Single-cell RNA sequencing identifies extracellular matrix gene expression by pancreatic circulating tumor cells.
Bottom Line: To define their composition, we compared genome-wide expression profiles of CTCs with matched primary tumors in a mouse model of pancreatic cancer, isolating individual CTCs using epitope-independent microfluidic capture, followed by single-cell RNA sequencing.Mouse as well as human pancreatic CTCs exhibit a very high expression of stromal-derived extracellular matrix (ECM) proteins, including SPARC, whose knockdown in cancer cells suppresses cell migration and invasiveness.The aberrant expression by CTCs of stromal ECM genes points to their contribution of microenvironmental signals for the spread of cancer to distant organs.
Affiliation: Massachusetts General Hospital Cancer Center, Harvard Medical School, Boston, MA 02114, USA; Department of Medicine, Harvard Medical School, Boston, MA 02114, USA.Show MeSH
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Mentions: CTCs are also likely to be enriched for metastatic precursors capable of initiating metastatic tumor deposits. The relationship between such precursor cells and cancer stem cells is uncertain, as is the relevance of established stem cell markers in identifying these cells. We evaluated putative pancreatic cancer stem cell genes (Rasheed and Matsui, 2012; Rasheed et al., 2010) in the single-cell RNA-seq reads (Figures 4A and S6). Among all candidate markers tested (Aldh1a1, Aldh1a2, Prom1/Cd133, Cd44, Met, EpCAM), only Aldh1a1 and Aldh1a2 were enriched in CTCs. Classical CTCs expressed predominantly the Aldh1a2 isoform, while Aldh1a1 was expressed in a variety of cell types (Figure S6). Within single CTCs, there was no correlation between expression of Aldh1 isoforms and either enrichment for the mesenchymal genes (Cdh11, Vim) or loss of epithelial genes (Cdh1, Muc1), suggesting that stem cell and EMT markers are not intrinsically linked in CTCs. Analysis of primary pancreatic tumors for Aldh1a2 using RNA in situ hybridization (RNA-ISH) identified rare epithelial tumor cells expressing this stem cell marker, but the majority of expression was present within the cancer associated stromal cells (Figure 4B), consistent with immunohistochemistry for ALDH protein in human PDAC (Rasheed et al., 2010).
Affiliation: Massachusetts General Hospital Cancer Center, Harvard Medical School, Boston, MA 02114, USA; Department of Medicine, Harvard Medical School, Boston, MA 02114, USA.