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Purpose: To determine NCX 470 (0.1%) and Lumigan® (bimatoprost ophthalmic solution, 0.01%-LUM) intraocular pressure (IOP)-lowering activity after single or repeated (5 days) dosing along with changes in aqueous humor (AH) dynamics. Methods: Ocular hypotensive activity of NCX 470 and LUM was compared with vehicle (VEH) in Beagle dogs using TonoVet®. Non-human primates (NHP) and bioengineered three-dimensional (3D) human Trabecular Meshwork/Schlemm's Canal (HTM/HSC™) constructs exposed to transforming growth factor-ß2 (TGFß2) were used to monitor NCX 470 and LUM-induced changes in AH dynamics. Results: NCX 470 (30 µL/eye) showed greater IOP reduction compared with LUM (30 µL/eye) following single AM dosing [maximum change from baseline (CFBmax) = -1.39 ± 0.52, -6.33 ± 0.73, and -3.89 ± 0.66 mmHg (mean ± standard error of the mean) for VEH, NCX 470, and LUM, respectively]. Likewise, repeated 5 days daily dosing of NCX 470 resulted in lower IOP than LUM across the duration of the study (average IOP decrease across tests was -0.45 ± 0.22, -6.06 ± 0.15, and -3.60 ± 0.22 mmHg for VEH, NCX 470, and LUM, respectively). NCX 470 increased outflow facility (Cfl) in vivo in NHP (CflVEH = 0.37 ± 0.09 µL/min/mmHg and CflNCX470 = 0.64 ± 0.17 µL/min/mmHg) as well as in vitro (CHTM/HSC) in HTM/HSC constructs (CHTM/HSC_VEH = 0.47 ± 0.02 µL/min/mm2/mmHg and CHTM/HSC_NCX470 = 0.76 ± 0.03 µL/min/mm2/mmHg). In addition, NCX 470 increased uveoscleral outflow (FuVEH = 0.62 ± 0.26 µL/min and FuNCX470 = 1.53 ± 0.39 µL/min with episcleral venous pressure of 15 mmHg) leaving unaltered aqueous flow (AHFVEH = 2.03 ± 0.22 µL/min and AHFNCX470 = 1.93 ± 0.31 µL/min) in NHP. Conclusions: NCX 470 elicits greater IOP reduction than LUM following single or repeated dosing. Data in NHP and 3D-HTM/HSC constructs suggest that changes in Cfl and Fu account for the robust IOP-lowering effect of NCX 470.
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Purpose: The purpose of this study was to assess the retinal protective activity and ocular hemodynamics after NCX 470 (0.1%) compared to bimatoprost administered as the US Food and Drug Administration (FDA)-approved drug (Lumigan - 0.01% ophthalmic solution, LUM) and at an equimolar dose (0.072%, BIM) to that released by NCX 470. Methods: Endothelin-1 (ET-1) induced ischemia/reperfusion injury model in rabbits was used. ET-1 was injected nearby the optic nerve head (ONH) twice/week for 6 weeks. Starting on week 3, the animals received vehicle (VEH), NCX 470, LUM, or BIM (30 µL/eye, twice daily, 6 days/week) until the end of ET-1 treatment. Intraocular pressure (IOP), ophthalmic artery resistive index (OA-RI), and electroretinogram (ERG) data were collected prior to dosing and at different time points postdosing. Reduced glutathione, 8-Hydroxy 2-deoxyguanosine, and Caspase-3 were determined in the retina of treated eyes. DNA fragmentation was determined by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) staining. Results: ET-1 increased IOP (VEHIOP_Baseline = 20.5 ± 0.8 and VEHIOP_Week6 = 24.8 ± 0.3 mmHg) and OA-RI (VEHOA-RI_Baseline = 0.36 ± 0.02 and VEHOA-RI_Week6 = 0.55 ± 0.01) and reduced rod/cone responses over time. Oxidative stress, inflammation, and apoptotic markers increased in ET-1-treated eyes. NCX 470 prevented IOP (NCX 470IOP_Week6 = 18.1 ± 0.6 mmHg) and OA-RI changes (NCX 470OA-RI_Week6 = 0.33 ± 0.01) and restored ERG amplitude leaving unaltered the respective latency; these effects were only partially demonstrated by LUM or BIM. Additionally, NCX 470 reduced oxidative stress, inflammation, and apoptosis in the retinas of treated eyes. BIM and LUM were numerically less effective on these parameters. Conclusions: NCX 470 repeated ocular dosing ameliorates ocular hemodynamics and retinal cell dysfunction caused by ischemia/reperfusion via nitric oxide- and bimatoprost-mediated mechanisms. Translational Relevance: If confirmed in clinical setting our data may open new therapeutic opportunities to reduce visual field loss in glaucoma.
