Die Entwicklung neuer Medikamente und Therapieverfahren wie die Hochdosischemotherapie und die Möglichkeit der Stammzelletransplantation haben die Heilungschance krebskranker Kinder in den letzen Jahrzehnten enorm verbessert. Aus diesem Grund erlangt die Berücksichtigung der Spätfolgen der Therapie eine größere Bedeutung. Es wurden die Zytostatika Cyclophosphamid und Thiotepa auf ihre Neurotoxizität im infantilen Rattengehirn untersucht. Dazu wurde Ratten im Alter von 7, 14, 21 oder 28 Tagen Cyclophosphamid (200-600mg/kg) oder Thiotepa (15-45 mg/kg) intraperitoneal injiziert. Nach einer Überlebenszeit von 4-24 Stunden wurden die Tiere getötet. Die Dichte degenerierter Zellen wurde lichtmikroskopisch in den nach De Olmos gefärbten Hirnschnitten mit Hilfe des stereologischen Dissektors ermittelt. Weiterhin wurden eine TUNEL-Färbung, elektronenmikroskopische sowie eine immunhistochemische Untersuchung für Caspase 3 und den Fas Rezeptor durchgeführt. Die Unterschiede zwischen den einzelnen Versuchsgruppen wurde mit Hilfe des Student`s t-Test auf ihre Signifikanz hin überprüft. Die Untersuchungen zur Zeit und Dosisabhängigkeit wurde mit Hilfe der ermittelten Gesamtscores und der Varianzanalyse (ANOVA) überprüft. Diese Untersuchung zeigte, dass eine Exposition mit den Zytostatika Cyclophosphamid und Thiotepa altersabhängig zu ausgeprägten Zellschädigungen im Gehirn führt. Besonders ausgeprägte Zelluntergänge fanden sich im Cortex, den thalamischen Kerngebieten, und dem Hippocampus. Ultrastrukturell ließen sich bereits kurz nach der Applikation des Zytostatikums anschwellende Dendriten als Hinweis auf einen exzitotoxischen Zelltodmechanismus nachweisen. Im Gegensatz dazu zeigten sich bei Tieren mit längerer Lebensdauer nach Exposition gegenüber dem Zytostatikum typische ultrastrukturelle Veränderungen wie man sie bei apoptotischem Zelltod finden kann. Mit dieser Untersuchung konnte gezeigt werden, dass die neurotoxische Wirkung der Zytostatika Cyclophosphamid und Thiotepa eine exzitotoxische und eine apoptotische Komponente aufweist. / Survival rates for children with cancer have increased dramatically over the past few decades. The expanded use of older agents, the development of new chemotherapeutic agents, the introduction of high dose chemotherapy and stem cell transplantation regimen have had a major impact on this improvement. These positive results have also focused increased attention on post-therapeutic effects of anticancer drugs. To investigate whether common cytotoxic drugs cause neurotoxic effects in the developing rat brain the following alkylated agents were administered to 7-day-old rats: cyclophosphamide (200–600mg/kg IP) and thiotepa (15– 45mg/kg IP). The brains were analysed at 4 to 24 hours. Quantitation of brain damage was performed in De Olmos cupric silver-stained sections using the stereological dissector method. Furthermore electron microscopy on plastic sections, TUNEL staining and immunohistochemistry for activated caspase 3 and Fas receptor was performed. Statistical analysis was performed by means of Student´s t test or one-way analysis of variance with subsequent pairwise comparison (Scheffé-test). Cytotoxic drugs produced widespread lesions within cortex, thalamus, hippocampal dentate gyrus, and caudate nucleus in a dose-dependent fashion. Early histological analysis demonstrated dendritic swelling and relative preservation of axonal terminals, which are morphological features indicating excitotoxicity. After longer survival periods, degenerating neurons displayed morphological features consistent with active cell death. These results demonstrate that anticancer drugs are potent neurotoxins in vivo; they activate excitotoxic mechanisms but also trigger active neuronal death.

