András Gyovai

In vitro tests of cellular activity form part of the diagnostic algorithm of drug hypersensitivity reactions. Because of the wide range of pharmacological mechanisms, clinical symptoms, genetic components, and laboratory tests involved, it is important to know how a particular test performs in the diagnostic procedure. We carried out a detailed retrospective analysis of more than 6000 measurements of numerous drug compounds tested in 738 serum samples over the past 6 years. Our cell viability-based lymphocyte transformation had a coefficient of variation of 10% and showed similar performance over the whole range of tested ages. With an adequate number of parallel measurements, the test can identify modest increases in stimulation indices with high confidence. Similar percentages of analytically positive responses (11.4%, 13.5%, and 9.7%) were observed for the three most frequently tested drug groups, namely, antibiotics, non-steroid anti-inflammatory agents, and anesthetics. These results confirm that cell viability tests are suitable alternatives for proliferation assays in drug allergy testing.

In vitro tests of cellular activity form part of the diagnostic algorithm of drug hypersensitivity reactions. Because of the wide range of pharmacological mechanisms, clinical symptoms, genetic components and laboratory tests, it is important to know how a particular test performs in the diagnostic procedure. We carried out the detailed retrospective analysis of more than 6,000 measurements of numerous drug compounds tested in the past 6 years. Our cell viability-based lymphocyte transformation had a coefficient of variation of 10% and showed similar performance over the whole range of tested ages. With an adequate number of parallel measurements, the test can identify modest increases in stimulation indices with high confidence. Similar percentages of analytically positive responses were observed for the three most frequently tested drug groups, namely antibiotics, non-steroid anti-inflammatory agents and anesthetics. These results confirm that cell viability tests are suitable alternatives of proliferation assays in drug allergy testing.

19-Nortestosterone C-17 epimers with prominent antiproliferative properties have been previously described. In our present study, five novel 17α-19-nortestosterones (3−7) were synthesized to increase their beneficial biological activities with no associated undesired hormonal effects. The compounds were screened by a viability assay against a panel of human adherent gynecological cancer cell lines. Three of the tested derivatives (3−5) exhibited a remarkable inhibitory effect on the proliferation of HeLa cells with IC₅₀ values lower than that of our reference agent cisplatin (CIS). These three active agents also displayed considerable cancer selectivity as evidenced by their weaker growth inhibitory effect on non-cancerous fibroblast cells compared to CIS. The most potent newly synthesized 17α-chloro derivative (3) was selected for additional experiments in order to characterize its mechanism of action. Since nandrolone (19-nortestosterone, 1) is a structural analog with selective antiproliferative action on cervical carcinoma cells, it was utilized as a positive control in these studies. A lactate dehydrogenase (LDH) assay demonstrated a moderate cytotoxic effect of the test compounds. Cell cycle disturbance and the elevation of the hypodiploid population elicited by the test agents were detected by flow cytometry following propidium staining. The proapoptotic effects of the tested steroids were confirmed by fluorescent microscopy and a caspase-3 activity assay. Treatment-related caspase-9 activation without a substantial change in caspase-8 activity indicates the induction of the intrinsic apoptotic pathway. The selected agents directly influence the rate of tubulin assembly as evidenced by a polymerization assay. Yeast-based reporter gene assay revealed that the androgenic activity of the novel 19-nortestosterone derivative 3 is by multiple orders of magnitude weaker than that of the reference agent 1. Based on the behavior of the examined compounds it can be concluded that a halogen substitution of the 19-nortestosterone scaffold at the 17α position may produce compounds with unique biological activities. The results of the present study support that structurally modified steroids with negligible hormonal activity are a promising basis for the research and development of novel anticancer agents.

One-hundred and sixty-eight aqueous and organic extracts of 42 selected bryophyte species were screened in vitro for antiproliferative activity on a panel of human gynecological cancer cell lines containing HeLa (cervix epithelial adenocarcinoma), A2780 (ovarian carcinoma), and T47D (invasive ductal breast carcinoma) cells using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and for antibacterial activity on 11 strains using the disc-diffusion method. A total of 99 extracts derived from 41 species exerted ≥25% inhibition of proliferation of at least one of the cancer cell lines at 10 μg/mL. In the cases of Brachythecium rutabulum, Encalypta streptocarpa, Climacium dendroides, Neckera besseri, Pleurozium schreberi, and Pseudoleskeella nervosa, more than one extract was active in the antiproliferative assay, whereas the highest activity was observed in the case of Paraleucobryum longifolium. From the tested families, Brachytheciaceae and Amblystegiaceae provided the highest number of antiproliferative extracts. Only 19 samples of 15 taxa showed moderate antibacterial activity, including the most active Plagiomnium cuspidatum, being active on 8 tested strains. Methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus aureus were the most susceptible to the assayed species. This is the first report on the bioactivities of these 14 species.

Novel D- and A-ring-fused quinolines in the estrone and 5α-androstane series were efficiently synthesized from the corresponding β-chlorovinyl aldehydes with different arylamines in DMF under microwave irradiation. The rates of the one-pot catalyst-free syntheses and the yields of the desired products were found to be affected significantly by the electronic and steric character of the substituents on the anilines and the different reactivities of rings D and A of the sterane skeleton. All the synthesized compounds were tested in vitro on human cervical (C33A, HeLa and SiHa) and breast (MCF-7, MDA-MB-231, MDA-MB-361 and T47D) cancer cell lines in order to investigate their antiproliferative activities in vitro. Evidence of cell cycle blockade and apoptosis induction was obtained for the most effective compound 14c by means of flow cytometry, caspase-3 activity determination and microscopic techniques.