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Publications (11)31.39 Total impact

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    ABSTRACT: Choroidal neovascularization (CNV) is a defining feature of wet age-related macular degeneration. We examined the functional role of CCR3 in the development of CNV in mice and primates. CCR3 was associated with spontaneous CNV lesions in the newly described JR5558 mice, whereas CCR3 ligands localized to CNV-associated macrophages and the retinal pigment epithelium/choroid complex. Intravitreal injection of neutralizing antibodies against endothelial growth factor receptor 2, CCR3, CCR3 ligand 11/eotaxin-1, and CCR3 ligand 24/eotaxin-2 all reduced CNV area and lesion number in these mice. Systemic administration of the CCR3 antagonists GW766994X and GW782415X reduced spontaneous CNV in JR5558 mice and laser-induced CNV in mouse and primate models in a dose-dependent fashion. Combination treatment with antivascular endothelial growth factor receptor 2 antibody and GW766994X yielded additive reductions in CNV area and hyperpermeability in mice. Interestingly, topical GW766994X and intravitreal anti-CCR3 antibody yielded strong systemic effects, reducing CNV in the untreated, contralateral eye. Contrarily, ocular administration of GW782415X in primates failed to substantially elevate plasma drug levels or to reduce the development of grade IV CNV lesions. These findings suggest that CCR3 signaling may be an attractive therapeutic target for CNV, utilizing a pathway that is at least partly distinct from that of endothelial growth factor receptor. The findings also demonstrate that systemic exposure of CCR3 antagonists may be crucial for CNV-targeted activity. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.
    American Journal Of Pathology 07/2015; 185(9). DOI:10.1016/j.ajpath.2015.04.029 · 4.59 Impact Factor
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    ABSTRACT: Purpose: To test the effect of pazopanib, a tyrosine kinase inhibitor that blocks VEGF and platelet-derived growth factor (PDGF) receptors and c-Kit, on vascular leakage and neovascularization (NV) in the retina. Methods: Pazopanib was tested to determine its effect on VEGF-induced vascular permeability via measurement of [(3)H]mannitol retina to lung (RLLR) and retina to renal leakage ratios (RRLR) and in rho/VEGF mice with subretinal NV. In rabbits, the effect of intravitreal, topical, and systemic pazopanib on VEGF-induced leakage was tested by vitreous fluorophotometry. Results: In mice, oral pazopanib (40 mg/kg twice a day [bid]) reduced RLLR (0.84 to 0.58, P = 0.0014) and RRLR (0.55 to 0.30, P = 0.0018) in VEGF-injected eyes. After intraocular injection of VEGF into both eyes, topical pazopanib (10 mg/mL three times a day [tid] for 14 days) reduced RLLR (0.85 vs. 0.56, P = 0.001), RRLR (0.44 vs. 0.28, P = 0.0075), and immunoreactive albumin in the retina compared to values in fellow eye controls. Treatment of one eye of rho/VEGF mice with 10 mg/mL, but not 5 mg/mL, pazopanib tid reduced the mean area of subretinal NV compared to that in fellow eyes (0.0055 vs. 0.0025 mm(2), P = 0.020). In rabbits, intravitreal pazopanib suppressed VEGF-induced fluorescein leakage, but topical (10 mg/mL four times a day [qid] or 12 mg/mL bid) had no significant effect. Systemic administration of pazopanib by osmotic pump with or without 10 mg/mL drops tid also failed to suppress VEGF-induced leakage. Conclusions: Administration of pazopanib topically or systemically suppressed retinal vascular leakage in mice, but not rabbits. These data suggest differences in the blood-retinal barrier (BRB) of mice and rabbits and indicate that penetration through the outer BRB may be needed for topically administered drugs to exert effects in the retina.