Das Ziel der vorliegenden Arbeit war die Evaluierung der Möglichkeit der Bildung von Einschlußverbindungen eines neuen Vitamin D3 Analogon mit nativem [beta]-Cyclodextrin, Dimethyl-[beta]-Cyclodextrin, [beta]-Cyclodextrin-Polymer, Maltosyl-[beta]-Cyclodextrin, Hydroxypropyl-[beta]-Cyclodextrin und Carboxyethyl-[beta]-Cyclodextrin. Die resultierenden Addukte wurden im Hinblick auf Zusammensetzung und Eigenschaften charakterisiert. Weiterhin wurden in vitro und in vivo Untersuchungen durchgeführt, um den Einfluß von [beta]-Cyclodextrinen, vor allem im Hinblick auf Retardierungseffekte, nach Einarbeitung in Salben für die Indikation Psoriasis zu interpretieren. Die Herstellung der festen Kavitate erfolgte in Abhängigkeit vom Cyclodextrin-Typ mittels Knetmethode oder Kopräzipitation. Die Assoziatbildung mit [beta]-Cyclodextrinen in Lösung führte zu einer deutlichen Verbesserung der Löslichkeit des Vitamin D3 Analogon. Zur Ermittlung von Löslichkeitsisothermen wurden von den Systemen Phasenlöslichkeitsdiagramme aufgenommen sowie Komplexstabilitätskonstanten berechnet. Zur Charakterisierung der festen dienten DSC-Untersuchungen. Die Anwesenheit von [beta]-Cyclodextrinen führte zu einer Reduktion der Freisetzungsrate der aktiven Verbindung in vitro, besonders im Falle von nativem [beta]-Cyclodextrin. Auch ein deutlich reduzierter Effekt im Hinblick auf die Epidermishyperplasie von Nacktmäusen ist in vivo demonstrierbar. Die Ergebnisse werden im Rahmen der Fragestellung eines Retardierungseffektes sowie einer besseren Hautverträglichkeit, systemischen Nebenwirkungen und lokaler Effektivität des neuen Vitamin D3 Analogon diskutiert. / The aim of the present work was to study the possibility of forming cavitates of a new Vitamin D3 analogue in native [beta]-cyclodextrin, dimethyl-[beta]-cyclodextrin, [beta]-cyclodextrin-polymer, maltosyl-[beta]-cyclodextrin, hydroxypropyl-[beta]-cyclodextrin and carboxyethyl-[beta]-cyclodextrin and to characterize the resulting adducts as regards their composition and properties. Furthermore, in vitro and in vivo studies will be conducted to examine the influence of the [beta]-cyclodextrins in view to retardation effects after incorporation in ointments chosen for the treatment of psoriasis. The solid inclusions are produced with the kneading method or by co-precipitation, depending on the type of cyclodextrin. Phase-solubility diagrams will be drawn and complex stability constants calculated. The formation of associates with the cyclodextrins leads to a substantial improvement of the solubility of the drug. DSC studies are used to characterize the solid associates. The presence of [beta]-cyclodextrins leads to a reduction of the release rate of the active substance in vitro, especially in the case of native [beta]-cyclodextrin. A much reduced effect with regard to epidermal hyperplasia of nude mice is also demonstrable in vivo. The findings are discussed within the framework of the question of retardation and better tolerance as regards skin irritation, systemic side effects and local efficacy of the Vitamin D3 analogue.

Da die Arzneistoffpermeation durch okulare Gewebe einen entscheidenden Einfluss auf die Heilung vieler Augenleiden hat, wurde die in vitro-Permeation hydrophiler und lipophiler Arzneistoffe durch okulare Gewebe und Zellkulturen in dieser Arbeit untersucht. Die Dissertation befasst sich vorrangig mit der Permeation des hydrophilen Modellarzneistoffs Pilocarpinhydrochlorid (P-HCl) durch isolierte Schweinecornea (SC), Schweinesklera, Kaninchenkonjunktiva und corneale bzw. konjunktivale Kaninchenepithelzellkultur. Der Einfluss verschiedener Formulierungsparameter wie Benzalkoniumchlorid (BAC), Natriumedetat, pH-Wert und Tonizität auf die P-HCl-Permeation wurde untersucht. Dabei konnte eine gute Korrelation zwischen isolierten Geweben und Zellkulturen in Reaktion auf die Variation der Formulierungsparameter festgestellt werden. Unter den getesteten Parametern zeigte BAC den größten Enhancereffekt. Weiterhin wurden vergleichende Permeationsstudien an gelaserter SC mit P-HCl und dem relativ lipophilen Diclofenac-Natrium (D-Na) durchgeführt. Das Entfernen von Epithelschichten der SC mittels Excimer-Laser sollte den Heilungsverlauf, vor