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Study of Membrane Transport of Albumin in the Kidney in vitro
Suchan, Martin ; Trejtnar, František (advisor) ; Lázníček, Milan (referee)
Mgr. Martin Suchan 2010 Hradec Králové Doc. PharmDr. František Trejtnar, CSc. Katedra farmakologie a toxikologie Abstrakt Cíl: Práce byla zaměřena na studium transportu albuminu in vitro pomocí akumulačních buněčných studií a s využitím dvou typů albuminu - albuminu značeného techneciem- 99m a FITC-albuminu značeného fluorescenční sondou. Součástí práce bylo ověření optimální přípravy a hodnocení kvality 99m Tc-albuminu. Cílem bylo porovnat akumulaci obou typů albuminu v ledvinných buňkách, zjistit vliv vybraných faktorů na akumulaci albuminu, stanovit typ mechanismu transmembránového transportu albuminu a zpracovat získané výsledky do formy odborného vědeckého sdělení. Metody: Ke značení albuminu techneciem-99 bylo použito optimalizovaného postupu za použití nízkého pH, vysoké koncentrace albuminu a optimální koncentrace cínatých iontů. Radiochemická čistota 99m Tc-albuminu byla ověřována chromatografií na papíře. Jako buněčné in vitro modely byly použity izolované nativní ledvinné buňky potkana a standardní linie ledvinných tubulárních buněk prasete LLC-PK1. V akumulačních studiích byl sledován uptake albuminu v závislosti na teplotě, čase, složení inkubační směsi, přítomnosti inhibitoru transportu a byl hodnocen vliv experimentálního modelu. U LLC-PK1 buněk byl též testován vliv stáří buněk na míru...
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Study of stability of receptor specific radiopharmaceuticals
Hanzalová, Lenka ; Lázníček, Milan (advisor) ; Trejtnar, František (referee)
Thesis: Determination of radiochemical purity and stability compared to two selected receptor-specific radiolabelled peptides. Charles University, Faculty of Pharmacy in Hradec Kralove Lenka Hanzalová, May 2011 Abstract: Radiopharmaceuticals are among medicinal substances which have very a limited application over time. Not only due to the fact that it gradually decreases their activity, depending on the half-life of the radionuclide applied, but it may also lead to destructive reactions in the molecule. In view of these reactions, there are decreases in the percentage of radiopharmaceutical degradation of products, which reduces the value of medicines. Degradation of products containing non-peptide nature radionuclides from the parent substances differ in their distribution in the body, increasing side effects, and radiopharmaceuticals, due to their nature, can accumulate in different tissues and subsequently damage them. It is therefore important to know and be able to assume representation of the evolution of the degradation of products over time. In the presented thesis, the problem of determining stability, or radiochemical purity at different time intervals in two days, is solved. Substances being compared were a synthetic somatostatin analogue, labelled with radionuclides two mA, pure beta-emitter 90...
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Analysis of the Biological Behavior of Receptor-Specific Radiopharmaceuticals in Vitro
Melicharová, Ludmila ; Lázníček, Milan (advisor) ; Pávek, Petr (referee) ; Komárek, Pavel (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Kralove Department of: Pharmacology and Toxicology Candidate: PharmDr. Ludmila Melicharová Supervisor: Prof. PharmDr. Ing. Milan Lázníček, PhD. Title of Doctoral Thesis: Analysis of the biological behavior of receptor-specific radiopharmaceuticals in vitro In the present work were studied receptor-specific radiopharmaceuticals and their biological behavior. Studies were performed with radiolabelled somatostatin and gastrin analogues. All of these substances are potentially useful in the diagnosis and therapy of neuroendocrine tumors. Somatostatin analogues under study: DOTA-TATE and DOTA-NOC labelled with 111 indium, 177 lutetium, 90 yttrium and 125 iodine. The studies were performed on AR42J cell line (rat pancreatic tumour cells). Internalization rate of these analogues and their specific binding to the cells with high density of appropriate receptors were determined. The data were compared with values obtained after intravenous administration of the same peptides to rats. The uptake of radioactive peptides in tissues with high density of somatostatin receptors in vivo were correlated with the results obtained in vitro conditions. We have compared four 111 In labelled gastrin analogues (so called minigastrins /MG/, namely MG11, MG45,...
