@phdthesis{discovery1310148,
            note = {Unpublished},
          school = {UCL (University College London)},
           title = {Whole genome RNA expression profiling for the identification of novel biomarkers in the diagnosis and prognosis of biliary tract cancer},
            year = {2011},
           month = {April},
          author = {Chapman, M. H.},
             url = {https://discovery.ucl.ac.uk/id/eprint/1310148/},
        abstract = {Biliary tract cancer (BTC) is difficult to diagnose, in part related to the lack of
reliable tumour markers. The aim of this project was to use whole genome
RNA expression profiling in order to identify novel biomarkers for diagnosis
and prognosis in biliary tract cancer.
Chapter 1 summarises clinical aspects of BTC as well as current diagnostic
and prognostic tests.
Chapter 2 addresses the identification of circulating tumour cells for the
diagnosis of BTC. It includes details of a study investigating measurement of
circulating cytokeratin 19 fragments (CYFRA 21-1), demonstrating that
CYFRA 21-1 is a more specific, but less sensitive diagnostic marker than
CA19-9, and predicts a poor prognosis in BTC.
Chapter 3 investigates the potential for using RNA isolated from archived
formalin fixed, paraffin embedded (FFPE) surgical and explanted liver tissues
from patients with primary sclerosing cholangitis (PSC) with and without
cholangiocarcinoma, for use in whole genome RNA expression analysis. We
demonstrate that, although technically possible, the rarity of samples and
RNA degradation that occurs as a result of the tissue processing, are such
that further evaluation using these materials is not feasible at this time.
Chapter 4 addresses and validates methodology for isolating RNA from
samples of biliary brushings taken at the time of endoscopic retrograde cholangiopancreatography (ERCP). We demonstrate that RNA isolated from
biliary brushings is of low quantity and degraded, and that this degradation
occurs in vivo. However, we demonstrate that such RNA is still useful for
downstream applications such as quantitative real time PCR and is therefore
suitable for whole genome RNA expression analysis using microarray
technology.
Chapter 5 describes the methods and results obtained from using whole
genome RNA expression analysis using microarray of RNA isolated from
ERCP biliary brushings. The results are presented as a shortlist of candidate
genes requiring further validation.
Chapter 6 provides results of qPCR studies performed in order to validate the
gene expression profile identified by microarray. A selection of candidate
genes are investigated using TaqMan Array and SYBR Green qPCR and
demonstrate a high correlation with the pattern of expression shown by
microarray.
Chapter 7 investigates whether a selection of the genes identified in
malignant biliary brushings are similarly upregulated in fresh frozen surgical
resection material from patients with benign and malignant biliary diseases. In
addition, we provide evidence for gene translation and upregulation at the
protein level by immunohistochemistry for a selection of the protein products.
Chapter 8 discusses the main conclusions drawn from the work as well as
potential future studies.}
}