Inhibitor of apoptosis proteins (IAPs) are important regulators of apoptosis and pro-survival signaling pathways whose deregulation is often associated with tumor genesis and tumor growth. IAPs have been proposed as targets for anticancer therapy, and a number of peptidomimetic IAP antagonists have entered clinical trials. Using our fragment-based screening approach, we identified nonpeptidic fragments binding with millimolar affinities to both cellular inhibitor of apoptosis protein 1 (cIAP1) and X-linked inhibitor of apoptosis protein (XIAP). Structure-based hit optimization together with an analysis of protein-ligand electrostatic potential complementarity allowed us to significantly increase binding affinity of the starting hits. Subsequent optimization gave a potent nonalanine IAP antagonist structurally distinct from all IAP antagonists previously reported. The lead compound had activity in cell-based assays and in a mouse xenograft efficacy model and represents a highly promising start point for further optimization.

XIAP and cIAP1 are members of the inhibitor of apoptosis protein (IAP) family and are key regulators of anti-apoptotic and pro-survival signaling pathways. Overexpression of IAPs occurs in various cancers and has been associated with tumor progression and resistance to treatment. Structure-based drug design (SBDD) guided by structural information from X-ray crystallography, computational studies, and NMR solution conformational analysis was successfully applied to a fragment-derived lead resulting in AT-IAP, a potent, orally bioavailable, dual antagonist of XIAP and cIAP1 and a structurally novel chemical probe for IAP biology.

Aberrant activation of the MAPK pathway drives cell proliferation in multiple cancers. Inhibitors of BRAF and MEK kinases are approved for the treatment of BRAF mutant melanoma, but resistance frequently emerges, often mediated by increased signaling through ERK1/2. Here, we describe the fragment-based generation of ERK1/2 inhibitors that block catalytic phosphorylation of downstream substrates such as RSK but also modulate phosphorylation of ERK1/2 by MEK without directly inhibiting MEK. X-ray crystallographic and biophysical fragment screening followed by structure-guided optimization and growth from the hinge into a pocket proximal to the C-α helix afforded highly potent ERK1/2 inhibitors with excellent kinome selectivity. In BRAF mutant cells, the lead compound suppresses pRSK and pERK levels and inhibits proliferation at low nanomolar concentrations. The lead exhibits tumor regression upon oral dosing in BRAF mutant xenograft models, providing a promising basis for further optimization toward clinical pERK1/2 modulating ERK1/2 inhibitors.

The DDR1 and DDR2 receptor tyrosine kinases are activated by extracellular collagen and have been implicated in a number of human diseases including cancer. We performed a fragment-based screen against DDR1 and identified fragments that bound either at the hinge or in the back pocket associated with the DFG-out conformation of the kinase. Modeling based on crystal structures of potent kinase inhibitors facilitated the "back-to-front" design of potent DDR1/2 inhibitors that incorporated one of the DFG-out fragments. Further optimization led to low nanomolar, orally bioavailable inhibitors that were selective for DDR1 and DDR2. The inhibitors were shown to potently inhibit DDR2 activity in cells but in contrast to unselective inhibitors such as dasatinib, they did not inhibit proliferation of mutant DDR2 lung SCC cell lines. KEYWORDS: discoidin domain receptor, fragment-based drug design, back to front kinase design D iscoidin domain receptors, DDR1 and DDR2, are transmembrane receptor tyrosine kinases that are activated by collagen binding to their extracellular domain. 1,2 DDR1 and DDR2 have been associated with extracellular remodeling, cell adhesion, proliferation and migration, and they have been linked to a number of human diseases, including fibrotic disorders, atherosclerosis and cancer. 3−5 There has recently been evidence suggesting that DDR2 inhibitors would be useful for the treatment of lung squamous cell cancer 6,7 although the role of DDR2 may be more complex than first realized. 8−10 Hammerman et al. 6,7 have shown that DDR2 is mutated in approximately 4% of lung squamous cell cancer and have presented data to suggest that these are gain of function mutations. Hammerman et al. 6 have also shown that cell lines harboring these mutations are selectively sensitized through knockdown of DDR2 by RNA interference or by treatment of the multitargeted kinase inhibitor dasatinib. 11 Selective DDR2 inhibitors would therefore be useful to probe the role of DDR2 and may have utility for the treatment of lung cancer. To date, most DDR1/2 inhibitors have been derived from cross-screening of existing kinase inhibitors. 12 Initial compounds often lacked selectivity over homologous enzymes such as Bcr-Abl, but a number of recent papers have described more selective inhibitors which all bind to the DFG-out form of the enzyme. 13−15 Gray and co-workers have identified DDR1/2 inhibitors by screening a library of compounds that had previously been designed by mixing and matching motifs from known Type II kinase inhibitors. 14 The resulting compounds were reported to be selective with IC 50 values versus DDR1 and DDR2 of approximately 100 nM, and experimental binding modes in DDR1 were determined. 14,16 Gao et al. have published a series of potent Type II DDR1 inhibitors that are selective over other kinases, including DDR2. 13 In this Letter we describe a fragment based approach to the discovery of potent and selective DDR1/2 inhibitors. We first obtained a soakable crystal form of DDR1 suitable for high-throughput cry...

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