Influenza A (IVA) continues to pose a growing global threat even as current medications are becoming less effective. One of the main avenues of research into new anti-IVA drugs is its homotetrameric Matrix 2 proton channel (M2A), without which the virus would be unable to release its viral RNA into the host cell. The drug amantadine used to bind and block M2A until near-ubiquitous resistance formed as an M2A-S31N mutation, starting around 1995 and proceeding to 2005 when amantadine was disallowed for use as an anti-IVA drug. The standard organic structure currently being used for M2A inhibitor research comprises an adamantyl foot group, a heterocyclic aryl body group, and a cyclic head group. A sample set of compounds with this standard structure was compared and reviewed, focusing on positive and negative moieties and modifications. Modifications on the foot group were all more or less detrimental, body groups with two heteroatoms were advantageous, and larger head groups appeared better. Four other scaffolds known to literature were proposed for further study due to beneficial aspects of each. Where most anti-M2A research deals exclusively with organic compounds, metals and their potential in drugs have been almost entirely ignored due to the increased toxicity they bring. Free copper was found in past research to be the only first-row transition metal to show significant M2A-inhibitory activity, proposed to do so by binding the H37 cluster that acts as a pH-dependent control switch for the channel. Six overall-neutral copper complexes were synthesized as a combination of amantadine, cyclooctylamine, and null scaffolds with two of either acetate or acetamide arms as chelators. The complexes were found to block both M2A-WT and M2A-S31N. Along with CuCl¬¬2, though, they had little to no effect on M2A-H37A, providing confirming evidence that the copper binds at the H37 tetrad. Only one complex, Cu(cyclooctylamineiminodiacetate), outperformed CuCl2 in channel block studies and efficacy against two IVA strains, but all of the complexes were found to have lower cytotoxicity. Because M2-H37 is highly conserved, these complexes show promise for further testing against all strains of influenza A. Five net-divalent copper complexes were then synthesized with multiple aza or amine groups as chelators. The complexes failed to show any significant activity against M2A, however, which was thought to be due to size or polarity rejection or electromagnetic repulsion. One of the ligands, though, an adamantyl derivative of a tetraaza macrocycle, was a novel compound, and its copper complex, along with two others, were unknown to the CCDC database. The three complexes were characterized by X-ray diffraction and discussed.

Studies were undertaken to investigate if gastric acidity and iron chelation to a meat component enhance nonheme iron absorption. Cereal meals, with and without added proteins, were gavaged into iron-deficient rats. The role of iron chelation was investigated by adding sodium phytate, an iron chelator implicated with decreased iron absorption, to the meals. The role of gastric acidity was investigated by treating the rats with cimetidine, which inhibits gastric acid production. In rats with normal acid production, beef, pork and chicken enhanced iron absorption when phytate had been added to the meals, suggesting a role for chelation in meat enhancement of iron absorption. However, the enhancement by beef and pork was insignificant in cimetidine-treated rats given the cereal + phytate meals, indicating that gastric acid production also plays a role in meat enhancement of iron absorption. Fish and egg white were sometimes inhibitory to iron absorption and, therefore, did not fit the pattern of enhancement demonstrated by beef, pork, and chicken. In a separate experiment, gastric acidity was not directly altered by the protein source included with cereal meals. No significant effects of the various proteins on iron absorption from cereal + phytate meals were observed in a final experiment involving iron-replete rats. In vitro iron solubilizing capacity of beef, pork, chicken, and egg white was positively correlated with enhanced iron absorption by iron-deficient rats. Studies were performed to 1) investigate if ferric iron bound in complex with iron-solubilizing meat components is absorbable, 2) compare the relative iron-solubilizing capacity of meats, and 3) investigate the physicochemical and compositional characteristics of the meat components responsible for the iron solubilizing capability of meat. Iron-solubilizing components of beef were isolated from pH 2 HCl homogenates into dialysis bags (MWCO, 6-8 K). Radiolabelled iron complexes were then generated using ferric iron and either the ILC (isolated low-molecular-weight components) from undigested beef or ascorbate. The bioavailabilities of radioiron in these complexes or as ferric iron were measured as radioiron absorption into the blood one hour after injection into ligated duodenal loops of rats. Iron absorption values were ascorbate-ferrous complexes > beef ILC-ferric complexes > ferric iron (p < .05). In separate experiments, ILC from 0.1 g of various dietary protein sources (beef, pork, chicken, fish, or egg white) were added to 400 μg ferric iron in pH 2 HCl, the pH raised to 7.2, and soluble iron determined in the supernatant after centrifugation at 2,500 g for 10 min. Iron solubilizing capabilities of ILC were pork > beef > chicken > fish > egg white (p < .05). In a final series of experiments, the compositional and physicochemical characteristics of the ILC from the various dietary proteins were investigated.

