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Inhibition of TET-1 protein by iron chelators
Antonyová, Veronika ; Jakubek, Milan (advisor) ; Mikula, Ivan (referee) ; Gumulec, Jaromír (referee)
EN The objective of this work was to study novel synthetic iron chelators and to study their potential use in inhibition of TET1 (ten-eleven translocation methylcytosine dioxygenase 1) protein. Epigenetic mechanisms, such as hydroxymethylation of DNA, are promising target in many serious pathologies, including oncological disorders. In the presented study, we intended to discover novel inhibitors by screening small libraries of heterocyclic molecules, such as pyrrolo[3,2-b]pyrrole derivatives with hydrazide (compound 1) or hydrazone (compound 2-6) iron-binding group and hydrazone-based iron chelators (compound 7-10). Within the scope of this study, we used various analytical and biochemical techniques for the purpose of characterization of novel cancer therapies based on TET1 protein inhibition. The absorbance and the complexation of tested compounds with Fe(II) ions was studied by UV-Vis spectroscopy. Inhibition of TET1 protein was studied by fluorometric assay based on ELISA and further supported by microscale thermoporesis and in silico docking. The cytotoxicity of compounds on cancer cell lines was measured by MTT assay and intracellular distribution was determined by live-cell imaging. This study presents a biochemical analysis of potential TET1 protein inhibitors and brings significant...
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HSP90 as a target for antitumor therapy
Drápalová, Kateřina ; Zatloukalová, Pavlína (advisor) ; Dibus, Nikol (referee)
Stability and correct tertiary structure of proteins are necessary for maintaining cellular homeostasis. The cell uses molecular chaperones, including Hsp90, to achieve this balance. Hsp90 is an essential protein for healthy and cancer cells. Overexpression of this chaperone is noticeable in many cancers. This thesis summarizes current understandings of the Hsp90 protein and its role in carcinogenesis. Hsp90 became a target of anticancer therapy in the 90s. Inhibition of this chaperone, though successful in many studies, has not yet reached the wanted results in clinical practice. Current failures are mostly caused by high hepatotoxicity and other side effects accompanying the therapy, or the inhibition having insufficient antitumor effects. Nevertheless, the inhibition of the Hsp90 protein represents an interesting approach in antitumor therapy. New inhibitors are constantly being developed and tested in monotherapy or in combination therapy, which demonstrates significantly higher efficacy and, thanks to the synergistic effect, enables the application of a lower concentration of therapeutics.
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Polymer-based therapeutics for immunooncotherapy
Kashmel, Pavel ; Etrych, Tomáš (advisor) ; Bárta, František (referee)
This master thesis describes the synthesis, physico-chemical characterization and preliminary biological testing of water-soluble polymer conjugates of the model drug ZM241385. This drug was first derivatized and then connected to a polymer carrier in order to prepare a system suitable for targeted drug transport to tumor tissues. Improved pharmacokinetic parameters of the prepared polymeric systems carrying ZM241385 should be the basis for increased therapeutic activity of the polymeric nanosystem in tumor immunotherapy. Polymeric precursors were synthesized by controlled RAFT polymerization in order to prepare highly defined carrier systems. In the framework of this master thesis, two derivatives of the selected drug were prepared, differing in the carboxylic acid used and the mode of their binding to the polymeric carrier. For derivatization, 4-(2-oxopropyl)benzoic acid was used, in which case the prepared derivative was bound to the carrier via a hydrazone bond. This bond is pH sensitive and shows high stability at pH 7.4 (corresponds to the pH of the blood stream), and at a slightly acidic pH 5.5 (corresponds to the microenvironment of the tumor, or endosomes of tumor cells), its rapid hydrolysis occurs. For the second derivative, 5-azidopentanoic acid was used. An uncatalysed click reaction...
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Metabolic alterations in cancer cells and their implications in the therapy of acute leukemias
Harárová, Alžbeta ; Starková, Júlia (advisor) ; Mrvová, Silvia (referee)
Cancer metabolism differs from that of the healthy cells in several aspects. Aerobic glycolysis (e.g. converting pyruvate to lactate under normoxic conditions) was the first described metabolic alteration of cancer cells. Metabolic alterations have since been described in the tricarboxylic acid cycle, oxidative phosphorylation, in the metabolism of amino acids (especially glutamine, asparagine and serine) and also in the metabolism of fatty acids and cholesterol. The common feature of these changes is the tendency to prefer anabolic pathways, thus enabling fast proliferation of cancer cells. The study of cancer metabolism is particularly important in the case of cancer cells that show resistance to treatment, as their aberrant metabolism is not only a potential diagnostic marker but also a potential therapeutic target. The majority of metabolic alterations have been described for the first time in solid tumors, whereas only recently has the metabolism of acute leukamias gained more attention. Asparaginase is an example of a chemotherapeutic agent that targets a metabolic alteration of leukemic cells. Distinct metabolic profile is also associated with the glucocorticoid resistance. Detailled study of the metabolic alterations of leukemic cells has elucitated the mechanisms of the asparaginase and...
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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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Metabolic alterations in cancer cells and their implications in the therapy of acute leukemias
Harárová, Alžbeta ; Starková, Júlia (advisor) ; Mrvová, Silvia (referee)
Cancer metabolism differs from that of the healthy cells in several aspects. Aerobic glycolysis (e.g. converting pyruvate to lactate under normoxic conditions) was the first described metabolic alteration of cancer cells. Metabolic alterations have since been described in the tricarboxylic acid cycle, oxidative phosphorylation, in the metabolism of amino acids (especially glutamine, asparagine and serine) and also in the metabolism of fatty acids and cholesterol. The common feature of these changes is the tendency to prefer anabolic pathways, thus enabling fast proliferation of cancer cells. The study of cancer metabolism is particularly important in the case of cancer cells that show resistance to treatment, as their aberrant metabolism is not only a potential diagnostic marker but also a potential therapeutic target. The majority of metabolic alterations have been described for the first time in solid tumors, whereas only recently has the metabolism of acute leukamias gained more attention. Asparaginase is an example of a chemotherapeutic agent that targets a metabolic alteration of leukemic cells. Distinct metabolic profile is also associated with the glucocorticoid resistance. Detailled study of the metabolic alterations of leukemic cells has elucitated the mechanisms of the asparaginase and...
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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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