Aukšto laipsnio prostatos intraepitelinė neoplazija (ALPIN, angl. – HGPIN – high grade prostatic intraepithelial neoplasia ) priskiriama prie priešvėžinių būklių ( arba prostatos vėžio prekursorių) ir susijusi su padidėjusia kartu esamo vėžio diagnozavimo rizika arba vėlesne jos progresija iki vėžio. Manoma, kad pacientai, kuriems nustatyta ALPIN prostatos biopsinėje medžiagoje, yra tinkami kandidatai chemoprevencijai. Šio darbo tikslas buvo nustatyti 5-alfa reduktazės inhibitoriaus dutasterido reikšmę prostatos vėžio prevencijai esant didelei šios ligos rizikai. Darbo uždaviniai : 1. Nustatyti prostatos vėžio dažnį esant didelei šios ligos rizikai (biopsinėje medžiagoje nustatyta aukšto laipsnio prostatos intraepitelinė neoplazija). 2. Įvertinti pakartotinių biopsijų reikšmę prostatos vėžio diagnozavimo dažniui. 3. Įvertinti tiriamojo vaisto reikšmę prostatos vėžio prevencijai (ar sumažina prostatos vėžio diagozavimo dažnį) esant didelei šios ligos rizikai. 4. Palyginti diagnozuoto prostatos vėžio diferenciacijos skirtumus tiriamosiose grupėse. / A high grade prostatic intraepithelial neoplasia (HPIN) is traditionally ascribed to pre-cancerous conditions or prostate cancer (PC) precursors, and associated with an increased risk of concomitant cancer or its later progression to malignant disease. After evaluation of the data indicating HPIN as a pre–cancerous condition, it is thought that the patients having HPIN in the prostate biopsy material are suitable candidates for chemoprevention.The aim of the study was to find out the significance of dutasteride, a 5–alpha reductase inhibitor, for the prevention of development of prostate cancer in case of a high risk for the disease. Objectives of the study: 1. To determine the rate of prostate cancer in case of a high risk for the disease (when a high grade prostatic intraepithelial neoplasia is found in biopsy material). 2. To find out the significance of repeat biopsies for the detection rate of prostate cancer. 3. To find out the significance of the investigatory medication for the prevention of development of prostate cancer in case of a high risk for the disease. 4. To compare the differences in the differentiation of diagnosed prostate cancer between the study groups.

A few abnormal cells found in a small piece of prostate tissue are most consequential for a man’s future. The prevalence of prostate cancer (PCa) is increasing globally. The main instigating factor for this cancer is not yet known, but it appears to be the consequence of many variables such as an increasingly older population, more frequent PSA-testing, and factors involving lifestyle. Prostate cancer screening, as an equivalent for breast cancer screening, has been suggested but unfortunately there are no accurate diagnostic tools available for this type of screening. The reason for this is simply that the prostate is one of the most difficult organs to diagnose and, consequently, PCa screening would generate far too many false-positive and false-negative results.  The prostate is not easily accessible as it is deeply-seated in the male pelvic area, wrapped around the urethra and surrounded by sensitive vital organs.  Furthermore, PCa is frequently multi-focal, and the cancer cells have a tendency of assimilating among normal cells and, thus, do not always form solid lumps.  Therefore, prostate tumors are often not felt by digital rectal examination (DRE) or identified by imaging.  The PSA-test is not reliable as it is more prostate-specific than cancer-specific.  Due to increasing prostate awareness, more early-stage and locally confined PCa are being detected. This is saving lives, although there is a high risk of over treatment and unnecessary side-effects.  The increased detection of PCa requires sophisticated diagnostic methods and highly skilled clinicians who can discern between indolent and aggressive cancers.  The current “gold-standard” for PCa diagnosis is biopsy grading by pathologists using the Gleason score system, which is a difficult task.  Therefore, innovative methods to improve the precision of prostate diagnosis, by increased biopsy sensitivity and tumor localization, are of essence. In light of these difficulties, the metabolomic approach using 1D and 2D high-resolution magic angle spinning (HRMAS) NMR spectroscopy combined with histopathology on intact prostatectomy specimens was evaluated in this research project.  The non-destructive nature of HRMAS NMR enables spectroscopic analysis of intact tissue samples with consecutive histological examinations under light microscope. Metabolomics aids in the unraveling and the discovery of organ-specific endogenous metabolites that have the potential to be reliable indicators of organ function and viability, extrinsic and intrinsic perturbations, as well as valuable markers for treatment response. The results may, therefore, be applied clinically to characterize an organ by utilizing biomarkers that have the capacity to distinguish between disease and health. The aim was to characterize the human and the rat prostate in terms of its intermediary metabolism, which I show here to differ between species and anatomical regions.  Furthermore, the aim is to seek the verification of HRMAS NMR derived metabolites which are known to be a part of the prostate metabolome such as, citrate, choline, and the polyamines which were performed, but also the identification of metabolites not previously identified as part of the local prostate metabolism, such as Omega-6, which was detected in tumors.  The extended aim was to elucidate novel bio-markers with clinical potential. In this study, the common phyto-nutrient, inositol, which appears to possess protective properties, was identified as being a potentially important PCa bio-marker for the distinction between the more indolent Gleason score 6 and the more aggressive Gleason score 7 in non-malignant prostate tissues with tumors elsewhere in the organ. Further studies in this area of PCa research are therefore warranted.

