A New Generation XIAP Inhibitor Induces Apoptosis of Human Myeloma Cells and Synergises with Conventional and Novel Anti-Myeloma Therapeutics.

We have demonstrated that a new generation of IAP inhibitor (Novartis) induces apoptosis of a range (n = 8) of genetically heterogeneous human myeloma cell lines (HMCLs) in a dose and time dependent fashion. Dose responsiveness to the IAP inhibitor (IAPI) was determined via MTS assays using IAPI at doses 5 to 50μM for 24 and 72 hours with IC50s of 25 to 50μM evident in 7 out of 8 HMCLs at 72 hours. Mechanistic studies were undertaken utilising 3 HMCLs with high (NCI H929), intermediate (LP-1) or low (OPM2) sensitivity to IAPI with QRT-PCR demonstrating the highest level of XIAP expression in the most IAPI sensitive HMCL NCI H929. Following IAPI exposure immunoblot analyses of all 3 HMCLs exhibited rapid falls in ICAD and CAD as well as cleavage of ROCK-1 and PARP, confirming apoptosis induction. IAPI - induced apoptosis was mediated via both caspase 7 and caspase 3 as demonstrated by rapid cleavage of both caspases following IAPI exposure. Studies combining IAPI with conventional therapies (bortezomib plus doxorubicin) and (bortezomib plus etoposide) against all 3 HMCLs demonstrated synergistic killing with the optimum sequence (IAPI administered 24 hours before or after drug partners) varying between the 3 HMCLs. Similarly, TNF-related-apoptosis-inducing-ligand (TRAIL) and IAPI also demonstrated synergistic killing irrespective of drug sequence. Finally, IAPI combined with LBY135 (Novartis), a chimeric agonistic TRAIL-R2 monoclonal antibody, demonstrated synergistic killing maximally with LBY135 for 24 hours followed by IAPI rather than the reverse sequence. We conclude that IAPI induces down-regulation of both ICAD/CAD signalling as well as ROCK-1 and PARP cleavage via a caspase 3 and 7 dependent process. Furthermore, our data suggest that IAPI is synergistic with a range of conventional and a novel therapeutic agent. While the mechanism of XIAP inhibition in MM warrants further clarification IAPI clearly represents a potentially novel therapeutic approach to MM.