The mechanism by which prostate cancer progresses from androgen dependence (AD) to androgen independence/castration resistance (AI/CR) is currently a major focus of prostate cancer-related research. Prostate cancers that progress to a state of AI/CR are typically resistant to most standard types of treatments. Due to its primary role in driving normal prostate cell growth and proliferation, the androgen receptor (AR) is believed to play a key role in progression. Coregulators, or any proteins which may either enhance or abrogate AR activity, are considered to be one of the potential mechanisms by which AR function may become impaired. Cyclin G-associated kinase (GAK) was initially identified as a potential coregulator of AR in a Tup 1 repressed transactivation system. A LNCaP cDNA library was screened for proteins which interacted with the NH2-terminus of AR. GAK was isolated from three independent library clones using two different AR baits (AR 1-549 and AR 1-646). This interaction was confirmed via GST pulldown and coimmunoprecipitation experiments, and preliminary luciferase assays suggested that GAK activates AR in a hormone dependent manner. In this study, my objectives were to validate GAK’s role as a coregulator of AR and to determine if overexpressing GAK affects progression to AI. In vitro luciferase assays whereby GAK was either overexpressed or knocked down in both LNCaP and PC3 cells did not significantly affect AR activity. Xenograft experiments utilizing a doxycycline (DOX) inducible lentiviral LNCaP-GAK overexpressing stable cell line demonstrated that while GAK may not play a significant role in modulating AR activity, it may adopt a more subtle role enhancing tumour take and tumour volume growth rate in vivo. While these results could not confirm GAK to be a direct coregulator of AR, it is entirely possible that GAK may influence prostate cancer progression, albeit indirectly. Recent publications report a growing amount of evidence suggesting GAK’s involvement in the critical cellular process of clathrin coated vesicle endocytosis, the dysregulation of which could potentially indirectly affect AR regulated genes.

LIV-1, a zinc transporter, is an effector molecule downstream from soluble growth factors. This protein has been shown to promote epithelial-to-mesenchymal transition (EMT) in human pancreatic, breast, and prostate cancer cells. Despite the implication of LIV-1 in cancer growth and metastasis, there has been no study to determine the role of LIV-1 in prostate cancer progression. Moreover, there is no clear delineation of the molecular mechanism underlying LIV-1 function in cancer cells. In this study, we found increased LIV-1 expression in a progresssive manner in benign, PIN, primary and bone metastatic human prostate cancer. We characterized the mechanism by which LIV-1 drives prostate cancer EMT in an androgen-refractory human prostate cancer cell (ARCaP) bone metastasis model. LIV-1, when overexpressed in ARCaPE cells (derivative cells of ARCaP with epithelial phenotype), promoted EMT irreversibly. LIV-1 overexpressed ARCaPE cells had elevated levels of HB-EGF and matrix metalloproteinase (MMP) 2 and MMP 9 proteolytic enzyme activities, without affecting intracellular zinc concentration. The activation of MMPs resulted in the shedding of heparin binding-epidermal growth factor (HB-EGF) from ARCaPE cells, eliciting constitutive epidermal growth factor receptor (EGFR) phosphorylation and its downstream extracellular signal regulated kinase (ERK) signaling. Further investigation of the HB-EGF promoter revealed that both Stat3 and AP-1 controlled HB-EGF promoter activity. Ectopic LIV-1 overexpression induced AP-1 and Stat3 activation. Blockade of both Stat3 and AP-1 by specific inhibitors or dominant negative expression vectors diminished the HB-EGF promoter activity induced by LIV-1 overexpression. These results suggest that LIV-1 is involved in prostate cancer progression as an intracellular target of growth factor receptor signaling which promotes EMT and cancer metastasis. LIV-1 could be an attractive therapeutic target for the eradication of pre-existing human prostate cancer and bone and soft tissue metastases.

