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The use of selected inhibitors to overcome anthracycline resistance in breast cancer therapy
Grofková, Kateřina ; Laštovičková, Lenka (advisor) ; Svrčková, Katarína (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Kateřina Grofková Supervisor: Mgr. Lenka Laštovičková, Ph.D. Title of diploma thesis: The use of selected inhibitors to overcome anthracycline resistance in breast cancer therapy The breast cancer is a heterogenic disease and a worldwide problem mainly for women. It has been treated with anthracycline antibiotic doxorubicine for decades. The greatest problem of this treatment is a cardiotoxicity usually associated with a metabolite of doxorubicine - doxorubicinole. This metabolite is produced in an enzymatic reaction catalysed by carbonyl reductases. A multidrug anthracycline resistance, caused by induced expression of carbonyl reductases and efflux ABC transporters, is another serious problem. Therefore, new inhibitors of carbonyl reductases and efflux ABC transporters have been searched for, to overcome multidrug resistance and reduce doxorubicine cardiotoxicity. The inhibitory effect of several proteinkinase inhibitors (afatinib, erlotinib hydrochloride, dacomitinib and tipifarnib) on carbonyl reductase activity (AKR1A1, 1B1, 1B10, 1C3 and CBR1) was investigated in the thesis. The greatest inhibitory potential was found for inhibitor tipifarnib and enzyme AKR1C3. The inhibition of 10μM...
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Plasmid constuction for expression of reductase from Haemonchus contortus
Klouček, David ; Matoušková, Petra (advisor) ; Laštovičková, Lenka (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: David Klouček Supervisor: doc. Ing. Petra Matoušková, Ph.D. Title of diploma thesis: Plasmid constuction for expression of reductase from Haemonchus contortus Haemonchus contortus is the most important pathogenic nematode of small ruminants, mostly sheep and goats. It causes haemonchosis leading to massive deaths of infected animals. Pharmacotherapy by approved anthelmintics is the main solution to the disease. Currently, the growing resistance to available antihelmintics is still bigger and bigger problem, leading to economical and ecological issues. There are many causes of resistance. Among the non-target site mechanisms of resistance belongs the increased activity of enzymes from the first biotransformation phase mediating carbonyl-reduction; specifically aldo/ketoreductases and dehydrogenases/reductases. Increased activity of these enzymes leads to faster drug metabolism, formation of inactive metabolites and therapy failure. The aim of the thesis was to prepare a plasmid for the expression of selected reductase, which can be used for further study of the activity and possibly design of new targeted drugs. Method, which were used during my experimental work, involved isolation of RNA, reverse...
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The potential role of inhibitor tipifarnib in the treatment of acute myeloid leukemia
Hrbáčová, Patricie ; Laštovičková, Lenka (advisor) ; Jirkovský, Eduard (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Patricie Hrbáčová Supervisor: Mgr. Lenka Laštovičková, Ph.D. Title of diploma thesis: The potential role of inhibitor tipifarnib in the treatment of acute myeloid leukaemia Anthracycline antibiotics, such as daunorubicin, are the gold standard in a treatment of acute myeloid leukaemia. However, they are not fully specific and besides cancer cells, they can damage also other structures, especially heart muscle cells. Moreover, the anthracycline resistance can develop. The daunorubicin is metabolized by carbonyl reducing enzymes to daunorubicinol, which has significantly worse therapeutic effect. This reaction can be prevented by using a specific inhibitor. In the thesis, the inhibitor tipifarnib and its effect on reactions catalysed either by aldo- ketoreductases (AKR1A1, 1B1, 1B10 and 1C3) or short-chain dehydrogenases/reductases (CBR1) were tested. The strongest inhibition was observed for AKR1C3. When using 10 μM tipifarnib in reaction, an inhibition of 88.0% was observed, and 50 μM inhibitor resulted in inhibition by 92.9%. That is why kinetic parameters were specified only for AKR1C3 in all other experiments. The IC50 value was experimentally determined to 0.51 ± 0.03 μM and the inhibition...
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Preparation of STAT3-knockout glioblastoma multiforme cell line using CRISPR-Cas9 editing
Papinčáková, Dominika ; Skálová, Lenka (advisor) ; Laštovičková, Lenka (referee)
Charles University Fakulty of Pharmacy in Hradec Králove Department of Biochemical Sciences Candidate: Bc. Dominika Papinčáková Supervisor: prof. RNDr. Lenka Skálová, Ph.D. Title of diploma thesis: Preparation of STAT3-knockout glioblastoma multiforme cell line using CRISPR-Cas9 editing Glioblastoma multiforme (GBM) is one of the most common primary brain tumours in adults and one of the most fatal cancers in humans A key oncogene in GBM development and progression is the signal transducer and activator of transcription 3 (STAT3). It is one of the mediators of tumorigenesis, plays a strong pro-survival role in GBM, and is permanently activated in GBM, leading to increased cell proliferation, invasiveness, and angiogenesis. The aim of this work was to establish a GBM STAT3-knockout (STAT3-KO) cell line. The U87MG cell line and the CRISPR-Cas9 gene knockout method were used for this purpose. In a first step, we investigated the possibility of silencing STAT3 expression by siRNA and the suitability of this cell type to pinpoint the mechanism for CRISPR gene editing into the cell. In the second step, we transfected the cells using the STAT3 gene knockout kit (CRISPR) and tested the knockout success. In the established STAT3-KO GBM cell line, we confirmed STAT3-KO at the DNA, RNA and protein levels. We...