Flavonols are hydroxyl-substituted flavonoids and naturally occur as secondary metabolites in plants. Several studies have discovered extensive medicinal properties of flavonols. The present work reports on structural and functional investigation of the B ring substituted flavonols based on spectroscopic and electrochemical techniques. The purpose of this study is to determine the influence of the B ring substitutions on the hydrogen bonding interactions, the electronic effects in ruthenium complexes and the Al3+ chelation of B ring substituted flavonols. The electronic effects of the B rings were changed by introducing methyl, methoxy and nitro groups at position 4ʹ on the B ring. The 3ʹ-methyl substitution was performed in order to increase the electronic density of the B ring via inductive effects. The 2ʹ-methyl and 2ʹ, 6ʹ-dimethyl substitutions increased the steric effects around the inter-ring bond between the B and the C rings, and the B ring was highly deconjugated from the AC rings. The intramolecular hydrogen bonding distances at 3-hydroxy-4-carbonyl units of the B ring substituted flavonols were elongated while the dihedral angles between the B and AC increased. Strong intermolecular hydrogen bonding interactions were also observed in the crystal structures of 4ʹ-methylflavonol, 4ʹ-methoxyflavonol, 4ʹ-nitroflavonol and 2ʹ,6ʹ-dimethylflavonol. Furthermore, several crystal packing patterns were observed, and it is postulated that dihedral angles and intramolecular hydrogen bonding distances are both affected by the intermolecular hydrogen bonding interactions and the crystal packing forces. In addition, the ruthenium complexes of B ring substituted flavonols were synthesized and characterized by spectroscopic and electrochemical techniques. B ring substitution effects were minimal in IR spectroscopy and X-ray crystallography. The levels of the conjugation of the rutheniumlavonolate complexes were demonstrated by electronic absorption spectra recorded in methanol at room temperature. The most positive oxidation potential was obtained with the electron withdrawing nitro group substitution, and the electron donating substitutions resulted in more negative oxidation potentials. The spectroscopic investigation of the complex formation of Al(III) with flavonols and 3-hydroxychromone is described. The stoichiometric composition and stability constants are also given. The comparison of the results obtained from Al(III) chelation shows significant effects of the B ring substitutions.

Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Botany Candidate: Nikola Holotíková Supervisor: PharmDr. Jana Karlíčková, Ph.D. Title of Thesis: Can essential amino acids be considered as copper chelators? Copper is a trace element playing an important role in the human organism. Copper levels in the body have to be carefully controlled because disruption of copper homeostasis can cause oxidative damage which may lead to various pathologies. Chelation of copper can potentially be used as a therapeutic tool. Medical chelators should ideally mimic chelators, especially peptides and proteins which are important for regulation of copper homeostasis. Their structure is formed by amino acids linked by peptide bonds. For this reason, the study is focused on selected amino acids. Proteinogenic amino acids are classified into essential and nonessential. Essential amino acids must be obtained from the diet. The aim of this study was to compare the copper chelation and reduction activity of essential amino acids L-histidine and L-methionine, and nonessential amino acids L-cysteine, its dimer L-cystine, L-aspartic acid and L-glutamic acid. Also a L-cysteine-derived compound, N-acetylcysteine, was tested. All of them were tested using spectrophotometric methods at pHs which...

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Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Lucie Liptáková Supervisor: RNDr. Pavlína Hašková, Ph.D. Title of master thesis: In vitro study of newly synthesized potential cardioprotective drugs Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are in an organism generated under normal or pathological conditions. There are antioxidant mechanisms, which protects the organism from their harmful effect. In case of imbalance between ROS/RNS production and antioxidant mechanisms, an oxidative stress is initiated. The oxidative stress is involved in the pathogenesis of many diseases, including cardiovascular desease. In consequence of higher presence of mitochondria and lower presence of antioxidants cardiomyocytes are more sensitive to the oxidative stress. Iron, by catalysing radical's reactions, significantly participates on formation and development of oxidative stress. Elimination of the free iron by iron chelators is one option how to prevent or moderate oxidative stress. The aim of this master theses was to study cardioprotective effect in presence of H2O2 and own toxicity of newly synthetized aroylhydrazone iron chelators (H21, H22, H23, H24, H25 and H26) on rat embryotic cardiomyoblasts H9c2. Protective and toxic...

