published_or_final_version / abstract / Paediatrics and Adolescent Medicine / Doctoral / Doctor of Philosophy

published_or_final_version / Medical Sciences / Master / Master of Medical Sciences

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No description available.

No description available.

No description available.

No description available.

Hypoxia, a state of low oxygen, is a feature of most solid tumours as a consequence of poor tumour vascularisation. The mechanisms, which allow cancer cells to survive and continue to grow in hypoxia, are coordinated by the transcription factor HIF. The tumour suppressor gene van Hippel-Lindau (vHL) that targets HIF for degradation is mutated in the vast majority of renal cell carcinomas (RCCs), highlighting the importance of hypoxia to tumour biology. There is, therefore, an important need to understand the adaptive changes mediated by hypoxia and to target this clinically. One class of genes regulated by HIF are microRNAs (miRNAs). MiRNAs are short, single stranded RNA that primarily inhibit protein expression from target m RNA. The first aim of this project was to identify novel hypoxia regulated miRNAs in bladder cancer and assess their functional significance. It was found that a number of miRNAs were induced in hypoxic conditions. The hypoxic induction of miR-210 was conserved in all cell lines tested. MiR-145 was found to be highly induced by hypoxia in RT4, a cell line derived from a low- grade, non-muscle invasive tumour. We showed that miR-145 was a novel, HIF target gene with two hypoxia response elements identified within the promoter. Functionally we found that miR-145 induces apoptosis in RT4 cells. MiR-100 was downregulated in hypoxia, but this downregulation did not involve HIF. Regulation of miR-100 was of interest, as it is known to target FGFR3, a gene commonly overexpressed or mutated in bladder cancer. Concomitant with a decrease in miR-100, both the mRNA and protein level of FGFR3 were found to increase in hypoxia in RT4 and RT112 cells. Increased FGFR3 expression in hypoxia was involved in sustaining activation of the downstream signaling targets phospho-PKB and phospho-ERK. In addition, we demonstrate a role for FGFR3 in regulating both 2D and 3D growth and of miR-100 in regulating 3D growth of RT4 cells. We also showed that miR-100 decreased the protein levels of mammalian target of rapamycin (mTOR). However, transfection of miR-l00 into RT4 cells did not affect the sensitivity of this cell line to rapamycin. The genetic and biochemical changes that occur in (hypoxic) tumours may alter their responsiveness to chemotherapeutic agents such as rapamycin. The second aim of this project was to investigate the responsiveness of RCCs to clinically approved chemotherapeutic agents, with the goal of correlating any differences in response to alterations in expression of specific genes. Although hypoxia regulated miR-100 did not affect sensitivity to rapamycin, we extended these studies and investigated the role of vHL status on response of renal cancer cell lines to sorafenib, sunitinib, rapamycin and metformin. We found that the presence of vHL increased resistance to rapamycin. Sensitivity to these drugs was also tested in 10 primary cell lines. There was varying sensitivity to these drugs across the cell lines representing the heterogeneity of renal cancer. We analysed the expression of a number of genes in the m TOR and hypoxic pathways in these tumours, we found the expression of a known hypoxic gene REDDl correlated with sensitivity to rapamycin. REDDl expression levels were also higher in tumour tissue when compared to normal renal parenchymal tissue and was associated with other prognostic markers such as CA9, miR-210 and vascular invasion suggesting a role as a diagnostic or prognostic marker and in patient selection for treatment with rapamycin.