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Glaucoma , Disco Óptico , Traumatismo por Reperfusão , Estados Unidos , Animais , Coelhos , Bimatoprost , Citoproteção , Artéria Oftálmica , Hemodinâmica , Retina , Traumatismo por Reperfusão/tratamento farmacológico , Traumatismo por Reperfusão/prevenção & controleRESUMO
Purpose: Determine whether NCX 470, a nitric oxide (NO)-donating bimatoprost with clinically demonstrated intraocular pressure (IOP)-lowering effects, improves ocular hemodynamics and retinal physiology. Methods: Endothelin-1 (ET-1)-induced ischemia/reperfusion model in New Zealand white rabbits was used. ET-1 was injected next to the optic nerve twice/week (Monday and Thursday) for 6 weeks. Starting on week 3, animals received NCX 470 (0.1% bid, 6 days/week Monday-Saturday) or vehicle until the end of ET-1 treatment. IOP, ophthalmic artery resistive index (OA-RI) and retina physiology (electroretinogram, ERG) were determined before dosing and at different times post-dosing. All measurements were taken on Mondays before the AM daily dosing (36 h treatment-free). Finally, oxidative stress markers were determined in dissected retina and iris/ciliary body of treated eyes. Results: Injection of ET-1 progressively increased IOP (20.7 ± 0.6, 24.9 ± 1.2, and 27.0 ± 0.6 mmHg at baseline, week 2 and 6, respectively) and OA-RI (0.30 ± 0.02, 0.39 ± 0.02, and 0.42 ± 0.03 at baseline, week 2 and 6, respectively) and reduced rods and/or cones response as indicated by changes in ERG amplitudes under different stimulating conditions. NCX 470 re-established baseline IOP (21.8 ± 1.0 mmHg), OA-RI (0.33 ± 0.02), and ERG amplitude by week 6 (mostly rod response, 0.01Dark_AVeh_6week = 32.2 ± 3.0 µV and 0.01Dark_ANCX470_6week 44.3 ± 4.5 µV; mostly cone response, 3.0Dark_AVeh_6week = 87.6 ± 10.1 µV and 3.0Dark_ANCX470_6week = 122.8 ± 11.4 µV; combined rod/cone response, 3.0Light_AVeh_6week = 49.8 ± 6.5 µV and 3.0Light_ANCX470_6week = 64.2 ± 6.8 µV). NCX 470 also reversed ET-1-induced changes in glutathione and manganese superoxide dismutase (oxidative stress markers) in retina and iris/ciliary body. Conclusions: Repeated ocular topical dosing with NCX 470 reverses ET-1-induced changes in IOP, OA-RI, and ERG suggesting improved ocular hemodynamics and retinal physiology likely independently from its demonstrated IOP-lowering effect.
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Hipertensão Ocular , Traumatismo por Reperfusão , Animais , Fenômenos Fisiológicos Celulares , Endotelina-1/farmacologia , Endotelina-1/uso terapêutico , Hemodinâmica , Pressão Intraocular , Hipertensão Ocular/tratamento farmacológico , Nervo Óptico , Coelhos , Traumatismo por Reperfusão/tratamento farmacológico , RetinaRESUMO
Purpose: NCX 667, a novel nitric oxide (NO) donor with an isomannide core, was characterized for its IOP-lowering ability in animal models of ocular hypertension and glaucoma. Bioengineered human trabecular meshwork/Schlemm's canal (HTM/HSC) constructs were used to explore the mode of action. Methods: Ocular normotensive New Zealand white (NZW) rabbits (ONT-rabbits), spontaneously ocular hypertensive pigmented Dutch-belted rabbits (sOHT-rabbits), hypertonic saline (5%)-induced transient ocular hypertensive NZW rabbits (tOHT-rabbits), ocular normotensive Beagle dogs (ONT-dogs), and laser-induced ocular hypertensive cynomolgus monkeys (OHT-monkeys) were used. NCX 667 or vehicle (30 µL) was instilled in a crossover, masked fashion and intraocular pressure (IOP) measured before dosing (baseline) and for several hours thereafter. The ONT-rabbits were used for cyclic guanosine monophosphate (cGMP) determination in ocular tissues after ocular dosing with NCX 667. Transforming growth factor-beta2 (TGFß2) (2.5 ng/mL, six days)-treated HTM/HSC constructs were used to address changes in outflow facility. Results: NCX 667 resulted in robust and dose-dependent IOP decrease in all models used. Maximal IOP-lowering efficacy at 1% was -4.1 ± 0.6, -12.2 ± 2.7, -10.5 ± 2.0, -5.3 ± 0.8, and -6.6 ± 1.9 mmHg, respectively, in ONT-dogs, sOHT-rabbits, tOHT-rabbits, ONT-rabbits, and OHT-monkeys. In ONT-rabbits NCX 667 (1%) increased cGMP in aqueous humor (AH) but not in retina and iris/ciliary body. NCX 667 concentration-dependently increased outflow facility in TGFß2-treated HTM/HSC constructs (outflow facility, 0.10 ± 0.06 and 0.30 ± 0.10 µL/min/mmHg/mm2, respectively, in vehicle- and NCX 667-treated constructs). Conclusions: NCX 667 leads to robust IOP lowering in several animal models. Evidence in HTM/HSC constructs indicate that the IOP reduction likely results from NO-mediated increase of the conventional outflow pathway. Other mechanisms including changes in AH production and episcleral vein pressure may not be excluded at this time.
