Perineural spread of pelvic tumors mechanism and diagnosis Abstract Neoplastic lumbosacral plexopathies are infrequent affections of the lumbosacral plexus. Cases with minimal or non-specific finding on imaging can be particularly puzzling to diagnose. We describe a series of patients with perineural spread from the site of the primary tumor along the visceral autonomous nerves to the lumbosacral plexus and further proximally to the spinal nerves and even intradurally and also distally to the sciatic nerve. On series of 17 patients diagnosed with perineural spread of pelvic malignancy we describe characteristic clinical presentation and imaging finding. In many of these cases a tissue biopsy is necessary to finalize the diagnosis. We describe operative technique of targeted fascicular biopsy of the sciatic nerve and our experience with this procedure. On series of 117 patients, we report the outcome and complication: diagnostic yield was 84.8% and complication rate was 2.7 %. If a tissue sample is needed to conclude the diagnosis, targeted fascicular biopsy does increase the yield at an acceptable complication rate. Perineural spread of pelvic cancer is a new clinical-pathological entity with an unknown natural history or ideal treatment strategy. Based on the imaging finding in this group we present a...

Natural bioactive compounds have drawn the interest of many researchers worldwide in their effort to find novel treatments, including prostate cancer (PC) treatment which is estimated to be 13.1% of all new cancer cases in the U.S. in 2021. Many of these bioactive compounds have been identified from treatments in traditional Chinese medicine (TCM), that often have multiple bioactive compounds present. However, in vitro studies frequently focus on the compounds in isolation, or in simple combinations of two compounds. We used mixture design response surface methodology (MDRSM) to assess changes in PC cell viability after 48 hours of treatment to identify the optimal mixture of all 35 three-compound combinations of seven bioactive compounds from TCM. We used Berberine, Wogonin, Shikonin, Curcumin, Triptolide, Emodin, and Silybin to treat PC-3, DU145, and LNCaP human PC cells, and a drug-resistant PC-3 cell line. Berberine and Wogonin most frequently contributed to the optimal combination to reduce cell viability in PC-3 and LNCaP cells; DU145 cells more frequently responded best to a single compound.

Hintergrund: Der Phospholipase A2 Rezeptor 1 (PLA2R1) ist ein Typ 1 Transmembranrezeptor, welcher der Mannose-Rezeptor-Familie zugeordnet werden kann. Die Bedeutung von PLA2R1 für physiologische und pathologische Vorgänge ist noch weitestgehend unbekannt. Jedoch wird die Regulation wichtiger zellulärer Prozesse, wie Proliferation, Apoptose/ Seneszenz, Adhäsion, Migration/ Invasion und Inflammation im Zusammenhang mit dem Rezeptor diskutiert. Darüber hinaus ist eine Änderung der PLA2R1-Expression bei der Entstehung verschiedenster Krebserkrankung nachweisbar. Hierbei wird der Rezeptor einerseits mit einer pro-onkogenen und pro-migratorischen Wirkung in Verbindung gebracht. Andererseits ist ein tumorsuppressiver Effekt von PLA2R1 und eine Induktion der mitochondrialen Apoptose in Tumorzellen beschrieben. Zudem ist die Expression von PLA2R1 durch epigenetische Mechanismen kontrolliert und eine Promotor-Hypermethylierung ist assoziiert mit einer Repression der Rezeptor-Expression in der Prostatakarzinom (PCa)-Zelllinie LNCaP und der pädiatrischen, akuten lymphoblastischen Leukämie (ALL)-Zelllinie Jurkat. Vorangegangene Arbeiten zeigten eine Hypermethylierung innerhalb eines definierten Bereiches des PLA2R1-Promotors bei adulten Patienten mit akuter myeloischer Leukämie und Myelodysplastischem Syndrom (MDS) sowie eine Korrelation der PLA2R1-Promotormethylierung mit dem Krankheitsstadium und der Klassifizierung nach dem Internationalen Prognostischen Scoring System (IPSS). Fragestellung/ Hypothese: Das Ziel der vorliegenden Arbeit war einerseits die Untersuchung der Funktion von PLA2R1 in den PCa-Zelllinien LNCaP und PC-3. Während in LNCaP die Rezeptor-Expression durch Promotor-Hypermethylierung unterdrückt ist, kann in PC-3-Zellen eine Hochregulation von PLA2R1 im Vergleich zu normalen Prostataepithelzellen nachgewiesen werden. Durch