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The effect of antitumor agents on the expression and activity of cytochrome P450 subfamily 2C
Uher, Tomáš ; Dračínská, Helena (advisor) ; Feglarová, Tereza (referee)
Targeted therapies, acting by blocking essential biochemical pathways required for tumor cell growth and survival, are being used lately in multiple cases of cancer treatment. Cabozantinib, a small molecule inhibitor of receptor tyrosine kinases, is an example of a targeted drug, regulating growth, angiogenesis and metastatic pro- gression of medullary thyroid cancer. Such drugs are also often complemented by cytotoxic agents, e.g. ellipticine; however, therapeutic usage of ellipticine itself is limited due to its poor solutibility and variety of adverse effects. In this bachelor thesis, effects of cabozantinib, ellipticine, or their combina- tion on gene and protein expression of cytochromes P450 2C6 and 2C11 have been studied in vivo. Aforementioned cytochromes have an important role in biotrans- formation of xenobiotics in rat liver. Their protein expression has been assessed by Western blot immunoassay technique, while the gene expression was evaluated by quantitative PCR method. Furthermore, the effects of studied substances and their combination on CYP2C6 and CYP2C11 specific activity have been determined by diclofenac 4'-hydroxylation and testosterone 16α-hydroxylation, respectively. Additionally, direct inhibitory effect of cabozantinib on recombinant CYP2C11 rat isoform has been studied in...
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Effect of cytochrome b5 on activity of cytochromes P450
Ličko, Vojtech ; Indra, Radek (advisor) ; Feglarová, Tereza (referee)
ABSTRACT Cytochrome b5 (CYB5) is heme protein capable of reduction of cytochromes P450 (CYP) or some other enzymes. However, his regulative capability was also observed by his apo form, i.e. in absence of heme prosthetic group in the active center. CYB5 can accept electron from cytochrome b5 reductase (CYB5R) or from cytochrome P450 reductase (CYPOR). CYPOR by itself is reduced by NADPH and is also able to forward electron to CYP independently of CYB5. CYB5R on the other hand is reduced by NADH. Efficiency of CYB5 to accept and forward an electron was studied in vitro with five different substrates - testosterone, Sudan I, aristolochic acid I (AAI), ellipticine and vandetanib. These substrates were chosen considering their characteristic reactions, which are catalyzed by their respective isoforms of CYP. The experiments with these substrates were carried out in the medium with recombinant CYPs prepared in insect cells or E. coli or in the medium with hepatic microsomes isolated from different organisms. Rats, from which the majority of these microsomes was isolated, were premedicated by different CYP inducers. The experiments were carried out in medium with NADH or NADPH in order to assess the capability of CYB5 to reduce CYP independently of CYPOR. The capability of CYB5 and CYB5R to act as a...
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