The interplay between cancer cells and the extracellular matrix (ECM) remains a critical regulator of both normal tissue organization and cancer cell invasion. Proteins that function as ECM receptors function to link the cell with the ECM. Abberations in either the structure of the ECM or the expression of ECM receptors leads to disrupted interaction and downstream signaling effects. Dystroglyan (DG) is an ECM receptor that is expressed in a variety of tissue types and functions to mediate sarcolemma stability, epithelial polarity, and is critical in the early formation of basement membranes. However, DG has primarily been studied in muscle where loss of its function is linked to a host of muscular dystrophies. In the epithelium, the role of DG remains enigmatic. While DG has repeatedly been shown to lose function during cancer development and progression, the mechanism and functional consequence of its loss are currently unknown. In order to increase our understanding of DG in cancer development, we analyzed its expression and glycosylation, a functional requirement for DG, in a range of prostate cancer cell lines. Previous work has shown DG to be downregulated in prostate cancer, but the mechanism by which this occurs has remained largely unclear. We found that DG expression is maintained while its glycosylation was heterogeneous in the cell lines. Further investigation revealed that lines with hypoglycosylated DG strongly associated with the loss of expression of the glycosyltransferase LARGE2. Further this enzyme is frequently downregulated in human cancers and appears to serve as a required enzyme in DG glycosylation within prostate epithelium. This is the first work to demonstrate the functional requirement of LARGE2 for DG, and the only work to implicate loss-of-function of LARGE2 in cancer progression. To determine whether loss of LARGE2 is found in other tumor types, we analyzed human clear cell renal cell carcinoma (ccRCC) samples by immunohistochemistry and via in silico analysis with the Cancer Genome Atlas (TCGA). Our work demonstrated a frequent and significant downregulation of LARGE2 expression and its association with DG hypoglycosylation. Additionally, we found the loss of LARGE2 strongly associated with increased mortality. Thus, we again demonstrated a functional requirement of LARGE2 but also found a clinical correlate with increased mortality. Finally, we examined the functional outcome of DG hypoglycosylation or loss of expression in both a mouse model of prostate cancer and a variety of cell lines models. We found that while loss of DG expression does not increase prostate cancer growth or metastasis in one model of cancer, loss of its glycosylation does seem to mediate downstream metabolic changes within cells. The mechanism for this change remains unclear. In summary, these studies have contributed to our understanding of DG glycosylation and function in both prostate and renal carcinoma. Additionally, we have shown a novel mechanism by which DG glycosylation is lost with downregulation of LARGE2 expression. Finally, while we were unable to demonstrate a clear mechanism by which signaling changes arose, we were able to demonstrate a strong correlation between DG hypoglycosylation and increased mortality in ccRCC. These insights could be used to improve treatment of multiple cancer types as our understanding of DG function continues to improve.

Ο μεταγραφικός παράγοντας PPARγ ανήκει στην οικογένεια των πυρηνικών ορμονικών υποδοχέων και διαδραματίζει σημαντικό ρόλο στην διαδικασία της καρκινογένεσης του προστάτη, συμμετέχοντας στην κυτταρική διαφοροποίηση, στη ρύθμιση της γονιδιακής έκφρασης, στην ενεργοποίηση ή καταστολή της μεταγραφής συγκεκριμένων γονιδίων, στη ρύθμιση του κυτταρικού κύκλου, στην απόπτωση, στην αναστολή της αγγειογένεσης, στην διεισδυτική ικανότητα και στο μεταστατικό δυναμικό του αδενοκαρκινώματος του προστάτη, καθώς επίσης και στο μεταβολικό μονοπάτι του αραχιδονικού οξέος μέσω αναστολής της κυκλο-οξυγενάσης 2. O ΡΡΑRγ φαίνεται να διαθέτει αντινεοπλασματική δράση, η οποία μπορεί να χρησιμοποιηθεί τόσο στην χημειοπροφύλαξη, όσο και στην χημειοθεραπευτική αντιμετώπιση του αδενοκαρκινώματος του προστάτη. / The transcription factor PPARγ is a member of the superfamily of the nuclear hormone receptors and plays a significant role in the carcinogenesis of the prostate. PPARγ participates and controls various cellular functions, necessary for the malignant transformation of the prostatic cell, such as the cellular differentiation, the gene expression, the activation or repression of the transcription of various genes, the control of the cell cycle, the apoptosis, the inhibition of aggiogenesis, the invasive and metastatic potential of the adenocarcinoma and finally the metabolic pathway of the arachidonic acid, through inhibition of the COX-2. The antiproliferative functions of PPARγ can be used for the chemoprevention and as an adjuvant in the chemotherapeutic regimens targetting the adenocarcinoma of the prostate.

