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An identification of selective antagonists of constitutive androstane receptor
Ondraszková, Karolína ; Smutný, Tomáš (advisor) ; Pourová, Jana (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Karolína Ondraszková Supervisor: PharmDr. Tomáš Smutný, Ph.D. Title of diploma thesis: An identification of selective antagonists of constitutive androstane receptor The constitutive androstane receptor (CAR) is an important ligand-regulated xenosensor that regulates the transcription of important drug biotransforming enzymes (e.g. CYP2B6). The pregnane X receptor (PXR) belongs to the same family of nuclear receptors as the CAR. The structure and function of both receptors share many common features. However, PXR has a larger and more flexible ligand-binding domain. At present, a selective, highly potent and non- toxic inhibitor of CAR is not practically available. This work aims to determine the affinity of previously identified compounds in our group with antagonistic activity towards CAR. This is a replication study. For this purpose, we tested the compounds using gene reporter and two-hybrid assay. Of all the tested substances, we obtained desired results with substance 1A. Using the gene reporter assay method, we found that substance 1A can reduce basal CAR expression but also suppresses CAR activity induced by the model agonist CITCO. The high affinity of the substance for the...
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Interactions of rifampicin derivatives with pregnane X receptor and OATP1B1 transporter
Krajníková, Zdeňka ; Smutný, Tomáš (advisor) ; Mladěnka, Přemysl (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Zdeňka Krajníková Supervisor: PharmDr. Tomáš Smutný, Ph.D. Title of diploma thesis: Interactions of rifampicin derivatives with pregnane X receptor and OATP1B1 transporter Pregnane X receptor (PXR) is a ligand-activated transcription factor, which regulates gene expression in the liver. Among PXR target genes, cytochrome P450 3A4 (CYP3A4) is the most important enzyme responsible for metabolism of clinically used drugs. Rifampicin (RIF) is a prototype PXR ligand. It enters hepatocytes across the basolateral membrane by OATP1B1 transporter. Noteworthy, RIF is a chemically unstable molecule. Additionally, it is also metabolized in the human body. In this diploma thesis, we decided to determine the affinity of RIF derivatives (i.e. rifampicin quinone, rifampicin N-oxide, 25-desacetylrifampicin, and 3-formylrifamycin SV) to PXR and OATP1B1, which has not been explored in details so far. For this, two hybrid and gene reporter assays were employed. As revealed by two hybrid assay, rifampicin quinone, rifampicin N-oxide, and 3- formylrifamycin SV (10 µM) activated PXR at a level comparable with RIF (positive control). Contrary, 25-desacetylrifampicin showed a lower affinity to PXR than that...
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Interactions of rifampicin derivatives with pregnane X receptor and OATP1B1 transporter
Krajníková, Zdeňka ; Smutný, Tomáš (advisor) ; Mladěnka, Přemysl (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Zdeňka Krajníková Supervisor: PharmDr. Tomáš Smutný, Ph.D. Title of diploma thesis: Interactions of rifampicin derivatives with pregnane X receptor and OATP1B1 transporter Pregnane X receptor (PXR) is a ligand-activated transcription factor, which regulates gene expression in the liver. Among PXR target genes, cytochrome P450 3A4 (CYP3A4) is the most important enzyme responsible for metabolism of clinically used drugs. Rifampicin (RIF) is a prototype PXR ligand. It enters hepatocytes across the basolateral membrane by OATP1B1 transporter. Noteworthy, RIF is a chemically unstable molecule. Additionally, it is also metabolized in the human body. In this diploma thesis, we decided to determine the affinity of RIF derivatives (i.e. rifampicin quinone, rifampicin N-oxide, 25-desacetylrifampicin, and 3-formylrifamycin SV) to PXR and OATP1B1, which has not been explored in details so far. For this, two hybrid and gene reporter assays were employed. As revealed by two hybrid assay, rifampicin quinone, rifampicin N-oxide, and 3- formylrifamycin SV (10 µM) activated PXR at a level comparable with RIF (positive control). Contrary, 25-desacetylrifampicin showed a lower affinity to PXR than that...
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Experimental approaches for studying hepatic enzyme induction mediated by pregnane X receptor
Dobečka, Kryštof ; Smutný, Tomáš (advisor) ; Carazo Fernández, Alejandro (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Kryštof Dobečka Supervisor: PharmDr. Tomáš Smutný, Ph.D. Advisor: prof. PharmDr. Petr Pávek, Ph.D. Title of diploma thesis: Experimental approaches for studying hepatic enzyme induction mediated by pregnane X receptor The thesis focuses on hepatic pregnane X receptor (PXR)-mediated induction of biotransformation enzymes. Emphasis is placed on experimental models and methods which are used for the assessment of enzyme induction. In addition to summarizing its well- established role as a xenobiotic-sensing receptor, PXR is also presented as a transcription factor with an important role in endogenous pathways. Furthermore, cell and animal models are evaluated in terms of expression and function of PXR and its target xenobiotic-metabolising enzymes. Primary human hepatocytes in 2D cultures are considered to be the gold standard of in vitro hepatic models. However, 3D technologies are expected to be increasingly used in the future. The use of animal models is limited due to pronounced interspecies differences in PXR activation. Thus, humanized models have been established to overcome these limitations. Next, this thesis comments screening methods for an assessment of interaction between PXR and...
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