allem aber die zeitabhängige Reepithelisierung der Cornea nach erfolgter Photorefraktiver Keratektomie, simulieren. Die Ergebnisse dieser Studie zeigten, dass unterschiedliche Epitheldicken einen signifikanten Einfluss auf die P-HCl-Permeation haben. Im Gegensatz dazu blieb die D-Na-Permeation nahezu unbeeinflusst. Ein weiteres Anliegen dieser Arbeit bestand darin, eine okular applizierbare Formulierung des Immunsuppressivums Mycophenolatmofetil (MMF) zu entwickeln. Sowohl für das Prodrug MMF als auch für dessen aktiven Metaboliten Mycophenolsäure (MPA) wurde die Permeabilität von SC getestet. Ausgewählt wurde eine Zubereitung, die 1% MMF in Glutathion-Bicarbonat-Ringerlösung enthält und mit 10% Hydroxypropyl-beta-Cyclodextrin versetzt ist. Diese Suspension wurde bei 121° C und 200 kPa für 15 min autoklaviert, um das schwerlösliche MMF in Lösung zu bringen. Da der Ester MMF bei der Herstellung der Testlösung einer teilweisen Hydrolyse zu MPA unterliegt, außerdem eine minimale in vitro-Verfügbarkeit aufweist und sehr schwer wasserlöslich ist, sollte MPA in Augentropfenformulierungen der Vorzug gegeben werden. Die Korrelierbarkeit mit in vivo-Resultaten ist jedoch im Rahmen dieser Arbeit nicht untersucht worden, so dass die Ergebnisse als Grundlage für Permeationsstudien am in vivo-Modell zu bewerten sind. / The permeation of drugs through ocular tissues plays an important role in healing of various eye diseases. The objective of this work was to investigate the in vitro permeability of hydrophilic and lipophilic drugs through ocular tissues and cell cultures. Mainly, the permeability of the hydrophilic model drug pilocarpine hydrochloride (P-HCl) through isolated pig cornea (PCr) and sclera, rabbit conjunctiva, and rabbit conjunctival or corneal epithelial cell culture was compared. Additionally, the study included investigations about the influence of the formulation parameters benzalkonium chloride (BAC), ethylene diamine tetra acetic acid disodium salt, pH value and tonicity on the permeability of the small drug. In summary, a good correlation between the isolated tissues and cell cultures with regard to P-HCl transport could be observed. In general, BAC caused the most facilitated drug transport within the formulation parameters. Furthermore, the permeation of P-HCl and the lipophilic diclofenac-sodium (D-Na) through lasered PCr was studied. To investigate the effects of photorefractive keratectomy on drug permeation, excimer laser ablations with varying depths were performed on isolated pig eyes. As a result, P-HCl demonstrated a significant enhancement of permeation in relation to the ablation depth. In contrast, corneal epithelial thickness scarcely influenced the permeation rate of D-Na. Another aim of this work was to develop an appropriate application form for topical ocular use of the immunomodulating substance mycophenolate mofetil (MMF) and to investigate the in vitro permeability of PCr for the prodrug MMF and its parent substance mycophenolic acid (MPA). The test formulation consisted of Glutathion-bicarbonate-Ringer's solution, 10% hydroxypropyl- beta-cyclodextrin and 1% MMF. To reach a concentration of 1% of the poor soluble MMF a treatment under autoclaving conditions at 121° C and 200 kPa for 15 min was needed. MPA should be preferred in eye drops because of an higher in vitro availability compared to MMF, an hydrolysis of MMF to MPA in the cornea during permeation and the poor water solubility of MMF. The correlation with in vivo results was not studied in this work but the findings can be assumed as basis for investigations on in vivo models.

