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Title: Use of genetically engineered mouse models in preclinical drug development
Author: Creedon, Helen
ISNI:       0000 0004 5916 5256
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2015
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The paucity of well validated preclinical models is frequently cited as a contributing factor to the high attrition rates seen in clinical oncological trials. There remains a critical need to develop models which are accurately able to recapitulate the features of human disease. The aims of this study were to use genetically engineered mouse models (GEMMs) to explore the efficacy of novel treatment strategies in HER2 positive breast cancer and to further develop the model to facilitate the study of mechanisms underpinning drug resistance. Using the BLG--HER2KI-PTEN+/- model, we demonstrated that Src plays an important role in the early stages of tumour development. Chemopreventative treatment with dasatinib delayed tumour inititation (p= 0.046, Wilcoxon signed rank test) and prolonged overall survival (OS) (p=0.06, Wilcoxon signed rank test). Dasatinib treatment also induced squamous metaplasia in 66% of drug treated tumours. We used 2 cell lines derived from this model to further explore dasatinib’s mechanism of action and demonstrated reduced proliferation, migration and invasion following in vitro treatment. Due to the prolonged tumour latency and the low metastatic rate seen in this model, further studies were undertaken with the MMTV-NIC model. This model also allowed us to study the impact of PTEN loss on therapeutic response. We validated this model by treating a cohort of MMTV-NIC PTEN+/- mice with paclitaxel and demonstrated prolonged OS (p=0.035, Gehan Breslow Wilcoxon test). AZD8931 is an equipotent signalling inhibitor of HER2, HER3 and EGFR. We observed heterogeneity in tumour response but overall AZD8931 treatment prolonged OS in both MMTV-NIC PTEN FL/+ and MMTV-NIC PTEN+/- models. PTEN loss was associated with reduced sensitivity to AZD8931 and failure to suppress Src activity, suggesting these may be suitable predictive biomarkers of AZD8931 response. To facilitate further studies exploring resistance, we transplanted MMTV-NIC PTEN+/- fragments into syngeneic mice and generated 3 tumours with acquired resistance to AZD8931. These tumours displayed differing resistance strategies; 1 tumour continued to express HER2 whilst the remaining 2 underwent EMT and lost HER2 expression reflecting to a very limited degree some of the heterogeneity of resistance strategies seen in human disease. To further explore resistance to HER2 targeting tyrosine kinase inhibitors, we generated a panel of human cell lines with acquired resistance to AZD8931 and lapatinib. Western blotting demonstrated loss of HER2, HER3 and PTEN in all resistant lines. Acquisition of resistance was associated with a marked change in phenotype and western blotting confirmed all lines had undergone EMT. We used a combination of RPPA and mass spectrometry to further characterise the AZD8931 resistant lines and identified multiple potential novel proteins involved in the resistant phenotype, including several implicated in EMT. In conclusion, when coupled with appropriate in vitro techniques, the MMTV-NIC model is a valuable tool for selection of emerging drugs to carry forward into clinical trials of HER2 positive breast cancer.
Supervisor: Brunton, Valerie Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: breast cancer ; Human Epidermal Growth Factor receptor 2 ; HER2 ; genetically engineered mouse models ; GEMM ; acquired resistance