Sexuality is a sensitive topic in health care and is often interpreted through a natural scientific lens as just corresponding to sexual dysfunction and fertility problems. The purpose of this thesis was to describe sexuality and its outcomes in two cancer populations. Women with breast cancer and men with prostate cancer in all stages were invited to participate. In this thesis, these two populations are restricted to age groups between 45 and 65 years, since there are reasons to believe that younger people are more vulnerable to sexuality changes. Lifeworld, gender, and sexuality are three concepts of importance in this thesis and they are used from the viewpoint of nursing care. Phenomenological interviews (I, III) and focus group interviews (II, IV) were carried out with a total number of 46 informants. The EPP-method (Empirical Phenomenological Psychological) was used (I, III) in order to grasp the lived experience, and qualitative content analysis was used to analyse the seven focus groups (II, IV). The lifeworld experiences of those women and men were comparable. The changes brought by the cancer and its treatment were a threat to their very existence, their existential base of knowledge had gone and alienation occurred (I, III). For the women, this was illustrated through the metaphor of a bird which is pinioned and unable to fly anymore. For the men it was expressed in the essential meaning “to lose the elixir of life”. Both women and men suffered, sexuality changed from one day to another and they handled it individually. Changed body appearance, and feeling old and unattractive were, for the women, the dominating features, whilst for the men changed desire and erection problems were their main concerns. The findings from the group discussions (II, IV) elucidate the gendered differences in these two contexts. The aim of the women was to look healthy and attractive and for the men the ability to have an erection was important. Neither of these two groups of people was able to meet their aims. On the other hand, being diagnosed with a life-threatening disease they were not in a position to claim preserved sexuality. This opens up existential questions that need to be confirmed in health care. To succeed in this, a change of perspective is required in health care. It should be possible to use human science to the same extent as natural science in health care.

Im Rahmen einer prospektiven Studie wurden zwischen Juli 2007 und Oktober 2008 Patienten mit Prostatakarzinom in der Klinik für Urologie des Universitätsklinikums Leipzig befragt. Die Lebensqualität und Lebenszufriedenheit wurden mit Hilfe von Selbstbeurteilungsfragebögen (EORTC QLQ-C30, SF-8 und FLZM) erfasst. Insgesamt lagen Daten von 276 Patienten vor. Ziel der Arbeit war es, den Verlauf von Lebensqualität und Lebenszufriedenheit über drei Monate nach dem Klinikaufenthalt zu beobachten, sowie diese Daten mit denen der Allgemeinbevölkerung zu vergleichen. Weiterhin wurden verschiedene Einflussfaktoren auf Lebensqualität und Lebenszufriedenheit bewertet. Prostatakarzinompatienten gaben die schlechteste Lebensqualität zum Zeitpunkt vierzehn Tage nach Entlassung aus der Klinik an. Innerhalb von drei Monaten erreichten sie wieder das Ausgangsniveau an Lebensqualität. Insgesamt zeigten sich kaum klinisch bedeutsame Unterschiede in der Lebensqualität zwischen Patienten und Allgemeinbevölkerung. Die Lebenszufriedenheit nahm im Verlauf zwar ab, jedoch gaben die Prostatakrebspatienten zu fast allen Zeitpunkten eine ähnliche oder sogar höhere Lebenszufriedenheit als die Vergleichsgruppe an. Die einzige, aber wichtige Ausnahme hiervon bildete der Bereich Sexualität, hier waren die Patienten nach 3 Monaten deutlich unzufriedener als die Männer der Allgemeinbevölkerung. Alter, Bildungsgrad und die seit Diagnosestellung vergangene Zeit stellten sich im Gegensatz zum Tumorstadium als Einflussfaktoren für die Beurteilung der Lebensqualität dar. Alle diese Faktoren zeigten jedoch keinen signifikanten Einfluss auf deren Lebenszufriedenheit. Die Korrelationen zwischen den drei eingesetzten Fragebögen waren fast ausnahmslos positiv, entsprechend einer gleichsinnigen Variabilität. Eine generelle psychoonkologische Betreuung aller Patienten scheint nicht notwendig, jedoch sollten Ärzte für die Probleme des Einzelnen sensibilisiert sein um rechtzeitig Hilfe anbieten zu können.

