Objective-Considerable evidence supports the early involvement of monocyte/macrophage recruitment to activated endothelial cells by leukocyte adhesion molecules during atherogenesis. AMP-activated protein kinase (AMPK) is highly expressed in vascular endothelial cells, but its impact on monocyte adhesion and the related mechanisms are not fully understood. The present study was designed to evaluate the impact of and gain mechanistic insight into the signaling coupling AMPK function to the antiinflammatory response. Methods and Results-5-Aminoimidazole-4-carboxamide-1-␤-D-ribonucleotide (AICAR) treatment or overexpression of constitutively active AMPK markedly reduced human monocytic human acute monocytic leukemia cell line-1 cell adhesion and the expression of vascular cell adhesion molecule-1 in tumor necrosis factor-␣-activated human aortic endothelial cells. Furthermore, AICAR or constitutively active AMPK overexpression strongly inhibited the histone acetyltransferase activity of the transcriptional coactivator p300 by phosphorylation of Ser89, which in turn decreased tumor necrosis factor-␣-activated p300-mediated acetylation of nuclear factor-B p65 on Lys221 and reduced the DNA binding activity of nuclear factor-B by inhibiting its recruitment to its target gene promoters. AMPK phosphorylates the transcriptional coactivator p300 via the atypical protein kinase C/. Conclusion-Our findings demonstrate that transcriptional coactivator p300 phosphorylation at Ser89 by AMPK is critical for the therapeutic effect of AMPK and may be a potential target for pharmaceutical intervention in inflammatory diseases such as atherosclerosis. (Arterioscler Thromb Vasc Biol. 2011;31:2897-2908.)

Previously it was found that a conformationally rigid analogue (2) of the epilepsy drug vigabatrin (1) did not inactivate gamma-aminobutyric acid aminotransferase (GABA-AT). A cyclic compound with an exocyclic double bond (6) was synthesized and was found to inactivate GABA-AT, but only in the absence of 2-mercaptoethanol. The corresponding difluoro-substituted analogue (14) was synthesized and was shown to be a very potent time-dependent inhibitor, even in the presence of 2-mercaptoethanol.

Recently, we found (Qiu, J.; Pingsterhaus, J. M.; Silverman, R. B. J. Med. Chem. 1999, 42, 4725-4728) that conformationally rigid analogues of the GABA aminotransferase (GABA-AT) inactivator vigabatrin were not inactivators of GABA-AT. To determine if this is a general phenomenon of GABA-AT inactivators, several mono- and di-halogen-substituted conformationally rigid analogues (7-15) of other GABA-AT inactivators, 4-amino-5-halopentanoic acids, were synthesized as potential inactivators of GABA-AT. Four of them, (+)-7, (-)-9, (+)-10, and (+)-15, were inactivators, although not as potent as the corresponding open-chain analogues. The maximal inactivation rate constants, k(inact), for the fluoro- and bromo-substituted analogues were comparable, indicating that cleavage of the C-X bond is not rate determining. Consistent with that observation is the finding that [3-(2)H]-10 exhibits a deuterium isotope effect on inactivation of 3.3, suggesting that C-H bond cleavage is the rate-determining step. The rate of inactivation of GABA-AT by the fluorinated analogue 7 is 1/15 that of inactivation by the corresponding open-chain analogue, 4-amino-5-fluoropentanoic acid (3a). Whereas inactivation by 3a releases only one fluoride ion, inactivation by 7 releases 148 fluoride ions, accounting for the less efficient inactivation rate. Inactivation leads to covalent attachment of 2 equiv of inactivator after gel filtration; upon urea denaturation, 1 equiv of radioactivity remains bound to the enzyme. This suggests that, unlike the open-chain anlogue, the conformationally rigid analogue becomes, at least partially, attached to an active-site residue. It appears that the conformational constraint has a larger effect on inactivators that inactivate by a Michael addition mechanism than by an enamine mechanism.

S-Nitrosoglutathione reductase (GSNOR) regulates S-nitrosothiols (SNOs) and nitric oxide (NO) in vivo through catabolism of S-nitrosoglutathione (GSNO). GSNOR and the anti-inflammatory and smooth muscle relaxant activities of SNOs, GSNO, and NO play significant roles in pulmonary, cardiovascular, and gastrointestinal function. In GSNOR knockout mice, basal airway tone is reduced and the response to challenge with bronchoconstrictors or airway allergens is attenuated. Consequently, GSNOR has emerged as an attractive therapeutic target for several clinically important human diseases. As such, small molecule inhibitors of GSNOR were developed. These GSNOR inhibitors were potent, selective, and efficacious in animal models of inflammatory disease characterized by reduced levels of GSNO and bioavailable NO. N6022, a potent and reversible GSNOR inhibitor, reduced bronchoconstriction and pulmonary inflammation in a mouse model of asthma and demonstrated an acceptable safety profile. N6022 is currently in clinical development as a potential agent for the treatment of acute asthma.