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Trotz großer Fortschritte in der Forschung und Therapie sind Krebserkrankungen die zweithäufigste Todesursache. Die Diagnose Bauchspeicheldrüsenkrebs ist, vor allem wenn das Karzinom spät erkannt wird, immer noch zumeist ein Todesurteil. Neue Hoffnungen der schwer zu behandelnden Krebserkrankung bringt aber die Immuntherapie und die Kombination dieser mit Strahlentherapie. Um Immuntherapie erfolgreich anwenden zu können, müssen der menschliche Körper, die Tumore und vor allem deren Reaktion auf unterschiedliche Behandlungen vollständig verstanden und erforscht werden. Da Pankreaskarzinome schwer zu heilen sind, ist es wichtig alle Parameter der Erkrankung vorweg zu analysieren, die Auswirkungen der komplizierten Behandlungen einzuschätzen, um anschließend die ideale zielorientierte Behandlung zu finden. Kompliziert deshalb, da es sich bei Bauchspeicheldrüsenkrebs um immunogene Tumore handelt, welche hochregulierte Immunnetze enthalten, einschließlich der an der erworbenen Immunsuppression beteiligte Signalwege. In der vorliegenden Arbeit wurden zwei Zelllinien des Pankreaskarzinoms untersucht, PANC-1 und MiA PaCa-2. Diese wurden nach Kultivierung mit Protonen- und Photonenstrahlung behandelt, was jedoch nur einen mäßigen Effekt auf die Immunogenität aufgezeigt hat. Bei den beiden ELISA´s von GAL-1 und HMGB1 konnte jedoch festgestellt werden, dass es einen Anstieg mit steigender Dosis gibt. Beide Analysen erreichten ihren Höchststand nach der fraktionierten Bestrahlung mit Röntgenstrahlung. Es gab keinen großen Unterschied zwischen den Ergebnissen der beiden Zelllinien. Western Blots wurden mit verschiedenen Proteinen durchgeführt, um einen Überblick der Reaktion der Zellen zu erhalten. Diese Ergebnisse müssen für weiterführende Aussagen mit weiteren Durchläufen wiederholt werden. Weiters müssen noch zahlreiche Details zu den molekularen Ereignissen geklärt werden. Da Tumore nicht nur aus Krebszellen bestehen ist es wichtig, die Experimente auch mit einer Mischung aus gesunden Zellen, zum Beispiel Bindegewebszellen, fortzuführen. VIIDas Ziel Bauchspeicheldrüsenkrebs im fortgeschrittenen Stadium zu heilen, erfordert den Mechanismus dahinter zu verstehen und kontrollieren zu können. Diese Form der Krebserkrankung am Ende besiegen zu können bedarf noch langer Forschungsarbeit, die jedoch auch einem vielversprechenden Weg scheint. Despite major advances in research and therapy, cancer is the second leading cause of death. The diagnosis of pancreatic cancer is still mostly a death sentence, especially if the carcinoma is detected late. However, new hope for this difficult-to-treat cancer is being brought by immunotherapy and the combination of this with radiotherapy. In order to successfully apply immunotherapy, the human body, tumors and, most importantly, their response to different treatments must be fully understood and researched. Since pancreatic cancers are difficult to cure, it is important to analyze all the parameters of the disease in advance, assess the effects of complicated treatments, and then find the ideal target-oriented treatment. Complicated because pancreatic cancers are immunogenic tumors, which contain highly regulated immune networks, including signaling pathways involved in acquired immunosuppression. In the present work, two pancreatic cancer cell lines were studied, PANC-1 and MiA PaCa-2, which were treated with proton and photon radiation after cultivation, but this revealed only a moderate effect on immunogenicity. However, both ELISA's of GAL-1 and HMGB1 were found to show an increase with increasing dose. Both analyses peaked after fractionated irradiation with X-rays. There was not much difference between the results of the two cell lines. Western blots were performed with different proteins to get an overview of the response of the cells. These results need to be repeated with additional runs for further conclusions. Furthermore, numerous details of the molecular events still need to be clarified. Since tumors do not only consist of cancer cells it is important to continue the experiments with a mixture of healthy cells, for example connective tissue cells. VIIIThe goal of curing advanced pancreatic cancer requires understanding and controlling the mechanism behind it. To be able to defeat this form of cancer in the end still requires long research work, which, however, also seems to be on a promising path.Despite major advances in research and therapy, cancer is the second leading cause of death. The diagnosis of pancreatic cancer is still mostly a death sentence, especially if the carcinoma is detected late. However, new hope for this difficult-to-treat cancer is being brought by immunotherapy and the combination of this with radiotherapy. In order to successfully apply immunotherapy, the human body, tumors and, most importantly, their response to different treatments must be fully understood and researched. Since pancreatic cancers are difficult to cure, it is important to analyze all the parameters of the disease in advance, assess the effects of complicated treatments, and then find the ideal target-oriented treatment. Complicated because pancreatic cancers are immunogenic tumors, which contain highly regulated immune networks, including signaling pathways involved in acquired immunosuppression. In the present work, two pancreatic cancer cell lines were studied, PANC-1 and MiA PaCa-2, which were treated with proton and photon radiation after cultivation, but this revealed only a moderate effect on immunogenicity. However, both ELISA's of GAL-1 and HMGB1 were found to show an increase with increasing dose. Both analyses peaked after fractionated irradiation with X-rays. There was not much difference between the results of the two cell lines. Western blots were performed with different proteins to get an overview of the response of the cells. These results need to be repeated with additional runs for further conclusions. Furthermore, numerous details of the molecular events still need to be clarified. Since tumors do not only consist of cancer cells it is important to continue the experiments with a mixture of healthy cells, for example connective tissue cells. VIIIThe goal of curing advanced pancreatic cancer requires understanding and controlling the mechanism behind it. To be able to defeat this form of cancer in the end still requires long research work, which, however, also seems to be on a promising path. Verfasserin: Lisa Komornik Abweichender Titel laut Übersetzung der Verfasserin/des Verfassers Masterarbeit FH JOANNEUM 2021

