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Design and synthesis of novel type of prodrugs of acyclic nucleoside phosphonates based on ProTides
Markušová, Katarína ; Janeba, Zlatko (advisor) ; Kundrát, Ondřej (referee)
This diploma thesis focuses on the synthesis of novel prodrugs of tenofovir, one of the main components of medications against human immunodeficiency virus (HIV) infection. The design of the compounds is based on the concept of ProTides and takes inspiration from the previous work of Filip Kalčic. In this work, the aromatic ester on the phosphonate moiety, typical for the ProTides, is replaced by an aliphatic ester. This aliphatic moiety is represented by a derivative of the amino acid L-serine. All of the target compounds were synthesized in a one-pot manner from three key components (L-serine, L-alanine, and tenofovir), except for one, the synthesis of which remains unsuccessful. The prepared compounds exhibit single-digit micromolar to low nanomolar potency against HIV-1. The most active compound (as a mixture of epimers) exhibited similar anti-HIV-1 activity to the reference ProTide prodrug, tenofovir alafenamide. Most importantly, none of the synthesized ProTides showed cytotoxicity. This was one of the key properties of the compounds from Kalčic's project that we wanted to improve. The results of this thesis represent a stepping stone to a new category of ProTide prodrugs of phosphonates. Key words: prodrugs, acyclic nucleoside phosphonates, ProTides, tenofovir, serine, TAF
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The influence of acyclic nucleotide phosphonates PMEG and PMEDAP on p38 kinase signaling in human leukemic cells
Nejedlá, Michaela ; Entlicher, Gustav (advisor) ; Slaninová, Jiřina (referee)
PMEG [9-(2-phosphonomethoxyethyl)guanine] and PMEDAP [9-phosphonomethoxy- ethyl)-2,6-diaminopurine] are acyclic nucleoside phosphonates possessing cytotoxic properties. Antiproliferative effect of PMEG was demonstrated in various tumor cell lines in vitro. PMEG also represents an active component of some experimental prodrugs with enhanced selectivity and efficacy (such as GS-9219). PMEDAP seems to have weaker effect in vitro compared to PMEG, however it exhibited pronounced antitumor effect in SD-rats with spontaneous lymphoma. Therefore it was included in the present study as well. The aim of this study was to describe the interactions of PMEG and PMEDAP with p38 MAP kinase signaling and its relationship to the apoptosis. We investigated the influence of these compounds on the expression of four genes encoding p38 MAPK isoforms and whether this change is translated into the protein. It was found that PMEG up-regulates p38β and γ mRNA in CCRF-CEM cells and p38 β and δ in HL-60 cells. The effect of PMEDAP was less pronounced than that of PMEG. However, total p38 protein level remained unaffected by PMEG and PMEDAP. Activation of p38 MAPK cascade was also measured in the cells exposed to these agents using phospho-specific antibodies. We found that neither PMEG nor PMEDAP activated p38 kinase...
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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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Acyclic nucleosides of 3-hydroxypyrazine-2-carboxamide bases
Chaloupecká, Ema ; Jindřich, Jindřich (advisor) ; Smrček, Stanislav (referee)
This thesis deals with the preparation of acyclic nucleosides and nucleoside phosphonates of compounds T-705 (6-fluoro-3-hydroxypyrazine-2-carboxamide) and T-1105 (3-hydroxypyrazine-2-carboxamide). Acyclic nucleoside phosphonates are substances that can terminate viral RNA or DNA replication, and some of them are used in the treatment of viral diseases. T-705 and T-1105 have shown activity against the influenza virus, and T-705 has already been approved for its treatment in Japan. Since both compounds mimic natural nucleobases in the body, their acyclic nucleosides and nucleoside phosphonates also have the potential to be biologically active. Methods for the synthesis of 3-fluoro-2-(phosphonomethoxy)propyl and 3-hydroxy-2-(phosphonomethoxy)propyl derivatives of T-705 and T-1105, their prodrugs containing lipophilic groups for the improvement of the pharmacokinetic properties and also their phosphonate diphosphates, suitable for the biological activity measurements, have been proposed. Some of these derivatives were subsequently prepared. Key words: acyclic nucleosides, acyclic nucleoside phosphonates, T-705, T-1105, favipiravir, antiviral activity, influenza
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The influence of acyclic nucleotide phosphonates PMEG and PMEDAP on p38 kinase signaling in human leukemic cells
Nejedlá, Michaela ; Entlicher, Gustav (advisor) ; Slaninová, Jiřina (referee)
PMEG [9-(2-phosphonomethoxyethyl)guanine] and PMEDAP [9-phosphonomethoxy- ethyl)-2,6-diaminopurine] are acyclic nucleoside phosphonates possessing cytotoxic properties. Antiproliferative effect of PMEG was demonstrated in various tumor cell lines in vitro. PMEG also represents an active component of some experimental prodrugs with enhanced selectivity and efficacy (such as GS-9219). PMEDAP seems to have weaker effect in vitro compared to PMEG, however it exhibited pronounced antitumor effect in SD-rats with spontaneous lymphoma. Therefore it was included in the present study as well. The aim of this study was to describe the interactions of PMEG and PMEDAP with p38 MAP kinase signaling and its relationship to the apoptosis. We investigated the influence of these compounds on the expression of four genes encoding p38 MAPK isoforms and whether this change is translated into the protein. It was found that PMEG up-regulates p38β and γ mRNA in CCRF-CEM cells and p38 β and δ in HL-60 cells. The effect of PMEDAP was less pronounced than that of PMEG. However, total p38 protein level remained unaffected by PMEG and PMEDAP. Activation of p38 MAPK cascade was also measured in the cells exposed to these agents using phospho-specific antibodies. We found that neither PMEG nor PMEDAP activated p38 kinase...
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Determination of enantiopurity of new types of acyclic nucleoside phosphonates by capillary electrophoresis with cyclodextrins-based chiral selectors
Šolínová, Veronika ; Kaiser, Martin Maxmilian ; Lukáč, Miloš ; Janeba, Zlatko ; Kašička, Václav
Capillary electrophoresis with neutral and cationic cyclodextrins as chiral selectors was applied for determination of enantiopurity analysis of new types of six acyclic nucleoside phosphonates, nucleotide analogues bearing ((3-hydroxypropan-2-yl)-1H-1,2,3-triazol-4-yl)phosphonic acid, 2-((diisopropoxyphosphonyl)methoxy)propanoic acid or 2 (phosphonomethoxy)propanoic acid moieties attached to adenine, guanine, 2,6-diaminopurine, uracil and 5-bromouracil nucleobases. All these compounds were found to be synthesized in pure enantiomeric forms. Employing the UV-absorption detection at 206 nm, their detection limits were in the low micromolar level.
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Tyrosine-based prodrugs of acyclic nucleoside phosphonates
Tichý, Tomáš ; Pomeisl, Karel ; Krečmerová, Marcela ; McKenna, Ch. E.
Prodrug approach based on masking of a phosphonate function by ester linkage to a tyrosine promoiety has been developed. Results demonstrate that tyrosine is a promoiety providing drug conjugates with good chemical stability, bioavailability and efficient activation to active drug species. Another properties like metabolic stability and antiviral activity can be tuned by modification of the carboxyl function of the promoiety. Phosphonate monoester prodrugs were prepared by PyBOP coupling of a protected tyrosine promoiety with suitably derivatized phosphonate function of the parent drug. Phosphonate diester prodrugs were prepared by "synthon" approach, emloying alkylation of purine nucleobase with pre-prepared PME synthons bearing two protected tyrosine promoieties.
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