Bladder symptoms 5-11 years after radiotherapy for cervical carcinoma were investigated by postal questionnaire in 66 women. 25% complained of severe bladder symptoms and 30% of moderate symptoms. The most common symptoms were urgency and urge incontinence. Symptoms in a group of postmenopausal women were significantly less and were not altered by taking hormone replacement or placebo for 16 years. Bladder function measured urodynamically in women with cervical carcinoma prior to treatment was normal, suggesting that cervical carcinoma does not alter bladder function. Following radiotherapy voiding was unaltered. Bladder capacity is reduced but only by 80ml. Detrusor pressure rises slowly for at least 5 years after radiotherapy, but only to a mean of 13 cmH20. unstable detrusor contractions were found in 30% of women 5 or more years after radiotherapy and appear to be the cause of symptoms in these women, not a fibrosed, contracted bladder as thought. Major bladder complications following radiotherapy had an incidence of 1.7% with fistulae in 1.4%. After radical hysterectomy 25% complained of stress incontinence and 75% had voiding problems. Maximum flow rate was reduced and residual volume increased. The prospective study found a reduction in functional urethral length and maximum urethral closure pressure at six months despite the posterior lcm of the cardinal ligament being preserved. The combination of radical postoperative radiotherapy caused detrusor pressure was increased detrusor instability. hysterectomy voiding with a to high and adjuvant be unchanged, incidence of Radiotherapy therefore may be a cause of detrusor instability. As bladder dysfunction is common after any treatment for cervical carcinoma patients should be followed up with regard to this and offered urodynamic investigation.

Background: Household contacts (HHCs) of pulmonary TB patients are at high risk of Mycobacterium tuberculosis (Mtb) infection and early disease development. Tuberculin skin test (TST) has been traditionally used to identify infected individuals; however, its use is limited by low specificity in populations with high levels of BCG vaccination or significant exposure to non-tuberculosis mycobacteria (NTM), and reduced sensitivity in immunocompromised individuals. Interferon-gamma release assays (IGRAs) such as QuantiFERON-TB Gold In-Tube (QFT-GIT) using Mtb specific antigens provide an alternative to TST for infection detection. IGRAs are now widely used for the detection of Mtb infection and are included in the guidelines of many countries with a low incidence of TB. Despite a growing body of literature on IGRAs, the relationship between the magnitude of T-cell Interferon-γ responses to Mtb specific antigens and risk of progression to disease has not been studied. Objective The main objective of this study was to determine whether HHCs with high (≥10 IU/ml) levels of IFN-γ in response to Mtb specific antigens (ESAT-6, CFP-10 or TB 7.7) in the QFT-GIT assay are at higher risk of developing TB compared to those with low (> 0.35-<10 IU/ml) levels. Other secondary objectives included to determine the following: the performance and operational characteristics of QFT-GIT in a field setting; risk factors associated with positive QFT-GIT results; concordance between the two tests; incidence rates of TB in HHCs with positive and negative QFT-GIT and TST results at baseline as well as positive and negative predictive values. Method: This study was nested within a large community