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Title: Acquired tamoxifen resistance and promotion of angiogenic responses in breast cancer
Author: Hayes, Edward
ISNI:       0000 0004 2735 0908
Awarding Body: Cardiff University
Current Institution: Cardiff University
Date of Award: 2012
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In the treatment of pre-menopausal women with oestrogen positive (ER+) breast cancer, tamoxifen represents a first line of adjuvant treatment with demonstrable benefits. Despite this, resistance is frequently acquired to tamoxifen with an associated poor prognosis. Breast cancer cell models have revealed the importance of growth factor signalling networks in sustaining growth of endocrine-resistant cancers and, more recently, their ability to promote a highly migratory and invasive phenotype, together with the expression of genes with pro-angiogenic ontology. The potential of endocrine-resistant cells to elicit angiogenic responses, however, remains unknown. Real-time PCR was used to validate results from preliminary Affymetrix-based gene profiling of pro-angiogenic gene expression in endocrine-sensitive MCF7wt cells and their endocrine resistant counterparts. The expression of pro-angiogenic factors in conditioned media (CM) from these cells was assessed by ELISA. The proliferative and migratory effects of conditioned media on vascular endothelial cells (HUVEC and HECV cells), was determined by MTS cell proliferation assay, wound closure assays and Matrigel tubule formation assays. Changes in endothelial cell migration following co-culture with endocrine-resistant cells were examined using Boyden-chamber chemotaxis assays. Growth factor signalling and migration pathway activation in endothelial cells in response to CM was determined by Western blotting. TamR cells were found to express high levels of IL-8 and VEGF at an mRNA level compared with expression in MCF7wt cells. High levels of VEGF protein were also confirmed in the conditioned media from TamR cells versus their endocrine-sensitive counterparts. TamR conditioned media promoted in vivo and ex vivo endothelial cell proliferation, as well as in vitro endothelial cell migration and the formation of tubules to a greater extent than that seen in MCF7wt CM treated cells. TamR conditioned media was found to stimulate VEGFR2 phosphorylation and downstream activation of MAPK and Akt in endothelial cells compared to MCF7wt CM. Pharmacological inhibition of VEGFR2 activity in endothelial cells suppressed TamR-induced endothelial cell proliferation and VEGFR phosphorylation. Further pharmacological manipulation of Src kinase in TamR cells revealed a Src kinase dependent mechanism of VEGF production in these cells. In addition, in vivo examination of TamR xenografts illustrated higher presence of endothelial cells in these tissues than in MCF7wt xenografts. These data suggest acquired tamoxifen resistance is accompanied by development of a Src kinase-dependant pro-angiogenic phenotype which, if recapitulated in vivo, may promote tumour progression. Therapeutic targeting of Src signalling may prove beneficial in such cases.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Q Science (General) ; RC0254 Neoplasms. Tumors. Oncology (including Cancer) ; RM Therapeutics. Pharmacology