The transcirptomic and methylomic changes caused by subtoxic doses of 5-Azacytidine and 5-Aza-2'-deoxycitide
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5-Aza-2'-deoxycytidine (5-Aza-CdR) and 5-Azacytydine (5-Aza-CR) are used to treat myelodysplastic syndrome (MDS) in the clinic. Additionally, a number of clinical trials have shown the efficacy of the drugs in solid tumors. However, the genomic changes after the drug treatments are unknown. With our study, we have shown the genomic changes after the treatment with 5-Aza-CdR and 5-Aza-CR in bladder cell lines while using subtoxic doses of the drugs. For the study, we have used one normal immortalized bladder cell line SV-HUC, and two bladder cancer cell lines T24, and SCaBER. We treated these cells with 100 nM and 1 μM of 5-Aza-CdR and 5-Aza-CR. To identify genomic changes after the treatment with the demethylating agents, we utilize Illumina 450K methylation arrays and RNA sequencing. Methylomic changes after the treatment with 5-Aza-CdR and 5-Aza-CR vary for each cell line. The clustering of the methylation data has shown the different efficacy of the drugs and unique demethylated CpG loci for each treatment. Furthermore, demethylation of the gene bodies and 3'UTRs were higher on average compared with demethylation of the promoter regions in all three-cell lines. Striking was that the 1 μM 5-Aza-CR treatment was more efficient in SV-HUC compared with T24 and SCaBER cell lines. The altered transcriptome of the SV-HUC cells after the drug treatments has shown an upregulation of immune modulatory pathways. Also, the difference in transcription between the two drug concentrations of 5-Aza-CdR was higher compared with the 5-Aza-CR treatments, but demethylation after 1 μM 5-Aza-CR in SV-HUC was almost as efficient as the 100 nM 5-Aza-CdR treatment. To understand the chemical demethylation of DNA and RNA expression, we have combined both data sets. We have shown that the chemical demethylation of promoter regions and gene bodies are in the minority of the differentially expressed genes, significantly increased and differently methylated. In summary, we have shown that demethylation after 5-Aza-CdR and 5-Aza-CR treatments was more prone in the gene bodies and 3'UTRs but the demethylation of the gene body was only for a small number of genes related to increased RNA expression.