RESUMEN
INTRODUCTION: The adenine model of kidney disease typically involves dietary delivery of adenine over several weeks. This model can be variable in its disease progression and can result in significant mortality. In the current study, the amount of adenine delivered to rats was controlled by utilizing oral gavage administration over a short period in an attempt to induce robust renal pathology while addressing variability and viability of the animals. METHODS: Adenine (150 or 200â¯mg/kg) was administered via oral gavage for 10 consecutive days, and assessed over a total of 20â¯days. RESULTS: Both adenine dose groups manifested pathophysiological features of kidney disease such as proteinuria, elevated serum creatinine and BUN, and tubulointerstitial fibrosis. The animals also displayed a decline in glomerular filtration rate. Renal mRNA expression of genes associated with injury, inflammation, and fibrosis (i.e., Col1a1, Acta2, Serpine1, Timp1, Fn-Eda, Tgfb1, Ccl2, Nlrp3, Aqp1 and Ccnd1) were elevated as were urinary biomarkers that have translational utility (i.e., clusterin, KIM-1, MCP-1, OPN, NGAL, B2M, calbindin, and cystatin C). All disease endpoints were more pronounced in the 200â¯mg/kg group, however, while measures of tissue fibrosis were sustained, there was partial recovery by day 20 in functional readouts. No mortality was observed in either dose group. DISCUSSION: Short-term delivery of adenine via precise gavage delivery induced a robust model with hallmarks of fibrotic kidney disease, had limited variance between animals, and no animal morbidity within the 20â¯days studied. This model represents a methodical alternative to long-term dietary dosing of adenine.
Asunto(s)
Adenina/administración & dosificación , Fibrosis/inducido químicamente , Enfermedades Renales/inducido químicamente , Riñón/efectos de los fármacos , Administración Oral , Animales , Biomarcadores/orina , Fibrosis/metabolismo , Fibrosis/orina , Tasa de Filtración Glomerular/efectos de los fármacos , Inflamación/inducido químicamente , Inflamación/metabolismo , Inflamación/orina , Riñón/metabolismo , Enfermedades Renales/metabolismo , Enfermedades Renales/orina , Masculino , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
Targeted delivery of a therapeutic agent to a site of pathology to ameliorate disease while limiting exposure at undesired tissues is an aspirational treatment scenario. Targeting diseased kidneys for pharmacologic treatment has had limited success. We designed an approach to target an extracellular matrix protein, the fibronectin extra domain A isoform (FnEDA), which is relatively restricted in distribution to sites of tissue injury. In a mouse unilateral ureteral obstruction (UUO) model of renal fibrosis, injury induced significant upregulation of FnEDA in the obstructed kidney. Using dual variable domain Ig (DVD-Ig) technology, we constructed a molecule with a moiety to target FnEDA and a second moiety to neutralize TGF-ß After systemic injection of the bispecific TGF-ß + FnEDA DVD-Ig or an FnEDA mAb, chemiluminescent detection and imaging with whole-body single-photon emission computed tomography (SPECT) revealed significantly higher levels of each molecule in the obstructed kidney than in the nonobstructed kidney, the ipsilateral kidney of sham animals, and other tissues. In comparison, a systemically administered TGF-ß mAb accumulated at lower concentrations in the obstructed kidney and exhibited a more diffuse whole-body distribution. Systemic administration of the bispecific DVD-Ig or the TGF-ß mAb (1-10 mg/kg) but not the FnEDA mAb attenuated the injury-induced collagen deposition detected by immunohistochemistry and elevation in Col1a1, FnEDA, and TIMP1 mRNA expression in the obstructed kidney. Overall, systemic delivery of a bispecific molecule targeting an extracellular matrix protein and delivering a TGF-ß mAb resulted in a relatively focal uptake in the fibrotic kidney and reduced renal fibrosis.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Enfermedades Renales/tratamiento farmacológico , Riñón/efectos de los fármacos , Factor de Crecimiento Transformador beta/antagonistas & inhibidores , Animales , Modelos Animales de Enfermedad , Matriz Extracelular/metabolismo , Fibronectinas/química , Fibrosis/tratamiento farmacológico , Humanos , Hibridomas/metabolismo , Riñón/diagnóstico por imagen , Riñón/patología , Masculino , Ratones , Tomografía Computarizada de Emisión de Fotón Único , Uréter/patologíaRESUMEN
Adenosine (ADO) is an important regulatory purine nucleoside that accumulates at sites of inflammation and tissue injury including in diseases associated with renal pathology. Endogenous levels of ADO may be increased by inhibiting the ADO-metabolizing enzyme, ADO kinase (AK). AK inhibitors have demonstrated protection in rodent models of diabetic nephropathy. To further investigate AK inhibition as a potential mechanism for renal protection, A-306989, a potent non-nucleoside AK inhibitor, was examined in both in vitro and in vivo assays of renal injury. A-306989 prevented podocyte damage (disruption of actin cytoskeleton) and increased podocyte survival following puromycin aminonucleoside (PAN) application in both mouse and human conditionally immortalized podocytes. Prophylactic oral administration of A-306989 (1.5, 5 and 15mg/kg) reduced proteinuria in a dose-dependent manner and repressed pro-inflammatory/fibrotic gene up-regulation; A-306989 was also efficacious when administered two days following the PAN-insult. A-306989 (10 and 30mg/kg) also significantly reduced proteinuria and macrophage infiltration in a rat model of glomerulonephritis. Finally, A-306989 (15 and 50mg/kg) reduced the expression levels of pro-inflammatory/fibrotic genes, and reduced macrophage infiltration (50mg/kg), but did not affect the deposition of interstitial collagen in fibrotic kidneys from mice with unilateral ureter obstruction. A-306989 also had beneficial actions on "quality of life" measures including improving body weight loss. Thus, these data indicate that enhancement of endogenous ADO levels by A-306989 can positively modulate renal pathology and mimic some of the previously reported beneficial actions of ADO A2A receptor agonists.