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Preparation of STAT3-knockout glioblastoma multiforme cell line using CRISPR-Cas9 editing
Papinčáková, Dominika ; Skálová, Lenka (advisor) ; Laštovičková, Lenka (referee)
Charles University Fakulty of Pharmacy in Hradec Králove Department of Biochemical Sciences Candidate: Bc. Dominika Papinčáková Supervisor: prof. RNDr. Lenka Skálová, Ph.D. Title of diploma thesis: Preparation of STAT3-knockout glioblastoma multiforme cell line using CRISPR-Cas9 editing Glioblastoma multiforme (GBM) is one of the most common primary brain tumours in adults and one of the most fatal cancers in humans A key oncogene in GBM development and progression is the signal transducer and activator of transcription 3 (STAT3). It is one of the mediators of tumorigenesis, plays a strong pro-survival role in GBM, and is permanently activated in GBM, leading to increased cell proliferation, invasiveness, and angiogenesis. The aim of this work was to establish a GBM STAT3-knockout (STAT3-KO) cell line. The U87MG cell line and the CRISPR-Cas9 gene knockout method were used for this purpose. In a first step, we investigated the possibility of silencing STAT3 expression by siRNA and the suitability of this cell type to pinpoint the mechanism for CRISPR gene editing into the cell. In the second step, we transfected the cells using the STAT3 gene knockout kit (CRISPR) and tested the knockout success. In the established STAT3-KO GBM cell line, we confirmed STAT3-KO at the DNA, RNA and protein levels. We...
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Determination of ovarian function parameters
Jará, Barbora ; Skálová, Lenka (advisor) ; Laštovičková, Lenka (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Thesis title: Determination of ovarian function parameters Supervisor: prof. RNDr. Lenka Skálová, Ph. D. Consultant: RNDr. Michala Vosátková, MBA Created by: Barbora Jará The levels of hormones that control of ovaria are important markers of development and health status of the female organism. Laboratory examination and correct interpretation of the results are important for proper diagnostic procedures. Among the hormones of ovarian function, the level of estradiol, progesterone, luteinizing and follicle-stimulating hormones is most often tested. Separation and immunochemical methods are used. For common clinical practice, immunochemical methods are particularly suitable due to their availability, relative accuracy, unpretentiousness, and speed of processing. The impossibility of standardizing individual methodological approaches represents the shortcoming of these methods. A series of examinations and interpretation of results can be performed using immunochemical methods only within one workplace. Keywords: estradiol, immunochemical methods, ECLIA, CMIA, RIA
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Liability for damage under civil law and under commercial law
Laštovičková, Lola ; Švestka, Jiří (advisor) ; Salač, Josef (referee)
The purpose of my thesis is to compare the conformity and diversity between legal provisions of liability for damages in czech civil and commercial law. The thesis is divided into four chapters and its core lies in Chapter Three. First chapter subdivided into three parts deals with characteristics of liability for damages in general, further contains the location of this liability among other types of liability and the last part concerns with the functions of it. Chapter Two is concentrated on the Principles of European Tort Law. Part One describes the europeisation of tort law, which is mostly connected with spontaneous projects of soft law. In Part Two there is reffered to general conception of the Principles and some particular provisions in detail. I have endeavoured to draw a comparison between almost every field of liability for damages in czech civil and commercial law and these Principles. As I have mentioned the main part of my thesis rests in Chapter Three. The chapter consists of eight parts. At first I have explained the relationship between Czech Civil Code and Commercial Code regarding liability for damages. Legal provisions in both codes are sometimes duplicate, on the other hand in some parts different but often without any relevant reason. Subsequent part highlights the importance...
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Influence of albendazole on the antioxidant system of alfalfa in real conditions
Lisoňková, Monika ; Szotáková, Barbora (advisor) ; Laštovičková, Lenka (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Monika Lisoňková Supervisor: prof. Ing. Barbora Szotáková, PhD. Title of diploma thesis: The effect of albendazole on the antioxidant system of alfalfa in real conditions Anthelmintics are drugs used to treat diseases caused by parasitics worms. In the Czech republic they are mainly used in veterinary medicine. Benzimidazole anthelmintic albendazole is used in cattle and sheep, during its biotransformation metabolites are formed, which are excreted together with the parent compound in faeces. Application of faeces from treated cattle to the field may affect non-target organisms, occurring in the field. An example of such organisms are agricultural crops, including alfalfa (Medicago sativa). Plants do not have the ability to eliminate the absorbed drugs and its metabolites, therefore they are deposited in plants and can affect the antioxidant system of plants. The aim of this study was to monitor the effect of albendazole on the antioxidant system of alfalfa (M. sativa) in real conditions. Plants were affected by faeces of sheep treated with albendazole. Subcellular fractions were prepared from harvested plants in the years 2019 and 2020. The concentrations of photosynthetic pigments...
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