Foxm1 expression in prostate epithelial cells is essential for prostate carcinogenesis.

The treatment of advanced prostate cancer (PCa) remains a challenge. Identification of new molecular mechanisms that regulate PCa initiation and progression would provide targets for the development of new cancer treatments. The Foxm1 transcription factor is highly up-regulated in tumor cells, inflammatory cells, and cells of tumor microenvironment. However, its functions in different cell populations of PCa lesions are unknown. To determine the role of Foxm1 in tumor cells during PCa development, we generated two novel transgenic mouse models, one exhibiting Foxm1 gain-of-function and one exhibiting Foxm1 loss-of-function under control of the prostate epithelial-specific Probasin promoter. In the transgenic adenocarcinoma mouse prostate (TRAMP) model of PCa that uses SV40 large T antigen to induce PCa, loss of Foxm1 decreased tumor growth and metastasis. Decreased prostate tumorigenesis was associated with a decrease in tumor cell proliferation and the down-regulation of genes critical for cell proliferation and tumor metastasis, including Cdc25b, Cyclin B1, Plk-1, Lox, and Versican. In addition, tumor-associated angiogenesis was decreased, coinciding with reduced Vegf-A expression. The mRNA and protein levels of 11β-Hsd2, an enzyme playing an important role in tumor cell proliferation, were down-regulated in Foxm1-deficient PCa tumors in vivo and in Foxm1-depleted TRAMP C2 cells in vitro. Foxm1 bound to, and increased transcriptional activity of, the mouse 11β-Hsd2 promoter through the -892/-879 region, indicating that 11β-Hsd2 was a direct transcriptional target of Foxm1. Without TRAMP, overexpression of Foxm1 either alone or in combination with inhibition of a p19(ARF) tumor suppressor caused a robust epithelial hyperplasia, but was insufficient to induce progression from hyperplasia to PCa. Foxm1 expression in prostate epithelial cells is critical for prostate carcinogenesis, suggesting that inhibition of Foxm1 is a promising therapeutic approach for prostate cancer chemotherapy.

Health technology assessment in evolution - focal therapy in localised prostate cancer.

Focal therapy in prostate cancer aims to treat only the part of the gland harboring clinically significant disease while preserving the rest of the tissue. This approach may substantially reduce treatment-related toxicity without compromising disease control outcomes. Short- to medium-term functional and oncological results in prospective interventional studies are promising, but comparative effectiveness research against standard of care is required to incorporate focal therapy among standard options. In this review, we discuss the actual stage of assessment and results of sources of energy commonly used to deliver focal therapy. We also provide our viewpoint on how the field will evolve in the near future.

Mécanismes moléculaires de la sécrétion d'insuline : implication de Slac2c/MyRIP, protéine effectrice de Rab27a, et rôle des phosphoinositides dans le processus d'exocytose