Predicting the prognosis of patients with prostate cancer is a clinically important problem. Previous studies have indicated that the expression of haptoglobin-related protein epitopes in samples of breast cancer in early stages was associated with earlier relapses and higher risk for tumor recurrence. Oncogenic antigen 519 (OA-519) is the new marker designation for molecules expressing haptoglobin-related protein epitopes. The objective of this immunohistochemical study was to examine OA-519 expression in prostate cancer samples and its relationship to the established prognostic indicators of tumor grade, tumor volume, and clinical stage. Forty-two consecutive tissue samples of prostate adenocarcinoma were examined using an affinity- purified anti-OA-519 antibody. Twenty specimens (48%) tested positive, whereas 22 (52%) tested negative. No staining was observed in normal or hyperplastic prostate tissue. Staining occurred in 6 of 9 (67%) grade III, 14 of 23 (61%) grade II, and in none of 10 (0%) grade I cases (I vs. II and/or III: Fisher exact test, P < 0.006). Twenty-three of the 42 samples were transurethral resection specimens with cancer; 11 (48%) of these tested positive. The mean percentage of tissue chips with tumor, a measure of tumor volume, was significantly higher in the positive group (57%) than in the negative group (15%) (P = 0.004). The proportion of positively stained cases increased with advancing clinical stage, with 25% of Stage A cases expressing OA-519, and 46%, 67%, and 64% of Stages B, C, and D, respectively, expressing OA-519. OA-519 expression correlates with higher tumor grades, larger tumors, and possibly with advanced stage, and thus, it is potentially of prognostic value in prostate cancer.

Prostate adenocarcinoma (PCa) is one of the most prevalent cancers in the world. Second only to lung cancer, the key to its successful treatment is in its early detection. With the introduction of prostate-specific antigen in the early 1990s, a screening test involving measuring levels of this protein was developed to detect PCa in asymptomatic individuals. This test is also known as the PSA test. PCa-specific mortalities have been in decline since the test's introduction. Despite this decline, recent studies have called the efficacy of the PSA test into question. Two large randomized controlled trials conducted in the US and Europe reveal contradicting results as to PSA's accuracy and usefulness. Concerns of overdiagnosis and overtreatment as the result of using PSA screening has led to many national organizations recommending caution or even recommending against its use. Through a thorough review of a large collection of current PCa literature, this study reviews the flaws of using PSA to screen for PCa and investigates alternative approaches currently being pursued through active research to make PCa early detection more accurate. These approaches include improving the accuracy of the PSA screen using PSA-derived testing methods, using PCa-induced epigenetic modifications as a new target for PCa screening, and using urine biomarkers. All of these methods were compared using area under the curve (AUC) values obtained via receiver operating characteristic analysis. Each method has its own flaws but by comparing each of the different approaches, I was able to conclude that out of the currently available screening methods, screening for Engrailed-2 protein in urine is the most promising screening method with the highest AUC values compared to the other methods. Although this method has been introduced in the UK, it has not been introduced in the US yet. Epigenetic screening methods hold the most promise for accurate PCa screening in the future as it confers the highest accuracy in detecting PCa. However, as it hasn't been shown that epigenetic modifications can be easily obtained in the urine or blood serum for easy and accurate screening, I believe more work has to be done in order for it to be successful in being applied as a screening test. By determining the most promising screening type, we can focus resources and efforts towards finding a way to detect PCa early, allowing for successful treatment.

