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Investigating the role of zinc transporter ZIP 6 and STAT3 in mitosis
Burgetová, Lenka ; Čečková, Martina (advisor) ; Štaud, František (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Lenka Burgetová Supervisor: PharmDr. Martina Čečková PhD. Specialized supervisor: Dr. Kathryn Taylor PhD. Title of diploma thesis: Investigating the role of zinc transporter ZIP 6 and STAT3 in mitosis It has been shown that STAT3 (signal transducer and activator of transcription 3) plays a role in the development of cancer. ZIP6 is the downstream target of this transcription factor. Previous research has focused mainly on the activation of STAT3 by tyrosine phosphorylation, while the effect of phosphorylation at a second site, serine 727, remained relatively uninvestigated. In this study, it is proposed that serine-phosphorylated STAT3 is activated throughout mitosis in tamoxifen-resistant (TamR) breast cancer cells and that zinc transporter ZIP6 and serine-phosphorylated STAT3 are involved in a zinc-mediated mitotic mechanism. After using nocodazole to induce mitotic arrest, the expression of tyrosine- phosphorylated STAT3 protein was observed to be reduced while the expression of serine- phosphorylated STAT3 was increased. Zinc supplementation after nocodazole treatment appeared to push cells through mitotic-arrest and cause proteolytic cleavage of STAT3 suggesting a novel...
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MRP transporters in the placenta
Dvořáková, Marie ; Čečková, Martina (advisor) ; Pávek, Petr (referee)
The MRP mebrane proteins, which belong to the ABC transporter family, comprise currently 9 members. The MRP transport proteins are expressed in various tissues of the organism, including placenta. The major physiological role of the multidrug transporters is the transport of many endogenous and exogenous compounds, including drugs, across the cell membrane. This thesis summarizes up to date information concerning expression and function MRP transporters in placenta a and in other tisssues in organism. Only five MRP transporters have been detected in placenta, namely MRP1, MRP2, MRP3, MRP5 a MRP8. Expression of all this proteins in placenta changes with progress of the pregnancy. MRP transporters help to protect fetus from potentially toxic substances, on the other hand some of them can facilitate the passage of substances across placenta. Some MRPs possess specific physiological functions in placenta. For example, MRP1 influences apoptosis, MRP5 participates in NO-dependent vasodilatation in fetal vessels.
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Interactions of selected anticancer drugs of the MAPK/ERK signaling pathway inhibitors group with the ABC drug transporters
Slatinský, Lukáš ; Čečková, Martina (advisor) ; Hofman, Jakub (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Lukáš Slatinský Supervisor: Assoc. prof. PharmDr. Martina Čečková, Ph.D. Title of diploma thesis: Interactions of selected anticancer drugs of the MAPK/ERK signaling pathway inhibitors group with the ABC drug transporters ABCB1 (Pgp, P-glycoprotein) and ABCG2 (BCRP, breast cancer resistance protein) are members of a transmembrane efflux ATP dependent transporter family, so called ATP-binding cassettes (ABC). Physiologicaly they are expressed in the cellular membrane and protect body tissues against potentially toxic xenobiotics including drugs. They represent also one of the tumor defense mechanisms when being able to efflux a wide variety of cytotoxic drugs out of the cancer cells leading to treatment failure. BRAF protein plays an important regulatory and signal role in MAPK/ERK pathway affecting cell division, differentiation and secretion. Mutations of BRAF lead to overactivity in MAPK/ERK pathway in many cancer cells and can be therefore targeted by anticancer therapy. Cobimetinib and dabrafenib are relatively new anticancer therapeutics inhibiting the signal pathway mentioned above and they are used in treatment of melanoma carrying the BRAF mutation. The aims of this project were to...
