Cellular, transcriptomic and isoform heterogeneity of breast cancer cell line revealed by full-length single-cell RNA sequencing

Tumor heterogeneity is generated through a combination of genetic and epigenetic mechanisms, the latter of which plays an important role in the generation of stem like cells responsible for tumor formation and metastasis. Although the development of single cell transcriptomic technologies holds promise to deconvolute this complexity, a number of these techniques have limitations including drop-out and uneven coverage, which challenge the further delineation of tumor heterogeneity. We adopted deep and full-length single-cell RNA sequencing on Fluidigm's Polaris platform to reveal the cellular, transcriptomic, and isoform heterogeneity of SUM149, a triple negative breast cancer (TNBC) cell line. We first validate the quality of the TNBC sequencing data with the sequencing data from erythroleukemia K562 cell line as control. We next scrutinized well-defined marker genes for cancer stem-like cell to identify different cell populations. We then profile the isoform expression data to investigate the heterogeneity of alternative splicing patterns. Though classified as triple-negative breast cancer, the SUM149 stem cells show heterogeneous expression of marker receptors (ER, PR, and HER2) across the cells. We identified three cell populations that express patterns of stemness: epithelial-mesenchymal transition (EMT) cancer stem cells (CSCs), mesenchymal-epithelial transition (MET) CSCs and Dual-EMT-MET CSCs. These cells also manifested a high level of heterogeneity in alternative splicing patterns. For example, CSCs have shown different expression patterns of the CD44v6 exon, as well as different levels of truncated EGFR transcripts, which may suggest different potentials for proliferation and invasion among cancer stem cells. Our study identified features of the landscape of previously underestimated cellular, transcriptomic, and isoform heterogeneity of cancer stem cells in triple-negative breast cancers.