Résumé : La sécrétion de l'insuline en réponse au glucose circulant dans le sang est la fonction principale de la cellule β. La perte de cette fonction est une des caractéristiques du diabète de type 2. L'exocytose est une fonction cellulaire indispensable au renouvellement des composants lipidiques et protéiques de la membrane cellulaire, à la communication entre les cellules et au maintien d'un environnement adéquat. On peut distinguer deux types d'exocytose : l'exocytose constitutive et l'exocytose régulée. Cette dernière est déclenchée par des stimuli externes. L'exocytose régulée est contrôlée au niveau de la fusion des vésicules de sécrétion avec la membrane plasmique. Certains composants moléculaires impliqués dans ce processus font partie de la famille des GTPases Rab. Les deux membres de cette famille impliqués sont Rab3 et Rab27. Nous avons étudié le rôle de la GTPase Rab27 dans les cellules INS-1E, une lignée cellulaire pancréatique β qui sécrète de l'insuline de façon régulée. Nous avons trouvé que la diminution d'expression de la protéine en utilisant le technique de « RNA interference » diminue la sécrétion stimulée, mais que la distribution des granules n'est nullement affectées par ce changement d'activité intrinsèque. Un des effecteurs identifiés de cette GTPase est Slac2c/MyRIP. Cette protéine possède plusieurs domaines fonctionnels dont un qui lui permet de se lier à l'actine, constituant du cytosquelette cellulaire. L'ensemble de nos résultats suggèrent que Rab27 et MyRIP font partie d'un complexe permettant l'interaction de la granule de sécrétion avec le cytosquelette d'actine corticale et participent à la régulation des dernières étapes de l'exocytose d'insuline. Ensuite, nous avons étudié les phosphoinositides (PI). Les phosphoinositides sont d'importantes molécules impliquées dans le régulation du trafic vésiculaire. Nous avons trouvé que le phosphatidylinosito1-4-phosphate (PI4P) et le phosphatidylinositol-4,5-biphosphate (PI(4,5)P2) augmentent la sécrétion sous l'action de 10µM de Ca2+ dans les cellules INS-1E perméabilisées avec la streptolysine-O. En plus, nous avons démontré que l'exocytose est diminuée dans les cellules intactes exprimant une protéine qui séquestre le PI(4,5)P2. Une diminution similaire est observée en diminuant l'expression de deux enzymes impliquées dans la production du PI(4,5)P2, la PI4Kinase β type III et la PIP5Kinase γ type I. Pour clarifier le mécanisme d'action des PI, nous avons investigué l'implication de trois cibles potentielles des PI, la PLD1, CAPS1 et Mint1. Pour ce faire, nous avons réduit le niveau d'expression endogène de ces protéines, ce qui inhibe la libération d'hormones provoquée par le glucose. Tout ceci indique donc que la production du PI(4,5)P2 est nécessaire pour le contrôle de la sécrétion et suggère qu'une partie de l'effet du PI sur la sécrétion pourrait être exercé par l'activation de la PLD1, CAPS1 et Mint1. Abstract Insulin release from pancreatic β-cells plays an essential role in the achievement of blood glucose homeostasis and defects in the regulation of this process lead to profound metabolic disorders and hyperglycaemia (eg. type 2 diabetes). Almost every cell in our organism releases proteins and other biological compounds using a fundamental cellular process known as constitutive exocytosis. In exocrine and endocrine glands, the cells are endowed with an additional and more refined release mechanism directly tuned by extracellular signals. This process, referred to as regulated exocytosis, ensures the timely delivery of molecules such as peptide hormones and digestive enzymes to match the moment¬-to-moment requirements of the organism. Some of the molecular components involved in this process have been identified, including Rab3 and Rab27, two GTPases that regulate the final steps of secretion in many cells. We investigated the involvement of Rab27 GTPase in the secretory process of the insulin-secreting cell line INS-1E. We found that selective reduction of Rab27 expression by RNA interference did not alter granule distribution but impaired exocytosis triggered by insulin secretagogues. Screening for potential effectors revealed that Slac2c/MyRIP is associated with granules and attenuation of Slac2c expression severely impaired hormone release. This protein contains several functional domains, including, a binding domain for the cellular cytoskeleton constituent actin. Taken together our data suggest the Rab27 and MyRIP are part of a complex mediating the interaction of secretory granules with cortical cytoskeleton and participate to the regulation of the final steps in insulin exoctytosis. In the second part of the thesis, we studied phosphoinositides (PI). Phosphoinositides are important molecules involved in the regulation of vesicular trafficking. We found that phosphatidylinosito1-4-phosphate (PI4P) and phosphatidylinosito1-4,5-biphosphate (PI(4,5)P2) increase the secretory response triggered by 10µM Ca2+ in streptolysin-O permeabilized insulin-secreting INS-1E cells. In addition, nutrient-induced exocytosis was diminished in intact cells expressing constructs that sequester PI(4,5)P2. A similar decrease was observed after silencing of two enzymes involved in PI(4,5)P2 production, type III PI4Kinase β and type I PIP5Kinase γ, by RNA interference. To clarify the mechanism of action of PI, we investigated the involvement in the regulation of exocytosis of three potential PI targets, PLD1, CAPS1 and Mint1. Transfection of cells with silencers capable of reducing the endogenous levels of these proteins inhibited hormone release elicited by glucose. Our data indicate that the production PI(4,5)P2 is necessary for proper control of p-cell secretion and suggest that at least part of the effects of PI on insulin exocytosis could be exerted through the activation of PLD1, CAPS1 and Mint1.