Objectives. To study whether birth weight (BW) was correlated to cardiovascular risk factors, coronary heart disease (CHD), cardiovascular disease (CVD), and prostate cancer (PCA) at adult ages, whether a possible relationship depended on mediating factors from birth time, hereditary circumstances, and adult life variables, and what importance possible associations might have for the rate of the complaint in the general population. Material and methods. Population-based cohorts of men born in 1913 and followed up until old age. Risk of outcome was estimated using Cox’s and Poisson regressions. The results were transformed to population attributable risk percentage (PAR%) of the complaint that could be attributed to low or high BW, given causality between exposure and outcome. Results. After adjustment for the influence of covariates, systolic blood pressure at age 50 decreased by 3.7 mmHg per 1000 g increase in BW, the prevalence of antihypertensive treatment decreased by 32%, diabetes by 53%, serum total cholesterol decreased by 0.20 mmol L-1, and being in top quintile of serum cholesterol decreased by 23%. The adjusted risks were somewhat more marked relative to the crude risks. CHD and CVD incidence and mortality were virtually unaffected by BW. In the general population, the risk percentage attributable to a BW ≤3000 g was 18% for diabetes, 2.5% for cholesterol, and ≤1% for antihypertensive treatment and CHD and CVD incidence and mortality. PCA incidence and mortality risk increased by 62% and 82%, respectively, among those whose BW was ≥4250 g compared with those whose BW was 3001-4249 g. The risk percentages attributable to a BW ≥4250 g in the general population for PCA incidence and mortality were 7.8% and 10.8%. Conclusions. Low BW seemed to affect cardiovascular risk factors but not incidence and mortality from CHD and CVD. A high proportion of diabetes on the community level could be attributed to low BW, while the proportional burden of other cardiovascular complaints that could be attributed to low BW was modest. PCA incidence and mortality seemed to be affected by high BW.

The natural history of localized prostate cancer is not fully understood. In most patients the tumor will never progress to a lethal disease, while a subset of patients will ultimately die of the disease. Efficient tools to separate indolent from lethal disease is currently lacking which means that many patients will be offered treatment without any benefit, but still be at risk of experiencing treatment related side effects. The aims of these studies were to get more insight into the natural history of untreated localized prostate cancer, to assess the prognostic value of established clinical parameters such as Gleason score, nuclear grade and tumor volume and, moreover, some new prognostic markers Ki-67, AMACR and MUC-1. We also aimed to study time trends in the detection of incidental tumors in Sweden. Patients with localized disease (n=223) and no initial treatment were followed for 21 years. Most patients had a favorable outcome. However, a subset of patients developed lethal disease even beyond 15 years of follow-up and these patients define the group that may benefit most from treatment with curative intent. Patients with poorly differentiated tumors experienced a 9 time higher risk of dying in prostate cancer. The studies on prognostic markers are based on a cohort of patients (n=253) with incidental prostate cancer detected by transurethral resection for presumed benign hyperplasia. All patients were left without initial treatment. Gleason grade, nuclear grade and tumor volume turned all out to be independent prognostic factors. MUC-1, AMACR and Ki-67 also carried prognostic information. However, after adjustment for Gleason grade, nuclear grade and tumor volume only MUC-1 and AMACR remained as statistically significant prognostic factors. When tested for sensitivity and specificity they all failed and, consequently, they seem to be of less value in daily practice for cancelling an individual patient regarding the choice of treatment. Time trends in incidental prostate tumors in Sweden were analyzed in a cohort of patients with prostate tumors detected by transurethral resection (TUR-P). Through linkage of the national registration number (NRN) with several registers, e.g. the Swedish Cancer Registry, the National Inpatient registry and the Cause of Death Registry we identified, during the period 1970 through 2003, in total 23288 patients with incidental prostate cancer, who constituted the study group. As comparison group we choose all patients diagnosed with prostate cancer between 1970-2003 excluding those with incidental cancer, in total 112204 patients. Our result confirms earlier findings that there has been a dramatic change over time in incidence of incidental prostate cancers in Sweden, which parallels the introduction of prostate specific antigen. We also found that the cumulative incidence of prostate cancer death is high in the incidental group, opposing earlier findings that incidental tumours are a non-lethal disease.

