Multiparametric magnetic resonance imaging of the prostate: current concepts

Multiparametric MR (mpMR) imaging is rapidly evolving into the mainstay in prostate cancer (PCa) imaging. Generally, the examination consists of T2-weighted sequences, diffusion-weighted imaging (DWI), dynamic contrast-enhanced (DCE) evaluation, and less often proton MR spectroscopy imaging (MRSI). Those functional techniques are related to biological properties of the tumor, so that DWI correlates to cellularity and Gleason scores, DCE correlates to angiogenesis, and MRSI correlates to cell membrane turnover. The combined use of those techniques enhances the diagnostic confidence and allows for better characterization of PCa. The present article reviews and illustrates the technical aspects and clinical applications of each component of mpMR imaging, in a practical approach from the urological standpoint.

Role and Function of c-Jun Protein Complex in Cancer Cell Behavior

Transcription factors play a crucial role in the regulation of cell behavior by modulating gene expression profiles. Previous studies have described a dual role for the AP-1 family transcription factor c-Jun in the regulation of cellular fate. In various cell types weak and transient activations of c-Jun N-terminal kinase (JNK) and c-Jun appear to contribute to proliferation and survival, whereas strong and prolonged activation of JNK and c-Jun result in apoptosis. These opposite roles played by c-Jun are cell type specific and the molecular mechanisms defining these antonymous c-Jun-mediated responses remain incompletely understood. c-Jun activity in transformed cells is regulated by signalling cascades downstream of oncoproteins such as Ras and Raf. In addition, the pro-proliferative role and the survival promoting function for c-Jun has been described in various cancer models. Furthermore, c-Jun was described to be overexpressed in different cancer types. However, the molecular mechanisms by which c-Jun exerts these oncogenic functions are not all clearly established. Therefore it is of primary interest to further identify molecular mechanisms and functions for c-Jun in cancer. Regulation of gene expression is tightly dependent on accurate protein-protein interactions. Therefore, co-factors for c-Jun may define the functions for c-Jun in cancer. Identification of protein-protein interactions promoting cancer may provide novel possibilities for cancer treatment. In this study, we show that DNA topoisomerase I (TopoI) is a transcriptional co-factor for c-Jun. Moreover, c-Jun and TopoI together promote expression of epidermal growth factor receptor (EGFR) in cancer cells. We also show that the clinically used TopoI inhibitor topotecan reduces EGFR expression. Importantly, the effect of TopoI on EGFR transcription was shown to depend on c-Jun as Jun-/- cells or cells treated with JNK inhibitor SP600125 are resistant to topotecan treatment both in regulation of EGFR expression and cell proliferation. Moreover, c-Jun regulates the nucleolar localization and the function of the ribonucleic acid (RNA) helicase DDX21, a previously identified member of c-Jun protein complex. In addition, c-Jun stimulates rRNA processing by supporting DDX21 rRNA binding. Finally, this study characterizes a DDX21 dependent expression of cyclin dependent kinase (Cdk) 6, a correlation of DDX21 expression with prostate cancer progression and a substrate binding dependency of DDX21 nucleolar localization in prostate cancer cells. Taken together, the results of this study validate the c-Jun-TopoI interaction and precise the c-Jun-DDX21 interaction. Moreover, these results show the importance for protein-protein interaction in the regulation of their cellular functions in cancer cell behavior. Finally, the results presented here disclose new exciting therapeutic opportunities for cancer treatment.

Fibroblast Growth Factor 8 and its Receptors in The Growth and Angiogenesis of Experimental Breast Cancer . With Special Reference to Thrombospondin-1 as a Novel Target Gene for FGF-8

