The Study Of Treatment By Anti-programmed Death Ligand 1 Combining With 5-azacytidine To Esophageal Cancer KYSE450 Cell Line

Objective: We used low doses of demathylation drugs of anti-programmed death ligand 1 combining with 5-Azacy tidine to act on esophageal cancer KYSE450 cell line for the treatm ent of Epigenetics immunotherapy, and then detected the relationship bet ween the e xpression of DNA methyltransferases, PD- L1 and the growth stat e in esophageal cancer KYSE450 cell line. W e aimed to explore the im pact of low doses of 5-Azacytidine which was demethylation drug to the expression of PD- L1 protein in esophageal cancer KYSE450 cell line, and the relationshi p between the expression of DNMTs and the growth status of the esophageal cancer line.Methods: Western blotting was used to detect expression of PD- L1 in the esophageal cancer KYSE450 cell line with the purpose of observing the im pact of 5-Azacytidine disposed by the dr ug dose of 1000nM/L. T umor specific lymphocytes and esophageal cancer KYSE450 cell line were induced compounded, and the com bined effect of 5-Azacytidine and PD- L1 in vitrol was analyzed. Methylation Specific Polymerase Chain Reaction was utilized to explore the expression of PD- L1 a nd 5-Azacytidine. Western blotting was carried out to validate the ex pressive change of PD- L1 and 5-Azacytidine which was untreated and treated w ith the drugs. Quantitative Real time Polymerase Chain Reaction was applied for m RNA expression of DNMT1 and PD-L1 after treated by the drug. ELISA was implem ented to check up the secretion of PD-L1 which was secreted out side of the cells after treated by the drug. Cell scratch test was put into us e to identify the invasion ability of the esophageal cancer KYSE450 cell line.Results: 1. The protein expression of PD-L1 increased after 72 h treatment of 5-Aza in the esophageal cancer KYSE450 cell line. 2. Th e expression of DNMT1 a nd PD-L1 in the treated group was lower than the untreated group. 3. The gene and prot ein expression of DNMT1 and PD-L1 in the treated group was lower than in the untreated group, presenting a descending tendency. 4. In the test of QR T- PCR, the mRNA expression of DNMT1 and PD-L1 in the group of 5-Aza treatment was higher than the control group, what,s more, the anti- PD-L1 treatment group and combined drugs treatment group were also higher than the control group. 5. The supernatant liquid of cells in t he treatment groups was less than the control group(P<0.05).Conclusions: 1. The proliferation and in vasion of esophageal cancer KYSE450 cell with mixed treatment of 5-Aza and anti- PD-L1 was significantly inhibited compared to only a single dr ug. 2. There was a close relationshi p between the expression of PD-L1 on the le vels of protein, mRNA, pro moter of the gene and the invasion, protein secre tion, metastasis. 3. By the ef fective immunotherapy, the anti- PD-L1 could induce non-DNA structure changes making the cells presenting demathylati on, which can lead to reprogram the genomic or increase the activity and the function of the esophageal cancer KYSE450 cell line. 4. It was also detect ed that anti- PD-L1 com bined 5-Aza could play a regulatory role i n the DNA demethylation in T lymphocytes, and could also accelerate the cell secretion.