randomized trial called ZAMSTAR implemented in 16 communities in Zambia and 8 communities in the Western Cape Province of South Africa. A cohort of HIV-positive and HIV-negative adult (≥ 15 years) HHCs were prospectively followed for 2-4 years. Consenting HHCs had blood drawn for HIV antibodies. QFT-GIT test was performed according to the manufacturer's instructions. TST were performed according to the standard IUATLD protocol. A standardized questionnaire was used to collect information on risk factors for TB and TB treatment information (for those with TB). Results: The feasibility studies showed three main findings. Firstly, the sensitivity of QFT-GIT was greater than that of TST overall, at all the standard TST cut-offs and when stratified by HIV status. The sensitivity of QFT-GIT was 85.6% (95%CI: 77.0-91.9) (indeterminate results excluded) compared to that of TST at 51.6% (95% CI: 40.9-62.2) at a cut-off of ≥ 10 mm. Secondly, test-retest reproducibility of QFT-GIT was high at 91.74% (ICC: 0.90; 95% CI 0.82-0.97). Thirdly, in this setting, some biological and operational factors that affected the performance of QFT-GIT were identified such as HIV positivity, low CD4+ T-lymphocytes, delayed incubation of blood samples and power outages. 8 For the main study, the study population at baseline consisted of 1,789 HHCs who were predominantly women (71%); median age was 28 years (IQR: 21-43); HIV positivity rate was 27.9%. Prevalence of tuberculous infection was 63.7% as measured by QFT-GIT and 39.6% by TST. There was a low level of agreement between the tests regardless of TST cut-off point (% agreement=59.7%; kappa=0.24). QFT+/TST- discordance (575/719; 80%) was more frequent than QFT-/TST+ discordance (144/719; 20%) at TST ≥10 mm. Risk factors associated with QFT-GIT positivity were identified at baseline. In multivariable analysis adjusted for sex, age, and community, HIV status was negatively associated with QFTGIT positivity (aOR: 0.48; 95% CI: 0.37–0.63; p<0.001) whereas residing in an urban area (aOR: 2.37; 95% CI: 1.10–5.13; p<0.03), smear status of index (OR: 1.26; 95% CI: 0.91-1.76; p=0.15) and country (aOR: 1.93; 95% CI: 1.48–2.51; p<0.001) were positively associated with QFT-GIT positivity. Similar results were obtained for TST. From a total of 1789 HHCs seen at baseline, 1113 (62.2%) HHCs entered follow-up and were included in the main analysis. The overall incidence rate of TB was 20.96/1000 pyrs (95% CI: 15.93-27.58). TB incidence rate was higher among test positive HHCs compared to those who were negative (IRR for QFT-GIT: 1.65; 95% CI: 0.86-3.37; p=0.06) and for TST (IRR: 1.88; 95%CI: 1.04-3.41; p=0.01). Results were similar in univariable analysis (QFT-GIT: 1.66 (95%CI: 0.88-3.11; p=0.11) and TST: 1.89 (95%CI: 1.09-3.28; p=0.02)) and multivariable analysis adjusted for sex, age and HIV (QFT-GIT: 2.20 (95%CI: 1.14-4.25; p=0.02) and TST: 2.19 (95%CI: 1.24-3.86; p=0.007)). Overall, PPV for QFT-GIT was 5.38% (95%CI: 3.84-7.31), compared to TST, 6.57% (95% CI: 4.41- 9.36). Overall for QFT-GIT, the IRR was higher among HIV negative HHCs (IRR: 3.85; 95%CI: 0.90-34.51; p=0.07) compared to HIV positives (IRR; 1.93; 95%CI: 0.88-4.57; p=0.04). Overall for TST, the IRR for HIV negatives (IRR: 2.21; 95%CI: 0.78-6.72; p=0.05) was similar to that among HIV positives (IRR: 2.32; 95%CI: 1.09-5.00; p=0.009). Univariable analysis showed similar results for both tests. In multivariable analyses adjusted for age, sex and country as an effect modifier, the HR