Die Umwandlung in wasserlösliche Verbindungen, die renal ausgeschieden werden können, ist ein grundlegendes Prinzip im Abbau von Fremdstoffen. Hierbei unterscheidet man Phase-I- und Phase-II-Reaktionen. Die Aktivität vieler Phase-I- und Phase-II-Enzyme ist genetisch beeinflusst und kann starke interindividuelle Unterschiede im Metabolismus von Fremdstoffen verursachen und dadurch das Krebsrisiko und das Risiko für Arzneimittelnebenwirkungen beeinflussen. Die Häufigkeitsverteilungen der Allele der Gene, die Phase-I- und Phase-II-Enzyme kodieren, zeigen eine große interethnische Varianz. Die Polymorphismen dieser Enzyme wurden bisher jedoch noch nicht in der größten slawischen Volksgruppe, der russischen, untersucht. An der vorliegenden Studie nahm eine Gruppe von 325 Personen russischer Abstammung teil - gesunde Probanden bzw. Patienten, die nicht an einer malignen Erkrankung litten. Die Polymorphismen von zwei Enzymen der Phase I, CYP1A1 und CYP2D6, und von einem Enzym der Phase II, NAT2, wurden mittels PCR-RFLP-Genotypisierung und Real-time-PCR-Verfahren komplett untersucht. Die Häufigkeit der CYP1A1 Allele mit hoher Aktivität, CYP1A1*2A und CYP1A1*2B, betrug 4,6% (3,1%-6,5%) bzw. 5,1% (3,5%-7,1%). Die Häufigkeiten der genetischen Varianten von CYP1A1 waren: m1 (3801T>C) - 9,8% (95% Vertrauensbereich, 7,7%-12,4%), m2 (2455A>G) - 5,0% (95% VB, 3,5%-7,1%), m4 (2453C>A) - 2,5% (1,4%-4,0%), m5 (-4335G>A) - 25,8% (22,5%-29,4%), m6 (-3219C>T) - 6,0% (4,3%-8,1%), und m7 (-3229G>A) - 2,9% (1,8%-4,5%). Die Mutation m3, die bisher nur bei Afrikaner gefunden wurde, konnten wir nicht nachweisen. 5,9% (3,5%-9,2%) aller Probanden waren CYP2D6 Langsam-Metabolisierer und 3,4% (1,7%-6,3%) wurden als Ultraschnell-Metabolisierer identifiziert (CYP2D6*1x1/*1). Bei der Genotypisierung von acht verschiedenen Punktmutationen im NAT2-Gen ergab sich für 59,7% (54,1%-65,1%) der Studienteilnehmer ein Genotyp, der mit einer Langsam-Acetylierer-Status einhergeht. 34,7% (29,6%-40,2%) der Probanden hatten ein und 5,6% (3,3%-8,6%) zwei für die Schnellacetylierung kodierende Allele. Die Allelverteilung der für die wichtigsten Enzyme im Arzneimittelstoffwechsel kodierenden Gene ist bei Russen ähnlich wie bei anderen Kaukasiern. Es kann deshalb erwartet werden, dass die genetisch-bedingten Unterschiede in der Wirksamkeit und im Auftreten von Arzneimittelnebenwirkungen in der russischen Bevölkerung vergleichbar sind mit denen in anderen europäischen Populationen. / The basic principle of drug and xenobiotic metabolism in the body is to make them more water soluble and thus more readily excreted in the urine. Genetic polymorphisms of phases I and II xenobiotic transformation reactions are known to contribute considerably to interindividual variations in the metabolism of numerous drugs and xenobiotics and to associate with altered risk of adverse drug reactions and some cancers. The frequency of functionally important mutations and alleles of genes coding for xenobiotic metabolizing enzymes shows a wide ethnic variation. However, little is known of the frequency distribution of the major allelic variants in the Russian population. In this study we investigated 325 individuals of Russian origin, who were healthy volunteers or patients without malignant diseases. Our study included the complete investigation of two enzymes of phase I, CYP1A1 and CYP2D6, and one phase II enzyme, NAT2, using PCR-RFLP genotyping and LightCycler method. The frequencies of the CYP1A1 high-activity alleles, CYP1A1*2A and CYP1A1*2B, were 4.6% (3.1%-6.5%) and 5.1% (3.5%-7.1%), respectively. The mutations m1 (3801T>C), m2 (2455A>G), m4 (2453C>A), m5 (-4335G>A), m6 (-3219C>T), and m7 (-3229G>A) of CYP1A1 occurred in 9.8% (95% confidence interval, 7.7%-12.4%), 5.0% (95% C. I., 3.5%-7.1%), 2.5% (1.4%-4.0%), 25.8% (22.5%-29.4%), 6.0% (4.3%-8.1%), and 2.9% (1.8%-4.5%) of alleles, respectively. We did not find the m3 mutation, which has only been detected in Africans up to now. 5.9% (3.5%-9.2%) of all subjects were CYP2D6 poor metabolizers, whereas 3.4% (1.7%-6.3%) were identified as ultra-rapid metabolizers (CYP2D6*1x1/*1). Genotyping eight different single nucleotide polymorphisms in the NAT2 gene provided a genotype associated with slow acetylation in 59.7% (54.1%-65.1%) of individuals, 34.7% (29.6%-40.2%) of participants carried at least one allele encoding rapid acetylation, and 5.6% (3.3%-8.6%) were homozygous for the rapid-acetylation allele (wild-type allele *4 or mutant allele *12A). The overview of allele distribution of the important drug and xenobiotic metabolizing enzymes among Russians shows that the allele frequency is similar to that of other Caucasians. Therefore it may be expected that drug side effects and efficacy problems due to an individual's genetic background are similar compared to those in other European populations.