High grade intraepithelial neoplasia (HGPIN) and atypical small acinar proliferation (ASAP) are two pathological lesions associated with prostate adenocarcinoma. HGPIN is an architectural finding, while ASAP is a term used to describe a lesion that cannot confidently be diagnosed as prostate adenocarcinoma. The mean incidence rate for HGPIN is 7.7% with a median of 5.2% and range of 0-24.6% and the cancer detection rate mean is 18.1%. The mean incidence rate for ASAP is 5.0% with a median of 4.4% and a range of 0.7-23.4%. The mean cancer detection rate is 40.2%. Currently, the incidence and cancer detection rates for HGPIN and ASAP for Winnipeg, Manitoba, have not been published. A retrospective study was conducted on all prostate biopsies collected from the Manitoba Cancer Care Prostate Centre (MCCPC) from 2008 and 2009. Prostate biopsies with a diagnosis of isolated HGPIN and or ASAP and no previous history of cancer were included in this study. In Winnipeg, Manitoba, from 2008-2009, the mean HGPIN incidence rate was 5.0% and the mean cancer detection rate was 46.1%. The mean ASAP incidence rate was 4.6% and the mean cancer detection rate of 48.2%. As a control, the cancer detection rate following a benign diagnosis was also calculated at 33.3%. The mean incidence and cancer detection rates for HGPIN and ASAP for Winnipeg, Manitoba are slightly lower than literature, but still fall within the published range. In addition, the mean ASAP cancer detection rate is similar to the cancer detection rate following a benign diagnosis indicating that, in our study, both a benign finding and a diagnosis of ASAP hold the same predictive value for cancer on a subsequent re-biopsy.

Σκοπός της μελέτης ήταν η ανάδειξη της νεοαγγειογένεσης και ο προσδιορισμός της έκφρασης των VEGF-A, FGF-2, COX-2, AR και BCL-2 στην καλοήθη υπερπλασία και το αδενοκαρκίνωμα του προστάτη. Το υλικό αφορούσε σε δείγματα προστατικού ιστού μονιμοποιημένα και εγκλεισμένα σε παραφίνη, από 24 περιστατικά καλοήθους υπερπλασίας και 139 περιστατικά προστατικού αδενοκαρκινώματος. Τα τελευταία χωρίστηκαν περαιτέρω σε 3 υποομάδες (Grade I, II και ΙΙΙ) ανάλογα με το βαθμό διαφοροποίησης του νεοπλάσματος κατά Gleason (2-4, 5-7 και 8-10 αντίστοιχα). Χρησιμοποιήθηκε ανοσοϊστοχημική μέθοδος Βιοτίνης-Στρεπταβιδίνης-Υπεροξειδάσης, και εφαρμόστηκε ημιποσοτική μέθοδος για την εκτίμηση της ανοσοϊστοχημικής χρώσης. Τα ευρήματά μας ανέδειξαν σαφή αύξηση της νεοαγγείωσης (MVD) στο αδενοκαρκίνωμα του προστάτη σε σχέση με την καλοήθη υπερπλασία, η οποία εμφάνισε στατιστικώς σημαντική θετική σχέση με το βαθμό κακοήθειας των προστατικών νεοπλασμάτων (ANOVA p<0.001) και με την έκφραση των VEGF-A και COX-2 (ANOVA p<0.001). Αναδείχθηκε αντίστροφη συσχέτιση της πυρηνικής έκφρασης του ανδρογονικού υποδοχέα (AR) με το βαθμό διαφοροποίησης των νεοπλασμάτων (p<0.0001) και την έκφραση του VEGF-A στο προστατικό στρώμα (p<0.001 Spearman r = -0.312). Η έκφραση της BCL-2 παρουσιάστηκε αυξημένη στα προστατικά αδενοκαρκινώματα και σχετίστηκε με το βαθμό διαφοροποίησης των νεοπλασμάτων (p<0.001) και την έκφραση των VEGF-A και COX-2 (p<0.001). Η έκφραση του VEGF-A σε όλα τα περιστατικά αδενοκαρκινώματος του προστάτη εμφάνισε στατιστικώς σημαντική σχέση με το βαθμό διαφοροποίησης των νεοπλασμάτων (p<0.0001). Ωστόσο στα πτωχής διαφοροποίησης αδενοκαρκινώματα η μέση τιμή της έκφρασης του VEGF-A παρουσίασε πτώση. Αντίθετα, η σημασία της COX-2 στον προστατικό καρκίνο αναδείχθηκε με την έκφρασή της τόσο στην καλοήθη υπερπλασία όσο και στα αδενοκαρκινώματα του προστάτη. Η έκφραση παρουσίασε σημαντική σχέση με το βαθμό διαφοροποίησης των νεοπλασμάτων (p<0.01). Η σαφώς αυξημένη έκφραση της COX-2 σε σχέση με τη μείωση της έκφρασης του VEGF-A στα πτωχής διαφοροποίησης νεοπλάσματα πιθανόν υποδηλώνει την ύπαρξη ενός αγγειογενετικού διακόπτη στα νεοπλάσματα αυτά όπου η COX-2 φαίνεται να παίζει σημαντικότερο ρόλο από τον VEGF-A. Η σημαντική αυτή πληροφορία θα μπορούσε να