    Investigative ophthalmology & visual science 11/2012; 54(1). DOI:10.1167/iovs.12-10473 · 3.40 Impact Factor
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    ABSTRACT: The aim of the study was to investigate the ocular hypotensive activity of a nitric oxide (NO)-donating latanoprost, BOL-303259-X, following topical administration. The effect of BOL-303259-X (also known as NCX 116 and PF-3187207) on intraocular pressure (IOP) was investigated in monkeys with laser-induced ocular hypertension, dogs with naturally-occurring glaucoma and rabbits with saline-induced ocular hypertension. Latanoprost was used as reference drug. NO, downstream effector cGMP, and latanoprost acid were determined in ocular tissues following BOL-303259-X administration as an index of prostaglandin and NO-mediated activities. In primates, a maximum decrease in IOP of 31% and 35% relative to baseline was achieved with BOL-303259-X at doses of 0.036% (9 μg) and 0.12% (36 μg), respectively. In comparison, latanoprost elicited a greater response than vehicle only at 0.1% (30 μg) with a peak effect of 26%. In glaucomatous dogs, IOP decreased from baseline by 44% and 10% following BOL-303259-X (0.036%) and vehicle, respectively. Latanoprost (0.030%) lowered IOP by 27% and vehicle by 9%. Intravitreal injection of hypertonic saline in rabbits increased IOP transiently. Latanoprost did not modulate this response, whereas BOL-303259-X (0.036%) significantly blunted the hypertensive phase. Following BOL-303259-X treatment, latanoprost acid was significantly elevated in rabbit and primate cornea, iris/ciliary body and aqueous humor as was cGMP in aqueous humor. BOL-303259-X lowered IOP more effectively than latanoprost presumably as a consequence of a contribution by NO in addition to its prostaglandin activity. The compound is now in clinical development for the treatment of glaucoma and ocular hypertension.
    Experimental Eye Research 03/2011; 93(3):250-5. DOI:10.1016/j.exer.2011.03.001 · 2.71 Impact Factor
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    ABSTRACT: Better control of intraocular pressure (IOP) is the most effective way to preserve visual field function in glaucomatous patients. While prostaglandin FP analogs are leading the therapeutic intervention for glaucoma, new target classes also are being identified with new lead compounds being developed for IOP reduction. One target class currently being investigated includes the prostaglandin EP receptor agonists. Recently PF-04217329 (Taprenepag isopropyl), a prodrug of CP-544326 (active acid metabolite), a potent and selective EP(2) receptor agonist, was successfully evaluated for its ocular hypotensive activity in a clinical study involving patients with primary open angle glaucoma. In the current manuscript, the preclinical attributes of CP-544326 and PF-0421329 have been described. CP-544326 was found to be a potent and selective EP(2) agonist (IC(50) = 10 nM; EC(50) = 2.8 nM) whose corneal permeability and ocular bioavailability were significantly increased when the compound was dosed as the isopropyl ester prodrug, PF-04217329. Topical ocular dosing of PF-04217329 was well tolerated in preclinical species and caused an elevation of cAMP in aqueous humor/iris-ciliary body indicative of in vivo EP(2) target receptor activation. Topical ocular dosing of PF-04217329 resulted in ocular exposure of CP-544326 at levels greater than the EC(50) for the EP(2) receptor. PF-04217329 when dosed once daily caused between 30 and 50% IOP reduction in single day studies in normotensive Dutch-belted rabbits, normotensive dogs, and laser-induced ocular hypertensive cynomolgus monkeys and 20-40% IOP reduction in multiple day studies compared to vehicle-dosed eyes. IOP reduction was sustained from 6 h through 24 h following a single topical dose. In conclusion, preclinical data generated thus far appear to support the clinical development of PF-04217329 as a novel compound for the treatment of glaucoma.
