Cytoprotective effect of selective small-molecule caspase inhibitors against staurosporine-induced apoptosis.
Bottom Line: The objective of this study was to discover small-molecule caspase inhibitors with which to achieve cytoprotective effect.Nineteen compounds were found to have significant cytoprotective effects in cell viability assays.DNA microarray analysis demonstrated that staurosporine treatment induced broad global gene expression alterations, and RBC1023 co-treatment significantly restored these changes, especially of the genes that are related to cell growth and survival signaling such as Egr1, Cdc25c, cdkn3, Rhob, Nek2, and Taok1.
Affiliation: Reaction Biology Corp, Malvern, PA, USA.
Caspases are currently known as the central executioners of the apoptotic pathways. Inhibition of apoptosis and promotion of normal cell survival by caspase inhibitors would be a tremendous benefit for reducing the side effects of cancer therapy and for control of neurodegenerative disorders such as Parkinson's, Alzheimer's, and Huntington's diseases. The objective of this study was to discover small-molecule caspase inhibitors with which to achieve cytoprotective effect. We completed the high-throughput screening of Bionet's 37,500-compound library (Key Organics Limited, Camelford, Cornwall, UK) against caspase-1, -3, and -9 and successfully identified 43 initial hit compounds. The 43 hit compounds were further tested for cytoprotective activity against staurosporine-induced cell death in NIH3T3 cells. Nineteen compounds were found to have significant cytoprotective effects in cell viability assays. One of the compounds, RBC1023, was demonstrated to protect NIH3T3 cells from staurosporine-induced caspase-3 cleavage and activation. RBC1023 was also shown to protect against staurosporine-induced impairment of mitochondrial membrane potential. DNA microarray analysis demonstrated that staurosporine treatment induced broad global gene expression alterations, and RBC1023 co-treatment significantly restored these changes, especially of the genes that are related to cell growth and survival signaling such as Egr1, Cdc25c, cdkn3, Rhob, Nek2, and Taok1. Collectively, RBC1023 protects NIH3T3 cells against staurosporine-induced apoptosis via inhibiting caspase activity, restoring mitochondrial membrane potential, and possibly upregulating some cell survival-related gene expressions and pathways.
No MeSH data available.
Related in: MedlinePlus
Mentions: Since the staurosporine/NIH3T3 apoptosis induction system is a very well-established model system for apoptosis-related mechanism studies, we next focused on the cytoprotection mechanism studies in the NIH3T3 cell line. We first investigated the protective effect of different concentrations of RBC1023 against different concentrations of staurosporine-induced cell death. NIH3T3 cells were treated with different dose combinations of RBC1023 (50 μM to 1 μM, three-fold dilution, ten doses) and staurosporine (0.768 μM or 2 μM) for 24 hours before the cell viability was measured. As shown in Figure 3A, RBC1023 started to show significant cytoprotection against the staurosporine-induced cell death at about 2 μM, when the concentration of staurosporine was at 0.768 μM. The cytoprotective effect by RBC1023 increased in a dose-dependent manner. To understand if the pretreatment or posttreatment of the caspase inhibitor would affect its cytoprotective effect, RBC1023 was added to cells at 1 hour before staurosporine treatment or 1 hour post-staurosporine treatment. The cell viability was measured after a total of 24 hours’ incubation. Interestingly, there were no significant differences of cytoprotective activity by RBC1023 between the 1-hour pretreatment and and 1-hour posttreatment group with staurosporine (Figure 3B).
No MeSH data available.