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Study on the role of selected cytochrome P450 isoforms in cytostatic resistance at apoptosis level
Moriová, Magdalena ; Hofman, Jakub (advisor) ; Novotná, Eva (referee)
Charles University Faculty of Pharmacy in Hradci Králové Departement of Pharmacology & Toxicology Student: Magdalena Moriová Supervisor: RNDr. Jakub Hofman, Ph.D. Title of diploma thesis: Study on the role of selected cytochrome P450 isoforms in cytostatic resistance at apoptosis level Cytostatic resistance is one of the most problematic obstacles in oncological treatment. Beside pharmacodynamic mechanisms, pharmacokinetic factors play an important role in drug resistance as well. Enzymatic transformation of active substance to inactive metabolite in tumor cells probably belongs to these mechanisms, however, evidences concerning the relevance of this phenomenon are predominantly either indirect and/or affected by interference elements. Using comparative experiments with HepG2 cell lines with/without CYP3A4 overexpression, we focused on the evaluation of the role of this clinically important enzyme in the resistance against docetaxel. Methodologically, it was the assessment of apoptosis induction (activation of caspases 3/7, 8 and 9) using commercial luminescent kits. Our results suggest significant participation of CYP3A4 enzyme on the reduction of docetaxel anticancer efficacy after 48 h from treatment, whereas this effect was not recorded in earlier intervals. These findings perfectly correlate...
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Use of fluorescence methods for the study of protein interactions
Johaníková, Klára ; Bezděková,, Jaroslava (referee) ; Pavelicová, Kristýna (advisor)
The diploma thesis "Use of fluorescence methods for the study of protein interactions" is focused on the use of fluorescence methods for the study of protein interactions using electromigration methods and Förster resonance energy transfer (FRET). The aim of this work was to create a bioconjugate of metallothionein (MT) protein with quantum dots (QDs) and commercial dyes. FRET was subsequently studied between these conjugates. QDs were synthesized under UV light and conjugation with MT was performed via a carbodiimide zerolengthcrooss-linker (EDC / sulfo-NHS), which serves to activate carboxyl groups and allows bioconjugation of the ligand by covalent bonding. Due to the high proportion of cysteines in MT, this protein has a very high affinity for metals. It is also involved in scavenging free radicals and there are studies that show that MT is overexpressed in cancer cells. Attention was also paid to the study of MT dimerization, which leads to an understanding of oxidative dimerization of MT and thus can contribute to understanding the formation of free radicals in the body and to deepen the knowledge about neurodegenerative disorders such as Parkinson's or Alzheimer's disease or amyotrophic lateral sclerosis. The formation of the MT dimer was confirmed by energy transfer between the donor (QDs) and the acceptor (commercial dye-cyanine) through the physical phenomenon of FRET and MALDI-TOF-MS.
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The significance of platinum in the environment
Brestovská, Marta ; Holubová, Zuzana (referee) ; Sommer, Lumír (advisor)
This bachelor thesis deals with monitoring of individual platinum compounds in the environment (water, soil, air) and is also reviewing methods used for their analysis. The no less important determination of platinum in tissues and body fluids, when the platinum is used in form of a cisplatin as an anticancer drug, is also mentioned. Afterwards the anticancer drug itself or its derivatives and metabolites on the base of platinum complexes in clinical sample (urine, plasma) are determined. Mainly the ICP-AES, ICP-MS, ETA-AAS, HPLC methods are highlighted from the methods used for the determination of Pt. The on-line connection between some of these methods such as ICP-MS with HPLC or ICP-AES with HPLC seems to be useful. The spectrofotometric methods using organic and inorganic agents can also be used for the determination of platinum. These methods are mentioned here rather marginally, just for completeness. In terms of practical use they are difficultly reproducible and not very sensitive. The hydrolysis of platinum metals to form insoluble hydrated oxides makes also the determination difficult.