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Pressão Intraocular/efeitos dos fármacos , Limbo da Córnea/efeitos dos fármacos , Doadores de Óxido Nítrico/uso terapêutico , Hipertensão Ocular/tratamento farmacológico , Malha Trabecular/efeitos dos fármacos , Animais , Humor Aquoso/fisiologia , Compostos Bicíclicos Heterocíclicos com Pontes/química , GMP Cíclico/metabolismo , Modelos Animais de Doenças , Cães , Feminino , Limbo da Córnea/metabolismo , Macaca fascicularis , Coelhos , Malha Trabecular/metabolismo , Fator de Crescimento Transformador beta2/farmacologiaRESUMO
Purpose: We studied the IOP-lowering effects of NCX 1741, a novel nitric oxide (NO)-donating derivative of the phosphodiesterase type-5 inhibitor, avanafil, in Cynomolgus monkey with laser-induced ocular hypertension (OHT-monkeys). NCX 1193 (NO-donating moiety), NCX 1744 (NCX 1741 without ester nitrate moiety), and travoprost (PGF2α analogue) were used for comparison. Ocular exposure after NCX 1741 dosing also was addressed. Methods: Vehicle (phosphate buffer pH 6.0, Kolliphor® 5%, DMSO 0.3%, benzalkonium chloride 0.02%), NCX 1741, NCX 1193, NCX 1744, or travoprost were instilled (30 µL; single dose) masked and conscious IOPs were measured by pneumatonometry. LC-MS/MS-based methods were employed to monitor ocular exposure of NCX 1741 and main metabolites after ocular dosing in New Zealand White rabbits. Results: NCX 1741 (2.2%, 0.8 µmol/eye) lowered IOP with an Emax (ΔΔIOP, IOP change vs. pre-dose and vehicle) between 5 and 8 h post-dosing (ΔΔIOP5h, -5.3 ± 2.0 mmHg and ΔΔIOP8h, -6.0 ± 2.1 mmHg). Conversely, equimolar (0.47%, 0.8 µmol/eye) NCX 1193 IOP-lowering effects were maximal 3 h post-dosing (ΔΔIOP3h, -4.7 ± 1.6 mmHg) and declined thereafter (ΔΔIOP5h, -1.6 ± 1.1 mmHg). In a follow-up study, NCX 1741 (1.5%, 0.5 µmol/eye) was more effective than NCX 1744 despite a similar duration. Further, NCX 1741 was as effective as travoprost (0.1%, 0.06 µmol/eye) at 5 and 8 h post-dosing (travoprost, ΔΔIOP5h, -3.4 ± 2.2 mmHg and ΔΔIOP8h, -4.9 ± 1.3 mmHg) but had shorter duration (NCX 1741, ΔΔIOP24h, -1.5 ± 1.1 mmHg; travoprost, ΔΔIOP24h, -7.1 ± 2.8 mmHg). NCX 1741 resulted in significant aqueous humor exposure, as determined by the levels of the main metabolite, avanafil. Conclusions: NCX 1741 rapidly and effectively lowers IOP in OHT-monkeys for several hours post-dosing. How these effects translate in humans is still to be defined.
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Dinoprosta/análogos & derivados , Pressão Intraocular/efeitos dos fármacos , Hipertensão Ocular/tratamento farmacológico , Pirimidinas/farmacologia , Animais , Anti-Infecciosos Locais/administração & dosagem , Anti-Hipertensivos/administração & dosagem , Anti-Hipertensivos/farmacologia , Humor Aquoso/efeitos dos fármacos , Humor Aquoso/metabolismo , Compostos de Benzalcônio/administração & dosagem , Cromatografia Líquida/métodos , Feminino , Seguimentos , Macaca fascicularis , Modelos Animais , Óxido Nítrico/metabolismo , Doadores de Óxido Nítrico/metabolismo , Inibidores da Fosfodiesterase 5/metabolismo , Pirimidinas/administração & dosagem , Pirimidinas/uso terapêutico , Coelhos , Espectrometria de Massas em Tandem/métodos , Tonometria Ocular/métodos , Travoprost/administração & dosagem , Travoprost/farmacologiaRESUMO
In patients with ocular hypertension or glaucoma, all treatments aim to lower intraocular pressure (IOP) by modulating aqueous humour (AH) production and/or uveoscleral and trabecular meshwork/Schlemm's canal AH drainage. PG analogues are considered to be the 'gold standard' treatment and are the most frequently used IOP-lowering agents. Recent data support an important role for NO in regulating IOP. Thus, novel PG analogues carrying a NO-donating moiety were recently advanced. Latanoprostene bunod (LBN) and NCX 470, NO-donating derivatives of latanoprost and bimatoprost, respectively, are examples of such compounds. LBN ophthalmic solution, 0.024% (Vyzulta™), showed greater IOP-lowering efficacy compared with that of Xalatan® (latanoprost ophthalmic solution, 0.005%) or 0.5% timolol maleate in clinical settings. NCX 470 was found to be more effective than bimatoprost in animal models of ocular hypertension and glaucoma. Selective EP2 receptor agonists (i.e. taprenepag isopropyl, omidenepag isopropyl and aganepag isopropyl) and non-selective prostanoid receptor agonists (i.e. ONO-9054, sepetaprost isopropyl) that concomitantly stimulate FP and EP3 receptors have also been shown to hold promise as effective IOP-lowering agents. LINKED ARTICLES: This article is part of a themed section on Eicosanoids 35 years from the 1982 Nobel: where are we now? To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.8/issuetoc.
