Background: This thesis is composed of three studies. In the first paper, we tested the association of metformin use with prostate cancer incidence. In the second paper, we examined the association of metformin use with all-cause and prostate cancer specific mortality. The final paper explored the benefit of detailed pathology review to predict mortality among diabetic men with prostate cancer. Methods: A total of 5306 incident diabetic men older than 66 who subsequently developed prostate cancer were identified using the Ontario Diabetes Database and the Ontario Cancer Registry between 1994-2008. The association of metformin use and risk of prostate cancer and its grade was tested with a nested case-control design using a conditional logistic regression model. We used a cohort design with a time dependent Cox-proportional hazard model to examine the association of metformin use and mortality. Finally, we employed a c-statistic and Net Reclassification Improvement analysis to study the impact of pathology abstraction on predicting mortality. Results: The data suggest metformin use was not associated with the risk of prostate cancer or its grade at presentation. However, each additional 6 month of metformin use was associated with a 24% decrease in prostate-cancer-specific and 8% decrease in all-cause mortality. Pathology abstraction improved the accuracy in predicting all-cause and prostate-cancer specific mortality. Conclusions: In our study metformin use was not associated with a decreased risk of prostate cancer, but had a significant impact on all-cause and prostate cancer specific mortality. These results may serve as proof of concept in designing an interventional study of metformin to delay progression in prostate cancer.

Background: This thesis is composed of three studies. In the first paper, we tested the association of metformin use with prostate cancer incidence. In the second paper, we examined the association of metformin use with all-cause and prostate cancer specific mortality. The final paper explored the benefit of detailed pathology review to predict mortality among diabetic men with prostate cancer. Methods: A total of 5306 incident diabetic men older than 66 who subsequently developed prostate cancer were identified using the Ontario Diabetes Database and the Ontario Cancer Registry between 1994-2008. The association of metformin use and risk of prostate cancer and its grade was tested with a nested case-control design using a conditional logistic regression model. We used a cohort design with a time dependent Cox-proportional hazard model to examine the association of metformin use and mortality. Finally, we employed a c-statistic and Net Reclassification Improvement analysis to study the impact of pathology abstraction on predicting mortality. Results: The data suggest metformin use was not associated with the risk of prostate cancer or its grade at presentation. However, each additional 6 month of metformin use was associated with a 24% decrease in prostate-cancer-specific and 8% decrease in all-cause mortality. Pathology abstraction improved the accuracy in predicting all-cause and prostate-cancer specific mortality. Conclusions: In our study metformin use was not associated with a decreased risk of prostate cancer, but had a significant impact on all-cause and prostate cancer specific mortality. These results may serve as proof of concept in designing an interventional study of metformin to delay progression in prostate cancer.

The purpose of the research was to investigate the potential for converting agricultural by-products, barley hull and flaxseed hull as well as their co-extract, into value-added functional food ingredients. Four varieties of barley hull and 3 types of flaxseed hull were hydrolyzed in calcium hydroxide solution in a water bath at 70°C for 4 hrs with shaking. The major phenolic compounds in barley hull, flaxseed hull and their co-extracts were identified by reversed phase high performance liquid chromatography (HPLC) coupled with photodiode array detection (PAD) and quadrupole - time of flight (Q-TOF) mass spectrometry (LC-MS). Ferulic acid, p-coumaric acid, vanillic acid and vanillin, and four ferulate dehydrodimers were detected in barley hull and their co-extracts. Quantitative analysis was conducted on the phenolic acids using the available standards. However, the phenolic compounds in flaxseed were found to be distinct from that of barley hull. Large amounts of secoisolariciresinol diglucoside (SDG), ferulic acid glucoside (FeAG), p-coumaric acid glucoside (CouAG) were found in flaxseed hull with minor content of caffeic acid glucoside (CAG) and flavonoids herbacitin glucoside (HDG), whereas the phytochemical profile of the co-extract was enriched by combining major phenolic compounds from both barley hull and flaxseed hull.The antioxidant activity of barley hull, flaxseed hull as well as their co-extract was evaluated using DPPH radical scavenging assay while total phenolic content was measured using the Folin-Ciocalteau method. After screening using chemical assays, the representative barley hull extract, flaxseed hull extract as well as their co-extract were tested for their intracellular antioxidant activity and the antiproliferative activity in PC-3 human prostate cancer cells. Both chemical assays and the cell culture assays indicated that barley and flaxseed hull had strong antioxidant activity and antiproliferative activity. Although the co-extract exhibited the strong antioxidant activity in the chemicals assay, it behaved differently in the cell culture assay, which may be attributed to the chemical and biological properties of the major phenolics in the co-extract.Following evaluation of the antioxidant activity and anticancer effect of barley hull extract, flaxseed hull extract as well as their co-extract, each type of extract was incorporated into Chinese steamed bread (CSB). The phytochemical profile of CSB was enriched by incorporating barley hull extract, flaxseed hull extract as well as their co-extract, which resulted in a significant enhancement in the antioxidant activity evaluated by DPPH and ORAC. Therefore, barley hull, flaxseed hull and their co-extract are suggested as promising sources of functional food ingredients.