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Therapy of difficult-to-treat pancreatic tumours with new metallopharmaceuticals and their mechanism of action
Švitelová, Marie ; Prachařová,, Jitka (referee) ; PhD, Vojtěch Novohradský, (advisor)
The issue of tumor diseases and anticancer therapy is extremely current nowadays, in which is all acquired knowledge a great benefit to society. Despite the extraordinary development of this field, the available treatment approaches for cancer are still very limited by a number of factors, that motivate research teams to find targeted, effective therapy. The main aim of this bachelor work was to test an in vitro cytotoxicity of newly synthesized enantiomers [Pt(OXA)(1,2-DACHEX)] derived from oxaliplatin. During the testing of the above mentioned substances we focused on the analysis of the antiproliferative action of these complexes and their mechanism of action. The results of the work show that the R,R-enantiomer has high therapeutic effects when it is applied to the PSN1 pancreatic adenocarcinoma cells. The experiments also proved that the mechanism of action of this complex in pancreatic cancer cells involves influencing the lipogenesis pathway, namely inhibition of de novo lipid synthesis. The antitumor action aimed at influencing the metabolism represents a new mechanism of action that has not yet been considered for clinically used antitumor platinum drugs. The obtained information suggests that R,R-enantiomer could represent a future promising cytostatic that would cause fewer adverse effects on the patient.
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Study and mechanism of new-generation heterocyclic cytostatic with an emphasis on IL-6 andIL-8
Talianová, Veronika ; Jakubek, Milan (advisor) ; Mikula, Ivan (referee) ; Babula, Petr (referee)
EN The purpose of this study was to screen for novel synthetic inhibitors of interleukins IL-6 and IL-8, which are promising targets for cancer therapy. Small molecules are an attractive approach to inhibit these signalling pathways, which are known to be important for tumour cell growth and survival. The study aimed to discover new inhibitors of IL-6 and IL-8 signalling pathways by screening a small library of chemical derivatives, including π-expanded naphthalimide derivatives, Tröger's base, and pentamethinium derivatives. The structures of the compounds were designed based on in silico studies focusing on blocking the IL-6 and IL-8 signalling pathways. The study involved the use of various analytical and bioanalytical methods for the in vitro analysis of new potential anticancer drugs. The compounds were tested for cytotoxicity and their effect on cell growth using an MTT proliferation assay. Their antitumor activity was observed in NU/NU mice. The invasiveness of the cells was monitored using the wound healing assay. Live cell imaging was used to detect the distribution of the substances at the intracellular level. Other methods such as ELISA, microscale thermophoresis, and in silico predictions have been used to determine the selectivity of inhibitors of the signalling pathways IL-6 and IL-8....
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Use of fluorescence methods for the study of protein interactions
Johaníková, Klára ; Bezděková,, Jaroslava (referee) ; Pavelicová, Kristýna (advisor)
The diploma thesis "Use of fluorescence methods for the study of protein interactions" is focused on the use of fluorescence methods for the study of protein interactions using electromigration methods and Förster resonance energy transfer (FRET). The aim of this work was to create a bioconjugate of metallothionein (MT) protein with quantum dots (QDs) and commercial dyes. FRET was subsequently studied between these conjugates. QDs were synthesized under UV light and conjugation with MT was performed via a carbodiimide zerolengthcrooss-linker (EDC / sulfo-NHS), which serves to activate carboxyl groups and allows bioconjugation of the ligand by covalent bonding. Due to the high proportion of cysteines in MT, this protein has a very high affinity for metals. It is also involved in scavenging free radicals and there are studies that show that MT is overexpressed in cancer cells. Attention was also paid to the study of MT dimerization, which leads to an understanding of oxidative dimerization of MT and thus can contribute to understanding the formation of free radicals in the body and to deepen the knowledge about neurodegenerative disorders such as Parkinson's or Alzheimer's disease or amyotrophic lateral sclerosis. The formation of the MT dimer was confirmed by energy transfer between the donor (QDs) and the acceptor (commercial dye-cyanine) through the physical phenomenon of FRET and MALDI-TOF-MS.