The growth of breast cancer is regulated by hormones and growth factors. Recently, aberrant fibroblast growth factor (FGF) signalling has been strongly implicated in promoting the progression of breast cancer and is thought to have a role in the development of endocrine resistant disease. FGFs mediate their auto- and paracrine signals through binding to FGF receptors 1-4 (FGFR1-4) and their isoforms. Specific targets of FGFs in breast cancer cells and the differential role of FGFRs, however, are poorly described. FGF-8 is expressed at elevated levels in breast cancer, and it has been shown to act as an angiogenic, growth promoting factor in experimental models of breast cancer. Furthermore, it plays an important role in mediating androgen effects in prostate cancer and in some breast cancer cell lines. We aimed to study testosterone (Te) and FGF-8 regulated genes in Shionogi 115 (S115) breast cancer cells, characterise FGF-8 activated intracellular signalling pathways and clarify the role of FGFR1, -2 and -3 in these cells. Thrombospondin-1 (TSP-1), an endogenous inhibitor of angiogenesis, was recognised as a Te and FGF-8 regulated gene. Te repression of TSP-1 was androgen receptor (AR)-dependent. It required de novo protein synthesis, but it was independent of FGF-8 expression. FGF-8, in turn, downregulated TSP-1 transcription by activating the ERK and PI3K pathways, and the effect could be reversed by specific kinase inhibitors. Differential FGFR1-3 action was studied by silencing each receptor by shRNA expression in S115 cells. FGFR1 expression was a prerequisite for the growth of S115 tumours, whereas FGFR2 expression alone was not able to promote tumour growth. High FGFR1 expression led to a growth advantage that was associated with strong ERK activation, increased angiogenesis and reduced apoptosis, and all of these effects could be reversed by an FGFR inhibitor. Taken together, the results of this thesis show that FGF-8 and FGFRs contribute strongly to the regulation of the growth and angiogenesis of experimental breast cancer and support the evidence for FGF-FGFR signalling as one of the major players in breast cancers.

First-Line Chemotherapy with Anthracycline and Taxane Combination in Metastatic Breast Cancer. Detection of bone metastases with TRACP 5b

Background: In Finland, breast cancer (BC) is the most common cancer among women, and prostate cancer (PC) that among men. At the metastatic stage both cancers remain essentially incurable. The goals of therapy include palliation of symptoms, improvement or maintenance of quality of life (QoL), delay of disease progression, and prolongation of survival. Balancing between efficacy and toxicity is the major challenge. With increasing costs of new treatments, appropriate use of resources is paramount. When new treatment regimes are introduced into clinical practice a comprehensive assessment of clinical benefit, adverse effects and cost is necessary. Both BC and PC show a predilection to metastasize to bone. Bone metastases cause significant morbidity impairing the patients´ QoL. Diagnosis of bone metastases relies mainly on radiological methods, which however lack optimal sensitivity and specificity. New tools are needed for detection and follow-up of bone metastases. Aims: Anthracyclines and taxanes are effective chemotherapeutic agents in the treatment of metastatic breast cancer (MBC) with different mechanisms of action. Therefore, evaluation of the combination of anthracyclines with taxanes was a justifiable approach in the treatment of MBC patients. We assessed the efficacy, toxicity, cost of treatment and QoL of BC patients treated with first-line chemotherapy for metastatic disease with the combination epirubicin and docetaxel. We also evaluated the diagnostic potential of tartrate-resistant acid phosphatase 5b (TRACP 5b) and carboxyterminal telopeptides of type I collagen (ICTP) in the diagnosis of bone metastases in BC and TRACP 5b in PC patients. Results: The combination of epirubicin and docetaxel was effective in this phase II study, but required individual dose adjustment to avoid neutropenic infections, and the use of growth factors to maintain a feasible dose level. The response rate was 54 % (95 % CI 37-71) and the median overall survival (OS) was 26 months. Of the patients, 87 % were treated for infections. The treatment of adverse events required additional use of health resources mainly due to neutropenic infections, thereby raising direct treatment costs by 20 %. Despite adverse events, the global QoL was not significantly compromised during the treatment. Clinically evident acute cardiac toxicity was not observed. The combination of serum TRACP 5b and ICTP was at least equally sensitive and specific in detection of of bone metastases as commonly used total alkaline phosphatise (tALP) in BC patients. In contrast, TRACP 5b was less specific and sensitive than tALP as a marker of skeletal changes in PC patients. Conclusions: Treatment with epirubicin and docetaxel showed high efficacy in first-line chemotherapy of MBC. The relatively high incidence of neutropenic infections requiring hospitalization increased the treatment costs. Despite adverse events, the global QoL of the patients was not significantly compromised. The combination of TRACP 5b and ICTP showed similar activity as tALP in detecting bone metastases in MBC. In contrast, TRACP 5b was less specific and sensitive than tALP as a marker of skeletal changes in PC.

