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Influence of V-ATPase inhibitors on chemoresistant neuroblastoma lines in vitro
Honzejková, Karolína ; Eckschlager, Tomáš (advisor) ; Martínková, Markéta (referee)
Tumor diseases are one of the most common causes of death worldwide. Despite the great advances in therapy in the last fifty years, this is still a serious health problem. Therefore, great efforts are still concentrated on development of new anti-cancer drugs and therapeutic approaches. Neuroblastoma (NBL) is the most common tumor in infants and the fourth most common in children. Successful treatment is greatly complicated by its heterogeneity. Chemoresistance is an undesirable phenomenon of chemotherapy. One of the chemoresistance mechanisms is the accumulation of weakly basic anticancer drugs in lysosomes. This work deals with the measurement of lysosomal uptake of these compounds in neuroblastoma cell lines UKF-NB-4 and derived, ellipticine-resistant, line (UKF-NB- 4ELLI ) under different conditions. A method for determining the cell lysosomal capacity (volume) by measuring fluorescence intensity of lysosome-specific LTR dye was introduced and the ability of bafilomycin A, a V-ATPase inhibitor, to potentiate the effects of an anticancer agent ellipticine by inhibiting its lysosomal accumulation was investigated. Keywords: neuroblastoma, lysosome, vacuolar ATPase, multidrug resistance
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Monitoring the effect of medically important drugs on activity of human P-glycoprotein using fluorescent probes
Veľas, Lukáš ; Gášková, Dana (advisor) ; Sigler, Karel (referee)
One of the main causes of failure in cancer treatment when using chemotherapy is the phenomenon of multidrug resistance - MDR. The most important protein mediating MDR in human cells is P-glycoprotein. The main goal of this thesis was the modification of fluorescent method - developed for studying yeast MDR pumps at the Department of Biophysics at the Institute of Physics of Charles University - to study the activity of P-glycoprotein in human tumor cells. The fluorescent method is based on the use of redistribution potentiometric probe diS-C3(3) which we found to be a substrate of P-glycoprotein. The optimalization of the method (experimental window) allowed sensitive monitoring of changes in the activity of P-glycoprotein caused by the effect of its several known inhibitors/substrates: oligomycin, amiodarone, verapamil, vinblastine, ketoconazole, itraconazole and FK506. New important results regarding the effect of these medically significant drugs on human cells were obtained. The developed method will undoubtedly be of great benefit in the search for new effective inhibitors and the study of their mechanisms in the future.
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