The nfkb1 and nfkb2 genes encode closely related products regulating immune and inflammatory responses. Their role during development and differentiation remains unclear. The generation of nfkb1 null mice (p50-/-) resulted in altered immune responses, but had no effect on development. Similarly, nfkb2 knockout mice (p52-/-) did not show developmental defects (J.C. et al., manuscript submitted). We have investigated the potential for in vivo compensatory functions of these genes by generating double-knockout mice. The surprising result was that the animals developed osteopetrosis because of a defect in osteoclast differentiation, suggesting redundant functions of NF-kappaB1 and NF-kappaB2 proteins in the development of this cell lineage. The osteopetrotic phenotype was rescued by bone marrow transplantation, indicating that the hematopoietic component was impaired. These results define a new mouse osteopetrotic mutant and implicate NF-kappaB proteins in bone development, raising new directions in the treatment of bone disorders.

Chemokines are a structurally related family of cytokines that are important for leukocyte trafficking. The C-C chemokine monocyte chemoattractant protein-1 (MCP-1) is a potent monocyte activator in vitro and has been associated with monocytic infiltration in several inflammatory diseases. One C-C chemokine receptor, CCR2, has been identified that mediates in vitro responses to MCP-1 and its close structural homologues. CCR2 has also recently been demonstrated to be a fusion cofactor for several HIV isolates. To investigate the normal physiological function of CCR2, we generated mice with a targeted disruption of the ccr2 gene. Mice deficient for CCR2 developed normally and had no hematopoietic abnormalities. However, ccr2 −/− mice failed to recruit macrophages in an experimental peritoneal inflammation model. In addition, these mice were unable to clear infection by the intracellular bacteria, Listeria monocytogenes. These results suggest that CCR2 has a nonredundant role as a major mediator of macrophage recruitment and host defense against bacterial pathogens and that MCP-1 and other CCR2 ligands are effectors of those functions.

␥-Secretase inhibitors are one promising approach to the development of a therapeutic for Alzheimer's disease (AD). ␥-Secretase inhibitors reduce brain ␤-amyloid peptide (A␤), which is believed to be a major contributor in the etiology of AD. Transgenic mice overexpressing the human ␤-amyloid precursor protein (APP) are valuable models to examine the dynamics of A␤ changes with ␥-secretase inhibitors in plaque-free and plaque-bearing animals. BMS-299897 2-[(1R)-1-[[(4-chlorophenyl)sulfony](2,5-difluorophenyl)amino]ethyl]-5-fluorobenzenepropanoic acid, a ␥-secretase inhibitor, showed doseand time dependent reductions of A␤ in brain, cerebrospinal fluid (CSF), and plasma in young transgenic mice, with a significant correlation between brain and CSF A␤ levels. Because CSF and brain interstitial fluid are distinct compartments in composition and location, this correlation could not be assumed. In contrast, aged transgenic mice with large accumulations of A␤ in plaques showed reductions in CSF A␤ in the absence of measurable changes in plaque A␤ in the brain after up to 2 weeks of treatment. Hence, CSF A␤ levels were a valuable measure of ␥-secretase activity in the central nervous system in either the presence or absence of plaques. Transgenic mice were also used to examine potential side effects due to Notch inhibition. BMS-299897 was 15-fold more effective at preventing the cleavage of APP than of Notch in vitro. No changes in the maturation of CD8ϩ thymocytes or of intestinal goblet cells were observed in mice treated with BMS-299897, showing that it is possible for ␥-secretase inhibitors to reduce brain A␤ without causing Notch-mediated toxicity.Diagnosis of AD is based on key pathological features at autopsy in the presence of dementia: loss of neuronal mass, intracellular tangles, and extracellular plaques. Plaques are composed predominantly of A␤. A␤ has both N-and C-terminal heterogeneity, with the C terminus usually ending at residues 40 or 42. Although A␤40 is 80 to 90% of the total, A␤42 is most associated with disease and is predominant in plaques. A␤ is produced by two cleavage events in APP (Wolfe, 2001). ␤-Secretase, now identified as ␤-site APP cleaving enzyme, makes the initial N-terminal cleavage, releasing a soluble N-terminal form of APP. The remaining, membrane-bound C-terminal fragment (CTF) of APP is cleaved by ␥-secretase to release soluble A␤ along with the APP intracellular domain. Alternatively, APP can This research was supported by Bristol-Myers Squibb and SIBIA Neurosciences, Inc.Article, publication date, and citation information can be found at

The discovery of asunaprevir (BMS-650032, 24) is described. This tripeptidic acylsulfonamide inhibitor of the NS3/4A enzyme is currently in phase III clinical trials for the treatment of hepatitis C virus infection. The discovery of 24 was enabled by employing an isolated rabbit heart model to screen for the cardiovascular (CV) liabilities (changes to HR and SNRT) that were responsible for the discontinuation of an earlier lead from this chemical series, BMS-605339 (1), from clinical trials. The structure-activity relationships (SARs) developed with respect to CV effects established that small structural changes to the P2* subsite of the molecule had a significant impact on the CV profile of a given compound. The antiviral activity, preclincial PK profile, and toxicology studies in rat and dog supported clinical development of BMS-650032 (24).

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