Quality-of-life and functional outcomes are important in the choice of treatment for men with localized prostate cancer. These issues were investigated in the present thesis. All living 400 men randomized to radical prostatectomy or watchful waiting from 1989 to 1999 in the Scandinavian Prostate Cancer Group Number 4 (SPCG-4) were included. An additional 281 men compromised an age-matched control group. Physical symptoms, symptom-induced stress, sense of well-being and self-assessed quality of life were evaluated by a study-specific questionnaire. Results showed that prostate cancer men, regardless if they were allocated to radical prostatectomy or watchful waiting were suffering of long term adverse effects, mainly erectile dysfunction, urinary leakage and voiding symptoms. In the prostatectomy group, erectile dysfunction and urinary leakage were often consequences of surgery; in the watchful waiting group the side-effects could be caused by tumor progression. The quality of life deteriorated over time. High self-assessed quality of life was reported by 35 % in the radical, 34 % in watchful-waiting, and 43 % in the control groups after a median follow-up time of 12.2 years. The SPCG-4 men significantly more often reported anxiety than did controls. Erectile dysfunction was associated with the most negative influence on quality of life in both SPCG-4 groups. Men in the prostatectomy group were more distressed by erectile dysfunction than watchful waiting. Androgen deprivation therapy had negative effects on all psychological parameters, including quality of life, for the watchful waiting but not for the prostatectomy group. Information about the prostate-cancer disease was significantly higher in the radical-prostatectomy group than in watchful waiting. Check-ups were associated with worry, especially for those on androgen deprivation therapy. Open radical prostatectomy led to an increased rate of inguinal hernia compared with robot-assisted technique. In conclusion, the data of this thesis emphasize that it takes more than a decade to understand the patterns of adverse effects and time dimension of their occurrence for each treatment. Consideration of quality of life has a high priority to aid the ageing man through the shifting scenarios of localized prostate cancer.

Background: Prostate Cancer (PC) is a common and highly variable disease. Using current diagnostic methods, the prostate specific antigen (PSA) blood test and histological grading of prostate tissue needle biopsies, it is often difficult to evaluate whether the patient has a PC that requires active treatment or not. The absolute majority of all 10,000 cases of PCs diagnosed annually in Sweden have tumours graded as Gleason score (GS) 6-7 and a PSA value in blood below 10. Many of these are harmless and can be left without active treatment and hence spared problematic post-therapy side-effects, others are highly malignant and require early diagnosis and treatment. Better prognostic markers are needed and the aim of this study was to evaluate prognostic markers and to test if these markers could identify patients with indolent tumours. Methods: We have studied tumour material from 419 men consecutively diagnosed with PC at transurethral resection (1975-1990). The majority of these patients (295) had no metastasis at diagnosis and was not given any curative treatment and only hormonal treatment upon symptoms from metastatic progression. Standard histological sections and tissue microarrays (TMA) from these tumours and surrounding normal prostate tissue were stained and evaluated for cell proliferation (Ki67), blood vessels (endoglin and von Willebrand factor, vWf) and the extracellular matrix component hyaluronan (HA). An orthotopic rat PC model was used to explore hyaluronan staining, hyaluronic acid synthase (HAS)-1 mRNA levels and the effect of local HA treatment on tumour growth. Results: Tumour cell proliferation (Ki67) and the density of intra-tumoural endoglin stained blood vessels were independent prognostic markers (i.e. they added prognostic information to the conventional prognostic markers; clinical stage and GS). None of the GS 6 patients with low staining for both Ki67 and endoglin died of PC within 15 years of follow-up. High HA staining in the tumour epithelium and stroma was a negative prognostic marker of cancer specific survival but they were not independent of GS. High HA staining and high vascular density in the stroma of the surrounding morphologically normal prostate were prognostic for short cancer specific survival. Implantation of tumour cells in the normal rat prostate resulted in an increase in HA and HAS-1 mRNA levels in the prostate tissue surrounding prostate tumours. Concurrently intra-prostatic injection of HA also stimulated tumour growth. Conclusions: By evaluating both tumour cell proliferation (Ki67) and vascular density, it is possible to identify patients with very low risk of cancer specific death in the absence of active treatment. Prostate tumours influence the surrounding non-malignant prostate tissue, for example they cause an increased angiogenesis and synthesis of hyaluronan. Such responses can possibly be used to diagnose PC and to evaluate PC aggressiveness.

