Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.706776
Title: Investigation of biomarkers associated with hypergastrinaemia and their responses to CCK2 receptor antagonism
Author: Lloyd, K. A.
ISNI:       0000 0004 6058 9339
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2016
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Abstract:
Gastrin is a hormonal regulator of gastric acid secretion, but also regulates other cellular functions in the stomach such as proliferation, migration and apoptosis. Elevated serum concentrations of gastrin may dysregulate these processes and contribute to the development of gastric neuroendocrine tumours (NETs) and adenocarcinomas. The inhibition of gastrin or its subsequent downstream signalling pathways may therefore have therapeutic value in gastrin-related disorders including gastric NETs. The aims of this PhD were to investigate the role of gastrin in gastric tumour development and to identify potential biomarkers for the detection of gastrin-associated cellular phenotypical changes which may contribute to the development of gastric pathologies. I also investigated whether three novel cholecystokinin type-2 receptor (CCK2R) antagonists inhibited gastrinassociated morphological changes in the stomach, in vitro. miRNA PCR arrays showed that gastrin significantly increased the expression of both miR-222 and miR-376c in human gastric cancer cells which express the cholecystokinin type-2 receptor (AGSGR). Of these two miRNAs, only the increase in miR-222 expression was confirmed using quantitative polymerase chain reaction (qPCR). Using chemical molecular pathway inhibitors, gastrininduced miR-222 overexpression was shown to occur via activation of the CCK2R and subsequent phosphokinase C (PKC) and phosphatidylinositol-3-kinase (PI3K) pathways. miR-222 expression significantly increased with age and further increased with Helicobacter felis (H. felis) infection in hypergastrinaemic INS-GAS mice. Similarly in patients with hypergastrinaemia and type-1 gastric NETs, miR-222 expression was increased at baseline relative to healthy control subjects and decreased whilst the patients were taking the CCK2R antagonist, netazepide. miR-222 overexpression caused p27 mislocalisation to the cytoplasm and resulted in decreased p27 mRNA and protein expressions, in vitro. Functional analyses using small interfering RNAs (siRNAs) identified that this decreased p27 expression resulted in cell migration and structural remodelling in AGSGR cells. Previous cDNA microarray studies identified additional gastrin-regulated genes in gastric biopsies from hypergastrinaemic patients with type-1 gastric NETs being treated with netazepide. Of eight potential gene candidates, the tissue remodelling protein pregnancy-associated plasma protein-A2 (PAPPA2) was selected for further investigated in vitro due to its previous association with tissue remodelling in the placenta. Gastrin treatment increased PAPPA2 mRNA and protein expressions in AGSGR cells and this resulted in increased cell migration and cellular remodelling. Several CCK2R antagonists have previously been developed for the treatment of acid-peptic disorders and hypergastrinaemic conditions including gastric NETs. However oral bioavailability, receptor selectivity and/or drug potency still remain an issue. The inhibition of gastrin-induced cellular phenomena by the novel CCK2R antagonists TR1, TR2 and TR3 was therefore assessed using cell-based assays and their potencies were compared against the previously established CCK2R antagonists, YM022 and netazepide. All compounds caused significant reductions in gastrin-induced cellular processes such as cell growth, migration, structural remodelling and clonogenic survival. TR2 showed the highest potency, but was equally or less potent than both YM022 and netazepide in all assays. These data have therefore identified two potential novel gastrin-regulated biomarkers and a potential novel CCK2R antagonist which may be further developed for the detection and treatment of gastrinrelated conditions.
Supervisor: Pritchard, D. M. ; Varro, A. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.706776  DOI:
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