Asunto(s)
Adenosina Quinasa/antagonistas & inhibidores , Membrana Basal/diagnóstico por imagen , Citoprotección/efectos de los fármacos , Riñón/citología , Riñón/lesiones , Podocitos/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Animales , Fibrosis , Riñón/efectos de los fármacos , Riñón/patología , Masculino , Ratones , Podocitos/citología , Podocitos/metabolismo , Puromicina Aminonucleósido/toxicidad , RatasRESUMEN
UNLABELLED: Voltage-gated Ca(2+) channels play an important role in nociceptive transmission. There is significant evidence supporting a role for N-, T- and P/Q-type Ca(2+) channels in chronic pain. Here, we report that A-1264087, a structurally novel state-dependent blocker, inhibits each of these human Ca(2+) channels with similar potency (IC50 = 1-2 µM). A-1264087 was also shown to inhibit the release of the pronociceptive calcitonin gene-related peptide from rat dorsal root ganglion neurons. Oral administration of A-1264087 produces robust antinociceptive efficacy in monoiodoacetate-induced osteoarthritic, complete Freund adjuvant-induced inflammatory, and chronic constrictive injury of sciatic nerve-induced, neuropathic pain models with ED50 values of 3.0, 5.7, and 7.8 mg/kg (95% confidence interval = 2.2-3.5, 3.7-10, and 5.5-12.8 mg/kg), respectively. Further analysis revealed that A-1264087 also suppressed nociceptive-induced p38 and extracellular signal-regulated kinase 1/2 phosphorylation, which are biochemical markers of engagement of pain circuitry in chronic pain states. Additionally, A-1264087 inhibited both spontaneous and evoked neuronal activity in the spinal cord dorsal horn in complete Freund adjuvant-inflamed rats, providing a neurophysiological basis for the observed antihyperalgesia. A-1264087 produced no alteration of body temperature or motor coordination and no learning impairment at therapeutic plasma concentrations. PERSPECTIVE: The present results demonstrate that the neuronal Ca(2+) channel blocker A-1264087 exhibits broad-spectrum efficacy through engagement of nociceptive signaling pathways in preclinical pain models in the absence of effects on psychomotor and cognitive function.
Asunto(s)
Analgésicos/farmacología , Compuestos de Azabiciclo/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Leucina/análogos & derivados , Neuronas/metabolismo , Nocicepción/efectos de los fármacos , Médula Espinal/efectos de los fármacos , Animales , Modelos Animales de Enfermedad , Inmunohistoquímica , Leucina/farmacología , Masculino , Neuronas/efectos de los fármacos , Dolor/metabolismo , Técnicas de Placa-Clamp , Ratas Sprague-Dawley , Médula Espinal/metabolismoRESUMEN
N-, T- and P/Q-type voltage-gated Ca(2+) channels are critical for regulating neurotransmitter release and cellular excitability and have been implicated in mediating pathological nociception. A-1264087 is a novel state-dependent blocker of N-, T- and P/Q-type channels. In the present studies, A-1264087 blocked (IC50 = 1.6 µM) rat dorsal root ganglia N-type Ca(2+) in a state-dependent fashion. A-1264087 (1, 3 and 10 mg/kg po) dose-dependently reduced mechanical allodynia in rats with a spinal nerve ligation (SNL) injury. A-1264087 (4 mg/kg iv) inhibited both spontaneous and mechanically evoked activity of spinal wide dynamic range (WDR) neurons in SNL rats but had no effect in uninjured rats. The inhibitory effect on WDR neurons remained in spinally transected SNL rats. Injection of A-1264087 (10 nmol/0.5 µl) into the spinal cord reduced both spontaneous and evoked WDR activity in SNL rats. Application of A-1264087 (300 nmol/20 µl) into the receptive field on the hindpaw attenuated evoked but not spontaneous firing of WDR neurons. Using electrical stimulation, A-1264087 (4 mg/kg iv) inhibited Aδ- and C-fiber evoked responses and after-discharge of WDR neurons in SNL rats. These effects by A-1264087 were not present in uninjured rats. A-1264087 moderately attenuated WDR neuron windup in both uninjured and SNL rats. In summary, these results indicate that A-1264087 selectively inhibited spinal nociceptive transmission in sensitized states through both peripheral and central mechanisms.