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Glaucoma/tratamento farmacológico , Glaucoma/metabolismo , Óxido Nítrico/metabolismo , Hipertensão Ocular/tratamento farmacológico , Hipertensão Ocular/metabolismo , Prostaglandinas Sintéticas/administração & dosagem , Animais , Ensaios Clínicos como Assunto/métodos , Humanos , Soluções Oftálmicas/administração & dosagem , Receptores de Prostaglandina/agonistas , Receptores de Prostaglandina/metabolismo , Resultado do TratamentoRESUMO
Nitric oxide (NO) donors may be useful for treating pulmonary hypertension (PH) complicating sickle cell disease (SCD), as endogenous NO is inactivated by hemoglobin released by intravascular hemolysis. Here, we investigated the effects of the new NO donor NCX1443 on PH in transgenic SAD mice, which exhibit mild SCD without severe hemolytic anemia. In SAD and wild-type (WT) mice, the pulmonary pressure response to acute hypoxia was similar and was abolished by 100 mg/kg NCX1443. The level of PH was also similar in SAD and WT mice exposed to chronic hypoxia (9% O2) alone or with SU5416 and was similarly reduced by daily NCX1443 gavage. Compared with WT mice, SAD mice exhibited higher levels of HO-1, endothelial NO synthase, and PDE5 but similar levels of lung cyclic guanosine monophosphate. Cultured pulmonary artery smooth muscle cells from SAD mice grew faster than those from WT mice and had higher PDE5 protein levels. Combining NCX1443 and a PDE5 inhibitor suppressed the growth rate difference between SAD and WT cells and induced a larger reduction in hypoxic PH severity in SAD than in WT mice. By amplifying endogenous protective mechanisms, NCX1443 in combination with PDE5 inhibition may prove useful for treating PH complicating SCD.
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Anemia Falciforme/tratamento farmacológico , Anti-Hipertensivos/farmacologia , Pressão Arterial/efeitos dos fármacos , Hipertensão Pulmonar/prevenção & controle , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico/metabolismo , Artéria Pulmonar/efeitos dos fármacos , Anemia Falciforme/complicações , Anemia Falciforme/metabolismo , Animais , Anti-Hipertensivos/metabolismo , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , GMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 5/metabolismo , Modelos Animais de Doenças , Heme Oxigenase-1/metabolismo , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/metabolismo , Hipertensão Pulmonar/fisiopatologia , Hipóxia/complicações , Masculino , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Doadores de Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Inibidores da Fosfodiesterase 5/farmacologia , Artéria Pulmonar/metabolismo , Artéria Pulmonar/fisiopatologiaRESUMO
A small series of water-soluble NO-donor furoxans bearing a basic center at the 4-position, having a wide lipophilic-hydrophilic balance range, and endowed with different NO-release capacities, were synthesized and characterized. Selected members were studied for their IOP-lowering activity in the transient ocular hypertensive rabbit model at 1% dose. The most effective IOP-lowering products were compounds 3 and 7, whose activity 60min after administration was similar to that of Timolol. Notably, 7 was characterized by a long-lasting action. The IOP-lowering activity in this series of products appeared to be modulated by the lipophilic-hydrophilic balance rather than by the NO-donor capacity.
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Oxidiazóis/química , Animais , Anti-Hipertensivos/química , Anti-Hipertensivos/farmacologia , Anti-Hipertensivos/uso terapêutico , Modelos Animais de Doenças , Glaucoma/tratamento farmacológico , Pressão Intraocular/efeitos dos fármacos , Óxido Nítrico/metabolismo , Hipertensão Ocular/tratamento farmacológico , Hipertensão Ocular/patologia , Oxidiazóis/farmacologia , Oxidiazóis/uso terapêutico , Coelhos , Solubilidade , Timolol/farmacologia , Timolol/uso terapêuticoRESUMO
NCX1404 [(3S)-5-methyl-3-(((1-(4-(nitrooxy)butanoyloxy)ethoxy)carbonylamino) methyl)hexanoic acid] is a novel nitric oxide (NO)-donating pregabalin that is readily absorbed and processed in vivo to pregabalin and NO. We determined the antiallodynic response of NCX1404 after acute or after 7, 14, and 21 days of repeated daily oral dosing in mice with streptozotocin (STZ)-induced painful diabetic neuropathy (PDN). Pregabalin and its combination with the NO donor isosorbide mononitrate (ISMN) were used for comparison. The blood levels of pregabalin and nitrites, used as surrogate marker of NO release, after NCX1404 or pregabalin dosing were monitored in parallel experiments using liquid chromatography with tandem mass spectrometry (LC-MS/MS). NCX1404 and pregabalin resulted in similar pregabalin levels as it was their antiallodynic activity after acute dosing in STZ mice. However, NCX1404 resulted in disease-modifying properties when administered daily for 21 days, as indicated by the time- and dose-dependent reversal of STZ-induced mechanical allodynia (paw withdrawal threshold [PWT]Veh_21d= 1.3 ± 0.15 g for vehicle; PWTNCX1404_21d= 1.4 ± 0.5 g, 2.9 ± 0.2 g* and 4.1 ± 0.2 g*, respectively for 19, 63, and 190µmol/kg, oral gavage [PO] of NCX1404; *P< 0.05 versus vehicle). This effect was not shared by pregabalin at equimolar doses (190µmol/kg, PO, PWTPregab_21d= 1.4 ± 0.1 g*, *P< 0.05 versus equimolar NCX1404). In addition, the NO donor ISMN (52.3µmol/kg, PO) alone or combined with pregabalin (63µmol/kg) was active at 7 days (PWTVeh_7d= 1.7 ± 0.16 g; PWTISMN_7d= 3.9 ± 0.34 g*; PWTPregab_7d= 1.3 ± 0.07 g; PWTISMN+pregab_7d= 3.8 ± 0.29 g*; *P< 0.05) but not at later time points. The long-term effect of NCX1404 was independent of residual drug exposure and lasted for several days after the treatment was stopped. In summary, like pregabalin, NCX1404 is an effective antiallodynic agent. Differently from pregabalin, repeated dosing of NCX1404 re-established normal nociceptive responses in STZ-induced PDN in mice.
