RESUMO
TNF-alpha converting enzyme (TACE) is a validated therapeutic target for the development of oral tumor necrosis factor-alpha (TNF-alpha) inhibitors. Here we report the pre-clinical results and characterization of a selective and potent TACE inhibitor, (2R, 3S)-2-([[4-(2-butynyloxy)phenyl]sulfonyl]amino)-N,3-dihydroxybutanamide (TMI-2), in various in vitro and in vivo assays. TMI-2 is a potent TACE inhibitor in an enzymatic FRET assay (IC50=2 nM). It is more than 250-fold selective over MMP-1, -7, -9, -14, and ADAM-10 in vitro. In cell-based assays and human whole blood, TMI-2 inhibits lipopolysaccharide (LPS)-induced TNF secretion with IC50s<1 uM. Importantly, TMI-2 inhibits the spontaneous release of TNF-alpha in human synovium tissue explants of rheumatoid arthritis patients with an IC50 of 0.8 microM. In vivo, TMI-2 potently inhibits LPS-induced TNF-alpha production in mice (ED50=3 mg/kg). In the adjuvant-induced arthritis (AIA) model in rats, treatment with TMI-2 at 30 mg/kg and 100 mg/kg p.o. b.i.d. was highly effective in reducing joint arthritis scores. In a semi-therapeutic collagen-induced arthritis (CIA) model in mice, TMI-2 is highly effective in reducing disease severity scores after oral treatment at 100 mg/kg twice per day. In summary, TMI-2 is a potent and selective TACE inhibitor that inhibits TNF-alpha production and reduces the arthritis scores in pre-clinical models. TMI-2 represents a novel class of TACE inhibitors that may be effective and beneficial in the treatment of rheumatoid arthritis as well as other TNF-mediated inflammatory autoimmune diseases.
Assuntos
Metaloendopeptidases/antagonistas & inibidores , Inibidores de Proteases/farmacologia , Sulfonamidas/farmacologia , Proteínas ADAM , Proteína ADAM17 , Animais , Artrite Experimental/tratamento farmacológico , Artrite Experimental/patologia , Artrite Reumatoide/tratamento farmacológico , Disponibilidade Biológica , Linhagem Celular , Colágeno , Humanos , Técnicas In Vitro , Lipopolissacarídeos , Metaloproteases/antagonistas & inibidores , Metaloproteases/biossíntese , Camundongos , Camundongos Endogâmicos DBA , Ensaios de Proteção de Nucleases , Inibidores de Proteases/farmacocinética , RNA Mensageiro/biossíntese , Ratos , Ratos Endogâmicos Lew , Sulfonamidas/farmacocinética , Membrana Sinovial/efeitos dos fármacos , Sinovite/patologia , Fator de Necrose Tumoral alfa/biossínteseRESUMO
A potent, selective series of MMP-13 inhibitors has been derived from a weak (3.2 microM) inhibitor that did not bear a zinc chelator. Structure-based drug design strategies were employed to append a Zn-chelating group to one end of the molecule and functionality to enhance selectivity to the other. A compound from this series demonstrated rat oral bioavailability and efficacy in a bovine articular cartilage explant model.
Assuntos
Inibidores de Metaloproteinases de Matriz , Inibidores de Proteases/química , Inibidores de Proteases/farmacologia , Administração Oral , Aminoácidos/química , Animais , Benzofuranos/química , Cartilagem/efeitos dos fármacos , Cartilagem/enzimologia , Bovinos , Quelantes/química , Colagenases/química , Colagenases/metabolismo , Cristalografia por Raios X , Desenho de Fármacos , Técnicas In Vitro , Concentração Inibidora 50 , Metaloproteinase 13 da Matriz , Modelos Moleculares , Inibidores de Proteases/administração & dosagem , Inibidores de Proteases/farmacocinética , Estrutura Terciária de Proteína , Ratos , Sensibilidade e Especificidade , Relação Estrutura-Atividade , Especificidade por Substrato , Zinco/metabolismoRESUMO
Tumor necrosis factor (TNF)-alpha is a well validated therapeutic target for the treatment of rheumatoid arthritis. TNF-alpha is initially synthesized as a 26-kDa membrane-bound form (pro-TNF) that is cleaved by a Zn-metalloprotease named TNF-alpha-converting enzyme (TACE) to generate the 17-kDa, soluble, mature TNF-alpha. TACE inhibitors that prevent the secretion of soluble TNF-alpha may be effective in treating rheumatoid arthritis (RA) patients. Using a structure-based design approach, we have identified a novel dual TACE/matrix metalloprotease (MMP) inhibitor 4-[[4-(2-butynyloxy)phenyl]sulfonyl]-N-hydroxy-2,2-dimethyl-(3S)thiomorpholinecarboxamide (TMI-1). This molecule inhibits TACE and several MMPs with nanomolar IC(50) values in vitro. In cell-based assays such as monocyte cell lines, human primary monocytes, and human whole blood, it inhibits lipopolysaccharide (LPS)-induced TNF-alpha secretion at submicromolar concentrations, whereas there is no effect on the TNF-alpha mRNA level as judged by RNase protection assay. The inhibition of LPS-induced TNF-alpha secretion is selective because TMI-1 has no effect on the secretion of other proinflammatory cytokines such as interleukin (IL)-1beta, IL-6, and IL-8. Importantly, TMI-1 potently inhibits TNF-alpha secretion by human synovium tissue explants of RA patients. In vivo, TMI-1 is highly effective in reducing clinical severity scores in mouse prophylactic collagen-induced arthritis (CIA) at 5, 10, and 20 mg/kg p.o. b.i.d. and therapeutic CIA model at 100 mg/kg p.o. b.i.d. In summary, TMI-1, a dual TACE/MMP inhibitor, represents a unique class of orally bioavailable small molecule TNF inhibitors that may be effective and beneficial for treating RA.