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Title: CSPG4 in osteosarcoma : functional roles and therapeutic potential
Author: Worrell, Harrison
ISNI:       0000 0004 7654 272X
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2018
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Osteosarcoma is the most common primary malignancy of bone. 5-year survival has remained stable at around 60-70% for 40 years. However, a number of patients will suffer from recurrent and/or metastatic disease representing a large unmet clinical need. CSPG4 is a transmembrane protein which is expressed on a number of progenitor cells and tumour types. Preliminary work had found CSPG4 present in osteosarcoma tumour samples. In this study, CSPG4 mRNA and protein expression was demonstrated in clinical samples and model cell lines. CSPG4 mRNA is overexpressed in osteosarcoma samples compared to mature osteoblast cells, the putative cell of origin for osteosarcoma. In a cohort of patients, CSPG4 protein expression was found on 86% of samples. Furthermore, CSPG4 expression was demonstrated in U2OS, MG63, HOS, HOS-MNNG and 143B osteosarcoma cell lines. CSPG4 protein expression was successfully deleted in 143B cells using CRISPR/Cas9 technology. Two stable CSPG4-negative cell lines were produced. CSPG4 expression was then reintroduced into negative cell lines, as well as the parental 143B cell line. This created a panel of 6 cell lines with differing CSPG4 expression. Furthermore, siRNA treatment of U2OS, MG63, 143B and U87MG cell lines reduced CSPG4 expression. These cells provided another panel with varying CSPG4 expression for in vitro investigation. In vitro experiments failed to demonstrate a role for CSPG4 in osteosarcoma tumorigenesis. The CRISPR/Cas9 cell panel found that CSPG4 expression did not influence cell proliferation, adhesion and spreading on fibronectin or collagen-I, cell migration, chemosensitivity or anchorage-independent growth. Similarly, the siRNA cell panel found that CSPG4 expression did not influence cell proliferation or anchorage-independent growth. In vivo experimentation did not demonstrate a role for CSPG4 in mediating osteosarcoma tumour growth or metastatic spread. Treatment with a sc-Fv antibody fragment failed to demonstrate specific toxicity of CSPG4-positive cell lines. These results indicate that CSPG4 plays no role in osteosarcoma tumour cell behaviour. However, due to its wide expression pattern it represents a viable therapeutic option for drug targeting.
Supervisor: Salter, Donald ; Brunton, Valerie Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: osteosarcoma ; CSPG4 ; transmembrane protein ; CSPG4 mRNA ; CRISPR/Cas9 ; CSPG4 expression ; osteosarcoma tumour cell behaviour