submitted by Florian Rathje Literaturverzeichnis: Blatt 45-52 Abstract in deutscher und englischer Sprache Paris-Lodron-University Salzburg, Masterarbeit, 2019 (VLID)5061259

Adenosine is a signaling molecule that exerts dual effects on tumor growth: while it inhibits immune cell function and thereby prevents surveillance by the immune system, it influences tumorigenesis directly via activation of adenosine receptors on tumor cells at the same time. However, the adenosine���mediated mechanisms affecting oncogenic processes particularly in head and neck squamous cell carcinomas (HNSCC) are not fully understood. Here, we investigated the role of adenosine receptor activity on HNSCC���derived cell lines. Targeting the adenosine receptor A2B (ADORA2B) on these cells with the inverse agonist/antagonist PSB���603 leads to inhibition of cell proliferation, transmigration as well as VEGFA secretion in vitro. At the molecular level, these effects were associated with cell cycle arrest as well as the induction of the apoptotic pathway. In addition, shRNA���mediated downmodulation of ADORA2B expression caused decreased proliferation. Moreover, in in vivo xenograft experiments, chemical and genetic abrogation of ADORA2B activity impaired tumor growth associated with decreased tumor vascularization. Together, our findings characterize ADORA2B as a crucial player in the maintenance of HNSCC and, therefore, as a potential therapeutic target for HNSCC treatment., What's new? The adenosinergic system plays an important role in the development of many types of cancer. However, the underlying mechanisms remain poorly understood. This study set to identify the oncogenic function of the adenosine receptor 2B (ADORA2B) in head and neck squamous cell carcinoma (HNSCC)���derived tumor cells. The results show that ADORA2B is upregulated and constitutively active in HNSCC���derived cell lines. This activity is sufficient to promote autonomous cell growth, migration, and angiogenesis in vitro and in vivo. The data suggest ADORA2B as an important biomarker and potential therapeutic target for the treatment of HNSCC and other ADORA2B���expressing solid tumors., publishedVersion

