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    A systems biology approach to target identification using three-dimensional multi-cellular tumour spheroids (MCTS). Regio-specific molecular dissection of gene expression, protein expression and functional activity in 3D MCTS.

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    PhD thesis- Kelly McMahon.pdf (3.807Mb)
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    Publication date
    2013-11-27
    Author
    McMahon, Kelly M.
    Supervisor
    Sutton, Chris W.
    Phillips, Roger M.
    Keyword
    Proteomics
    Transcriptomics
    Multi-Cellular Tumour Spheroids (MCTS)
    Autophagy
    Metabolism
    Hypoxia
    Solid tumours
    Target identification
    Rights
    Creative Commons License
    The University of Bradford theses are licenced under a Creative Commons Licence.
    Institution
    University of Bradford
    Department
    Institute of Cancer Therapeutics
    Awarded
    2011
    
    Metadata
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    Abstract
    Within solid tumours, a microenvironment exists that causes resistance to chemotherapy. New drugs that target cells within this microenvironment are required, the first step in this process being the identification of new targets. The aim of this thesis was to characterise changes in the transcriptome and proteome within specific regions of multicell-tumour spheroids (MCTS), an experimental model that mimics many of the features of the tumour microenvironment. HT29 MCTS were separated by sequential trypsinisation into 3 main regions; the outer surface layer (SL) the peri-necroric region (PN) and the necrotic core (NC). Using an iTRAQ quantitative proteomics approach, the proteome of the different MCTS regions was investigated. A 2 dimensional separation approach using Agilent¿s OffGel system and RP-nano HPLC was incorporated prior to MS analysis. MS analysis was done using both MALDI-TOF-TOF (Bruker Ultraflex II) and ESI-Q-TOF (Agilent 6530 QTOF LC/MS) instruments. Gene expression profiles of the different MCTS were investigated and compared using Agilent¿s one-color oligonucleotide based microarrays. Transcriptomic and proteomic analysis identified several key differences in the proteins involved in cell metabolism between the SL and PN/NC regions. Similar metabolic changes were also noted between autophagic and normal monolayer cells. Many highlighted proteins represented established cancer associated proteins. Interestingly, a number of proteins were highlighted which have no previous association with cancer and may upon further validation, provide attractive leads for therapeutic intervention.
    URI
    http://hdl.handle.net/10454/5722
    Type
    Thesis
    Qualification name
    PhD
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    Theses

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