Prostate cancer (PCa) represents the most frequently diagnosed cancer and the second leading cause of cancer death in males. Initial development and progression of the disease is mainly regulated by androgens. However, the pathology of the disease may progress to a loss of hormone dependence, resulting in rapid growth and a metastatic phenotype. Invasion and metastasis of tumour cells results from the degradation of the basement membrane (BM) and extracellular matrix (ECM). The degradation of the BM and ECM is in part mediated by a family of proteinases called the matrix metalloproteinases (MMPs). Currently more than 20 members of the MMP family have been identified and they are further divided in to sub-classes according to their protein structure. Collectively, MMPs are capable of degrading essentially all ECM components. High expression of some MMPs correlates with a malignant phenotype of various tumours. This study focused on the expression and regulation of a sub-class of MMPs called the membrane-type MMPs (MT-MMPs) in PCa. To date 6 MT-MMPs have been identified and they are characterized by a transmembrane domain, followed by a short cytoplasmic tail (MT1-, MT2-, MT3- and MT5-MMPs) or a glycosylphosphatidylinositol (GPI) moiety (MT4- and MT6-MMPs). MT-MMPs are thought to play a key role in tumour cell invasion by virtue of their ability to activate MMP-2 (a secreted MMP, which is implicated in many metastatic tumours) and their direct degradation activity on ECM components. Elevated MT-MMP expression has been shown in breast, colon, skin, stomach, lung, pancreas and brain cancers. Until very recently there had been no studies conducted on MT-MMPs in PCa. The few studies preceding or occurring in parallel with this one, have mainly reported the mRNA expression of these enzymes in PCa. Most studies have focused on MT1-MMP. Thus, at the commencement of this project there were many unexplored aspects of the expression and regulation of the broader MT-MMP family in PCa. The aims of this study were to examine: 1 a) The expression of MT-MMPs in prostate cancer cell lines using RT-PCR and western blot analysis and b) expression of MT1-MMP and MT5-MMP in BPH (benign prostatic hyperplasia) and PCa clinical tissue sections by immunohistochemistry. 2) The regulation of MT1-MMP, MT3-MMP and MT5-MMP in PCa cell lines by Concanavalin A (Con A), phorbol-12-myristate 13-acetate (PMA), dihydrotestosterone (DHT) and insulin-like growth factors I and II (IGF I and IGF II) using western blot analysis. In this study RWPE1, a transformed but non-tumorigenic prostate cell line was used as a "normal" prostate cell model, ALVA-41 and LNCaP as androgen-dependent PCa cell models and DU-145 and PC-3 as androgen-independent PCa cell models. The mRNA expression for the 6 MT-MMPs was determined by RT-PCR. The results indicate that MT1- and MT3-MMP were detected in all cell lines. This is the first study to report MT1-MMP mRNA expression in LNCaP cells and MT3-MMP mRNA in DU-145 cells. MT2-MMP mRNA was detected in only LNCaP and DU-145 cells, whilst MT5-MMP was detected in PC-3, DU-145 and LNCaP cells. nterestingly, MT2-, MT4-, MT5- or MT6-MMP mRNA expression was not detected in the "normal" cell line RWPE1, perhaps indicating an induction in gene transcription in tumour cells. MT4-MMP mRNA was only detected in the androgen-independent cell lines, indicating a potential role in the invasion and metastasis processes of the aggressive androgen-independent PCa. In this study, very low expression of MT6-MMP was detected only in LNCaP and DU-145 cells. Previously there had been no reports on the expression of MT6-MMP in the normal or cancerous prostate. Due to the mRNA of MT1-, MT3- and MT5-MMPs being the predominant MT-MMPs expressed in the current study, and the availability of suitable antibodies against them, the protein expression of these three