The long-term fate of biomedical nanoparticles after endocytosis is often only sparsely addressed in vitro and in vivo, while this is a crucial parameter to conclude on their utility. In this study, dual-fluorescent polyisobutylene-polyethylene glycol (PiB-PEG) polymersomes were studied for several days in vitro and in vivo. In order to optically track the vesicles' integrity, one fluorescent probe was located in the membrane and the other in the aqueous interior compartment. These non-toxic nanovesicles were quickly endocytosed in living A549 lung carcinoma cells but unusually slowly transported to perinuclear lysosomal compartments, where they remained intact and luminescent for at least 90 h without being exocytosed. Fluorescence-assisted flow cytometry indicated that after endocytosis, the nanovesicles were eventually degraded within 7-11 days. In zebrafish embryos, the polymersomes caused no lethality and were quickly taken up by the endothelial cells, where they remained fully intact for as long as 96 h post-injection. This work represents a novel case-study of the remarkable potential of PiB-PEG polymersomes as an in vivo bio-imaging and slow drug delivery platform.

eingereicht von Theresa Haslauer, BSc Literaturverzeichnis: Blatt 94-107 Zusammenfassung/Abstract in deutscher und englischer Sprache Paris-Lodron-Universität Salzburg, Masterarbeit, 2018 (VLID)5003171

Mit weltweit mehr als 1 Millionen Neuerkrankungen pro Jahr ist das kolorektale Karzinom eine der häufigsten Krebserkrankung des Menschen und somit ein wichtiges Krankheitsbild für medizinische Studien. Um die Früherkennung dieser Krebserkrankungen und damit die Überlebensrate der Patienten zu erhöhen, wurden im Rahmen dieser Arbeit mittels Raman Mikrospektroskopie die Veränderungen des Proteoms in Zellen und in Gewebe untersucht. Am Beispiel des kolorektalen Karzinoms wurde die Raman basierte Spektrale Histopathologie (SHP) gewählt, mittels welcher man zuvor definierte Gewebetypen aufgrund ihrer spektralen Ähnlichkeiten automatisiert zuordnen und somit den Mediziner bei der Diagnose unterstützen kann. Die Messungen an Darmkrebsgewebe zeigen dabei, dass die hohe Auflösung in der Raman basierten SHP von Nutzen ist, um zelluläre Strukturen, wie mobile Zellen im Gewebe aufzulösen und sogar Zellkompartimente einzelner Zellen einer Darmkrebszelllinie zu visualisieren

Cisplatin (CDDP) is a principal compo­ nent of standard ovarian cancer chemother­ apy. However, the frequent development of cellular resistance to CDDP severely restricts the clinical therapeutic success. The molecu­ lar mechanisms of resistance are manifold and not completely understood [1]. Previously, we reported that liposo­ mal CDDP overcomes CDDP resistance of A2780cis human ovarian cancer cells [2]. However, the superior cytotoxic activi­ ty of liposomal CDDP vs. the free drug in A2780cis cells did not correlate with DNA platination levels, thus suggesting a different mode of action of liposomal CDDP com­ pared to free CDDP. To gain an insight into liposomal CDDP activity, a transcriptome analysis of A2780cis cells, either untreated, or in response to ex­ posure with IC50 values of free or liposomal CDDP has been performed [3]. A process network analysis of up­regulated genes indi­ cated that liposomal CDDP induced a highly different gene expression profile compared to the free drug. p53 was identified as play­ ing a key role in directing transcriptional responses to free or liposomal CDDP. Most interestingly, the transcriptome data showed that liposomal CDDP, in contrast to the free drug, induces expression of genes from the extrinsic pathway of apoptosis, which indi­ cates how liposomal CDDP can overcome chemoresistance of A2780cis cells. The aim of this study was to gain further insight into liposomal CDDP activity and to confirm prior findings on apoptosis induction on a protein and functional level. Materials and methods