for developing TB was 4.72 (95%CI: 1.35-16.46; p=0.01) in HIV positive QFT-GIT positives compared to 2.13 (95%CI: 0.81-5.60; p=0.12) in HIV positives TST positive HHCs. Risk factors for TB were identified. In multivariable analyses, adjusted for age, sex , HIV status and country there was strong evidence that occasional smoking, (HR: 4.07; 95%CI:1.31-12.63), HIV positivity (HR: 4.60; 95%CI:2.48-8.56), smear positivity of the index (HR: 2.00 ; 95%CI:1.04- 3.87) and country (HR: 1.79 ; 95%1.02-3.15; p=0.04) ) were associated with incidence of TB. Out of the 1,113 HHCs who entered follow-up, 406 HHCs had IFN-γ levels <0.35 IU/ml and were excluded leaving 707 HHCs in analysis for the primary objective. Out of these 536 (75.8%) had IFN-gamma levels ≥ 0.35 and <10 IU/ml (low IFN-γ levels) while 171 (24.2%) HHCs had ≥ 10 IU/ml (high IFN-γ levels). Out of the 707 HHCs that entered follow-up, 38 (5.4%) HHCs developed active TB over 1558.0 person-years (pyrs) of follow-up, giving an incidence rate of 24.39/1000 pyrs (95% CI: 17.75- 33.52).TB incidence rates were 24.51/1000 pyrs (9 cases/367.2 pyrs) in HHCs with high levels and 24.35 (29 cases/1190.7 pyrs) among those with low levels of IFN-γ, giving an IRR of 1.0 (95% CI: 0.42-2.18; p=0.48). Overall, unadjusted HR in HHCs with high IFN-γ levels was 1.02 (95%CI: 0.48-2.15; p=0.96) while in multivariable analysis adjusted for age, sex, country and HIV as an effect modifier, HR was 1.74 (95%CI: 0.63-4.79; p=0.29). TB incidence rates in HIV positives was 51.94/1000 pyrs (3 cases/57.8 pyrs) in HHCs with high levels and 65.29/1000 pyrs (19 cases/291.0 pyrs) among those with low levels of IFN-γ, giving an IRR of 0.79 (95%CI: 0.15-2.70; p=0.38).TB incidence rates in HIV negatives were 19.56/1000 pyrs (6 cases/306.7 pyrs) in HHCs with high levels and 11.47 (10 cases/871.7 pyrs) among those with low levels of IFN-γ, giving an IRR of 1.70 (95%CI: 0.51-5.18, p=0.16). Unadjusted HR among HIV negative HHCs was 1.73 (95%CI: 0.63-4.77; p=0.29) and 0.75 (95%0.22-2.55; p=0.65) among HIV positive ones respectively. In multivariable analysis adjusted for age, sex and country, the HR remained similar as unadjusted analysis for both HIV negatives and positives. For all the groups used for sensitivity analysis of the primary question, HHCs with the highest IFN-γ levels had increased IRRs ranging from 1.5 to 2 compared to the reference sub-group. For HIV negatives, HHCs with the highest IFN-γ levels had the highest IRRs in all groups apart from one group. HIV negative HHCs with the highest IFN-γ levels had increased IRRs ranging from 4 to 5-fold compared to the reference sub-group. In comparison, HIV positive HHCs with the highest IFN-γ levels had increased IRRs ranging from 1.6 to 2.6 compared to the reference sub-group. Conclusions: The principal finding in this study is that there was no difference in incidence rates between HHCs with low and high levels (overall IRR: 1.0 (95% CI: 0.42-2.18)). Another principal finding was that there was strong evidence of a five-fold increased risk of TB in HIV positive QFT-GIT positive HHCs compared to HIV positive QFT-GIT negative ones (aHR : 4.72; 95%CI: 1.35-16.46; P=0.01). For all the groups used in the sensitivity analysis of the primary question, HHCs with the highest IFN-γ levels had increased IRRs ranging from 1.5 to 2 compared to the reference sub-group. The feasibility studies emphasized the need for stringent sample collection and processing techniques to ensure the accuracy of QFT-GIT results.

