RESUMO
Inhibitors of phosphodiesterases (PDEs) have been widely studied as therapeutics for the treatment of human diseases, but improvement of inhibitor selectivity is still desirable for the enhancement of inhibitor potency. Here, we report identification of a water-containing subpocket as a PDE4-specific pocket for inhibitor binding. We designed against the pocket and synthesized two enantiomers of PDE4 inhibitor Zl-n-91. The ( S)-Zl-n-91 enantiomer showed IC50 values of 12 and 20 nM for the catalytic domains of PDE4D2 and PDE4B2B, respectively, selectivity several thousand-fold greater than those of other PDE families, and potent neuroprotection activities. Crystal structures of the PDE4D2 catalytic domain in complex with each Zl-n-91 enantiomer revealed that ( S)-Zl-n-91 but not ( R)-Zl-n-91 formed a hydrogen bond with the bound water in the pocket, thus explaining its higher affinity. The structural superposition between the PDE families revealed that this water-containing subpocket is unique to PDE4 and thus valuable for the design of PDE4 selective inhibitors.
Assuntos
Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Desenho de Fármacos , Furanos/química , Furanos/farmacologia , Éteres Fenílicos/química , Éteres Fenílicos/farmacologia , Inibidores da Fosfodiesterase 4/química , Inibidores da Fosfodiesterase 4/farmacologia , Animais , Sítios de Ligação/efeitos dos fármacos , Domínio Catalítico/efeitos dos fármacos , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/química , Furanos/farmacocinética , Humanos , Ligação de Hidrogênio/efeitos dos fármacos , Masculino , Memória/efeitos dos fármacos , Camundongos Endogâmicos ICR , Simulação de Acoplamento Molecular , Éteres Fenílicos/farmacocinética , Inibidores da Fosfodiesterase 4/farmacocinética , Rolipram/análogos & derivados , Rolipram/farmacocinética , Rolipram/farmacologia , Estereoisomerismo , Água/químicaRESUMO
Brain imaging is considered one of the most fruitful applications of radioisotope scanning. Rolipram, a selective phospodiesterase-4 inhibitor, has been labeled using [125 I] with chloramine-T (Ch-T) as an oxidizing agent. Factors, such as the amount of substrate, pH, the amount of oxidizing agent, temperature, and the reaction time, have been systematically studied to optimize the iodination process. In addition, bio-distribution studies have indicated that the brain uptake of [125 I]iodorolipram is 7.6 ± 0.33 injected dose/g organ at 10 minutes post-injection, which cleared from the brain with time until it reaches 1.30 ± 0.17% at 1 hour post-injection. Therefore, iodorolipram could be considered as a potential, new selective radiotracer for brain imaging.
Assuntos
Encéfalo/diagnóstico por imagem , Radioisótopos do Iodo/química , Inibidores de Fosfodiesterase/síntese química , Compostos Radiofarmacêuticos/síntese química , Rolipram/análogos & derivados , Animais , Raios gama , Camundongos , Inibidores de Fosfodiesterase/farmacocinética , Cintilografia/métodos , Compostos Radiofarmacêuticos/farmacocinética , Distribuição TecidualRESUMO
Phosphodiesterase (PDE) 4 is the most prevalent PDE in the central nervous system (CNS) and catalyzes hydrolysis of intracellular cAMP, a secondary messenger. By therapeutic inhibition of PDE4, intracellular cAMP levels can be stabilized, and the symptoms of psychiatric and neurodegenerative disorders including depression, memory loss and Parkinson's disease can be ameliorated. Radiotracers targeting PDE4 can be used to study PDE4 density and function, and evaluate new PDE4 therapeutics, in vivo in a non-invasive way, as has been shown using the carbon-11 labeled PDE4 inhibitor R-(-)-rolipram. Herein we describe a small series of rolipram analogs that contain fluoro- or iodo-substituents that could be used as fluorine-18 PET or iodine-123 SPECT PDE4 radiotracers. This series was evaluated with an in vitro binding assay and a 4-(fluoromethyl) derivative of rolipram, MNI-617, was identified, with a five-fold increase in affinity for PDE4 (Kd = 0.26 nM) over R-(-)-rolipram (Kd = 1.6 nM). A deutero-analogue d2 -[(18) F]MNI-617 was radiolabeled and produced in 23% yield with high (>5 Ci/µmol) specific activity and evaluated in non-human primate, where it rapidly entered the brain, with SUVs between 4 and 5, and with a distribution pattern consistent with that of PDE4.
Assuntos
Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Radioisótopos de Flúor , Rolipram/análogos & derivados , Rolipram/metabolismo , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Animais , Feminino , Ligantes , Macaca mulatta , Radioquímica , Rolipram/químicaRESUMO
The lack of selective inhibitors toward the long, short, or supershort phosphodiesterases (PDE4s) prevented researchers from carefully defining the connection between different enzyme isoforms, their brain localization, and their role in neurodegenerative diseases such as Alzheimer's disease (AD). In the search for new therapeutic agents for treating memory and learning disorders, we synthesized new rolipram related PDE4 inhibitors, which had some selectivity toward the long form PDE4D3. The first series was synthesized as racemate and then resolved by semipreparative HPLC on chiral supports. Herein we report the synthetic pathways to obtain compounds 1a-c, 2a-c, 3a-c, 4a-f, 5a,b, 6a,b, 7a,b, the chiral analytical study to resolve compounds 1a-c, 2a-c, 3a-c, the molecular docking study for compound 1c, and the biological results and some SAR considerations that provide some insights and hints for the structural requirements for PDE4D subtype selectivity and enzyme inhibition.