Lethal weapons : novel approaches for receptor-targeted cancer cell elimination

The currently used forms of cancer therapy are associated with drug resistance and toxicity to healthy tissues. Thus, more efficient methods are needed for cancer-specific induction of growth arrest and programmed cell death, also known as apoptosis. Therapeutic forms of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) are investigated in clinical trials due to the capability of TRAIL to trigger apoptosis specifically in cancer cells by activation of cell surface death receptors. Many tumors, however, have acquired resistance to TRAIL-induced apoptosis and sensitizing drugs for combinatorial treatments are, therefore, in high demand. This study demonstrates that lignans, natural polyphenols enriched in seeds and cereal, have a remarkable sensitizing effect on TRAIL-induced cell death at non-toxic lignan concentrations. In TRAIL-resistant and androgen-dependent prostate cancer cells we observe that lignans repress receptor tyrosine kinase (RTK) activity and downregulate cell survival signaling via the Akt pathway, which leads to increased TRAIL sensitivity. A structure-activity relationship analysis reveals that the γ-butyrolactone ring of the dibenzylbutyrolactone lignans is essential for the rapidly reversible TRAIL-sensitizing activity of these compounds. Furthermore, the lignan nortrachelogenin (NTG) is identified as the most efficient of the 27 tested lignans and norlignans in sensitization of androgen-deprived prostate cancer cells to TRAIL-induced apoptosis. While this combinatorial anticancer approach may leave normal cells unharmed, several efficient cancer drugs are too toxic, insoluble or unstable to be used in systemic therapy. To enable use of such drugs and to protect normal cells from cytotoxic effects, cancer-targeted drug delivery vehicles of nanometer scale have recently been generated. The newly developed nanoparticle system that we tested in vitro for cancer cell targeting combines the efficient drug-loading capacity of mesoporous silica to the versatile particle surface functionalization of hyperbranched poly(ethylene imine), PEI. The mesoporous hybrid silica nanoparticles (MSNs) were functionalized with folic acid to promote targeted internalization by folate receptor overexpressing cancer cells. The presented results demonstrate that the developed carrier system can be employed in vitro for cancer selective delivery of adsorbed or covalently conjugated molecules and furthermore, for selective induction of apoptotic cell death in folate receptor expressing cancer cells. The tested carrier system displays potential for simultaneous delivery of several anticancer agents specifically to cancer cells also in vivo.

A Farewell to Flat Biology. Three-dimensional Cell Culture Models in Cancer Drug Target Identification and Validation

