Molecularne spremembe v celicah rezistentnih na citostatike

The major obstacle to the ultimate success in cancer therapy is the ability oftumor cells to develop resistance to anticancer drugs. Several molecular mechanisms have been suggested to be involved in drugresistance: a) decrease in the intracellular drug accumulation (increased activity of membrane transporters such as P-glycoprotein or multidrug-resistance-associated protein), b) changes in intracellular detoxification system (increased concentrations of glutathione or metallothioneins, or increased activity of related enzymed), c) alteration in nuclear enzymes (enhanced DNA repair and/orbetter tolerance of DNA damage, decreased acitivty of topoisomerases), d) altered expression of oncogenes (inducing increased level of protective molecules in cells or the inhibition of apoptosis). Drug resistance is a multifactorial phenomenon. The complexity of molecular alterations in drug-resistant cells is and will stay the main problem for the successful treatment of cancer

Total tissue lactate dehydrogenase activity in endometrial carcinoma

Lactate dehydrogenase (LDH) is essential for continuous glycolysis necessary for accelerated tumor growth. The aim of this study was to reconsider if assay of total tissue activity of this enzyme could be useful as marker for endometrial carcinoma (EC). Activity of LDH was measured spectrophotometrically in homogenate supernatants of uterine tissue samples of 40 patients (10 normal endometria, 27 normal myometria, and 33 EC), including 30 matched pairs. Data obtained were analyzed in relation to clinical and histopathologic findings and compared with our previously published results on the tissue levels of the same enzyme in ovarian cancer and on the proteolytic activity of dipeptidyl peptidase III (DPP III) in EC (suggested biochemical indicator of this malignancy). Significantly increased (1.8-3.0 times; P < 1 x 10(-4)) LDH activity was observed in EC samples if compared with normal uterine tissues. This rise was not related to the clinicopathologic findings, however. In contrast to previous results on LDH in ovarian carcinomas, a significant rise in LDH activity was found already in grade 1 EC. Using the cutoff value of 1.06 U/mg, diagnostic sensitivity of 82%, specificity of 100%, and accuracy of 91% for total tissue LDH assay have been calculated. A correlation of tissue's LDH and DPP III activities was found, and their combined assay for EC showed increased diagnostic sensitivity (94%) and accuracy (96%)

Genotoksični učinak ekstrakta zelenog čaja na ljudske stanice raka grkljana u kulturi

Green tea (Camellia sinensis) contains several bioactive compounds which protect the cell and prevent tumour development. Phytochemicals in green tea extract (mostly flavonoids) scavenge free radicals, but also induce pro-oxidative reactions in the cell. In this study, we evaluated the potential cytotoxic and prooxidative effects of green tea extract and its two main flavonoid constituents epigallocatechin gallate (EGCG) and epicatechin gallate (ECG) on human laryngeal carcinoma cell line (HEp2) and its crossresistant cell line CK2. The aim was to see if the extract and its two flavonoids could increase the sensitivity of the cisplatin-resistant cell line CK2 in comparison to the parental cell line. The results show that EGCG and green tea extract increased the DNA damage in the CK2 cell line during short exposure. The cytotoxicity of EGCG and ECG increased with the time of incubation. Green tea extract induced lipid peroxidation in the CK2 cell line. The pro-oxidant effect of green tea was determined at concentrations higher than those found in traditionally prepared green tea infusions.Zeleni čaj, koji je vrlo popularno piće, proizvodi se iz biljke Camellia sinensis i bogat je fl avonoidima za koje se smatra da imaju važnu ulogu u sprečavanju nastanka raka. Kao najvažniji mehanizmi djelovanja fl avonoida najčešće se spominju vezanje slobodnih radikala te sprečavanje nastanka reaktivnih kisikovih skupina. Cilj ovog rada bio je istražiti potencijalni genotoksični učinak ekstrakta zelenog čaja te epigalokatehin galata (EGCG) i epikatehin galata (ECG), fl avonoida koji se u zelenom čaju nalaze u najvišoj koncentraciji. Dobiveni su rezultati pokazali da EGCG i ekstrakt zelenog čaja izazivaju povećanu citotoksičnost otporne stanične linije raka grkljana CK2 nakon kraće inkubacije. Produljenjem vremena inkubacije povećava se citotoksičnost istraživanih spojeva. Također, ekstrakt zelenog čaja izaziva lipidnu peroksidaciju u CK2-stanicama. Prooksidativni učinak ekstrakta zelenog čaja u koncentracijama višim od onih prisutnih u infuzijskoj otopini dobivenoj tradicionalnim načinom, imaju prooksidativno djelovanje