Die vorliegende Arbeit hatte die Aufgabe, die humane Hepatomzellinie Hep G2 hinsichtlich ihrer Biotransformationskapazität zu charakterisieren, um Aussagen über ihre Eignung als in vitro-Testsystem treffen zu können. Dazu wurden die Aktivitäten und die Induzierbarkeit von unterschiedlichen Cytochrom P450 Isoenzymen (CYP) bestimmt und mit Aktivitäten von isolierten Hepatozyten der Wistarratte verglichen. Als Vertreter der Phase II-Reaktionen wurde die Konjugierung von p-Nitrophenol untersucht. Hep G2-Zellen enthielten detektierbare CYP 1 A1 und 2-Aktivitäten, was mit Hilfe des 7-Ethoxyresorufin- und des 7-Ethoxycoumarin-Assays festgestellt werden konnte. Die Enzymaktivitäten waren durch 3-Methylcholanthren und Phenobarbital induzierbar. Die Umsatzraten waren höher als in Monolayerkulturen von Rattenhepatozyten. Die Umsatzraten der Azoreduktion von 4-(N,N-Dimethylamino)azobenzen waren in Hep G2-Zellen ebenfalls höher als in Hepatozyten der Wistarratte. Hep G2-Zellen zeigten sich hinsichtlich der Demethylierung von Aminophenazon, katalysiert durch CYP 3A1 und 2, und der Konjugierung von p-Nitrophenol den Rattenhepatozyten unterlegen. Die Konjugierung war durch 3-Methylcholanthren und Phenobarbital induzierbar. Des weiteren wurde die Biotransformation von 3 potentiellen Arzneistoffen in Hep G2-Kulturen untersucht. Dabei handelte es sich um AWD 100-041(3-(2-Mercaptoethyl)chinazolin-2, 4(1H,3H)-dion), AR 12463 (5-Piperidino-7-[N-pentyl-N (ß-hydroxyethyl)]amino-s-triazolo(1,5a)-pyrimidin) und dem Lipoxygenaseinhibitor FLM 5011(2-Hydroxy-5-methyllaurophenon -oxim). In allen drei Fällen wurden zwar die gleichen Hauptmetaboliten wie in Rattenhepatozyten gebildet, die Umsatzraten waren aber wesentlich geringer. Um die Toxizität dieser drei Verbindungen und die von Solanum lycopersicon- Mazeraten zu untersuchen, wurde der Proteingehalt und der DNA-Gehalt mit Hilfe von Amidoschwarz bzw. bisBenzimid der Kulturen bestimmt. Membranschäden wurden durch den LDH Cytotoxicity Test von Boehringer Mannheim detektiert. Unter anderem konnte gezeigt werden, daß die Toxizität von FLM 5011 in Hep G2-Zellen auf die Induktion apoptotischer Prozesse zurückzuführen ist, welche durch die sinkende Konzentration von 5(S)-Hydroxyeikosatetraensäure in der Zelle ausgelöst wird. Insgesamt stellen Hep G2-Zellen ein brauchbares in vitro-Modell für Biotransformations- und Zytotoxizitätsuntersuchungen dar. / This investigations had the intention to characterise the capacity of biotransformation of the human hepatoblastoma- derived cell line Hep G2 and to draw conclusions about its suitability as in vitro-model. The enzyme activities and inducabilities of cytochrome P450 isoenzymes (CYP) as phase I reactions were measured and compared with the activity of monolayer primary cultures of rat hepatocytes. As a phase II-reaction the conjugation of p-nitrophenol was examined. Hep G2 contained detectable activities of CYP 1A1 and 2 measured by the 7-ethoxyresorufin assay and the 7-ethoxycoumarin assay and which were inducable by 3-methylcholanthrene and phenobarbitone. The turnover was higher than in rat hepatocytes. Also reductive activities, detected by the azoreduction of 4-(N,N-dimethylamino)- azobenzene, had a higher level than rat hepatocytes. Hep G2 cells were inferior compared to rat hepatocytes concerning the demethylation of aminophenazone catalysed by CYP 3A1 and 2 and the conjugation of p-nitrophenol. The latter was highly inducable by phenobarbitone. The biotransformation of the three active substances AWD 100-041 (3-(2-mercaptoethyl) chinazoline-2,4(1H,3H)-dione), AR 12463(5-piperidino-7-[N- pntyl-N (ß-hydroxyethyl)]amino-s-triazolo[1,5a)-pyrimidin) and the lipoxygenase-inhibitor FLM 5011(2-hydroxy-5- methyllaurophenone-oxim) in Hep G2 cell were also examined. In all cases the major metabolites were the same as in rat hepatocytes but the turnover was much lower than in rat hepatocytes. To study the toxicity of these three compounds and of Solanum lycopersicon mazerates the protein and the DNA content of the Hep G2 cultures were measured with amido black and bisbenzimid respectively. Membrane damages were detected by the LDH Cytotoxicity Test of Boehringer Mannheim. It could be proved that the toxicity of FLM 5011 is due to apoptotic activities aroused by the down regulation of 5-(S)hydroxyeicosatetraenoic acid. Hep G2 cells are a useful model for assessing the metabolism and toxicity of xenobiotics.