Cells of epithelial origin, e.g. from breast and prostate cancers, effectively differentiate into complex multicellular structures when cultured in three-dimensions (3D) instead of conventional two-dimensional (2D) adherent surfaces. The spectrum of different organotypic morphologies is highly dependent on the culture environment that can be either non-adherent or scaffold-based. When embedded in physiological extracellular matrices (ECMs), such as laminin-rich basement membrane extracts, normal epithelial cells differentiate into acinar spheroids reminiscent of glandular ductal structures. Transformed cancer cells, in contrast, typically fail to undergo acinar morphogenic patterns, forming poorly differentiated or invasive multicellular structures. The 3D cancer spheroids are widely accepted to better recapitulate various tumorigenic processes and drug responses. So far, however, 3D models have been employed predominantly in the Academia, whereas the pharmaceutical industry has yet to adopt a more widely and routine use. This is mainly due to poor characterisation of cell models, lack of standardised workflows and high throughput cell culture platforms, and the availability of proper readout and quantification tools. In this thesis, a complete workflow has been established entailing well-characterised 3D cell culture models for prostate cancer, a standardised 3D cell culture routine based on high-throughput-ready platform, automated image acquisition with concomitant morphometric image analysis, and data visualisation, in order to enable large-scale high-content screens. Our integrated suite of software and statistical analysis tools were optimised and validated using a comprehensive panel of prostate cancer cell lines and 3D models. The tools quantify multiple key cancer-relevant morphological features, ranging from cancer cell invasion through multicellular differentiation to growth, and detect dynamic changes both in morphology and function, such as cell death and apoptosis, in response to experimental perturbations including RNA interference and small molecule inhibitors. Our panel of cell lines included many non-transformed and most currently available classic prostate cancer cell lines, which were characterised for their morphogenetic properties in 3D laminin-rich ECM. The phenotypes and gene expression profiles were evaluated concerning their relevance for pre-clinical drug discovery, disease modelling and basic research. In addition, a spontaneous model for invasive transformation was discovered, displaying a highdegree of epithelial plasticity. This plasticity is mediated by an abundant bioactive serum lipid, lysophosphatidic acid (LPA), and its receptor LPAR1. The invasive transformation was caused by abrupt cytoskeletal rearrangement through impaired G protein alpha 12/13 and RhoA/ROCK, and mediated by upregulated adenylyl cyclase/cyclic AMP (cAMP)/protein kinase A, and Rac/ PAK pathways. The spontaneous invasion model tangibly exemplifies the biological relevance of organotypic cell culture models. Overall, this thesis work underlines the power of novel morphometric screening tools in drug discovery.

The Effects of Fibroblast Growth Factor 8b on Reproductive Organs and Prostate Tumorigenesis

Fibroblast growth factors (FGFs) are involved in the development and homeostasis of the prostate and other reproductive organs. FGF signaling is altered in prostate cancer. Fibroblast growth factor 8 (FGF8) is a mitogenic growth factor and its expression is elevated in prostate cancer and in premalignant prostatic intraepithelial neoplasia (PIN) lesions. FGF8b is the most transforming isoform of FGF8. Experimental models show that FGF8b promotes several phases of prostate tumorigenesis - including cancer initiation, tumor growth, angiogenesis, invasion and development of bone metastasis. The mechanisms activated by FGF8b in the prostate are unclear. In the present study, to examine the tumorigenic effects of FGF8b on the prostate and other FGF8b expressing organs, an FGF8b transgenic (TG) mouse model was generated. The effect of estrogen receptor beta (ERβ) deficiency on FGF8binduced prostate tumorigenesis was studied by breeding FGF8b-TG mice with ERβ knockout mice (BERKOFVB). Overexpression of FGF8b caused progressive histological and morphological changes in the prostate, epididymis and testis of FGF8b-TG-mice. In the prostate, hyperplastic, preneoplastic and neoplastic changes, including mouse PIN (mPIN) lesions, adenocarcinomas, sarcomas and carcinosarcomas were present in the epithelium and stroma. In the epididymis, a highly cancer-resistant tissue, the epithelium contained dysplasias and the stroma had neoplasias and hyperplasias with atypical cells. Besides similar histological changes in the prostate and epididymis, overexpression of FGF8b induced similar changes in the expression of genes such as osteopontin (Spp1), connective tissue growth factor (Ctgf) and FGF receptors (Fgfrs) in these two tissues. In the testes of the FGF8b-TG mice, the seminiferous epithelium was frequently degenerative and the number of spermatids was decreased. A portion of the FGF8b-TG male mice was infertile. Deficiency of ERβ did not accelerate prostate tumorigenesis in the FGF8b-TG mice, but increased significantly the frequency of mucinous metaplasia and slightly the frequency of inflammation in the prostate. This suggests putative differentiation promoting and anti-inflammatory roles for ERβ. In summary, these results underscore the importance of FGF signaling in male reproductive organs and provide novel evidence for a role of FGF8b in stromal activation and prostate tumorigenesis.

