From the authors'650,000 compound collection, they have selected approximately 15,000 potential small-molecule protease inhibitors, which were subjected to high-throughput screening against caspase-3. The screening yielded a series of hits that belong to 11 different scaffolds. Based on the structure of one of the hits, a new class of the small-molecule inhibitors with a double electrophilic warhead, 8-sulfonyl-pyrrolo[3,4-c]quinoline-1,3-diones (SPQ), was synthesized and tested in follow-up mechanistic and antiapoptosis assays. Mechanistic analysis of a representative compound of this class, CD-001-0011, showed that the compound exhibited a high potency (IC 50 = 130 nM), was reversible though noncompetitive, and had a broad selectivity profile to other caspases belonging to groups I to III. The compound was effective in preventing staurosporineinduced apoptosis in a few cell lines and retinoic acid-induced apoptosis in zebrafish. (Journal of Biomolecular Screening 2006:277-285)

No abstract

When studying cysteinyl proteases in general and caspases in particular, it is generally accepted that a reaction buffer must contain a reducing agent to prevent essential cysteinyl groups from spontaneous oxidation. Dithiothreitol (DTT) and beta-mercaptoethanol (beta-MCE) are 2 of the most broadly used reducing agents. While screening a library of small molecules against caspase-3, the authors have found that the nature of the reducing agent used, DTT or beta-MCE, dramatically affects screening results and leads to identification of nonoverlapping hits. Screening in DTT-containing buffer revealed few novel classes of small molecules that selectively and reversibly inhibit caspase-3 but failed to identify isatin sulfonamides recently found to be potent and selective caspase-3 inhibitors (false negatives). On the other hand, screening in the presence of beta-MCE failed to identify a series of hit compounds, 1,3-dioxo-2,3-dichloro-1H-pyrrolo[3,4-c]quinolines, discovered with DTT, whereas isatin sulphonamides in these conditions exhibited strong caspase-3 inhibition. In this work, the authors show that thiol-containing reducing agents can affect catalytic activity of caspase-3 and modify its thermostability in a redox-potential-independent manner. The authors speculate that the differential structural modifications of caspase-3 seen with different reducing agents represent structurally different caspase-3 conformations and are responsible for its differential sensitivity to small molecules of different chemotypes. Hence, selection of the reducing agent may dramatically affect the quality of high-throughput screening campaigns.

No abstract

The results of in vitro studies demonstrate cytotoxic activity alantolactone and isoalantolactone, presented in the elecampane plant (Inula helenium) of the natural sesquiterpene lactones. These compounds show toxicity against human cancer cell lines: MS, A549, HCT116, MCF7, RD and K562. The involvement of the p53 signaling pathway in the death of tumor cells, as well as the contribution of reactive oxygen species (ROS) formation during cell death, was studied.