The Effects Of The Epigenetic Treatment To Nonsmall-cell Lung Cancer On The Expression Of DNMT1

Objective:To explore the characteristics and biological functions of the epigetics treatment to DNMT1expressions in1NSCLC.Methods:This research is divided into two parts. Part I:research of the effects and function mechanism of the epigenetic treatment to the NSCLC DNMT1. l.The effects of the epigenetic treatment on DNMT1expressions in NSCLC. Lung cancer cells are divided into control group (not using medicine),5-AZA-CDR treated group (1to3days),MS275treated group (the fourth day) and mixed treated group (first three days,5-AZA-CDR; the fourth day, MS275), and cells will be collected, protein will be extracted after treatment, and DNMT1expressions will be evaluated through Western blotting;2.The epigenetic treatment suppresses the function mechanism of DNMT1expressions. According to the therapeutic schedule of Part One, the cell samples will be collected, total RNA will be extracted, cDNA will be conducted and then real-time fluorogenic quantitative PCR will be evaluated. Part Two:research of the drug concentration of the epigenetic treatment drugs and changes of DNMT1expressions in lung cancer cells after the epigenetic treatment.1. Research of the best drug concentration of the epigenetic treatment to lung cancer. Lung cancer cells are divided into control group (not using drug),5-AZA-CDR treated group (1to3days), MS275treated group (the fourth day) and mixed treated group (first three days,5-AZA-CDR; the fourth day, MS275). Then add in drugs with different concentrations, and explore the best drug concentration of curing lung cancer cells through MTT and Western blotting.2. Changes of DNMT1expression in lung cancer cells after the epigenetic treatment. Collect lung cancer cells in different time periods after drug withdrawl, evaluate them through Western blotting, and test the changes of DNMT1expressions in lung cancer cells after drug withdrawl.Results:Part I:1. After the epigenetic treatment, the level of DNMT1expression in the treated group is less than the untreated group, the combined treated group has more effects than the5-AZA-CDR treated group does, and the5-AZA-CDR treated group has more obvious effects than the MS-275treated group.2. Prove that the reduction of DNMT1production is the main cause of its reduced expressions in cells through real-time fluorogenic quantitative PCR. In Part II, it is found that too little drug concentration of the epigenetic treatment drugs has unapparent effects on suppressing DNMT1, but too much drug concentration will produce cytotoxicity; the epigenetic treatment drugs have the strongest suppressing function to DNMT1in lung cancer cells when the epigenetic treatment is over, however, with the extension of drug withdrawl time, the level of DNMT1expressions will recover a little; and different cell lines have different sensitivity to the epigenetic treatment.Conclusions1. The epigenetic treatment can effectively suppress DNMT1expression in NSCLC and control the increase of lung cancer cells to some extent and even induce the lung cancer cells to death. Compared with histone deacetylase inhibitor MS-275, DNA methylation transferases inhibitor5-AZA-CDR has more direct and effective suppressing function to DNMT1genes.Combined drug use lead to better effects.2. Low dose drug concentration of the epigenetic treatment drugs has apparent effects on suppressing DNMT1, and different cell lines have different sensitivity to the epigenetic treatment.3. After the epigenetic treatment is over, with the extension of drug withdrawl time, the level of DNMT1expression recovers gradually, it is further clarified that the epigenetic can be reversed.