Analysis of p53 expression and proliferative assessment using PCNA in localized prostate carcinoma

The surgical specimens from 51 men submitted to radical prostatectomy for localized prostate cancer were examined by immunohistochemistry using proliferation cell nuclear antigen (PCNA) monoclonal antibody to evaluate the proliferative index (PI). The relationship between PI, biological variables and p53 protein expression was evaluated by immunohistochemistry. PI was low in invasive localized prostate carcinoma (mean, 12.4%) and the incidence of PCNA-positive cells was significantly higher in tumors with p53 expression (P = 0.0226). There was no statistical difference in PCNA values when biological parameters such as Gleason score, tumor volume, extraprostatic involvement, seminal vesicle infiltration or lymph node metastasis were considered. We conclude that proliferative activity is usually low in prostate carcinoma but is correlated with p53 immune staining, indicating that p53 is important in cell cycle control in this neoplasm.

Profile of thyroid hormones in breast cancer patients

Estrogen involvement in breast cancer has been established; however, the association between breast cancer and thyroid diseases is controversial. Estrogen-like effects of thyroid hormone on breast cancer cell growth in culture have been reported. The objective of the present study was to determine the profile of thyroid hormones in breast cancer patients. Serum aliquots from 26 patients with breast cancer ranging in age from 30 to 85 years and age-matched normal controls (N = 22) were analyzed for free triiodothyronine (T3F), free thyroxine (T4F), thyroid-stimulating hormone (TSH), antiperoxidase antibody (TPO), and estradiol (E2). Estrogen receptor ß (ERß) was determined in tumor tissues by immunohistochemistry. Thyroid disease incidence was higher in patients than in controls (58 vs 18%, P < 0.05). Subclinical hyperthyroidism was the most frequent disorder in patients (31%); hypothyroidism (8%) and positive anti-TPO antibodies (19%) were also found. Subclinical hypothyroidism was the only dysfunction (18%) found in controls. Hyperthyroidism was associated with postmenopausal patients, as shown by significantly higher mean T3 and T4 values and lower TSH levels in this group of breast cancer patients than in controls. The majority of positive ERß tumors were clustered in the postmenopausal patients and all cases presenting subclinical hyperthyroidism in this subgroup concomitantly exhibited Erß-positive tumors. Subclinical hyperthyroidism was present in only one of 6 premenopausal patients. We show here that postmenopausal breast cancer patients have a significantly increased thyroid hormone/E2 ratio (P < 0.05), suggesting a possible tumor growth-promoting effect caused by this misbalance.

Lycopene and ß-carotene protect in vivo iron-induced oxidative stress damage in rat prostate

It has been suggested that iron overload may be carcinogenic. In the present study, we evaluated the effect of plasma and prostate carotenoid concentration on oxidative DNA damage in 12-week-old Wistar rats treated with intraperitoneal (ip) ferric nitrilotriacetate (Fe-NTA) (10 mg Fe/kg). Plasma ß-carotene and lycopene concentrations were measured as a function of time after ip injection of carotenoids (10 mg kg-1 day-1 ß-carotene or lycopene) in rats. The highest total plasma concentration was reached 3 and 6 h after ip injection of lycopene or ß-carotene, respectively. After 5 days of carotenoid treatment, lycopene and ß-carotene were present in the 0.10-0.51 nmol/g wet tissue range in the prostate. Using a sensitive method to detected 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) by HPLC/EC, the level of 8-oxodGuo in rat prostate DNA was significantly higher (6.3 ± 0.6 residues/10(6) dGuo) 3 h after Fe-NTA injection compared with control rats (1.7 ± 0.3 residues/10(6) dGuo). Rats supplemented with lycopene or ß-carotene for 5 days prior to Fe-NTA treatment showed a reduction of about 70% in 8-oxodGuo levels to almost control levels. Compared with control rats, the prostate of Fe-NTA-treated animals showed a 78% increase in malondialdehyde accumulation. Lycopene or ß-carotene pre-treatment almost completely prevented lipid damage. Epidemiological studies have suggested a lower risk of prostate cancer in men reporting a higher consumption of tomato products. However, before associating this effect with tomato sauce constituents, more information is required. The results described here may contribute to the understanding of the protective effects of carotenoids against iron-induced oxidative stress.