in vitro Transfektionsexperimente sollte der Effekt einer Re-expression von PLA2R1 in LNCaP-Zellen sowie die Auswirkungen einer Reduktion der PLA2R1-Expression in PC-3-Zellen untersucht werden. Der Einfluss der veränderten PLA2R1-Expressionslevel auf wichtige Zellparameter wurde evaluiert. Die in vitro Daten der PCa-Zelllinien wurden mit den in vivo Ergebnissen des Tumorwachstums von transfizierten LNCaP- und PC-3-Zellen in Xenograft-Mausmodellen verglichen. Andererseits sollte basierend auf den Ergebnissen von adulten Patienten mit AML- und MDS-Diagnose und der dabei festgestellten Hypermethylierung des Rezeptor-Promotors der Methylierungsstatus des Rezeptors bei der pädiatrischen ALL untersucht werden. Überdies sollte die Eignung der PLA2R1-Methylierungsanalyse als sensitiver Biomarker für die Therapiekontrolle, Überwachung der minimalen Resterkrankung (MRD) und Risikostratifizierung der pädiatrischen ALL evaluiert werden. Die Funktion des Rezeptors im Kontext der pädiatrischen ALL wurde durch eine transfektionsbasierte Re-expression von PLA2R1 in der Jurkat-ALL-Zelllinie untersucht. Durch in vitro Experimente wurden die Auswirkungen der verschiedenen PLA2R1-Expressionslevel auf Proliferation und Apoptose/ Nekrose in transfizierten Jurkat-Zellen analysiert. Material und Methoden: Durch Transfektion mit einem PLA2R1-Expressionsvektor konnte eine stabile Überexpression des Rezeptors in LNCaP- (LNCaP-PLA2R1) und Jurkat-Zellen (Jurkat-PLA2R1) erreicht und die Ergebnisse mit Kontrollvektor-transfizierten LNCaP- (LNCaP-Ctrl) und Jurkat-Zellen (Jurkat-Ctrl) verglichen werden. Mittels CRISPR/Cas9-Knockdown konnte eine Verminderung der PLA2R1-Expression in PC-3-Zellen (PC-3-KD) im Vergleich zu Kontrollvektor-transfizierten PC-3-Zellen (PC-3-Ctrl) erreicht werden. Genexpressionsanalysen wurden mittels quantitativer PCR nach reverser Transkription (RT-qPCR) durchgeführt und die Proteinsynthese des Rezeptors durch Western Blot Analyse überprüft. In vitro sollten die Auswirkungen der differenziellen PLA2R1-Expression der transfizierten Zellen auf wichtige proliferative und metastatische Zellparameter untersucht werden. Die Zellviabilität/ Proliferation wurde mittels WST-1 Assay für adhärente Zellen und Zellwachstumskurven-Analyse mit Trypanblau-Färbung bei Suspensionszellen analysiert. Zellmotilität und Proliferation wurden bei transfizierten PCa-Zelllinien mithilfe des Wundheilungsassays beurteilt. Apoptose konnte durch Wasserstoffperoxid stimuliert und mittels Caspase-Glo® 3/7 Assay und RealTime-Glo™ Annexin V Apoptosis and Necrosis Assay für transfizierte PCa-Zelllinien sowie durchflusszytometrische Analysen nach Annexin-V-FLUOS/ Hoechst 33258 Färbung für transfizierte Jurkat-Zellen untersucht werden. Die klonogene Überlebensrate und das Koloniewachstum der transfizierten PCa-Zelllinien sollten mithilfe des klonogenen Assays analysiert werden. In einer in vivo Pilotstudie wurde der Effekt von PLA2R1 auf das Tumorwachstum mittels Xenograft-Mausmodellen (männliche SCID/beige Mäuse) durch subkutane Injektion der transfizierten LNCaP- (n = 5) und PC-3-Zellen (n = 9) überprüft. Die PLA2R1-Promotormethylierung als sensitiver Biomarker für die pädiatrische ALL wurde durch Isolation und Bisulfit-Behandlung der genomischen DNA von Knochenmark (KM)-Aspiraten und Leukozyten des peripheren Blutes (PB) von ALL-diagnostizierten Kindern (n = 44) sowie einer anschließenden Analyse mittels digitaler PCR (dPCR) evaluiert. Die Ergebnisse konnten mit dem Methylierungsstatus einer gesunden Kontrollgruppe (n = 20) verglichen werden. Ergebnisse und Schlussfolgerungen: In LNCaP-PLA2R1 und Jurkat-PLA2R1 konnte im Gegensatz zu den dazugehörigen Kontrollzellen eine stabile Überexpression des Rezeptors auf Ebene der Genexpression und Proteinsynthese detektiert werden. Bei PC-3-KD-Zellen war eine Reduktion der PLA2R1-Genexpression und eine Repression der Proteinsynthese unterhalb der Nachweisgrenze des Western Blot Assays zu verzeichnen, während PC-3-Ctrl-Zellen eine Genexpression und Proteinsynthese des Rezeptors zeigten. Die Zellviabilität/ Proliferation und Motilität war signifikant erhöht in LNCaP-PLA2R1 und PC-3-Ctrl im Vergleich zu LNCaP-Ctrl- und PC-3-KD-Zellen. Demgegenüber war eine Verminderung von Apoptose und Koloniewachstum in LNCaP-PLA2R1 