No abstract

Certain leukocytes release serine proteases that sustain inflammatory processes and cause disease conditions, such as asthma and chronic obstructive pulmonary disease. We identified ␤-ketophosphonate 1 (JNJ-10311795; RWJ-355871) as a novel, potent dual inhibitor of neutrophil cathepsin G (K i ‫؍‬ 38 nM) and mast cell chymase (K i ‫؍‬ 2.3 nM). The x-ray crystal structures of 1 complexed with human cathepsin G (1.85 Å) and human chymase (1.90 Å) reveal the molecular basis of the dual inhibition. Ligand 1 occupies the S 1 and S 2 subsites of cathepsin G and chymase similarly, with the 2-naphthyl in S 1 , the 1-naphthyl in S 2 , and the phosphonate group in a complex network of hydrogen bonds. Surprisingly, however, the carboxamido-N-(naphthalene-2-carboxyl)piperidine group is found to bind in two distinct conformations. In cathepsin G, this group occupies the hydrophobic S 3 /S 4 subsites, whereas in chymase, it does not; rather, it folds onto the 1-naphthyl group of the inhibitor itself. Compound 1 exhibited noteworthy antiinflammatory activity in rats for glycogen-induced peritonitis and lipopolysaccharide-induced airway inflammation. In addition to a marked reduction in neutrophil influx, 1 reversed increases in inflammatory mediators interleukin-1␣, interleukin-1␤, tissue necrosis factor-␣, and monocyte chemotactic protein-1 in the glycogen model and reversed increases in airway nitric oxide levels in the lipopolysaccharide model. These findings demonstrate that it is possible to inhibit both cathepsin G and chymase with a single molecule and suggest an exciting opportunity in the treatment of asthma and chronic obstructive pulmonary disease.

The preclinical antiinflammatory effects of RWJ-355871 in these animal models of inflammation indicate that this dual inhibitor may have therapeutic utility for treating airway inflammatory diseases involving mechanisms that depend on Cat G and/or chymase.

Time- and vehicle-related variability of bladder and urethral rhabdosphincter (URS) activity as well as cardiorespiratory and blood chemistry values were examined in the acetic acid-induced bladder irritation model in α-chloralose-anesthetized female cats. Additionally, bladder and urethra were evaluated histologically using Mason trichrome and toluidine blue staining. Urodynamic, cardiovascular and respiratory parameters were collected during intravesical saline infusion followed by acetic acid (0.5%) to irritate the bladder. One hour after starting acetic acid infusion, a protocol consisting of a cystometrogram, continuous infusion-induced rhythmic voiding contractions, and a 5 min “quiet period” (bladder emptied without infusion) was precisely repeated every 30 minutes. Administration of vehicle (saline i.v.) occurred 15 minutes after starting each of the first 7 cystometrograms and duloxetine (1mg/kg i.v.) after the 8th. Acetic acid infusion into the bladder increased URS-EMG activity, bladder contraction frequency, and decreased contraction amplitude and capacity, compared to saline. Bladder activity and URS activity stabilized within 1 and 2 hours, respectively. Duloxetine administration significantly decreased bladder contraction frequency and increased URS-EMG activity to levels similar to previous reports. Cardiorespiratory parameters and blood gas levels remained consistent throughout the experiment. The epithelium of the bladder and urethra were greatly damaged and edema and infiltration of neutrophils in the lamina propria of urethra were observed. These data provide an ample evaluation of the health of the animals, stability of voiding function and appropriateness of the model for testing drugs designed to evaluate lower urinary tract as well as cardiovascular and respiratory systems function.

SK&F 107647, a novel synthetic low-molecular-weight peptide, has demonstrated potent antiinfective activities in murine models of fungal and viral infection. To determine if the hematoregulatory activities of SK&F 107647 could offer protection over conventional antibiotic therapy or as a single agent in animal models of bacterial sepsis, rats were implanted intraperitoneally with a live bacteria-containing fibrin-thrombin clot. Rats pretreated subcutaneously or orally with SK&F 107647 and then infected with either a gram-negative (Escherichia coli) or a gram-positive (Staphylococcus aureus) bacteria-containing clot demonstrated significantly improved survival over control formulation-treated animals. Treated animals showed increased effector cell activation, measured by CD11b expression on neutrophils and monocytes, and up to 1000-fold reduction in the number of E. coli recovered from blood. Thus, the hematoregulatory activities of SK&F 107647 can increase natural host resistance to infections caused by both gram-negative and gram-positive bacteria.