Asunto(s)
Anestésicos/farmacología , Compuestos de Azabiciclo/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Leucina/análogos & derivados , Neuralgia/fisiopatología , Nocicepción/efectos de los fármacos , Nervios Periféricos/efectos de los fármacos , Nervios Espinales/efectos de los fármacos , Potenciales de Acción , Administración Cutánea , Anestésicos/administración & dosificación , Anestésicos/uso terapéutico , Animales , Compuestos de Azabiciclo/uso terapéutico , Bloqueadores de los Canales de Calcio/administración & dosificación , Bloqueadores de los Canales de Calcio/uso terapéutico , Relación Dosis-Respuesta a Droga , Hiperalgesia/tratamiento farmacológico , Hiperalgesia/fisiopatología , Concentración 50 Inhibidora , Inyecciones Espinales , Leucina/farmacología , Leucina/uso terapéutico , Masculino , Neuralgia/tratamiento farmacológico , Neuronas/efectos de los fármacos , Neuronas/fisiología , Nervios Periféricos/fisiología , Ratas , Ratas Sprague-Dawley , Nervios Espinales/fisiologíaRESUMEN
A novel series of N-type calcium channel inhibitors have been discovered. Optimization of potency and HT-ADME properties provides 4-aminocyclopentapyrrolidines with analgesic efficacy after oral dosing.
Asunto(s)
Analgésicos/química , Analgésicos/uso terapéutico , Bloqueadores de los Canales de Calcio/química , Bloqueadores de los Canales de Calcio/uso terapéutico , Canales de Calcio Tipo N/metabolismo , Neuralgia/tratamiento farmacológico , Administración Oral , Analgésicos/síntesis química , Analgésicos/metabolismo , Animales , Bloqueadores de los Canales de Calcio/síntesis química , Bloqueadores de los Canales de Calcio/metabolismo , Humanos , Microsomas/metabolismo , Ratas , Relación Estructura-ActividadRESUMEN
A novel 4-aminocyclopentapyrrolidine series of N-type Ca(2+) channel blockers have been discovered. Enantioselective synthesis of the 4-aminocyclopentapyrrolidines was enabled using N-tert-butyl sulfinamide chemistry. SAR studies demonstrate selectivity over L-type Ca(2+) channels. N-type Ca(2+) channel blockade was confirmed using electrophysiological recording techniques. Compound 25 is an N-type Ca(2+) channel blocker that produces antinociception in inflammatory and nociceptive pain models without exhibiting cardiovascular or motor liabilities.
Asunto(s)
Acetamidas/síntesis química , Analgésicos/síntesis química , Bloqueadores de los Canales de Calcio/síntesis química , Canales de Calcio Tipo N/química , Pirrolidinas/química , Pirrolidinas/síntesis química , Acetamidas/farmacología , Acetamidas/uso terapéutico , Analgésicos/farmacología , Analgésicos/uso terapéutico , Animales , Conducta Animal/efectos de los fármacos , Bloqueadores de los Canales de Calcio/farmacología , Bloqueadores de los Canales de Calcio/uso terapéutico , Canales de Calcio Tipo N/metabolismo , Modelos Animales de Enfermedad , Masculino , Dolor/tratamiento farmacológico , Pirrolidinas/farmacología , Pirrolidinas/uso terapéutico , Ratas , Ratas Sprague-Dawley , Relación Estructura-ActividadRESUMEN
Blockade of voltage-gated Ca²âº channels on sensory nerves attenuates neurotransmitter release and membrane hyperexcitability associated with chronic pain states. Identification of small molecule Ca²âº channel blockers that produce significant antinociception in the absence of deleterious hemodynamic effects has been challenging. In this report, two novel structurally related compounds, A-686085 and A-1048400, were identified that potently block N-type (IC50=0.8 µM and 1.4 µM, respectively) and T-type (IC50=4.6 µM and 1.2 µM, respectively) Ca²âº channels in FLIPR based Ca²âº flux assays. A-686085 also potently blocked L-type Ca²âº channels (EC50=0.6 µM), however, A-1048400 was much less active in blocking this channel (EC50=28 µM). Both compounds dose-dependently reversed tactile allodynia in a model of capsaicin-induced secondary hypersensitivity with similar potencies (EC50=300-365 ng/ml). However, A-686085 produced dose-related decreases in mean arterial pressure at antinociceptive plasma concentrations in the rat, while A-1048400 did not significantly alter hemodynamic function at supra-efficacious plasma concentrations. Electrophysiological studies demonstrated that A-1048400 blocks native N- and T-type Ca²âº currents in rat dorsal root ganglion neurons (IC50=3.0 µM and 1.6 µM, respectively) in a voltage-dependent fashion. In other experimental pain models, A-1048400 dose-dependently attenuated nociceptive, neuropathic and inflammatory pain at doses that did not alter psychomotor or hemodynamic function. The identification of A-1048400 provides further evidence that voltage-dependent inhibition of neuronal Ca²âº channels coupled with pharmacological selectivity vs. L-type Ca²âº channels can provide robust antinociception in the absence of deleterious effects on hemodynamic or psychomotor function.
