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Development of the new method for in vitro selection of DNA aptamers
Bláhová, Kamila ; Míšek, Jiří (advisor) ; Moserová, Michaela (referee)
Aptamers are short sequences of single-stranded DNA or RNA that are able to specifically bind various molecules (drugs, lipids, sugars, proteins, etc.). These aptamers are isolated from large libraries of random oligonucleotide sequences by SELEX (Systematic Evolution of Ligands by Component Enrichment) or in vitro selection. Despite the success of this method, in vitro selection often requires more than ten rounds of affinity selection as well as optimization of selection conditions. To increase the efficiency of aptamer selection, several methods have been developed that use an increase in the number of secondary structures in random oligonucleotide libraries. These methods, based on increasing the possibility of canonical base pairing in single-stranded oligonucleotides, increased the efficiency of the in vitro selection method. In this work, it was tested whether increasing the probability of occurrence of G-quadruplexes, as structural motifs in random sequences, will lead to increased efficiency of aptamer selection. Four single-stranded DNA libraries with different numbers of guanine (25 %, 35 %, 45 %, 55 %) in a random sequence were used. Streptavidin was chosen as the model molecule for selection, against which several aptamers that are not rich in guanine (G) have previously been selected....
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Synthesis of novel types of acyclic nucleoside phosphonates and preparation of prodrugs and drug delivery systems
Kalčic, Filip ; Janeba, Zlatko (advisor) ; Míšek, Jiří (referee) ; Krečmerová, Marcela (referee)
First part of this thesis was focused on the previously overlooked field of C1'-branched acyclic nucleoside phosphonates (ANPs). Five diverse synthetic approaches were developed/optimized affording key 6-chloropurine intermediates bearing N9 -phosphonomethoxyethyl (PME) branched at C1' position in 2-4 steps. It was demonstrated that these intermediates can be further vastly diversified into ANPs bearing both natural and unnatural nucleobases. Single enantiomers as well as racemates of final C1'-branched ANPs (overall 48 final compounds) were prepared and selected compounds were evaluated with respect to their biological properties. The aforementioned ANPs showed no antiviral potency against studied viruses and only weak to moderate cytostatic activity. Adenine C1'-branched ANPs proved to be the most potent currently known inhibitors of Trypanosoma brucei adenine phosphoribosyl transferase (TbrAPRT), an enzyme involved in purine salvage pathway (PSP) of T. brucei. Further biological evaluation of prepared compounds is in progress. Second part of this thesis was focused on development of novel prodrug moieties with higher selectivity index (i.e. toxicity/potency ratio - SI) based on so-called ProTide prodrugs where phenol (present in ProTides) was replaced by tyrosine derivatives. Tenofovir was...
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The preparation of chiral NHC carbene precatalysts
Krejčířová, Kateřina ; Veselý, Jan (advisor) ; Míšek, Jiří (referee)
This bachelor thesis is focused on the preparation of chiral NHC carbene precatalysts, which are used as organocatalysts in a variety of organic transformations leading to the formation of optically active compounds. In the course of this work, several triazolium salts were prepared from suitable starting materials using six reaction steps. Triazolium salts derived from oxazolidinone were obtained from well available starting materials, arylhydrazines and phenylalanine.
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The preparation of chiral model aromatics for the study of biologically important charge/spin transfer processes.
Vilím, Vojtěch ; Starý, Ivo (advisor) ; Míšek, Jiří (referee)
This bachelor thesis is dedicated to synthesis of 1-ethynyl-7-((triisopropylsilyl)ethynyl)- 2-naphthaldehyde 61, 15-((triisopropylsilyl)ethynyl)benzo-[5,6]phenanthro[3,4- h]isoquinoline 62 and 1,4-bis(benzo[5,6]phenanthro[3,4-h]isoquinolin-15-yl)buta-1,3-diyne 63. The helical structure is a common feature of biological structures in which the CISS effect has been observed. To study it, it is necessary to find suitable model substances with a helical structure, which have a large variability of functional groups and for which it has already been proven that the CISS effect takes place on them. Therefore, helicenes are good model molecules. Key words: helicene, chirality, spin filtration
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On Azahelicenes: Synthesis, Resolution, Properties and Applications
Míšek, Jiří
Mgr. Jiří Míšek PhD Thesis Abstract A novel and modular approach to azahelicenes has been developed. Using [2+2+2] cyclotrimerisation of appropriate triyne as a key step, azahelicenes 83, 84 and 85 have been prepared (Scheme 1). Scheme 1. Aza[6]helicenes 84 and 85 have shown to be sufficiently configurationally stable and have been resolve into individual enantiomers by the diastereoisomeric salt crystalisation and chiral HPLC. This allowed to determine exact values of racemisation barriers for both aza[6]helicenes 84 and 85. In general, both hexacyclic derivatives 84 and 85 are stable enough to be used in asymmetric transformations or chiral sensing. The ability of aza[6]helicenes 84 and 85 to serve as chiral ligands for transition metals has been proven by the preparation and X-ray analysis of corresponding silver complexes. Interesting chiral self-recogniton properties of aza[6]helicenes 84 in gas phase have been described using mass spectrometry techniques. In order to complement information about azahelicenes basicities, gas phase proton affinities of derivatives 84 and 85 and solution phase dissociation constants of azahelicenes 83, 84 and 85 have been determined. Eventually, the first utilization of azahelicene in asymmetric catalysis has been demonstrated by the successful use of optically pure...