There is a growing demand to involve patients in their own healthcare. Personal Health Records are among the most promising tools for this purpose. However, these tools need to meet patients’ needs and interests in order to be fully adopted and successfully used. This study takes a user centered design approach to design and develop a personal health record for prostate cancer patients by involving them in two main activities of a user centered design: requirements gathering and evaluation. The first phase of the study uses content analysis to analyze interviews with patients and elicit their needs and concerns. Results of this phase showed that patient’s information needs are different depending on the stage of the disease. Before starting treatment, patients are more interested in information about different methods of treatment and their potential side effects. However, after treatment, patients mostly need information about the management of treatment complications and the long term follow ups of their disease. Results also showed that the Internet is the most common information source for patients to find information. However, patients expressed concerns regarding the credibility and reliability of information they found on the Internet. The majority of patients also showed interest in accessing their medical records. However, some patients were concerned about the understandability of the information. Also, there was some concern regarding electronic access to medical records and security of personal data. The findings from phase one are used in phase two to modify a preliminary prototype of the system. In phase three, the modified prototype is evaluated by undergoing usability testing. Overall, the results of usability testing showed that the system was generally useful and easy to use. However, a number of issues were identified that could be resolved in the next iteration of its design and development. / Graduate / 0984

Temporal and spatial molecular imaging of disease biomarkers using non-invasive MRI with high resolution is largely limited by lack of MRI contrast agents with high sensitivity, high specificity, optimized biodistribution and pharmacokinetics. In this dissertation, I report my Ph. D. work on the development of protein-based MRI contrast agents (ProCAs) specifically targeting different cancer biomarkers, such as grastrin-releasing peptide receptor (GRPR), prostate specific membrane antigen (PSMA), and vascular endothelial growth factor receptor-2 (VEGFR-2). Similar to non-targeted ProCAs, these biomarker-targeted ProCAs exhibit 5 - 10 times higher r1 and r2 relaxivites than that of clinical MRI contrast agents. In addition, these biomarker-targeted ProCAs have high Gd3+ binding affinities and metal selectivities. The highest binding affinity of the three GRPR-targeted contrast reagents obtained by grafting a GRPR ligand binding moiety into ProCA32 for GRPR is 2.7 x 10-9 M. We further demonstrate that GRPR-targeted ProCAs were able to semi-quantitatively evaluate GRPR expression levels in xenograft mice model by MRI. In addition, we have also created a PSMA-targeted ProCA which has a binding affinity to PSMA biomarker of 5.2 x 10-7 M. Further, we developed VEGFR-targeted contrast agent which is able to image VEGFR2 in mice models using T1-weighted and T2-weighted sequences. Moreover, the relaxivities and coordination water numbers of ProCAs can be tuned by protein design of ProCA4. Since disease biomarkers are expressed in various tumors and diseases, our results may have strong preclinical and clinical implications for the diagnosis and therapeutics of cancer and other type of diseases.