Asunto(s)
Analgésicos/administración & dosificación , Bloqueadores de los Canales de Calcio/administración & dosificación , Hemodinámica/fisiología , Neuronas/fisiología , Dimensión del Dolor , Piperidonas/administración & dosificación , Piperidonas/química , Administración Oral , Animales , Relación Dosis-Respuesta a Droga , Células HEK293 , Hemodinámica/efectos de los fármacos , Humanos , Masculino , Neuronas/efectos de los fármacos , Dimensión del Dolor/efectos de los fármacos , Dimensión del Dolor/métodos , Ratas , Ratas Sprague-DawleyRESUMEN
Positive modulation of the neuronal nicotinic acetylcholine receptor (nAChR) α4ß2 subtype by selective positive allosteric modulator NS-9283 has shown to potentiate the nAChR agonist ABT-594-induced anti-allodynic activity in preclinical neuropathic pain. To determine whether this benefit can be extended beyond neuropathic pain, the present study examined the analgesic activity and adverse effect profile of co-administered NS-9283 and ABT-594 in a variety of preclinical models in rats. The effect of the combined therapy on drug-induced brain activities was also determined using pharmacological magnetic resonance imaging. In carrageenan-induced thermal hyperalgesia, co-administration of NS-9283 (3.5 µmol/kg, i.p.) induced a 6-fold leftward shift of the dose-response of ABT-594 (ED(50)=26 vs. 160 nmol/kg, i.p.). In the paw skin incision model of post-operative pain, co-administration of NS-9283 similarly induced a 6-fold leftward shift of ABT-594 (ED(50)=26 vs. 153 nmol/kg). In monoiodo-acetate induced knee joint pain, co-administration of NS-9283 enhanced the potency of ABT-594 by 5-fold (ED(50)=1.0 vs. 4.6 nmol/kg). In pharmacological MRI, co-administration of NS-9283 was shown to lead to a leftward shift of ABT-594 dose-response for cortical activation. ABT-594 induced CNS-related adverse effects were not exacerbated in presence of an efficacious dose of NS-9283 (3.5 µmol/kg). Acute challenge of NS-9283 produced no cross sensitization in nicotine-conditioned animals. These results demonstrate that selective positive allosteric modulation at the α4ß2 nAChR potentiates nAChR agonist-induced analgesic activity across neuropathic and nociceptive preclinical pain models without potentiating ABT-594-mediated adverse effects, suggesting that selective positive modulation of α4ß2 nAChR by PAM may represent a novel analgesic approach.
Asunto(s)
Analgésicos/uso terapéutico , Azetidinas/uso terapéutico , Agonistas Nicotínicos/uso terapéutico , Oxadiazoles/uso terapéutico , Dolor/tratamiento farmacológico , Piridinas/uso terapéutico , Receptores Nicotínicos/metabolismo , Regulación Alostérica , Analgésicos/administración & dosificación , Analgésicos/efectos adversos , Animales , Azetidinas/administración & dosificación , Azetidinas/efectos adversos , Conducta Animal/efectos de los fármacos , Temperatura Corporal/efectos de los fármacos , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Modelos Animales de Enfermedad , Quimioterapia Combinada , Imagen por Resonancia Magnética , Masculino , Agonistas Nicotínicos/administración & dosificación , Agonistas Nicotínicos/efectos adversos , Osteoartritis/tratamiento farmacológico , Osteoartritis/metabolismo , Oxadiazoles/administración & dosificación , Oxadiazoles/efectos adversos , Dolor/metabolismo , Piridinas/administración & dosificación , Piridinas/efectos adversos , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND AND PURPOSE: Cannabinoid CB2 receptor activation by selective agonists has been shown to produce analgesic effects in preclinical models of inflammatory and neuropathic pain. However, mechanisms underlying CB2-mediated analgesic effects remain largely unknown. The present study was conducted to elucidate the CB2 receptor expression in 'pain relevant' tissues and the potential sites of action of CB2 agonism in rats. EXPERIMENTAL APPROACH: Expression of cannabinoid receptor mRNA was evaluated by quantitative RT-PCR in dorsal root ganglia (DRGs), spinal cords, paws and several brain regions of sham, chronic inflammatory pain (CFA) and neuropathic pain (spinal nerve ligation, SNL) rats. The sites of CB2 mediated antinociception were evaluated in vivo following intra-DRG, intrathecal (i.t.) or intraplantar (i.paw) administration of potent CB2-selective agonists A-836339 and AM1241. KEY RESULTS: CB2 receptor gene expression was significantly up-regulated in DRGs (SNL and CFA), spinal cords (SNL) or paws (CFA) ipsilateral to injury under inflammatory and neuropathic pain conditions. Systemic A-836339 and AM1241 produced dose-dependent efficacy in both inflammatory and neuropathic pain models. Local administration of CB2 agonists also produced significant analgesic effects in SNL (intra-DRG and i.t.) and CFA (intra-DRG) pain models. In contrast to A-836339, i.paw administration of AM-1241 dose-relatedly reversed the CFA-induced thermal hyperalgesia, suggesting that different mechanisms may be contributing to its in vivo properties. CONCLUSIONS AND IMPLICATIONS: These results demonstrate that both DRG and spinal cord are important sites contributing to CB2 receptor-mediated analgesia and that the changes in CB2 receptor expression play a crucial role for the sites of action in regulating pain perception.