The frequent development of cellular resistance to cisplatin in cancer patients is a serious limitation for clinical drug therapy. However, cisplatin resistance is incompletely understood. We have shown that cisplatin-resistant A2780 ovarian cancer cells (A2780cis) can efficiently be eliminated by liposomal cisplatin, which displayed similar cytotoxicity towards both A2780 and A2780cis cells. This may, at least in part, be related to a higher intracellular accumulation of the drug within the resistant cells after liposomal entry. However, the superior cytotoxicity of the liposomal drug was not reflected by DNA platination. This suggests a more complex mode of action of liposomal cisplatin, most likely affecting different signaling pathways. To gain insight into the resistance gene signature, a whole-genome gene expression analysis was performed for A2780cis cells, untreated or treated with half-minimal inhibitory concentration (IC50) of free and liposomal cisplatin. Strong differences in the functional networks affected by free and liposomal cisplatin became evident. p53 was identified as a key factor directing differences in the apoptotic processes. While free cisplatin induced the intrinsic pathway of apoptosis, liposomal cisplatin induced expression of genes of DNA damage pathways and of the extrinsic pathway of apoptosis. These predictions from gene expression data were confirmed at the protein and function level. This sheds new light on liposomal drug carrier approaches in cancer and suggests liposomal cisplatin as a promising strategy for the treatment of cisplatin-resistant ovarian carcinoma.

The present PhD thesis describes the development of new organic selenium and tellurium containing compounds as synthetic redox-modulating surfactants and their biological activities. Ten organic selenide and telluride compounds (in publication 1) were designed, successfully synthesized and chemically characterized. These compounds are amphiphilic. The tellurides, in contrast to their selenide analogous, are more effective against not only RAW 264.7 macrophages, S. feltiae and E. coli but also against HCT116 cancer cells. The tellurides can modulate the intracellular redox balance by generating the superoxide anion radicals, which trigger the intrinsic apoptotic pathway. Further tests (in publication 2) show that the selenide compound DP31 has no effect on either HCT116 cancer cells or ARPE-19 cells, while the telluride analog DP41 induces the activation of the PERK/eIF2α/ATF4 signaling pathway by modulating ROS, which triggers the intrinsic apoptotic pathway in HCT116 cancer cells, but not in ARPE-19 cells. Based on the success of tellurium compounds such as DP41, a series of surfactants containing seleninic acid head groups has been designed, synthesized, purified and characterized chemically (in publication 3). These compounds form micelles and interact strongly with the cell membrane of RBCs. Hence they promote Ca2+ influx, which induces eryptosis. Furthermore, the series of these surfactants with CMC properties also shows anti-funginal activity against S. cerevisiae. Diese Dissertation beschreibt die Entwicklung der neuen organischen Selen- und Tellur-Verbindungen als Redox modulierende Tensiden und ihre biologischen Aktivitäten. Zehn organische Selenid- und Tellurid-Verbindungen (in der Veröffentlichung 1) wurden erfolgreich synthetisiert und chemisch charakterisiert. Sie sind amphiphil. Die Telluriden, im Gegensatz zu den analogen Seleniden, sind nicht nur wirksamer gegen RAW 264.7 Makrophagen, S. feltiae und E. coli sondern auch gegen HCT116 Krebszellen. Diese Telluriden können das Redoxgleichgewicht durch die Erzeugung von Superoxid Radikalen so modulieren, dass der intrinsische apoptose ausgelöst wird. Weitere Experimente sind in der Veröffentlichung 2 zusammengefasst. Die Selenid DP31 zeigt keinen Effekt auf HCT116 Krebszellen oder ARPE-19 Zellen. Hingegen aktiviert das analoge Tellurid, DP41, die Signalweg von PERK/eIF2α/ATF4 durch Generierung von ROS, wodurch die intrinsische Apoptose in HCT116 Krebszellen ausgelöst wird. Dies geschieht aber nicht in ARPE-19 Zellen. Basierend auf dem Erfolg der Telluriden, wie beispielsweise DP41, wurde eine Reihe von Seleninsäure-enthaltender Tenside synthetisiert und chemisch charakterisiert (in der Veröffentlichung 3). Sie formen Mizellen, interagieren stark mit der Zellmembran von Erythrozyten und fördern den Ca2+ Einstrom. Dies induziert wiederum Eryptose. Interessanterweise zeigen diese Seleninsäure-enthaltenden Tenside mit amphiphiler Eigenschaft auch fungizide Aktivität gegen S. cerevisiae.