und PC-3-Ctrl-Zellen nachweisbar. Durch Genexpressionsanalysen konnte eine Induktion der Expression von Fibronektin 1 (FN1), TWIST Homolog 1 (TWIST1) und Cyclin-abhängige Kinase 6 (CDK6) in LNCaP-PLA2R1-Zellen identifiziert werden. In vivo schien die PLA2R1-abhängige negative Regulation des Koloniewachstums die pro-onkogenen Eigenschaften des Rezeptors zu überwiegen. Dies resultierte in einem verminderten Tumorwachstum von LNCaP-PLA2R1 und einer tumorsuppressiven Rolle des Rezeptors in dieser PCa-Zelllinie. Im Gegensatz dazu zeigten PC-3-Ctrl-Zellen ein schnelleres Tumorwachstum im Xenograft-Mausmodell, was für einen pro-onkogenen Effekt der endogenen PLA2R1-Expression in PC-3-Zellen sprechen würde. Der differenzielle Einfluss von PLA2R1 auf die Regulierung des Tumorzellwachstums könnte im Zusammenhang mit der veränderten Expression von FN1, TWIST1 und CDK6 stehen, jedoch sind weiterführende Experimente nötig, um die Beteiligung dieser Gene in der PLA2R1-Signaltransduktion zu untersuchen. Die Analyse der Zellwachstumskurve der transfizierten Jurkat-Zellen zeigte eine Abnahme der Proliferationsrate und eine Zunahme des Anteils an toten Zellen bei Jurkat-PLA2R1 im Vergleich zu Jurkat-Ctrl-Zellen. Durchflusszytometrische Analysen bestätigten eine Abnahme des Anteils gesunder sowie eine vermehrte Repräsentation von apoptotischen und nekrotischen Jurkat-PLA2R1-Zellen im Vergleich zur Kontrolle, was einen tumorsuppressiven Einfluss des Rezeptors bei der pädiatrischen ALL suggeriert. Die Funktion von PLA2R1 als Tumorsuppressor steht im Einklang mit der festgestellten Hypermethylierung des Rezeptor-Promotors in KM-Aspiraten und PB-Proben von pädiatrischen Patienten mit prä-B und common ALL zum Zeitpunkt der Diagnose der primären Krebserkrankung und des ALL-Rezidives im Vergleich zu der Kontrollgruppe. Der parallele Abfall der PLA2R1-Promotormethylierung und der relativen Blastenzahl im Verlauf der ALL-Induktionstherapie sowie eine signifikante, positive Korrelation beider Größen in KM- und PB-Proben ließen auf die leukämischen Blasten als Quelle der Hypermethylierung des PLA2R1-Promotors schließen. Überdies wiesen Hochrisikopatienten der pädiatrischen ALL eine signifikant höhere PLA2R1-Promotormethylierung am Tag 15 der ALL-Induktionstherapie auf im Vergleich zu Patienten mit einem geringeren Risiko. Zusammenfassend deuteten die in vitro und in vivo Daten auf eine wichtige Funktion des Rezeptors bei der Regulation von Proliferation und Apoptose bei der pädiatrischen ALL hin. Die Analyse der PLA2R1-Promotormethylierung könnte als sensitiver Biomarker zu einer verbesserten ALL-Therapiekontrolle, MRD-Überwachung und Risikostratifizierung während der ALL-Induktionstherapie beitragen.:Inhaltsverzeichnis 1 Zusammenfassung 4 2 Abstract 8 3 Einführung in die Thematik 11 4 Publikation 1: “Diverse Effects of Phospholipase A2 Receptor Expression on LNCaP and PC-3 Prostate Cancer Cell Growth in vitro and in vivo” 24 5 Publikation 2: “Methylation of the Phospholipase A2 Receptor 1 Promoter Region in Childhood B Cell Acute Lymphoblastic Leukaemia” 25 6 Diskussion und Ausblick 26 7 Literaturverzeichnis 32 8 Danksagung 41 9 Anlagen 42 / Background: The phospholipase A2 receptor 1 (PLA2R1) is a type I transmembrane receptor and a member of the mannose receptor family. Physiological and pathophysiological functions of PLA2R1 are still not completely understood. However, PLA2R1 expression is discussed to have an impact on proliferation, apoptosis/ senescence, adhesion, migration/ invasion as well as inflammatory cell responses and divergent PLA2R1 expression is detectable in different types of cancer compared to corresponding normal tissues. In this context, receptor expression is linked to both a pro-oncogenic/ pro-migratory and a tumour-suppressive/ pro-apoptotic impact in different cancer cells. Moreover, PLA2R1 expression is controlled by epigenetic mechanisms and hypermethylation of the PLA2R1 promoter is associated with silenced expression of the receptor in the prostate carcinoma (PCa) cell line LNCaP and the paediatric, acute lymphocytic leukaemia (ALL) cell line Jurkat. Previous work revealed a defined hypermethylated region of the PLA2R1 promoter in adult patients with acute leukaemia and myelodysplastic syndrome (MDS). PLA2R1 promoter