A series of new Smac mimetics have been designed, synthesized and evaluated. The most potent compound 10 binds to XIAP, cIAP-1 and cIAP-2 BIR3 proteins with K i values of 3.9, 0.37 and 0.25 nM, respectively. Compound 10 antagonizes XIAP in a cell-free functional assay and induces rapid cIAP-1 degradation in cancer cells. Compound 10 inhibits cell growth in the MDA-MB-231 cancer cell line with an IC 50 value of 8.9 nM.Apoptosis is a critical cell process in normal development and homeostasis of multicellular organisms to eliminate unwanted or damaged cells. Evasion of apoptosis is now recognized as a hallmark of all cancers. 1-2 Targeting key apoptosis regulators with a goal to promote apoptosis in cancer cells is a new strategy for anticancer drug design. [3][4] Inhibitor of apoptosis proteins (IAPs) a are a class of key apoptosis regulators. 5,6 Among them, cellular IAP-1 (cIAP-1) and cIAP-2 play a critical role in regulation of tumor necrosis factor (TNF) receptor-mediated apoptosis, 7 and X-linked IAP (XIAP) is a central regulator of both death-receptor-mediated and mitochondria-mediated apoptosis pathways. 8 XIAP inhibits apoptosis through direct binding to and inhibition of three cysteine proteases, an initiator caspase-9 and the two effectors caspase-3 and -7. 5,6 XIAP contains three Baculoviral IAP Repeats (BIR) domains. While the third BIR domain (BIR3) of XIAP selectively targets caspase-9, the BIR2 domain, together with the immediate preceding linker, inhibits both caspase-3 and caspase-7. 6,8 Since these caspases play a critical role in the execution of apoptosis, XIAP functions as an efficient inhibitor of apoptosis. 6,8 Accordingly, these IAP proteins, especially XIAP, are considered as promising cancer therapeutic targets. [8][9] Smac (Second Mitochondria-derived Activator of Caspases) is a potent pro-apoptotic protein and an endogenous antagonist of IAP proteins. 10,11 Previous studies have firmly established that Smac interacts with XIAP and cIAP-1/2 proteins via its AVPI tetra-peptide motif. 15 Smac, in its dimeric form, interacts with both BIR2 and BIR3 domains in XIAP and effectively removes the inhibition of XIAP to not only caspase-9 but also caspase-3/-7.In the last few years, intense research efforts have been devoted to the design and development of small molecules to mimic the AVPI binding motif as antagonists of IAP proteins and as a new class of anticancer drugs. 4,[16][17][18][19][20][21][22][23] These small molecules are called Smac mimetics. To date, two different types of Smac mimetics have been reported, namely monovalent and bivalent Smac mimetics ( Figure 1). 4,[16][17][18][19][20][21][22][23] Monovalent Smac mimetics such as 1-4 are designed to mimic a single AVPI binding motif, [16][17][18][19] whereas the bivalent compounds, such as 5 and 6, contain mimetics of two AVPI binding motifs tethered together through a linker. 4,[20][21][22][23] Bivalent Smac mimetics can achieve much higher affinities to XIAP and can be much more potent than the corresponding monovalent Sma...