HDR brachytherapy boost and external radiotherapy for unfavorable prostate cancer patients

Purpose/Objective: To evaluate the outcome of prostate cancer patients treated with a combination of HDR Brachytherapy boost (HDR-BT) and 3D conformal external pelvic radiotherapy (EBRT) in a dose escalation study. Materials and Methods: 162 patients were followed between November 2004 and December 2010 . Two different dose escalation groups were done: group 1 (n= 92), 1 fraction HDR boost (9-10 Gy ) followed by EBRT (60 Gy in 6 weeks) - BED: 203-216 Gy and group 2 (n=70): 2 fraction HDR boost (18-19 Gy), 6 hours interval between fractions, followed by EBRT (46 Gy in 4.5 weeks) - BED: 233.3 -247 Gy; 116 pts (71.6%) received concomitant androgen deprivation. Patients were classified according to the MSKCC criteria into high (N=137) and intermediate (N=25) risk. Phoenix biochemical failure definition was used. Toxicity was scored by Radiation Morbidity Scoring Criteria (RTOG) Results: The mean follow-up was 41 (range 7-84) months. The 7- years cancer-specific and overall survival was 100% an 92%, respectively. The 7 years actuarial biochemical control rate was 89% and 100% for group 1 and 2, respectively. One patient from group 1 and two patients from group 2 never reached a low nadir. Two patients developed distant metastases 12 and 16 months after the treatment. In a multivariate Cox-regression analysis neither treatment nor risk group (intermediate vs. high risk) were associated with increased risk for biochemical failure. The RTOG grade 3 genitourinary early toxicity was 1.0% and 8.5% while gastrointestinal/genitourinary late toxicity was 7.6% and 1.4% for group 1 and 2, respectively Conclusions: HDR BT boost followed by EBRT appears to be a safe, feasible and effective treatment for patients with unfavorable localized prostate cancer. This study shows a beneficial effect on biochemical control in group 2 pts, however without statistical significance. Higher radiation doses (BED 233.3-247 Gy) do not seem to carry extra toxicity.

Combination of androgen deprivation therapy and radiotherapy for localized prostate cancer in the contemporary era.

Currently, androgen deprivation therapy (ADT) has a well-defined role when administered together with radiotherapy (RT): neo-adjuvant and concurrent combination for intermediate risk-disease and adjuvant therapy for high risk disease. Evidence of this association was generated by randomized trials designed and led approximately 30 years ago; thus the question which arises is how relevant and portable are these data in our current clinical practice? In the present review, we examine the pitfalls of these published randomized controlled trials, their relevance to present daily clinics, where high-dose external beam RT or brachytherapy is applied, as well as the adoption of ADT in patients with concomitant cardiovascular disorders.

First imaging results of an intraindividual comparison of (11)C-acetate and (18)F-fluorocholine PET/CT in patients with prostate cancer at early biochemical first or second relapse after prostatectomy or radiotherapy.

PURPOSE: (18)F-Fluorocholine (FCH) and (11)C-acetate (ACE) PET are widely used for detection of recurrent prostate cancer (PC). We present the first results of a comparative, prospective PET/CT study of both tracers evaluated in the same patients presenting with recurrence and low PSA to compare the diagnostic information provided by the two tracers. METHODS: The study group comprised 23 patients studied for a rising PSA level after radical prostatectomy (RP, 7 patients, PSA ≤ 3 ng/ml), curative radiotherapy (RT, 7 patients, PSA ≤ 5 ng/ml) or RP and salvage RT (9 patients, PSA ≤ 5 ng/ml). Both FCH and ACE PET/CT scans were performed in a random sequence a median of 4 days (range 0 to 11 days) apart. FCH PET/CT was started at injection (307 ± 16 MBq) with a 10-min dynamic acquisition of the prostate bed, followed by a whole-body PET scan and late (45 min) imaging of the pelvis. ACE PET/CT was performed as a double whole-body PET scan starting 5 and 22 min after injection (994 ± 72 MBq), and a late view (45 min) of the prostate bed. PET/CT scans were blindly reviewed by two independent pairs of two experienced nuclear medicine physicians, discordant subgroup results being discussed to reach a consensus for positive, negative end equivocal results. RESULTS: PET results were concordant in 88 out of 92 local, regional and distant findings (Cohen's kappa 0.929). In particular, results were concordant in all patients concerning local status, bone metastases and distant findings. Lymph-node results were concordant in 19 patients and different in 4 patients. On a per-patient basis results were concordant in 22 of 23 patients (14 positive, 5 negative and 3 equivocal). In only one patient was ACE PET/CT positive for nodal metastases while FCH PET/CT was overall negative; interestingly, the ACE-positive and FCH-negative lymph nodes became positive in a second FCH PET/CT scan performed a few months later. CONCLUSION: Overall, ACE and FCH PET/CT showed excellent concordance, on both a per-lesion and a per-patient basis, suggesting that both tracers perform equally for recurrent prostate cancer staging.