Background: Androgen deprivation therapy (ADT) has known adverse effects (AEs). Intermittent (INT) ADT may reduce AEs, improve quality of life, and lower costs compared to continuous (CONT) treatment. Objective: To evaluate the cost-effectiveness of INT vs CONT ADT in men with advanced prostate cancer. Methods: A lifetime Markov individual simulation model was developed to evaluate the incremental cost per quality adjusted life month (QALM) of INT vs CONT ADT. Results: INT dominated CONT ADT (mean total costs $94,460 vs $109,431; mean total QALMs 47.0 vs 46.4). INT ADT resulted in less time on therapy (22.4 vs 56.8 months), fewer hip fractures (0.080 vs 0.093 per patient), and fewer total cases of sexual dysfunction (72.5% vs 87.0% of patients) and cardiovascular disease (38.7% vs 44.6% of patients). Conclusions: These results suggest INT ADT is cost-effective compared to CONT ADT however, differences were small. Additional research is required to confirm these findings.

Aberrant DNA methylation in gene promoters causes gene silencing and is a common event in prostate cancer development and progression. While commonly identified methylated genes have been analyzed for their potential clinical utility in a variety of cancers, few studies have attempted a genome-wide methylation approach to discover new and possibly improved biomarkers for prostate cancer. In order to identify DNA methylation changes associated with aggressive prostate cancer, we performed a genome-wide analysis of 40 prostate cancers using Agilent human CpG island microarrays. Methylation profiles of candidate genes were validated using quantitative MethyLight technology in an independent series of 219 radical prostatectomies and compared to clinicopathological parameters. The effects of methylation on expression of HOXD3 and HOXD8 and the possible role of HOXD8 in progression of PCa were also investigated. We discovered previously unidentified methylation in the HOXD cluster of genes, namely HOXD3 and HOXD8, as well as TGFβ2 and GENE X as potential prognostic biomarkers. Furthermore, unsupervised clustering of samples by methylation signature indicated ERG oncogene expression as significantly different between clusters. Within the independent cohort, we observed strong correlations between Gleason score (GS) and HOXD3 as well as GENE X, while HOXD3 and HOXD8 methylation were associated with ERG expresson. TGFβ2 was an independent predictor of disease recurrence using Cox multivariate regression analysis. In gene expression studies, both HOXD3 and HOXD8 were elevated in cancers with poor prognosis, while DNA methylation did not correlate with expression levels. Both genes were found to contain alternative transcription start sites, explaining the poor correlation between methylation and expression. Finally, knockdown of HOXD8 expression did not have any effect on viable cells or cell motility in an in vitro model. These results indicate that a panel of novel DNA methylation markers distinguish indolent prostate cancers from aggressive ones, and that expression of HOXD3 and HOXD8 is regulated by mechanisms including, but not dependent on, DNA methylation.

Background: Androgen deprivation therapy (ADT) has known adverse effects (AEs). Intermittent (INT) ADT may reduce AEs, improve quality of life, and lower costs compared to continuous (CONT) treatment. Objective: To evaluate the cost-effectiveness of INT vs CONT ADT in men with advanced prostate cancer. Methods: A lifetime Markov individual simulation model was developed to evaluate the incremental cost per quality adjusted life month (QALM) of INT vs CONT ADT. Results: INT dominated CONT ADT (mean total costs $94,460 vs $109,431; mean total QALMs 47.0 vs 46.4). INT ADT resulted in less time on therapy (22.4 vs 56.8 months), fewer hip fractures (0.080 vs 0.093 per patient), and fewer total cases of sexual dysfunction (72.5% vs 87.0% of patients) and cardiovascular disease (38.7% vs 44.6% of patients). Conclusions: These results suggest INT ADT is cost-effective compared to CONT ADT however, differences were small. Additional research is required to confirm these findings.