Palladium-catalysed coupling reactions have been embraced by synthetic chemists as one of the preferred means for smooth formation of new carbon-carbon bonds: a truly ubiquitous methodology of synthesizing complex molecules. This thesis describes the study of a series of palladium(0)-catalysed C2-arylations of a 1-cyclopentenyl ether, equipped with a chiral (S)-N-methyl-pyrrolidine auxiliary. The investigated olefin was demonstrated to undergo Si-face insertion, providing (R)-configuration of the arylated C2-carbon. In addition, the mild and novel palladium(II)-catalysed dominoHeck/Suzuki β,α-diarylation-reduction of a dimethylaminoethyl-substituted chelating vinyl ether was developed using arylboronic acids as arylating agents in combination with 1,4-benzoquinone (BQ). Further, highly regioselective palladium(II)-catalysed α-and β-monoarylation of the chelating vinyl ether was achieved using either a bidentate ligand or by employing ligand-less conditions. These studies demonstrate that the choice of ligands has a profound effect on the reaction outcome, as productive β,α-diarylation could only be obtained by suppressing the competing β-hydride elimination using BQ as the stabilising ligand and terminal reoxidant. The pivotal role of BQ in the reaction was studied using computer-aided density functional theory calculations. The calculations highlight the crucial role of BQ as a Pd(II)-ligand. In addition of serving as an oxidant of palladium, the calculations support the view that the coordination of BQ to the Pd(II)-centre in the key σ-alkyl complex leads to a low-energy pathway, aided by a strong η2 Pd-BQ donation-back-donation interaction. Furthermore, an investigation of the scope and limitations of novel stereoselective and BQ-mediated palladium(II)-catalysed domino Heck/Suzuki β,α-diarylation reactions, involving metal coordinating cyclic methylamino vinyl ethers and a number of electronically diverse arylboronic acids, conducted. In addition, a set of 4-quinolone-3-carboxylic acids, structurally related to elvitegravir and bearing different substituents on the condensed benzene ring, was designed and synthesized as potential HIV-1 integrase inhibitors. Finally, in an effort to identify a new class of HIV-1 protease inhibitors, four different stereopure β-hydroxy γ-lactam-containing inhibitors were synthesized, biologically evaluated, and co-crystallized with the enzyme. / The time 12:05 for the public defense mentioned in the thesis is incorrect. It will take place at 09:15, 2012-06-08.

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Several lines of evidence link Alzheimer's disease (AD) to atherosclerosis (CVD), including that elevated low density lipoprotein (LDL)-cholesterol is a common risk factor. Development of genomic instability could also link the two diseases. Previous fluorescence in situ hybridization (FISH) analyses revealed a clonal expansion of aneuploid smooth muscle cells underlying atherosclerotic plaques. Likewise, cellular and mouse models of AD revealed tau-dependent mitotic defects and subsequent aneuploidy partly resulting from amyloid-beta (A&beta) interference with microtubule (MT) stability, and specific MT motors function. Moreover, AD patients develop aneuploid/hyperploid cells in brain and peripheral tissues, implicating similar mechanism that may lead to apoptosis and neurodegeneration. This dissertation tested the hypothesis that elevated lipoproteins and cholesterol may contribute to genomic instability in AD and CVD and showed that: (1) treatment with oxidized LDL (OX-LDL), LDL and water soluble cholesterol, but not high density lipoprotein (HDL), induced chromosome mis-segregation, including trisomy and tetrasomy 12, 21, and 7 in human epithelial cells (hTERT-HME1), primary aortic smooth muscle cells, fibroblasts, mouse splenocytes and neural precursors; (2) LDL-induced aneuploidy may depend on a functional LDL receptor (LDLR), but not amyloid precursor protein (APP) gene; (3) fibroblasts and brain cells of patient with the mutation in the Niemann-Pick C1 gene (NPC1) characterized by impaired intracellular cholesterol trafficking and changed intracellular cholesterol distribution harbored trisomy 21 cells; (4) young wild-type mice fed high and low cholesterol diets developed aneuploidy in spleen but not in brain cells within 12 weeks; (5) like with the studies on A&beta-induced aneuploidy, calcium chelation reduced OX-LDL and LDL-mediated chromosomal instability; and (6) altering plasma membrane fluidity with ethanol attenuated OX-LDL and LDL-induced aneuploidy. These results suggest a novel biological mechanism by which disrupted cholesterol homeostasis may promote both atherosclerosis and AD by inducing chromosome mis-segregation and development of aneuploid cells. Understanding the cause and consequence of chromosomal instability as a common pathological trait in AD and CVD may be beneficial to designing therapies relevant for both diseases.