Effect of the consumption of tomato paste on plasma prostate-specific antigen levels in patients with benign prostate hyperplasia

The consumption of tomatoes and tomato products has been associated with a reduced risk of prostate cancer. We observed a decrease of 10.77% in prostate-specific antigen (PSA) levels in patients with benign prostate hyperplasia who were submitted to daily ingestion of tomato paste. This was an experimental rather than a controlled study with a sample of 43 men ranging in age from 45 to 75 years, all with histological diagnoses of benign prostate hyperplasia and plasma PSA levels of 4-10 ng/mL. All patients received 50 g of tomato paste once a day for 10 consecutive weeks and PSA levels were analyzed before, during and after the consumption of tomato paste. ANOVA for repeated measures was used to compare PSA levels before, during and after the consumption of tomato paste. The mean ± SD PSA level was 6.51 ± 1.48 ng/mL at baseline and 5.81 ± 1.58 ng/mL (P = 0.005) after 10 weeks. Acceptance was good in 88.3, regular in 9.3, and poor in 2.3% of the patients. Dietary ingestion of 50 g of tomato paste per day for 10 weeks significantly reduced mean plasma PSA levels in patients with benign prostate hyperplasia, probably as a result of the high amount of lycopene in tomato paste. This was not a prostate cancer prevention study, but showed some action of tomato paste in prostate biology. The development of prostate cancer is typically accompanied by an increase in plasma PSA levels, thus any intervention that affects plasma PSA levels can suggest an impact in the progression of disease.

Construction of a recombinant adenovirus co-expressing truncated human prostate-specific membrane antigen and mouse 4-1BBL genes and its effect on dendritic cells

Our aim was to construct a recombinant adenovirus co-expressing truncated human prostate-specific membrane antigen (tPSMA) and mouse 4-1BBL genes and to determine its effect on dendritic cells (DCs) generated from bone marrow suspensions harvested from C57BL/6 mice for which the effect of 4-1BBL on DCs is not clear, especially during DCs processing tumor-associated antigen. Replication deficient adenovirus AdMaxTM Expression System was used to construct recombinant adenovirus Ad-tPSMA-internal ribosome entry site-mouse 4-1BBL (Ad-tPSMA-IRES-m4-1BBL) and Ad-enhanced green fluorescent protein. Day 7 proliferating DC aggregates generated from C57BL/6 mice were collected as immature DCs and further mature DCs were obtained by lipopolysaccharide activated immature DCs. After DCs were exposed to the recombinant adenovirus with 250 multiplicity of infection, the expression of tPSMA and m4-1BBL proteins were detected by Western blot, and the apoptosis and phenotype of DCs were analyzed by flow cytometry. Cytokines (IL-6 and IL-12) in the supernatant were detected by enzyme-linked immunosorbent assay (ELISA). Proliferation of T cells was detected by allogeneic mixed lymphocyte reactions. The tPSMA and m4-1BBL proteins were expressed correctly. The apoptosis rate of DCs transfected with Ad-tPSMA-IRES-m4-1BBL was 14.6%, lower than that of control DCs. The expression of co-stimulatory molecules [CD80 (81.6 ± 5.4%) and CD86 (80.13 ± 2.81%)] up-regulated in Ad-tPSMA-IRES-m4-1BBL-pulsed DCs, and the level of IL-6 (3960.2 ± 50.54 pg/mL) and IL-12 (249.57 ± 12.51 pg/mL) production in Ad-tPSMA-IRES-m4-1BBL-transduced DCs were significantly higher (P < 0.05) than those in control DCs. Ad-tPSMA-IRES-m4-1BBL induced higher T-cell proliferation (OD450 = 0.614 ± 0.018), indicating that this recombinant adenovirus can effectively enhance the activity of DCs.