βρει πιθανή θεραπευτική εφαρμογή, καθώς σε πτωχής διαφοροποίησης αδενοκαρκινώματα του προστάτη ίσως η θεραπεία με COX-2 εκλεκτικούς αναστολείς να είχε πολύ καλύτερα αποτελέσματα από τις αντι-αγγειογενετικές θεραπείες με αναστολείς του VEGF. / The aim of this study was to immunohistochemically evaluate the expression of VEGF-A, FGF-2, COX-2, AR and BCL-2 in benign prostatic hyperplasia (BPH) and prostate carcinoma in relation to microvessel density (MVD) and the Gleason grade of the neoplasms. A total of 139 cases of primary prostate carcinoma and 24 cases of benign hyperplasia were included in the study. Tumors were graded according to the Gleason grading system and further divided into 3 subgroups (GRADE I, II and III). The immunostaining was performed according to the Streptavidin-Biotin Complex Peroxidase method, in formalin fixed paraffin-embedded tissue. Mean micro vessel density (MVD) was strongly related to tumor grade, VEGF-A and COX-2 histoscore (ANOVA, p<0.001). The androgen receptor was localized in the nuclei of prostate epithelial cells in 97% of cases. The comparison of AR staining with tumor grade revealed an inverse relationship between these two parameters (ANOVA, p<0,0001). An interesting finding was the inverse relationship of stromal AR expression in relation to VEGF-A immunoreactivity. BCL-2 expression was correlated with tumor grade in prostate carcinoma cases (p<0.001) and was strongly correlated with COX-2 and VEGF-A expression (p<0.001). These findings suggest that BCL-2 may play a dual role in tumorigenesis, possibly through an angiogenetic axis. VEGF-A expression was detected in only 17% of BPH cases but all prostate cancer specimens demonstrated some degree of immunoreactivity. COX-2 immunopositivity was present in 54% of BPH specimens and in 99% of primary prostate carcinomas. The increased COX-2 expression correlated significantly with Gleason grade. In our study, high-grade neoplasms presented low to moderate VEGF staining intensity compared to COX-2 expression. These results suggest the activation of an angiogenic switch in poorly differentiated neoplasms, where COX-2 may play a crucial role compared to VEGF and the possible key role of COX-2 in poorly differentiated cancers. According to our findings, anti-VEGF therapy could prove to be more beneficial in patients with low-grade disease, while patients with high-grade prostate carcinoma are more likely to respond to selective COX-2 inhibitors. Immunohistochemical determination of VEGF-A and COX-2 content might prove a useful tool in the design of patient-tailored, anti-angiogenic treatments.

Prostate cancer is one of the most abundant types of cancer among men and the demand for a specific treatment is very high. In this thesis, I have focused on using Glutamate Carboxypepti- dase II (GCPII), as a target for a proof-of-principle delivery system. GCPII is a transmembrane protein that internalizes after a binding of a ligand and is overexpressed in prostate cancer. Virus-like particles from Murine polyomavirus (VLPs) are a suitable nanocarrier for the delivery of imaging agents and drugs. Here I describe modifying these VLPs with inhibitors of GCPII and fluorescent dyes and characterize their binding to GCPII on surface plasmon resonance and to cells expressing GCPII on confocal microscopy. VLPs carrying a GCPII inhibitor show specific binding to GCPII on surface plasmon reso- nance, however they bind non-specifically to cells that don't express GCPII. Several approaches have been tried to avoid that. The substitution of BC loop on the exterior surface of VLPs that is partially responsible for the binding of sialic acid did not seem to affect specificity on cells. Another approach tested was coating of the wild-type VLPs with large polymer carrying a flu- orescent label and a GCPII inhibitor. After the conjugation of the polymer to the VLP, specific binding and internalization in GCPII-positive...