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Asymmetric Tandem Lithium Amide Conjugate Addition/Radical Reactions and Their Application in the Total Synthesis of Natural Products
Hidasová, Denisa ; Jahn, Ullrich (advisor) ; Kočovský, Pavel (referee) ; Míšek, Jiří (referee)
This thesis deals with single-electron transfer (SET) radical processes mediated by ferrocenium hexafluorophosphate and TEMPO and their application in the total synthesis of natural products. Asymmetric aminooxygenation methodology for the synthesis of anti-β-amino-α-hydroxy acid derivatives has been developed by utilizing a highly diastereoselective aza-Michael addition of chiral lithium amides to various α,β-unsaturated esters or amides/SET oxidation/radical α-oxygenation. The potential of this methodology was demonstrated in short total syntheses of the anti-β-amino-α-hydroxy acid fragments of the macrocyclic (depsi)peptides perthamide C and largamide H, and (-)-cytoxazone, which is a selective modulator of TH2 cytokine secretion. The SET-catalyzed asymmetric tandem lithium amide conjugate addition/5-exo radical cyclization/oxygenation reactions were applied in the synthesis of highly substituted pyrrolidines, azabicyclo[n.3.0]alkanes and spiropyrrolidines. An enantioselective total synthesis of the pyrrolidine alkaloid (-)-α-kainic acid was accomplished by employing the SET-catalyzed 5-exo radical cyclization/oxygenation.
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Use of alkenylboronic acids in the tandem cyclisation/Suzuki cross-coupling
Klanicová, Kristýna ; Matoušová, Eliška (advisor) ; Míšek, Jiří (referee)
This thesis deals with the synthesis of polycyclic compounds using tandem cyclisation/Suzuki cross-coupling, by modification of the method developed by our research group. This thesis mainly investigates the scope of the tandem reaction with aliphatic boronic acids or their esters. The products of the palladium catalyzed reaction then underwent electrocyclization to form an aromatic ring. The polycyclic skeleton, containing the aromatic core, is found in a considerable amount of natural substances with biological activity. The structure of the compounds prepared herein is similar to miltiorin D or commiphorane A.
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Using Heck reaction for aromatic ring formation in the synthesis of polycyclic compounds
Bažíková, Ema ; Matoušová, Eliška (advisor) ; Míšek, Jiří (referee)
This bachelor's thesis focuses on the synthesis of a polycarbocyclic compound with naphthalene core and the following oxidation to the corresponding ortho-naphthoquinone. The naphthalene and naphthoquinone skeleton can be found in many natural products with interesting biological properties. This thesis extends the scope of a method developed by our group for the synthesis of naturally occurring ortho-naphthoquinones. The key step of the synthesis was to form an aromatic ring using the Heck reaction. Keywords: synthesis, polycyclic compounds, naphthalenes, catalysis, Heck reaction
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Study of DNA-compatible Reactions and Their Utilisation for DNA Encoded Libraries
Havelka, Václav ; Míšek, Jiří (advisor) ; Kožíšek, Milan (referee)
DNA-encoded peptide libraries are the basis for in vitro selection methods that use various biological systems (phage display; yeast display; mRNA display). Despite the great success of these selection methods, their obvious disadvantage is the limited number of building blocks, which consist of only twenty proteinogenic amino acids. The involvement of other non- proteinogenic amino acids and other building blocks could significantly expand the range of possible applications of these selection methods. For example, the introduction of chemical modifications in amino acid side chains in such libraries would allow the effective study of post-translational modifications (phosphorylation, acylation, glycosylation, methylation, etc.) in living organisms. The aim of this work was to develop a method for preparation of a fully synthetic DNA encoded library of peptides. The basic steps for the preparation were the chemical synthesis of the peptide and associated enzymatic synthesis of encoding DNA. Compatibility of chemical reactions with DNA is essential for the synthesis of DNA-encoded peptide libraries. Because the final acidic deprotection of the side chains in the peptide is not compatible with DNA, two approaches have been tested to overcome this problem. The first was an attempt to develop finer...
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