methylation correlated with disease stage and International Prognostic Scoring System (IPSS) classification. Aim: The aim of the present study was to evaluate the function of PLA2R1 in PCa cell lines LNCaP and PC-3. The receptor expression is silenced in LNCaP but upregulated in PC-3 cells compared to normal prostate epithelial cells. A pilot in vivo study addressed the effects of PLA2R1 in mice xenografted with transfected LNCaP and PC-3 cells. Based on previous findings of PLA2R1 promoter hypermethylation in adult ALL and MDS patients, the aim of the present study was to analyse the methylation status of the PLA2R1 promoter in paediatric ALL patients compared to healthy individuals. PLA2R1 methylation analysis was evaluated as sensitive biomarker for ALL treatment response, minimal residual disease (MRD) monitoring, and risk stratification. The impact of the receptor in childhood ALL was investigated by transfection-based re-expression of PLA2R1 in the paediatric ALL cell line Jurkat and the effect of different PLA2R1 expression levels on proliferation and apoptosis/ necrosis was analysed in in vitro experiments. Material and Methods: Stable PLA2R1 overexpression was achieved by transfection of LNCaP (LNCaP-PLA2R1) and Jurkat cells (Jurkat-PLA2R1) with a PLA2R1 plasmid vector. Results were compared to control vector transfected LNCaP (LNCaP-Ctrl) and Jurkat cells (Jurkat-Ctrl). Alternatively, PLA2R1 was knocked down using CRISPR/Cas9 in PC-3 cells (PC-3-KD) and compared to the corresponding control-transfected cells (PC-3-Ctrl). Gene expression analysis was conducted by quantitative reverse transcription PCR (RT-qPCR). PLA2R1 protein synthesis was analysed by western blot. The impact of the differential PLA2R1 expression on proliferative and metastatic parameters of transfected cancer cells was investigated in vitro. Cell viability/ proliferation was assessed by means of WST-1 Assay for adherent cells and via cell growth curve analysis after trypan blue staining for suspension cells. Cell motility and proliferation of transfected PCa cell lines were estimated by wound healing assay. Hydrogen peroxide-stimulated apoptosis was analysed by Caspase-Glo® 3/7 Assay and RealTime-Glo™ Annexin V Apoptosis and Necrosis Assay for transfected PCa cell lines and flow cytometric analysis after Annexin-V-FLUOS/ Hoechst 33258 staining for transfected Jurkat cells. Colony formation of transfected PCa cell lines was evaluated by clonogenic assay. A pilot in vivo study addressed the effects of PLA2R1 in mice xenografted with transfected LNCaP (n = 5) and PC-3 cells (n = 9). Evaluating PLA2R1 promoter methylation as sensitive biomarker for paediatric ALL, genomic DNA was isolated from bone marrow (BM) and peripheral blood (PB) of 44 paediatric ALL patients. After bisulfite treatment of isolated DNA samples, PLA2R1 methylation was analysed using digital PCR and compared to 20 healthy controls. Results and Conclusions: PLA2R1 gene expression and protein synthesis were detectable in LNCaP-PLA2R1, PC-3-Ctrl, and Jurkat-PLA2R1 cells but not in LNCaP-Ctrl and Jurkat-Ctrl cells. In PC-3-KD cells, PLA2R1 gene expression was significantly reduced compared to PC-3-Ctrl and PLA2R1 protein synthesis of PC-3-KD cells was below the limit of detection of western blot analysis. Cell viability/proliferation and motility were significantly increased in LNCaP-PLA2R1 and PC-3-Ctrl compared to LNCaP-Ctrl and PC-3-KD cells, respectively. However, levels of apoptosis and clonogenicity were reduced in LNCaP-PLA2R1 and PC-3-Ctrl cells. Gene expression analysis revealed an up-regulation of fibronectin 1 (FN1), TWIST homolog 1 (TWIST1), and cyclin-dependent kinase 6 (CDK6) in LNCaP-PLA2R1 compared to control cells. In LNCaP xenografts, PLA2R1-dependent regulation of clonogenicity appeared to outweigh the receptor’s pro-oncogenic properties, resulting in decreased tumour growth, supporting the tumour-suppressive role of PLA2R1. Alternatively, PC-3-Ctrl xenografts exhibited faster tumour growth compared to PC-3-KD cells, suggesting a pro-oncogenic effect of endogenous PLA2R1 expression. The differential growth-regulatory effects of PLA2R1 may be mediated by FN1, TWIST1, and CDK6 expression, although further investigation is required. Cell growth curve analyses of