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Title: A novel method to quantify gamma H2AX foci in circulating tumour cells in patients receiving chemotherapy for colorectal cancer
Author: Saggese, Matilde
ISNI:       0000 0004 7232 3976
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2018
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Colorectal cancer (CRC) is the third most common cancer in males and females. Circulating tumour cells (CTCs) are epithelial cancer cells that mediate haematogenous metastases and can be used as predictive and prognostic markers. Gamma-H2AX (γ- H2AX) foci represent double strand DNA breaks and DNA damage. Assessing γ-H2AX foci in CTCs could be utilised as a biomarker to measure patient response to DNAinteractive anti-cancer treatments in real time, aiding treatment decisions. The aim of this study was to develop a method to quantify changes in γ-H2AX in CTCs from metastatic CRC patients undergoing treatment with FOLFOX (oxaliplatin with fluorouracil 5FU and folinic acid chemotherapy) or FOLFORI (irinotecan with 5FU and folinic acid). Human CRC cell lines (HT-29; HCT-116) treated with oxaliplatin, SN-38 and topotecan alone or spiked into healthy donor blood were evaluated to assess γ-H2AX signal using both the CellSearch® System (Janssen Diagnostics) and the DEPArrayTM System (Silicon Biosystems). The fluorescent signal in cells could not be quantified using CellSearch followed by DEPArray analysis, but when DEPArray was used alone, treated cells demonstrated a significantly increased intensity of fluorescein isothiocyanate-conjugated (FITC) anti-γ-H2AX antibody staining compared with control cells. This indicated the DEPArray system was able to quantify differences in signal intensity caused by induction of γ-H2AX in CTCs. To determine if this could be applied clinically, the effect of CellSearch scanning on FITC intensity detected by DEPArray was evaluated using topotecan treated HT-29 cells that were scanned or unscanned with CellSearch followed by DEPArray analysis; scanned cells expressed a statistically significant lower FITC signal intensity compared with unscanned cells. Evaluation of γ-H2AX in CTCs from CRC patients was inconclusive due to small patient numbers. This study suggests a potential barrier for clinical application using the method of DEPArray following CellSearch analysis, therefore alternative methods should be evaluated to determine a suitable assay for use in the clinic.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available