The five-­‐year survival rate of patients with head and neck squamous cell carcinoma (HNSCC) has remained stable at 50% over the past five decades. Consequently, new treatment options are required. In approximately 90% of cases, over-­‐expression of the tumour associated antigen ErbB1 is seen. T4 immunotherapy retargets T-­‐cells against the extended ErbB-­‐receptor family and could be beneficial for HNSCC patients. T4 immunotherapy comprises the combined expression of the ErbB-­‐targeting chimeric antigen receptor T28ζ and the chimeric cytokine receptor 4αβ. Human T4+ T-­‐cells have a potent anti-­‐tumour effect. However, ErbB expression is not exclusive to tumour tissue, raising the concern of toxicity in healthy tissue. Here, I have investigated the potential toxicity of T4 immunotherapy in a SCID/Beige immunodeficient mouse model. Human T4+ T-­‐cells are activated by mouse ErbB receptors and consequently destroy both healthy and transformed mouse cells. Intravenous or intra-­‐tumoural T4+ T-­‐cell administration did not result in any clinical or histopathological toxicity. However, intraperitoneal T4+ T-­‐cell administration resulted in severe cytokine release syndrome (CRS). Target recognition in the peritoneal cavity resulted in elevated levels of serum human IL-­‐2 and IFNγ, as well as mouse IL-­‐6. The severity of CRS is hypothesized to be due to a combination of the T4+ T-­‐cell dose, magnitude of target recognition, and macrophage content within the peritoneal cavity. In keeping with this, macrophage depletion ameliorates both IL-­‐6 production and toxicity. Together, these data show that the SCID/Beige mouse is an adequate model to study T4 immunotherapy related toxicity. Furthermore, these results suggest that there may be a window for therapeutic application of T4+ T-­‐cells since anti-­‐tumour efficacy has been demonstrated at lower cell doses without the induction of toxicity. These findings, support progression to a Phase-­‐I clinical trial in which patients with locally recurrent HNSCC are treated with intra-­‐tumoural T4+ T-­‐cells.

End of life approaches in patients with cancer bodily deterioration and difficulties with everyday life can evoke psychological-existential distress. Dignity therapy (DT), a form of psychotherapeutic intervention, has been shown to alleviate psychological-existential distress in dying patients in Western culture. This thesis explores the potential application of DT in Taiwan. Three distinct, but linked phases of research, were conducted. Phase one elucidated the concept of dignity in Western culture through the technique of meta-ethnography. This revealed that psychological-existential adjustment strategies aimed at helping patients accept the inevitability of dying might preserve patients’ sense of dignity. In order to identify the conceptualisations of dignity in Taiwan, phase two comprised semi-structured interviews with Taiwanese dying patients (n=9) and palliative care professionals (n=10). These revealed that intrinsic to preserving a sense of dignity in this context were maintenance of moral life, having peace of mind, and a sense of existence. Findings supported the assertion that a modification of DT entitled dignity intervention (DI), making a life story legacy via a narrative approach, might help patients gain peace of mind and a sense of existence. Phase three consisted of a pre-test post-test single group evaluation with measurement of dignity, suffering, psychological symptoms, existential well-being and spiritual relationships, and interviews with terminally ill patients (n=14), their families (n=10) and palliative care professionals (n=9) in order to test the feasibility and acceptability of DI. No improvement in scores on outcome measures was recorded, but all participants reported DI to have been helpful. Patients used DI to convey messages regarding love, regret, gratitude and hope.

This thesis analyses the organisational response to resurgent tuberculosis (TB) across London. Tuberculosis in modern London reached its lowest recorded rate in 1987, but since that time there have been almost annual year on year increases, with the rate climbing by 50% between 1999 and 2009 alone. The rate of drug resistant TB is also increasing at a worrying pace. This research uses a novel complexity theory approach to analysis but empirically finds that positive features of complexity theory were crowded out by an embedded New Public Management paradigm. This study is qualitative and narrative-based, using complexity theory as the main theoretical framework, but also applies the theory of professional dominance and the paradigm of New Public Management (NPM) as possible alternatives. Institutionalism/archetype theory and Kingdon’s (1995) theory of public policy development are also introduced to help theorise the findings. This research found that complexity theory offers a useful, but partial, means of understanding the system responsible for TB control in London. Self-organisation, the key feature of complexity theory, was evident, but often resulted in maintaining the status quo and resisting change, in addition to infrequently resulting in innovation. The effects of highly embedded NPM practices and principles were wide-spread and powerful; its relentless preoccupation with risk aversion and control may have thwarted potentially positive benefits from self-organisation at the system level. Further, extensive NPM-inspired fragmentation almost eliminated co-adaptation (another complexity theory precept) by the TB control system to its changing environment, and reduced system fitness and robustness. TB control was also found to occupy a lowly place in terms of public health priorities. Finally, and rather surprisingly, there was little evidence of professional (medical) dominance observed. Medical consultants, stretched for time and faced with competing priorities, often acquiesced to NHS management efforts.

