Conclusion
Our data highlight the likelihood that resistant cells emerge from cancer-initiating cells/cancer stem-like cells and imply that these cells may gain further advantage in growth via epigenetic mechanisms. Illuminating the expression and DNA methylation features of putative cancer-initiating cells/cancer stem cells may suggest novel strategies to overcome tamoxifen resistance.
Methods
To evaluate the contribution of these possible tamoxifen resistance mechanisms, we applied modified DNA methylation-specific digital karyotyping (MMSDK) and digital gene expression (DGE) in combination with massive parallel sequencing to analyze a well-established tamoxifen-resistant cell line model (TAM(R)), consisting of 4 resistant and one parental cell line. Another tamoxifen-resistant cell line model system (LCC1/LCC2) was used to validate the DNA methylation and gene expression
Results
Significant differences were observed in global gene expression and DNA methylation profiles between the parental tamoxifen-sensitive cell line and the 4 tamoxifen-resistant TAM(R) sublines. The 4 TAM(R) cell lines exhibited higher methylation levels as well as an inverse relationship between gene expression and DNA methylation in the promoter regions. A panel of genes, including NRIP1, HECA and FIS1, exhibited lower gene expression in resistant vs. parental cells and concurrent increased promoter CGI methylation in resistant vs. parental cell lines. A major part of the methylation, gene expression, and pathway alterations observed in the TAM(R) model were also present in the LCC1/LCC2 cell line model. More importantly, high expression of SOX2 and alterations of other SOX and E2F gene family members, as well as RB-related pocket protein genes in TAMR highlighted stem cell-associated pathways as being central in the resistant cells and imply that cancer-initiating cells/cancer stem-like cells may be involved in tamoxifen resistance in this model.