Antigen binding to secretory immunoglobulin A results in decreased sensitivity to intestinal proteases and increased binding to cellular Fc receptors.

In intestinal secretions, secretory IgA (SIgA) plays an important sentinel and protective role in the recognition and clearance of enteric pathogens. In addition to serving as a first line of defense, SIgA and SIgA x antigen immune complexes are selectively transported across Peyer's patches to underlying dendritic cells in the mucosa-associated lymphoid tissue, contributing to immune surveillance and immunomodulation. To explain the unexpected transport of immune complexes in face of the large excess of free SIgA in secretions, we postulated that SIgA experiences structural modifications upon antigen binding. To address this issue, we associated specific polymeric IgA and SIgA with antigens of various sizes and complexity (protein toxin, virus, bacterium). Compared with free antibody, we found modified sensitivity of the three antigens assayed after exposure to proteases from intestinal washes. Antigen binding further impacted on the immunoreactivity toward polyclonal antisera specific for the heavy and light chains of the antibody, as a function of the antigen size. These conformational changes promoted binding of the SIgA-based immune complex compared with the free antibody to cellular receptors (Fc alphaRI and polymeric immunoglobulin receptor) expressed on the surface of premyelocytic and epithelial cell lines. These data reveal that antigen recognition by SIgA triggers structural changes that confer to the antibody enhanced receptor binding properties. This identifies immune complexes as particular structural entities integrating the presence of bound antigens and adds to the known function of immune exclusion and mucus anchoring by SIgA.

N-Glycans on secretory component: mediators of the interaction between secretory IgA and gram-positive commensals sustaining intestinal homeostasis.

Human beings live in symbiosis with billions of microorganisms colonizing mucosal surfaces. The understanding of the mechanisms underlying this fine-tuned intestinal balance has made significant processes during the last decades. We have recently demonstrated that the interaction of SIgA with Gram-positive bacteria is essentially based on Fab-independent, glycan-mediated recognition. Results obtained using mouse hybridoma- and colostrum-derived secretory IgA (SIgA) consistently show that N-glycans present on secretory component (SC) play a crucial role in the process. Natural coating may involve specific Gram-positive cell wall components, which may explain selective recognition at the molecular level. More widely, the existence of these complexes is involved in the modulation of intestinal epithelial cell (IEC) responses in vitro and the formation of intestinal biofilms. Thus, SIgA may act as one of the pillars in homeostatic maintenance of the microbiota in the gut, adding yet another facet to its multiple roles in the mucosal environment.

Automatic prostate segmentation in cone-beam computed tomography images using rigid registration.

We propose to evaluate automatic three-dimensional gray-value rigid registration (RR) methods for prostate localization on cone-beam computed tomography (CBCT) scans. In total, 103 CBCT scans of 9 prostate patients have been analyzed. Each one was registered to the planning CT scan using different methods: (a) global RR, (b) pelvis bone structure RR, (c) bone RR refined by local soft-tissue RR using the CT clinical target volume (CTV) expanded with a 1, 3, 5, 8, 10, 12, 15 or 20-mm margin. To evaluate results, a radiation oncologist was asked to manually delineate the CTV on the CBCT scans. The Dice coefficients between each automatic CBCT segmentation - derived from the transformation of the manual CT segmentation - and the manual CBCT segmentation were calculated. Global or bone CT/CBCT RR has been shown to yield insufficient results in average. Local RR with an 8-mm margin around the CTV after bone RR was found to be the best candidate for systematically significantly improving prostate localization.