MT-MMPs was studied by western blot analysis. MT1-, MT3- and MT5-MMP protein expression was detected in the cell lysates and conditioned medium (CM) of RWPE1, LNCaP and PC-3 cells. For each MT-MMP, various protein species were detected including putative proforms, mature (active) forms, processed or fragmented forms as well as soluble or shed forms. The presence of soluble or shed forms of MT-MMPs in the CM of cultures of "normal" and PCa cells could imply one of the following mechanisms: ectodomain shedding by either extracellular sheddases, the secretion of intracellular processed proteins without the transmembrane domain, the release of membrane vesicles containing membrane-bound enzymes, or the presence of alternatively spliced mRNA, which gives rise to MT-MMPs without a transmembrane domain. Further characterization of these various forms, including their amino acid sequence, is required to fully elucidate their structural composition. Despite the detection of the mRNA, we did not detect the cell-associated proteins of MT1-MMP and MT5-MMP and only very low expression of MT3-MMP in DU-145 cells (CM of DU-145 cells were not screened for soluble forms of the enzymes). This is the first study to report MT5-MMP expression at the protein level in prostate derived cell lines. Immunohistochemistry was carried out on benign prostatic hyperplasia (BPH) and PCa clinical tissues using MT1- and MT5-MMP antibodies to determine their cellular localisation in benign and cancer glands. MT1- and MT5-MMPs were expressed in BPH and moderate and high grade PCa. MT1-MMP expression was highest in moderate grade cancer compared to BPH and high grade cancer. MT1-MMP expression was predominantly observed in the cytoplasm of secretory epithelial cells of both benign and cancer glands, although in cancer glands, some nuclear staining was also observed. Stromal expression of MT1-MMP was only observed in high grade cancer. This study is the first to report the immunolocalization of MT5-MMP outside the brain and in kidneys of diabetic patients. MT5-MMP was predominantly expressed in the cytoplasm of the secretory cells in benign glands. In the cancer glands, staining was heterogeneous with low to intense staining, mainly in the nuclei, plasma membrane and cytoplasm of secretory epithelial cells. Stromal expression of MT5-MMP was only observed in cancer tissues, particularly in high grade cancer. To study the regulation of MT-MMPs in PCa, we treated LNCaP and PC-3 cells, with either Con A, PMA, DHT or IGF-I and -II and studied the protein expression of MT1-, MT3- and MT5-MMPs by western blot analysis. Con A and PMA have been shown to stimulate MMP expression in other cell systems. Con A treatment showed a general increase in the protein expression of MT1-, MT3- and MT5-MMPs. By far the greatest induction by Con A observed was the nearly 4 fold increase in MT5-MMP expression caused by 40μg/mL Con A treatment of PC-3 cells. PMA treatment of LNCaP and PC-3 cells appeared to increase shedding or secretion of all three MT-MMPs in to the CM. This increase in the soluble forms corresponded to a decrease in cell-associated forms in LNCaP cells. Treatment of LNCaP with DHT alone and treatment of LNCaP and PC-3 cells with IGF-I and -II alone failed to detect any change in expression of MT1-MMP. The information gathered in this study on MT-MMPs with respect to cellular localization, expression levels and regulation by growth factors or chemicals that mimic their actions, will aid in our understanding of the role of MT-MMPs in PCa. This study provides strong preliminary data for further research, particularly with respect to functional studies of MT-MMPs in PCa. Understanding the processes which govern the actions of such proteins as these will provide potential insights into development of new management and therapeutic regimens to prevent cancer progression.