The Mutagenic Potential of Chloroform, Orange Oil, Eucalyptus Oil and Halothane by Salmonella/Microsome Assay

Svrha rada bila je ispitati mutagenu aktivnost četiriju komercijalno dostupnih otapala gutaperke Salmonella/mikrosomskim testom. Ispitana su bila otopala: kloroform, narančino ulje, eukaliptusovo ulje i halotan u količinama od 10 μl, 30 μl, 50 μl, 100 μl i 200 μl. Upotrijebljen je standardni Ames test inkorporacije na ploči pri čemu su rabljene bakterije Salmonella typhimurium soja TA 98 i TA 100 s metaboličkom aktivacijom enzimom S9. Rezultati istraživanja pokazuju citotoksičnu aktivnost eukaliptusova ulja u svim koncentracijama, narančina ulja u koncentracijama od 50 μl i većima. Kloroform je pokazao citotoksičnost u koncentracijama od 100 μl i 200 μl. Niti jedno otapalo gutaperke nije pokazala mutagenu aktivnost Ames testom u ispitivanim koncentracijama.The aim of this study was to examine mutagenic activity of four commercially available gutta-percha solvents by means of the Salmonella/ microsome assay. The examined solvents were: chloroform, orange oil, eucalyptus oil and halothane in amounts of 10 μl, 30 μl, 50 μl, 100 μl and 200 μl. Standard plate incorporation Ames test was preformed by using two tester strains of Salmonella typhimurium, TA 98 and TA 100, with metabolic activation of S9. The results showed toxicity of eucalyptus oil in all aliquots, orange oil in aliquots of 50 μl and above and chloroform in aliquots of 100 μl and 200 μl, but all four substances responded negative to the Ames test. These results indicate that the tested solvents do not possess mutagenic activity toward the Salmonella strains used

Katepsin D in inhibitorji ter aktivatorji plasminogena tipa 1 v normalnem, benignem in malignem ovarijskem tkivu

Background. The aim of the present study was to determine the concentration ofcathepsin D (Cath D) and plasminogen activator inhibitor type 1 (PAI-1) in normal ovarian tissues, benign and malignant ovarian tumor tissues, and to asses relationship between Cath D and PAI-1 content, and some clinical and pathohistological parameters. Materials and methods. Cath D contents and PAI-1concentrations were determined (using immunoradiometric ELSA-Cath D assayand commercial IMUDIND R ELISA immunoassay, respectively) in 35 samples: 10 normal ovarii, 10 benign, 10 primary malignant and 5 metastatic ovarian tumors. Results. The concentrations of Cath D were significantly higher in malignant (32.89+-14.26 pmol/mg protein ) and metastatic (31.42+-10.24 pmol/mgprotein), than in normal (13.68+-4.03 pmol/mg protein) and benign (17.89+-13.13 pmol/mg protein) ovarian tissues. There was no statistical differences in the concentrations of PAI-1 between normal, benign, malignant and metastatic tumor specimens. The concentrations of Cath D as well as PAI-1 did not correlate to the age of patients, menopausal status, parity, GOG risk group, clinical stage or pathohistological grading. Conclusion. Concentrationsof Cath D (but not PAI-1) were significantly increased in malignant and metastatic ovarian tumor tissues when compared to normal and benign ovarian tumor samplesthey were independent from pathohistological andclinical parameters.Izhodišča. Študijo smo opravili z namenom, da bi določili koncentracijo katepsina D in aktivatorjev ter inhibitorjev plasminogena tipa 1 (PAI-1) v normalnem ovarijskem tkivu ter v tkivu benignega in malignega tumorja ter ocenili razmerje med vsebnostjo katepsina D in PAI-1 in še druge klinične ter patohistološke parametre. Material in metode. V 35 vzorcih (10 vzorcev normalnega ovarijskega tkiva, 10 vzrocev benignega, 10 vzrocev malignega tumorskega tkiva in 5 vzorcev metastatskega ovarijskega tkiva) smo izmerili vsebnost katepsina D (izmerjeno z imunoradiometrično metodo ELISA-Cath D) in koncentracijo PAI-1 (izmerjeno s komercialno imunsko metodo IMUDINDR ELISA). Rezultati. Koncentracija katepsinov D so bile bistveno višje v malignem (32.89+- 14.26 pmol/mg protein) in metastatskem tkivu (31.42 +- 10.24 pmol/mg protein) kot pa v normalnem ovarijskem tkivu (13.68 +- 14.03 pmol/mg protein) ali tkivu benignega ovarijskega tumorja (17.89 +- 13.13 pmol/mg protein). Med koncentracijami PAI-1 v vzorcih normalnega tkiva in vzorcih benignega in malignega tumorskega tkiva ni bilo ugotovljenih statistično pomembnih razlik. Koncentracije katepsina D in PAI-1 tudi niso bile v korelaciji s starostjo bolnic, menopavzo, gravidnistjo, rizičnim GOG skupinam, kliničnemu stadije in patohistološko stopnjo tumorja. Zaključki. Koncentracije katepsina D so bile pomembno večje v malignem in metastatskem ovarijskem tkivu, medtem ko koncentracije PAI-1 niso bile povečane. Koncentracije katepsinov niso bile odvisne od patohistoloških in kliničnih parametrov