The Fanconi Anemia (FA) DNA repair pathway is essential for the recognition and repair of DNA interstrand crosslinks (ICL). Inefficient repair of these ICL can lead to leukaemia and bone marrow failure. The vital step of the pathway is the monoubiquitination of FANCD2 and FANCI by the RING E3 ligase FANCL, part of a core complex of proteins. This process requires the co-ordination of 3 proteins, the E1 activating enzyme, E2-conjugating (Ube2T) and E3 ligase (FANCL). Once FANCD2 and FANCI are modified with ubiquitin, they co-localise on the chromatin at the site of damage with other repair factors such as BRCA2, and the ICL is removed. A further DNA repair process, translesion synthesis (TLS), works alongside the FA pathway. The key step in TLS is the monoubiquitination of PCNA by the E2/E3 pairing of Ube2B(Rad6) and Rad18. However, the molecular and mechanistic details of how these two DNA repair pathways converge are poorly understood. The data presented will demonstrate that the E2-like fold (ELF) domain within FANCL is able to interact non-covalently with ubiquitin. This interaction is not necessary for the recognition of ubiquitin on the Ube2T or on FANCL itself, and is not required for catalysis in vitro. Mutation of the interaction in cells lead to a loss of FANCD2 monoubiquitination, but this is not due to a loss of core complex formation. Finally I show that the interaction between the ELF domain and ubiquitin may mediate the enhancement of FANCD2 monoubiquitination seen by the inclusion of monoubiquitinated PCNA. The observations provide critical mechanistic insights into the processes involved in monoubiquitination and the molecular basis for the observed link between TLS and activation of the FA pathway.

Prostate specific antigen (PSA) has been used in the diagnosis and monitoring of prostate cancer for almost 20 years. Most men who present with metastatic prostate cancer have markedly elevated serum levels of PSA. However, approximately 1% of cases have serum PSA levels that are much lower than the tumour burden would suggest - so-called "PSA-Negative" tumours. Their diagnosis may be delayed, and management compromised. Little is known about this patient group. The aim of this study was to improve the understanding and management of "PSA-negative" prostate cancer. The clinical history and tissue from 33 patients who presented with treatment-naive metastatic prostate cancer and a serum PSA < 10 ng/ml were included in this study, the largest series so far reported. Clinical and immunohistochemical features were defined and alternative biomarkers investigated. Potential mechanisms underlying PSA-negativity were explored using prostate cancer cell lines and archival tissue. From the clinical case notes review, patients presenting with low serum PSA and metastatic prostate cancer have a similar pattern of disease to men with high PSA prostate cancer. However, response duration to first line hormonal treatment and overall survival were shorter. Immunohistochemistry performed on archival prostatic tissue has shown that the majority of the cancers are positive for PSA, despite low serum levels. The extent of PSA immunostaining is patchy and could be missed on biopsy. PSMA and AR are expressed, however, and represent alternative diagnostic aids. The study indicates that PSMA and PAP should be explored as potential serum biomarkers in this patient group. The androgen receptor (AR) remains expressed in over 90 % of these cases and therefore defects in this pathway are unlikely to explain the low serum PSA levels. Neither loss of heterozygosity nor gene methylation of AR or PSA appear to be mechanisms underlying low serum PSA levels.

The central aim of cell division is the accurate transmission of replicated genetic material to daughter cells. To enable this segregation, centimetre-long DNA molecules must be organised into condensed micrometre-sized chromosomes. This critical but poorly understood process is principally effected by the chromosomal condensin complex. Condensin is a multisubunit protein complex, comprising a core dimer of ATPases of the structural maintenance of chromosomes (SMC) family. However, the role of the condensin ATPase in chromosome condensation has remained unclear. Using specific structure-based point mutations, along with quantitative measurements of chromosome condensation, and novel conditional alleles of condensin in the eukaryotic model budding yeast Saccharomyces cerevisiae, I show that the ATPase activity of condensin is crucial for its function. Mutations in the ATPase domain alter the dynamic DNA binding properties of condensin, and compromise its ability to form compact mitotic chromosomes. Taken together, these results shed light on critical events in the assembly and faithful segregation of mitotic chromosomes.