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Study of plasma protein binding of receptor-specific peptides
Křivková, Šárka ; Lázníček, Milan (advisor) ; Trejtnar, František (referee)
SUMMARY: Drug binding to plasma proteins is one of the primary pharmacological parameters. The binding rate has significant effects on drug distribution and drug elimination. In vitro investigation of plasma protein binding helps us foresee their destiny after its administration to a living organism. This dissertation focused on studying plasma protein binding of DOTA-NOC marking by 90 Y and 111 In in term of interspecies comparison. The binding was determined by methods of equilibrium dialysis at 37řC. Results showed binding of 90 Y-DOTA-NOC increases respectively: human < rabbit < bovine < rat plasm. Binding of 111 In-DOTA-NOC increases respectively: pork < human < rat < rabbit plasm. KEYWORDS: protein binding, DOTA-NOC, 111 In, 90 Y, interspecies comparison
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Determination of plasma protein binding of new radiopharmaceuticals for Alzheimer's disease imaging
Hyršová, Lucie ; Lázníček, Milan (advisor) ; Melicharová, Ludmila (referee)
CHARLES UNIVERSITY IN PRAGUE Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Lucie Hyršová Supervisor: Prof. PharmDr. Ing. Milan Lázníček CSc. Title of thesis: Determination of plasma protein binding of new radiopharmaceuticals for Alzheimer`s disease imaging Summary: Determination of plasma protein binding of new potential drugs belongs to basic parameters which are made at early stages of clinical trials. The drug-protein binding affects all the pharmacokinetic factors and it influences pharmacodynamics as well. The aim of this thesis is to examine binding of 99m Tc-5C-2-BOC and 99m Tc-5C-5-BOC which are potential radiodiagnostics of Alzheimer`s disease, is investigated in this work. Blood plasma of four species (namely human, rabbit, bovine and sewer rat`s plasma) is tested and findings are mutually compared. The binding in both cases is determined by means of two methods - equilibrium dialysis and ultrafiltration. The binding of both substances is the lowest in human and bovine plasma with the use of equilibrium dialysis. The binding of 99m Tc-5C-2-BOC determined by ultrafiltration is the lowest in bovine plasma and a slightly higher in human one, the binding of 99m Tc-5C-5-BOC is the lowest in human plasma. These findings show that binding on plasma proteins...
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Study of plasma protein binding of two bifunctional chelates labelled with In-111
Drymlová, Petra ; Lázníček, Milan (advisor) ; Trejtnar, František (referee)
Summary: Distribution of many drugs in an organism is significantly influenced by their binding to plasma proteins. Determination of the extent of plasma binding for a concrete drug is necessary for prediction of its pharmacokinetics after administration to the organism. The aim of this thesis was to determine binding of a new bifunctional chelating agent DTPA-oxn labelled by 111 In to the plasma proteins of human and tree animal species and to compare these results with the plasma protein binding of routinely used radiopharmaceutical 111 In-DTPA. For measurement, a method of equilibrium dialysis at 37řC was used. The results show, that binding of 111 In-DTPA-oxn to the proteins of human, bovine, rabbit and rat plasma is similarly very low as at the compared chelate 111 In-DTPA and impossible to determine by equilibrium dialysis and pharmacokinetically unimportant. Radiochemical purity was also determined for both complexes by the method of thin layer chromatography ITLC-SG. Measured value was higher than 98% for each compound.
