Investigation On The Enhanced Inhibitory Effect Of Cardamonin And Its Mechanism On Paclitaxel-Resistant Ovarian Cancer Cells

AimTo clarify the effect of cardamonin（CAR）to enhance the anti-proliferation of paclitaxel（PTX）on PTX-resistant ovarian cancer cells（SKOV3/Taxol）and explore the underling mechanism,which is providing a novel drug chioce for the treatment of PTX resistant ovarian cancer.Methods1 Cell cultureThe normal human ovarian cancer SKOV3 and the PTX resistant ovarian cancer SKOV3-taxol cell lines were cultured in McCoy’s 5A and high glucose DMEM medium with 10%FBS,100 ng·mL^-1 streptomycin and 100 U·mL^-1 penicillin at 37℃with 5%CO₂,respectively,20 ng·mL^-1 paclitaxel was added into the high glucose DMEM medium to maintain drug resistance.PTX was deprived for one week before the experiment,and the culture was detoxified.2 Groups and drugs intervention2.1 Improving PTX resistance groups:Cells were devided into the following groups:control group,CAR groups（5μM,10μM,20μM）,PTX group（100 nM）,CAR+PTX groups（5μM,10μM,20μM+100 nM）and VER+PTX groups（20μM+100 nM）.2.2 Exploring mechanism groups:Cells were devided into the following groups:control group,CAR groups（5μM,10μM,20μM）,VER group（20μM）.3 Measurements and methodology3.1 Cell viability and proliferation was measured by CCK8 and EdU;3.2 Cell apoptosis and cell cycle was detected by flow cytometry;3.3 Rhodamine-123 cumulative test was used to test the inhibition ability of CAR on p-gp efflux pump;3.4 The expression of MDR1 mRNA was detected by qRT-PCR;3.5 Western blotting was used to detect the expression of P-gp,NF-κB p65（total,nuclear）,IкB,p-IкB（Ser32）,Akt,p-Akt（Ser473）.Results1 CAR combined with PTX had a synergistic anti-proliferation effect on the SKOV3/-Taxol cells,and the Combination Index（CI）was less than 1;2 CAR enhanced the apoptotic effect of PTX on the SKOV3-Taxol cells in a concentration-dependent way.（P<0.05）;3 CAR enhanced the cell cycle arrest effect of PTX on the SKOV3-Taxol cells,the cell cycle was stagnated in G2/M phase significantly in a concentration-dependent way after PTX combined with cardamonin.（P<0.05）;4 CAR significantly increased drug accumulation in the SKOV3-Taxol cells in a concentration-dependent way.（P<0.05）;5 CAR reduced the expression of MDR1 mRNA and P-gp in a concentration-dependent way.（P<0.05）;6 CAR inhibited the activity of NF-κB signaling（P<0.05）.Combined with the NF-κB inhibitor PDTC,cardamonin further inhibited the activity of NF-κB and the protein expression of P-gp;CAR siginificantly inhibited the NF-κB stimulator TNF-αinduced upregulation of the activity of NF-κB and the protein expression of P-gp.（P<0.05）.ConclusionsCAR enhanced the effect of PTX on cell anti-proliferation,inducing cell apoptosis and cell cycle arrest,thereby enhanced the inhibition of PTX on the SKOV3-Taxol cells.The mechanism might be that CAR inhibited the efflux function of P-gp and reduced the mRNA and protein expression of MDR1/P-gp by inhibiting the NF-κB activity of SKOV3-Taxol cells.