<p>Soja predstavlja jedan od osnovnih hraniva koje se koristi u ishrani pilića u tovu, za<br />zadovoljenje potreba u proteinima i esencijalnim aminokiselinama. Po ukupnom<br />sadržaju proteina i njihovoj biolo&scaron;koj vrednosti soja se svrstava u jedno od<br />najkvalitetnijih proteinskih hraniva, čija aminokiselinska struktura može potpuno da<br />zadovolji potrebe pilića u skoro svim esencijalnim aminokiselinama. Prisustvo<br />Kunitz tripsin inhibitora (KTI), i drugih termolabilnih antinutritivnih faktora<br />ograničava upotrebu soje uz obaveznu termičku obradu. Selekcijski napredak<br />omogućio je stvaranje novih sorti koje imaju niži nivo antinutritivnih faktora, među<br />kojima je i sorta &bdquo;Lana&ldquo;. Prema ostvarenim rezultatima upotreba soje sorte &bdquo;Lana&ldquo; u<br />sme&scaron;ama (grover i fini&scaron;er) za ishranu brojlera u koncentraciji od 30% dala je<br />značajno lo&scaron;ije proizvodne parametre u odnosu na termički tretiranu, ali i značajno<br />bolje u odnosu na termički neobrađenu sortu sa standardnim nivoom KTI.<br />Zabeležena ekonomska efikasnost i indeks cena u SL grupi je skoro isti kao i kod<br />grupa sa termičkim tretmanom. Najveća povr&scaron;ina i visina crevnih resica, kao i<br />dubina kripte je zabeležena u SL grupi. Konstatovana je statistički vrlo značajna<br />hipertrofija pankreasa i uvećanje jetre kod SS grupe ali i značajno uvećanje<br />pankreasa u SL grupi u odnosu na grupe sa termičkim tretmanom. Evidentna je i<br />značajno lo&scaron;ija svarljivost hranljivih materija (marker metoda, i metoda totalne<br />kolekcije) u SL i SS grupama nasuprot onih grupa koje su bile na termičkom<br />tretmanu. Kod upotrebe različitih nivoa sorte &bdquo;Lana&ldquo; u fini&scaron;er (peletiranim)<br />sme&scaron;ama za ishranu brojlera, kada su u pitanju proizvodni parametri, dobijene<br />vrednosti su zadovoljavajuće uz sporadična odstupanja u grupama SL14% i SL21%.<br />Takođe Indeks cena i ekonomske efikasnosti je vrlo približan kontrolnoj grupi.<br />Svarljivost u distalnom ileumu je bila značajno bolja u kontrolnoj i 7%SL grupi, za<br />razliku od proksimalnog ileuma gde ne postoji statistička značajnost. Svarljivost<br />azota u distalnom ileumu je bila značajno lo&scaron;ija u 21% SL grupi, dok u<br />proksimalnom ileumu nije bilo statistički značajnih razlika. Retencija azota je bila<br />značajno bolja u kontrolnoj i 7%SL grupi. Razmatrajući sve rezultate, upotreba soje<br />&bdquo;Lana&ldquo; u ishrani brojlera bez prethodne termičke obrade nije preporučljiva u ishrani<br />od 11 dana u koncentraciji od 30% , ali je moguća u peletiranim fini&scaron;er sme&scaron;ama u<br />nivou do 14% udela u sme&scaron;i.</p> / <p>Soybean is one of the basic feed used in the chickens diet for , to satisfy the protein<br />and essential amino acids needs. By total protein content and their biological value,<br />the soybean is classified as one of the best protein sources, whose amino acid<br />structure can fully meet the needs of chickens in almost all essential amino acids.<br />The presence of Kunitz trypsin inhibitor (KTI), and other thermolabile<br />antinutritional factors limiting the use of soy with necessary thermal processing.<br />Selection progress has enabled the creation of new varieties that have lower levels of<br />anti-nutritional factors, including the variety &quot;Lana&quot;. Our results showed that the use<br />of soybean variety &quot;Lana&quot; in mixtures (grower and finisher) for broilers at the<br />concentration of 30% gave a significantly lower production parameters, as compared<br />to heat-treated, but also a significant better, compared to the thermally non-treated<br />soybean variety with standard level of KTI . Noted Economic efficiency and the<br />Price index in the SL group was almost the same as in the group with thermal<br />treatment. Largest surface and height of the villi and crypt depth was observed in the<br />SL group. We found a statistically significant hypertrophy of the pancreas and<br />enlargement of the liver in the SS group and a significant increase in pancreatic at<br />SL group compared to the groups with thermal treatment. Evident is significantly<br />worse digestibility of nutrients (marker method , and the method of total collection )<br />in the SL and SS groups opposed to those groups that were on the thermal treatment.