Increased Adenovirus Type 5 Mediated Transgene Expression Due to RhoB Down-regulation

Adenovirus type 5 (Ad5) is a non-enveloped DNA virus frequently used as a gene transfer vector. Efficient Ad5 cell entry depends on the availability of its primary receptor, coxsackie and adenovirus receptor, which is responsible for attachment, and integrins, secondary receptors responsible for adenovirus internalization via clathrin-mediated endocytosis. However, efficacious adenovirus-mediated transgene expression also depends on successful trafficking of Ad5 particles to the nucleus of the target cell. It has been shown that changes occurring in tumor cells during development of resistance to anticancer drugs can be beneficial for adenovirus mediated transgene expression. In this study, using a study model consisting of a parental cell line, human laryngeal carcinoma HEp2 cells, and a cisplatin-resistant clone CK2, we investigated the cause of increased Ad5-mediated transgene expression in CK2 as compared to HEp2 cells. We show that the primary cause of increased Ad5- mediated transgene expression in CK2 cells is not modulation of receptors on the cell surface or change in Ad5wt attachment and/or internalization, but is rather the consequence of decreased RhoB expression. We show that RhoB plays an important role in Ad5 post-internalization events and more particularly in Ad5 intracellular trafficking. To the best of our knowledge, this is the first study showing changed Ad5 trafficking pattern between cells expressing different amount of RhoB, indicating the role of RhoB in Ad5 intracellular trafficking

Half-Sandwich Ir(III) and Os(II) Complexes of Pyridyl-Mesoionic Carbenes as Potential Anticancer Agents

A series of cationic chlorido arene-iridium(III) and arene-osmium(II) complexes with bidentate pyridyl functionalized mesoionic carbenes (MIC) of the 1,2,3-triazol-5-ylidene type have been prepared. The variations in the ligand structures include the position of the pyridyl substituent relative to the triazolylidene ring (N-wingtip vs C-wingtip), phenyl versus ethyl substituents, and incorporation of several functional groups at the phenyl substituents. Five complexes have been characterized by X-ray structural analysis. All complexes, including osmium(II) and ruthenium(II) analogues having a pyrimidyl in place of the pyridyl group, have been studied for their cytotoxic activity on a human cervical carcinoma HeLa cell line. Two of the compounds, Ir5 and Ir9, were the most cytotoxic with IC50 values of 7.33 μM and 2.01 μM, respectively. Examination of their cytotoxic effect on different cell lines revealed that they preferentially kill cancer over normal cells. The Ir5 and Ir9 compounds arrested cells in G2 and induced a dose-dependent increase in SubG0/G1 cell population. Apoptosis, as the primary mode of cell death, was confirmed by Annexin V/PI staining, detection of cleaved PARP, and caspases 3 and 7 activity upon treatment of HeLa cells with both compounds. The higher toxicity of Ir9 is probably due to its increased accumulation in the cells compared to Ir5. The role of glutathione (GSH) in the protection of cells against Ir5 and Ir9 cytotoxicity was confirmed by pretreatment of cells either with buthionine sulfoximine (inhibitor of GSH synthesis) or N-acetyl-cysteine (precursor in GSH synthesis)