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The effect of lipid signaling pathway interference on sorafenib cytotoxic efficacy and function of efflux transporters in mouse hepatocellular carcinoma cells
Sagandykova, Aigul ; Čečková, Martina (advisor) ; Novotná, Eva (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Aigul Sagandykova Supervisor: PharmDr. Martina Čečková, Ph.D. Consultant: Dr. Mikko Gynther Ph.D. Title of diploma thesis: The effect of lipid signalling pathway interference on sorafenib cytotoxic efficacy and function of efflux transporters in mouse hepatocellular carcinoma cells. Nowadays cancer remains one of the most challenging health issues worldwide. Chemotherapy represents one of the essential approaches in the treatment of malignant diseases. However, multidrug resistance (MDR), a multifactorial phenomenon described as a loss of sensitivity of cancer cells to several diverse chemotherapeutic agents at the same time, often compromises the therapy outcomes. A well-known cause of MDR is an increased expression or/and an enhanced activity of efflux drug transporters of ATP binding cassette (ABC) superfamily, which has been found in many types of cancer. In the last decade, an expanding body of literature suggested a new hallmark of cancer cells - inflammation. An inflammatory microenvironment potentiates tumorigenesis and upregulation of transporters. Moreover, several observations show that ABC transporters mediate the transport of some signalling lipids. This new insight provided...
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Interaction of selected anti-HCV drugs with placental OCTN2 transport protein
Machalová, Vanda ; Čečková, Martina (advisor) ; Smutná, Lucie (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Mgr. Vanda Machalová Supervisor: doc. PharmDr. Martina Čečková, Ph.D. Title of rigorous thesis: Interaction of selected anti-HCV drugs with placental OCTN2 transport protein. The aim of this rigorous work was to test the interaction of paritaprevir and daclatasvir with the placental OCTN2 transporter, which ensures the transport of L-carnitine as a cofactor of the fatty acid oxidation process to obtain sufficient energy for correct intra-arterial fusion growth. No drugs in this group have been approved for the treatment of chronic hepatitis C in pregnancy, therefore examining their effect on the transport of L-carnitine via OCTN2 can create certain image concerning to the fetal and maternal safety. In the first part of my thesis the transport and accumulation of radioactively labeled L- carnitine via monolayers of BeWo b30 cells in the presence of paritaprevir and daclatasvir was assayed and the concentration of radiolabeled L-carnitine inside the cell layer. In the second part of my thesis we focused on the evaluation of the presence of paritaprevir and daclatasvir on its gene expression of the OCTN1 and OCTN2 carnitine transporters (or their coding genes SLC22A4 and SLC22A5) by qRT-PCR. The...
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Monitoring of ABC-transporter expression at the transcription level
Kalmanová, Milada ; Štaud, František (advisor) ; Čečková, Martina (referee)
Human ABC (ATP-binding cassette) family of active transporters contains about 50 functionally diverse transmembrane proteins. They utilize energy from the hydrolysis of ATP and transport a variety of endogenous and exogenous compounds across the membranes against a concentration gradient. ABC transporters play important role in absorption, distribution and excretion of drugs. Some are able to confer multidrug resistance in cancer cells. Nowadays, to observe expression of ABC transporters on transcription level, it is possible to use several methods as Northern blotting, RNase protection assay, real-time RTPCR and microarray analysis. This thesis summarizes up to date information about quantification of ABC drug transporters. It was found that ABC transporters are expressed in the liver, kidney, gastrointestinal tract, blood-brain barrier, placenta and other tissues.
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The study of interaction of platinum cytostatics with BCRP drug transporter using MTT cytotoxicity test
Horká, Markéta ; Čečková, Martina (advisor) ; Štaud, František (referee)
AAbbssttrraacctt The resistance of oncological diseases to cytostatic treatment is considered as one of the most serious problems of cancer treatment. One of the mechanisms by which cancers develop resistance is the overexpression of efflux transport proteins that limit intracellular concentrations of substrate agents. This work is focused on study of the interaction between clinically used platinum cytostatics (cisplatinum, carboplatinum and oxaliplatinum) and the efflux transport protein - BCRP. The experiments were made on MDKCII (parental cell line) and MDCKII-BCRP (cells were transfected by the gene of BCRP) cell lines. The results of the experiment with oxaliplatinum did not confirm any effect of BCRP on viability of BCRP transfected cells. Cisplatinum and carboplatinum tests provided evidence that BCRP transporter plays an important role in resistance development. The cells that were transfected by the gene of BCRP showed remarkably enhanced viability than parental MDCKII cells. However, the inhibition of BCRP transporter by specific inhibitor fumitremorgin C did not affect the viability of MDCK-BCRP cells. Such results suggest that the resistance of transfected cells was developed by other mechanism. Enhanced resistance could be caused by EGFP which is used as a reporting protein in preparation of...