    Experimental Eye Research 03/2011; 93(3):256-64. DOI:10.1016/j.exer.2011.02.015 · 2.71 Impact Factor
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    ABSTRACT: The IOP lowering effects of NCX 139, a new chemical entity comprising latanoprost amide and a NO-donating moiety, were compared to those of the respective des-nitro analog in in vitro assays and in rabbit and dog models of ocular hypertension. The NO donor, molsidomine as well as the prostamide bimatoprost (Lumigan(®)) and the prostaglandin agonist, latanoprost (Xalatan(®)) were also investigated for comparison. NCX 139 but not its des-nitro analog resulted in NO-mediated vascular relaxant effect in pre-contracted rabbit aortic rings (EC(50)=0.70±0.06 μM; E(max)=80.6±2.9%). Like bimatoprost (IC(50)=3.07±1.3 μM) or latanoprost (IC(50)=0.48±0.15 μM), NCX 139 displaced (3)H-PGF2α binding on recombinant human prostaglandin-F (FP) receptors with an estimated potency of 0.77±0.13 μM. In transient ocular hypertensive rabbits, bimatoprost and latanoprost were not effective while molsidomine elicited a dose-dependent reduction of IOP confirming the responsiveness of rabbits to NO but not to FP receptor agonists. NCX 139 tested at a therapeutically relevant dose, significantly lowered IOP while the des-nitro analog was not effective (0.03% NCX 139, Δ(max)=-12.8±2.0 mmHg). In glaucomatous dogs, 0.03% NCX 139 decreased IOP to a greater extent compared to an equimolar dose of the respective des-nitro derivative (Δ(max)=-4.6±1.0 and -2.7±1.3 mmHg, respectively for NCX 139 and its des-nitro analog). Albeit with low potency, NCX 139 also resulted effective in normotensive dogs while it did not reduce IOP in normotensive rabbits. NCX 139, a compound targeting two different and important mechanisms, is endowed with ocular hypotensive effects more evident in hypertensive conditions which may be of interest in the search of more effective treatments for hypertensive glaucoma.
    Experimental Eye Research 02/2011; 93(3):243-9. DOI:10.1016/j.exer.2011.02.006 · 2.71 Impact Factor
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    ABSTRACT: Nitric oxide (NO) is involved in a variety of physiological processes including ocular aqueous humor dynamics by targeting mechanisms that are complementary to those of prostaglandins. Here, we have characterized a newly synthesized compound, NCX 125, comprising latanoprost acid and NO-donating moieties. NCX 125 was synthesized and tested in vitro for its ability to release functionally active NO and then compared with core latanoprost for its intraocular pressure (IOP)-lowering effects in rabbit, dog, and nonhuman primate models of glaucoma. NCX 125 elicited cGMP formation (EC(50) = 3.8 + or - 1.0 microM) in PC12 cells and exerted NO-dependent iNOS inhibition (IC(50) = 55 + or - 11 microM) in RAW 264.7 macrophages. NCX 125 lowered IOP to a greater extent compared with equimolar latanoprost in: (a) rabbit model of transient ocular hypertension (0.030% latanoprost, not effective; 0.039% NCX 125, Delta(max) = -10.6 + or - 2.3 mm Hg), (b) ocular hypertensive glaucomatous dogs (0.030% latanoprost, Delta(max)= -6.7 + or - 1.2 mm Hg; 0.039% NCX 125, Delta(max) = -9.1 + or - 3.1 mm Hg), and (c) laser-induced ocular hypertensive non-human primates (0.10% latanoprost, Delta(max) = -11.9 + or - 3.7 mm Hg, 0.13% NCX 125, Delta(max) = -16.7 + or - 2.2 mm Hg). In pharmacokinetic studies, NCX 125 and latanoprost resulted in similar latanoprost-free acid exposure in anterior segment ocular tissues. NCX 125, a compound targeting 2 different mechanisms, is endowed with potent ocular hypotensive effects. This may lead to potential new perspectives in the treatment of patients at risk of glaucoma.