<p>Abstract</p><p>The trefoil factor family of secreted proteins contains three members; trefoil factor 1 or TFF1, trefoil factor 2 or TFF2, and trefoil factor 3 or TFF3. These three proteins share a conserved 42-43 amino acid domain containing 6 cysteine residues resulting in three disulfide bonds that holds the protein in a characteristic three-loop or "trefoil structure" known as the P domain. TFF1 is primarily localized to the stomach and secreted by the gastric mucosa while TFF2 and TFF3 are primarily localized to the colon and duodenum and secreted by the goblet cells. All three of these proteins play a protective role in the gastrointestinal tract where they are normally localized and have been identified as possible tumor suppressors, however, these proteins are also upregulated in cancer within tissues where they are not normally expressed including the breast, pancreas, prostate, and liver. The mechanisms by which two of these factors, TFF1 and TFF3, promote tumorigenesis remain largely undefined. In this dissertation we will attempt to elucidate these mechanisms as well as the regulation of these two proteins in both pancreatic and prostate cancer. Many of the underlying genetic and molecular mechanisms involved in the development of both pancreatic and prostate cancer remain largely unknown and as a result, therapeutic and diagnostic tools for treating these diseases are not as effective as they could be. By deciphering the role of TFF1 and TFF3 in these cancers, they could potentially serve as new therapeutic targets or biomarkers for treating both diseases.</p><p>Chapter 2 of this dissertation will examine the functional role of TFF1 promoting tumorigenesis in pancreatic and prostate cancer. We will show that TFF1 expression is critical for the viability of both pancreatic and prostate cancer cells and that reduction of TFF1 expression in these cells results in decreased tumorigenicity when implanted in immunocompromised mice. It will also be demonstrated that TFF1's function in promoting tumorigenicity is its ability to assist tumor cells overcome the tumor suppressive barrier of senescence. Thirdly, we show that the form of senescence that TFF1 assists in allowing the cells overcome is oncogene-induced senescence (OIS). Lastly, a cell cycle array identifies the potential downstream target p21CIP, a cyclin-dependent kinase inhibitor and OIS marker, whose expression is induced by loss of TFF1 expression.</p><p>In Chapter 3 of this work, we examine the role of another trefoil factor family member, TFF3, and its role in promoting prostate tumorigenesis. Just as with TFF1, it appears that TFF3 3 expression is critical for prostate cancer cell viability and tumorigenicity using the same experimental techniques used in Chapter 2. Using a genetically defined model of prostate cancer, a PI3-kinase-dependent regulatory mechanism of TFF3 emerges in this prostate cancer context. Using this system we begin to see a divergence in both regulation and function of TFF1 and TFF3 in prostate cancer. Finally, a mouse model expressing TFF3 was developed to monitor the histopthological changes associated with expression of this protein. Initial characterization of this model suggests a hyperplastic phenotype coinciding with TFF3 expression in the prostate.</p><p>The two studies in this dissertation establish a role of TFF1 and TFF3 in both prostate and pancreatic tumorigenesis and demonstrate that ablation of expression of both proteins is a potent inhibitor of tumorigenesis. With this knowledge, it is possible that TFF1 and TFF3 may become a potential therapeutic target or diagnostic marker for better treatment of prostate and pancreatic cancer.</p> / Dissertation

<p>Glucose metabolism remains an intensely explored topic of cancer biology since the initial discoveries of Otto Warburg nearly 80 years ago. Many solid tumors metabolize glucose primarily to lactate despite the availability of oxygen, revealing a dependence on glycolysis that may serve as a basis for targeted therapy. In particular, a diet devoid of carbohydrate may minimize the growth capabilities of glucose-dependent cancers. As our interests lie in prostate cancer, we examined whether a ketogenic diet devoid of carbohydrate (NCKD) would reduce the growth rate of tumors derived from human prostate cancer cell lines in a murine xenograft model.</p> <p>Our initial experiments utilized the LAPC-4 cell line, a human androgen-sensitive prostate cancer cell line, in a SCID-mouse xenograft model to determine the effects of an NCKD on tumor growth and animal survival relative to two other diets: (1.) a Western-type diet (WD) reflecting consumptions patterns of men diagnosed with prostate-cancer in the Western world and (2.) a low-fat diet (LFD) representing the present standard of care. Following this study, we conducted a second study utilizing a different human prostate cancer cell line (LNCaP) in order to assess whether our initial observations were robust across multiple prostate cancer tumor models and to also further explore the molecular underpinnings of our observations. Both studies revealed the NCKD leads to a reduction in tumor growth rate and greater overall mouse survival relative to the WD. In addition, the NCKD was equivalent in these parameters to the LFD. We also observed key associations between survival and extent of urinary ketosis as well as favorable changes in insulin and insulin-like growth factor-1 (IGF-1) and gene expression that would be predictive of prolonged survival in mice consuming the NCKD.</p> <p>We believe these data provide compelling evidence to consider a potential therapeutic role for dietary carbohydrate restriction in prostate cancer. We hope these results ultimately serve as a basis to conduct future clinical trials assessing whether dietary carbohydrate restriction, either alone or in combination with more conventional therapies, provides clinicians with an additional weapon against prostate cancer.</p> / Dissertation

