Prostate cancer is the second most commonly diagnosed cancer in men worldwide. Because prognosis can vary depending on tumour stage, precise diagnosis is vital. Circulating tumour cells (CTC) detach from primary and secondary tumour sites into the bloodstream. Changes in three-dimensional (3D) nuclear organization are associated with different types of cancer and were examined in this study in CTCs of high-risk prostate cancer patients. CTCs were isolated from 3mL of patient blood samples of 20 high-risk prostate cancer patients before treatment; after neoadjuvant androgen deprivation therapy (ADT) but before radiotherapy (RT); and after completing RT. Telomere-specific fluorescence in situ hybridization was performed on filters containing cells, 3D images of 30 CTCs per filter were analysed. Changes in telomere organization were observed post ADT and RT; patients fell into three groups depending on the change in CTC telomeric profiles in response to ADT. These groups displayed responses characteristic to each group upon delivery of RT. 3D nuclear telomere profiles in CTCs post-ADT may indicate both ADT response and predict RT response in high-risk prostate cancer. / October 2016

Prostate cancer is the most common cancer occurring in males. The identification of novel microRNAs (miRs) that contribute to tumor progression represents prospective treatment targets. miRs are small non-coding RNAs important in gene regulation with specific tissue expression patterns. Each miR is thought to affect the expression of hundreds of different RNA targets. Two putative oncomiRs, miR-155 and miR-146a, were shown to be differentially expressed in the human derived, prostate cell sublines M12 and F6. Quantification of endogenous miR expression showed high levels in the metastatic M12 cell line versus low in its weakly tumorigenic F6 variant. The restoration of miR expression to M12 levels was evaluated on F6 growth, morphology, and in vitro behavior. F6 plus miR-155 or miR-146a displayed increased growth, motility and invasiveness when compared to M12, with less organized structural morphology when grown embedded in matrigel. Altogether these results suggest that the overexpression of miRs 155 and 146a could contribute to tumor progression in vivo.

Master of Science / Department of Kinesiology / Steven W. Copp / Cancer patients have a reduced exercise capacity compared to age-matched healthy counterparts which contributes to premature fatigue. The reductions in exercise capacity are multifactorial and vary depending on the type of treatments and the specific cancer. Given that cancer treatments have been shown to impair cardiovascular and/or skeletal muscle function, it is difficult to determine if cancer itself reduces exercise capacity. We used a rat prostate tumor model to test the hypothesis that cancer independently reduces endurance exercise capacity. Methods: In male Copenhagen rats (COP/CrCrl), an initial treadmill test to exhaustion was used to determine endurance exercise capacity. Subsequently, the prostates of the rats were injected with either prostate carcinoma cells (R-3327 AT-1) in Matrigel (cancer: n = 9) or Matrigel only (sham: n = 7). Treadmill tests to exhaustion were repeated four and eight weeks post-surgery. Results: Time to exhaustion decreased over the course of the experimental protocol in both the sham and cancer groups. However, the overall reduction in time to exhaustion in the cancer group (-16.7 ± 1.9 min) was significantly greater (p = 0.038) than the sham group (-10.1 ± 2.2 min). Despite no differences in total body mass at the end of the experimental protocol, heart, left ventricle, and gastrocnemius muscle mass were significantly lower in the cancer group compared to the sham group (p < 0.05 for all). Moreover, within the cancer group heart and left ventricle mass, but not gastrocnemius mass, were significantly inversely correlated with prostate tumor mass. Conclusion: Endurance exercise capacity was reduced in rats with untreated prostate cancer to a greater extent than it was reduced in sham operated rats. Although multiple mechanisms likely contributed to the reduced exercise capacity, reductions in heart and gastrocnemius muscle mass likely played an important role.