Diese Arbeit beschäftigt sich mit der prognostischen und prädiktiven Bedeutung der Serummessung des shed antigens des Onkoproteins HER-2/neu (s-HER-2/neu) für Mammakarzinompatientinnen in unterschiedlichen Erkrankungsstadien sowie unter verschiedenen Formen der Systemtherapie inklusive der Behandlung mit dem monoklonalen Antikörper Herceptin. Hierbei wurde der Stellenwert von s-HER-2/neu mit weiteren biochemischen Serummarkern (s-EGFR, s-uPA, CA 27.29) und etablierten Prognosefaktoren des Mammakarzinoms verglichen. Da HER-2/neu zu verschiedenen Zeitpunkten der Erkrankung mit verschiedenen Methoden an unterschiedlichen Materialien bestimmt werden kann, mußte schwerpunktmäßig auch die differentielle Wertigkeit von s-HER-2/neu im Vergleich zu Methoden der HER-2/neu-Bestimmung am Tumormaterial untersucht werden. Hierbei wurde insbesondere hinterfragt, ob die verschiedenen Methoden der HER-2/neu-Diagnostik stets zum gleichen Ergebnis führen, welche biologischen und technischen Ursachen möglichen Diskordanzen zugrunde liegen, und welche Konsequenzen diese Diskordanzen für die Therapiewahl (vor allem mit Herceptin beim metastasierten Mammakarzinom) mit sich bringen. Es wurden folgende Ergebnisse erzielt: - Grenzwerte: s-HER-2/neu: 50 ng/ml mit großer Wahrscheinlichkeit auch eine HER-2/neu-Überexpression am Tumorgewebe aufweisen. - Insgesamt 70% der Patientinnen mit HER-2/neu-negativen Tumoren hatten bei der späteren Fernmetastasierung einen erhöhten s-HER-2/neu-Spiegel. Diese Zunahme des Nachweises der HER-2/neu-Positivität zwischen Gewebenachweis am Primärtumor und dem Serumnachweis im Stadium IV ist nicht zufällig (p=0,001) und allenfalls teilweise durch den Einfluß der Tumormasse erklärbar. Klonale Veränderungen müssen in Erwägung gezogen werden. / This work focuses on the prognostic and predictive impact of the determination of the shed antigen of the oncoprotein HER-2/neu in serum (s-HER-2/neu) for breast cancer patients in different stages of the disease as well as under different forms of systemic therapy including treatment with the monocloncal antibody Herceptin. The biological meaning of s-HER-2/neu was compared to other serum markers (s-EGFR, s-uPA, CA 27.29) and established prognostic markers in breast cancer. As HER-2/neu can be measured at different times during the course of the disease using different methods and material, the differential meaning of s-HER-2/neu had to be investigated in relation to methods of HER-2/neu determination in tissue. Key questions were whether different methods of HER-2/neu testing lead to the same result, what biological and/or technical reason may lead to discordances, and if so, which are the consequences of these discordances for therapeutic decision making (before all Herceptin therapy in metastatic breast cancer). The results of the investigations are as follows: - Cut-offs of normal: s-HER-2/neu: 50 ng/ml are most probably HER-2/neu-positive in tissue. - Altogether, 70% of the patients with HER-2/neu negative tumors showed elevated s-HER-2/neu concentrations at later metastatic spread. This increase of the number of HER-2/neu positivity between tissue determination at the primary tumor and the serum results at stage IV disease cannot be explained by pure coincidence (p=0.001) and can only in part be explained by the influence of tumor burden . Clonal changes during the course of the disease must be considered.

A doença do enxerto contra o hospedeiro (DECH) é uma complicação que ocorre após o transplante alogênico de medula óssea. A doença é mediada por células T do enxerto que atacam as células e os tecidos do receptor; um processo inflamatório, que ataca vários órgãos e tecidos, sendo este processo responsável por 15% das causas de óbito após o acometimento da doença. As citocinas estão envolvidas na ativação das células T citotóxicas e células natural killer (NK), que causam as lesões nas glândulas salivares e exibem características de morte celular com infiltração de linfócitos conduzindo à apoptose. É conhecido que as unidades secretoras das glândulas salivares, após serem acometidas pela DECH, não recuperam o seu status funcional e a presença do infiltrado inflamatório pode ser um fator importante na fibrose das glândulas salivares e outros tecidos. Há evidências de uma redução significativa do fluxo salivar nos pacientes com DECH e de alterações importantes na proteômica salivar (IgA, IgG, lactoferrina) e polimorfismos que modifiquem a síntese de proteínas que devem ser consideradas para a evolução do paciente. Consequentemente os pacientes apresentam redução de fluxo salivar, com perda variável das unidades secretoras glandulares, comprometendo a qualidade de vida e os resultados do transplante. E para melhor compreensão da DECH, contribuindo para o diagnóstico precoce e na conduta destes pacientes, o presente estudo determinou o perfil proteômico dos pacientes com a doença do enxerto contra o hospedeiro crônica e comparou com o perfil de proteínas dos pacientes submetidos ao transplante de células tronco hematopoiéticas em cinco diferentes períodos, por meio da coleta de saliva, uma técnica não invasiva, e análise proteômica; além de comparar o perfil com os grupos controles (Líquen plano oral e indivíduos saudáveis). Foram incluídos neste estudo o total de 23 pacientes submetidos ao transplante de células-tronco hematopoiéticas, na Unidade de Transplante de Medula