Prostate cancer is the most commonly diagnosed cancer and second leading cause of death in American men. Patients with localized prostate cancer may experience unique and multidimensional symptoms that are distressful from treatment and thereafter. This cross-sectional correlational study aimed to investigate the relationships among stress, symptoms, symptom distress, and symptom self-management and identify the effective strategies of symptom self-management in men with localized prostate cancer following prostatectomy or radiation therapy.Eight saliva samples and 3 questionnaires (Perceived Stress Scale, Symptom Indexes, and Strategy and Effectiveness of Symptom Self-Management) were obtained from each participant between 1 and 3 months following their first prostate cancer treatment. The sample consisted of 53 men with localized prostate cancer. Mean salivary cortisol concentrations for the entire sample ranged from 0.3 to 0.08 ug/dL. Cortisol was secreted in a circadian rhythm with heightened activity in the early morning and lowered activity late in the day. The circadian pattern of cortisol secretion was similar in both the prostatectomy and radiation therapy groups, although the values were slightly different. Two areas Under the Curve (AUC) of salivary cortisol were calculated. Three cortisol circadian rhythms were identified, but the majority of the sample had a typical negative consistent circadian rhythm.Patients with localized prostate cancer who underwent radical prostatectomy or radiation therapy had low perceived stress. Perceived stress was positively correlated with AUCg, noon salivary cortisol concentrations, and afternoon salivary cortisol concentrations. Subjects reported a moderate degree of symptoms and symptom distress on urinary, bowel, and sexual dysfunction 1-3 months following treatments. The most effective strategies of urinary symptom management were pad and kegel exercise; the most effective strategy of bowel symptom management was rest or endure; the most effective strategies of sexual dysfunction management included express their feelings or find alternative ways to express their affection. The symptom self-management strategies were significantly and positively correlated with symptom self-management effectiveness.Symptom distress and AUCg were significant and strong predictors of symptom self-management. Findings can help health care providers develop effective strategies for symptom self-management that enhance health related quality of life among men with localized prostate cancer.

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Vinclozolin (VCZ), an antiandrogenic fungicide, is an endocrine disrupting chemical that is known to possess high affinity for the androgen receptor (AR) and modulate expression of critical androgen-dependant genes in the prostate. In this study, viability and expression of AR, NKX3.1 and CYP3A4 genes were measured in androgen-sensitive prostate cells LNCaP after exposure to VCZ and VCZ treated with S9 microsomes in a time and dose dependent manner. NKX3.1 is an androgen regulated gene that plays a vital role in prostate development. CYP3A4 is involved in xenobiotic metabolism. VCZ decreased the viability at high doses after 48 hours which was slightly mitigated by treatment with S9 metabolites. Expression of NKX3.1 and CYP3A4 was upregulated while an initial downregulation of AR was observed. NKX3.1 upregulation corroborates with possibility of antiandrogens to act as androgens in LNCaP. The results illustrate that VCZ can interfere with the expression of critical prostate genes.