Replication-selective oncolytic adenoviruses represent a promising anticancer approach with proven efficacy in cancer cell lines and tumour xenografts in vivo. Anti-tumour efficacy, both in preclinical studies and clinical trials, was significantly improved in combination with chemotherapeutics in numerous cancers, including prostate cancer. It has been established that expression of the viral E1A gene is essential for the enhancement of cell killing in combination with cytotoxic drugs. The overall goal of this project is to identify specific E1A gene regions involved in the sensitization to the cytotoxic drugs mitoxantrone and docetaxel, the current standard of care for late stage prostate cancers, to enable the development of improved anti-cancer therapies. Specific regions in the E1A proteins bind to numerous cellular factors to regulate the host cell function and the viral life cycle, including the p300, p400 and pRb family proteins. This work was aimed at determining the mechanisms involved in the synergistic cell killing in prostate cancer cells in response to the combination of the replication-selective (oncolytic) mutant AdΔΔ with cytotoxic drugs. Previous findings suggested an enhancement of drug-induced apoptosis. I found that the small E1A12S protein, unable to induce viral replication, is sufficient to sensitize the prostate cancer cells, 22Rv-1 (AR+), and PC-3 and DU145 (AR-), to drugs. The non-replicating AdE1A12S-mutant AdE1A1104 (defective in p300-binding) could not sensitize the cells while mutants with intact E1A-p300 binding (AdE1A12S, AdE1A1102, AdE1A1108) and defective in p400- (AdE1A1102) or pRb-binding (AdE1A1108) potently sensitized all tested cell lines. In fact, all mutants except AdE1A1104 potently synergised with mitoxantrone and docetaxel to kill the prostate cancer cells. When comparing the non-replicating E1A12S mutants with the corresponding replicating E1A-deletion mutants (expressing E1A12S and 13S) synergy was demonstrated with all replicating mutants except dl1104, which caused an additive effect with mitoxantrone. We hypothesised that the synergistic cell killing is the result of pathway convergence through E1A-p300 and mitoxantrone-activated DNA-damage/apoptosis events. To address this I employed an extensive miRNA array screen to identify potential pathways. Several miRNAs were found to be differentially regulated in response to the combination of AdE1A12S with mitoxantrone compared to each single agent treatment. The majority of these miRNAs are reported to be part of cell death and survival pathways (e.g. apoptosis and autophagy) and to be differentially regulated in prostate cancer. To further investigate the role of these pathways, I determined changes in expression levels of key proteins that had previously been suggested to be targeted by the identified miRNAs, thereby preventing translation of the respective mRNAs. The greatest changes in protein levels in response to AdE1A12S and mitoxantrone were observed for Bcl-2, p-Akt, LC3BII and p62. Finally, I verified similar mechanisms of action when the oncolytic AdΔΔ was combined with mitoxantrone under synergistic conditions. These findings will direct future investigations aimed at dissecting the mechanisms of action for virus-induced sensitization to cytotoxic drugs and may aid in the development of improved therapies for prostate cancer by design of novel oncolytic mutants and combination strategies and/or identification of targets for small molecules inhibitors.

Androgen receptor (AR) cell signalling is active in most castration-resistant prostate cancer (PCa) tumours and suppression is hypothesized to impede cell proliferation. Hey1, a corepressor of AR is being investigated as a therapeutic transgene for late-stage PCa. A replication-defective recombinant adenovirus deleted for E1 and E3 and expressing Hey1 under a CMV promoter was constructed (Ad5Hey1). A dual luciferase reporter system demonstrated that Ad5Hey1 repressed AR activity in a dose dependent manner in miboleronestimulated 22Rv1 cells. Ad5Hey1 was cytotoxic in both AR-positive 22Rv1 and LNCaP and AR-negative DU145 cells. The doses required to kill 50% of cells (EC50) were comparable to those of AdE1A12S expressing the cytotoxic E1A12S gene from an identical vector. The mechanisms of Ad5Hey1-induced cell killing were investigated in 22Rv1 and DU145 cells. Using RNA interference towards AR or p53 in 22Rv1 cells we concluded both proteins were required for optimal cell killing by Ad5Hey1. In DU145 cells, with non-functional p53, Ad5Hey1 decreased levels of phospho- STAT3 and total STAT3 suggesting Ad5Hey1 might inhibit STAT3 signalling while the JAK1/2 inhibitor, AZD1480 was ineffective at sensitising DU145 cells to Ad5Hey1. Preliminary data therefore suggests Ad5Hey1 may interfere with JAK/STAT signalling in these cells. Cell-killing efficacy with Ad5Hey1 in combination with cytotoxic drugs currently used in the clinic for the treatment of late-stage PCa, mitoxantrone and docetaxel, resulted in a synergistic enhancement of cell death in 22Rv1 and DU145 cells. LNCaP cells were also sensitised to the drugs. Characterisation of the mode of cell killing demonstrated augmented mitochondrial membrane depolarisation and caspase-3 activation when combined with docetaxel in all cell lines and with mitoxantrone in 22Rv1 and LNCaP cells, typical of apoptotic death. In DU145 cells, the combination of Ad5Hey1 with mitoxantrone decreased the proportion of apoptotic cells suggesting cells are dying by alternative cell death mechanisms. In this thesis I have demonstrated that Ad5Hey1 potently eliminates PCa cells both in the presence and absence of functional AR or p53, and that cell killing is 6 improved in combination with cytotoxic drugs. I demonstrate that the mechanisms by which Ad5Hey1 acts as a cell death enhancer is mainly through cooperation with drugs on apoptotic pathways while other factors such as inhibition of survival are also involved. In conclusion, these data suggest that it is feasible to develop a future replication-selective adenovirus expressing Hey1 as a cytotoxic transgene to improve antitumour efficacy in vitro and in vivo, especially in combination with apoptosis-inducing drugs.

