Rapid Identification of Novel Inhibitors of the Human Aquaporin-1 Water Channel.

Aquaporins (AQPs) are a family of membrane proteins that function as channels facilitating water transport in response to osmotic gradients. These play critical roles in several normal physiological and pathological states and are targets for drug discovery. Selective inhibition of the AQP1 water channel may provide a new approach for the treatment of several disorders including ocular hypertension/glaucoma, congestive heart failure, brain swelling associated with a stroke, corneal and macular edema, pulmonary edema, and otic disorders such as hearing loss and vertigo.

Preclinical pharmacology, ocular tolerability and ocular hypotensive efficacy of a novel non-peptide bradykinin mimetic small molecule.

We sought to characterize the ocular pharmacology, tolerability and intraocular pressure (IOP)-lowering efficacy of FR-190997, a non-peptidic bradykinin (BK) B2-receptor agonist. FR-190997 possessed a relatively high receptor binding affinity (Ki = 27 nM) and a high in vitro potency (EC50 = 18.3 ± 4.

FR-190997, a nonpeptide bradykinin B2-receptor partial agonist, is a potent and efficacious intraocular pressure lowering agent in ocular hypertensive cynomolgus monkeys.

Preclinical Research FR-190997 (8-[2,6-dichloro-3-[N-[(E)-4-(N-methylcarbamoyl) cinnaminoacetyl]-N-methylamino]benzyloxy]-2-methyl-4- (2-pyridylmethoxy) quinoline), a nonpeptide bradykinin (BK) B2-receptor-selective agonist, represents a novel class of ocular hypotensive agents. FR-190997 exhibited a high affinity for the human cloned B2-receptor (Ki = 9.8 nM) and a relatively high potency (EC50 = 155 nM) for mobilizing intracellular Ca(2+) ([Ca(2+)]i) in human ocular cells from nonpigmented ciliary epithelium; trabecular meshwork [h-TM]; ciliary muscle [h-CM] that are involved in regulating intraocular pressure (IOP).

Human non-pigmented ciliary epithelium bradykinin B2-receptors: receptor localization, pharmacological characterization of intracellular Ca(2+) mobilization, and prostaglandin secretion.

Purpose: To characterize the bradykinin (BK) receptor system in human non-pigmented ciliary epithelium (NPCE) using immunohistochemistry and functional cell-based techniques.

Methods: B2-receptor protein expression was studied in sections of human donor eyes and in Cynomolgus monkey eyes using immunohistochemical methods. The pharmacological characteristics of intracellular Ca(2+) ([Ca(2+)]i) mobilization in response to BK and related peptides, and blockade by two antagonists, was studied in primary human (p-h-NPCE) and in immortalized human NPCE (imh-NPCE) cells.

Trabecular meshwork bradykinin receptors: mRNA levels, immunohistochemical visualization, signaling processes pharmacology, and linkage to IOP reduction.

Purpose: To localize mRNA and protein of bradykinin (BK) receptors, BK precursor polypeptide (kininogen) mRNA, and to study functional biochemical pharmacology of the signal transduction processes mediated by B2-receptors in isolated human trabecular meshwork (h-TM) cells. Intraocular pressure (IOP) lowering effects of 2 kinins were also investigated.

Methods: Previously documented procedures were utilized throughout these studies.

Protein expression, biochemical pharmacology of signal transduction, and relation to intraocular pressure modulation by bradykinin B₂ receptors in ciliary muscle.

Purpose: To examine the bradykinin (BK) B₂-receptor system in human and monkey ciliary muscle (CM) using immunohistochemical techniques, and to pharmacologically characterize the associated biochemical signal transduction systems in human CM (h-CM) cells. BK-induced modulation of intraocular pressure (IOP) in pigmented Dutch-Belt rabbits and cynomolgus monkeys was also studied.

Methods: Previously published procedures were used throughout these studies.

Human conjunctival epithelial cell responses to platelet-activating factor (PAF): signal transduction and release of proinflammatory cytokines.

Purpose: The aims of the study were to characterize the signal transduction responses to platelet-activating factor (PAF) and to monitor the downstream effects of PAF on the production of proinflammatory cytokines in human conjunctival epithelial cells (HCECs).

Methods: The generation of inositol phosphates ([(3)H]IPs) from [(3)H]phosphoinositide (PI) hydrolysis and the mobilization of intracellular calcium ([Ca(2+)](i)) were evaluated using ion exchange chromatography and Fura-2 fluorescence techniques, respectively. The production of the cytokines (interleukin-6 [IL-6], interleukin-8 [IL-8], and granulocyte macrophage colony-stimulating factor [GM-CSF]) from PAF-stimulated HCECs was quantified using specific ELISA assays.

Preclinical pharmacology of AL-12182, a new ocular hypotensive 11-oxa prostaglandin analog.

Purpose: The aim of this study was to determine selected in vivo ocular properties of AL-12182 (5,6-dihydro-4,5-didehydro-11-deoxy-11-oxa-16-(3-chlorophenoxy)-omega-tetranor-PGF(2alpha) isopropyl ester) and the in vitro profile of its free acid, AL-12180.

Methods: Previously documented radioligand binding and functional assays involving human ciliary muscle cells (h-CM), human trabecular meshwork (h-TM) and other cells, and porcine ocular arteries were utilized. For in vivo procedures, we utilized rabbits, cats, and nonhuman primates to measure hyperemia, pupil diameter, and intraocular pressure (IOP), respectively.

Molecular pharmacology of the DP/EP2 class prostaglandin AL-6598 and quantitative autoradiographic visualization of DP and EP2 receptor sites in human eyes.

DP-class prostaglandins and prostaglandin analogs (collectively, prostaglandins or PGs) such as PGD2, BW245C, ZK110841, and ZK118182, lower intraocular pressure (IOP) in animal models of ocular hypertension. A new analog of ZK118182 (AL-6556; 13,14-dihydro-ZK118182) was synthesized, and the isopropyl ester of AL-6556 (AL-6598) was shown recently to lower IOP in the ocular hypertensive cynomolgus monkey model of glaucoma and in human subjects. AL-6556 and AL-6598 had an affinity (Ki) of 2.