RESUMO
The alpha4beta1 integrin (very late antigen-4, VLA-4) plays an important role in the migration of lymphocytes, monocytes, and eosinophils, but not neutrophils, to sites of inflammation. Pharmacological antagonism of VLA-4 is an attractive prospect for the treatment of predominantly eosinophil mediated diseases such as asthma and allergic rhinitis. We report here on a potent and selective, small molecule VLA-4 inhibitor, (2S)-3-(2', 5'-dichlorobiphenyl-4-yl)-2-({[1-(2-methoxybenzoyl)piperidin-3-yl]carbonyl}amino) propanoic acid, compound 1, and characterize the antagonist activities of this molecule in various cell-based assays and in an animal model of eosinophil migration. Compound 1 inhibited VLA-4/ vascular cell adhesion molecule-1(VCAM-1) interactions with in vitro potencies (IC50 value of 210 nM) in VLA-4-expressing Ramos cells, although the compound did not inhibit cell adhesion to fibronectin via alpha5beta1 integrin (very late antigen-5, VLA-5). Blockade of phorbol-12-myristate-13-acetate (PMA)- or Mn2+-stimulated VLA-4 interactions with compound 1 was observed in human T lymphocytes (IC50 value of 230 nM), human eosinophils (IC50 value of 4.0 microM) and mouse eosinophils (IC50 value of 1.6 microM). Furthermore, compound 1 administered by intraperitoneal injection inhibited eosinophil infiltration in a dose-dependent manner by up to 80% in an air pouch model. These data support the use of small molecule VLA-4 antagonists in the treatment of relevant diseases, such as asthma, atopic dermatitis, or allergic rhinitis.
Assuntos
Antialérgicos/farmacologia , Anti-Inflamatórios/farmacologia , Quimiotaxia de Leucócito/efeitos dos fármacos , Eosinofilia/prevenção & controle , Eosinófilos/efeitos dos fármacos , Integrina alfa4beta1/antagonistas & inibidores , Bifenilos Policlorados/farmacologia , Dermatopatias/prevenção & controle , Animais , Antialérgicos/farmacocinética , Antialérgicos/uso terapêutico , Anti-Inflamatórios/farmacocinética , Anti-Inflamatórios/uso terapêutico , Linfócitos B/efeitos dos fármacos , Linfócitos B/metabolismo , Adesão Celular/efeitos dos fármacos , Quimiocina CCL11 , Quimiocinas CC , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Eosinofilia/induzido quimicamente , Eosinofilia/metabolismo , Eosinofilia/fisiopatologia , Eosinófilos/metabolismo , Feminino , Fibronectinas/metabolismo , Humanos , Integrina alfa4beta1/metabolismo , Integrina alfa5beta1/metabolismo , Interleucina-5/biossíntese , Interleucina-5/genética , Células Jurkat , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Bifenilos Policlorados/farmacocinética , Bifenilos Policlorados/uso terapêutico , Dermatopatias/induzido quimicamente , Dermatopatias/metabolismo , Dermatopatias/fisiopatologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Acetato de Tetradecanoilforbol , Fatores de Tempo , Molécula 1 de Adesão de Célula Vascular/metabolismoRESUMO
To investigate the circadian rhythm of inducible cytokine release and a potential pacemaker role of endogenous cortisol, cortisol levels as well as cytokine release from ex vivo LPS-stimulated blood were assessed at 4-h intervals over 24 h in 11 volunteers. We found a significant diurnal variation for IFN-gamma and IL-8, and a tendency for TNF, all inversely correlated to the serum cortisol levels, but no evidence for such a rhythm for IL-1beta and IL-6. In vitro IC(50) values for cytokine inhibition by hydrocortisone (HC) corresponded to the observed rank order for circadian rhythmicity. mRNA analyses revealed that this was due to a reduction of gene transcription. These effects of HC were significantly reversed by the glucocorticoid receptor antagonist RU486. Supplementation of HC in vivo to maintain morning cortisol levels throughout the day blunted the circadian rhythm of ex vivo LPS-induced cytokines. Surprisingly, no significant diurnal variation for any investigated cytokine was found in the same volunteer group upon stimulation with lipoteichoic acid (LTA), the gram-positive counterpart to LPS. Furthermore, 10-50-fold higher HC concentrations as compared to LPS were required to block LTA-induced cytokine release. LTA, in contrast to LPS, failed to activate Jun kinase, a reported target for HC action.
Assuntos
Células Sanguíneas/imunologia , Ritmo Circadiano/imunologia , Hidrocortisona/imunologia , Lipopolissacarídeos/farmacologia , Ácidos Teicoicos/farmacologia , Abortivos Esteroides/farmacologia , Adulto , Células Sanguíneas/citologia , Células Sanguíneas/metabolismo , Células Cultivadas , Ritmo Circadiano/efeitos dos fármacos , Citocinas/biossíntese , Citocinas/imunologia , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/imunologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/imunologia , Humanos , Hidrocortisona/sangue , Lipopolissacarídeos/imunologia , Masculino , Mifepristona/farmacologia , Proteína Oncogênica p65(gag-jun)/imunologia , Proteína Oncogênica p65(gag-jun)/metabolismo , Receptores de Glucocorticoides/antagonistas & inibidores , Receptores de Glucocorticoides/imunologia , Receptores de Glucocorticoides/metabolismo , Ácidos Teicoicos/imunologia , Transcrição Gênica/efeitos dos fármacos , Transcrição Gênica/imunologiaRESUMO
Terephthalic acid based derivatives containing beta- and gamma-amino acid residues were prepared as antagonists of the leukocyte cell adhesion process that is mediated through the interaction of the very late antigen 4 (VLA-4) and the vascular cell adhesion molecule 1 (VCAM-1). The compounds 2, 10-12, 14, and 16-17 inhibited the adhesion in a cell based assay in the low and sub micromolar range.