Abstract | Razvoj lijekova složen je proces. Započinje probirom molekula pomoću računalnog modeliranja čime se izdvajaju molekule s potencijalnim ljekovitim djelovanjem. Slijede in vitro ispitivanja koja se najčešće provode na specifičnim tkivima ili stanicama. Od velike su važnosti jer ukoliko molekule ne pokažu željenu aktivnost, daljni razvoj molekule se obustavlja, ne provode se in vivo i klinička ispitivanja. Test citotoksičnosti spada u skupinu in vitro ispitivanja i ključan je u razvoju citostatika. Citostatici djeluju na način da ubijaju tumorske stanice. Neselektivni su, odnosno toksični i za zdrave stanice pa izazivaju teške nuspojave. Smanjenjem doze citostatika, smanjuju se i nuspojave. Zbog nedovoljne učinkovitosti postojećih terapija tumora, znanstvenici su u potrazi za novim, učinkovitijim lijekovima. Tema ovog diplomskog rada bila je ispitati citotoksičnost derivata itakonske kiseline kao potencijalnih citostatika. Ispitivanje je provedeno na stanicama hepatocelularnog karcinoma HepG2. Stanice su tretirane otopinom sintetskih spojeva u različitim koncentracima (100 μM, 50 μM, 25 μM, 12,5 μM, 6 μM, 3 μM, 1,5 μM, 0,8 μM). Vijabilnost stanica određena je pomoću MTT testa u kojem metabolički aktivne stanice pretvaraju MTT u ljubičasto obojen spoj formazan, dok mrtve nemaju tu sposobnost. Količina nastalog formazana koja je proporcionalna broju živih stanica određena je spektrofotometrijski. Radi lakše usporedbe citotoksičnosti, rezultati su prikazani u obliku IC50 vrijednosti. Svi ispitivani spojevi citotoksični su za stanice hepatocelularnog karcinoma HepG2. Najučinkovitiji je spoj V4 koji je toksičan već pri niskim koncentracijama. To saznanje je bitno jer bi se mogao primjenjivati u niskim dozama ukoliko prođe ostale faze razvoja. |
Abstract (english) | Drug development is a complex process. It begins with the screening of molecules by computer modeling, by which the molecules with potential therapeutic effects are isolated. It is followed by in vitro experiments which are usually carried out on specific tissues or cells. These experiments are of great importance because if the tested molecules do not show the desired activity, further development of the molecules is halted and in vivo and clinical trials are not performed. The cytotoxicity test, which is crucial in the development of cytostatics, belongs to the group of in vitro studies. Cytostatics act by killing tumor cells. They are non-selective, i.e. toxic to healthy cells and cause severe side effects. Therefore, by reducing the dose of cytostatics, side effects are also reduced. Due to the lack of effectiveness of existing tumor therapies, scientists are looking for new, more effective drugs. The topic of this thesis was to test the cytotoxicity of itaconic acid derivatives as potential cytostatics. The study was performed on hepatocellular carcinoma HepG2 cells. Cells were treated with a solution of synthetic compounds in various concentrations (100 μM, 50 μM, 25 μM, 12.5 μM, 6 μM, 3 μM, 1.5 μM, 0.8 μM). Cell viability was determined by using an MTT assay in which metabolically active cells convert MTT to a purple-colored compound formazan, whereas the dead ones lack this ability. The amount of produced formazan, which is proportional to the number of living cells, is determined spectrophotometrically. To simplify cytotoxicity comparisons, the results are presented in the form of IC50 values. All test compounds were cytotoxic for hepatocellular carcinoma cells HepG2. The most effective compound, toxic even at low concentrations, is V4. This finding is important because if the compound passes other stages of the development, it could be administered at low doses. |