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Sesquiterpenes in cancer therapy
Ambrož, Martin ; Skálová, Lenka (advisor) ; Hodek, Petr (referee) ; Souček, Pavel (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Mgr. Martin Ambrož Supervisor: Prof. RNDr. Lenka Skálová, Ph.D. Title of Doctoral Thesis: Sesquiterpenes in cancer therapy. Chemotherapy is an important part of treatment of cancer. Currently, the use of several anticancer drugs in combination seem to be more advantageous. This combination therapy prevents drug resistance development and make it possible to lower the doses of single drugs, which prevents the side effects in non-cancerous tissues. Large pool of potential drugs for combination therapy can be found among natural compounds, for example among sesquiterpenes. Our project was designed to evaluate the effect of Myrica rubra essential oil (MEO) and selected sesquiterpenes in combination cancer therapy. We focused on the potential ability of MEO and sesquiterpenes to increase the antiproliferative efficacy of classical cytostatics (doxorubicin, oxaliplatin, 5-fluorouracil). In addition, we also studied the possible interaction of sesquiterpenes with other drugs in combination therapy, which could be caused by sesquiterpene-mediated modulation of drug-metabolizing enzymes. For these purposes, a panel of various cell lines (both cancerous and non-cancerous) and primary culture of hepatocytes...
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Drug Interactions with Nuclear Receptors in the Regulation of Drug Metabolizing Enzymes and Drug Transporters
Nová, Alice ; Pávek, Petr (advisor) ; Skálová, Lenka (referee) ; Souček, Pavel (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department: Pharmacology and Toxicology Candidate: Alice Nová, MSc Supervisor: Assoc. Prof. PharmDr. Petr pávek, Ph.D. Title of Doctoral Thesis: Drug interactions with nuclear receptors in the regulation of drug metabolizing enzymes and drug transporters Drug metabolizing enzymes play a key role in metabolism, elimination and detoxification of xenobiotics exposed to the body and protect the organism from potentially harmful effect of xenobiotics as well as certain endobiotics. Phase I and II drug metabolizing enzymes and drug transporters are expressed in most tissues in basal level or may be induced after exposure to xenobiotics. Affecting of expression or activity of drug metabolizing enzymes and drug transporters may result in unpredictable tissue and systemic drug distribution, to side effects or therapy failure. Up-regulation of drug metabolizing enzymes and drug transporters mediates mainly orphan nuclear receptors - pregnane X receptor (PXR) and constitutive androstane receptor (CAR) and transcription factor aryl hydrocarbon receptor (AhR). This doctoral thesis presents the results of three studies dealing with the gene regulation of drug metabolizing enzymes and drug transporters via nuclear receptors. In the first part of...
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Site-directed mutagenesis of human cytochromes P450 family 1 and their interacting partners
Milichovský, Jan ; Martínek, Václav (advisor) ; Befekadu, Asfaw (referee) ; Souček, Pavel (referee)
Cytochromes P450 represent a large group of proteins metabolizing variety of substrates. Many of them are responsible for metabolism of xenobiotics including drugs and chemical carcinogens. Heme-protein cytochrome b5 is a single-electron donor cooperating with a NADPH:cytochrome P450 reductase and NADH:cytochrome b5 reductase 3 enzyme. Cytochrome b5 can affect the xenobiotic metabolism via modulation of the cytochromes P450 activity. One of the goals of the Ph.D. thesis was to utilize site directed mutagenesis of cytochromes P450 family 1 to elucidate the mechanism of their nitroreductase activity. Another aim was to study the interaction between cytochrome b5 and cytochromes P450 of the 1A subfamily using site directed mutagenesis on presumed protein-protein contact interface. Another goal was to utilize the combination of theoretical and experimental approaches to explain variance in the reduction state of several human cytochromes P450 heterologously expressed in intact bacterial cells. The results found in the thesis show that nitroreductase activity of CYP1A1, CYP1A2 and CYP1B1 is mediated by the presence of a particular hydroxyl group in their active centre. Single mutation introducing a hydroxyl group to the specific part of CYP1B1 active site to the active site turned on its artificial...
