Resistance to endocrine therapy in breast cancer is associated with poor prognosis. Cell models of acquired tamoxifen resistance have implicated altered growth factor receptor signalling, especially the ErbB family of receptor tyrosine kinases, in development of the accompanying aggressive phenotype. Microarray analysis of an in vitro wtMCF-7 based model of acquired tamoxifen resistance (‘Tam-R’) identified upregulation of CD44, a transmembrane glycoprotein, known to interact with ErbB receptors and influence breast cancer progression. We investigated the hypothesis that CD44 overexpression in Tam-R cells can modulate ErbB activity and promote an adverse phenotype. CD44 gene overexpression was validated by RT-PCR and its protein expression determined by Western blotting and immunocytochemistry. CD44 contribution to intracellular signalling and phenotype of Tam-R cells (migration, invasion and growth), both endogenous and in response to hyaluronan (HA), was determined using Western blotting, immunocytochemistry and functional assays including wound healing, Boyden chamber migration, Matrigel™ invasion and growth assays in the presence or absence of siRNA-mediated CD44 knockdown. Interactions between CD44 and ErbB receptors were investigated using immunofluorescence and immunoprecipitation. CD44 was overexpressed at gene and protein level in Tam-R versus wtMCF-7 cells and whilst this did not influence the endogenous phenotype of Tam-R cells, it enhanced their sensitivity to HA as evidenced by HA-induced MAPK, EGFR and HER2 activation and increased migration. HA-induced migration was attenuated following treatment with the MAPK inhibitor PD098059, gefitinib as well as trastuzumab. CD44 was found to associate with HER2 and HER3 at the cell surface whilst HA stimulation appeared to modulate ErbB dimerisation patterns. Our data suggest that CD44 overexpression sensitises tamoxifen-resistant cells to HA thereby modulating ErbB dimerisation and enhancing migration. These observations may have importance in vivo where the tumour microenvironment can provide a rich source of HA to promote the progression of tamoxifen-resistant tumours.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:585323 |
| Date | January 2013 |
| Creators | Baruah, Bedanta Prakash |
| Publisher | Cardiff University |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://orca.cf.ac.uk/53654/ |
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