| Bz-423 is a immunomodulatory 1,4-benzodiazepine that exerts its therapeutic effects in autoimmune mice by selectively inducing apoptosis of pathogenic B and T cells. The overall goal of the studies presented in this dissertation is to understand the cellular and molecular basis for the selectivity observed in vivo. Not only will the findings of these experiments be useful in helping advance Bz-423 to the clinic, but will also provide one of the first comprehensive outlines of the apoptotic signaling pathways induced by a small molecule.; Structure-activity studies identified the aromatic side chain at C2 and the C'4 position phenolic hydroxyl group are key elements for target binding and cytotoxic activity. Bz-423 induces an early, rapid production of superoxide (O2-) from mitochondria, and this response is the critical signal initiating apoptosis. The oligomycin sensitivity-conferring protein (OSCP) component of the mitochondrial F1F0-ATPase is the physiologically relevant target for both O2- production and subsequent cell death. JNK is a key molecule involved in Bz-423-induced apoptosis in fibroblasts. JNK activation occurs by a MAPK signaling cascade involving sequential activation of ASK1/MEKK/JNK resulting in activation of the transcription factor, c-Jun. JNK activation causes increased expression of BH3-only proteins triggering the activation of the multidomain pro-apoptotic proteins Bax and Bak. These results provide a detailed description of events during Bz-423-induced apoptosis, from initial O2- production to cytochrome c release and caspase activation.; Bz-423 induces apoptosis in a cellular model of pathogenic lupus through a mechanism unique from that observed in fibroblasts. In all cell types examined apoptotic signaling pathways induced by Bz-423 converge on the Bcl-2 family of proteins resulting in activation of Bax and Bak by BH3-only proteins culminating in cytochrome c release from the mitochondria. However, Bz-423-induced apoptosis in pathogenic lupus cells is independent of MAPK signaling and de novo protein synthesis. These differences result in increased kinetics of cell death. The difference in kinetics of Bz-423-induced cell death in different cell lines coupled with its pharmacokinetic profile could help explain why Bz-423 is selective to pathogenic B cells in vivo . |
