GX015-070 inhibits Mcl-1/Bax interaction, up-regulates Bim protein expression, and induces cytochrome c release. (A) HMCLs were incubated with vehicle control (−) or 1 μM GX015-070 (GX; +) for 24 hours. Cells were lysed in lysis buffer (KMS12PE, JJN3) or CHAPS buffer (MY5) and protein immunoprecipitated with anti-Bak or anti-Bax, where indicated, and immunocomplexes were analyzed by Western with either anti–Mcl-1 or anti–Bcl-2. GX015-070 inhibits Mcl-1/Bak but not Bcl-2/Bax interaction. (B) HMCLs were treated with 1 μM GX015-070 for times indicated. Total cell lysates were probed by Western blot with anti-Bim. A time-dependent increase in Bim protein was observed in JJN3, KMS11, and MY5 cells. (C) Cytosolic extracts from GX015-070–treated (GX; +) or vehicle–treated (−) HMCLs were probed with anti–cytochrome c and demonstrate release of cytochrome c after 24-hour exposure to GX015-070. An increase in cleaved caspase-3 was also observed after 24- and 48-hour treatment with GX015-070.