<br />When using a different levels of variety &quot; Lana&quot; in the finisher ( pelleted ) diets for<br />broilers , in terms of production parameters, values obtained were satisfactory with<br />sporadic deviations in groups and SL14% SL21% . Also index price and economic<br />efficiency is highly approximate to the control group. The digestibility at the distal<br />ileum was significantly better in the control and the SL 7%, group in contrast to the<br />proximal ileum, where is no statistical significance. The digestibility of nitrogen in<br />the distal ileum was significantly inferior to 21% SL group, and in the proximal<br />ileum, there is no statistically significant differences. Nitrogen retention was<br />significantly better control, and in 7%SL group. Considering all the results, the use<br />of variety &quot;Lana&quot; in diet for broilers without heat treatment is not recommended in<br />the grower mixture (from 11 day) at the concentration of 30%, but it is freely<br />available for use in pellet for of finisher mixture, at levels up to 14% of mixture.</p>

The p300 transcriptional coactivator has been implicated in the development of a large number of malignancies; however, the precise mechanism of p300-associated tumorigenesis remains unclear. Here, we demonstrate the functional impact of p300 in human melanomas using both genetic and chemical approach. Depletion of p300 in human melanoma cells was associated with cellular growth arrest and senescence. Microarray analysis identified the Microphthalmia-associated transcription factor (MITF), a critical lineage-specific transcription factor in melanocytes and melanomas, as a major downstream target of p300 in human melanoma. Ectopic expression of MITF in p300-depleted melanoma cells allowed rescue of the p300-silencing phenotype, suggesting a critical regulatory axis involving p300 and MITF. Chromatin immunoprecipitation studies revealed direct regulation of MITF transcription through p300 acetylation of proximal regulatory domains. Critically, we identified that Forkhead Box M1 (FOXM1), a potent pro-proliferation transcription factor, is a target of the p300-MITF signaling axis. Further evaluation of p300 regulation of melanoma cell growth was performed using a highly selective p300/CBP HAT inhibitor, 228-1. Inhibition of p300/CBP histone acetyltransferase (HAT) activity was found to significantly inhibit proliferation of multiple melanoma lines in an MITF-dependent fashion. Together, these data support the role of p300 as a promising therapeutic target in human melanoma and suggest particular therapeutic efficacy of small molecule inhibitors of p300 HAT activity in tumors expressing high levels of MITF. / 2018-12-14T00:00:00Z

Many patients suffering from depressive disorders are refractory to treatment with currently available antidepressant medications, while many more exhibit only a partial response. These factors drive research to discover new pharmacological approaches to treat depression. Numerous studies demonstrate evidence of inflammation and elevated oxidative stress in major depression. Recently, major depression has been shown to be associated with elevated levels of DNA oxidation in brain cells, accompanied by increased gene expression of the nuclear base excision repair enzyme, poly(ADP-ribose) polymerase-1. Given these findings and evidence that drugs that inhibit poly(ADP-ribose) polymerase-1 activity have antiinflammatory and neuroprotective properties, the present study was undertaken to examine the potential antidepressant properties of poly(ADP-ribose) polymerase inhibitors.