Small intestinal neuroendocrine tumours (SI NETs) are the most common malignancy of the small intestine, however they remain poorly characterised and the underlying pathogenic mechanisms driving disease development have yet to be elucidated. Whole genome and exome sequencing has suggested SI NETs to be mutationally quiet, with the most frequent mutation in Cyclin Dependent Kinase 1B occurring in only 8% of tumours, suggesting mechanisms other than genetic mutations may be responsible for driving SI NET tumourigenesis. Using integrated genomic and epigenomic analysis three distinct somatic copy number alteration (SCNA) profiles of SI NET were identified. The largest subgroup characterised by loss of heterozygosity at chromosome 18, negative CpG island methylator phenotype (CIMP) status, and the presence of CDKN1B mutations, is associated with improved clinical outcomes. A novel Multiple-SCNA signature has been described which defines a smaller subgroup of SI NETs and is characterized by significantly (p=0.04) reduced progression-free survival. A panel of 21 recurrently epigenetically dysregulated genes has been identified, and these represent putative novel pathogenic drivers for SI NET tumourigenesis and candidate novel biomarkers. Epigenetically dysregulated genes identified at a recurrence rate of 80-100% include gastric inhibitory polypeptide receptor (GIPR)(73.5%) – a target for novel imaging techniques in NETs, CDX1 (85.7%), CELSR3 (83.7%), FBP1 (83.7%), PCSK1 (67.3%) and TRIM15 (63.3%). The utility of methylated circulating tumour DNA analysis, and molecular profiling of circulating tumour cells as novel non-invasive biomarkers in SI NETs have been demonstrated. This is the first comprehensive integrated molecular analysis of SI NETs, providing evidence for epigenetic rather than mutational events in addition to SCNAs as drivers of SI NET development. These findings will facilitate improved patient management, treatment selection and prognostication.

Human cells employ a variety of mechanisms to repair DNA damage and maintain genomic integrity. Structure-specific endonucleases play crucial roles in the repair of DNA lesions by removing secondary DNA structures that can lead to chromosome mis-segregation, aneuploidy and cancer. MUS81 is the catalytic subunit of two human structure-specific endonucleases, MUS81-EME1 and MUS81-EME2. MUS81 nuclease has been shown to function in the resolution of homologous recombination (HR) intermediates, in the repair of stalled replication forks (RFs) and in the maintenance of telomere length in alternative lengthening of telomeres (ALT)-positive cells. However, it is unknown whether the two MUS81 complexes differ in their biological functions. To address this question, we carried out in vivo and in vitro studies to determine the roles of MUS81-EME1 and MUS81-EME2 in human cells. We found that EME2 interacts with MUS81 preferentially during the S-phase of the cell cycle, when MUS81 is important for the repair of stalled RFs. Cells treated with HU and depleted of MUS81 or EME2, but not of EME1, showed high levels of chromosomal aberrations, suggesting that the MUS81-EME2 complex is important for the maintenance of genomic stability following HU exposure. Also, depletion of EME2 or MUS81, but not of EME1, resulted in telomere loss, decreased rate of telomere sister chromatid exchanges (T-SCEs) and increased telomere fragility in ALT-positive cells. Consistent with the in vivo functional differences, we found that purified MUS81 complexes have different DNA substrate specificities in vitro, with the activity of MUS81-EME2 being 10-fold greater than that of MUS81-EME1. Together, these results indicate that MUS81-EME1 and MUS81-EME2 have different biochemical properties and distinct, non-overlapping functions, with MUS81-EME2 being important in the repair of stalled RFs and in telomere maintenance of ALT cells, whereas MUS81-EME1 plays an important role in the resolution of HJs in the context of HR-mediated DNA repair.