s observed using cell lines we observed higher expression of LEF1, suggesting WNT activation in Luminal, ER+ve breast cancer patients. A number of genes were differentially expressed between Luminal and Basal/ErbB2 breast cancer, such as WNT5B, WNT10B and DKK1. A number of genes were highly expressed specifically within the basal subtype of breast cancer, LRP5, LRP6, and sFRP1. Analysis of these genes showed significant associations to recurrence free survival within breast cancer patients. The most significant association was with sFRP1, where decreased expression predicts early recurrence in ER+ve breast cancer, most likely due to lack of Wnt inhibition. Lower levels of Lef1 were also predictive of recurrence in breast cancer within the ER+ve subgroup. It might be expected that higher levels of Lef1 would correlate with recurrence, however lower Lef1 expression may identify a subgroup of ER+ve SAR 405 site tumours that have a basal-type phenotype, based upon their Wnt gene expression. This subgroup within ER+ve tumours can also be observed with analysis of Wnt5b, where high expression predicts early recurrence, and where high expression is a marker of the basal-type phenotype. LRP5 and LRP6 although demonstrating a very similar expression profile within breast tumours have very different effects on recurrence. Low expression of LRP6 in ER+ve tumours was predictive of early recurrence, whilst low levels of LRP5 was predictive of early recurrence in ER-ve breast cancer. LRP5/6 are co-receptors for WNT ligands, previously thought to act similarly to activate downstream signalling. Recent research suggests that they may actually function differently, dependant upon physiological conditions and the type and availability of WNT ligand. Further investigations will be needed to determine their precise roles in breast cancer. for each cell line were calculated and displayed as a heatmap. Importantly, analysis of the change in gene expression between monolayer 
cultured 21836025 and AR cells showed differences between breast cancer and normal cell lines. WNT ligands WNT1, WNT10a and WNT4 were expressed at higher levels in normal AR cells compared to monolayer cells, however in the breast cancer cell lines, their expression was lower in the AR cells compared to monolayer cells. sFRP1 and b-catenin were highly expressed in BCSCs from ER+ve breast cancer cell lines. No increase in expression was observed in the ER-ve breast cancer cell line tested suggesting that canonical WNT signalling in BCSCs may correlate with 22038495 ER. Most importantly, we found higher levels of the WNT signalling gene LEF1 in BCSCs compared to monolayer cells, irrespective of ER expression, while normal breast stem cell populations showed lower levels compared to normal monolayer cells. The data demonstrates specific upregulation of WNT pathway signalling in a population enriched for BCSCs. Modulation of Wnt Pathway Signalling Affects Stem Celllike Activity To test the potential of targeting the WNT pathway we investigated the effects of WNT pathway modulation in cell lines. Cells were cultured as mammospheres with increasing concentrations of either DKK1 to inhibit or WNT3A to activate the WNT pathway. Recombinant WNT3A treatment significantly increased the number of MS in both normal and breast cancer cell lines. DKK1 treatment significantly decreased the number of MS in all cell lines tested with the most significant decrease observed using the highest concentration of DKK1. Using patient-derived ce