Background and Objectives: Integrase strand transfer inhibitors (INSTIs), dolutegravir, elvitegravir, and raltegravir, have become integral in the treatment of HIV, with close monitoring of continued efficacy and tolerability. As side effect occurrence varies among subjects receiving these drugs, we sought to perform an exploratory analysis examining the role of several single-nucleotide polymorphisms (SNPs) on drug concentration changes, selected clinical outcomes, and the occurrence of subject-reported adverse events. Methods: Adults (aged ≥ 18 years) receiving INSTI-based regimens for treatment of HIV were recruited and genotyped with an iPLEX ADME PGx Pro v1.0 Panel. Multiple linear or logistic regression with covariates [age, sex, BMI, regimen (in the across-regimen group), regimen duration, and baseline variables (for continuous parameters)] was used to detect significant association of selected clinical data with genetic variants within the study population. Results: In a sample with a median age of 52.5 years (IQR 45.7–57.2) being predominately Caucasian (88.6%) and male (86.4%), this exploratory study discovered several associations between variables and SNPs, when using INSTIs. Abnormal dream occurrence was statistically different between regimens. Additionally, several SNPs were found to be associated with adverse event profiles primarily when all regimens were grouped together. Conclusions: The associations found in this study point to a need for further assessment, within the population living with HIV, of factors contributing to unfavorable subject outcomes. These exploratory findings require confirmation in larger studies, which then may investigate pharmacogenetic mechanisms.

Over a decade, proteasome inhibitors (PIs), bortezomib, carfilzomib (Cfz) and ixazomib, have contributed to a significant improvement in the overall survival for multiple myeloma (MM) patients. However, the response rate of PI was fairly low, leaving a huge gap in MM patient care. Given this, mechanistic understanding of PI resistance is crucial towards developing new therapeutic strategies for refractory/relapsed MM patients. In this dissertation work, we found H727 human bronchial carcinoid cells are inherently resistant to Cfz, yet susceptible to other PIs and inhibitors targeting upstream components of the ubiquitin-proteasome system (UPS). It indicated H727 cells may serve as a cell line model for de novo Cfz resistance and remains UPS dependent for survival. To examine the potential link between proteasome catalytic subunit composition and cellular response to Cfz, we altered the composition of proteasome catalytic subunits via interferon-γ treatment or siRNA knockdown in H727 cells. Our results showed alteration in composition of proteasome catalytic subunits results in sensitization of H727 cells to Cfz. It supported that proteasome inhibition by alternative PIs may still be a valid therapeutic strategy for patients with relapsed MM after having received treatment with Cfz. With this in mind, we designed and synthesized a small library of epoxyketone-based PIs by structural modifications at the P1′ site. We observed that a Cfz analog, harboring a hydroxyl substituent at its P1′ position was cytotoxic against cancer cell lines with de novo or acquired resistance to Cfz. These results suggested that peptide epoxyketones incorporating P1′-targeting moieties may have the potential to overcome Cfz resistance mechanisms in cells. The immunoproteasome (IP), an inducible proteasome variant which is harboring distinct catalytic subunits, LMP2, MECL1 and LMP7 of the proteasome typically expressed in cells of hematopoietic origin, plays a role in immune response and is closely linked to inflammatory diseases. It has been reported that the IP is upregulated in reactive glial cells surrounding amyloid β (Aβ) deposits in brains of Alzheimer’s disease (AD) patients and AD animal models. To investigate whether the IP is involved in the pathogenesis of AD, we examined the impact of IP inhibition on cognitive function in AD mouse models. We observed that YU102, an epoxyketone peptide targeting the IP catalytic subunit LMP2, improved cognitive dysfunction in AD mice without clearance of Aβ deposition or tau aggregation. Our cell line model study also showed a potential mode of action of YU102 which is suppressing pro-inflammatory cytokine production in microglial cells. It suggested that LMP2 contributes to microglia-mediated inflammatory response. These findings supported that LMP2 may offers a valuable therapeutic target for treatment of Alzheimer’s disease, expanding the therapeutic potential of the LMP2-targeting strategy.