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Diabetes Mellitus Experimental/complicações , Neuropatias Diabéticas/tratamento farmacológico , Neuralgia/tratamento farmacológico , Nitratos/uso terapêutico , Doadores de Óxido Nítrico/uso terapêutico , Nociceptividade/efeitos dos fármacos , Pregabalina/análogos & derivados , Pregabalina/uso terapêutico , Animais , Glicemia/análise , Diabetes Mellitus Experimental/metabolismo , Neuropatias Diabéticas/metabolismo , Relação Dose-Resposta a Droga , Hiperalgesia/tratamento farmacológico , Dinitrato de Isossorbida/uso terapêutico , Masculino , Camundongos , Neuralgia/etiologia , Neuralgia/metabolismo , Medição da Dor/efeitos dos fármacos , Pregabalina/metabolismoRESUMO
PURPOSE: The prostaglandin F2alpha (PGF2α) analogue bimatoprost lowers intraocular pressure (IOP) by increasing uveoscleral outflow at doses shown to elicit redness of the eye. With the aim to enhance the IOP-lowering effect of bimatoprost we studied NCX 470 [(S,E)-1-((1R,2R,3S,5R)-2-((Z)-7-(ethylamino)-7-oxohept-2-enyl)-3,5-dihydroxycyclopentyl)-5-phenylpent-1-en-3-yl 6-(nitrooxy)hexanoate], a dual-acting compound combining bimatoprost with nitric oxide (NO) known to mainly act via relaxation of trabecular meshwork and Schlemm's canal. METHODS: New Zealand white rabbits with transient hypertonic saline-induced IOP elevation (tOHT-rabbits), cynomolgus monkeys with laser-induced ocular hypertension (OHT-monkeys), and normotensive dogs (ONT-dogs) were used. The levels of NCX 470, bimatoprost, and bimatoprost acid were determined in aqueous humor (AH), cornea (CR), and iris/ciliary body (ICB) by liquid chromatography-mass spectrometry/mass (LC-MS/MS), while cGMP in AH and ICB was monitored using an enzyme immunoassay (EIA) kit in pigmented Dutch Belted rabbits. RESULTS: NCX 470 (0.14%, 30 µL) lowered IOP in tOHT-rabbits with an E(max) of -7.2 ± 2.8 mm Hg at 90 minutes. Bimatoprost at equimolar dose (0.1%, 30 µL) was noneffective in this model. NCX 470 (0.042%, 30 µL) was more effective than equimolar (0.03%, 30 µL) bimatoprost in ONT-dogs (IOP change, -5.4 ± 0.7 and -3.4 ± 0.7 mm Hg, respectively, P < 0.05) and in OHT-monkeys (IOP change, -7.7 ± 1.4 and -4.8 ± 1.7 mm Hg, respectively, P < 0.05) at 18 hours post dosing. NCX 470 (0.042%, 30 µL) or bimatoprost (0.03%, 30 µL) resulted in similar bimatoprost acid exposure in AH, CR, and ICB while cGMP was significantly increased in AH and ICB at 18 and 24 hours after NCX 470 dosing. CONCLUSIONS: NCX 470 lowers IOP more than equimolar bimatoprost in three animal models of glaucoma by activating PGF2α and NO/cGMP signaling pathways.
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Humor Aquoso/metabolismo , Bimatoprost/farmacocinética , Corpo Ciliar/metabolismo , Pressão Intraocular/efeitos dos fármacos , Doadores de Óxido Nítrico/farmacocinética , Hipertensão Ocular/tratamento farmacológico , Animais , Anti-Hipertensivos/farmacocinética , Modelos Animais de Doenças , Cães , Macaca fascicularis , Masculino , Hipertensão Ocular/metabolismo , Hipertensão Ocular/fisiopatologia , Coelhos , Espectrometria de Massas em TandemRESUMO
The predominant risk factor for the progression of glaucoma is an increase in IOP, mediated via a reduction in aqueous outflow through the conventional (trabecular meshwork and Schlemm's canal) outflow pathway. Current IOP lowering pharmacological strategies target the uveoscleral (nonconventional) outflow pathway or aqueous humor production; however, to date no therapy that primarily targets the conventional pathway exists. Nitric oxide (NO) is an intracellular signaling molecule produced by endogenous NO synthases, well-known for its key role in vasodilation, through its action on smooth muscle cells. Under physiological conditions, NO mediates a multitude of diverse ocular effects, including maintenance of IOP. Nitric oxide donors have been shown to mediate IOP-lowering effects in both preclinical models and clinical studies, primarily through cell volume and contractility changes in the conventional outflow tissues. This review is focused on evaluating the current knowledge of the role and mechanism of action of endogenous NO and NO donors in IOP regulation. Data on key additional functions of NO in glaucoma pathology (i.e., ocular blood flow and effects on optic neuropathy) are also summarized. The potential for future therapeutic application of NO in the treatment of glaucoma is then discussed.
