Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.784213
Title: EGFR and related therapeutic targets in malignant pleural mesothelioma
Author: Oguh, Lily I.
ISNI:       0000 0004 7969 7743
Awarding Body: University of Hull and University of York
Current Institution: University of Hull
Date of Award: 2016
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Abstract:
Introduction Malignant pleural mesothelioma (MPM) is a rare but aggressive disease and the current first line treatment is associated with a survival rate of 40%. There is currently no second line therapy. This study aimed to explore the expression of proteins in the arachidonic acid pathway, VEGFR-2 pathway, EGFR/HER2 pathway, c-MET pathway and the PI3K/AKT/MTOR pathway in MPM archival tissue samples and cell lines. We also investigated the cytotoxic effect of inhibitors for each individual pathway as single agents and in some instances as combinations. Materials and Methods Immunohistochemical analysis was performed in 93 archival MPM tissue samples to determine the expression of the HER2, 5-LOX, 12-LOX, VEGFR-2 and c-MET proteins. Mesothelioma cell lines NCI-H2452, NCI-H2052, MSTO-211H and a non-small cell lung cancer cell line A549 were used for western blot analysis, MTS assay and in vitro scratch assay. Western blot analysis was used to evaluate 5-LOX, 12-LOX, VEGFR-2, EGFR, p-ERK, ERK, c-MET, PTEN and p70S6K protein expression in the cell lines. The antiproliferative effect of Baicalein (12-LOX), Zileuton (5-LOX), MK886 (FLAP inhibitor), Celecoxib (COX-2), Cediranib (VEGFR-2), MGCD265 (c-MET/VEGFR inhibitor), Afatinib (EGFR/HER2), Gefitinib (EGFR), Selumetinib (MEK), Tivantinib (c-MET), Crizotinib (c-MET/ALK), SU11274 (c-MET), Onartuzumab (MET monoclonal antibody), NVPBEZ235 (PI3K/AKT/MTOR), VS5584 (PI3K/AKT/MTOR), Ku0063794 (MTOR1/MTOR2), XL388 (MTOR1/MTOR2) was assessed as single agents and in combinations and analysed using the MTS proliferation assay. Results Positive 5-LOX and 12-LOX protein expression was seen in 73% (56/77) and 83% (69/83) of archival MPM tissue samples respectively. NCI-H2452, NCI-H2052, MSTO-211H and A549 cells also expressed 5-LOX and 12-LOX proteins. Baicalein was effective in all cell lines. Combination of celecoxib (3 μM) and baicalein (10 μM) was synergistic in the MSTO-211H cell line. Positive VEGFR-2 protein expression was seen in 93.8% (75/80) of archival tissue samples. Cediranib demonstrated cytotoxic effect at doses higher than the clinical relevant dose. MGCD265 also reduced cell proliferation in all cell lines. Positive HER2 expression was seen in 86.2% (69/80) of archival tissue samples. All cell lines expressed EGFR, p-ERK and ERK protein. Gefitinib and Afatinib demonstrated cytotoxic effects at doses significantly higher than their therapeutically relevant doses. Positive c-MET expression was seen in 82% (58/71) of archival tissue samples. NCI-H2452, NCI-H2052, MSTO-211H and A549 cells also expressed c-MET protein. Crizotinib inhibited cell growth by 50% in MSTO-211H cells within its clinically relevant dose. SU11274 also reduced cell growth by 50%. Tivantinib reduced cell growth by 50% in all cell lines at doses significantly lower than its clinically achievable dose of 4 μM. A549 and MSTO-211H cells positively expressed the p70S6K protein and loss of PTEN was also observed in the MSTO-211H cells. PI3K/AKT/MTOR inhibitors, NVPBEZ235 and VS-5584 significantly reduced cell growth by 50% at low nanomolar IC50 values. VS-5584 was combined in turn with Tivantinib and Afatinib. The combination of VS-5584 with Tivantinib demonstrated enhanced growth inhibition in all cell lines compared to either inhibitors alone. Combination of Tivantinib and Afatinib also enhanced the inhibition of cell growth in all cell lines compared to either inhibitors alone. The addition of cisplatin to the tyrosine kinase combinations produced a synergistic effect. Conclusions Our findings suggest that multiple signalling pathways are active in a significant proportion of MPM samples. Co-targeting the c-MET and PI3K/MTOR pathway might be a potential therapeutic strategy for mesothelioma patients. Further work is required to explore the combination of an EGFR inhibitor and a PI3K/MTOR inhibitor when the EGFR inhibitor is fixed at a therapeutically relevant dose. In addition, understanding the molecular mechanism of Tivantinib, Afatinib and VS-5584, through the use of comparative proteomic platforms, could potentially identify predictive biomarkers of response to these anti-cancer agents in mesothelioma patients.
Supervisor: Cawkwell, Lynn ; Lind, Michael Sponsor: University of Hull
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.784213  DOI: Not available
Keywords: Medicine
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