Asunto(s)
Analgésicos/farmacología , Neuralgia/tratamiento farmacológico , Dolor/tratamiento farmacológico , Receptor Cannabinoide CB2/metabolismo , Analgesia , Analgésicos/uso terapéutico , Animales , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Cannabinoides/farmacología , Cannabinoides/uso terapéutico , Modelos Animales de Enfermedad , Ganglios Espinales/efectos de los fármacos , Ganglios Espinales/metabolismo , Inflamación/inducido químicamente , Inflamación/tratamiento farmacológico , Masculino , Neuralgia/inducido químicamente , Péptidos Opioides/metabolismo , Dolor/metabolismo , Percepción del Dolor , ARN Mensajero/análisis , ARN Mensajero/genética , Ratas , Ratas Sprague-Dawley , Receptor Cannabinoide CB2/agonistas , Médula Espinal/efectos de los fármacos , Médula Espinal/metabolismo , Tiazoles/farmacología , Tiazoles/uso terapéuticoRESUMEN
The histamine H(4) receptor (H(4)R) is expressed primarily on cells involved in inflammation and immune responses. To determine the potential role of H(4)R in pain transmission, the effects of JNJ7777120, a potent and selective H(4) antagonist, were characterized in preclinical pain models. Administration of JNJ7777120 fully blocked neutrophil influx observed in a mouse zymosan-induced peritonitis model (ED(50)=17 mg/kg s.c., 95% CI=8.5-26) in a mast cell-dependent manner. JNJ7777120 potently reversed thermal hyperalgesia observed following intraplantar carrageenan injection of acute inflammatory pain (ED(50)=22 mg/kg i.p., 95% CI=10-35) in rats and significantly decreased the myeloperoxide activity in the carrageenan-injected paw. In contrast, no effects were produced by either H(1)R antagonist diphenhydramine, H(2)R antagonists ranitidine, or H(3)R antagonist ABT-239. JNJ7777120 also exhibited robust anti-nociceptive activity in persistent inflammatory (CFA) pain with an ED(50) of 29 mg/kg i.p. (95% CI=19-40) and effectively reversed monoiodoacetate (MIA)-induced osteoarthritic joint pain. This compound also produced dose-dependent anti-allodynic effects in the spinal nerve ligation (ED(50)=60 mg/kg) and sciatic nerve constriction injury (ED(50)=88 mg/kg) models of chronic neuropathic pain, as well as in a skin-incision model of acute post-operative pain (ED(50)=68 mg/kg). In addition, the analgesic effects of JNJ7777120 were maintained following repeated administration and were evident at the doses that did not cause neurologic deficits in rotarod test. Our results demonstrate that selective blockade of H(4) receptors in vivo produces significant anti-nociception in animal models of inflammatory and neuropathic pain.
Asunto(s)
Analgésicos/farmacología , Modelos Animales de Enfermedad , Inflamación/tratamiento farmacológico , Enfermedades del Sistema Nervioso Periférico/tratamiento farmacológico , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Analgésicos/uso terapéutico , Animales , Masculino , Ratones , Ratones Endogámicos BALB C , Ensayo de Unión Radioligante , Ratas , Receptores Histamínicos , Receptores Histamínicos H4RESUMEN
Several 3-acylindoles with high affinity for the CB(2) cannabinoid receptor and selectivity over the CB(1) receptor have been prepared. A variety of 3-acyl substituents were investigated, and the tetramethylcyclopropyl group was found to lead to high affinity CB(2) agonists (5, 16). Substitution at the N1-indole position was then examined. A series of aminoalkylindoles was prepared and several substituted aminoethyl derivatives were active (23-27, 5) at the CB(2) receptor. A study of N1 nonaromatic side chain variants provided potent agonists at the CB(2) receptor (16, 35-41, 44-47, 49-54, and 57-58). Several polar side chains (alcohols, oxazolidinone) were well-tolerated for CB(2) receptor activity (41, 50), while others (amide, acid) led to weaker or inactive compounds (55 and 56). N1 aromatic side chains also afforded several high affinity CB(2) receptor agonists (61, 63, 65, and 69) but were generally less potent in an in vitro CB(2) functional assay than were nonaromatic side chain analogues.
Asunto(s)
Indoles/farmacología , Cetonas/farmacología , Receptor Cannabinoide CB2/agonistas , Diseño de Fármacos , Humanos , Indoles/síntesis química , Indoles/química , Cetonas/síntesis química , Cetonas/química , Ligandos , Estructura Molecular , Receptor Cannabinoide CB1/agonistas , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
Studies demonstrating the antihyperalgesic and antiallodynic effects of cannabinoid CB(2) receptor activation have been largely derived from the use of receptor-selective ligands. Here, we report the identification of A-836339 [2,2,3,3-tetramethyl-cyclopropanecarboxylic acid [3-(2-methoxy-ethyl)-4,5-dimethyl-3H-thiazol-(2Z)-ylidene]-amide], a potent and selective CB(2) agonist as characterized in in vitro pharmacological assays and in in vivo models of pain and central nervous system (CNS) behavior models. In radioligand binding assays, A-836339 displays high affinities at CB(2) receptors and selectivity over CB(1) receptors in both human and rat. Likewise, A-836339 exhibits high potencies at CB(2) and selectivity over CB(1) receptors in recombinant fluorescence imaging plate reader and cyclase functional assays. In addition A-836339 exhibits a profile devoid of significant affinity at other G-protein-coupled receptors and ion channels. A-836339 was characterized extensively in various animal pain models. In the complete Freund's adjuvant model of inflammatory pain, A-836339 exhibits a potent CB(2) receptor-mediated antihyperalgesic effect that is independent of CB(1) or mu-opioid receptors. A-836339 has also demonstrated efficacies in the chronic constrain injury (CCI) model of neuropathic pain, skin incision, and capsaicin-induced secondary mechanical hyperalgesia models. Furthermore, no tolerance was developed in the CCI model after subchronic treatment with A-836339 for 5 days. In assessing CNS effects, A-836339 exhibited a CB(1) receptor-mediated decrease of spontaneous locomotor activities at a higher dose, a finding consistent with the CNS activation pattern observed by pharmacological magnetic resonance imaging. These data demonstrate that A-836339 is a useful tool for use of studying CB(2) receptor pharmacology and for investigation of the role of CB(2) receptor modulation for treatment of pain in preclinical animal models.