Prostate cell lines were derived from two regions of prostate tissue from the same patient. The objective was to produce cell lines (as a useful in vitro model) from these two different regions which exhibit different properties for carcinoma development. The tissue was obtained from patients suffering from benign prostate hyperplasia undergoing trans-urethral resection. Tissue was taken from the deep (peripheral) and superficial (peri-urethral) areas. The cells were immortalised by transduction with constructs over expressing the cdk4 and hTERT genes. These cell lines were then characterised for their cellular phenotypes utilized for radiation transformation studies and utilized to investigate the role of plant derived polyphenols on normal and tumour cells. The cell line from the superficial region (P21s) was treated to fractionated doses of gamma radiation and a transformed cloned cell line was derived (P21s 40Gy (clone-a)). The cell line from the deep region (P21d) was found to consist of a mixed population of abnormal cells and a transformed cloned cell line was derived from it (P21d 0Gy (clone-a). In an attempt to obtain a normal P21d cell line cloned cell lines from early passage P21d cells were established. All seven cloned lines were abnormal with an average of 80 chromosomes per cell, invasive using a Matrigel assay and produced anchorage independent colonies. All cell lines were fully characterised with immunocytochemistry, chromosome analysis, invasion assays, and anchorage independent colony formation. P21s expressed basal cell markers (cytokeratin 5 (CK5) and 14), were positive for stem cell markers (prostate specific stem cell antigen PSCA, CK6), positive for p16, p63 and telomerase expression and negative for c-Myc expression. P21s was not invasive in a Matrigel assay and did not produce anchorage independent colony formation. P21d and P21d 0Gy (clone-a) also expressed CK5, CK14, PSCA, CK6, and telomerase but not p16 or p63 and showed an increase in expression of nuclear c-Myc, highly invasive and produced anchorage independent colonies. P21s 40Gy (clone-a) expressed CK5, CK14, PSCA, CK6, telomerase and p63, produced anchorage independent colonies, and was weakly positive for c-Myc expression. Spectral karyotyping analysis (SKY) showed P21s had a normal chromosome complement except an additional chromosome 20 whereas the P21s 40Gy (clone-a), P21d and P21d 0Gy (clone-a) cell lines had an abnormal chromosome complement with P21d and P21d 0Gy (clone-a) cell lines expressing multiple copies of every chromosome including loss of the Y chromosome. These results were echoed in the single nucleotide polymorphism chip (SNP) results which showed P21s as normal but P21d and P21d 0Gy (clone-a) to have large deletion and amplification regions that correlated with the SKY analysis. No differential cytotoxic response was noted between normal and abnormal cell lines including prostatic carcinoma cell lines LNCaP and PC-3 following treatment with strawberry polyphenol compounds. Most reports of a cytotoxic response to tumour cells in the literature did not compare the response to normal cells and used established cell lines. Human lymphocytes were also tested and all compounds were toxic in high doses. Polyphenol and ellagitannin rich polyphenol fractions were very cytotoxic and the anthocyanin rich fraction less toxic. In contrast to the lack of a direct differential cytotoxic effect, plant polyphenols did produce a protective effect to a carcinogenic insult. However a protective effect was noted via micronucleus assay with 3 hour incubation with the polyphenol rich fraction prior to radiation treatment. Finally, the expression and association of metabolic enzymes within the cells cytosol were investigated. The P21s cells were found to express both isoforms of LDH and so thought to be able to metabolise anaerobically and aerobically. P21d and P21d 0Gy (clone-a) cells were found to only express one isoform in the complex and so it was assumed that these cells favoured anaerobic metabolism of ATP in correlation to the Warburg effect. c-Myc association with compounds in the cell cytosol of P21s cells existed whereas, abnormal cells lost this association along with up-regulation of c-Myc expression and down stream targets of c-Myc in the nuclei. Thus these newly established human prostate cell lines provide a useful model system for investigating the biology of the prostate and prostate cancer.

Tumour hypoxia has been demonstrated to cause development of an aggressive tumour phenotype and is associated with increased patient mortality and poorer response to treatments such as chemotherapy and radiotherapy. Previous studies have established that hypoxia exists within a nidus of prostate cancer. Based on the importance of the tumour microenvironment, especially hypoxia, in prostate cancer, the major aims of this thesis were to establish: (a) the role of a novel putative survival factor, dermcidin, in prostate cancer survival under hypoxia/oxidative stress; and (b) the effect of nitric oxide-donating non-steroidal anti-inflammatory drugs (NO-NSAIDS), a new class of drugs, on the killing of prostate cancer cells subjected to hypoxia. A wide-range of confirmatory, cellular and molecular biology techniques were employed in this thesis. The PC-3 hormone-insensitive prostate cancer cell line was used for the majority of studies as this cell line represents hormone-independent prostate cancer, treatment of which is currently palliative. Cell incubation at 0.2% oxygen for 48 hours was established as suitable conditions to stimulate the development of the hypoxia response. Upregulation of nuclear hypoxia-inducible factor-1α protein was the main marker used to assess the hypoxia response. Dermcidin messenger RNA production was found to occur in a range of prostate cancer cell lines; was upregulated in cell lines by both hypoxic and oxidative stress; and found to act as a proliferation, survival and pro-invasion factor under hypoxia and oxidative stress in immortalised prostate cancer cell lines. Furthermore, the portion of the dermcidin molecule responsible for the survival advantage was localised to the proteolysis-inducing factor core peptide subunit. However, subsequent analysis of primary cancer samples from prostate cancer patients revealed that dermcidin was not expressed in these tumours, although dermcidin mRNA was identified in analysis of other primary tumours. As such, the role of dermcidin in prostate cancer was not evaluated further in this thesis. Investigation of NO-sulindac (a NO-NSAID drug) in hypoxic PC-3 cells showed that these agents were significantly more pro-necrotic, pro-apoptotic and anti-invasive than traditional, unnitrated sulindac. NO-sulindac was found to downregulate the hypoxia response mounted by PC-3 cells under hypoxia via the Akt signalling pathway. Finally, analysis of the role of NO-sulindac in radiosensitising hypoxic PC-3 cells showed that NO-sulindac caused significant radiosensitisation under normoxia, but particularly in hypoxic conditions. As such, NO-NSAIDs show great promise as neoadjuvant, concurrent and adjuvant treatments for patients with hypoxic prostate cancer. The findings of this thesis illustrate several potential novel strategies for treatment of hormone-independent prostate cancer.