Metastases to bone are observed in around 80% of prostate cancer patients and represent the most critical complication of advanced prostate cancer. Unlike other solid tumors that are associated with osteolytic bone metastases, prostate cancer bone metastases stimulate osteoblastic activity with sclerosis in the bone lesions as a consequence. Osteoprotegerin (OPG) is part of a system with three proteins that play a key role in bone remodeling; namely OPG, RANK and RANKL. RANKL regulates osteoclast activity by binding to RANK on the osteoclasts surface, and this interaction is interrupted by OPG. OPG also plays a role in the lifecycle of tumor cells by blocking TNF-related apoptosis-inducing ligand (TRAIL) making it possible for them to evade cell death. The aim of this thesis was to investigate the interaction between the OPG/RANK/RANKL system and prostate cancer. Data showed that there was production of OPG from prostate cancer cell lines in vitro. This expression was under the influence of cytokines that are present in the microenvironment of bone. Further, there was documented a previously unnoticed cell surface expression of RANKL. Co-culturing the prostate cancer with human osteoblasts increased the expression of RANKL. To connect these findings with in vivo studies, OPG-gene single nucleotide polymorphisms (SNP) were investigated. To evaluate OPG SNPs association with bone, a cohort of elderly men was used. OPG SNPs was shown to be correlated to bone mineral density at hip and spine. There was also an association to fragility fractures. Then there was examined the association of the same SNPs to the incidence of prostate cancer but after a four-year follow-up there was no association to the genetic variants. To summarize this research, we hereby present data that the OPG/RANK/RANKL system might be relevant for prostate cancer growth in bone, and for the skeletal related morbidity in this disease. Future in vitro and in vivo studies will demonstrate the relative importance of this crosstalk, and whether pharmacological interference with the system might be used as a therapeutic tool aiming to decrease skeletal morbidity and possibly also prolong survival in prostate cancer.

This work presents an investigation of the potential development of phenoxodiol, a novel isoflavone, for the treatment of prostate cancer. The literature concerning isoflavone epidemiology, pharmacology, clinical use and their effect on prostate cancer is reviewed. Isoflavone impact on signal transduction pathways is also reviewed with a particular focus on the Akt / mTOR signal transduction pathway, a key signaling pathway in prostate cancer cells. In vitro experiments and xenograft nude mice studies show that phenoxodiol inhibits the growth of DU145 and PC3 prostate cancer cells, when used alone and in combination with cisplatin or carboplatin. Phenoxodiol in combination with cisplatin is highly synergistic when inhibiting the growth of DU145 cells in particular. We show for the first time, that phenoxodiol and cisplatin or carboplatin, inhibits phosphorylation of Akt and mTOR in DU145 and PC3 cells. A first-in-human study of single dose phenoxodiol investigating its pharmacokinetic properties in cancer patient volunteers was performed. Extensive conjugation of phenoxodiol and a short half-life was noted in this study. A Phase I study of intravenous phenoxodiol administered weekly to patients with advanced cancer was then performed to assess toxicity of a repeated dose schedule, as well as the maximum tolerated dose. This schedule was well tolerated in patients, with lymphocytopenia noted as the main toxicity. Given the short half-life of phenoxodiol and our preclinical studies suggesting that combination with cisplatin and carboplatin synergistically inhibited prostate cancer cell growth, we also undertook a Phase I dose escalation study of oral phenoxodiol administered in combination with cisplatin or carboplatin to patients with advanced cancer. The main adverse events include hyperglycemia, hypocalcemia, mild transaminase rises, as well as nausea, constipation, infusion site reactions and lethargy. Three patients with ovarian cancer responded to treatment according to CA125 criteria, and there was a 45% reduction in prostate specific antigen level in one man with androgen independent prostate cancer. The bioavailability of the oral formulation of phenoxodiol was calculated to be 17.5%. Phenoxodiol shows promise for further development in the potential treatment of prostate cancer.