Mouse mammary stem cells express prognostic markers for triple-negative breast cancer.

INTRODUCTION: Triple-negative breast cancer (TNBC) is a heterogeneous group of tumours in which chemotherapy, the current mainstay of systemic treatment, is often initially beneficial but with a high risk of relapse and metastasis. There is currently no means of predicting which TNBC will relapse. We tested the hypothesis that the biological properties of normal stem cells are re-activated in tumour metastasis and that, therefore, the activation of normal mammary stem cell-associated gene sets in primary TNBC would be highly prognostic for relapse and metastasis. METHODS: Mammary basal stem and myoepithelial cells were isolated by flow cytometry and tested in low-dose transplant assays. Gene expression microarrays were used to establish expression profiles of the stem and myoepithelial populations; these were compared to each other and to our previously established mammary epithelial gene expression profiles. Stem cell genes were classified by Gene Ontology (GO) analysis and the expression of a subset analysed in the stem cell population at single cell resolution. Activation of stem cell genes was interrogated across different breast cancer cohorts and within specific subtypes and tested for clinical prognostic power. RESULTS: A set of 323 genes was identified that was expressed significantly more highly in the purified basal stem cells compared to all other cells of the mammary epithelium. A total of 109 out of 323 genes had been associated with stem cell features in at least one other study in addition to our own, providing further support for their involvement in the biology of this cell type. GO analysis demonstrated an enrichment of these genes for an association with cell migration, cytoskeletal regulation and tissue morphogenesis, consistent with a role in invasion and metastasis. Single cell resolution analysis showed that individual cells co-expressed both epithelial- and mesenchymal-associated genes/proteins. Most strikingly, we demonstrated that strong activity of this stem cell gene set in TNBCs identified those tumours most likely to rapidly progress to metastasis. CONCLUSIONS: Our findings support the hypothesis that the biological properties of normal stem cells are drivers of metastasis and that these properties can be used to stratify patients with a highly heterogeneous disease such as TNBC.

Developmental stage-specific contribution of LGR5(+) cells to basal and luminal epithelial lineages in the postnatal mammary gland.

The leucine-rich repeat-containing heterotrimeric guanine nucleotide-binding protein-coupled receptor 5 (LGR5) has been identified as a marker of cycling stem cells in several epithelial tissues, including small intestine, colon, stomach and hair follicle. To investigate whether LGR5 also marks mammary epithelial stem cells, we performed in situ lineage-tracing studies and mammary gland reconstitutions with LGR5-expressing mammary epithelial cells. Interestingly, the LGR5 progeny population in mammary epithelium switches from the luminal to the myoepithelial compartment during the first 12 days of postnatal development, likely reflecting local changes in Wnt signalling. Together, our findings point to a stage-specific contribution of LGR5-expressing cells to luminal and basal epithelial lineages during postnatal mammary gland development.

Intestinal stem cells and their defining niche.

The intestinal epithelium is a classic example of a rapidly self-renewing tissue fueled by dedicated resident stem cells. These stem cells reside at the crypt base, generating committed progeny that mature into the various functional epithelial lineages while following a rapid migratory path toward the villi. Two models of intestinal stem cell location were proposed half a century ago and data have been presented in support of both models, dividing the scientific community. Molecular markers have been identified and validated using new techniques such as in vivo lineage tracing and ex vivo organoid culture. The intestinal stem cell niche comprises both epithelial cells, in particular the Paneth cell, and the stromal compartment, where cell-associated ligands and soluble factors regulate stem cell behavior. This review highlights the recent advances in identifying and characterizing the intestinal stem cells and their defining niche.

FRMD4A upregulation in human squamous cell carcinoma promotes tumor growth and metastasis and is associated with poor prognosis.

New therapeutic strategies are needed to improve treatment of head and neck squamous cell carcinoma (HNSCC), an aggressive tumor with poor survival rates. FRMD4A is a human epidermal stem cell marker implicated previously in epithelial polarity that is upregulated in SCC cells. Here, we report that FRMD4A upregulation occurs in primary human HNSCCs where high expression levels correlate with increased risks of relapse. FRMD4A silencing decreased growth and metastasis of human SCC xenografts in skin and tongue, reduced SCC proliferation and intercellular adhesion, and stimulated caspase-3 activity and expression of terminal differentiation markers. Notably, FRMD4A attenuation caused nuclear accumulation of YAP, suggesting a potential role for FRMD4A in Hippo signaling. Treatment with the HSP90 inhibitor 17-DMAG or ligation of CD44 with hyaluronan caused nuclear depletion of FRMD4A, nuclear accumulation of YAP and reduced SCC growth and metastasis. Together, our findings suggest FRMD4A as a novel candidate therapeutic target in HNSCC based on the key role in metastatic growth we have identified.

Actin and serum response factor transduce physical cues from the microenvironment to regulate epidermal stem cell fate decisions.

Epidermal homeostasis depends on a balance between stem cell renewal and differentiation and is regulated by extrinsic signals from the extracellular matrix (ECM). A powerful approach to analysing the pathways involved is to engineer single-cell microenvironments in which individual variables are precisely and quantitatively controlled. Here, we employ micropatterned surfaces to identify the signalling pathways by which restricted ECM contact triggers human epidermal stem cells to initiate terminal differentiation. On small (20 microm diameter) circular islands, keratinocytes remained rounded, and differentiated at higher frequency than cells that could spread on large (50 microm diameter) islands. Differentiation did not depend on ECM composition or density. Rather, the actin cytoskeleton mediated shape-induced differentiation by regulating serum response factor (SRF) transcriptional activity. Knockdown of SRF or its co-factor MAL inhibited differentiation, whereas overexpression of MAL stimulated SRF activity and involucrin expression. SRF target genes FOS and JUNB were also required for differentiation: c-Fos mediated serum responsiveness, whereas JunB was regulated by actin and MAL. Our findings demonstrate how biophysical cues are transduced into transcriptional responses that determine epidermal cell fate.