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Study of the Effect of EGFP on Cytotoxicity of Platinum-based Drugs
Musilová, Markéta ; Čečková, Martina (advisor) ; Štaud, František (referee)
Charles University in Prague Faculty of Pharmacy in Hradci Králové Department of pharmacology and toxicology Mgr. Markéta Musilová Master thesis Study of the effect of EGFP on cytotoxicity of platinum-based drugs Abstract The aim of the present study is to investigate possible impact of EGFP on the resistance of cancer cells to platinum-based drugs (cisplatin, carboplatin and oxaliplatin). The work follows on our previous study with BCRP efflux transporter, which suggested that higher resistance of cancer cells to platinum-based drugs is caused rather by the presence of EGFP than by the efflux transporter. To confirm this hypothesis series of cytotoxic MTT tests were performed on human Hep2 parent cell line and Hep2-EGFP cell line transfected for the expression of EGFP. Our results demonstrated higher tolerance of transfected Hep2-EGFP cells to platinum-based drugs compared with the parent Hep2 cell lines. Based on the obtained results we conclude that EGFP interferes with the toxicity of platinum-based drugs, which can cause misinterpretation of results obtained in the cytotoxicity studies.
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Study of Platinum Cytotoxic Drug Interactions with ABCG2 Membrane Transporter
Bouška, Petr ; Čečková, Martina (advisor) ; Vacková, Zuzana (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Candidate: Mgr. Petr Bouška Consultant: PharmDr. Martina Čečková, Ph.D. Title of Thesis: Study of platinum cytotoxic drug interactions with ABCG2 membrane transporter Platinum cytotoxic drugs pertain among most frequently used cytotoxic drugs. However, their usage is complicated with development of resistance which can be caused by few mechanisms. The aim of this study is to evaluate possible interactions of cisplatin, carboplatin and oxaliplatin with ABCG2 membrane efflux transporter which causes resistance to many cytotoxic drugs. Hoechst 33342 assay was performed on cell lines MDCKII (parent cell line) and MDCKII-ABCG2 (cell line genetically modified for expression of human ABCG2 gene). Intracellular concentration of Hoechst 33342 was not increased in presence of any tested cytotoxic drug; significant change of Hoechst 33342 intracellular fluorescence was observed only in presence of ABCG2 control substrate mitoxantron and ABCG2 inhibitor fumitremorgin C. Final results did not demostrate interactions of ABCG2 with any studied platinum cytotoxic. It shows insignificant interaction between ABCG2 and platinum cytotoxic drugs and so improbable effect of this transporter on toxicity reduction...
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Platinum cytostatics in the pharmacotherapy of cancer
Kritman, Gleb ; Čečková, Martina (advisor) ; Štaud, František (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Gleb Kritman Supervisor: PharmDr. Martina Čečková, Ph.D. Title of diploma thesis: Platinum cytostatics in the pharmacotherapy of cancer Platinum cytostatics belong currently to the most widely used anticancer drugs. They are involved as a part of chemotherapy regiments in many indications. Cisplatin, as a first substance discovered in this group of cytostatics, was originally registered for the treatment of testicular cancer and ovarian cancer, but it is currently used in many other indications. The limiting factor associated with the administration of cisplatin is represented by adverse effects and the development of resistance in previously sensitive tumor cells. These factors stimulated further research in the field of platinum compounds in order to create a substance with similar mechanism of action, but better properties. To date, more than thirty agents entered clinical trials, but only two of them are used worldwide - carboplatin and oxaliplatin. The aim of this thesis is to create a publication that summarizes current knowledge on platinum cytostatics, which are used in the pharmacotherapy of cancer. This work focuses especially on the comparison between the various representatives...
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