Óssea do Hospital das Clínicas da Universidade de São Paulo. As amostras foram submetidas a análises prévias em gel de poliacrilamida contendo dodecil sulfato de sódio (SDS) e gel bidimensional (2-DE) para o conhecimento do perfil proteômico destes pacientes. As análises foram feitas por espectrometria de massa e os dados obtidos foram confrontados com banco de dados humanos. As proteínas do grupo dos pacientes com DECHc foram comparadas com o grupo controle de indivíduos saudáveis, por meio do heatmap, e ressaltamos as proteínas de maior abundância. Portanto 16 proteínas mostraram uma maior expressão, Ig gamma-1 chain C region; Isoform H14 of Myeloperoxidase; Isoform 2 of 14-3-3 protein Sigma; Fibrinogen beta chain; Alpha- 1-acidic glycoprotein 1; Isoform gamma-A of Fibrinogen gamma chain; Hemopexin; Keratin type I CK 14; Alpha-2-Macroglobulin; Thioredoxin; Complement C3; Vitamin D; Keratin type I CK16; Lactoperoxidase; Cystatin B e Neutrophil elastase. No entanto, ao comparamos estas proteínas com os diversos períodos analisados e os grupos controles, ressaltamos a importância de três proteínas para a doença do enxerto contra o hospedeiro crônica: Isoform 2 of 14-3-3 protein sigma, Hemopexin e CK14 / Graft versus host disease (GVHD) is a complication that occurred after allogeneic bone marrow transplantation. The disease is mediated by graft T cells that attack as cells and tissues of the recipient; An inflammatory process, which attacks various organs and tissues, being responsible for 15% of the causes of death after the disease. Cytokines are involved in the activation of cytotoxic T cells and natural killer (NK) cells, which cause lesions in salivary glands and exhibit cell death characteristics with lymphocyte infiltration leading to apoptosis. It is known as the secretory units of the salivary glands, after being affected by GVHD, do not recover their functional state and the presence of the inflammatory infiltrate can be an important factor in the fibrosis of the salivary glands and other tissues. There is evidence of a significant reduction in salivary flow in patients with GVHD and significant changes in salivary protein (IgA, IgG, lactoferrin) and polymorphisms that modify a protein symptom that are considered for patient evolution. Consequently, patients with salary reduction, with variable loss of glandular secretory units, compromise quality of life and transplant results. In order to better understand GVHD, contributing to the early diagnosis and in the management of these patients, the present study determined the proteomic profile of patients with graft versus chronic host disease and compared them with the protein profile of patients submitted to cell transplantation Hematopoietic stem cells in five different periods, through the collection of saliva, a noninvasive technique and proteomic analysis; In addition to a profile with the control groups (Lichen planus oral and healthy individuals). Included in this study were the 23 patients submitted to hematopoietic stem cell transplantation in the Bone Marrow Transplantation Unit of the Hospital das Clínicas of the University of São Paulo. As samples were submitted to previous analyzes on polyacrylamide gel containing sodium dodecyl sulfate (SDS) and 2-dimensional gel (2-DE) to know the proteomic profile of these patients. The analyzes were done by mass spectrometry and the data obtained were compared with human data. The proteins of the group of patients with GVHDc were compared to the control group of healthy individuals by means of the Heatmap, and we emphasized as proteins of greater abundance. Therefore, 16 proteins show a greater expression, Ig gamma-1 chain C region; Isoform H14 of Myeloperoxidase; Isoform 2 of 14-3-3 protein Sigma; Fibrinogen beta chain; Alpha-1-acidic glycoprotein 1; Isoform gamma-A of Fibrinogen gamma chain; Hemopexin; Keratin type I CK 14; Alpha-2-Macroglobulin; Thioredoxin; Complement C3; Vitamin D; Keratin type I CK16; Lactoperoxidase; Cystatin B e Neutrophil elastase. Isoform 2 of 14-3-3 protein sigma, Hemopexin and CK14. Isoform 2 of 14-3-3 protein sigma, Hemopexin and CK14. However, when comparing these proteins with the various types of analyzes and the control groups, we emphasize the relevance of three proteins for the graft versus host disease: Isoform 2 of 14-3-3 protein sigma, Hemopexin and CK14

Europe is less dynamic than other areas in the world. The European Union will lose its position as the largest economic region, with the share in world output decreasing to less than 15% in 2050. This article designs a strategy based on which Europe can increase its internal dynamics, connect with its dynamic neighborhood and stabilize its eastern and southern regions. These regions are currently searching for development paths different from those in the US and China. The envisaged strategy learns from past errors, counteracts "my country first calls", balances uneven trade and investment pacts, and prevents land and resource grabbing. A new partnership strategy with neighbors in Eurasia, the Middle East and Africa will enable Europe to overcome its growth fatigue. It could help to reshape globalization and convert the current "ring of fire" into a "ring of friends". Then, together with its neighbors, Europe could still be an important economic region in 2050, on par with China and larger than the US plus its neighbors.

