CD44-Activated HER-2 Signalling in Tamoxifen Resistant Breast Cancer Cells Promotes a Migratory Phenotype

Background: Acquired endocrine resistance in breast cancer cells is accompanied by altered growth factor receptor signalling. In tamoxifen-resistant (9TamR9) MCF7 cells, our microarray analysis has demonstrated elevated levels of CD44, a transmembrane glycoprotein known to interact with, and modulate the function of, a number of growth factor receptors including members of the erbB family and plays a major role in tumor metastasis. Here, we have explored the role of CD44 as a modulator of erbB activity in TamR cells. Methods: Western blotting and immunocytochemistry were used to investigate expression of CD44 together with the activity of erbB signalling pathways following stimulation of CD44 by its ligand, hyaluronan (HA), or stimulation with erbB ligands (heregulin β1) after knockdown of CD44 expression using siRNA. Immunofluorescence microscopy was used to determine association between CD44 and erbB family members. Cell migration was assessed by seeding cells onto fibronectin-coated microporous inserts for 24 hours and also by performing scratch wound assays. Cell proliferation was determined by WST assay in response to trastuzumab (0-100nM) ± HA Results: CD44 was found to be over-expressed in TamR cells where it co-localized with HER-2. Whilst stimulation of HER-2 with heregulin beta-1 promoted an increase in cell migration, heregulin action was significantly attenuated in the absence of CD44 using siRNA. Moreover, treatment of TamR cells with the natural CD44 ligand, HA, promoted HER-2 signalling through MAPK and an increase in cell migration. Furthermore, whilst TamR cells were sensitive to the growth-inhibitory effects of trastuzumab, these effects were diminished in the presence of HA. Conclusions: Overexpression of CD44 sensitizes tamoxifen-resistant cells to heregulin and HA, factors commonly found within the stromal environment, with a resultant increase in migratory behaviour. Interestingly, activation of CD44 by HA appears to suppress the effects of trastuzumab on cell proliferation. Thus, CD44 may prove a valuable therapeutic target in breast cancer that expresses HER-2 and CD44. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5137.