GSE171429
scRNA-seq of DAC-treated and mock-treated HCT116.
Epigenetic reprogramming using demethylating drugs is a promising approach for cancer therapy, but its efficacy is highly dependent on the dosing regimen.  Low-dose treatment for a prolonged period shows a high therapeutic efficacy, despite its small demethylating effect.  Here, we aimed to reveal the mechanisms of how such low-dose treatment shows high efficacy by focusing on epigenetic reprograming at the single-cell level.  Single-cell RNA-sequencing of HCT116 cells treated with decitabine (DAC) revealed that up-regulated genes were highly variable at the single-cell level.  To analyze functional consequences at the single-cell level, DAC-treated HCT116 cells were cloned.  While only partial reduction of methylation levels was observed in bulk cells, complete demethylation of specific cancer-related genes was observed, depending upon clones.  For example, p16 was completely demethylated in the H3-32 clone out of 9 clones, and this clone showed slower proliferation than other clones without demethylation.  In addition, in this clone, the fraction of cells with tetraploid became much larger, indicating that cellular senescence was induced.  These results showed that epigenetic reprogramming of specific cancer-related pathways at the single-cell level is likely to underlie the high efficacy of low-dose DNA demethylating therapy. Overall design: scRNA-seq was conducted for two samples, DAC-treated HCT116 cells and mock-treated HCT116 cells, using the Chromium Controller (10x Genomics ) and Illumina HiSeq platform.