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Study of Biological Properties of Hyaluronic Acid Derivatives
Svanovský, Evžen ; Lázníček, Milan (advisor) ; Tilšer, Ivan (referee) ; Mičuda, Stanislav (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of pharmacology and toxikology Candidate: Mgr. Evžen Svanovský Supervisor: Prof. PharmDr. Ing. Milan Lázníček, CSc. Title of Doctoral Thesis: Study of biological properties of hyaluronic acid derivatives This PhD thesis, is dealing with physiological properties of Hyaluronic Acid labelled with 111 In isotope. HA is naturally occurring Glycosaminoglycan within organisms with many important functions on the cellular and organ level, including physiological and pathological processes. Its character of physiologically occurring substance makes its tracing in the organism very difficult. Thus preparation of suitable derivative and repeatable procedure of labelling of this molecule became the most important point of the whole work. During the experimental part DTPA-HA molecule created by CPN Dolni Dobrouč has been labelled using well-known method used for labelling of DTPA molecules with 111 In. A reproducible procedure has been created where 0,4 M Acetate buffer pH 5,5 is mixed with an equal volume of DTPA-HA purified by gel filtration and required amount of isotope 111 In and the whole mixture is mixed for 0,5 h. This method was used for labelling of DTPA-HA molecules of three different molecular weights 10, 100 and 400kDa,...
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Study of Transport Mechanisms of Receptor-Specific Radiopharmaceuticals in the Kidney Cells.
Cihlo, Jan ; Lázníček, Milan (advisor) ; Šimůnek, Tomáš (referee) ; Komárek, Pavel (referee)
Renal accumulation of radiolabelled receptor specific substances can represent clinical problems for diagnosis and therapy of some typical malignancies due to radionephrotoxicity of these substances. The aim of this work was to design a suitable in vitro model for evaluating cumulative disposals of studied radiopharmaceuticals in the kidney and for finding possible mechanisms reducing the reabsorption of these substances by renal tubular cells. The suggested model designed for this purpose provides another opportunity for evaluation of their radionephrotoxical potential. The aforementioned in vitro method was standardised and used for the study of transport mechanism of some representatives of radiolabelled somatostatin analogues (111In/125I/90Y/177Lu-DOTA-Tyr3-octreotate, 111In/90Y/177Lu-DOTA-1-Nal3-octreotide), antibodies (111In/90Y/99mTc-AntiCD66) and albumin (FITC-albumin, 99mTc-albumin). All studied radioactive substances were radiolabelled with good practice and analyzed using HPLC and/or ITLC-SG methods. The radiolabelling technique of all studied specifically acting substances (radiolabelled peptides and antibodies) and also any concrete radionuclide showed no significant influence over the internalization into the renal OK cells. The difference of the internalized amount of FITC-albumin and...
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Study on Renal Accumulation of Receptor-specific Radiopeptides
Kubačková, Tereza ; Trejtnar, František (advisor) ; Lázníček, Milan (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Candidate: Tereza Kubačková Consultant: doc. PharmDr. František Trejtnar, CSc. Title of Thesis: Study on renal accumulation of receptor-specific radiopeptides Radiolabeled peptides targeted to cholecystokinin/gastrin receptors are promising compounds for radiodiagnostics and radiotherapy of some malignancies. Shorter chain derivates of gastrin called minigastrins are particularly interesting. The use of these agents is limited by their nephrotoxicity caused by accumulation of the radiopeptides in the kidney. The aim of the study was to investigate accumulation rate and renal uptake mechanisms of new receptor specific peptides from the group of gastrin analogs using in vitro models. The investigation was aimed at a comparison of the renal uptake of DOTA-minigastrin 48 (DOTA-MG48) and DOTA-minigastin 11 (DOTA-MG11), both labeled with indium-111. In addition, a potential influence of the radiolabel on the renal accumulation was also investigated using DOTA- MG11 labeled with indium-111 or with lutecium-177. A comparison of the renal accumulation of the radiolabeled minigastrins with compounds from another group of receptor specific radiopeptides - somatostatin analogs such as 111 In-DOTA-NOC...
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