Abstract
NGS (Next Generation Sequencing) technologies allows us to determine key gene expression signatures that correlate with resistance (and responsiveness) to anti-cancer therapeutics. We have undertaken a transcriptomic and chromatin immunoprecipitation followed by sequencing (ChIP-seq) approach to describe differences in gene expression and the underlying chromatin landscape between two representative HER2+ cell lines, one of which is sensitive (SKBR3) and the other which is resistant (JIMT1) to trastuzumab. We identified differentially expressed genes (DEGs) and differentially expressed transcripts (DETs) between SKBR3 and JIMT1 cells. Several of the DEGs are components of the Polycomb Repressing Complex 2 (PRC2), and they are expressed higher in JIMT1 cells. In addition, we utilized ChIP-seq to identify H3K18ac, H3K27ac and H3K27me3 histone modifications genome-wide. We identified key differences of H3K18ac and H3K27ac enrichment in regulatory regions, found a correlation between these modifications and differential gene expression and identified a transcription factor binding motif for LRF near these modifications in both cell lines. Lastly, we found a small subset of genes that contain repressive H3K27me3 marks near the gene body in SKBR3 cells but are absent in JIMT1. Taken together, our data suggests that differential gene expression and trastuzumab responsiveness in JIMT1 and SKBR3 is determined by epigenetic mechanisms.
Highlights
HER2-positive (HER2+) breast cancer accounts for 20–25% of all breast cancers[1]
We determined that there were ~3200 differentially expressed genes (DEGs) and ~2800 differentially expressed transcripts (DETs) that were statistically significant (p-value < 0.05) between JIMT1 and SKBR3 cells (Fig. 1a). This greatly expands the number of DEGs and DETs ever reported between JIMT1 and SKBR3 cells[12,13]
By conducting Gene ontology (GO) analysis of the top-50 DEGs in both cell lines, we determined that the top DEGs in JIMT1 are involved in motility and migration (Fig. 1c)
Summary
HER2-positive (HER2+) breast cancer accounts for 20–25% of all breast cancers[1]. Prior to the clinical approval of trastuzumab, patients diagnosed with HER2+ breast cancer exhibited the worst prognosis and highest mortality[2]. Some groups have conducted comparisons between SKBR3 and JIMT1 cells and have used systems biology approach[12] which uses established sub-pathway identification and network permutation method They identified 32 upregulated KEGG sub-pathway genes that were common to trastuzumab resistant cells versus trastuzumab sensitive cells. Another excellent review byMartin-Castillo et al.[13] suggest interesting roles for Cancer Stem Cell (CSC) and non-Cancer Stem Cells (non-CSC) within HER2-overexpressing breast carcinomas. The authors have discussed extensively the biological significance of CSC features and the EMT on the molecular effects and efficacy of trastuzumab in HER2-positive breast cancer cells They have focused on the genetic heterogeneity that differentiates trastuzumab-responders from non-responders in terms of CSC cellular states. We conducted a transcriptomic and ChIP-seq interrogation of JIMT1 and SKBR3 cells to determine differential gene expression and epigenomic differences
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