Background: The Selenium and Vitamin E Cancer Prevention Trial (SELECT) was a randomized, double-blind, placebo-controlled, prostate cancer prevention study funded by the National Cancer Institute and conducted by SWOG (Southwest Oncology Group). A total of 35,533 men were assigned randomly to one of four treatment groups (vitamin E + placebo, selenium + placebo, vitamin E + selenium, placebo + placebo). At the time of the trial's development, NIH had invested substantial resources in evaluating the potential benefits of these antioxidants. To capitalize on the knowledge gained from following a large cohort of healthy, aging males on the effects of selenium and/or vitamin E, ancillary studies with other disease endpoints were solicited. Methods: Four ancillary studies were added. Each drew from the same population but had independent objectives and an endpoint other than prostate cancer. These studies fell into two categories: those prospectively enrolling and following participants (studies of Alzheimer's disease and respiratory function) and those requiring a retrospective medical record review after a reported event (cataracts/age-related macular degeneration and colorectal screening). An examination of the challenges and opportunities of adding ancillary studies is provided. The impact of the ancillary studies on adherence to SELECT was evaluated using a Cox proportional hazards model. Results: While the addition of ancillary studies appears to have improved participant adherence to the primary trial, this did not come without added complexity. Activation of the ancillary studies happened after the SELECT randomizations had begun resulting in accrual problems to some of the studies. Study site participation in the ancillary trials varied greatly and depended on the interest of the study site principal investigator. Procedures for each were integrated into the primary trial and all monitoring was done by the SELECT Data and Safety Monitoring Committee. The impact of the early closure of the primary trial was different for each of the ancillary trials. Conclusions: The ancillary studies allowed study sites to broaden the research opportunities for their participants. Their implementation was efficient because of the established infrastructure of the primary trial. Implementation of these ancillary trials took substantial planning and coordination but enriched the overall primary trial.

CTCs have emerged as a potential source of clinical significance. But with numerous isolating systems currently available, the numbers of captured CTCs vary widely. At this point, CellSearch remains the only FDA-approved system with clinical significance whereby the results could be used to monitor patients with metastatic colon, breast, or prostate cancer. However, its inability to isolate CTCs from non-high risk prostate cancer patients or CTCs that are EpCAM-negative has led to criticism. In this study, we have shown that size-based filtration successfully isolates CTCs from patients with localized and metastatic prostate cancer. We have also shown that CTCs can be successfully isolated from low and intermediate risk groups. Additionally, clusters of CTCs were preserved and isolated in all localized risk groups and metastatic patients. Furthermore, we enumerated the isolated CTCs using automated and manual methods in low risk, intermediate risk, high risk, and metastatic prostate cancer. The automated and manual counts were comparable. Moreover, the amounts of clusters and the size of clusters correlated with the status and stage of prostate cancer. / October 2016

