The Study Of Influence Of Cisplatin (DDP) On Lung Cancer Cell Resistant Genes Expression Levels

Objective To study the influence of cisplatin (DDP) on multidrug resistance-associated protein1 ( MRP1 ) , O⁶-methylguanine methytransferase (MGMT) and Xeroderma pigmentosun group A (XPA) mRNA expression levels. To explore effect of multidrug resistance mechanism, enzyme direct repair mechanism and nucleotide excision repair mechanism in lung cancer cells resistance to DDP.Methods Survival rate of lung cancer cell A549 with treatment of different concentration of DDP was measured using MTT metabolic viability assay. Expression levels of MRP1, MGMT and XPA mRNA in lung cancer cell A549 after exposed to different concentrations of DDP were measured by real time fluorescent quantitative reverse-transcription polymerase chain reaction (real time RT-PCR) in different time.Results The survival of lung cancer cell A549 decreased from 74.7% to 14.7% 48h after treatment ranging 1.25-40μg/ml of DDP. There were a correlation between dose of DDP and lung cancer cells survival rate(Correlation analysis: r=-0.930,P<0.01).The change of lung cancer cell survival rate was the most notable in concentration of 2.5-5μg/ml of DDP, the concentration of IC₅₀ is 4.0μg/ml. The cell survival decreased to 96.1%, 73%, 48.5% respectively, 12h, 24h, 48h after treatment with 4μg/ml DDP. Cell survival rate had been declining gradually with the time-dependent (Correlation analysis: r=-0.930, P<0.01)After lung cancer cells were treated with DDP of differentconcentrations, there were significant different expression levels ofMRP1 mRNA ( F=5.76,P<0.05) . MRP1mRNA expression level in 4.0μg/ml (IC₅₀) dose group is lower than that in 0.8μg/ml (1/5 IC₅₀) and 20μg/ml (5 IC₅₀) dose group. In the same dose group, MRP1mRNAexpression level was increased with the extension of DDP processingtime (F = 6.7, P <0.05). In the 4.0μg/ml (IC₅₀) dose group, 12h and 24 hafter DDP treatment, the level of MRP1mRNA expression is lower than before the treatment of DDP.After lung cancer cells were treated with DDP of different concentrations, there were significant different MGMT mRNA expression levels (F=5.76,P<0.05) . The expression levels of MGMT mRNA in groups of different dose DDP: the expression level of MGMT mRNA in 4μg/ml (IC₅₀) group was the lowest; the expression level of MGMT mRNA in 20μg/ml (5 IC₅₀) group was the highest. Comparison of different time after treatment of DDP: there were different MGMT mRNA expression levels with extension of DDP treatment time (F=6.76,P<0.01), of which expression levels below before DDP treatment in 4μg/ml (IC₅₀) group; and decreased at first in 0.8μg/ml (1/5 IC₅₀) group, then increased; however, in 20μg/ml (5 IC₅₀) group had a persistent increase.After lung cancer cells were treated with DDP of different concentrations, the expression levels of XPA mRNA were significant different statistically (F=9.78, P<0.01). The expression level of XPA mRNA in three dose groups increased altogether after DDP treatment. But the expression level of XPA mRNA in 4μg/ml (IC₅₀) group was lower than that in 0.8μg/ml (1/5 IC₅₀) group and 20μg/ml (5 IC₅₀) group. The level of expression in 20μg/ml (5 IC₅₀) was the highest. There are different expression levels of XPA mRNA with different time of DDP treatment. In three dose groups, expression levels of XPA mRNA significant increased with extension of DDP treatment time (F=8.9, P<0.01).Conclusions DDP can lead to lung cancer cell A549 drug-resistance associated gene MRP1,MGMT and XPA mRNA expression levels change with different dose DDP treatment in different time. Results of this study indicate that multidrug resistant mechanisms, enzyme direct repair mechanism and nucleotide excision repair mechanism play an important role in development of drug-resistance in lung cancer cell to DDP.