Asunto(s)
Amidas/farmacología , Ciclopropanos/farmacología , Inflamación/fisiopatología , Dolor/fisiopatología , Receptor Cannabinoide CB1/fisiología , Receptor Cannabinoide CB2/fisiología , Animales , Células CHO , Línea Celular , Cricetinae , Cricetulus , Procedimientos Quirúrgicos Dermatologicos , Miembro Posterior , Humanos , Hiperalgesia/fisiopatología , Riñón/embriología , Imagen por Resonancia Magnética/métodos , Masculino , Dolor Postoperatorio/fisiopatología , Desempeño Psicomotor/efectos de los fármacos , Desempeño Psicomotor/fisiología , Ratas , Ratas Sprague-Dawley , Receptor Cannabinoide CB2/agonistasRESUMEN
TRPV1 receptors are activated and/or modulated by noxious heat, capsaicin, protons and other endogenous agents released following tissue injury. There is a growing appreciation that this molecular integrator may also have a role in mechanosensation. To further understand this role, we investigated the systemic and site-specific effects of a selective TRPV1 receptor antagonist, A-889425, on low-intensity mechanical stimulation in inflamed rats. Systemic administration of A-889425 (30 and 100 micromol/kg po) reduced mechanical allodynia in complete Freund's adjuvant (CFA)-inflamed rats. Systemic A-889425 (3 and 10 micromol/kg iv) also decreased the responses of spinal wide dynamic range (WDR) neurons to low-intensity mechanical stimulation in CFA-inflamed but not uninjured rats. This effect of A-889425 was likely mediated via multiple sites since local injection of A-889425 into the spinal cord (1-3 nmol), ipsilateral hindpaw (200 nmol), and cerebral ventricles (30-300 nmol) all attenuated WDR responses to low-intensity mechanical stimulation. In addition to an effect on mechanotransmission, systemic administration of A-889425 reduced the spontaneous firing of WDR neurons in inflamed but not uninjured rats. Spontaneous firing is elevated after injury and may reflect ongoing pain in the animal. Local injection experiments indicated that this effect of A-889425 on spontaneous firing was mainly mediated via TRPV1 receptors in the spinal cord. Thus the current data demonstrate that TRPV1 receptors have an enhanced role after an inflammatory injury, impacting both low-intensity mechanotransmission and possibly spontaneous pain. Furthermore this study delineates the differential contribution of central and peripheral TRPV1 receptors to affect spontaneous or mechanically evoked firing of WDR neurons.
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Potenciales de Acción/fisiología , Inflamación/patología , Neuronas/fisiología , Médula Espinal/patología , Canales Catiónicos TRPV/fisiología , Potenciales de Acción/efectos de los fármacos , Animales , Conducta Animal/efectos de los fármacos , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Potenciales Evocados Somatosensoriales/efectos de los fármacos , Adyuvante de Freund/farmacología , Inflamación/inducido químicamente , Masculino , Neuronas/efectos de los fármacos , Umbral del Dolor/efectos de los fármacos , Estimulación Física/métodos , Piridinas/química , Piridinas/farmacología , Ratas , Ratas Sprague-Dawley , Canales Catiónicos TRPV/antagonistas & inhibidoresRESUMEN
The synthesis and pharmacological characterization of a novel furan-based class of voltage-gated sodium channel blockers is reported. Compounds were evaluated for their ability to block the tetrodotoxin-resistant sodium channel Na(v)1.8 (PN3) as well as the Na(v)1.2 and Na(v)1.5 subtypes. Benchmark compounds from this series possessed enhanced potency, oral bioavailability, and robust efficacy in a rodent model of neuropathic pain, together with improved CNS and cardiovascular safety profiles compared to the clinically used sodium channel blockers mexiletine and lamotrigine.
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Analgésicos no Narcóticos/química , Analgésicos no Narcóticos/farmacología , Furanos/química , Furanos/farmacología , Neuralgia/tratamiento farmacológico , Piperazinas/química , Piperazinas/farmacología , Bloqueadores de los Canales de Sodio/química , Bloqueadores de los Canales de Sodio/farmacología , Canales de Sodio/efectos de los fármacos , Analgésicos no Narcóticos/síntesis química , Animales , Canales de Potasio Éter-A-Go-Go/antagonistas & inhibidores , Furanos/síntesis química , Humanos , Masculino , Ratones , Piperazinas/síntesis química , Ratas , Ratas Sprague-Dawley , Bloqueadores de los Canales de Sodio/síntesis química , Relación Estructura-ActividadRESUMEN
A series of potent indol-3-yl-tetramethylcyclopropyl ketones have been prepared as CB 2 cannabinoid receptor ligands. Two unsubstituted indoles ( 5, 32) were the starting points for an investigation of the effect of indole ring substitutions on CB 2 and CB 1 binding affinities and activity in a CB 2 in vitro functional assay. Indole ring substitutions had varying effects on CB 2 and CB 1 binding, but were generally detrimental to agonist activity. Substitution on the indole ring did lead to improved CB 2/CB 1 binding selectivity in some cases (i.e., 7- 9, 15- 20). All indoles with the morpholino-ethyl side chain ( 32- 43) exhibited weaker binding affinity and less agonist activity relative to that of their tetrahydropyranyl-methyl analogs ( 5- 31). Several agonists were active in the complete Freund's adjuvant model of chronic inflammatory thermal hyperalgesia ( 32, 15).