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Modulation of metabolic activation of ellipticine by components of the mixed function monooxygenase system
Mrázová, Barbora ; Stiborová, Marie (advisor) ; Tichá, Marie (referee) ; Souček, Pavel (referee)
CHARLES UNIVERSITY IN PRAGUE FACULTY OF SCIENCE, DEPARTMENT OF BIOCHEMISTRY Modulation of metabolic activation of ellipticine by components of the mixed function monooxygenase system Summary of PhD Thesis RNDr. Barbora Mrázová Supervisor: Prof. RNDr. Marie Stiborová, DrSc. Praha 2010 RNDr. Barbora Mrázová Introduction 1 IINNTTRROODDUUCCTTIIOONN Ellipticine (5,11-dimethyl-6H-pyrido [4,3-b] carbazole, figure 1) and its more soluble derivates are alkaloids isolated from Apocyanaceae plants, which exhibit significant antitumor activities[15, 21, 24, 31] . Ellipticine was first isolated in 1959 from the leaves of the evergreen tree Ochrosia elliptica Labil[6] . Nevertheless, its pharmacological efficiencies (and efficiencies of some of its derivates) was found in 1967, when they were prepared by chemical syntheses[7] . Ellipticine and its more soluble derivates, 9-methoxy- ellipticine and 2-methyl-9-hydroxyellipticine in the form of acetate, have been utilized pharmacologically since 1970s. They are highly efficient against osteolytic breast cancer with metastases, acute myeloblastic leukemia, kidney sarcoma and thyroid carcinoma[1, 17, 18, 19, 23] . Ellipticines exhibit also significant anti-HIV activity because of their ability to inhibit retroviral integrase. This is the reason, why ellipticine is also...
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Combination of ellipticine chemotherapy and a5b1 integrin-targeted therapy in human glioblastoma
Martínková, Eva ; Stiborová, Marie (advisor) ; Souček, Pavel (referee) ; Eckschlager, Tomáš (referee) ; Barberi-Heyob, Muriel (referee)
Charles University Prague Faculty of Science Department of Biochemistry SUMMARY OF THE THESIS COMBINATION OF ELLIPTICINE CHEMOTHERAPY AND α5β1 INTEGRIN-TARGETED THERAPY IN HUMAN GLIOBLASTOMA by MARTÍNKOVÁ Eva Supervisors: Prof. RNDr. Marie STIBOROVÁ, DrSc. Dr. Monique DONTENWILL 2 TableofContents INTRODUCTION............................................................................................................................... 3 AIMSOFTHESTUDY ...................................................................................................................... 5 RESULTS............................................................................................................................................. 6 PARTA ................................................................................................................................................................6 PARTB.................................................................................................................................................................6 DISCUSSION....................................................................................................................................... 8...
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Histopathology and Molecular Biology of Gastrointestinal Tumours
Červenková, Lenka ; Souček, Pavel (advisor) ; Eckschlager, Tomáš (referee) ; Pivovarčíková, Kristýna (referee)
The incidence and mortality of gastrointestinal cancers are at a level that represents a serious problem. These tumours have non-specific clinical symptoms, which often results in late diagnosis. In recent years, there have been significant advances in the identification and monitoring of molecular markers of tumour changes in the diagnosis, prognosis and treatment of these diseases. However, clinical oncology still faces a shortage of such biomarkers. The aim of this study was to find new biomarkers that correlate especially with prognostic indicators and predictors of treatment response or chemoresistance in gastrointestinal tract cancers. This thesis is based on 5 studies addressing ductal adenocarcinoma of the pancreas, colorectal cancer and hepatocellular carcinoma using IHC and molecular methods. All studies were performed using archival formalin-fixed paraffin-embedded samples. The main results include immunohistochemical evidence of protein expression of MRP2, SLC22A3 and SUR1/ABCC8 transporters and its significant association with the prognosis of pancreatic adenocarcinoma. Patients without SLC22A3 protein expression in the apical membrane were found to have significantly shorter disease-free and overall survival. We found longer disease-free survival in patients with positive membrane...
