RESUMO
OBJECTIVE: Autotaxin is a secreted lysophospholipase that mediates the conversion of lysophosphatidyl choline (LPC) to lysophosphatidic acid (LPA), a bioactive lipid mediator. Autotaxin levels in plasma and synovial fluid correlate with disease severity in patients with knee osteoarthritis (OA). The goal of this study was to develop and characterize a novel small molecule inhibitor of autotaxin to inhibit LPA production in vivo and determine its efficacy in animal models of musculoskeletal pain. DESIGN: Compound libraries were screened using an LPC coupled enzyme assay that measures the amount of choline released from LPC by the action of autotaxin. Hits from this assay were tested in a plasma assay to assess inhibition of endogenous plasma autotaxin and subsequently tested for their ability to lower plasma LPA levels upon oral dosing of rats. The best compounds were then tested in animal models of musculoskeletal pain. RESULTS: Compound screening led to the identification of compounds with nanomolar potency for inhibition of autotaxin activity. Studies in rats demonstrated a good correlation between compound exposure levels and a decrease in LPA levels in plasma. The leading molecule (compound-1) resulted in a dose dependent decrease in joint pain in the mono-sodium iodoacetate (MIA) and meniscal tear models and a decrease in bone fracture pain in the osteotomy model in rats. CONCLUSION: We have identified and characterized a novel small molecule inhibitor of autotaxin and demonstrated its efficacy in animal models of musculoskeletal pain. The inhibitor has the potential to serve as an analgesic for human OA and bone fracture.
Assuntos
Artralgia/metabolismo , Artrite Experimental/metabolismo , Osteoartrite do Joelho/metabolismo , Inibidores de Fosfodiesterase/farmacologia , Diester Fosfórico Hidrolases/efeitos dos fármacos , Animais , Artralgia/etiologia , Artralgia/fisiopatologia , Artrite Experimental/induzido quimicamente , Artrite Experimental/complicações , Artrite Experimental/fisiopatologia , Cães , Humanos , Ácido Iodoacético/toxicidade , Lisofosfatidilcolinas/metabolismo , Lisofosfolipídeos/metabolismo , Masculino , Meniscos Tibiais/cirurgia , Osteoartrite do Joelho/induzido quimicamente , Osteoartrite do Joelho/complicações , Osteoartrite do Joelho/fisiopatologia , Osteotomia , Diester Fosfórico Hidrolases/metabolismo , Ratos , Ratos Endogâmicos Lew , Lesões do Menisco TibialRESUMO
OBJECTIVE: Emerging evidence suggests that osteoarthritis (OA) has a neuropathic component; however, the identity of the molecules responsible for this peripheral neuropathy is unknown. The aim of this study was to determine the contribution of the bioactive lipid lysophosphatidic acid (LPA) to joint neuropathy and pain. DESIGN: Male Lewis rats received an intra-articular injection of 50 µg of LPA into the knee and allowed to recover for up to 21 days. Saphenous nerve myelination was assessed by g-ratio calculation from electron micrographs and afferent nerve damage visualised by activation transcription factor-3 (ATF-3) expression. Nerve conduction velocity was measured electrophysiologically and joint pain was determined by hindlimb incapacitance. The effect of the LPA antagonist Ki-16425 was also evaluated. Experiments were repeated in the sodium monoiodoacetate (MIA) model of OA. RESULTS: LPA caused joint nerve demyelination which resulted in a drop in nerve conduction velocity. Sensory neurones were ATF-3 positive and animals exhibited joint pain and knee joint damage. MIA-treated rats also showed signs of demyelination and joint neuropathy with concomitant pain. Nerve damage and pain could be ameliorated by Ki-16425 pre-treatment. CONCLUSION: Intra-articular injection of LPA caused knee joint neuropathy, joint damage and pain. Pharmacological blockade of LPA receptors inhibited joint nerve damage and hindlimb incapacitance. Thus, LPA is a candidate molecule for the development of OA nerve damage and the origin of joint neuropathic pain.