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Humor Aquoso/metabolismo , Glaucoma , Pressão Intraocular/efeitos dos fármacos , Óxido Nítrico/uso terapêutico , Estresse Oxidativo , Animais , Fatores Relaxantes Dependentes do Endotélio/uso terapêutico , Glaucoma/tratamento farmacológico , Glaucoma/metabolismo , Glaucoma/fisiopatologia , HumanosRESUMO
BACKGROUND: NCX 434 is a nitric oxide (NO)-donating triamcinolone acetonide (TA), shown to enhance optic nerve head (ONH) oxygen saturation in non-human primate eyes. Here, the effects of a single intravitreal (IVT) injection of TA were compared with those of NCX 434 on intraocular pressure (IOP), retinal function and retrobulbar haemodymamics in endothelin-1 (ET-1) induced ONH ischaemia/reperfusion in rabbits. Biochemical changes were also assessed in the aqueous humour and in retinal biopsies. METHODS: IOP and resistivity index of ophthalmic artery (RI-OA) were recorded using TonoPen and ecocolor Doppler, respectively. Retinal function was assessed using photopic electroretinography. Cytokine expression and oxidative stress markers were evaluated with immunoassay techniques. RESULTS: At 4 weeks post IVT treatment, TA increased IOP and RI-OA while NCX 434 did not (IOP(Vehicle)=13.6±1.3, IOP(NCX 434)=16.9±2.2, IOP(TA)=20.9±1.9 mm Hg; p<0.05 vs vehicle; RI-OA(Vehicle)=0.44±0.03; RI-OA(NCX 434)=0.47±0.02; RI-OA(TA)=0.60±0.04). Both NCX 434 and TA reversed ET-1 induced decrease in electroretinography amplitude to similar extents. NCX 434 attenuated ET-1 induced oxidative stress markers and nitrotyrosine in retinal tissue, and interleukin-6 and tumour necrosis factor-α in aqueous humour more effectively than TA. CONCLUSION: NCX 434 attenuates ET-1 induced ischaemia/reperfusion damage without increasing IOP, probably due to NO release. If data are confirmed in other species and models, this compound could represent an interesting new therapeutic option for retinal and ONH diseases, including diabetic retinopathy.
Assuntos
Endotelina-1/toxicidade , Pressão Intraocular/efeitos dos fármacos , Nitratos/farmacologia , Doadores de Óxido Nítrico/farmacologia , Traumatismo por Reperfusão/prevenção & controle , Retina/efeitos dos fármacos , Doenças Retinianas/prevenção & controle , Triancinolona Acetonida/análogos & derivados , Animais , Caspase 3/metabolismo , Citocinas/metabolismo , Eletrorretinografia , Glutationa/metabolismo , Hemodinâmica , Injeções Intravítreas , Fluxometria por Laser-Doppler , Masculino , Artéria Oftálmica/fisiologia , Disco Óptico/irrigação sanguínea , Coelhos , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/fisiopatologia , Retina/metabolismo , Doenças Retinianas/metabolismo , Doenças Retinianas/fisiopatologia , Superóxido Dismutase/metabolismo , Tonometria Ocular , Triancinolona Acetonida/farmacologia , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMO
PURPOSE: 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. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.
Assuntos
Anti-Hipertensivos/farmacologia , Modelos Animais de Doenças , Glaucoma/tratamento farmacológico , Pressão Intraocular/efeitos dos fármacos , Óxido Nítrico/metabolismo , Prostaglandinas F Sintéticas/farmacologia , Prostaglandinas Sintéticas/farmacologia , Animais , Humor Aquoso/metabolismo , Corpo Ciliar/metabolismo , GMP Cíclico/metabolismo , Cães , Feminino , Glaucoma/metabolismo , Iris/metabolismo , Macaca fascicularis , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Óxido Nítrico Sintase Tipo II/antagonistas & inibidores , Hipertensão Ocular/tratamento farmacológico , Hipertensão Ocular/metabolismo , Soluções Oftálmicas/farmacologia , Prostaglandinas F Sintéticas/síntese química , Coelhos , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/metabolismoRESUMO
Adenosine A2A receptor antagonists provide a promising nondopaminergic approach to the treatment of Parkinson's disease (PD). Initial clinical trials of A2A antagonists targeted PD patients who had already developed treatment complications known as L-3,4-dihydroxyphenylalanine (L-DOPA)-induced dyskinesia (LID) in an effort to improve symptoms while reducing existing LID. The goal of this study is to explore the effect of A2A antagonists and targeted A2A receptor depletion on the actual development of sensitized responses to L-DOPA in mouse models of LID in PD. Hemiparkinsonian mice (unilaterally lesioned with 6-OHDA) were treated daily for 3 weeks with a low dose of L-DOPA (2 mg/kg) preceded by a low dose of selective A2A antagonist (KW-6002 [(E)-1,3-diethyl-8-(3,4-dimethoxystyryl)-7-methyl-3,7-dihydro-1H-purine-2,6-dione] at 0.03 or 0.3 mg/kg, or SCH58261 [5-amino-7-(2-phenylethyl)-2-(2-furyl)-pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine] at 0.03 mg/kg) or vehicle intraperitoneally. In control mice, contralateral rotational responses to daily L-DOPA gradually increased over the initial week before reaching a persistent maximum. Both A2A antagonists inhibited the development of sensitized contralateral turning, with KW-6002 pretreatment reducing the sensitized rotational responses by up to threefold. The development of abnormal involuntary movements (a measure of LID) as well as rotational responses was attenuated by the postnatal depletion of forebrain A2A receptors in conditional (Cre/loxP system) knock-out mice. These pharmacological and genetic data provide evidence that striatal A2A receptors play an important role in the neuroplasticity underlying behavioral sensitization to L-DOPA, supporting consideration of early adjunctive therapy with an A2A antagonist to reduce the risk of LID in PD.