In vitro combination of human and bovine free secretory component with IgA of various species.

The free form of the secretory component usually associated with secretory IgA can be isolated from human and bovine milk. These free secretory components of different origin combine in vitro with human polymeric myeloma IgA, with mouse myeloma IgA, and with the serum IgA of nine different mammalian species.

The reasons for discrepancies in target volume delineation: a SASRO study on head-and-neck and prostate cancer

But : comprendre les raisons des divergences observées dans le tracé des volumes cibles entre radio-oncologues. Matériel et méthodes : 18 centres suisses de radio-oncologie ont été invités à tracer les volumes pour un cas de cancer de la prostate et pour un cas de cancer de la sphère ORL. Nous avons également envoyé un questionnaire pour évaluer d'une part les différences dans la définition- des volumes (GTV [ gross tumor volume= volume tumoral macroscopique ], CTV [clinical tumor volume= volume de la tumeur clinique, macroscopique et microscopique], PTV [planning target volume= volume cible pour la planification], et d'autre part des variations dans les marges appliquées et les ganglions considérés à risque. Pour chaque centre, on a calculé la corrélation entre les marges qui ont été dessinées et celles qui ont été annoncées. Nous avons inclus dans le questionnaire une série de questions non spécifiques pour évaluer les méthodes de planification utilisées dans les différents centres. Résultats : Dans les 2 situations cliniques, on a mis en évidence de relativement grandes différences tant au niveau des volumes dessinés par les différents centres que dans leur définition des volumes. La corrélation entre les marges dessinées et définies était assez correcte dans le cas de la prostate mais médiocre dans le cas ORL. Le questionnaire a révélé d'importantes différences dans les méthodes de planification utilisées par les centres. Conclusion : Ces grandes différences peuvent être expliquées par, (1) une connaissance/interprétation variable des définitions ICRU, (2) des appréciations différentes de l'extension tumorale microscopique potentielle, (3) des difficultés dans l'identification du GTV, (4) des différences de concept, et (5) des différences entre la théorie (par ex. la description des marges) et la pratiqué (par ex. les marges dessinées).

Ultrastructural, morphometrical and immunocytochemical analyses of the exocrine pancreas in a hibernating dormouse.

Pancreatic acinar cells of euthermic, hibernating and arousing individuals of the hazel dormouse Muscardinus avellanarius (Gliridae) have been observed at the electron-microscopic level and analysed by means of ultrastructural morphometry and immunocytochemistry in order to investigate possible fine structural changes of cellular components during periods of strikingly different degrees of metabolic activity. During hibernation, the cisternae of the rough endoplasmic reticulum (RER) flatten assuming a parallel pattern, the Golgi apparatus is extremely reduced and the mitochondria contain many electron-dense particles. The cell nuclei appear irregularly shaped, with deep indentations containing small zymogen granules. They also contain abundant coiled bodies and unusual constituents, such as amorphous bodies and dense granular bodies. Large numbers of zymogen granules occur in all animals. However, the acinar lumina are open and filled with zymogen only in euthermic animals, whereas, in hibernating and arousing individuals, they appear to be closed. Morphometrical analyses indicate that, in pancreatic acinar cells, nuclei and zymogen granules significantly decrease in size from euthermia to hibernation, probably reflecting a drastic decrease of metabolic activities, mainly protein synthesis and processing. In all the studied animals, immunocytochemistry with specific antibodies has revealed an increasing gradient in alpha-amylase content along the RER-Golgi-zymogen granule pathway, reflecting the protein concentration along the secretory pathway. Moreover, during deep hibernation, significantly larger amounts of alpha-amylase accumulate in RER and zymogen granules in comparison to the other seasonal phases analysed. Upon arousal, all cytoplasmic and nuclear constituents restore their euthermic aspect and all morphometrical and immunocytochemical parameters exhibit the euthermic values, thereby indicating a rapid resumption of metabolic activities.

Pharmacotherapeutic management of locally advanced prostate cancer: current status.

