Inhibitor of apoptosis (IAP) proteins are antiapoptotic regulators that block cell death in response to diverse stimuli. They are expressed at elevated levels in human malignancies and are attractive targets for the development of novel cancer therapeutics. Herein, we demonstrate that small-molecule IAP antagonists bind to select baculovirus IAP repeat (BIR) domains resulting in dramatic induction of auto-ubiquitination activity and rapid proteasomal degradation of c-IAPs. The IAP antagonists also induce cell death that is dependent on TNF signaling and de novo protein biosynthesis. Additionally, the c-IAP proteins were found to function as regulators of NF-kappaB signaling. Through their ubiquitin E3 ligase activities c-IAP1 and c-IAP2 promote proteasomal degradation of NIK, the central ser/thr kinase in the noncanonical NF-kappaB pathway.

Glyburide, a sulfonylurea drug commonly used to treat type 2 diabetes, shuts down IL-1β secretion by preventing Cyropyrin activation.

Cellular IAP12 and IAP2 (c-IAP1 and c-IAP2) were identified in a search for proteins associated with TNF receptors (TNFRs) (1). Through binding to TNFR-associated factor 2 (TRAF2), c-IAP1 and c-IAP2 are recruited to TNFR signaling complexes, where they regulate the activation of caspase-8 (1, 2). c-IAP1 and c-IAP2 were also proposed to modulate activation of the canonical NF-B pathway, although most of these studies relied on overexpression (3, 4). In contrast, however, targeted deletion of c-IAP1 or c-IAP2 genes in mice did not reveal any abnormalities in TNF␣-induced NF-B (nuclear factor B) activation (5, 6). The absence of any appreciable phenotype in single c-IAP knock-out mice has been attributed to the putative redundancy of c-IAP1 and c-IAP2 due to their high level of sequence and functional similarities (7). Thus, combined deficiency of cellular IAPs might expose their role in this signaling pathway. In support of this possibility, a null mutation in the sole cellular IAP in zebrafish results in severe defects in NF-B activation (8). c-IAP1 and c-IAP2 are also RING domain-containing ubiquitin ligases capable of promoting ubiquitination of several of their binding partners, including TRAF2 and SMAC (second mitochondrial activator of caspases) (4, 5, 9 -12).TNFR1 mediates activation of several signaling pathways, among them the canonical NF-B pathway (13). Binding of TNF␣ to TNFR1 induces recruitment of the adaptor protein TNFR-associated death domain (TRADD) to the death domain of the receptor (14). Through its death domain and amino-terminal region, TRADD recruits RIP1 (receptor-interacting protein), TRAF2, and through its interaction with TRAF2, c-IAP1 and c-IAP2 (13). Following binding to TRADD, TRAF2 was thought to mediate non-degradative Lys-63-linked polyubiquitination of RIP1 via its RING E3 ligase domain (15,16). This RIP1 modification induces assembly of two RIP1-associated kinase complexes, TAK1-TABs (transforming growth factor ␤-activated kinase 1-TAK1-binding proteins) and IB kinase (IKK) (17)(18)(19). Binding of these two complexes to Lys-63-linked polyubiquitin chains on RIP1 leads to phosphorylation of IKK␤ and subsequent phosphorylation and proteasomal degradation of IB (20). Loss of IB allows translocation of p50/RelA dimer to the nucleus and induction of gene expression (20).In the present study, we investigate the role of c-IAP1 and c-IAP2 in TNF␣-induced NF-B activation. We discover that c-IAP proteins are important mediators of canonical NF-B signaling and demonstrate that the absence of c-IAPs severely attenuates TNF␣-induced NF-B activation. Finally, we show that c-IAPs are ubiquitin ligases capable of promoting polymerization of Lys-63-linked polyubiquitin chains on the critical adapter in the canonical NF-B signaling pathway, RIP1. was from Genentech, Inc. The primary antibodies against mouse c-IAP1 were kindly provided by Drs. John Silke and David Vaux; anti-human c-IAP1 antibodies were purchased from R&D (affinity-purified goat antibody) or Protein Tech Group Inc.; pan c-IAP1/2 ...

The prosurvival BCL-2 family protein BCL-X(L) is often overexpressed in solid tumors and renders malignant tumor cells resistant to anticancer therapeutics. Enhancing apoptotic responses by inhibiting BCL-X(L) will most likely have widespread utility in cancer treatment and, instead of inhibiting multiple prosurvival BCL-2 family members, a BCL-X(L)-selective inhibitor would be expected to minimize the toxicity to normal tissues. We describe the use of a high-throughput screen to discover a new series of small molecules targeting BCL-X(L) and their structure-guided development by medicinal chemistry. The optimized compound, WEHI-539 (7), has high affinity (subnanomolar) and selectivity for BCL-X(L) and potently kills cells by selectively antagonizing its prosurvival activity. WEHI-539 will be an invaluable tool for distinguishing the roles of BCL-X(L) from those of its prosurvival relatives, both in normal cells and notably in malignant tumor cells, many of which may prove to rely upon BCL-X(L) for their sustained growth.

No abstract