transfected Jurkat cells revealed a decreased proliferation and increased cell death of Jurkat-PLA2R1 compared to Jurkat-Ctrl cells. Flow cytometry confirmed the reduced fraction of healthy cells and an increase of the apoptotic and necrotic fractions in Jurkat-PLA2R1 cells compared to control cells, suggesting a tumour-suppressive effect of the receptor in paediatric ALL. PLA2R1’s tumour-suppressive function is in accordance with hypermethylation of the receptor promoter in BM aspirates and PB samples of paediatric patients diagnosed with pre-B and common ALL as well as in patients with disease relapse in comparison to healthy controls. PLA2R1 methylation decreased along with leukaemic blast cell reduction during ALL induction treatment and significant positive correlations between PLA2R1 methylation and leukaemic blast cell numbers of BM and PB samples were observable. Therefore, our data suggests that leukaemic blasts are the origin of PLA2R1 hypermethylation in BM and PB samples. Moreover, high risk paediatric ALL patients exhibited increased levels of PLA2R1 promoter methylation compared to non-high risk groups on day 15 of ALL induction treatment. Collected data indicates that PLA2R1 promoter methylation quantitation can be used as biomarker for ALL induction treatment control, risk stratification, and early detection of ALL relapse.:Inhaltsverzeichnis 1 Zusammenfassung 4 2 Abstract 8 3 Einführung in die Thematik 11 4 Publikation 1: “Diverse Effects of Phospholipase A2 Receptor Expression on LNCaP and PC-3 Prostate Cancer Cell Growth in vitro and in vivo” 24 5 Publikation 2: “Methylation of the Phospholipase A2 Receptor 1 Promoter Region in Childhood B Cell Acute Lymphoblastic Leukaemia” 25 6 Diskussion und Ausblick 26 7 Literaturverzeichnis 32 8 Danksagung 41 9 Anlagen 42

Le cancer de la prostate est le cancer le plus fréquemment diagnostiqué et la troisième cause de mortalité liée au cancer chez les hommes au Canada. Un quart des patients diagnostiqués développeront une forme plus agressive de ce cancer. Bien que nous possédions plusieurs indices cliniques pronostiques dans les cancers localisés (score de Gleason, taux sérique d’antigène prostatique spécifique (APS), stade, etc.), ceux-ci sont insuffisants pour adéquatement distinguer les patients à faible risque de progression de ceux à haut risque. A ce jour, aucun biomarqueur pronostique n’est encore utilisé en clinique. Les cliniciens ont donc besoin de nouveaux outils plus efficaces pour stratifier ce cancer et pour s’assurer d’adapter au mieux le traitement à chaque patient. En nous basant sur la littérature et sur des études préliminaires (cohortes de moins de 65 patients), notre hypothèse est que les protéines PUMA-NOXA et les récepteurs membranaires de la famille ERBB seraient, lorsqu’utilisés en combinaison, des biomarqueurs prédictifs de la progression du cancer de la prostate. Les objectifs de cette thèse sont : 1) identifier et valider de bons anticorps pour chaque biomarqueur d’intérêt, 2) définir les niveaux d’expression de chaque biomarqueur sur une cohorte de 285 patients, et 3) établir les corrélations entre les niveaux d’expression et les données cliniques des patients. Dans l’optique d’une utilisation en clinique, des anticorps de type monoclonal ont été choisis pour identifier les biomarqueurs d’intérêts. Ces anticorps ont été testés et validés pour leur spécificité par immunobuvardage de type western blot et par immunofluorescence. La localisation de la protéine d’intérêt a été validée sur des échantillons de tissus de patients suivie de l’optimisation du multi-marquage sur les cellules épithéliales et basales. Après perfectionnement de l’analyse d’images, nous avons montré qu’une expression extrême (faible ou forte) de PUMA couplée à une forte expression de NOXA dans les glandes bénignes est associée à la rechute biochimique des patients. La présence de ces biomarqueurs dans les glandes bénignes permet d’envisager d’améliorer l’identification lors des premières biopsies des patients se qualifiant pour la surveillance active. Par ailleurs, le suivi de l’expression des récepteurs de la famille ERBB dans les glandes tumorales permet une stratification des patients atteints d’un cancer de la prostate en fonction des risques de rechute