The Study Of The Mechanism Of Cisplatin-induced SCLC Acquired Multidrug Resistance

Background and Object:Lung cancer is the most common and one of the highest incidence malignant tumorin the world, it is rising year by year, and, this phenomenon brings extremely seriousthreat to our human health. In all types of lung cancer, small cell lung cancer accounts for20-25%. The treatment of small cell lung cancer is systemic chemotherapy mainly, andcisplatin is an effective and widely used first-line drugs. However, in recent years, theemergence of multidrug resistance brings great challenges to the clinical chemotherapy forpatients with SCLC. In order to make the effectiveness of chemotherapy and extend thepatients’ survival time, it’s essential to clarify the mechanism of MDR. In this experiment,the human SCLC cells H446and its MDR cells H446/CDDP are our research objects, andwe design our experiment from two aspects: one is cell metabolism and another is the stability of the genome. All of these are made to find the MDR cells H446/CDDP’s changein the two aspects, so we can explore the mechanism of the formation of MDR andprovide theoretical and experimental basis for reversing drug resistance.Methods:（1）After the SCLC H446cells and its MDR H446/CDDP cells were treated withthe same concentrations of cisplatin (0.5μg/ml) and different time, the lactic acidconcentration and ROS levels were detected by dry chemical method and laser confocalmicroscopy respectively.（2）Try to use the polymorphic microsatellite loci which cancover the whole genome to amplify the cells H446and its MDR cells H446/CDDP’sgenomic DNA(In our experiment we selected the chromosome1、2、3、4、5、8、9、14、17、20、21’s many sites). The obtained PCR product was purified by polyacrylamidegel electrophoresis and stained with silver nitrate, after that, the results were observed bythe FluorChem E chemiluminescent gel imaging system, the MSI and LOH were analysedby Image-pro Plus software. To the results show positive at the first time, it’s essential torepeat in the same way, in order to ensure the accuracy of experimental results.Results:（1）Within24h, there is no lactate concentration difference between the MDR cellsH446/DDP and its parental cells H446. However, with the time extending, the lactic acidproduction of H446was significantly higher than that of H446/CDDP. In addition, whentreated by cisplatin within35min, the ROS production of H446/CDDP cells decreasedgradually, while that of H446cells increased and reached its highest at15min. After that,the ROS production gradually decreased, accompanied with cell shrinkage.（2）In theMDR cells H446/DDP，we found two LOH sites near3p14.2in chromosome3’s short arm,they are D3S1234and D3S1295. In addition, there are two MSI sites near8p22inchromosome8’s short arm and in chromosome20, they are D8S1145and D20S82,respectively.Conclusion:（1）The results of our experiment reveal that relatively strong anti-oxidative ability might be an important factor in SCLC acquired MDR against the damage of cisplatin.（2）We identified four microsatellite loci mutation in the MDR cells H446/DDP, and thesemutations may be related to the formation of acquired MDR.