Assuntos
Discinesia Induzida por Medicamentos/metabolismo , Levodopa/toxicidade , Transtornos Parkinsonianos/metabolismo , Prosencéfalo/fisiologia , Receptor A2A de Adenosina/fisiologia , Antagonistas do Receptor A2 de Adenosina , Animais , Discinesia Induzida por Medicamentos/tratamento farmacológico , Levodopa/farmacologia , Levodopa/uso terapêutico , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Oxidopamina/toxicidade , Transtornos Parkinsonianos/induzido quimicamente , Transtornos Parkinsonianos/tratamento farmacológico , Prosencéfalo/efeitos dos fármacos , Prosencéfalo/metabolismo , Purinas/farmacologia , Purinas/uso terapêutico , Receptor A2A de Adenosina/genéticaAssuntos
Ascaríase , Ascaris lumbricoides , Diverticulite/parasitologia , Divertículo Ileal/parasitologia , Adulto , Animais , Humanos , MasculinoRESUMO
To improve protein delivery to the CNS following intracerebroventricular administration, we compared the distribution of a human Cu/Zn superoxide dismutase:tetanus toxin fragment C fusion protein (SOD1:TTC) in mouse brain and spinal cord with that of tetanus toxin fragment C (TTC) or human SOD1 (hSOD1) alone, following continuous infusion into the lateral ventricle. Mice infused with TTC or SOD1:TTC showed intense anti-TTC or anti-hSOD1 labeling, respectively, throughout the CNS. In contrast, animals treated with hSOD1 revealed moderate staining in periventricular tissues. In spinal cord sections from animals infused with SOD1:TTC, the fusion protein was found in neuron nuclear antigen-positive (NeuN+) neurons and not glial fibrillary acidic protein-positive (GFAP+) astrocytes. The percentage of NeuN+ ventral horn cells that were co-labeled with hSOD1 antibody was greater in mice treated with SOD1:TTC (cervical cord = 73 +/- 8.5%; lumbar cord = 62 +/- 7.7%) than in mice treated with hSOD1 alone (cervical cord = 15 +/- 3.9%; lumbar cord = 27 +/-4.7%). Enzyme-linked immunosorbent assay for hSOD1 further demonstrated that SOD1:TTC-infused mice had higher levels of immunoreactive hSOD1 in CNS tissue extracts than hSOD1-infused mice. Following 24 h of drug washout, tissue extracts from SOD1:TTC-treated mice still contained substantial amounts of hSOD1, while extracts from hSOD1-treated mice lacked detectable hSOD1. Immunoprecipitation of SOD1:TTC from these extracts using anti-TTC antibody revealed that the recovered fusion protein was structurally intact and enzymatically active. These results indicate that TTC may serve as a useful prototype for development as a non-viral vehicle for improving delivery of therapeutic proteins to the CNS.
Assuntos
Sistema Nervoso Central/citologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fragmentos de Peptídeos/farmacologia , Superóxido Dismutase/líquido cefalorraquidiano , Toxina Tetânica/farmacologia , Animais , Western Blotting/métodos , Contagem de Células/métodos , Sistema Nervoso Central/efeitos dos fármacos , Humanos , Imuno-Histoquímica/métodos , Injeções Intraventriculares/métodos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fragmentos de Peptídeos/metabolismo , Fosfopiruvato Hidratase , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia , Superóxido Dismutase-1 , Toxina Tetânica/metabolismo , Distribuição Tecidual/efeitos dos fármacosRESUMO
In the pursuit of improved treatments for Parkinson's disease (PD), the adenosine A(2A) receptor has emerged as an attractive nondopaminergic target. Based on the compelling behavioral pharmacology and selective basal ganglia expression of this G-protein-coupled receptor, its antagonists are now crossing the threshold of clinical development as adjunctive symptomatic treatment for relatively advanced PD. The antiparkinsonian potential of A(2A) antagonism has been boosted further by recent preclinical evidence that A(2A) antagonists might favorably alter the course as well as the symptoms of the disease. Convergent epidemiological and laboratory data have suggested that A(2A) blockade may confer neuroprotection against the underlying dopaminergic neuron degeneration. In addition, rodent and nonhuman primate studies have raised the possibility that A(2A) receptor activation contributes to the pathophysiology of dyskinesias-problematic motor complications of standard PD therapy--and that A(2A) antagonism might help prevent them. Realistically, despite being targeted to basal ganglia pathophysiology, A(2A) antagonists may be expected to have other beneficial and adverse effects elsewhere in the central nervous system (e.g., on mood and sleep) and in the periphery (e.g., on immune and inflammatory processes). The thoughtful design of new clinical trials of A(2A) antagonists should take into consideration these counterbalancing hopes and concerns and may do well to shift toward a broader set of disease-modifying as well as symptomatic indications in early PD.