Cancer of the prostate (CaP) is the most common malignancy diagnosed in men in the Western society. During the last years, prostate specific antigen (PSA) has been used as a biomarker for CaP, although a high PSA value is not specific for CaP. Thus, there is an urgent need for new and improved diagnostic markers for CaP. In this thesis, the aim was to find a miRNA signature for diagnosis of CaP and to elucidate if differences in behavior between transition zone and peripheral zone tumors are reflected in miRNA expression. One of the major findings is anexpression signature based on nine miRNAs that with high accuracy (85%) could classify normal and malignant tissues from the transition zone of the prostate. The results furthermore show that the major differences in miRNA expression are found between normal and malignant tissues, rather than between the different zones. In addition, tumors arising in the peripheral zone have fewer changes in miRNA expression compared to tumors in the transition zone, indicating that the peripheral zone is more prone to tumor development compared to the transition zone of the prostate. A crucial step in pre-processing of expression data, in order to differentiate true biological changes, is the normalization step. Therefore, an additional aim of this thesis was to compare different normalization methods for qPCR array data in miRNA expression experiments. The results show that data-driven methods based on quantile normalization performs the best. The results also show that in smaller miRNA expression studies, only investigating a few miRNAs, RNU24 is the most suitable endogenous control gene for normalization. Taken together, the results in this thesis show the importance of miRNAs and the possibility of their future use as biomarkers in the field of prostate cancer.

Introduction: Cancer-related fatigue is often described by patients as a lack of energy, mental or physical tiredness, diminished endurance, and prolonged recovery after activity. Etiologic mechanisms underlying CRF are not well understood. Methods: A literature review was conducted to examine studies that had investigated the association of mitochondrial dysfunction with fatigue. The major conclusion from this review was that alterations in energy metabolism may contribute to fatigue. Therefore, the dissertation study focused on laboratory techniques for measuring mitochondrial oxidative phosphorylation enzymes (complexes I-V) and a mitochondrial-specific oxidative stress marker (superoxide dismutase 2 [SOD2]). The primary aim of the dissertation research was to describe levels of biomarkers of mitochondrial function, fatigue, and health-related quality of life (HRQOL) before and at the completion of external beam radiation therapy (EBRT) in men with nonmetastatic prostate cancer (NM-PC). To achieve this aim a secondary analysis of a descriptive, longitudinal study was conducted (#10-NR-0128). Results: A total of n = 22 men with NM-PC were included in this study. There were significant increases in fatigue and a significant decrease HRQOL from baseline to the completion of EBRT. However, there was no significant change in the biomarkers of mitochondrial function from baseline to EBRT completion. Given the exploratory nature of the study, it was decided to further investigate the patient sample to understand the relationship of fatigue and mitochondrial function in a well-characterized fatigue phenotype. There was preliminary evidence to support the possibility of distinct patterns of mitochondrial enzyme levels between those with a high intensification of fatigue and those with a low intensification of fatigue during EBRT; however, these differences were not statistically significant. Discussion: To our knowledge, this is the first study to describe the relationship between mitochondrial enzymes and fatigue before and during EBRT in men with NM-PC. The most important preliminary finding from this study is the possibility that mitochondrial enzymes might be related to fatigue intensification during EBRT. Future studies will be critical to determine if these preliminary findings are replicable, and if so, whether there are potential therapeutic targets in individuals at highest risk for fatigue intensification during EBRT.

The chemokine receptor CCR5 has been implicated in the pathogenesis of prostate cancer. A novel natural product, anibamine, was isolated and found to be a micromolar inhibitor of the receptor. Anibamine was used as a new anti-prostate cancer lead compound. To discover the pharmacophore, analogs of anibamine were designed using the “deconstruction-reconstruction-elaboration” approach and synthesized. The establishment of a stereoselective route to only one isomer was explored, to increase yield and eliminate elaborate purification procedures. Analogs were found to have anti-prostate cancer activity at levels higher than the parent compound. The molecular modeling studies of the deconstructed analogs indicate that due to the psuedo-symmetry of the parent compound, the binding conformation of the deconstructed analogs may not be very different from each other. All this information together may help identify a next generation lead compound for anti-prostate cancer treatment.