A series of 5′-phenyl-3′H-spiro[indoline-3,2′-[1,3,4]thiadiazol]-2-one analogs were synthesized and their Bcl-2 protein inhibitory activities were studied. The lead compound was originally identified using a fluorescence polarization-based competitive binding assay. Among the 10 compounds investigated, 1k showed good binding affinities to Bcl-xL and Mcl-1, with inhibition constants of 8.9 μmol/L and 3.4 μmol/L, respectively. While compound 1c achieved tight binding affinities to Bcl-xL (Ki = 0.16 μmol/L), has the potential to be a new lead compound.A series of 5′-phenyl-3′H-spiro[indoline-3,2′-[1,3,4] thiadiazol]-2-one analogs were synthesized and screened for their Bcl-2 protein inhibitory activities. The lead compound was originally identified using a fluorescence polarization-based competitive binding assay. Among the 10 compounds investigated, 1k showed good binding affinities to Bcl-xL and Mcl-1, with inhibition constants of 8.9 μmol/L and 3.4 μmol/L, respectively. While compound 1c achieved tight binding affinities to Bcl-xL (Ki = 0.16 μmol/L), has the potential to be a new lead compound.