biochimique, de développement de métastases et de mort liée au cancer. Ainsi, les patients présentant la combinaison d’une forte expression de EGFR et d’une faible expression de ERBB3 sont les plus susceptibles de mourir spécifiquement de leur cancer de la prostate, en particulier si les cellules tumorales présentes en plus une faible expression de ERBB2 entrainant un fort risque de développer des métastases. Mon projet de doctorat aura donc permis d’identifier et de valider des biomarqueurs d’intérêt pour prédire l’évolution du cancer de la prostate et démontrer l’intérêt de suivre ces biomarqueurs en combinaison afin d’obtenir une meilleure stratification des patients. Ces résultats devront être validés sur une cohorte indépendante et multicentrique en vue de fournir aux cliniciens un plus grand nombre d’outils pour leur permettre de réaliser une stratification fine des patients atteints d’un cancer de la prostate, et ouvrirait la voie à de nouvelles stratégies thérapeutiques plus ciblées. / Prostate cancer is the most frequently diagnosed cancer and the third leading cause of cancer-related death in men in Canada. A quarter of patients will develop a more aggressive form of this cancer. While there are several clinical prognostic variables for localized prostate cancer (Gleason score, prostate specific antigen (PSA) levels, stage, etc.), these are insufficient to adequately distinguish between low and high-risk of progression cases. As a result, clinicians need new, more effective tools to stratify this cancer and to ensure that treatments are best tailored to each patient. To date, no prognostic biomarker has yet been used clinically. Based on the literature and preliminary studies of small cohorts (less than 65 patients), we hypothesize that the protein expression of PUMA-NOXA and ERBB family members could help with the prediction of prostate cancer progression. The objectives of this thesis are: 1) to identify and validate antibodies for each biomarker of interest, 2) to define the expression levels of each biomarker on a 285 patient cohort, 3) to evaluate the correlation between marker expression levels and patient clinical data. For clinical use, monoclonal-type antibodies were chosen to identify the biomarkers of interest. These antibodies were validated for specificity by western blot and immunofluorescence techniques. The localization of the protein of interest was further identified within samples of patient tissues and additional optimization involving combinatorial staining for epithelial and basal cells. After refining the imaging and statistical analysis of PUMA and NOXA in benign glands, we found that extreme (weak or strong) PUMA expression coupled with high NOXA expression was associated with biochemical relapse. In addition, these proteins have significant potential for predicting disease evolution based on the initial radical prostatectomy sample. The presence of these proteins in benign glands would allow the identification of patients less suitable for active surveillance. Additionally, statistical analysis of ERBB family receptors in tumor glands, when used alone, allow stratification of prostate cancer patients for the prediction of biochemical relapse, development of metastases and also specific death from prostate cancer. Moreover, patients expressing a combination of high EGFR expression and low ERBB3 expression are at high risk of biochemical relapse and are at higher risk of prostate cancer specific mortality. In addition, coupling this high EGFR – low ERBB3 combination to a low ERBB2 expression helps classify patients at high risk of developing metastases. My doctoral research project will have made it possible to identify and validate biomarkers of interest for predicting the progression of prostate cancer and demonstrating the interest of combining these biomarkers in order to achieve better stratification of patients with prostate cancer. In the context of clinical utility, these results need to be validated on an independent and multicenter cohort in order to confirm these findings. This would eventually provide clinicians with a greater number of tools at their disposal to correctly anticipate patient trajectories and possibly identify new therapeutic targets for the control of the disease.