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Effect of selected cytostatics for the treatment of leukemia on the activity of human carbonyl reducing enzymes
Šmídlová, Monika ; Novotná, Eva (advisor) ; Wsól, Vladimír (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical sciences Candidate: Bc. Monika Šmídlová Supervisor: RNDr. Eva Novotná, Ph.D. Title of diploma thesis: Effect of selected cytostatics for the treatment of leukemia on the activity of human carbonyl reducing enzymes Key words: reductases, leukemia, cytostatics, inhibition Anthracycline antibiotics, especially daunorubicin, are widely used for the treatment of acute myeloid leukemia (AML) and acute lymphocytic leukemia (ALL). Although the efficacy of these drugs is high, treatment is still limited due to cardiotoxicity and tumor cell resistance to anthracyclines. Mechanisms that contribute to the formation of anthracycline resistance include metabolic biotransformation (reduction) to less efficient secondary alcohols. The reduction is calatyzed by carbonyl reducing enzymes belonging to aldo-keto reductase (AKR) and short chain dehydrogenase/reductase (SDR) superfamilies. The discovery of AKR and SDR inhibitors could help to overcome anthracycline resistance and also reduce cardiotoxicity caused by these drugs. The aim of the diploma thesis was to find out whether all-trans-retinoic acid, cyclophosphamide, cytarabine, cladribine and prednisolone are able to inhibit anthracycline reductases AKR1A1, AKR1B10, AKR1C3, AKR7A2...
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Depletion of Treg cells for potentiation of cancer treatment with HPMA copolymer-bound cytostatic drug conjugates"
Dvořáková, Barbora ; Kovář, Marek (advisor) ; Reiniš, Milan (referee)
Tumor diseases are severe problem worldwide with increasing number of patients suffering from various types of malignancies. Many of approved therapeutics cause serious side toxicities. Therefore, there are intensive efforts to improve cancer treatment protocols. The aim of this study was to deplete regulatory T (Treg) cells without affecting other immunocompetent cells playing a positive role in tumor eradication. Treg cells were reported to hamper anti-tumor immunity and promote tumor growth and survival. Thus, their selective elimination could lead to induction of anti-tumor responses and tumor rejection if combined with chemotherapy with selected N-(2- hydroxypropyl)methacrylamide (HPMA) copolymer-bound drug conjugates. Original approach was to deplete of Treg cells without the use of anti-CD25 mAb that has been widely exploited for Treg cell elimination; however, its long-term persistence in circulation together with inhibitory effect on activated effector cells (CD25+ ) are its main disadvantages. Thus, Treg cells were sensitized to cell cycle-specific cytostatic drugs via application of IL-2/anti-IL-2 JES6.1 mAb immunocomplexes that induce vigorous selective proliferation of this cell population. Subsequent application of cell cycle-specific cytostatics showed steep decrease of Treg cell...
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Nanotoxikologie pokročilých materiálů =: Nanotoxicology of advanced materials /
Michálková, Hana
Nowadays, a wide range of nanoparticles is used in different fields of human acti-vities. These relatively are well-researched. However, with the development of new nanomaterials, new possibilities for their use both in nanomedicine and in other fields are possible. This phenomenon is accompanied by potential toxic risks of these mate-rials. The presented thesis deals with the extensive topic of cytotoxicity of both nanoma-terials and their modification for better binding of cytostatics to increase their effi-cacy. Experimental work is focused on the interaction of nanomaterials with the cells and their environment, the determination of the toxicity of nanomaterials and their potential use in nanomedicine. The main aim of my thesis was to prepare a comprehensive summary about nano-materials, their physico-chemical properties, to modify nanoparticles surface to provi-de an entity for targeted drug delivery and to test cytotoxicity on selected cell lines.
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