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Antiinflamatorios no Esteroideos/farmacología , Indoles/farmacología , Cetonas/farmacología , Receptor Cannabinoide CB2/efectos de los fármacos , Animales , Antiinflamatorios no Esteroideos/síntesis química , Antiinflamatorios no Esteroideos/química , Unión Competitiva , Línea Celular , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos , Humanos , Hiperalgesia/inducido químicamente , Hiperalgesia/tratamiento farmacológico , Indoles/síntesis química , Indoles/química , Cetonas/síntesis química , Cetonas/química , Ligandos , Conformación Molecular , Ratas , Receptor Cannabinoide CB1/efectos de los fármacos , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
Activation of metabotropic glutamate (mGlu) receptors has previously been shown to play a role in inflammatory or neuropathic pain states. However, the role of mGlu type 1 receptors in post-operative pain remains to be investigated. In the present study, effects of potent and selective mGlu1 receptor antagonists A-841720, A-794282, A-794278, and A-850002 were evaluated in a skin incision-induced post-operative pain model in rats. Post-operative pain was examined 2 h following surgery using weight-bearing difference between injured and uninjured paws as a measure of spontaneous pain. In this model, A-841720, A-794282, A-794278, and A-850002 induced significant attenuation of spontaneous post-operative pain behavior, with ED50s of 10, 50, 50, and 65 micromol/kg i.p., respectively. Depending on the compound, significant motor side effects were also observed at 3 to 10 fold higher doses. These results support the notion that mGlu1 receptor activation plays a significant role in nociceptive transmission in post-operative pain, though motor impairment may be a limiting factor in developing mGlu1 receptor antagonists as novel analgesics.
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Analgésicos no Narcóticos/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Dolor Postoperatorio/prevención & control , Receptores de Glutamato Metabotrópico/antagonistas & inhibidores , Analgésicos no Narcóticos/química , Animales , Calcio/metabolismo , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Cerebelo/citología , Cerebelo/efectos de los fármacos , Cerebelo/metabolismo , Dimetilaminas/química , Dimetilaminas/farmacología , Relación Dosis-Respuesta a Droga , Antagonistas de Aminoácidos Excitadores/química , Conducta Exploratoria/efectos de los fármacos , Fluorometría/métodos , Glicina/análogos & derivados , Glicina/farmacología , Compuestos Heterocíclicos con 3 Anillos/química , Compuestos Heterocíclicos con 3 Anillos/farmacología , Miembro Posterior/cirugía , Masculino , Estructura Molecular , Morfina/farmacología , Dolor Postoperatorio/etiología , Piridinas/química , Piridinas/farmacología , Pirimidinonas/química , Pirimidinonas/farmacología , Ensayo de Unión Radioligante , Ratas , Ratas Sprague-Dawley , Resorcinoles/farmacología , Prueba de Desempeño de Rotación con Aceleración Constante/métodos , Tiofenos/química , Tiofenos/farmacología , TritioRESUMEN
ATP-sensitive P2X(7) receptors are localized on cells of immunological origin including glial cells in the central nervous system. Activation of P2X(7) receptors leads to rapid changes in intracellular calcium concentrations, release of the proinflammatory cytokine interleukin-1beta (IL-1beta), and following prolonged agonist exposure, cytolytic plasma membrane pore formation. P2X(7) knockout mice show reduced inflammation as well as decreased nociceptive sensitivity following peripheral nerve injury. A-740003 (N-(1-{[(cyanoimino)(5-quinolinylamino) methyl] amino}-2,2-dimethylpropyl)-2-(3,4-dimethoxyphenyl)acetamide) is a novel competitive antagonist of P2X(7) receptors (IC(50) values = 40 nM for human and 18 nM for rat) as measured by agonist-stimulated changes in intracellular calcium concentrations. A-740003 showed weak or no activity (IC(50) > 10 muM) at other P2 receptors and an array of other neurotransmitter and peptide receptors, ion channels, reuptake sites, and enzymes. A-740003 potently blocked agonist-evoked IL-1beta release (IC(50) = 156 nM) and pore formation (IC(50) = 92 nM) in differentiated human THP-1 cells. Systemic administration of A-740003 produced dose-dependent antinociception in a spinal nerve ligation model (ED(50) = 19 mg/kg i.p.) in the rat. A-740003 also attenuated tactile allodynia in two other models of neuropathic pain, chronic constriction injury of the sciatic nerve and vincristine-induced neuropathy. In addition, A-740003 effectively reduced thermal hyperalgesia observed following intraplantar administration of carrageenan or complete Freund's adjuvant (ED(50) = 38-54 mg/kg i.p.). A-740003 was ineffective in attenuating acute thermal nociception in normal rats and did not alter motor performance at analgesic doses. These data demonstrate that selective blockade of P2X(7) receptors in vivo produces significant antinociception in animal models of neuropathic and inflammatory pain.