    Journal of ocular pharmacology and therapeutics: the official journal of the Association for Ocular Pharmacology and Therapeutics 04/2010; 26(2):125-32. DOI:10.1089/jop.2009.0120 · 1.47 Impact Factor
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    ABSTRACT: Purpose. p38 mitogen-activated protein kinase (MAPK) is known to play a regulatory role in inflammatory processes in disease. Inflammation has been linked also to the development of diabetic retinopathy in rodents. This study was conducted to evaluate the effect of a p38 MAPK inhibitor on the development of early stages of diabetic retinopathy in rats. Methods. Streptozotocin-diabetic rats were assigned to two groups-treated with the p38 MAPK inhibitor PHA666859 (Pfizer, New York, NY) and untreated-and compared with age-matched nondiabetic control animals. Results. At 2 months of diabetes, insulin-deficient diabetic control rats exhibited significant increases in retinal superoxide, nitric oxide (NO), cyclooxygenase (COX)-2, and leukostasis within retinal microvessels. All these abnormalities were significantly inhibited by the p38 MAPK inhibitor (25 mg/kgBW/d). At 10 months of diabetes, significant increases in the number of degenerate (acellular) capillaries and pericyte ghosts were measured in control diabetic rats versus those in nondiabetic control animals, and pharmacologic inhibition of p38 MAPK significantly inhibited all these abnormalities (all P < 0.05). This therapy also had beneficial effects outside the eye in diabetes, as evidenced by the inhibition of a diabetes-induced hypersensitivity of peripheral nerves to light touch (tactile allodynia). Conclusions. p38 MAPK plays an important role in diabetes-induced inflammation in the retina, and inhibition of p38 MAPK offers a novel therapeutic approach to inhibiting the development of early stages of diabetic retinopathy and other complications of diabetes.
    Investigative ophthalmology & visual science 04/2010; 51(4):2158-64. DOI:10.1167/iovs.09-3674 · 3.40 Impact Factor
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    ABSTRACT: Steroids are used in a diverse range of conditions in clinical ophthalmology and one of the most significant complications is corticosteroid-induced glaucoma, which is characterized by an increase in intraocular pressure (IOP). 11beta-Hydroxysteroid dehydrogenase-1 (11beta-HSD1) is known to catalyze the interconversion of hormonally inactive cortisone to hormonally active cortisol and is widely expressed in the eye, particularly ciliary epithelium. Carbenoxolone (CBX), an 11beta-HSD1 inhibitor, has been shown to reduce IOP in healthy volunteers and patients with ocular hypertension (OHT). The purpose of this study was to: (1) develop an in vivo model for the assessment of cortisone to cortisol conversion in the eye, that is, 11beta-HSD1 activity and (2) assess the pharmacokinetic/pharmacodynamic relationship following topical treatment with 11beta-HSD1 inhibitors using an in vivo rabbit model. Potent and selective 11beta-HSD1 inhibitors were topically administered to the rabbit eye and exogenous cortisone to endogenous cortisol conversion in the eye was assessed in rabbits. Tissues were then evaluated for cortisone, cortisol, and 11beta-HSD1 inhibitor levels by LC/MS/MS. Concomitantly cortisol activity in ocular tissue samples was determined using a secondary mechanistic pLuc-GRE assay. Topical treatment with potent and selective 11beta-HSD1 inhibitors resulted in complete inhibition in the conversion of cortisone to cortisol in the rabbit eye as well as decreased pLuc-GRE luciferase activity. The reduction of cortisone conversion was time- and dose-dependent as well as dependent on dosing volume (suggestive of increased spillover and washout with greater dosing volume). In conclusion, topical delivery of 11beta-HSD1 inhibitors can reduce or inhibit the conversion of cortisone to cortisol in the eye, indicating that the rabbit eye possesses an active enzyme for glucocorticoid synthesis. Dosing concentration and volume play an important role in the pharmacokinetic and pharmacodynamic effects of topically delivering an 11beta-HSD1 inhibitor. The rabbit model is useful for mechanistically assessing the conversion of cortisone to cortisol in the eye.
