Rational design of novel diaryl ether-linked benzimidazole derivatives as potent and selective BACE1 inhibitors.

Due to the large size and high flexibility of the catalytic active site of BACE1 enzyme, the development of nonpeptide inhibitors with optimal pharmacological properties is still highly demanding. In this work, we have discovered 2-aminobenzimidazole-containg ether scaffolds having potent and selective inhibitory potentials against BACE1 enzyme. We have synthesized novel 29 compounds and optimization of aryl linker region resulted in highly potent BACE1 inhibitory activities with EC50 values of 0.05-2.71 µM. The aryloxy-phenyl analogs 20j showed the EC50 value as low as 0.07 µM in the enzyme assay, whereas, the benzyloxyphenyl dervative 24b was comparatively less effective in the enzyme assay. But interestingly the latter was more effective in the cell assay (EC50 value 1.2 µM). While comparing synthesized derivatives in the cell assay using PC12-APPSW cell, compound 27f appeared as the most potent BACE1 inhibitor having EC50 value 0.7 µM. This scaffold also showed high selectivity over BACE2 enzyme and cathepsin D. Furthermore, the research findings were bolstered through the incorporation of molecular docking, molecular dynamics, and DFT studies. We firmly believe that these discoveries will pave the way for the development of a novel class of small-molecule selective BACE1 inhibitors.

Suppression of NFAT5-mediated Inflammation and Chronic Arthritis by Novel κB-binding Inhibitors.

Nuclear factor of activated T cells 5 (NFAT5) has been implicated in the pathogenesis of various human diseases, including cancer and arthritis. However, therapeutic agents inhibiting NFAT5 activity are currently unavailable. To discover NFAT5 inhibitors, a library of >40,000 chemicals was screened for the suppression of nitric oxide, a direct target regulated by NFAT5 activity, through high-throughput screening. We validated the anti-NFAT5 activity of 198 primary hit compounds using an NFAT5-dependent reporter assay and identified the novel NFAT5 suppressor KRN2, 13-(2-fluoro)-benzylberberine, and its derivative KRN5. KRN2 inhibited NFAT5 upregulation in macrophages stimulated with lipopolysaccharide and repressed the formation of NF-κB p65-DNA complexes in the NFAT5 promoter region. Interestingly, KRN2 selectively suppressed the expression of pro-inflammatory genes, including Nos2 and Il6, without hampering high-salt-induced NFAT5 and its target gene expressions. Moreover, KRN2 and KRN5, the latter of which exhibits high oral bioavailability and metabolic stability, ameliorated experimentally induced arthritis in mice without serious adverse effects, decreasing pro-inflammatory cytokine production. Particularly, orally administered KRN5 was stronger in suppressing arthritis than methotrexate, a commonly used anti-rheumatic drug, displaying better potency and safety than its original compound, berberine. Therefore, KRN2 and KRN5 can be potential therapeutic agents in the treatment of chronic arthritis.

Synthesis of (3S,4S)-4-aminopyrrolidine-3-ol derivatives and biological evaluation for their BACE1 inhibitory activities.

Synthesis, SAR study and BACE1 inhibitory activity of (3S,4S)-4-aminopyrrolidine-3-ol derivatives (2) were described. The compound 7c exhibited more inhibition activity than 11a (IC50: 0.05µM vs 0.12µM, respectively), but the latter was more effective in cell-based assay (IC50: 1.7µM vs 40% inhibition by 7c @ 10µM) due to the relatively higher cell permeability. Most of the compounds showed high selectivity over BACE2 and cathepsin D. This work will provide useful information for further structural modifications to develop potent BACE1 inhibitors in cell.

Sphingosylphosphorylcholine attenuated ß-amyloid production by reducing BACE1 expression and catalysis in PC12 cells.

Abnormal accumulation of ß-amyloid (Aß) is the main characteristic of Alzheimer's disease (AD) brain and Aß peptides are generated from proteolytic cleavages of amyloid precursor protein (APP) by ß-site APP-converting enzyme 1 (BACE1) and presenilin 1 (PS1). Sphingosylphosphorylcholine (SPC), a choline-containing sphingolipid, showed suppressive effect on Aß production in PC12 cells which stably express Swedish mutant of amyloid precursor protein (APPsw). SPC (> 3 µM) significantly lowered the accumulation of Aß40/42 and the expression of BACE1. However, the transcriptions of other APP processing enzymes like ADAM10 and PS1 were not affected by the SPC addition. Meanwhile, phosphocholine (PC) or other lysophospholipids, such as lysophosphatidylcholine (LPC), lysophosphatidic acid (LPA), sphingosyl-1-phosphate (S1P), did not alter BACE1 expression. Down-regulatory effect of SPC on BACE1 expression appeared to be mediated by NF-κB which is known to suppress the trans-activation of BACE1 promoter in PC12 cells. Here, the nuclear tanslocation of NF-κB was enhanced by SPC treatment in immune-fluorescent image analysis and NF-κB reporter assay. Furthermore, the catalytic activities of BACE1 and BACE2 were dose-dependently inhibited by SPC displaying IC50 values of 2.79 µM and 12.05 µM, respectively. Overall, these data suggest that SPC has the potential to ameliorate Aß pathology in neurons by down-regulating the BACE1-mediated amyloidogenic pathway.

Oral administration of 1,4-aryl-2-mercaptoimidazole inhibits T-cell proliferation and reduces clinical severity in the murine experimental autoimmune encephalomyelitis model.

T cells play a pivotal role in the initiation and progression of multiple sclerosis. We have found that 1,4-aryl-2-mercaptoimidazole (KRM-III) inhibited T-cell antigen receptor- and phorbol myristate acetate/ionomycin-induced activation of nuclear factor of activated T cells (NFAT) and T-cell proliferation with an IC(50) of 5 microM. The KRM-III-mediated inhibitory effect was specific for NFAT activation but not for nuclear factor kappaB. Oral administration of 90 mg/kg KRM-III resulted in complete abrogation of anti-CD3 antibody-induced T-cell activation and a 45.8% reduction in footpad swelling in bovine serum albumin-induced delayed-type hypersensitivity. In the murine experimental autoimmune encephalomyelitis (EAE) model, oral administration of KRM-III significantly attenuated the severity of disease when given before or after disease onset. Draining lymph node cells from KRM-III-treated mice showed markedly reduced proliferation in response to myelin oligodendrocyte glycoprotein peptide. Histological analysis indicated that KRM-III reduced the infiltration of inflammatory cells to the white matter of spinal lumbar cords. These results demonstrate that KRM-III efficiently inhibits T-cell activation and inflammatory responses and lessens EAE clinical signs, which suggest KRM-III as a potential lead compound for the treatment of T-cell-driven autoimmune diseases.

T-cell-targeted signaling inhibitors.

Recent advances in our understanding of the mechanisms of T-cell activation, migration to inflammatory sites, and pathologic disease processes triggered the development of a wide variety of T-cell-targeted signaling inhibitors, which have different targets and modes of action. Depending on the distribution and the role of targets in disease processes, T-cell inhibitors exhibit different levels of efficacy and potential side effects. This review outlines target molecules to which T-cell inhibitors have been developed, their efficacy, and potential safety concerns of T-cell inhibitors.