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Information × Registration Number 0213U007124, 0110U001192 , R & D reports Title Study of the effects of antimitotic antineoplastic drugs upon proliferation, apoptosis and cell cycle regulatory mechanisms in the malignant thyroid cells. popup.stage_title Head Tron’ko Mykola Dmytrovych; Pushkarev Volodymyr Mikhailovich, Registration Date 05-04-2013 Organization State Institution "V.P. Komisarenko Institute of Endocrinology and Metabolism оf the Academy of Medical Scienses of Ukraine" popup.description2 Object of research: biochemical mechanisms of cell cycle regulation and apoptosis in cells of malignant tumors of the thyroid gland. Aim: To study the effect of taxanes on anaplastic thyroid cancer cells. Methods and equipment: cell culture, extraction of nucleic acids, DNA electrophoresis in agarose gel, electrophoresis of proteins in polyacrylamide gel, Western blotting, flow cytometry, laboratory ionomer-160M, spectrophotometer СФ-46, pipettes. Results: The tested anaplastic thyroid cancer (ATC) cells are sensitive to taxanes, which caused a dose-dependent cell death of anaplastic thyroid cancer cells. More sensitive were cells of line KTC-2, possibly due to the activity of unmutated p53 gene. Important in practical terms, may be evidence that the cytotoxicity of taxanes significantly increased in the enriched culture medium. Determination of clonogenicity of the ATC cells showed that after incubation of the cells with the compound starting from 50 nmol/l of paclitaxel colonies were not formed. Flow cytometry study showed that paclitaxel, even at concentrations as low as 1 nmol/l after 48 h of incubation caused a marked apoptotic effects. At a concentration of the drug 6 nmol/l most of the KTC-2 cells are in the apoptosis, and from the concentration of 15 nmol/l - almost all cells. The findings have practical importance because it shows that low concentrations of paclitaxel by prolonged exposure to exercise maximum cytotoxic effect on the ATC cells. The inversion of the cell membranes of the cells in the presence of paclitaxel accompanied by corresponding changes in the cells, characteristic of apoptosis. Observed activation of the main effector caspase - caspase-3 and caspase cleavage of the protein poly (ADP-ribose) polymerase (PARP). Studied the participation of protein kinases glycogen synthase kinase 3beta (GSK-3beta), PI3K and Akt in mediating the action of anti-cancer drug, paclitaxel in the cells of thyroid cancer. It is shown that in the presence of paclitaxel activation of Akt in tumor cells is inhibited. Inhibitor of PI3K, LY294002, increased the activation of caspase-3 and inhibits the expression of an inhibitor of apoptosis - survivin, indicating a possible mechanism of tumor cell resistance to paclitaxel with PI3K/Akt-cascade. Therefore, the combined effect of inhibitors of this cascade and paclitaxel is one of the ways to strengthen carcinostatic action of the drug on the anaplastic thyroid cancer cells. Exploring the role of GSK-3beta in mediating of the cytotoxic effects of paclitaxel upon the ATC cells showed that paclitaxel causes dose-dependent cell death, which is determined by the time of incubation of the cells with the compound. Inhibition of GSK-3beta by specific inhibitor of the kinase, Li+, leads to a significant weakening of the cytotoxic effect of paclitaxel. These data suggest that GSK-3beta is a one of major protein kinases that mediate paclitaxel-induced apoptosis in ATC. Determined the effect of paclitaxel on the activation of the cell cycle inhibitor pRB, cycle check-point kinase CHK2 in human ATC cells. Activation pRB in KTC-3 cells, showed multidirectional processes. Paclitaxel activates CHK2 in ARO cell line, but not in the FRO cell line. At low concentrations taxanes exhibit mitogenic properties, and cause of the cell cycle arrest when exposed to taxanes can be activation of CHK2 and peptidyl-prolyl cis/trans isomerase Pin1. At concentrations of 1 - 25 nmol/l taxanes caused radioprotective effect on the ATC cells. The key mechanism that links cell cycle disorders caused be these compounds with taxane-induced apoptosis is the activation of CDK. Phosphorylation by cyclin-dependent kinases of antiapoptotic protein Bcl-2 is the trigger mechanism of apoptosis. Theoretical and practical novelty: the findings represent new and important information that will be used in future preclinical studies for the development of new treatments for anaplastic cancer. Novelty and advantages over similar products: distinction from the existing data is a detailed study of the mechanisms of cell cycle, which react to the effects of ionizing radiation and taxanes. In the prototype, this analysis is missing. Efficiency of implementation: The data published in the scientific literature and presented at scientific meetings. Patented. Field of use: scientific and practical institutions of endocrinologic and oncologic profiles, medical universities. Product Description popup.authors Калініченко Олена Вікторівна Левчук Наталія Іванівна Мишуніна Тамара Мар'янівна Пушкарьов Віктор Володимирович Пушкарьов Володимир Михайлович Сологуб Неля Віталіївна Тронько Микола Дмитрович popup.nrat_date 2020-04-02 Close