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Acetamidas/farmacología , Analgésicos , Dolor/tratamiento farmacológico , Dolor/etiología , Enfermedades del Sistema Nervioso Periférico/complicaciones , Antagonistas del Receptor Purinérgico P2 , Quinolinas/farmacología , Animales , Antineoplásicos Fitogénicos/toxicidad , Calcio/metabolismo , Línea Celular , Colorantes , Relación Dosis-Respuesta a Droga , Edema/inducido químicamente , Edema/tratamiento farmacológico , Adyuvante de Freund/farmacología , Humanos , Hiperalgesia/inducido químicamente , Hiperalgesia/tratamiento farmacológico , Inflamación/inducido químicamente , Inflamación/complicaciones , Inflamación/tratamiento farmacológico , Interleucina-1beta/metabolismo , Masculino , Actividad Motora/efectos de los fármacos , Nociceptores/efectos de los fármacos , Equilibrio Postural/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Receptores Purinérgicos P2X7 , Neuropatía Ciática/prevención & control , Nervios Espinales/lesiones , Vincristina/toxicidadRESUMEN
Gabapentin and pregabalin have been demonstrated, both in animal pain models and clinically, to be effective analgesics particularly for the treatment of neuropathic pain. The precise mechanism of action for these two drugs is unknown, but they are generally believed to function via initially binding to the alpha2delta subunit of voltage-gated Ca2+ channels. In this study, we used a pharmacological approach to test the hypothesis whether high affinity interactions with the alpha2delta subunit alone could lead to attenuation of neuropathic pain in rats. The anti-allodynic effects of gabapentin and pregabalin, along with three other compounds--(L)-phenylglycine, m-chlorophenylglycine and 3-exo-aminobicyclo[2.2.1]heptane-2-exo-carboxylic acid (ABHCA)--discovered to be potent alpha2delta ligands, were tested in the rat spinal nerve ligation model of neuropathic pain. Gabapentin (Ki = 120 nM), pregabalin (180 nM) and (L)-phenylglycine (180 nM) were shown to be anti-allodynic, with respective ED50 values of 230, 90 and 80 micromol/kg (p.o.). (L)-Phenylglycine was as potent as pregabalin and equi-efficacious in reversing mechanical allodynia. In contrast, two ligands with comparable or superior alpha2delta binding affinities, m-chlorophenylglycine (Ki = 54 nM) and ABHCA (150 nM), exhibited no anti-allodynic effects at doses of 30-300 micromol/kg (p.o.), although these compounds achieved substantial brain levels. The data demonstrate that, at least in the rat spinal nerve ligation model of neuropathic pain, (L)-phenylglycine has an anti-allodynic effect, but two equally potent alpha2delta subunit ligands do not. These results suggest that additional mechanisms, besides alpha2delta interactions, may contribute to the effects of compounds like gabapentin, pregabalin and (L)-phenylglycine in neuropathic pain.
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Encéfalo/metabolismo , Canales de Calcio/efectos de los fármacos , Canales de Calcio/metabolismo , Glicina/análogos & derivados , Neuralgia/metabolismo , Neuralgia/prevención & control , Analgésicos/administración & dosificación , Animales , Encéfalo/efectos de los fármacos , Bloqueadores de los Canales de Calcio/administración & dosificación , Canales de Calcio Tipo L , Relación Dosis-Respuesta a Droga , Glicina/administración & dosificación , Hiperestesia/metabolismo , Hiperestesia/prevención & control , Masculino , Ratas , Ratas Sprague-Dawley , Resultado del TratamientoRESUMEN
Previous studies have demonstrated that Fos-like immunoreactivity is increased in spinal dorsal horn neurons in several pain models, and have suggested that Fos-like immunoreactivity could be used as a marker of neurons activated by painful stimulation. In the present study, we evaluated nociceptive behaviors and spinal Fos-like immunoreactivity in a rat skin incision model of post-operative pain. In this model, evoked and non-evoked pain behaviors were observed at least for 2 days after paw surgery, an increased number of Fos-like immunoreactive neurons was observed in the spinal dorsal horn at lumbar levels 4-5 two-hour post-surgery. The number of Fos-like immunoreactive neurons was significantly greater in animals with skin-muscle incision compared to animals with skin-alone incision. Interestingly, spinal Fos-like immunoreactivity was quickly normalized in rats with paw surgery at later time points (8 and 24 h post-surgery), whereas nociceptive behaviors were still observed. Furthermore, at 24 h post-surgery, spinal Fos-like immunoreactivity induced by thermal stimulation (42, 44, 46, 48, 52 degrees C for 15 s) was not significantly different between sham animals and animals with surgery. In both groups, an increase in spinal Fos-like immunoreactive neurons was observed with increasing temperatures, with similar laminar distribution. Finally, systemic morphine reduced post-operative pain and Fos-like immunoreactivity in a naloxone reversible manner, with greater potency and efficacy on behavioral endpoints than on Fos-like immunoreactivity. These results demonstrate a different profile of nociceptive behaviors and spinal Fos-like immunoreactivity in the rat skin incision model, suggesting a limited potential of spinal Fos-like immunoreactivity to study post-surgical pain and its pharmacology.