Le dépistage du cancer de la prostate est largement controversé de par l’équilibre de la balance bénéfices-risques de cette procédure. L’enjeu de santé publique est alors d’accompagner au mieux les hommes susceptibles de se faire dépister pour qu’ils puissent être informés et participer au choix. L’objectif de ce doctorat est de contribuer à comprendre pourquoi les hommes participent au dépistage du cancer de la prostate, pour pouvoir améliorer dans un second temps leur accompagnement. Nous avons tout d’abord interrogé des hommes sur leurs connaissances, leurs sources d’informations et leur rôle dans le choix de participer au dépistage du cancer de la prostate. Ces entretiens ont abouti à la nécessité de concevoir un nouvel outil d’information pour améliorer les connaissances des hommes, en particulier sur les risques de la procédure de dépistage. Enfin, nous avons estimé comment les bénéfices et les risques étaient intégrés dans le choix des individus selon si ces derniers avaient eu accès à notre outil d’information. Les résultats de ce travail montrent que les hommes semblent moins favorables à une proposition de dépistage après avoir eu accès à notre outil d’information. Cependant, cela ne modifie pas leur appréciation des bénéfices et des risques du dépistage. Il est nécessaire de poursuivre les efforts pour permettre aux hommes de faire un choix éclairé. / Prostate cancer screening is highly controversial because of the benefit risk ratio. An issue is to support men susceptible to perform this screening towards informed choice and shared decision-making. The aim of this doctorate is to understand why men adhere to prostate cancer screening, and in a second time to improve their support. We interviewed men about their knowledge, their information seeking behaviour, and their implication in prostate cancer screening decision. This highlighted the necessity to create a new information tool to improve men’s knowledge, in particular regarding risks of the screening procedure. Finally, we investigate benefit risk trade-off with and without our information tool access. Results of this work shows that men are less favourable to a screening option after taking into account our information tool. However, benefit and risk of this screening appreciation was not changed. We need to continue efforts to allow men to make an informed choice.

Orientador: Marco Antônio Rodrigues Fernandes / Resumo: O sucesso da radioterapia está intimamente ligado à razão terapêutica que representa o quociente entre a quantidade de tecido tumoral irradiado e o volume de tecido sadio atingido. A Probabilidade de Complicação em Tecidos Normais (NTCP) e a Probabilidade de Controle do Tumor (TCP) são parâmetros fornecidos por Sistemas de Planejamentos de Tratamentos (TPS) computadorizados, usados na rotina da radioterapia que auxiliam na interpretação da qualidade do tratamento. Neste trabalho são analisados os planejamentos de radioterapia de 03 pacientes portadores de câncer de próstata. Os planejamentos dos tratamentos foram realizados no TPS XiO, simulando as técnicas de radioterapia por intensidade modulada de feixe (IMRT) e radioterapia tridimensional conformada (3D-CRT). A dose de radiação preconizada para o volume de tratamento planejado (PTV) foi de 7.600 cGy, as simulações foram realizadas para um arranjo de 6 campos de radiação com feixes de raios X de megavoltagem e energia de 10 MV. Os volumes prostáticos variaram entre 107 cm3 e 143 cm3. A dose de cobertura D98% do PTV variou de 6.940 cGy a 7.570 cGy com IMRT e de 6.410 cGy a 7.250 cGy com 3D-CRT. Os valores obtidos para o TCP ficaram entre 73,5% a 81,1% com IMRT e entre 70,6% a 75,9% com 3D-CRT. Considerando os valores de NTCP para o reto e a bexiga, os maiores valores encontrados foram 6,9% para o reto e 6,1% para a bexiga, ambos planejados com a técnica de 3D-CRT. Para os casos analisados, os resultados mostram que a técnic... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The success of radiotherapy is closely related to the therapeutic ratio which represents the ratio of the amount of irradiated tumor tissue to the volume of healthy tissue achieved. Normal Tissue Complication Probability (NTCP) and Tumor Control Probability (TCP) are parameters provided by computerized treatment planning systems (TPS), used in radiotherapy routine and also allow the interpretation of treatment quality. The aim of this work is analyze the planning of 03 cases of patients submitted to prostate cancer radiotherapy. The treatment plans were performed in TPS XiO, simulating the techniques of beam intensity modulated radiotherapy (IMRT) and tree-dimensional conformal radiation therapy (3D-CRT). The recommended radiation dose for the planned treatment volume (PTV) was 7600 cGy, the simulations were performed for an arrangement of 6 radiation fields with megavoltage X-ray beams and 10 MV energy. Prostatic volumes ranged from 107cm3 to 143cm3 . The D98% PTV coverage dose ranged from 6,940 cGy to 7,570 cGy with IMRT and from 6,410 cGy to 7,250 cGy with 3D-CRT. The values obtained for TCP were between 73.5% to 81.1% with IMRT and between 70.6% to 75.9% with 3D-CRT. Considering the NTCP values for the rectum and bladder, the highest values found were 6.9% for the rectum and 6.1% for the bladder, both planned using the 3D-CRT technique. For the analyzed cases, the results show that the IMRT technique presents better NTCP and TCP values than the 3D-CRT technique. These par... (Complete abstract click electronic access below) / Mestre

