Title:
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Bladder carcinogenesis and the biological activity of sulforaphane and iberin
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In this thesis, I sought to explore the biological mechanisms that may underlie the
chemoprotective effects of broccoli consumption towards bladder cancer. In particular, I was
interested in the effects of diet on the transition from superficial bladder cancer to a more
invasive form of this disease. Initially, I characterised global gene expression profiles of
superficial and invasive bladder carcinomas through tissues obtained from the Norfolk and
Norwich University Hospital Tissue Bank. Non hierarchical cluster analyses was used to
identify a suite of genes that characterised these two tumour types. Of particular interest was
variation in genes involved in the synthesis of the extra cellular matrix, such as COL6A1 that
was upregulated in the more invasive tumour type. This was subsequently verified by RT
PCR with independent tissue samples. I then explored how the broccoli isothiocyanates,
sulforaphane and iberin and their N-acetylcysteine conjugates were able to perturb gene
expression in two bladder cell lines that differed in their cancer phenotype. I showed that not
only were these ITCs able to upregulate phase II detoxification genes, which has previously
been reported in other cell lines, but they, and their conjugates, were able to down regulate
the COL61A gene. Studies on the effects of ITCs on gene expression were complemented
with studies on cell migration and invasiveness. I then sought to see if any of the changes in
gene expression observed in the cell lines could be observed in tissues obtained as part of a human intervention study. I designed and executed a small pilot study that enabled bladder
tissue biopsies to be obtained before and after a four day intervention with broccoli. Global
gene expression analyses again suggested alterations in genes determining the extracellular
matrix, such as Tenascin-C. This study demonstrated the proof-of-principle that these types
of intervention studies are possible, but was of an insufficient size to draw definite
conclusions. Finally, I investigated in more details the expression of splice variants of
Tenascin-C in relationship to bladder cancer grade. In conclusion, this study has suggested
that dietary ITCs may perturb genes involved in the extracellular matrix and this may be an
important component of understanding their chemopreventive activities, and has
demonstrated the feasibility of human intervention studies with the analyses of target tissues
as opposed to peripheral biomarkers.
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