Abstract
Abstract The analysis of average gene expression in cell populations may obscure the heterogeneity of rare cells such as cancer stem cells (CSCs). The use of CSC markers to enrich for these cell populations has proven valuable in their molecular characterization. However, conventional gene expression analysis of these CSC enriched populations is still not conducive to elucidating their heterogeneity. We exploited single-cell resolution methods using C1 and BioMark HD platform and TaqMan assays in the analysis of CSCs and non-CSCs/differentiated tumor cells. The expression of 96 target genes was studied at single cell level in a multiplex RT-qPCR setting. Breast cancer cell lines including MCF7 (luminal), BT474 (HER2+), SUM149 (Basal), and MDA-MB-231 (Basal-Claudin low) that represent major different subtypes of breast cancers and normal mammary epithelial cells (NM) were studied at single cell resolution. Despite similarities, expression profiles of 96 target genes were distinctly different between subtypes of breast cancers and also with that of NM cells. Interestingly, results of single cell studies revealed significant levels of hierarchical heterogeneities within and between each analyzed population of cells. The observed heterogeneous signatures were due to both different expression levels as well as presence/absence of certain genes. We could identify CSCs with EMT (i.e. CD44+/CD24-/ALDH-/CDH2+/Vimentin+), MET (i.e. ALDH+/CD44-/CD24+/CDH1+/Vimentin-), and dual EMT-MET (CD44+/CD24-/ALDH+/CDH1+ or CDH2+/Vimentin+) characteristics. This methodology enabled us to also identify a range of several different non-CSCs/differentiated single cells with heterogeneous expression patterns for CD44, CD24, ALDH, ESR, PGR, HER2, Vimentin, CDH1, CDH2, EpCAM, and Cytokeratins that were similar to luminal, HER2 overexpressed, basal, and basal-Claudin low breast cancer cells. Additionally, NM single cells also showed significant heterogeneity that was similar to the wide spectrum patterns of some single breast cancer cells described above. This study highlights the use of single cell technologies to redefine tumor cell characterization, revealing heterogeneity of single tumor cells with respect to stem cell properties, metastatic and drug resistance characteristics. This will further improve the identification of new markers/pathways that can be used to develop new target therapies for better therapeutic responses in cancer patients. Citation Format: Ebrahim Azizi, Justin Colacino, Shamileh Fouladdel, Max S. Wicha. Molecular subtypes of breast cancers and normal mammary epithelial cells show hierarchical heterogeneity and complexity at single-cell resolution. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1585.
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