Calpain activation is hypothesized to be an early occurrence in the sequence of events resulting in neurodegeneration, as well as in the signaling pathways linking extracellular accumulation of Aβ peptides and intracellular formation of neurofibrillary tangles. In an effort to identify small molecules that prevent neurodegeneration in Alzheimer’s disease by early intervention in the cell death cascade, a cell-based assay in differentiated Sh-SY5Y cells was developed utilizing calpain activity as a read-out for the early stages of death in cells exposed to extracellular Aβ. This assay was optimized for high-throughput screening, and a library of approximately 120,000 compounds tested. It was expected that the compounds identified as calpain inhibitors would include those that act directly on the enzyme and those that prevented calpain activation by blocking an upstream step in the pathway. In fact, of the compounds that inhibited calpain activation by Aβ with IC50 values < 10 μM and showed little or no toxicity at concentrations up to 30 μM, none inhibit the calpain enzyme directly. Studies to identify the targets of these compounds in the cell death pathway are ongoing.
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