Tissue inhibitor of metalloproteinase-3 (TIMP-3) is a protein with multiple functions that include regulating the turnover of the extracellular matrix (ECM) by inhibiting members of the metzincin family. Extracellular levels of soluble TIMP-3 are low, reflecting its binding to components of the ECM including sulfated glycosaminoglycans (SGAGs) and its endocytosis by low density lipoprotein receptor-related protein 1. Because TIMP-3 inhibits ECM-degrading enzymes, the ability of SGAG mimetics to elevate extracellular concentrations of TIMP3 is of interest for osteoarthritis treatment. However, previous studies of such interactions have utilized immobilized forms of the protein or ligands. Here we have quantified the thermodynamics of the interactions of the inhibitory domain of TIMP-3 with chondroitin sulfate (CS), pentosan polysulfate (PPS) and suramin in solution using isothermal titration calorimetry. All three interactions are driven by a (favorable) negative enthalpy ychange combined with an unfavorable decrease in entropy. The heat capacity change (ΔCp) for the interaction of N-TIMP-3 with CS, PPS, or suramin is essentially zero, indicating an insignificant contribution from the hydrophobic effect. Based on the effects of ionic strength on the interaction of N-TIMP-3 with suramin, their interaction appears to be driven by electrostatic interactions. Modeling supports the view that the negatively charged sulfates of CS, PPS, and suramin interact with a cationic region on N-TIMP-3 that includes Lys -26, -27, -30, and -possibly 76 on the opposite face of TIMP-3 from its reactive site for metalloproteases. / Includes bibliography. / Dissertation (Ph.D.)--Florida Atlantic University, 2019. / FAU Electronic Theses and Dissertations Collection

Die Zahl der Tuberkuloseerkrankungen ist in den letzten Jahrzehnten weltweit gestiegen. Da es an innovativen Antituberkulotika mangelt, werden nach wie vor Medikamente der ersten Generation eingesetzt. Das wachsende Problem sind multi-resistente und extrem-resistente Bakterienstämme, die kaum oder gar nicht auf die medikamentöse Therapie ansprechen. Charakteristisch für M. tuberculosis ist eine dicke Zellwand. Der Aufbau der Zellwand ermöglicht es dem Bakterium in den Makrophagen zu persistieren und sich dort zu vermehren. Die Zellwand ist reich an Mykolsäuren und so wenig durchlässig für Fremdstoffe. Das mykobakterielle Zellwandskelett kann man in zwei Teile unterteilen, den Zellwandkern und die äußere Lipidhülle. Die freien Lipide der äußeren Lipidhülle dienen als Signalmoleküle im Krankheitsverlauf und interkalieren mit den Mykolsäuren des Zellwandkerns. M. tuberculosis besitzt für die Fettsäurebiosynthese zwei Enzymkomplexe: Die Typ-I-Fettsäuresynthase, die auch in Säugetieren zu finden ist, produziert Fettsäuren von C16- bis C26-Kettenlänge, die dann in der Typ-II-Fettsäuresynthase (FAS-II) zu Meromykolsäuren verlängert werden. Im Synthesezyklus des FAS-II sind mehrere monofunktionale Enzyme hintereinander geschaltet. Wird eines dieser Enzyme in seiner Funktion gestört, kumulieren Zwischenprodukte und benötigte Zellwandlipide können nicht synthetisiert werden. In der Folge wird die Zellwand instabil und das Bakterium stirbt. Die mykobakterielle Lipidbiosynthese ist somit ein ideales Target für die Entwicklung neuer Antituberkulotika. Ziel dieser Arbeit war es, eine neue Inhibitorklasse des FAS-II Enzyms InhA des M. tuberculosis mittels virtuellem Screening zu finden. Für das virtuelle Screening wurden drei aufeinander aufbauende Pharmakophorhypothesen