The androgen receptor (AR) fulfils important roles for both sexes. By mediating the biological function of androgens, the AR has remained the target for endocrine therapies treating prostate cancer. The AR also determines the effectiveness of medroxyprogesterone acetate (MPA) in treating AR positive breast cancer. Every man will be affected by prostate cancer if he lives long enough. Prostate cancer continues to be a leading cause of death for males despite research into this cancer covering more than 60 years since Huggins' seminal 1941 study showing that androgens play a key role in this cancer. Unfortunately, significant advances have not been forthcoming and the effect of treatment has remained largely the same over past decades, whereby initial treatment provides temporary remission but eventually advanced cases become refractory to further intervention and the disease recurs in a more aggressive form. A plethora of factors are exquisitely sensitive to minute changes in the AR's structural profile, which can be altered by a single mutation, resulting in aberrant activity. A principal feature of these variant ARs associated with prostate cancer, is enhanced capacity to bind a number of molecules other than its cognate ligand, dihydrotestosterone (DHT). The promiscuous activity of this receptor leads to continued AR signalling and stimulus for the cancer despite low androgen levels induced by treatment regimes. A key question is whether mutations occurring within the AR occur as a result of cancer or contribute to the propagation of the cancer. Recent research has demonstrated that treatments incorporating anti-androgens such as flutamide, which are designed to impede prostate cancer progression by inhibiting AR activity, may actually provide selective pressure favouring somatic mutation of the receptor to take place. The specific changes to the AR which are responsible for gains of function have not been resolved as their crystal structures, which are used to provide conformational analysis of proteins, are tremendously problematic to produce with little success found in literature. Generating representative crystals of the AR protein involves producing soluble recombinant protein. Unfortunately the AR is prone to aggregation and is highly unstable, especially in the presence of antagonistic molecules or absence of a stabilising ligand, preventing the protein from being maintained in the soluble state required for crystallization. In order to produce sufficient quantities of soluble material for crystallization, the androgen receptor's ligand binding domain (LBD) was produced as a recombinant protein in Escherichia coli bacteria strain BL21 (DE3) in the presence of DHT, flutamide, as well as in the absence of ligand. Since soluble unbound AR-LBD has not been produced until now, the bacterial culture containing no ligand was further processed to the stage of cleaving the purification tag from the recombinant protein and represents considerable progress into producing soluble material for crystallizing the troublesome yet considerably important AR in the absence of ligand. Although distinct from prostate cancer in males, AR activity in breast tissue is also a factor determining the action of drugs, such as MPA, included in therapies aimed at breast cancer. The use of MPA has declined primarily due to its adverse effects including unsuccessful generation of a biological response, as well as the advent of other drugs administered for hormonal therapies treating breast cancer. Alternative drugs are needed when breast cancer therapies fail as tumours develop resistance to primary drugs. Although there are a number of drugs on the market, success would be maximised if the determined therapy is matched with the patient, based for example, on their genetic makeup. There is a conundrum whereby some patients with an AR do not respond to MPA, a drug normally recognised by the receptor. In clinical trials it was discovered that an AR with threonine instead of methionine at residue 780 (M780T) fails to activate in response to MPA, but the exact mechanism has remained elusive and needs to be answered at the molecular level. The X-ray crystallographic studies that generate 3D images of macromolecules and wet chemistry, which have traditionally been used to provide insight into science in these dimensions, are incorporated with computer based molecular simulation. This is both complementary and distinct to traditional scientific methodologies, enabling further elucidation of protein-protein interactions, and the influence applied to such inter-relations by natural and drug ligands. This approach has been used, and is continually developed, to understand the binding mechanisms of current drugs as well as designing new drugs. In order to produce a receptor representing the M780T variant, the crystal structure representing the AR-LBD was mutated in silico, into which MPA was then docked. It was found that MPA binds into the M780T AR-LBD with considerably more spatial displacement compared to the position of DHT in the crystal structure, and is predicted to be the primary reason for the inability of MPA to activate this variant AR. The clarification of MPA binding and failure to elicit a response from the variant AR is significant for a cohort of breast cancer patients, as not only does the presence of an AR in the tumour determine the effectiveness of MPA, but correct composition of the AR, specifically, the absence of a M780T mutation. In the absence of this AR mutation, MPA could effectively be used either as an alternative to primary drugs, or in secondary therapies when primary therapies fail. Aberrant activity of variant ARs in response to MPA should also be taken into consideration when analysing drug studies about the effectiveness of MPA. The findings on the loss of response to MPA by the M780T variant AR have been included in the journal article &quotDecreased Androgen Receptor Levels and Receptor Function in Breast Cancer Contribute to the Failure of Response to Medroxyprogesterone Acetate" appearing in the September 2005 issue of Cancer Research journal.