Made available in DSpace on 2016-04-04T18:27:57Z (GMT). No. of bitstreams: 1 Francisco Ronald Capoulade Nogueira.pdf: 739486 bytes, checksum: a94c39cca51acd5d59aab56b20beda3d (MD5) Previous issue date: 2011-02-21 / The present dissertation discusses an investigation in the area of the Social Psychology of Work. Specifically, the author studied how five different psychoanalysts of Lacanian orientation listen to and deal with patients who bring with them some type of complaint related to the world of work. First, a theoretical investigation was carried out to determine into what field this dissertation should be classified, concentrating especially on a discussion related to the ontological aspects of work, its psychic consequences in the context of capitalism, possible contributions from the field of mental health as related to work, and theoretical elements from psychoanalysis. In the second phase of the research, five psychoanalysts were interviewed individually, their statements being classified as narratives. The content was established through semi-structured interviews that called for the narration of experiences in treating patients affected by some type of mental suffering related to work. In dealing with their patients the respondents tended to give greater weight to aspects related to family dynamics and histories than to aspects related to the world of work. This fact brings once again to the fore the old debate over the differences between individuals and collectivity in society. / A presente pesquisa visou investigar, a partir de uma perspectiva da Psicologia Social do Trabalho, como alguns psicanalistas de orienta??o lacaniana escutam e lidam com pacientes que t?m algum tipo de queixa relacionada ao mundo do trabalho. Primeiramente, foi realizada uma investiga??o te?rica que teve por intento situar em que campo se daria esta disserta??o, priorizando uma discuss?o dos aspectos ontol?gicos do trabalho, suas consequ?ncias ps?quicas dentro de um contexto capitalista, as contribui??es do campo da Sa?de Mental relacionada ao Trabalho e alguns elementos te?ricos da Psican?lise. No segundo momento da pesquisa, foram entrevistados cinco psicanalistas e suas falas classificadas como narrativas. Tal conte?do foi obtido por meio de entrevistas semiestruturadas, orientadas para a narra??o de experi?ncias no tratamento com pacientes acometidos por algum tipo de sofrimento mental relacionado ao trabalho. Observouse que os entrevistados privilegiaram, nos tratamentos que empreenderam, mais os aspectos das din?micas da hist?ria familiar do que as din?micas do mundo do trabalho, reavivando um antigo debate entre o individual/coletivo.

Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro / A proposta deste trabalho é realizar uma análise da figura feminina nos romances Jangada de pedra, História do cerco de Lisboa e Ensaio sobre a cegueira, de José Saramago. Iniciando por uma breve pesquisa da história da mulher como forma de traçar a longa trajetória por que passou até a sua emancipação, o estudo revela, através da análise de diferentes identidades femininas, a abertura de uma nova concepção de mulher na literatura portuguesa.O escritor propõe em sua obra uma reflexão sobre a situação do mundo através do indivíduo e suas atitudes, responsabilidades e anseios. Os ensinamentos que seus romances costumam transmitir aos leitores, na maioria das vezes, partem de personagens femininas. A mulher se completa realizando o papel de mãe, protetora e companheira e, ao mesmo tempo, de amante e guerreira. Considerada pelo presente trabalho uma heroína contemporânea, ela engloba estas duas vertentes que a sociedade por muito tempo insistiu em separar, cuidando da casa e da família ao mesmo tempo em que trabalha e busca seu sustento fora de casa Também é a mulher que, nesta obra, tem o poder da sensibilidade e da compaixão ajudando àqueles que precisam, mesmo quando mal os conhece. E também é ela quem consegue, encantadoramente, metamorfosear-se de menina a mulher, intercalando as imagens de santa e prostituta, sedutora externamente e pura em sua essência.Entretanto, a escrita saramagueana não trata apenas de inibir a matriz patriarcal, ignorando com isso o papel do homem. O que há é o resgate das culturas primitivas matriarcais, com a mulher e o homem governando em igualdade e harmonia. Por este motivo, o homem também é muito importante em seus romances já que, em união com a mulher, incentivado e guiado por ela, partem juntos, através do amor, para uma travessia (humana) existencial, na busca do conhecimento / The proposal of this work is to carry out an analysis of the feminine figure in the romances novels Jangada de Pedra, História do cerco de Lisboa and Ensaio sobre a Cegueira of the writer José Saramago. Beginning with a short research of the woman history as a way to trace her trajectory along the time for her emancipation, and with the analysis of the different feminine identities, the study reveals a new concept of the woman in Portuguese literature.The writer proposes in his work a reflection of the world situation through the individuals and their attitudes, responsibilities and desires. The lessons of his novels to the readers, most of the time have start from feminine figures. The woman completes herself playing a mothers role, protector and partner, and at the same time lover and warrior. Considered at the present work as an actual heroine, she takes the mission of the two distinct activities that the society insisted in making them apart, taking care of home and family while she works to provide their needs.In this work the woman also has the power of sensibility and compassion to help those in needs even when she barely knows them. She is also the one who can, with charm, become herself from girl to woman, fitting in as saint and prostitute, externally pure and seducer in her essence.However, Saramagos writings are not only about the matrixs patriarchal inhibition, ignoring the mans role. There is the rescue of matriarchal primitive cultures, with woman and man governing in equality and peacefully. For this reason, the man is also very important in his novels, since together with the woman, motivated and guided by her, they go trough love for an existential cross over for the search of knowledge