entwickelt und mit diesen zwei unabhängige Datenbanken durchsucht. Als Grundlage für die Berechnungen des virtuellen Screenings diente die PDB Röntgenkristallstruktur 2h7m mit dem Liganden 1-Cyclohexyl-N-(3,5-dichlorophenyl)-5-oxopyrrolidin-3-carboxamid. Für die Erstellung der Pharmakophorhypothesen wurden zuerst die Strukturen des Enzyms mit und ohne Ligand bezüglich ihrer Konformationsunterschiede vor allem im Bereich der Bindetasche analysiert. Als nächstes wurden die Wechselwirkungen des Liganden mit den Aminosäuren der Bindetasche und dem Cofaktor näher analysiert und die verschiedenen Wechselwirkungsarten hinsichtlich ihrer Relevanz für eine inhibitorische Aktivität beurteilt. Schließlich wurde eine Bindetaschenanalyse durchgeführt und Hotspots für unterschiedliche chemische Funktionalitäten berechnet. Für das Datenbankenscreening wurden das ZINC 'drug-like' Subset (2005) und CCGs MOE 2006 Vendor Compound 3D Collection verwendet, beides Datenbanken exklusiv kommerziell erhältlicher Verbindungen. Das ZINC 'drug-like' Subset wurde über einen für InhA individuell angepassten hierarchischen Filter numerisch reduziert. Von den verbleibenden Verbindungen wurde eine Konformerendatenbank berechnet. Die MOE 2006 Vendor Compound 3D Collection lag bereits als Konformerendatenbank vor und wurde für das Screening 'as-is' verwendet. Mit den Pharmakophorhypothesen I und II wurde das reduzierte ZINC 'drug-like' Subset gescreent. Für die Treffer wurden Fingerprints berechnet, sie danach mithilfe des Tanimotokoeffizienten nach ihrer Ähnlichkeit in Cluster eingeteilt und visuell analysiert; 149 Verbindungen wurden für die Dockingsimulationen ausgewählt. Die MOE Konformerendatenbank wurde ebenso über einen für InhA individuell angepassten hierarchischen Filter numerisch reduziert und mit der Pharmakophorhypothese III gescreent, 28 Verbindungen wurden für die Dockingsimulationen ausgewählt. Die Dockingsimulationen wurden mit den Programmen MOE Dock und Autodock durchgeführt. Die Ergebnisse wurden numerisch ausgewertet und innerhalb der Bindetasche relativ zur jeweiligen zugrunde liegenden Pharmakophorhypothese visuell analysiert; 27 Substanzen wurden schließlich für die Testungen ausgewählt. Die Testungen erfolgten mit einem enzymatischen Assay und einem Assay an attenuierten M. tuberculosis Für die Etablierung des enzymatischen Assays wurde das Enzym InhA mittels Vektortransformation in E. coli überexprimiert und säulenchromatographisch aufgereinigt. Das Substrat 2-trans-Octenoyl-Coenzym A wurde synthetisiert. Von den 27 ausgewählten Substanzen waren 9 im Handel erhältlich und wurden schließlich auf ihre inhibitorische Aktivität getestet. Es wurden ein Thiazolidin-2,4-dion, ein 2-Thioxoimidazolidin-4-on und ein Sulfonamid als aktive Substanzen gefunden. / Worldwide the number of tuberculosis cases has increased in the decades. Since there is a lack of innovative anti-tuberculosis drugs, the first-generation drugs are still used as gold standard. Therefore, strains of mycobacteria, that respond only little or not at all to drug therapy, picture a growing problem. Characteristic of M. tuberculosis is its thick cell wall. The structure of the cell wall allows the bacterium to persist in the macrophages and to multiply there. The cell wall is rich in mycolic acids and, in this, little permeable to xenobiotics. The mycobacterial cell wall skeleton can be divided into two parts, the cell wall core and the outer lipid envelope. The free lipids of the outer lipid envelope serve as signalling molecules in