submitted by Liisa Andersen Zsfassung in dt. und engl. Sprache Graz, Univ., Masterarb., 2013

Mit Hilfe der Festphasen-Peptidsynthese (SPPS) wurden Azid-funktionalisierte Ferrocenyl- und Phosphin-Gold(I)-Metallkomplexe mit Alkin-funktionalisierten MPP (mitochondria penetrating peptides) in [3+2]-Cycloadditionsreaktionen verbunden, mittels HPLC, \(^{1}H-\), \(^{31}P-NMR\) und MS charakterisiert und auf ihre Zytotoxizität gegen Krebszelllinien untersucht. Die organometallischen Gold(I)-Peptidkonjugate zeigten dabei vielversprechende Aktivität. Für die fluoreszenzmikroskopische, intrazelluläre Detektion der Gold(I)-Peptidkonjugate wurde Thiazolorange (TO) an den N-terminus der Peptidkonjugate gekoppelt. Mittels Atomabsorptionsspektroskopie (AAS) wurde die Zellaufnahme und durch Isolieren der Mitochondrien auch die mitochondriale Aufnahme der Gold(I)-Peptidkonjugate quantitativ bestimmt. Weitere Untersuchungen wurden \(\textit {in vitro}\) am Redoxenzym Thioredoxin-Reduktase (TrxR), isolierten, funktionalen Mitochondrien und cisplatin-resistenten Zellen durchgeführt.

For cancer therapy TRAIL is of special interest because TRAIL has been shown to preferentially kill cancer cells but sparing normal cells. Importantly, combined treatment of cells with TRAIL together with chemotherapy or gamma-irradiation has been demonstrated to synergize and to achieve higher anticancer activity in cancer cell lines and in vivo tumor models. However, pancreatic cancer, a commonly known very aggressive cancer type, has been shown to be frequently TRAIL-resistant. In the study presented here the TRAIL-resistant cell line Panc-1 was used as a model and Panc-1 cells were investigated whether they could be sensitized to TRAIL-induced apoptosis. In order to characterize TRAIL resistance in Panc-1 cells analyses of factors that may contribute to TRAIL resistance in Panc-1 cells were carried out. Adenovirus vectors were established to express shRNA to down-regulate anti-apoptotic proteins such as c-FLIP, Bcl-xl, XIAP or alternatively to over express pro-apoptotic factors such as the death receptor TRAIL-R1. Moreover, the expression of caspase-8 was induced in these targeted cells by the addition of interferon-gamma. These treatments were verified to yield a balance between over-expression of pro-apoptotic factors and down-regulation of anti-apoptotic factors. This optimization of the combined treatments was proven to be necessary to overcome TRAIL resistance in Panc-1 cells. Finally, by analyzing the long-term effect of down-regulation of XIAP in those cells we observed a change in phenotype and in proliferation. The analyses of these cells revealed that these stable XIAP knock-out Panc-1 cells yielded cells with a phenotype, which is senescence-like. Thus these cells are no longer capable to proliferate and the phenotype of these aggressive pancreatic cancer cells is dramatically changed since they lost their proliferation capabilities. Forcing such aggressive pancreatic cells into senescence may reveal a new avenue for future pancreatic cancer patient treatment

