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Title: Generation and characterisation of novel monoclonal antibodies towards ovarian tumour stem cells
Author: Luk, Louisa Tien Sze
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2012
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Tumour stem cells (TSCs) are hypothesised to be a rare population of tumour cells which possess stem cell-like properties and are resistant to conventional therapy. Although cell surface markers have been widely used to characterise TSCs, previous literature suggests that no specific marker has been found for ovarian TSCs. We aimed to identify and characterise novel antibodies specific to ovarian epithelial TSCs, particularly towards populations with Hoechst efflux (Side Population, SP) and aldehyde dehydrogenase activity which are associated with a stem cell phenotype and drug resistance. Putative TSC subpopulations from ovarian tumour cell lines isolated by fluorescence activated cell sorting (FACS) using differential Hoechst dye uptake and Aldefluor activity assays displayed stem cell-like characteristics, including the upregulation of stem cell markers, increased anchorage-independent growth and increased invasive properties. A panel of monoclonal antibodies (mAbs) was then generated by injecting Aldefluor-positive ovarian tumour IGROV1 cells into female BALB/c mice. 34 antibodies were found to be specific to Aldefluor-positive cells and 2 of these enriched for SP cells. The 2 mAbs demonstrated cross-reactivity on human embryonic stem cells but no cross-reactivity to normal ovarian cell lines. Subpopulations of ovarian cell lines positive for the mAbs displayed stem cell-like characteristics, including upregulation of stem cell markers - CD133, ABCB1 and ALDH1A1. Sorted mAb populations were injected into non-obese diabetic/severe combined (NOD/SCID) mice and differential in vivo tumour formation was observed. Finally, the target antigen which both mAbs recognised was identified by mass spectrometry to be clathrin heavy chain (CHC1). We conclude that rare subpopulations with tumour-sustaining capability and stem cell-like characteristics can be identified in ovarian cancer using the 2 novel antibodies generated. Both mAbs target CHC1 on tumour-sustaining populations which are enriched for multiple stem cell markers and are therefore potential novel diagnostic markers and/or therapeutic agents.
Supervisor: Oh, Steve ; Brown, Bob ; Choo Boon Hwa, Andre Sponsor: Agency for Science, Technology and Research, Singapore
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available