Cyclin D1 expression in prostate carcinoma

The purpose of this study was to investigate the relationship between cyclin D1 expression and clinicopathological parameters in patients with prostate carcinoma. We assessed cyclin D1 expression by conventional immunohistochemistry in 85 patients who underwent radical prostatectomy for prostate carcinoma and 10 normal prostate tissue samples retrieved from autopsies. We measured nuclear immunostaining in the entire tumor area and based the results on the percentage of positive tumor cells. The preoperative prostate-specific antigen (PSA) level was 8.68±5.16 ng/mL (mean±SD). Cyclin D1 staining was positive (cyclin D1 expression in >5% of tumor cells) in 64 cases (75.4%) and negative (cyclin D1 expression in ≤5% of tumor cells) in 21 cases (including 15 cases with no immunostaining). Normal prostate tissues were negative for cyclin D1. Among patients with a high-grade Gleason score (≥7), 86% of patients demonstrated cyclin D1 immunostaining of >5% (P<0.05). In the crude analysis of cyclin D1 expression, the high-grade Gleason score group showed a mean expression of 39.6%, compared to 26.9% in the low-grade Gleason score group (P<0.05). Perineural invasion tended to be associated with cyclin D1 expression (P=0.07), whereas cyclin D1 expression was not associated with PSA levels or other parameters. Our results suggest that high cyclin D1 expression could be a potential marker for tumor aggressiveness.

Accuracy of dose planning for prostate radiotherapy in the presence of metallic implants evaluated by electron spin resonance dosimetry

Radiotherapy is one of the main approaches to cure prostate cancer, and its success depends on the accuracy of dose planning. A complicating factor is the presence of a metallic prosthesis in the femur and pelvis, which is becoming more common in elderly populations. The goal of this work was to perform dose measurements to check the accuracy of radiotherapy treatment planning under these complicated conditions. To accomplish this, a scale phantom of an adult pelvic region was used with alanine dosimeters inserted in the prostate region. This phantom was irradiated according to the planned treatment under the following three conditions: with two metallic prostheses in the region of the femur head, with only one prosthesis, and without any prostheses. The combined relative standard uncertainty of dose measurement by electron spin resonance (ESR)/alanine was 5.05%, whereas the combined relative standard uncertainty of the applied dose was 3.35%, resulting in a combined relative standard uncertainty of the whole process of 6.06%. The ESR dosimetry indicated that there was no difference (P>0.05, ANOVA) in dosage between the planned dose and treatments. The results are in the range of the planned dose, within the combined relative uncertainty, demonstrating that the treatment-planning system compensates for the effects caused by the presence of femur and hip metal prostheses.

From epithelial to mesenchymal: regulation of invasive cancer cell motility

Metastasis is the main cause of death among cancer patients. In order to initiate the metastatic cascade cancer cells have to undergo epithelial-to-mesenchymal transition (EMT). In EMT epithelial cells lose their cell-cell and cell-extracellular matrix (ECM) contacts and become more motile. The expression of the transcription factor Slug and of the mesenchymal intermediate filament vimentin is induced during EMT. Vimentin is often overexpressed in malignant epithelial cancers but the functional role of vimentin remains incompletely understood. In addition, kinases such as AKT and ERK are known to be involved in the regulation of EMT and cancer cell motility but the mechanisms underlining their functions are often unclear. Integrins are heterodimeric receptors that attach cells to the surrounding tissue and participate in regulating cell migration and invasion. Changes in integrin activity are linked to increased cell motility and further cancer metastasis. The aim for my PhD studies was to investigate the role of cellular signalling pathways and vimentin in the regulation of cancer cell motility and EMT. Our results revealed that in prostate cancer the downregulation of AKT1 and AKT2, but not AKT3, induces activation of cell surface 1-integrins leading to enhanced cell adhesion, migration and invasion. In addition, our findings demonstrated a reciprocal regulatory interaction between vimentin and ERK2 facilitating ERK-mediated phosphorylation of Slug at serine-87 (S87) in breast cancer. Surprisingly, Slug S87 phosphorylation is dispensable for E-cadherin repression but essential for the induction of vimentin and Axl expression in early onset of EMT. Our findings reveal previously unknown mechanistic information of how prostate and breast cancer cell motility and disease progression is regulated