Mit Hilfe der Festphasen-Peptidsynthese (SPPS) wurden Azid-funktionalisierte Ferrocenyl- und Phosphin-Gold(I)- Metallkomplexe mit Alkin-funktionalisierten MPP (mitochondria penetrating peptides) in [3+2]-Cycloadditionsreaktionen verbunden, mittels HPLC, 1H-, 31P-NMR und MS charakterisiert und auf ihre Zytotoxizität gegen Krebszelllinien untersucht. Die organometallischen Gold(I)-Peptidkonjugate zeigten dabei vielversprechende Aktivität. Für die fluoreszenzmikroskopische, intrazelluläre Detektion der Gold(I)-Peptidkonjugate wurde Thiazolorange (TO) an den N- terminus der Peptidkonjugate gekoppelt. Mittels Atomabsorptionsspektroskopie (AAS) wurde die Zellaufnahme und durch Isolieren der Mitochondrien auch die mitochondriale Aufnahme der Gold(I)-Peptidkonjugate quantitativ bestimmt. Weitere Untersuchungen wurden in vitro am Redoxenzym Thioredoxin-Reduktase (TrxR), isolierten, funktionalen Mitochondrien und cisplatin-resistenten Zellen durchgeführt.

To obtain more evidence for intracellular magnetic fluid hyperthermia (MFH), endocytosis and hyperthermia efficacy of silan and dextran magnetite was investigated. Differential endocytosis was observed in dependence of nanoparticle and cell type. Clonogenic survival was 3-fold lower after MFH versus waterbath hyperthermia. The selective “remote inactivation” of cancer cells by an AC magnetic field has been demonstrated in vitro.

Es wurden Metallocen-Peptid-Biokonjugate der Peptide Octreotat, Neurotensin, Polyarginin, sowie der SS-Peptide und einer Mitochondrialen-Targeting-Sequenz erfolgreich mithilfe der Festphasen-Peptidsynthese hergestellt. Für die mikroskopischen Untersuchungen konnten die Biokonjugate mit einem Fluorophor funktionalisiert werden, indem der Fluoreszenzmarker FITC nach selektiver Entschützung einer Lysin-Seitenkette an diese gekuppelt wurde. Die synthetisierten Biokonjugate wurden mithilfe von HPLC, Massenspektrometrie und \(^{1}H\)-, \(^{13}C\)- und 2D-NMR-Spektroskopie charakterisiert. Zusätzlich wurden ausgewählte Verbindungen mittels IR- und Raman-Spektroskopie und Cyclovoltammetrie untersucht. Die Verbindungen wurden auf ihre Zellaufnahme, Lokalisation in der Zelle und antiproliferative Aktivität hin untersucht. Die Biokonjugate zeigen eine zum Teil vielversprechende zytotoxische Aktivität, die sowohl von dem Metallkomplex als auch von dem verwendeten Peptid abhängig ist.

