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Synthesis of fluorinated nucleosides
Nguyen, Van Hai ; Hocek, Michal (advisor) ; Baszczyňski, Ondřej (referee)
The key intermediate 6-amino-7-iodo-7-deazapurine 3'-deoxy-3'-fluororibonucleoside was synthesized using multistep sequence of several reactions, which started from the commercially available D-xylose and 6-chloro-7-deazapurine. The synthetic strategy was based on fluorination of sugar and glycosylation with corresponding nucleobase afterwards. The fluorination of 5-protected-1,2-isopropylidine xylose with different protecting groups at position 5 always led to elimination. It was later discovered that isopropylidine forces the conformation, which is unfavorable for substitution. During the extensive optimization it was also found out that DAST appears to be an optimal fluorinating agent. Fluorination was performed on 2,3-unprotected xylose, which was subsequently used for glycosylation. After several unsuccessful attempts on "protection group free" glycosylation, Vorbrüggen glycosylation was successful and gave desired 3'-fluoro nucleoside in good yield. However, benzoyl group had to be introduced into position 2'. The protected nucleoside was then aminated and simultaneously deproctected with solution of aqueous NH3 and 1,4-dioxane. The obtained key intermediate was used for synthesis of a small series of desired 6-amino-7-hetaryl nucleoside using Pd-catalyzed Suzuki reaction under aqueous...
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Study of adenosine effects on proliferation of BeWo cell line
Papírník, Josef ; Červený, Lukáš (advisor) ; Vokřál, Ivan (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Josef Papírník Supervisor: doc. PharmDr. Lukáš Červený, Ph.D. Title of diploma thesis: Studium of adenosine on proiferation activity in BeWo cell line The placenta is a rapidly developing organ that provides nutrition, protection and environment for the growing fetus. Fetal development is dependent on the supply of nutrients from the mother's blood either by passive diffusion or mediated by transporters. One of the essential nutrients are nucleosides, which are known to promote DNA synthesis and thus the growth of certain tissues. In order to pass across the plasma membrane they need nukleoside transporters (NTs) because of its hydrophilic feature. Additionally Ado takes part in cell signaling. Its wide- ranging effects, including influencing proliferation, are mediated by its binding to adenosine receptors (ARs). The placenta expresses NTs and ARs, which means that it is equipped to uptake nucleosides from maternal blood and has ability to receive signals from the external environment via the adenosine molecule. However, the importance of nucleosides for placental growth has not been investigated yet. The aim of this thesis is to test the effect of nucleosides on the proliferation of...
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Study of adenosine effects on proliferation of JEG-3 cell line
Nguyen, Ngoc Duong ; Červený, Lukáš (advisor) ; Jirkovský, Eduard (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Nguyen Ngoc Duong Supervisor: doc. PharmDr. Lukáš Červený, Ph.D. Title of diploma thesis: Effect of adenosine on the proliferation of JEG-3 cell line Adenosine is a purinergic signaling molecule that is used in nucleic acid synthesis. Transport of hydrophilic nucleosides through the plasma and/or organelle membranes is provided by equilibrative nucleoside transporters (ENTs), members of the SLC29A transporter family, and concentrative nucleoside transporters (CNTs), members of the SLC28A transporter family. The placenta is a complex and rapidly growing organ. It shows some patterns similar to tumors except for the fact that the placenta's growth is fully controlled. It was found that extracellular nucleosides support the proliferation of cancerous and some non-cancerous cells. The placenta expresses high levels of NTs which indicates the placenta's ability to take up the nucleosides from circulation, however, the contribution of this process to placental growth is known. The diploma thesis aims to test the effect of adenosine and other nucleosides on trophoblast proliferation. We evaluated whether adenosine and other nucleosides increase the proliferation of the choriocarcinoma-derived JEG-3 cell...