course of the disease, and intercalate with the mycolic acids of the cell wall core. For fatty acid biosynthesis M. tuberculosis has two enzyme complexes: the type I fatty acid synthase, which is also found in mammals, produces fatty acids of C16 to C26 chain length; subsequently, these are extended to meromycolic acids in the type II fatty acid synthase (FAS II). The synthesis cycle of FAS-II consists of mono-functional enzymes that build up on each other. If one of these enzymes is disturbed in its functionality, intermediates accumulate and required cell wall lipids can not be synthesized. As a result, the cell wall turns unstable and the bacterium dies. Therefore, the mycobacterial lipid biosynthesis is an ideal target for developing new antituberculous drugs. The aim of this study was to develop a new inhibitor class of the mycobacterial FAS-II enzyme InhA by means of virtual screening. For the virtual screening three consecutive pharmacophore hypotheses were developed, and with these two independent databases were screened. As a basis for the calculations of the virtual screening the PDB X-ray crystal structure 2h7m with the ligand 1-cyclohexyl-N-(3,5-dichlorophenyl)-5-oxopyrrolidine-3-carboxamide was used. In order to construct the pharmacophore hypotheses, first, the structures of the enzyme with and without a ligand were analyzed for their conformational differences, in particular with respect to the geometry of the binding pocket. Next, the interactions of the ligand with the amino acids of the binding pocket and with the cofactor were analyzed in detail; thereby, the different types of interactions were assessed in terms of their relevance for the inhibitory activity. Finally, a hot spot analysis of the active site was carried out for different chemical functionalities. The ZINC 'drug-like' subset (2005) and CCG's 2006 Vendor MOE 3D compound collection were used for the database screening, both being databases of commercially available compounds. The ZINC 'drug-like' subset was numerically reduced by a hierarchical filter customized for InhA; of the remaining compounds a database of conformers was calculated. The MOE 2006 Vendor 3D Compound Collection was already available as a conformer database. The reduced ZINC 'drug-like' subset was screened with the pharmacophore hypotheses I and II. After calculating fingerprints the hits were clustered according to their similarity using the Tanimoto coefficient and visually analyzed; 149 compunds were selected for the docking simulations. The MOE conformers database also was numerically reduced by a hierarchical filter customized for InhA, and then screened with the pharmacophore hypothesis III, 28 compounds were chosen for the docking simulations. The docking simulations were performed with the programs MOE Dock and Autodock. The results were evaluated numerically, and analyzed visually within the binding pocket relative to the respective underlying pharmacophore hypothesis. Finally, 27 substances were selected for testing. The tests were carried out using an enzymatic assay and an assay on attenuated M. tuberculosis. For establishing the enzymatic assay, the enzyme InhA was overexpressed using vector transformation into E. coli and purified by column chromatography. The substrate 2-trans-octenoyl-coenzyme A was synthesized. Of the 27 selected compounds 9 substances were commercially available and were tested for their inhibitory activity. A thiazolidine-2,4-dione, a 2-thioxoimidazolidine-4-one and a sulfonamide were found to be active.