Locally advanced prostate cancer (LAPC) is a heterogeneous entity usually embracing T3-4 and/or pelvic lymph-node-positive disease in the absence of established metastases. Outcomes for LAPC with single therapies have traditionally been poor, leading to the investigation of adjuvant therapies. Prostate cancer is a hormonally sensitive tumour, which usually responds to pharmacological manipulation of the androgen receptor or its testosterone-related ligands. As such, androgen deprivation therapy (ADT) has become an important adjuvant strategy for the treatment of LAPC, particularly for patients managed primarily with radiotherapy. Such results have generally not been replicated in surgical patients. With increased use of ADT has come improved awareness of the numerous toxicities associated with long-term use of these agents, as well as the development of strategies for minimizing ADT exposure and actively managing adverse effects. Several trials are exploring agents to enhance radiation cell sensitivity as well as the application of adjuvant docetaxel, an agent with proven efficacy in the metastatic, castrate-resistant setting. The recent work showing activity of cabazitaxel, sipuleucel-T and abiraterone for castrate-resistant disease in the post-docetaxel setting will see these agents investigated in conjunction with definitive surgery and radiotherapy.

Acute toxicity of curative radiotherapy for intermediate- and high-risk localised prostate cancer in the EORTC trial 22991.

INTRODUCTION: This trial randomly assessed short-term adjuvant hormonal therapy added to radiotherapy (RT) for intermediate- and high-risk (UICC 1997 cT2a or cT1b-c with high PSA or Gleason score) localised prostate cancer. We report acute toxicity (CTCAE v2) assessed weekly during RT in relation to radiation parameters. PATIENTS AND METHODS: Centres selected the RT dose (70, 74 or 78Gy) and RT technique. Statistical significance is at 0.05. RESULTS: Of 791 patients, 652 received 3D-CRT (70Gy: 195, 74Gy: 376, 78Gy: 81) and 139 received IMRT (74Gy: 28, 78Gy: 111). During RT, grade 3 gastrointestinal (GI) and genitourinary (GU) toxicities were reported by 7 (0.8%) and 50 (6.3%) patients, respectively. No grade 4 was reported. The risk of grade 2 GI toxicity increased significantly with increasing D50%-rectum (p=0.004) and that of grade 2 GU toxicity correlated only to Dmax-bladder (p=0.051). 3D-RT technique, increasing total dose and V95% >400 cc increased D50% and Dmax. One month after RT, only 14 patients (1.8%) reported grade 3 toxicity. AST did not seem to influence the risk of GU or GI acute toxicity. CONCLUSION: RT up to 78Gy was well tolerated. Dmax-bladder and D50%-rectum influenced the risk of grade 2 GU toxicity and GI toxicity, respectively. Both were lower with IMRT but remained high for an irradiated RT volume>400 cc for 3D-RT and for a dose of 78Gy. Hormonal treatment did not influence acute toxicity.

Can 3T multiparametric magnetic resonance imaging accurately detect prostate cancer extracapsular extension?

BACKGROUND: Accurate staging is essential to determine the correct management of patients diagnosed with prostate cancer. We assess the accuracy of 3T multiparametric magnetic resonance imaging (MRI) with endorectal coil (3TemMRI) in detecting prostate cancer local extension. METHODS: We retrospectively reviewed charts from January 2008 to July 2012 from all patients undergoing radical prostatectomy. Patients were only included if 3TemMRI and radical prostatectomy were performed at our institution. Based on the presence of extracapsular extension (ECE) at 3TemMRI, prostate cancer was dichotomized into locally advanced or organ-confined disease. The accuracy of 3TemMRI local staging was then evaluated using definitive pathology as a reference. RESULTS: Overall, 177 radical prostatectomies were performed within the timeframe. After applying exclusion criteria, 60 patients were included in the final analysis. The mean patient age was 67 ± 7 (standard deviation) years. Mean prostate-specific antigen value was 12.7 ± 12.7 ng/L. Based on preoperative characteristics, we considered 38 of the 60 patients (63%) patients high risk. 3TemMRI identified an organ-confined tumour in 46 patients and locally advanced disease in 14 patients. When correlated to final pathology, 3TemMRI specificity, sensitivity, negative and positive predictive values, and accuracy in detecting locally advanced prostate cancer were 90%, 35%, 57%, 79% and 62%, respectively. INTERPRETATION: This study shows that the use of preoperative 3TemMRI can be used to identify organ-confined prostate cancer when locally advanced disease is suspected.