In this study, we have explored the combination of a novel Purine Nucleoside Phosphorylase mediated Gene-Directed Enzyme Prodrug Therapy (PNP-GDEPT) with chemotherapeutics, Taxotere and/or Carboplatin to target prostate and ovarian cancer (PC & OC). PNP converts the prodrug (Fludarabine-phosphate) to a toxic purine, 2-fluoroadenine (2FA) that inhibits RNA/DNA synthesis. Taxotere is active against late stage PC whilst carboplatin is first line therapy for OC. Neither modality is adequately effective. We expect that a combination will target heterogeneity via cytotoxicity to diverse cancer cell populations leading to effective synergies, which may improve efficacy and quality of life. For PC, Synergy between Ad-PNP-GDEPT and Taxotere were assessed in vitro and in vivo. Cell killing effects of combination led to significant synergistic killing of human PC-3 & murine RM1 PC cells accompanied by enhanced apoptosis. A lower individual dose (by up to 8 fold) led to enhanced efficacy. In vivo, the combination regimen given at the suboptimal doses led to reduction in local tumour (PC-3 & RM1) growth in nude and in C57BL/6 mice, respectively. A significant reduction in lung RM1 colony numbers indicated enhanced systemic efficacy. Combination treated mice also displayed significantly improved survival (25 days vs 15 days for control mice). Importantly, the condition of combination treated mice (e.g. weight loss) was better than those given individual treatments. The possible involvement of the immune system in this enhanced effect is under investigation. For OC, three-way synergy between Ad-PNP-GDEPT, Taxotere and carboplatin was effectively demonstrated in SKOV-3 and OVCAR-3 cells. This was significantly greater than bimodal or individual treatments. A 10-50 fold dose reduction of individual treatments was effective when combined, accompanied by enhanced apoptosis. Western-blotting analyses revealed a shift in the expression of anti-apoptotic and proapoptotic proteins upon treatment with various combinations. This is the first demonstration of synergy between these modalities.

ABSTRACT The morphological premalignant changes in prostate epithelium such as high grade prostatic intraepithelial neoplasia (HGPIN) precede invasive prostate cancer (PC) by several decades. The overall aim of this project was to identify patterns of gene expression in HGPIN and early PC which increase our understanding of the early biology of PC and identify genes and pathways that correlate with an aggressive phenotype. A comprehensive tissue cohort of premalignant prostate lesions was collected in a tissue microarray (TMA) platform that was utilised for high-throughput validation of target genes. Using this unique resource, the expression of the tumour suppressor gene PTEN was assessed using immunohistochemistry in an initial candidate gene approach based on mouse models implicating PTEN in carcinogenesis. No significant difference in expression of PTEN was detected in premalignant and benign epithelium. A transcript profiling approach was undertaken by integrating laser capture microdissection, linear RNA amplification and oligonucleotide microarrays to perform a screen of matched patient samples of normal, HGPIN and PC cells. The expression patterns of two genes encoding secreted proteins, neuropeptide Y (NPY) and macrophage inhibitory cytokine (MIC-1) were validated using immunohistochemistry on TMAs representing the progression model of early PC. Increased expression of these proteins in PC was confirmed to occur early in the disease process and altered expression of NPY and MIC-1 was associated with worse clinical outcome. Further analysis of global gene expression patterns using a structured network knowledge base identified a notable aberration in the expression of extracellular matrix and extracellular matrix associated proteins in HGPIN and provided novel evidence for the role of this class of molecules in the development of PC. In summary, contrary to current dogma based on work in animal models, altered PTEN expression is unlikely to represent an important event in the development of malignancy in the human prostate. In contrast, the expression patterns and prognostic value of NPY and MIC-1 in HGPIN support their further evaluation as biomarkers for the development and progression of PC. The aberrant expression of genes and networks of genes detected in HGPIN will assist in further identification of biological pathways which may be targeted in therapeutic strategies against the development and progression of PC.