Assuntos
Antagonistas do Receptor A2 de Adenosina , Encéfalo/efeitos dos fármacos , Cafeína/uso terapêutico , Estimulantes do Sistema Nervoso Central/uso terapêutico , Doença de Parkinson/tratamento farmacológico , Purinas/uso terapêutico , Animais , Encéfalo/anatomia & histologia , Encéfalo/metabolismo , Ensaios Clínicos como Assunto , Humanos , Doença de Parkinson/fisiopatologia , Doença de Parkinson/prevenção & controle , Receptores A2 de Adenosina/fisiologiaRESUMO
Adenosine A(2A) receptors (A(2A)Rs) are well positioned to influence the maladaptive CNS responses to repeated dopaminergic stimulation in psychostimulant addiction. Expression of A(2A)Rs in brain is largely restricted to the nucleus accumbens and striatum, where molecular adaptations mediate chronic effects of psychostimulants such as behavioral sensitization. Using a novel forebrain-specific conditional (Cre/loxP system) knockout of the A(2A)R in coordination with classical pharmacological approaches, we investigated the involvement of brain A(2A)Rs in amphetamine-induced behavioral sensitization. Tissue-specific, functional disruption of the receptor was confirmed by autoradiography, PCR, and the loss of A(2A) antagonist-induced motor stimulation. Daily treatment with amphetamine for 1 week markedly enhanced locomotor responses on day 8 in control mice and the sensitization remained robust after a week of washout. Their conditional knockout littermates however showed no sensitization to amphetamine on day 8 and only a modest sensitization following the washout. Pharmacological blockade of adenosine A(2A)Rs also was able to block the development (but not the expression) of sensitization in multiple mouse strains. Thus activation of brain A(2A)Rs plays a critical role in developing augmented psychomotor responses to repeated psychostimulant exposure.
Assuntos
Anfetamina/farmacologia , Estimulantes do Sistema Nervoso Central/farmacologia , Prosencéfalo/efeitos dos fármacos , Receptor A2A de Adenosina/efeitos dos fármacos , Animais , Autorradiografia , Genótipo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Atividade Motora/efeitos dos fármacos , Desempenho Psicomotor/efeitos dos fármacos , Desempenho Psicomotor/fisiologia , Receptor A2A de Adenosina/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Interactions between dopaminergic and glutamatergic systems in the striatum are thought to underlie both the symptoms and adverse effects of treatment of Parkinson's disease. We have previously reported that activation of the dopamine D1 receptor triggers a rapid redistribution of striatal N-methyl-d-aspartate (NMDA) receptors between intracellular and postsynaptic sub-cellular compartments. To unravel the signaling pathways underlying this trafficking, we studied mice with targeted disruptions of either the gene that encodes the dopamine- and cAMP-regulated phosphoprotein (DARPP-32), a potent and selective inhibitor of protein phosphatase-1, or the protein tyrosine kinase Fyn. In striatal tissue from DARPP-32-depleted mice, basal tyrosine and serine phosphorylation of striatal NMDA receptor subunits NR1, NR2A, and NR2B was normal, and activation of dopamine D1 receptors with the agonist SKF-82958 [(+/-)-6-chloro-7,8-dihydroxy-3-allyl-1-phenyl-2,3,4,5-tetra-hydro-1H-benzazepine] produced redistribution of NMDA receptors from vesicular compartments (P3 and LP2) to synaptosomal membranes (LP1). In the Fyn knockout mice, basal tyrosine phosphorylation of NR2A and NR2B was drastically reduced, whereas serine phosphorylation of these NMDA subunits was unchanged. In the Fyn knockout mice, the dopamine D1 receptor agonist failed to induce subcellular redistribution of NMDA receptors. In addition, Fyn-depleted mice lesioned with 6-hydroxydopamine also failed to exhibit l-DOPA-induced behavioral sensitization, but this may be caused, at least in part, by resistance of these mice to the neurotoxic lesion. These findings suggest a novel mechanism for the trafficking of striatal NMDA receptors by signaling pathways that are independent of DARPP-32 but require Fyn protein tyrosine kinase. Strategies that prevent NMDA receptor subcellular redistribution through inhibition of Fyn kinase may prove useful in the treatment of Parkinson's disease.
Assuntos
Dopamina/metabolismo , Proteínas do Tecido Nervoso , Fosfoproteínas/metabolismo , Receptores de Dopamina D1/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Animais , Corpo Estriado/efeitos dos fármacos , Corpo Estriado/metabolismo , Fosfoproteína 32 Regulada por cAMP e Dopamina , Expressão Gênica , Levodopa/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais , Oxidopamina/farmacologia , Doença de Parkinson/metabolismo , Fosforilação , Proteínas Tirosina Quinases , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-fyn , Frações SubcelularesRESUMO
The A2A receptor has recently attracted considerable interest as a potential target for Parkinson's disease (PD) therapy based on the motor-enhancing and neuroprotective effects of A2A antagonists in animal models of PD. The unique neuronal localization of the adenosine A2A receptor in the basal ganglia and its extensive interactions with dopaminergic and glutamatergic systems led the authors to investigate a potential role of the A2A receptor in the development of behavioral sensitization in response to repeated dopaminergic stimulation. Because dopamine-induced behavioral sensitization shares several neurochemical and behavioral features with dyskinesia, characterizing this novel aspect of A2A receptor function may enhance understanding and management of dyskinesia in PD. Recent studies from several laboratories suggest that the A2A receptor may be an important mediator of maladaptive changes in response to long-term dopamine stimulation. The authors summarize their investigation of the role of A2A receptors in two paradigms of behavioral sensitization elicited by daily treatment with either L-dopa in hemiparkinsonian mice or amphetamine in naive mice. The results demonstrate that the A2A receptor is required for the development of behavioral sensitization in response to repeated L-dopa treatment in hemiparkinsonian mice and repeated amphetamine administration in normal mice. Together with pharmacologic studies, these results raise the possibility that the maladaptive dyskinetic responses to long-term L-dopa management of PD may be attenuated by A2A receptor blockade. Potential presynaptic, postsynaptic (cellular), and trans-synaptic (network) mechanisms are discussed.