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Receptor interacting protein kinase 1 (RIPK1) plays a key role in necroptosis, which is a type of programmed necrosis that is involved in ocular diseases, including glaucoma and dry age-related macular degeneration (AMD). We previously introduced RIPK1-inhibitory compound (RIC), which has biochemical characteristics and a mode of action that are distinct from those of the prototype RIPK1 inhibitor necrostatin-1. The intraperitoneal administration of RIC exerts a protective effect on retinal ganglion cells against a glaucomatous insult. In this study, we examined the protective effect of RIC on retinal pigment epithelium (RPE) against sodium iodate (SI) insult, which is associated with dry AMD pathogenesis. The eye drop administration of RIC that reached on the retina prevented RPE loss in SI-induced retinal degeneration. RIC consistently demonstrated retinal protection in the funduscopy and electroretinogram analyses in SI-injected rabbits and iodoacetic acid-treated mini-pigs. Moreover, the in vivo protective effects of RIC were superior to those of ACU-4429 and doxycycline, which are other medications investigated in clinical trials for the treatment of dry AMD, and RIC did not induce retinal toxicity following topical administration in rats. Collectively, RIC displayed excellent retinal penetration and prevented retinal degeneration in the pathogenesis of dry AMD with a high in vivo efficacy.

CKD-712 is a potential treatment for sepsis, as it exhibits protective effects against lipopolysaccharide-mediated platelet aggregation, inducible nitric oxide synthase expression, and cecum-ligation puncture-induced septic mortality in mice. In this study, we develop a rapid and sensitive LC-MS/MS method for determining CKD-712 in rat plasma. CKD-712 and papaverine hydrochloride (an internal standard) were analyzed using an LC-MS/MS system consisting of an Agilent HPLC system (HP-1100) equipped with an Atlantis HILIC Silica (2.1×50mm, 3μm) column and a API 4000 (Applied Biosystems/MDS Sciex, USA) in a positive ESI mode. We utilized multiple reaction monitoring (MRM) at m/z transitions of 306.2-164.0 to analyze CKD-712, and 340.3-202.1 m/z for IS, with a mobile phase of acetonitrile (0.025% trifluoroacetic acid):20mM ammonium acetate (94:6, v/v) at a flow rate of 0.25mL/min. The lower limit of quantification (LLOQ) was 5ng/mL, with a linearity ranging from 5 to 1000ng/mL (r>0.999). Validation parameters including specificity, precision, accuracy, matrix effect, recovery, dilution effect and stability results were well within acceptance criteria, and applied successfully on a pharmacokinetic study in rats.

Motolimod (VTX-2337) is an agonist of toll-like receptor 8. In this study, a novel and sensitive liquid chromatography-tandem mass spectrometry method was developed for quantifying motolimod in rat plasma and subsequently used in a pharmacokinetic study. Proteins were precipitated from plasma samples using acetonitrile prior to the analysis. GS-9620 was used as an internal standard. High-performance liquid chromatography was performed using a Spursil C18-EP column (3 μm, 50 × 2.1 mm). Aqueous ammonium formate and acetonitrile were used as the mobile phase. Motolimod was detected using an electrospray ionization interface under multiple reaction monitoring conditions in the positive ion mode. The developed method produced a linear correlation over a concentration range of 1-1000 ng/mL (r = 0.9944). Intra- and inter-day precision values ranged from 4.8%-10.7% (the lower limit of quantification precision value was 16.3 %), whereas intra- and inter-day accuracy values ranged from 0.3%-9.1 %. The mean recovery of motolimod from rat plasma was 109.4 %. Additionally, motolimod was found to be stable under various conditions (three freeze-thaw cycles, 6-h storage at room temperature, short- and long-term stability tests, and processing). The developed method was successfully used in a pharmacokinetic study in rats. Motolimod showed non-linear pharmacokinetics following its intravenous administration to rats at 0.6-6 mg/kg. Additionally, very low exposure (1 %) was obtained following oral administration of the drug to rats. The results also showed that motolimod has a low metabolic stability in the liver microsomes and exhibits extensive binding to the plasma proteins.

The objective of this study was to characterize the extent of the formation of the active ( trans -alcohol form) and inactive ( cis -alcohol) metabolites of loxoprofen and to compare the kinetics after its intragastric, intravenous, and intramuscular administrations in rats. After intravenous administration of the drug at doses of 5–20mg/kg, the clearance and the volume of distribution for loxoprofen, and the ratios of the AUC for the metabolites to the parent drug were not statistically different with the dosage; the formation clearances were 1.08 and 0.87mL/min/kg for the active and its isomeric metabolite, respectively. After the intragastric, intravenous, or intramuscular administration, AUC for loxoprofen and the metabolites at a dose of 10mg/kg were not statistically different for the different routes of administration. The formation of the metabolites with the concomitant loss of loxoprofen was found in incubations with liver homogenates and blood but not with a muscle homogenate or plasma, indicating that the conversion of loxoprofen to the metabolites may occur both in the liver and extraheptic tissue(s). Thus, approximately 22% of the loxoprofen may have been converted to the active metabolite in the liver and the extraheptic tissue(s) and the pharmacokinetics of the active metabolite was independent of the route of administration. © 2005 Wiley-Liss, Inc. and the American Pharmacists Association