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Modified ribonucleotides as building blocks for enzymatic construction of functionalized RNA or as antiviral compounds
Milisavljević, Nemanja ; Hocek, Michal (advisor) ; Baszczyňski, Ondřej (referee) ; Krečmerová, Marcela (referee)
The aim of this thesis was to study the steric influence of the base-modified nucleoside triphosphates (NTPs) on the enzymatic incorporation into RNA, as well as to study their inhibitory effect on different viral RNA polymerases in vitro. Their parent nucleosides and prodrug derivatives were also prepared and their antiviral activity evaluated. In the first part of the thesis, NTPs bearing groups varying in size from small methyl and ethynyl substituents via medium-size phenyl and benzofuryl groups, up to large dibenzofuran ring were prepared. Aromatic substituents were installed via Suzuki coupling on iodinated triphosphates or, in the case of modified guanosines, by the phosphorylation of modified nucleosides. Methyl and ethynyl NTPs were prepared via Pd-catalyzed coupling with AlMe3 and Sonogashira coupling, respectively, followed by the phosphorylation of modified nucleoside. To examine their incorporation into RNA by T7 RNA polymerase, templates coding for 35mer RNA containing one, three or seven modifications were designed. Modified pyrimidine triphosphates worked well for all the sequences, while the biggest dibenzofuryl group was not accepted in the difficult sequence with seven modifications. In the case of AR TPs dibenzofuryl modification did not incorporate at all, while other...
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Study of redox reactions and adsorption of 4-nitrophenyl triazole labeled nucleosides and cresols on boron doped diamond electrodes for development of electroanalytical methods
Vosáhlová, Jana ; Schwarzová, Karolina (advisor) ; Havran, Luděk (referee) ; Navrátil, Tomáš (referee)
In this work, the application of boron-doped diamond electrodes with various surface pre- treatment were tested on selected oxidizable and reducible model compounds, specifically para and ortho cresols and 2'-deoxycytidine and 7-deazaadenosine labeled by 4-nitrophenyl triazole. The aim of the study was the investigation of electrochemical and adsorptive processes with respect to the development of electroanalytical methods of detection of the selected compounds, or utilization of boron-doped diamond electrodes as detectors in liquid-flow systems. Cresols were used as the model oxidizable compound. On O-terminated and polished boron- doped diamond electrodes, cresols provide oxidation signal owing to their oxidation to methylphenoxy radical at comparable potentials as on other carbon-based electrodes used as a reference in this work, i.e., glassy carbon electrode, pyrolytic graphite electrode, and carbon paste electrode. These carbon electrode materials show relatively high propensity to adsorption of reaction products, while such adsorption is minimal on boron-doped diamond. In situ anodic activation allows for a rapid regeneration of boron-doped diamond surface prior to each scan. A differential pulse voltammetric method that was developed with a detection limit of 0.61 mol∙l-1 to 2.97 mol∙l-1...
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Synthesis of fluorinated nucleosides
Nguyen, Van Hai ; Hocek, Michal (advisor) ; Baszczyňski, Ondřej (referee)
The key intermediate 6-amino-7-iodo-7-deazapurine 3'-deoxy-3'-fluororibonucleoside was synthesized using multistep sequence of several reactions, which started from the commercially available D-xylose and 6-chloro-7-deazapurine. The synthetic strategy was based on fluorination of sugar and glycosylation with corresponding nucleobase afterwards. The fluorination of 5-protected-1,2-isopropylidine xylose with different protecting groups at position 5 always led to elimination. It was later discovered that isopropylidine forces the conformation, which is unfavorable for substitution. During the extensive optimization it was also found out that DAST appears to be an optimal fluorinating agent. Fluorination was performed on 2,3-unprotected xylose, which was subsequently used for glycosylation. After several unsuccessful attempts on "protection group free" glycosylation, Vorbrüggen glycosylation was successful and gave desired 3'-fluoro nucleoside in good yield. However, benzoyl group had to be introduced into position 2'. The protected nucleoside was then aminated and simultaneously deproctected with solution of aqueous NH3 and 1,4-dioxane. The obtained key intermediate was used for synthesis of a small series of desired 6-amino-7-hetaryl nucleoside using Pd-catalyzed Suzuki reaction under aqueous...
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