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The investigation of Rab25 protein expression in early breast cancer

Kelly, Claire (2011) The investigation of Rab25 protein expression in early breast cancer. MD thesis, University of Glasgow.

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Abstract

More than a million women worldwide are diagnosed with breast cancer every year and breast cancer is the most common cancer in women in the UK. Despite improvements in the treatment of breast cancer, it remains the second most common cause of death from cancer in women after lung. Research into the aberrant molecular pathways which characterise neoplasia is necessary to highlight potential therapeutic targets with the aim of improving the survival of women with breast cancer. Rab25 is a member of the Rab superfamily of small GTPases, which are involved in the regulation of intracellular vesicular trafficking. A number of studies have suggested that dysregulation of Rab25 protein expression may be implicated in breast, ovarian and colorectal cancer. However, the published data are conflicting and further research is required to improve the understanding of the role of Rab25 in neoplasia. At the commencement of this project, there was no commercially available anti-Rab25 antibody. The data presented here describe the affinity purification and validation of a rabbit polyclonal anti-Rab25 antibody suitable for use in Western blotting, immunofluorescence and immunohistochemistry. Furthermore, an immunohistochemical scoring system was devised and validated prior to the investigation of Rab25 expression in a large breast cancer cohort. The data presented here show that loss of Rab25 expression correlates with decreased breast cancer related survival of patients who have tumours which have also lost the expression of the oestrogen receptor. Furthermore, loss of Rab25 expression has maximal negative effect on the survival of patients with tumours which have lost both oestrogen and HER2 receptor expression. To investigate the effect of Rab25 knockdown in vitro, the MCF7 cell line was stably transfected with a short hairpin siRNA vector targeted against Rab25. Knockdown of Rab25 had no demonstrable effect on cellular proliferation or colony formation, but Rab25 knockdown cells can migrate to heal a scratch wound at a greater rate than cells transfected with a non-targeted control vector. Knockdown of Rab25 protein expression increases total surface and intracellular β1 integrin protein levels in MCF7 cells, but this increase is not as a result of increased transcription.

Item Type: Thesis (MD)
Qualification Level: Doctoral
Keywords: Rab25, breast cancer,
Subjects: R Medicine > RC Internal medicine > RC0254 Neoplasms. Tumors. Oncology (including Cancer)
Q Science > Q Science (General)
Colleges/Schools: College of Medical Veterinary and Life Sciences > Institute of Cancer Sciences
Supervisor's Name: Norman, Dr. J. and Evan, Prof. J.
Date of Award: 2011
Depositing User: Dr Claire Kelly
Unique ID: glathesis:2011-2808
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 05 Sep 2011
Last Modified: 10 Dec 2012 14:00
URI: http://theses.gla.ac.uk/id/eprint/2808

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