Orientador: Silvia Regina Rogatto / Banca: Spencer L. M. Payão / Banca: Carla Rosemberg / Banca: José Carlos de S. Trindade / Banca: Maria Aparecida M. Rodrigues / Resumo: O câncer de próstata (CaP) é a neoplasia mais comumente diagnosticada entre homens no ocidente. Embora tratamentos efetivos para a doença localizada estejam disponíveis atualmente, não há terapia curativa para tumores metastáticos. Além disso, os marcadores diagnósticos utilizados na clínica não conseguem discriminar totalmente a evolução diferencial da doença. Desta forma, o conhecimento das diferenças biológicas entre tumores primários confinados ao órgão e metástases é essencial para o desenvolvimento de novos marcadores e identificação de alvos terapêuticos. Neste estudo a análise baseada na metodologia de HR-CGH cromossômico foi realizada para identificar alterações de ganhos e perdas genômicas em três grupos de amostras: o grupo I, que compreende amostras pareadas de tumor primário e respectivas metástases (11 casos); o grupo II, constituído de pacientes que apresentaram seguimento clínico favorável por mais de 10 anos (5 casos); e o grupo III, constituído por diferentes biópsias do mesmo paciente (5 pacientes com 2 biópsias cada). As amostras foram microdissecadas (amostras a fresco: a partir de lâminas de referência; em blocos de parafina: a laser) e após a obtenção de DNA foram amplificadas (amostras de arquivo: PCR-SCOMP) ou marcadas por nick-translation para a realização de HR-CGH. Os resultados de HR-CGH foram comparados com os dados obtidos da análise de CGH-array num subgrupo de amostras e revelaram concordâncias significativas. Os resultados obtidos na presente investigação revelaram perdas dos cromossomos 1p, 2, 3q, 4p, 5q, 7, 8, 9q, 10q, 11q, 12q, 14q, 15q, 16q, 17q, 18q, 19, 20q e 22q em 80% dos casos avaliados. Além disso, perdas em 17q11.2-25, por exemplo, foram detectadas exclusivamente nos tumores do grupo I e nas suas metástases, e não nos tumores do grupo II, sugerindo que esta alteração deve ser importante... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Prostate cancer (PCa) is the most commonly diagnosed non-cutaneous malignancy and the second leading cause of cancer mortality in men from Occident. Although effective treatments for the localized disease are available, there is no efficient therapy for metastatic tumors. Additionally, clinical diagnostic markers are not able to completely discriminate the differential evolution of the disease. The knowledge of biological differences between localized primary tumors and metastasis can establish new molecular markers and therapeutic targets. In this study, an analysis based on HR-CGH methodology was performed to identify imbalances genomic in three groups of samples: group I, paired samples of primary tumors and its metastasis (11 cases); group II, patients that exhibited favorable follow-up over 10 years (5 cases); and group III, different biopsies from the same patient (5 patients with 2 biopsies each). The tumor samples were submitted to microdissection procedures (fresh samples: from reference slides; paraffin embedded samples: laser), DNA extracted and amplified (archive sample: PCR-SCOMP) or labeled by nick-translation to HR-CGH. The HRCGH results were compared with data obtained from CGH-array analysis of a subgroup of samples and revealed significant concordances. In the present investigation, there were observed losses on chromosomes 1p, 2, 3q, 4p, 5q, 7, 8, 9q, 10q, 11q, 12q, 14q, 15q, 16q, 17q, 18q, 19, 20q and 22q in 80% of the cases. Losses in 17q11.2-25, for instance, were detected exclusively in tumor from group I and its metastasis, but were not found in tumors from group II, suggesting that this alteration must be important in the progression of the disease. Five genes were selected after the comparison between the HR-CGH and CGH-array data. The tumor suppressor genes ARID1A, MTSS1, NME1 and S100A4 and TOP2A (oncogenes) were evaluated by quantitative real time... (Complete abstract click electronic access below) / Doutor

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Xu, Dandan. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 209-231). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

