhibitors,are used to study the function of PI3K in cells frequently.Studies reported that LY294002 or Wortmannin could reverse multidrug resistance mediated by P-glycoprotein(P-gp)or multidrug resistance associated proteins(MRPs).The present study was to investigate the reversing effects and mechanisms of two PI3K inhibitors 2-(4-Morpholinyl)-8-phenylchromone(LY294002)and wortmannin on human breast cancer protein mediated resistance to mitoxantrone in a mitoxantrone resistant cell line MCF-7/MXT.MCF-7/MXT cells were selected in medium with MXT.The effects of PI3K on breast cancer cells were detected by several technologies. After being treated with mitoxantrone(MXT),wortmannin and/or LY294002(LY),cellular viability and proliferation were measured using the MTT assay and morphological changes were recorded by microscopy.Intracellular accumulation and export of MXT in MCF-7/MXT cells,mitochondrial membrane potential and distribution of cell cycle were detected by flow cytometry.Expression and activity of some proteins associated with apoptosis were measured by Western blots.The results showed that wortmannin had no effect on MCF-7/MXT cells survival to mitoxantrone.The accumulation and efflux of MXT in MCF-7/MXT cells,the mitochondrial membrane potential and the distribution of cell cycle of MCF-7/MXT cells were not effected by wortmannin.While LY reduced the IC50 of MCF-7/MXT cells to MXT about 35.4-fold.MXT accumulation in MCF-7/MXT cells increased about 14-fold when cotreated with LY for 2h.MXT efflux was reduced about 4-fold when cotreated with LY for 1h.MCF-7/MXT cells were mostly arreast at G2/M phase and slightly at S phase when treated with MXT.LY could induce G0/G1 phase arreast in MCF-7/MXT cells. MCF-7/MXT cells were arreast at G2/M phase and S phase significantly when cotreated with MXT and LY.LY or MXT had no effects on the mitochondrial membrane potential of MCF-7/MXT cells when treated for 24h.While the mitochondrial membrane potential of MCF-7/MXT cells reduced significantly when cotreated with LY and MXT for 24h.LY could reduce the activity of Akt.The activity of caspase 8,9,3 and 7 and PARP increased when cotreated with LY and MXT.The expression of BCL-2 reduced significantly when treated with LY or MXT.We conclude that the PI3K inhibitor LY can overcome MCF-7/MXT cells resistance to mitoxantrone and the mechanism underlined might be that LY induced the inhibition of PI3K/Akt pathway and the function of BCRP,which induced apoptosis through both death receptor-dependent(extrinsic)and independent(intrinsic or mitochondrial)pathway. Wortmannin,another PI3K inhibitor,had no effects on MCF-7/MXT cells survival to mitoxantrone,which may because the half-life period of wortmannin is about 4-6h in solution.And may because wortmannin can not induce the inhibition of the function of BCRP.LY294002 can overcome MCF-7/MXT cells resistance to mitoxantrone may also associated with other functions of LY in cells,such as induce the inhibition of the functions of calcium,kalium ion channel,DNA-dependent protein kinase and calcium protein kinase. This study suggests that combination of LY294002 and MXT may provide an effective approach to reverse breast cancer resistance protein(BCRP)-mediated multidrug resistance. |
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