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dc.contributor.advisorO'Sullivan, Jacintha
dc.contributor.authorBUCKLEY, AMY
dc.date.accessioned2019-08-07T12:28:39Z
dc.date.available2019-08-07T12:28:39Z
dc.date.issued2019en
dc.date.submitted2019
dc.identifier.citationBUCKLEY, AMY, Enhancing treatment response to neo-adjuvant radiation therapy in oesophageal cancer using novel dual action drugs targeting tumour energy metabolism and angiogenesis, Trinity College Dublin.School of Medicine, 2019en
dc.identifier.otherYen
dc.identifier.urihttp://hdl.handle.net/2262/89158
dc.descriptionAPPROVEDen
dc.description.abstractOesophageal cancer is an aggressive malignancy associated with a poor 5-year survival of <20%, accounting for over 400,000 deaths each year. The current standard of care for oesophageal adenocarcinoma (OAC) is neo-adjuvant therapy with chemotherapy or chemoradiation therapy to debulk the tumourprior to surgery. However, only 30% of patients achieve a complete pathological response to neo-adjuvant treatment. Currently there are no approved biomarkers which can be used in the clinic to stratify patients into good and poor responders prior to treatment. In addition, there are currently no approvedradiosensitisersin the neo-adjuvant space which can be administeredwith the standard of care to enhance radioresponse. The radioresistant phenotype is polymodal and influenced bymultiple biological processes. Altered energy metabolism and mitochondrial function were previously linked with the radioresistant phenotype in OAC. In this thesis we evaluated the actionof our novel small molecule patented compounds onangiogenesis, energy metabolism and inflammatory secretions to enhance radioresponse. Through phenotype-based screens we identified Pyrazinib (P3), a pyrazine phenol small molecule drug with anti-angiogenic and anti-metabolic activity in-vivo inzebrafish andin-vitro isogenic models of OAC radioresistance. Pyrazinib (P3) significantly inhibited intersegmental vessel development in zebrafish embryos. In-vivo, in zebrafish Pyrazinib (P3) significantly inhibited both oxygen consumption rate (OCR) and extracellular acidification rate (ECAR), measures of oxidative phosphorylation and glycolysis respectively, when evaluated using seahorse technology. In-vitro, in an isogenic model of OAC radioresistance, Pyrazinib (P3) significantly reduced OCR in both OE33P (radiation-sensitive) and OE33R (radiation-resistant) cells, in addition Pyrazinib (P3) produced a simultaneous reduction in ECAR in OE33R cells. The ability of Pyrazinib (P3) to alter radiosensitivity was evaluated by clonogenic assay. Pyrazinib (P3)significantly reduced the surviving fraction in OE33P; radiation-sensitive and OE33R; radiation-resistant cells following 2, 4 and 6 Gy irradiation. Under hypoxic conditions Pyrazinib (P3), significantly reduced the surviving fraction of OE33R cells following 4 Gy irradiation.Pyrazinib (P3) did not significantly alter the sensitivity of cisplatin sensitive OE33 Cis P and cisplatin resistant OE33 Cis R cells to cisplatin treatment nor did it alter sensitivity of OE33P or OE33R cells to 5-flouracil. Inflammation haspreviouslybeenlinked with the radioresistant phenotype in OAC. Proteomic profiling of 184 proteins found leukaemia inhibitory factor (LIF), to be significantly elevated in radioresistant OAC cells. Furthermore, significantly higher circulating levels of LIF were present ivin the serum of treatment-naive OAC patients who had a subsequent poor pathological response to neo-adjuvant therapy. Quantitative PCR analysis revealed expression of LIF receptor (LIFR) may function as a predictive indicator of response to neo-adjuvant chemoradiation therapy in OAC. LIF was demonstrated to be actively secreted from human OAC treatment-naïve biopsies and significantly correlated with the secretion of bFGF, VEGF-A and IL-8. Importantly, LIF secretion negatively correlated withthe percentage oftumour infiltrating lymphocytes inmatchedpre-treatment OAC patient biopsies. Elevated circulating LIF is a marker of poor response to neo-adjuvant treatment in OAC and secretion of this chemokine from the tumour is tightly linked with pro-tumorigenic mediators including bFGF, VEGF-A and IL-8. Targeting this pathway may reveal a novel mechanism to enhance neo-adjuvant treatment responses in OAC.Further investigation of the inflammatory secretions from our isogenic model of OAC radioresistance by multiplex ELISA showed11 of 30 detected inflammatory and angiogenic factors were secreted at significantly higher levels in OE33R than OE33P cells; IL-2, IL-4, IL-6, IL-8, IL-12p70, IL-10, MCP-1, IP-10, ICAM. Pyrazinib (P3) significantly reduced the secretions of IL-4, IL-6, IL-8 and IL-13 in OE33R cells.Furthermore, in this thesis for the first time we have demonstrated in real-time using the Seahorse Biosciences XFe24 analyser and human ex-vivobiopsies, OAC tumour biopsies have a significantly higher OCR, a measure of oxidative phosphorylation compared to ECAR, a measure of glycolysis highlighting the importance of aerobic respiration in OAC. Pyrazinib (P3), an anti-metabolic small molecule radiosensitiser significantly inhibited oxygen consumption rate in OAC biopsies. Furthermore, biopsies from OAC tumours can quickly adapt their metabolic rate to their environment to cope with hypoxia. Under conditions of hypoxia, Pyrazinib (P3) produced a simultaneous and significant reduction in both OCR and ECAR in OAC treatment-naïvebiopsies. We demonstrated that the inflammatory secretome profile of OAC treatment-naïve biopsies is highly heterogeneous. OCR was significantly correlated with three secreted factors in the tumour conditioned media: VEGF-A, IL-1RA and TSLP. ECAR was significantly correlated with VEGF-A, IL-1RA, TSLP, IL-13, MIP-3α and TNF-α. Pyrazinib (P3) significantly inhibited IL-1β secretion and increased IL-3 and IL-17B secretions from treatment-naïve OAC patient tumour biopsies. Critically, Pyrazinib (P3) did not directly alter the expression of dendritic cell maturation markers. The findings of this thesis have identified the small molecule compound Pyrazinib (P3) with anti-angiogenic, anti-metabolic and anti-inflammatory action which can enhance radiosensitivity both under normoxic and hypoxic conditions in-vitroin isogenic model of OAC radioresistance. Further pre-clinical development of this small molecule compound willbeenrequiredin different in-vivomodels to track radiosensitivity more precisely.en
dc.language.isoenen
dc.publisherTrinity College Dublin. School of Medicine. Discipline of Surgeryen
dc.rightsYen
dc.subjectOesophageal Canceren
dc.subjectRadiationen
dc.subjectMetabolismen
dc.subjectAngiogenesisen
dc.titleEnhancing treatment response to neo-adjuvant radiation therapy in oesophageal cancer using novel dual action drugs targeting tumour energy metabolism and angiogenesisen
dc.typeThesisen
dc.contributor.sponsorIrish Cancer Societyen
dc.contributor.sponsorIrish Cancer Societyen
dc.type.supercollectionthesis_dissertationsen
dc.type.supercollectionrefereed_publicationsen
dc.type.qualificationlevelDoctoralen
dc.identifier.peoplefinderurlhttps://tcdlocalportal.tcd.ie/pls/EnterApex/f?p=800:71:0::::P71_USERNAME:BUCKLEA6en
dc.identifier.rssinternalid205922en
dc.rights.ecaccessrightsopenAccess


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