The SARS virus is the etiological agent of the severe acute respiratory syndrome, a deadly disease that caused more than 700 causalities in 2003. One of its viral proteins, the SARS coronavirus main protease, is considered as a potential drug target and represents an important model system for other coronaviruses. Despite extensive knowledge about this enzyme, it still lacks an effective anti-viral drug. Furthermore, it possesses some unusual features related to its active-site region. This work gives atomistic insights into the SARS coronavirus main protease and tries to reveal mechanistic aspects that control catalysis and inhibition. Thereby, it applies state-of-the-art computational methods to develop models for this enzyme that are capable to reproduce and interpreting the experimental observations. The theoretical investigations are elaborated over four main fields that assess the accuracy of the used methods, and employ them to understand the function of the active-site region, the inhibition mechanism, and the ligand binding. The testing of different quantum chemical methods reveals that their performance depends partly on the employed model. This can be a gas phase description, a continuum solvent model, or a hybrid QM/MM approach. The latter represents the preferred method for the atomistic modeling of biochemical reactions. A benchmarking uncovers some serious problems for semi-empirical methods when applied in proton transfer reactions. To understand substrate cleavage and inhibition of SARS coronavirus main protease, proton transfer reactions between the Cys/His catalytic dyad are calculated. Results show that the switching between neutral and zwitterionic state plays a central role for both mechanisms. It is demonstrated that this electrostatic trigger is remarkably influenced by substrate binding. Whereas the occupation of the active-site by the substrate leads to a fostered zwitterion formation, the inhibitor binding does not mimic this effect for the employed example. The underlying reason is related to the coverage of the active-site by the ligand, which gives new implications for rational improvements of inhibitors. More detailed insights into reversible and irreversible inhibition are derived from in silico screenings for the class of Michael acceptors that follow a conjugated addition reaction. From the comparison of several substitution patterns it becomes obvious that different inhibitor warheads follow different mechanisms. Nevertheless, the initial formation of a zwitterionic catalytic dyad is found as a common precondition for all inhibition reactions. Finally, non-covalent inhibitor binding is investigated for the case of SARS coranavirus main protease in complex with the inhibitor TS174. A novel workflow is developed that includes an interplay between theory and experiment in terms of molecular dynamic simulation, tabu search, and X-ray structure refinement. The results show that inhibitor binding is possible for multiple poses and stereoisomers of TS174. / Das Schwere Akute Respiratorische Syndrom (SARS) wird durch eine Infektion mit dem SARS Virus ausgelöst, dessen weltweite Verbreitung 2003 zu über 700 Todesfällen führte. Die SARS Coronavirus Hauptprotease stellt ein mögliches Wirkstoffziel zur Behandlung dar und hat Modellcharakter für andere Coronaviren. Trotz intensiver Forschung sind bis heute keine effektiven Wirkstoffe gegen SARS verfügbar. Die vorliegende Arbeit gibt Einblicke in die mechanistischen Aspekte der Enzymkatalyse und Inhibierung der SARS Coronavirus Hauptprotease. Hierzu werden moderne computerchemische Methoden angewandt, die mittels atomistischer Modelle experimentelle Ergebnisse qualitativ reproduzieren und interpretieren können. Im Zuge der durchgeführten theoretischen Arbeiten wird zunächst eine Fehlereinschätzung der Methoden durchgeführt und diese nachfolgend auf Fragestellungen zur aktiven Tasche, dem Inhibierungsmechanismus und der Ligandenbindung angewandt. Die Einschätzung der quantenchemischen Methoden zeigt, dass deren Genauigkeit teilweise von der Umgebungsbeschreibung abhängt, welche als Gasphasen, Kontinuum, oder QM/MM Modell dargestellt werden kann. Letzteres gilt als Methode der Wahl für die atomistische Modellierung biochemischer Reaktionen. Die Vergleiche zeigen für semi-empirische