Ac-DNLD-CHO is a novel caspase-3 specific peptide inhibitor that was rationally designed by our computational strategy. The specificity was shown to be due to the specific interaction of NLD moiety with the active site of caspase-3 on the basis of docking mode and site-directed mutagenesis analyses. Here, we computationally screened non-peptidic small molecular inhibitors of caspase-3 from our chemical library using a reliable pharmacophore derived from the specific binding mode of NLD. Through in vitro enzyme assay of the screened candidate compounds, we discovered a novel caspase-3 specific small molecular inhibitor, CS4566, which has a unique scaffold structure. The binding mode of CS4566 to caspase-3 mimics that of NLD, especially LD moiety. This represents a promising lead compound for creating non-peptidic pharmaceuticals for caspase-mediated diseases, such as neurodegenerative disorders.