Thesis

Preclinical pharmacological and toxicological study of novel inhibitory kappa B kinase alpha (IKK?) inhibitor compounds in prostate cancer

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Awarding institution
  • University of Strathclyde
Date of award
  • 2016
Thesis identifier
  • T14414
Person Identifier (Local)
  • 201251857
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Abstract
  • Prostate cancer (PCa) is the most frequently diagnosed cancer and the second most common cause of cancer deaths in men in the UK, despite the progress that has been made in PCa treatment. Nuclear Factor kappa B (NF-?B) regulates several genes involved in immune response, inflammation,proliferation and apoptosis. Overexpression of NF-?B has been shown to be involved in tumour progression and radiation therapy resistance of PCa. One potential approach in PCa cancer treatment therefore involves targeting I?Bkinases (IKKs) which are key regulators of the NF-?B signaling pathway. Whilst IKK? regulates the canonical NF-?B pathway involving degradation of I?B-? and phosphorylation of p65, IKK? regulates the non-canonical NF-?B pathway by mediating the processing of p100 to p52. Inhibition of NF-?B signaling by targeting IKK? is a sub-optimal treatment choice as it has been shown to be associated with multiple serious toxicities. However, IKK? is of interest in PCa therapy as it has been implicated in PCa survival, proliferation, invasion and metastasis. In addition, the non-canonical NF-?B pathway, which is regulated mainly by IKK?, has been demonstrated to drive the development of radioresistance in PCa cells. Therefore, targeting IKK? is a new approach for the development of novel therapies for PCa. In Chapter 3, the pharmacological characteristics of novel first-in-class IKK? inhibitors (SU compounds) developed in-house were investigated in terms of selectivity against IKK? and IKK? in PCa cell lines using Western blot analysis.In Chapter 4, the phenotypic impact of the selective IKK? inhibitors (SU compound) in PCa and normal prostate cell lines was interrogated using cell growth assay, cell viability assay, clonogenic assay, caspase-3 assay and cell cycle analysis, and the radiosensitisation potential of selective IKK? inhibitor SU compounds in PCa cell lines were investigated using clonogenic assay, cellcycle assay and combination index methods. In Chapter 5, the cytotoxicity and radiosensitisation effects of non-pharmacological inhibition of IKK? and IKK? using small interfering RNAs (siRNAs) in PCa cell lines were also investigated using cell growth assay, clonogenic assay, cell cycle analysis and linear quadratic analysis.;These studies demonstrated that among investigated SU compounds, SU1261 most selectively inhibits the non-canonical NF-?B pathway biomarker p52 at lower concentrations than the canonical NF-?B pathway biomarkers, I?B-? degradation and phosph-p65, demonstrating selectivity of this compound for IKK? over IKK?. Furthermore, SU1261 showed a highly cytotoxic profile in term of inhibition of growth rate, reduction in cell viability and clonogenicity, and induced apoptosis in both PCa (PC3M-luc-C6 and PC3) and normal prostate (PNT2A) cell lines. However, SU1261 exhibited lower cytotoxicity against normal prostate epithelium cells (PNT2A) compared with PCa cells (PC3M-luc-C6 andPC3), suggesting that inhibition of IKK? may offer a therapeutic differential in cancer versus normal cells. Furthermore, a combination of SU1261 and X-ray irradiation delivered to PC3M-luc-C6s showed a synergistic interaction which resulted in an enhanced radiation-induced colony survival inhibition arrest of thecell cycle at the G2/M phase. On the other hand, knockdown of the IKK? proteinin PC3M-luc-C6s using siRNA only impacted the growth rate of cells, but silencing of IKK? inhibited both growth rate and clonogenicity of PC3M-luc-C6 cells as well as inducing apoptosis. In addition, knockdown of both kinases did not show significant enhancement in radiation cytotoxicity in PC3M-luc-C6 cells.These data taken together represent one of the first interrogations of the phenotypic effect of IKK? inhibition and suggest that SU1261 is a first-in-class novel selective IKK? inhibitor compound. Pharmacological inhibition of IKK? using SU1261, but not with siRNA against IKK?, as a single agent or incombination with radiation may be a useful approach for PCa treatment.
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DOI
Date Created
  • 2016
Former identifier
  • 9912533812802996

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