D. Litt et Phil. / It is well known that high levels of anxiety and/or depression often accompany the diagnosis and treatment of cancer. Literature from various sources, but in particular from the fairly new field of research, Psychoneuroimmunology, also provides ample evidence that excessive anxiety and/or depression can be immunosuppressive. It makes sense, therefore, that any intervention restoring balance to the immuno-regulatory system, thereby allowing the body’s innate healing processes to focus on eliminating cancer, is highly desirable. In line with current thinking based on the mind-body connection as well as cognitive behavioural techniques utilised in many therapeutic settings, various psychological interventions have been found to help the patient gain a better sense of control over distressing symptoms and side-effects of cancer. Some of these include: basic cognitive restructuring, hypnotherapy, relaxation-meditation techniques, art and music therapy, and guided imagery. Substantial international research illustrates the beneficial effect that relaxation and/or guided imagery provides in such diverse settings including work, sport and health. In this regard, it was decided to run a pilot study to ascertain whether a customised tape recording with a relaxation and guided imagery dialogue aimed at helping patients manage and cope with negative symptoms of cancer, could significantly reduce anxiety levels in patients with cancer receiving radiotherapy. To operationalise the above, 30 men and women, aged between 20 and 80, with Stages 1, 2 or 3 breast, prostrate, gynaecological cancers, and head and neck cancers, who were about to commence radical (minimum 25 fractions) radiotherapy, were randomly selected to an experimental and a control group. A consecutive sample, pre-test post-test experimental design was applied to this study in which the experimental and control groups were subjected to pre- and post radiotherapy Hospital Anxiety & Depression (HAD) Scale, Institute for Personality Assessment and Training (IPAT) Anxiety Scale and blood pressure measurements during their 1st, 3rd, 6th week cycle of treatments, as well as a final measurement 12 weeks after commencement of therapy. The main hypothesis of this pilot study was that there would be statistically significant decreases in levels of anxiety as a result of the intervention of guided imagery tape recording in patients with non-metastatic cancer undergoing curative radiotherapy. For the intervention, each experimental participant was taught a relaxation technique and then following an interview a customised guided imagery dialogue developed for the participant’s sole use. The participant was requested to listen to this tape at least once a day. The control group had the same pre- and post tests as the experimental group, but did not receive any intervention. Statistical analysis of the data revealed that the experimental group showed a tendency towards decreased blood pressure and anxiety over the course of radiotherapy. The most significant change, however, was noted in terms of diastolic blood pressure, suggesting that the intervention corresponded to a physiological decrease in anxiety. There was not a statistically significant difference in terms of the measured psychological variables. A general conclusion to this pilot study suggests that whilst guided imagery may contribute to a lowering of anxiety, additional cognitive intervention would probably affect a more substantial and sustained change in the patient. Although this pilot study revealed some methodological weaknesses the results are sufficiently encouraging to warrant further in-depth research regarding the use of guided imagery as a cost-effective, easy method for individuals to learn and utilise as part of their integrative cancer treatment programme.

Inflammatory Breast Cancer (IBC) is a rare, lethal, and understudied form of breast cancer. Although affecting 1-2% of the population, the remission rate is half that of the spectrum of other breast cancers, and most cases present in the advanced stages due to rapid undetectable development. Of the diagnosed cases, systemic chemotherapeutics are relatively ineffective in comparison to non-IBC breast cancer cases, indicating other unique mechanisms driving IBC progression. Historically, the specific sensitivities of a particular tumor type or subtype have been linked to genetic alterations that represent addiction hubs, such as hyperactivation of oncogenes due to mutation. Although some efforts have been made to characterize the molecular fingerprint of inflammatory breast cancers (IBCs), unfortunately, no clinical application has emerged from these studies. Thus, we decided to utilize a different strategy to identify the Achilles' heel of IBC cells. Using shRNA libraries, we performed an unbiased genome-wide loss-of-function screen comparing the gene functions required for survival of IBC and non-IBC cells. Histone deacetylase 6 (HDAC6) emerged as one of the top genes required for IBC cell survival and was further validated. HDAC6 is vital in the cell's unfolded protein response (UPR) to clear misfolded or toxic protein, and IBC cells proved to be preferentially sensitive to this aspect of HDAC6 inhibition, displaying increased protein accumulation, ER stress indicators, and subsequent apoptosis upon failure to clear or refold accumulated proteins. These data indicate HDAC6 is a crucial gene required for IBC cell line survival, in part due to its function in IBC cell UPR. Furthermore, emerging orally bioavailable agents for HDAC6 inhibition make it a promising candidate towards tailored therapeutic implementation in IBC patient trials.