IOP-Lowering Effect of ONO-9054, A Novel Dual Agonist of Prostanoid EP3 and FP Receptors, in Monkeys.

PURPOSE: The purpose of this study was to determine whether a better IOP reduction can be observed in conscious, normotensive monkeys treated with ONO-9054, a novel dual EP3 and FP receptor agonist, compared with prostaglandin F2α analogs. METHODS: The binding affinities and agonistic activities of ONO-AG-367, a carboxylic acid of ONO-9054, to prostanoid receptors were assessed. The IOP-lowering effect of ONO-9054 in monkeys was analyzed after a single (0.3, 3, or 30 µg/mL) or 7-day repeated (30 µg/mL, every day) topical ocular administration. Ophthalmologic and histopathologic evaluations of the eye were performed after 4-week ocular administration of ONO-9054 (30 µg/mL, twice a day) in monkeys. RESULTS: The ONO-AG-367 exhibited high affinity for both EP3 and FP receptors and potent agonist activity, with EC50 values of 28.6 nM for the EP3 receptor and 22.3 nM for the FP receptor. Single and repeated topical ocular administration of ONO-9054 caused IOP reductions in normotensive monkeys. The maximum IOP reductions on day 7 observed with ONO-9054 (7.3 ± 0.8 mm Hg) were significantly greater than those observed with latanoprost (50 µg/mL, 4.9 ± 0.4 mm Hg) or travoprost (40 µg/mL, 5.1 ± 0.6 mm Hg). In ophthalmologic and histopathologic evaluations, slight and transient mydriasis was occasionally observed and no histopathologic lesions attributable to ONO-9054 were noted. CONCLUSIONS: A more profound and longer-lasting reduction in IOP in normotensive monkeys can be observed with ONO-9054, which simultaneously stimulates both EP3 and FP receptors, compared with prostaglandin analogs.

Role of lipid-derived free radical in bleomycin-induced lung injury in mice: availability for ESR spin trap method with organic phase extraction.

Bleomycin is well known as causative molecule for acute lung injury and interstitial pneumonia. The free radical production from bleomycin is thought to play an important role in the pathogenesis of acute lung injury and interstitial pneumonia. However, there was no direct evidence of free radical production in this model. Therefore, we examined in vivo radical production by mice treated with a bleomycin using electron spin resonance with the spin trap, alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone. Six hours after instillation of bleomycin, the lung exposed to bleomycin gave a lipid-derived free radical adduct, which would support evidence for in vitro lipid peroxidation resulting from bleomycin administration. In the treatment of deferoxamine, chelating agent for iron and other metals, to reduce the bleomycin induced free radical production, parallel to decrease the lipid-derived free radical production by deferoxamine, pathophysiological findings of lung injuries were improved by deferoxamine. In conclusion, this is a first paper of in vivo direct evidence of production of free radical from bleomycin-induced lung injury. It is suggested that this method may be used in many kinds of lung disease models, which have potentials of free radical production to cause lung damage.

Superoxide scavenging activity of pirfenidone-iron complex.

Pirfenidone (PFD) is focused on a new anti-fibrotic drug, which can minimize lung fibrosis etc. We evaluated the superoxide (O2*-) scavenging activities of PFD and the PFD-iron complex by electron spin resonance (ESR) spectroscopy, luminol-dependent chemiluminescence assay, and cytochrome c reduction assay. Firstly, we confirmed that the PFD-iron complex was formed by mixing iron chloride with threefold molar PFD, and the complex was stable in distilled water and ethanol. Secondary, the PFD-iron complex reduced the amount of O2*- produced by xanthine oxidase/hypoxanthine without inhibiting the enzyme activity. Thirdly, it also reduced the amount of O2*- released from phorbor ester-stimulated human neutrophils. PFD alone showed few such effects. These results suggest the possibility that the O2*- scavenging effect of the PFD-iron complex contributes to the anti-fibrotic action of PFD used for treating idiopathic pulmonary fibrosis.

Applicability of new spin trap agent, 2-diphenylphosphinoyl-2-methyl-3,4-dihydro-2H-pyrrole N-oxide, in biological system.

Electron spin resonance using spin-trapping is a useful technique for detecting direct reactive oxygen species, such as superoxide (O2.-). However, the widely used spin trap 2,2-dimethyl-3,4-dihydro-2H-pyrrole N-oxide (DMPO) has several fundamental limitations in terms of half-life and stability. Recently, the new spin trap 2-diphenylphosphinoyl-2-methyl-3,4-dihydro-2H-pyrrole N-oxide (DPhPMPO) was developed by us. We evaluated the biological applicability of DPhPMPO to analyze O2.- in both cell-free and cellular systems. DPhPMPO had a larger rate constant for O2.- and formed more stable spin adducts for O2.- than DMPO in the xanthine/xanthine oxidase (X/XO) system. In the phorbol myristate acetate-activated neutrophil system, the detection potential of DPhPMPO for O2.- was significantly higher than that of DMPO (k(DMPO)=13.95M(-1)s(-1), k(DPhPMPO)=42.4M(-1)s(-1)). These results indicated that DPhPMPO is a potentially good candidate for trapping O2.- in a biological system.