Methoden gravierende Probleme bei der Beschreibung von Proton-Transfer Reaktionen auf. Diese wurden für die katalytische Cys/His Dyade betrachtet, um Einblicke in Substratspaltung und Inhibierung zu erhalten. Dem Wechsel zwischen neutralem und zwitterionischem Zustand konnte hierbei eine zentrale Bedeutung für beide Prozesse zugeordnet werden. Es zeigt sich, dass dieser „electrostatic trigger“ von der Substratbindung, nicht aber von der Inhibitorbindung beeinflusst wird. Folglich beschleunigt ausschließlich die Substratbindung die Zwitterionbildung, was im Zusammenhang mit der Abschirmung der aktiven Tasche durch den Liganden steht. Dies gibt Ansatzpunkte für die Verbesserung von Inhibitoren. Aus in silico screenings werden genauere Einblicke in die reversible und irreversible Inhibierung durch Michael-Akzeptor Verbindungen gewonnen. Es wird gezeigt, dass unterschiedlichen Substitutionsmustern unterschiedliche Reaktionsmechanismen in der konjugierten Additionsreaktion zugrunde liegen. Die vorangehende Bildung eines Cys-/His+ Zwitterions ist allerdings für alle Inhibierungsmechanismen eine notwendige Voraussetzung. Letztendlich wurde die nicht-kovalente Bindung eines Inhibitors am Beispiel des TS174-SARS Coronavirus Hauptprotease Komplexes untersucht. Im Zusammenspiel von Theorie und Experiment wurde ein Prozess, bestehend aus Molekulardynamik Simulation, Tabu Search und Röntgenstruktur Verfeinerung ausgearbeitet, der eine Interpretation der Bindungssituation von TS174 ermöglicht. Im Ergebnis zeigt sich, dass der Inhibitor gleichzeitig in mehreren Orientierungen, als auch in beiden stereoisomeren Formen im Komplex vorliegt.

Untersuchung des Wirkmechanismus von AEZS 126 auf drei triple negative Mammakarzinomzelllinien HCC1937, HCC1806 und MDA-MB468 und eine Oestrogenrezeptor positive Zelllinie MCF-7 mittels Kristallviolett assay, FACS und Western Blot. Es konnte gute Antitumorwirkung des Inhibitors in vitro gezeigt werden. / Of more than one million global cases of breastcancer diagnosed each year, a high percentage are characterized as triple-negative, lacking the oestrogen, progesterone and Her2/neu receptors. The incidence exceeds the incidence of malignancies like CML by far. Lack of effective therapies, younger age at onset and early metastatic spread have contributed to the poor prognosis and outcomes associated with these malignancies. Here, we investigate the ability of the PI3 K/AKT inhibitor AEZS 126 to selectively target the triple negative breast cancer (TNBC) cell proliferation and survival in vitro by MTT-assays and FACS-based analysis. Furthermore, the mechanism of cytotoxicity is analysed by FACS-based assays and Western blots. Results AEZS 126 showed good antitumour activity in in vitro models of TNBC as well as in MCF-7 cells. We demonstrated the highly efficient antitumour activity of AEZS 126 in in vitro models of TNBC. Due to the good anti-tumour activity and the expected favourable toxicity profile, AEZS 126 in combination with chemotherapy seems to be a promising candidate for clinical testing in TNBC especially in the basal-like subgroup of TNBC.

The potential use of sodium propionate as a corrosion inhibitor for mild steel in aqueous media is investigated using a range of electrochemical and surface analytical techniques. The use of sodium propionate for the inhibition of mild steel corrosion is discussed, and the effective pH range of sodium propionate using various buffers is investigated. The effectiveness of sodium propionate as an inhibitor for mild steel pitting corrosion in the presence of various concentrations of CI- is studied. The effect of some oxidants, IO3-, BrO3-, NO32- on the anodic behaviour of mild steel in deaerated 0.01M carboxylate solutions of acetate, propionate, formate, succinate and salicylate is investigated. The critical temperature for effective inhibition of mild steel corrosion with sodium propionate is established, and the chemical composition of the film formed on mild steel surface in sodium propionate solution is studied using surface sensitive Fourier transform infrared spectroscopy FTIR. The efficiency of sodium propionate is compared to that of conventional inhibitors and a mechanism for the inhibition of mild steel corrosion with sodium propionate is proposed. / Doctor of Philosophy (PhD)