Natural sulfur compounds from plants, bacteria, fungi and animals frequently exhibit interesting biological activities, such as antioxidant, antimicrobial and anticancer activity. Considering the recent developments in medicine (e.g. oxidative stress in ageing, antibiotic resistant bacteria, and selective anticancer agents) and Agriculture (e.g. 'green'; pesticides), several of these compounds have become the focus of interdisciplinary research. Among the various sulfur agents isolated to date, polysulfides, such as diallyltrisulfide, diallyltetrasulfide (from garlic) and dipropyltrisulfide, dipropyltetrasulfide (from onion), are of particular interest, since they combine an unusual chemistry and biochemical mode(s) of action with a distinct biological activity, which includes antimicrobial activity and cytotoxicity against certain cancer cells. As part of this PhD thesis, the activity of diallyltrisulfide and diallyltetrasulfide against the fairly 'robust'; Caco-2 colon cancer cell line and induction of apoptosis and cell cycle arrest in U937 cells have been confirmed. Accordingly, diallyltetrasulfide triggered the programmed cancer cell death- both via the extrinsic and intrinsic pathway. Similarly, these polysulfides showed a good activity in nematode toxicity assays considering them as potential green nematicides. Controls with the long chain carbon analogue 1,9-decadiene eliminate the possibility of solely lipophilic effects of diallyltetrasulfide and, together with the 'ranking'; of activity, point toward a special sulfur redox chemistry which emerges when shifting from the di- to the trisulfide. The electrochemical studies and thiol oxidation assays, however, count against the notion of diallyltrisulfide and diallyltetrasulfide as effective oxidants. On the contrary, the rather negative oxidation and reduction potentials associated with these agents point toward a reducing chemistry, which is confirmed in the Nitrotetrazolium Blue assay. It is therefore likely that diallyltrisulfide and diallyltetrasulfide are reduced inside the cancer cells to perthiols and hydropolysulfides, which in turn trigger a lethal oxidative burst via superoxide radical anion formation. Further interdisciplinary studies are required to investigate in more detail the rather complicated chemical and biochemical processes which ultimately may explain the biological activity which is clearly associated with many natural polysulfides. Natürliche Schwefelverbindungen von Pflanzen, Bakterien, Pilzen und Tieren zeigen oft interessante biologische Aktivitäten, wie antioxidative, antimikrobiale und Antikrebs-Wirkungen. Angesichts der derzeitigen Entwicklung in Medizin (z.B. oxidativer Stress beim Alterungsprozess, gegen Antibiotika resistente Bakterien und selektive Antikrebs-Mittel) und Landwirtschaft (z.B. "grüne" Pestizide) haben verschiedene dieser Verbindungen ein besondeses Augenmerk in interdisziplinärer Forschung erhalten. Unter diversen bislang isolierten Schwefel-Agenzien sind Polysulfide, wie Diallyltrisulfid, Diallyltetrasulfid (von Knoblauch), so wie Dipropyltrisulfid und Dipropyltetrasulfid (von Zwiebeln) von besonderem Interesse, und diese Verbindungen vereinen eine ungewöhnliche Chemie und biochemische Wirkungsweise mit einer ausgeprägten biologischen Aktivität, welche antimikrobiale Aktivität and Zytotoxizität gegenüber bestimmten Krebszellen umfasst, Als Teil dieser Dissertation wurde die Aktivität von Diallyltrisulfid und Diallyltetrasulfid gegen die "robuste" Caco-2 Darmkrebs-Zellinie, so wie die Induktion von Apoptose und Zellzyklus-Arrest in U937-Zellen bestätigt. Dementsprechend löst Diallyltetrasulfid programmierten Zelltod über den extrinsischen als auch den intrinsischen Weg aus. Gleichermaßen zeigten diese Polysulfide eine gute Aktivität im Nematoden-Toxizitäts-Assay, welches sie als potenzielle grüne Nematizide im Betracht ziehen lässt. Durch Kontrollen mit dem langkettigen Kohlenstoff-Analogen 1,9-Decadien lässt sich die Möglichkeit der alleinigen lipophilen Effekte von Diallytetrasulfid ausschließen, was zusammen mit dem "Ranking" der Aktivitäten auf eine besondere Schwefel-Redoxchemie hindeutet, wie sie beim Übergang vom Di- zum Trisulfid auftritt. Die elektrochemischen Studien und Thiol-Oxidations-Assays sprechen gegen Diallyltrisulfid und Diallyltetrasulfid als effektive Oxidantien. Im Gegensatz, die eher negativen Oxidations- und Reduktionspotentiale deuten auf eine Reduktionschemie hin, welches durch das Nitrotetrazoliumblau-Assay bestätigt wurde. Deshalb ist es wahrscheinlich, dass Diallyltrisulfid und Diallyltetrasulfid in Krebszellen zu Perthiolen und Hydropolysulfiden reduziert werden, die wiederum einen letalen oxidativen Burst durch Superoxid-Radikalanionen-Bildung auslösen.

