RESUMO
SAR studies on a series of thiophene amide derivatives provided CB(2) receptor agonists. The activity of the compounds was characterized by radioligand binding determination, multiple functional assays, ADME, and pharmacokinetic studies. A representative compound with selectivity for CB(2) over CB(1) effectively produced analgesia in behavioral models of neuropathic, inflammatory, and postsurgical pain. Control experiments using a CB(2) antagonist demonstrated the efficacy in the pain models resulted from CB(2) agonism.
Assuntos
Amidas/síntese química , Analgésicos/síntese química , Hiperalgesia/tratamento farmacológico , Neuralgia/tratamento farmacológico , Receptor CB2 de Canabinoide/agonistas , Tiofenos/síntese química , Amidas/farmacocinética , Amidas/farmacologia , Analgésicos/farmacocinética , Analgésicos/farmacologia , Animais , Disponibilidade Biológica , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Humanos , Hiperalgesia/metabolismo , Neuralgia/metabolismo , Ensaio Radioligante , Ratos , Ratos Sprague-Dawley , Receptor CB1 de Canabinoide/metabolismo , Receptor CB2 de Canabinoide/metabolismo , Relação Estrutura-Atividade , Tiofenos/farmacocinética , Tiofenos/farmacologiaRESUMO
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.
Assuntos
Indóis/farmacologia , Cetonas/farmacologia , Receptor CB2 de Canabinoide/agonistas , Desenho de Fármacos , Humanos , Indóis/síntese química , Indóis/química , Cetonas/síntese química , Cetonas/química , Ligantes , Estrutura Molecular , Receptor CB1 de Canabinoide/agonistas , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
Wnt ligands conduct their functions in canonical Wnt signaling by binding to two receptors, the single transmembrane low density lipoprotein receptor-related proteins 5 and 6 (LRP5/6) and seven transmembrane (7TM) Frizzled receptors. Subsequently, phosphorylation of serine/threonine residues within five repeating signature PPPSP motifs on LRP6 is responsible for LRP6 activation. GSK3beta, a cytosolic kinase for phosphorylation of a downstream effector beta-catenin, was proposed to participate in such LRP6 phosphorylation. Here, we report a new class of membrane-associated kinases for LRP6 phosphorylation. We found that G protein-coupled receptor kinases 5 and 6 (GRK5/6), traditionally known to phosphorylate and desensitize 7TM G protein-coupled receptors, directly phosphorylate the PPPSP motifs on single transmembrane LRP6 and regulate Wnt/LRP6 signaling. GRK5/6-induced LRP6 activation is inhibited by the LRP6 antagonist Dickkopf. Depletion of GRK5 markedly reduces Wnt3A-stimulated LRP6 phosphorylation in cells. In zebrafish, functional knock-down of GRK5 results in reduced Wnt signaling, analogous to LRP6 knock-down, as assessed by decreased abundance of beta-catenin and lowered expression of the Wnt target genes cdx4, vent, and axin2. Expression of GRK5 rescues the diminished beta-catenin and axin2 response caused by GRK5 depletion. Thus, our findings identify GRK5/6 as novel kinases for the single transmembrane receptor LRP6 during Wnt signaling.
Assuntos
Quinase 5 de Receptor Acoplado a Proteína G/metabolismo , Quinases de Receptores Acoplados a Proteína G/metabolismo , Receptores de LDL/metabolismo , Proteínas Wnt/metabolismo , Sequência de Aminoácidos , Animais , Proteína Axina , Células Cultivadas , Fibroblastos/citologia , Fibroblastos/fisiologia , Receptores Frizzled/genética , Receptores Frizzled/metabolismo , Quinase 5 de Receptor Acoplado a Proteína G/genética , Quinases de Receptores Acoplados a Proteína G/genética , Células HeLa , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo , Fosforilação , Receptores de LDL/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transdução de Sinais/fisiologia , Proteínas Wnt/genética , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo , beta Catenina/genética , beta Catenina/metabolismoRESUMO
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).
