CXCR7 binds chemokines CXCL11 (I-TAC) and CXCL12 (SDF-1) but does not act as a classical chemoattractant receptor. Using CCX771, a novel small molecule with high affinity and selectivity for CXCR7, we found that, although CXCR7 is dispensable for “bare filter” in vitro chemotaxis, CXCR7 plays an essential role in the CXCL12/CXCR4-mediated transendothelial migration (TEM) of CXCR4+CXCR7+ human tumor cells. Importantly, although CXCL11 is unable to stimulate directly the migration of these cells, it acts as a potent antagonist of their CXCL12-induced TEM. Furthermore, even though this TEM is driven by CXCR4, the CXCR7 ligand CCX771 is substantially more potent at inhibiting it than the CXCR4 antagonist AMD3100, which is more than 100 times weaker at inhibiting TEM when compared with its ability to block bare filter chemotaxis. Far from being a “silent” receptor, we show that CXCR7 displays early hallmark events associated with intracellular signaling. Upon cognate chemokine binding, CXCR7 associates with β-arrestin2, an interaction that can be blocked by CXCR7-specific mAbs. Remarkably, the synthetic CXCR7 ligand CCX771 also potently stimulates β-arrestin2 recruitment to CXCR7, with greater potency and efficacy than the endogenous chemokine ligands. These results indicate that CXCR7 can regulate CXCL12-mediated migratory cues, and thus may play a critical role in driving CXCR4+CXCR7+ tumor cell metastasis and tissue invasion. CXCR7 ligands, such as the chemokine CXCL11 and the newly described synthetic molecule CCX771, may represent novel therapeutic opportunities for the control of such cells.

Necrotizing and crescentic GN (NCGN) with a paucity of glomerular immunoglobulin deposits is associated with ANCA. The most common ANCA target antigens are myeloperoxidase (MPO) and proteinase 3. In a manner that requires activation of the alternative complement pathway, passive transfer of antibodies to mouse MPO (anti-MPO) induces a mouse model of ANCA NCGN that closely mimics human disease. Here, we confirm the importance of C5aR/CD88 in the mediation of anti-MPO-induced NCGN and report that C6 is not required. We further demonstrate that deficiency of C5a-like receptor (C5L2) has the reverse effect of C5aR/CD88 deficiency and results in more severe disease, indicating that C5aR/CD88 engagement enhances inflammation and C5L2 engagement suppresses inflammation. Oral administration of CCX168, a small molecule antagonist of human C5aR/CD88, ameliorated anti-MPO-induced NCGN in mice expressing human C5aR/CD88. These observations suggest that blockade of C5aR/CD88 might have therapeutic benefit in patients with ANCA-associated vasculitis and GN.

Microtubules are linear polymers of ␣-and ␤-tubulin heterodimers and are the major constituents of mitotic spindles, which are essential for the separation of chromosomes during mitosis. Here we describe a synthetic compound, 2-fluoro-1-methoxy-4-pentafluorophenylsulfonamidobenzene (T138067), which covalently and selectively modifies the ␤ 1 , ␤ 2 , and ␤ 4 isotypes of ␤-tubulin at a conserved cysteine residue, thereby disrupting microtubule polymerization. Cells exposed to T138067 become altered in shape, indicating a collapse of the cytoskeleton, and show an increase in chromosomal ploidy. Subsequently, these cells undergo apoptosis. Furthermore, T138067 exhibits cytotoxicity against tumor cell lines that exhibit substantial resistance to vinblastine, paclitaxel, doxorubicin, and actinomycin D. T138067 is also equally efficacious in inhibiting the growth of sensitive and multidrug-resistant human tumor xenografts in athymic nude mice. These observations suggest that T138067 may be clinically useful for the treatment of multidrug-resistant tumors.

The chemokine system represents a diverse group of G proteincoupled receptors responsible for orchestrating cell recruitment under both homeostatic and inflammatory conditions. Chemokine receptor 9 (CCR9) is a chemokine receptor known to be central for migration of immune cells into the intestine. Its only ligand, CCL25, is expressed at the mucosal surface of the intestine and is known to be elevated in intestinal inflammation. To date, there are no reports of small-molecule antagonists targeting CCR9. We report, for the first time, the discovery of a small molecule, CCX282-B, which is an orally bioavailable, selective, and potent antagonist of human CCR9. CCX282-B inhibited CCR9-mediated Ca 2ϩ mobilization and chemotaxis on Molt-4 cells with IC 50 values of 5.4 and 3.4 nM, respectively. In the presence of 100% human serum, CCX282-B inhibited CCR9-mediated chemotaxis with an IC 50 of 33 nM, and the addition of ␣1-acid glycoprotein did not affect its potency. CCX282-B inhibited chemotaxis of primary CCR9-expressing cells to CCL25 with an IC 50 of 6.8 nM. CCX282-B was an equipotent inhibitor of CCL25-directed chemotaxis of both splice forms of CCR9 (CCR9A and CCR9B) with IC 50 values of 2.8 and 2.6 nM, respectively. CCX282-B also inhibited mouse and rat CCR9-mediated chemotaxis. Inhibition of CCR9 with CCX282-B results in normalization of Crohn's disease such as histopathology associated with the TNF ⌬ARE mice. Analysis of the plasma level of drug associated with this improvement provides an understanding of the pharmacokinetic/ pharmacodynamic relationship for CCR9 antagonists in the treatment of intestinal inflammation.

Novel non-nucleoside inhibitors of the HCV RNA polymerase (NS5b) with sub-micromolar biochemical potency have been identified which are selective for the inhibition of HCV NS5b over other polymerases. The structures of the complexes formed between several of these inhibitors and HCV NS5b were determined by X-ray crystallography, and the inhibitors were found to bind in an allosteric binding site separate from the active site. Structure-activity relationships and structural studies have identified the mechanism of action for compounds in this series, several of which possess drug-like properties, as unique, reversible, covalent inhibitors of HCV NS5b.