Bladder cancer is the second common malignant urological neoplasia. Most of the tumors are superficial (80%) at first diagnosis and reccur frequently. In order to understand the initial genetic aberrations reflecting growth advantage in bladder cancer we investigated first chromosomal aberrations and validated their biological potential at single cell level. Using multi-colour fluorescence in situ hybridisation (FISH; Urovysion) and Ki-67 immunohistochemistry first data was aquired and completed by laser-microdissecting single cells for single-cell comparative genomic hybridisation (CGH) analy-ses. Double staining of fluorescence in situ hybridisation (Urovysion, Vysis/Abbott) and Ki-67 immunohistochemistry was carried out on frozen tissue sections from 25/40 patients with pre-cancerous lesions of the bladder (13 hyperplasia, 12 dysplasia; those with preferably primary diagnosis; and further specimen from 7 carcinoma in situ, 3 pTaG1, 4 pT1G3). In addition 55 single cells of these precancerous lesions were laser-microdissected and analysed with single cell comparative genomic hybridisation (CGH). Focussing on the proliferating cells versus their non-proliferative neighbourhood in precan-cerous lesions of the bladder, chromosomal aberrations were detected in both types of cells. Proliferating hyperplastic cells showed almost a normal, diploid FISH and no further loss of chromosomal loci in the CGH-analysis. The CGH data of dysplasia cells showed mainly a loss of the chromosomal region 9p21 in proliferating cells, like expected from FISH results. Other chromosomal aberrations, depicted in dysplasia cells, were deletion of chromosome 9 and 13q as well as amplifications of the chromosomes 9p, 12p, 17q, 18p, 22, X and Y. In this regions many candidate genes, involved in regulation of cell proliferation and differentiation, apoptosis and metabolism, are located. These methods established are apt to show that genetic aberrations detected in early bladder lesions or normal urothelium are biologically relevant since found in proliferating cells. This work has been supported by the German Science Foundation (DFG, grant no: Kn263/9-2).

Background Glyoxalases (Glo1 and Glo2) are involved in the glycolytic pathway by detoxifying the reactive methylglyoxal (MGO) into D-lactate in a two-step reaction using glutathione (GSH) as cofactor. Inhibitors of glyoxalases are considered as anti-inflammatory and anti-carcinogenic agents. The recent finding that various polyphenols modulate Glo1 activity has prompted us to assess curcumin's potency as an Glo1 inhibitor. Methodology/Principal Findings Cultures of whole blood cells and tumor cell lines (PC-3, JIM-1, MDA-MD 231 and 1321N1) were set up to investigate the effect of selected polyphenols, including curcumin, on the LPS-induced cytokine production (cytometric bead-based array), cell proliferation (WST-1 assay), cytosolic Glo1 and Glo2 enzymatic activity, apoptosis/necrosis (annexin V-FITC/propidium iodide staining; flow cytometric analysis) as well as GSH and ATP content. Results of enzyme kinetics revealed that curcumin, compared to the polyphenols quercetin, myricetin, kaempferol, luteolin and rutin, elicited a stronger competitive inhibitory effect on Glo1 (Ki = 5.1±1.4 µM). Applying a whole blood assay, IC50 values of pro-inflammatory cytokine release (TNF-a, IL-6, IL-8, IL-1v) were found to be positively correlated with the Ki-values of the aforementioned polyphenols. Moreover, whereas curcumin was found to hamper the growth of breast cancer (JIMT-1, MDA-MB-231), prostate cancer PC-3 and brain astrocytoma 1321N1 cells, no effect on growth or vitality of human primary hepatocytes was elucidated. Curcumin decreased D-lactate release by tumor cells, another clue for inhibition of intracellular Glo1. Conclusions/Significance The results described herein provide new insights into curcumin's biological activities as they indicate that inhibition of Glo1 by curcumin may result in non-tolerable levels of MGO and GSH, which, in turn, modulate various metabolic cellular pathways including depletion of cellular ATP and GSH content. This may account for curcumin's potency as an anti-inflammatory and anti-tumor agent. The findings support the use of curcumin as a potential therapeutic agent.

eingereicht von Leander Breitkreuz Literaturverzeichnis: Seite 93-99 Masterarbeit Paris-Lodron-Universität Salzburg 2020 Abstract in englischer und deutscher Sprache