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Study of the Sadova area in Brno
Souček, Petr ; Radimský, Michal (referee) ; Matuszková, Radka (advisor)
The diploma thesis focuses on the assessment of transport infrastructure in Brno, the Sadová area and presents a solving proposal for selected problems. The work is based on the site's survey and measurement, as well as data collected from all available sources. The CDV methodology on road safety inspections in accordance with EU Directive 2008/96/EC was used as a basis for processing the analysis of individual sites. A detailed risk assessment is performed at four sites in the area where problems are identified and a proposal for their solution is given. In relation to public transport, the work also evaluates the individual bus stops, identifies major drawbacks, and contains suggestions for a comprehensive solution of bus stops emplacement. The proposal also includes suitable places for the implementation of pedestrian crossings on Kociánka.
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Dynamic Behaviours of the C Axis Drive for Multifunction Cutting Center
Křepela, Jan ; Marek, Jiří (referee) ; Souček, Pavel (referee) ; Brychta, Zdeněk (referee) ; Singule, Vladislav (advisor)
his Disertation thesis involves the creating of the simulation model of the C axis drive over mentioned machine and them verification on the prototype of this machine. C axis is controlled with position feedback. Simulation model was created before the realisation of the machine prototype for the preliminary identification of the dynamic behaviours in the working cycles and them opportunity of the realisation this conception. C axis is constructed with worm gear and is controlled with help of Master-Slave drive. This torque drive eliminates the production backlash in the worm gear. The multifunction turning center, where is used this C axis, is determinate for heavy duty roughing cutting of the forged peaces, where is problem with dynamic stability of the cutting process. Simulation model includes the problems with multi-body mass system, friction on the worm gear, self locking, damping on the worm gear and the optimization of the parametrs for many regulators. Simulation model was verified on the prototype of the machine. Achieved results bring the new knowledge, which are used for simulation complicated machine nodes and this knowledge is used for research and developing of the similar mechatronics system.
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Development of methodology for detection of circulating nucleic acids in cancer patients
Kobrle, Lukáš ; Souček, Pavel (advisor) ; Valihrach, Lukáš (referee)
Cancer diagnosis represents one of the most common causes of death in the world, also linked to an ever-increasing incidence of even some more difficult-to-treat variants. One of the most common types of cancer in the world is colorectal cancer, which ranks second in terms of mortality. For these reasons, various methods have been developed for an early diagnosis of the disease. One of the modern and non-invasive methods is the so-called liquid biopsy, based on the detection of circulating DNA and RNA from the blood of patients. This method is also suitable for choosing a therapy, monitoring its effectiveness, and observing the relapse of the disease. In this work, a methodical workflow was created for the monitoring of circulating nucleic acids for its further use in a cohort of patients with colorectal cancer. Various commercial kits were used for the isolation of circulating nucleic acids from plasma of healthy controls, among which the best ones, in terms of quality and quantity, was chosen. The threshold for hemolysis detection by qPCR was also determined using samples from healthy donors. Everything was then applied to RNA samples extracted from plasma of cancer patients, from whichna library of short RNA molecules was prepared for subsequent sequencing. After processing the results, the...
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Heme sensor proteins as potential biomarkers of cellular and oxidative stress processes induced by ionizing radiation
Vávra, Jakub ; Martínková, Markéta (advisor) ; Souček, Pavel (referee) ; Tichý, Aleš (referee)
[IN CZECH] Ionizing radiation is a potential inducer of the oxidative stress processes in cells. As a result, reactive radicals are formed in the intracellular space modifying the essential biomolecules. Ionizing radiation has either direct (radiation sickness) or indirect (malignant processes) effect on the organism. Therefore, a fast determination of the dose is required when suspected irradiation of the organism occurs. However, a method routinely applicable, fast enough and at the same time suitable for dose estimation based on biomarkers has not been developed so far. The aim of this thesis is to describe new properties of the selected heme sensor proteins and discuss their potential importance for the cellular adaptation to oxidative stress conditions. Specifically, the thesis is focused on two eukaryotic proteins, heme regulated inhibitor (HRI) and transcription factor p53. The study of functional regulation as well as the conformational changes of these proteins induced by heme is greatly emphasized. Besides, the optimization of the key experimental methods was conducted. Specifically, Phos-tag electrophoresis was applied for the kinetics study of HRI wild type and its Gly202Ser mutant form, which is a characteristics of lung cancer development. Unsurprisingly, for both HRI forms studied,...
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