Assuntos
Fator 3 Ativador da Transcrição/efeitos dos fármacos , Artrite Experimental/fisiopatologia , Lisofosfolipídeos/farmacologia , Condução Nervosa/efeitos dos fármacos , Osteoartrite/fisiopatologia , Nervos Periféricos/efeitos dos fármacos , Fator 3 Ativador da Transcrição/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Artralgia , Artrite Experimental/induzido quimicamente , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Comportamento Animal , Estudos de Casos e Controles , Cromatografia Líquida , Inibidores Enzimáticos/toxicidade , Feminino , Humanos , Injeções Intra-Articulares , Ácido Iodoacético/toxicidade , Isoxazóis/farmacologia , Lisofosfolipídeos/antagonistas & inibidores , Lisofosfolipídeos/metabolismo , Masculino , Pessoa de Meia-Idade , Neuralgia , Osteoartrite/induzido quimicamente , Osteoartrite/metabolismo , Osteoartrite/patologia , Osteoartrite do Joelho/metabolismo , Nervos Periféricos/metabolismo , Nervos Periféricos/ultraestrutura , Propionatos/farmacologia , Ratos Endogâmicos Lew , Líquido Sinovial/químicaRESUMO
OBJECTIVE: Investigate a role for calcitonin gene-related peptide (CGRP) in osteoarthritis (OA)-related pain. DESIGN: Neutralizing antibodies to CGRP were generated de novo. One of these antibodies, LY2951742, was characterized in vitro and tested in pre-clinical in vivo models of OA pain. RESULTS: LY2951742 exhibited high affinity to both human and rat CGRP (KD of 31 and 246 pM, respectively). The antibody neutralized CGRP-mediated induction of cAMP in SK-N-MC cells in vitro and capsaicin-induced dermal blood flow in the rat. Neutralization of CGRP significantly reduced pain behavior as measured by weight bearing differential in the rat monoiodoacetate model of OA pain in a dose-dependent manner. Moreover, pain reduction with neutralization of CGRP occurred independently of prostaglandins, since LY2951742 and NSAIDs worked additively in the NSAID-responsive version of the model and CGRP neutralization remained effective in the NSAID non-responsive version of the model. Neutralization of CGRP also provided dose-dependent and prolonged (>60 days) pain reduction in the rat meniscal tear model of OA after only a single injection of LY2951742. CONCLUSIONS: LY2951742 is a high affinity, neutralizing antibody to CGRP. Neutralization of CGRP is efficacious in several OA pain models and works independently of NSAID mechanisms of action. LY2951742 holds promise for the treatment of pain in OA patients.
Assuntos
Anticorpos Neutralizantes/farmacologia , Peptídeo Relacionado com Gene de Calcitonina/efeitos dos fármacos , Osteoartrite/tratamento farmacológico , Dor/prevenção & controle , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Peptídeos Catiônicos Antimicrobianos , Catelicidinas/metabolismo , Modelos Animais de Doenças , Humanos , Masculino , Ratos , Ratos Endogâmicos Lew , Fluxo Sanguíneo Regional , Pele/irrigação sanguíneaRESUMO
OBJECTIVE: The purpose of this study was to use microarray technology to: (1) understand the early molecular events underlying the damage of articular cartilage initiated by this surgical procedure, and (2) determine whether these changes mimic those that are occurring in human osteoarthritic (OA) cartilage. DESIGN: Cartilage was harvested from both medial and lateral sides of the tibial plateaus and femoral condyles of both meniscal tear (MT) and sham surgery groups on days 3, 7 and 21 post-surgery. mRNA prepared from these rat cartilage samples was used for microarray analysis. RESULTS: Statistical analysis identified 475 genes that were differentially expressed between the sham and MT groups, at one or more of the time points that were analyzed. By integrating these genes with OA-related genes reported previously in a rat OA model and in human OA array studies, we identified 20 commonly changed genes. Six out of these 20 genes (Col5A1, Col6A2, INHBA, LTBP2, NBL1 and SERPINA1) were differentially expressed in two animal models and in human OA. Pathway analysis identified some key features of OA pathology, namely cartilage extracellular matrix remodeling, angiogenesis, and chondrocyte cell death that were recapitulated in the animal models. The rat models suggested increased inflammation and cholesterol metabolic pathways may play important role in early cartilage degeneration. CONCLUSION: We identified a large number of differentially expressed genes in the articular cartilage of the MT model. While there was lack of overall identity in cartilage gene expression between the rat models and human OA, several key biological processes were recapitulated in the rat MT OA model.