The aim of this thesis is to describe how men with localised prostate cancer make decisions about their medical treatments, to describe the psychological and decision-related adjustment of these men over time, and to identify what variables predict decision-related adjustment. Chapter 1 reviews the medical context of localised prostate cancer and factors that influence men's decision making in this context. It is concluded that owing to ongoing uncertainty about the optimal medical treatment for this cancer and the substantial negative quality of life effects of treatments, how men make decisions in this context is an important research question. Further, although men with prostate cancer are high seekers of medical treatment information, knowledge about how men use such information and actually make this treatment decision is limited. Chapter 2 discusses research approaches currently applied to patient decision making: first, a social interaction approach encompassing the interaction between the patient and their physician and the social context influencing this interaction (Charles, Gafni, & Whelan, 1999); and second, normative decision theory (Shafir & Tversky, 1992; von Neumann & Morgenstern, 1947). The Heuristic-Systematic Processing model (Chaiken, 1980) is then proposed as a theoretical framework for investigating patient decision making that includes both systematic and non-systematic decision strategies. Chapter 3 reviews applied decision research in cancer, and presents an overview of research findings regarding patients' preferences for involvement in decision making, the relationship between decisional involvement and psychological adjustment, and decisional support interventions. Research on adjustment to cancer is discussed and the need for further research about men's psychological and decision-related adjustment after localised prostate cancer is identified. Finally, a multivariate analysis of decision-related adjustment for men with localised prostate cancer based on the stress and coping framework of Lazarus and Folkman (1984) is proposed. Chapter 4 describes Study 1 that was an experiment to investigate the utility of the Heuristic-Systematic Processing model (HSM) in explaining low desire for involvement in decision making about prostate cancer treatments as an example of use of the expert opinion heuristic. Using a hypothetical decision scenario about localised prostate cancer it was found that a low desire for involvement in decision making by men was predicted by a high belief in powerful others controlling health, a low belief in the self being responsible for good health, a high preference for black and white thinking, and a lower level of education. This study provides preliminary support for use of the HSM in this context and for the conceptualisation of decision deferral as the expert opinion heuristic. Chapter 5 introduces and describes the method of Study 2 that was a descriptive, prospective study of men's decision making after an actual diagnosis of localised prostate cancer. This method allowed for an analysis of men's decision making that includes both systematic and non-systematic processes, and for further investigation of the utility of the HSM in explaining decision behaviour. In addition, a multivariate approach was used to describe men's physical, psychological and decision-related adjustment over time, and to identify psychological predictors of decision-related adjustment. Chapters 6 describes men's use of systematic processing as limited and the use of non-systematic processes, such as lay beliefs and heuristics, as pervasive. It is concluded that patients do not utilise information about medical treatments in a comprehensive or systematic way when making treatment decisions and that patients' decision making is biased by their prior beliefs about cancer and health. A framework is outlined to demonstrate how the results of Study 1 and 2 support the application of the HSM to decision making about prostate cancer with particular reference to the role of beliefs about the physician, health locus of control, and uncertainty about the treatment decision in influencing decision strategies. Chapter 7 describes men's physical, psychological and decision-related adjustment over time, and concludes that decision-related distress is high but psychological distress in general is low. Decisional conflict at diagnosis and at twelve months concurrently, and at two months prospectively, was predicted by dispositional optimism; and this effect was mediated by the man's proximal cognitive appraisal of the impact of the cancer. It is concluded that decisional conflict is a person's cognitive judgment of the treatment decision that is generated by similar processes to that of the psychological distress that follows a cancer diagnosis. Conclusions and implications of these studies for future research in this area are summarised in Chapter 8.

Prostate cancer is one of the most prominent diseases in men’s health. It is inherently 'male', given the exclusivity of the prostate gland to men’s bodies and its physiological connection to testosterone and male sexuality. The biomedical complexities of prostate cancer continue to be unravelled and researched and are often connected to identifying causes, the virtues of screening and treatment modalities. However, despite the biological male 'sex' link, most of the prostate cancer research is not connected with research on gender relations, men and masculinities. The net outcome is that men’s lives and illness experiences are absent in much of the prostate cancer research. This PhD thesis Prostate cancer: Anglo-Australian heterosexual perspectives, is an ethnographic study of thirty-five Anglo-Australian men diagnosed with prostate cancer. Participants shared their experiences of living with prostate cancer in the context of health promotion, health services and in relation to their sexuality and intimate relationships. Through participant photographic novella and in-depth semi-structured interviews, rich cultural insights are provided. A social constructionist gender analysis is used in this research that shows how the social constructions of masculinity interconnect and occasionally collide with prostate cancer throughout the illness trajectory.