    Journal of ocular pharmacology and therapeutics: the official journal of the Association for Ocular Pharmacology and Therapeutics 07/2009; 25(3):215-22. DOI:10.1089/jop.2008.0120 · 1.47 Impact Factor
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    ABSTRACT: A novel class of timolol derivatives with nitric oxide (NO)-donating moieties achieved chemical stability yet under physiologically relevant conditions released timolol and NO. Hindered esters A were designed and synthesized, whose 'triggered' release relied on enzymatic hydrolysis of the nitrate ester in A to B, that in turn cyclized to liberate timolol.
    Bioorganic & medicinal chemistry letters 04/2009; 19(10):2785-8. DOI:10.1016/j.bmcl.2009.03.111 · 2.42 Impact Factor
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    ABSTRACT: The recently introduced Clonetics human corneal epithelium (cHCE) cell line is considered a promising in vitro permeability model, replacing excised animal cornea to predict corneal permeability of topically administered compounds. The purpose of this study was to further characterize cHCE as a corneal permeability model from both drug metabolism and transport aspects. First, good correlation was found in the permeability values (P(app)) obtained from cHCE and rabbit corneas for various ophthalmic drugs and permeability markers. Second, a previously established real-time quantitative polymerase chain reaction method was used to profile mRNA expression of drug-metabolizing enzymes (major cytochromes P450 and UDP glucuronosyltransferase 1A1) and transporters in cHCE in comparison with human cornea. Findings indicated that 1) the mRNA expression of most metabolizing enzymes tested was lower in cHCE than in excised human cornea, 2) the mRNA expression of efflux transporters [multidrug resistant-associated protein (MRP) 1, MRP2, MRP3, and breast cancer resistance protein], peptide transporters (PEPT1 and PEPT2), and organic cation transporters (OCTN1, OCTN2, OCT1, and OCT3) could be detected in cHCE as in human cornea. However, multidrug resistance (MDR) 1 and organic anion transporting polypeptide 2B1 was not detected in cHCE; 3) cHCE was demonstrated to possess both esterase and ketone reductase activities known to be present in human cornea; and 4) transport studies using probe substrates suggested that both active efflux and uptake transport may be limited in cHCE. As the first detailed report to delineate drug metabolism and transport characteristics of cHCE, this work shed light on the usefulness and potential limitations of cHCE in predicting the corneal permeability of ophthalmic drugs, including ester prodrugs, and transporter substrates.
    Drug metabolism and disposition: the biological fate of chemicals 03/2009; 37(5):992-8. DOI:10.1124/dmd.108.026286 · 3.25 Impact Factor
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    ABSTRACT: Studies were designed to quantitatively assess the mRNA expression of 1) 10 cytochrome P450 (P450) enzymes in human cornea, iris-ciliary body (ICB), and retina/choroid relative to their levels in the liver, and of 2) 21 drug transporters in these tissues relative to their levels in human small intestine, liver, or kidney. Potential species differences in mRNA expression of PEPT1, PEPT2, and MDR1 were also assessed in these ocular tissues from rabbit, dog, monkey, and human. P450 expression was either absent or marginal in human cornea, ICB, and retina/choroid, suggesting a limited role for P450-mediated metabolism in ocular drug disposition. In contrast, among 21 key drug efflux and uptake transporters, many exhibited relative expression levels in ocular tissues comparable with those observed in small intestine, liver, or kidney. This robust ocular transporter presence strongly suggests a significant role that transporters may play in ocular barrier function and ocular pharmacokinetics. The highly expressed efflux transporter MRP1 and uptake transporters PEPT2, OCT1, OCTN1, and OCTN2 may be particularly important in absorption, distribution, and clearance of their drug substrates in the eye. Evidence of cross-species ocular transporter expression differences noted in these studies supports the conclusion that transporter expression variability, along with anatomic and physiological differences, should be taken into consideration to better understand animal ocular pharmacokinetic and pharmacodynamic data and the scalability to human for ocular drugs.
    Drug metabolism and disposition: the biological fate of chemicals 08/2008; 36(7):1300-7. DOI:10.1124/dmd.108.021121 · 3.25 Impact Factor