Adjuvant hormonal therapy has been demonstrated to be able to delay disease progression in nonmetastatic prostate cancer. To date, however, a favorable impact on survival has only been demonstrated in lymph-node-positive disease and in external-beam radiotherapy series with locally advanced and probably mainly micrometastatic tumors. The Bicalutamide Early Prostate Cancer Program is the largest study under way to define the role of adjuvant treatment in early prostate cancer and identify subgroups of patients likely to benefit from immediate hormonal therapy. At the time of the most recently published analysis, the risk of objective clinical progression was significantly reduced in the bicalutamide arm (hazards ratio 0.58, 95% confidence interval 0.51–0.66, p < 0.0001). However, further maturation of data is needed to see whether this difference will lead to a survival advantage. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Objectives: To investigate the effects of serotonin and melatonin (MLT) on the regulation of malignant growth and the activity of serotonin receptors (5HTR1a/-1b) in prostate cancer (PCa) cell lines. Materials and Methods: In four PCa cell lines (LNCaP, 22RV1, PC3, DU145) and two reference cell lines 5HTR1a and -1b, relative mRNA expression levels were assessed. Different serotonin and MLT receptor agonists and antagonists were used in stimulation and inhibition experiments. Results: mRNA expression of 5HTR1b was higher than that of 5HTR1a in all PCa cell lines. Serotonin showed a significant growth stimulatory effect in all PCa lines. The 5HTR1a and -1b agonists/antagonists did not significantly affect viability. MLT inhibited viability only in PC3 cells. Similarly, the 5HTR1a antagonist induced apoptotic changes in PC3 cells only at 10–4M, while the 5HTR1b antagonist induced necrosis at 10–4M in all cell lines. Cell cycle alterations were seen in PC3 and DU145 cells under the influence of the 5HTR1a antagonist. Conclusions: Serotonin receptor antagonists and agonists as well as MLT influence viability and apoptosis of PCa cell lines at supraphysiologic concentrations. In contrast to other reports, our results do not support a regulatory role of serotonin or MLT receptor activation or inhibition in PCa growth. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

The treatment of clinically locally advanced prostate carcinoma (stage cT3) remains controversial. One of the main reasons for this controversy results from the substantial staging error attached to the clinical diagnosis cT3 with overstaged T2 tumors and understaged node-positive cases. Treatment options in this situation include radical prostatectomy, external beam radiotherapy, immediate or delayed androgen deprivation treatment and the so-called ‘watchful waiting’. Acceptable and often surprisingly good tumor-specific survival rates have been reported for radical prostatectomy in pT3 series – based on good clinical case selection – approaching those of pT2 series. In lymph node-positive pT3 cases, adjuvant hormone deprivation seems to prolong survival which it does not in lymph node-negative pT3 disease. A benefit of adjuvant external beam radiotherapy after radical prostatectomy for pT3 cases in prolonging overall survival has not been shown, despite the fact that it can prevent or delay biochemical and local recurrence. External beam radiotherapy as the only treatment for cT3 disease results in unfavorable tumor-specific survival rates, which can be significantly improved with adjuvant hormonal treatment with LHRH agonists. If, in case of advanced age and/or significant comorbidity, primary hormonal treatment is chosen, early hormonal deprivation therapy seems to offer marginal benefits in survival compared to delayed treatment. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Background: To compare the clinical value of the measurement of complex and total PSA in the discrimination between benign prostatic hyperplasia (BPH) and prostate cancer. Methods: In serum samples collected from 166 men with histopathologically proven clinically localized prostate cancer and of 97 men with BPH, total prostate-specific antigen (PSA), complexed PSA and the free to total PSA ratio were determined. The statistical analysis was done by the comparison of the receiver operator characteristic (ROC) curves. Results: The areas under the ROC curves were 0.776 for total PSA, 0.799 for complexed PSA (total PSA vs. cPSA: p < 0.0001) and 0.812 for the free to total PSA ratio. With a cut-off of 3.0 ng/ml for complexed PSA, the sensitivity was 90%, the specificity 58%, the positive and the negative predictive values 79 and 78%, respectively. With a cut-off of 4.0 ng/ml for total PSA, the sensitivity was 87%, the specificity 59%, the positive and the negative predictive values were 78 and 72%, respectively. Conclusions: There was a statistically significant advantage for complexed PSA compared to total PSA in the discrimination between BPH and prostate cancer. The difference was, however, small and its clinical relevance is questionable. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.