Title from title page of PDF (University of Missouri--St. Louis, viewed February 9, 2010). Includes bibliographical reference (p. 165-182).

p38 mitogen-activated protein kinases (MAPKs) control the endocytic trafficking of various growth-related cell surface receptors and transporters. Herein, I demonstrate that p38 MAPKs also regulate autophagy, or the process of self-cannibalism. In my studies, inhibition of p38 MAPKs triggered rapid formation of autophagosomes in prostate cancer cells, even under nutrient-rich conditions, and remarkably, the autophagosomal membranes emanated from endoplasmic reticulum exit sites via the concerted actions of the small GTPases, ARF1 and SAR1. Once formed, the autophagosomes fused with late endosomes and/or lysosomes, in a Rab7-dependent manner, to form “hybrid organelles” that were co-labeled with ER, autophagic, late endosomal, and lysosomal markers. Unlike other inducers of autophagy, however, inhibition of p38 MAPKs suppressed the fission of hybrid organelles, resulting in a profound but reversible accumulation of large cytoplasmic vacuoles. Thus, in addition to their previously reported roles in endocytosis, p38 MAPKs appear to coordinately regulate autophagy and the downstream biogenesis and fission of hybrid organelles. / text

Prostate cancer (PCa) is heterogeneous containing phenotypically diverse cells. It is unclear whether these phenotypically different PCa cells are functionally distinct and possess divergent tumorigenic potential. Androgen signaling plays important roles in differentiation and survival of malignant PCa cells, and prostate specific antigen (PSA) as one of the androgen signaling target genes is used as a biomarker of AR signaling to assess tumor progression and evaluate therapeutic efficiency in clinic. Here we present evidence for discordant AR and PSA expression resulting in AR⁺/PSA⁺, AR⁺/PSA⁻, AR⁻/PSA⁻, and AR⁻/PSA⁺ PCa cells in human tumors. We also show that prostate tumor PSA mRNA levels inversely correlate with poor clinical outcomes and patient survival. By employing a lentiviral reporter system, we have fractionated bulk PCa cells into PSA⁺ and PSA⁻[superscript '/lo'] cell populations, with the former being AR⁺/PSA⁺ and the latter containing both AR⁺/PSA⁻ and AR⁻/PSA⁻ cells. The PSA⁺ and PSA⁻[superscript '/lo'] PCa cells demonstrate distinct molecular, cellular, and tumor-propagating properties. PSA⁻[superscript '/lo'] PCa cells are quiescent and refractory to stresses including androgen deprivation, exhibit high clonogenic potential, and possess long-term tumor-propagating capacity. They preferentially express stem cell genes and can undergo asymmetric cell division to generate PSA⁺ cells. Of great clinical interest, PSA⁻[superscript '/lo'] PCa cells can initiate robust tumor development and resist androgen ablation in castrated hosts, and they harbor highly tumorigenic castration resistant PCa cells. In contrast, PSA⁺ PCa cells possess more limited tumor-propagating capacity, undergo symmetric division, and are sensitive to castration. Systemic androgen levels dynamically regulate the relative abundance of PSA⁺/PSA⁻[superscript '/lo'] PCa cells in the tumors, which in turn impact the kinetics of tumor growth. Further studies reveal that the PSA⁻[superscript '/lo'] PCa cell population harbors several overlapping but nonidentical tumorigenic subsets including ALDH⁺, CD44⁺, and [alpha]2[beta]1⁺ cells and ALDH⁺CD44⁺[alpha]2[beta]1⁺ can further enrich castration resistant PCa cells. These observations together suggest that heterogeneous PCa cells are organized as a tumorigenic hierarchy. Our results have important implications in understanding how different subpopulations of PCa cells manifest differential responses to current androgen deprivation therapy (ADT). / text

published_or_final_version / Medical Sciences / Master / Master of Medical Sciences