Assuntos
Benzaldeídos/síntese química , Ciclopentanos/síntese química , Inibidores da Fosfodiesterase 4 , Rolipram/análogos & derivados , Rolipram/síntese química , Benzaldeídos/química , Sítios de Ligação , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/química , Ciclopentanos/química , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/química , Modelos Moleculares , Conformação Molecular , Morfolinas/síntese química , Morfolinas/química , Proteínas Recombinantes/antagonistas & inibidores , Proteínas Recombinantes/química , Rolipram/química , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
INTRODUCTION: To complement recent studies using the high-affinity (11)C-labeled phosphodiesterase-4 (PDE4) inhibitor (R)-rolipram and the less active enantiomer (S)-[(11)C]rolipram for in vivo quantification of PDE4 levels, we evaluated the presence of radiolabeled metabolites and their potential binding to PDE4 in the rat plasma, brain, heart, pancreas, skeletal muscle and brown adipose tissue. METHODS: A reverse-phase capture and analytical HPLC column-switch method was used to detect (R)-[(11)C]rolipram, (S)-[(11)C]rolipram and their radiolabeled metabolites in rat plasma and tissue extracts. The relative proportion of PDE4-specific binding of the radiotracers and their labeled metabolites was analyzed following co-injections with a saturating dose of unlabeled (R)-rolipram at 45 min post-tracer injection in tissue extracts. RESULTS: Radiolabeled metabolites were found in the plasma (72-75% of total radioactive signal), and in the heart, skeletal muscle, pancreas and brown adipose tissue (44-52%), but not in the brain. In comparison to polar labeled metabolites, the proportion of unchanged (R)-[(11)C]rolipram was reduced in PDE4-rich organs by co-injection of unlabeled (R)-rolipram. Conversely, no changes were obtained in brown adipose tissue, or with (S)-[(11)C]rolipram, suggesting that radiolabeled metabolites of (R)-[(11)C]rolipram display no specific binding to PDE4. CONCLUSIONS: Radiolabeled hydrophilic metabolites are unlikely to compete with (R)-[(11)C]rolipram for PDE4-specific retention. However, due to the high proportion of the radioactive metabolites in the total radioactive signal, any kinetic modeling calculations in the peripheral tissues will need to take into account the presence of labeled metabolites.
Assuntos
Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/análise , Rolipram/análogos & derivados , Rolipram/farmacocinética , Tecido Adiposo Marrom/diagnóstico por imagem , Animais , Ligação Competitiva , Encéfalo/diagnóstico por imagem , Radioisótopos de Carbono/farmacocinética , Cromatografia Líquida de Alta Pressão , Coração/diagnóstico por imagem , Interações Hidrofóbicas e Hidrofílicas , Masculino , Músculo Esquelético/diagnóstico por imagem , Pâncreas/diagnóstico por imagem , Inibidores da Fosfodiesterase 4 , Inibidores de Fosfodiesterase/farmacocinética , Plasma/diagnóstico por imagem , Cintilografia , Compostos Radiofarmacêuticos/farmacocinética , Ratos , Ratos Sprague-Dawley , Estereoisomerismo , Distribuição Tecidual , Contagem Corporal TotalRESUMO
The discovery, synthesis and biological activity of a series of triarylethane phosphodiesterase 4 inhibitors is described. Structure-activity relationship studies are presented for CDP840 (29), a potent, chiral, selective inhibitor of PDE 4 (IC(50) 4nM). CDP840 is non-emetic in the ferret at 30mgkg(-1) (po), active in models of inflammation and reverses ozone-induced bronchial hyperreactivity in the guinea pig.
Assuntos
3',5'-AMP Cíclico Fosfodiesterases/antagonistas & inibidores , Anti-Inflamatórios não Esteroides/síntese química , Anti-Inflamatórios não Esteroides/farmacologia , Inibidores de Fosfodiesterase/síntese química , Inibidores de Fosfodiesterase/farmacologia , Piridinas/síntese química , Piridinas/farmacologia , Administração Oral , Animais , Anti-Inflamatórios não Esteroides/sangue , Asma/tratamento farmacológico , Broncoconstrição/efeitos dos fármacos , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4 , Relação Dose-Resposta a Droga , Furões , Cobaias , Humanos , Concentração Inibidora 50 , Inibidores de Fosfodiesterase/sangue , Piridinas/sangue , Ratos , Rolipram/análogos & derivados , Saccharomyces cerevisiae/enzimologia , Relação Estrutura-Atividade , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/metabolismoRESUMO
This communication describes the synthesis and in vitro evaluation of a novel and potent series of phthalazine phosphodiesterase type (IV) (PDE4) inhibitors. The interaction with two distinct polar binding sites allowed us to eliminate the cyclopentyloxy substitution from rolipram-like analogues.
Assuntos
3',5'-AMP Cíclico Fosfodiesterases/antagonistas & inibidores , Inibidores de Fosfodiesterase/síntese química , Ftalazinas/síntese química , Sítios de Ligação , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4 , Desenho de Fármacos , Avaliação Pré-Clínica de Medicamentos , Humanos , Ligação de Hidrogênio , Neutrófilos/enzimologia , Inibidores de Fosfodiesterase/química , Inibidores de Fosfodiesterase/farmacologia , Ftalazinas/química , Ftalazinas/farmacologia , Rolipram/análogos & derivados , Relação Estrutura-AtividadeRESUMO
Structure activity studies of N-phenylrolipram derivatives have led to the identification of highly potent PDE4 inhibitors. The potential of these inhibitors for cellular activity was routinely assessed in an assay of fMLP induced oxidative burst in human eosinophils. Since first generation PDE4 inhibitors have been plagued with a number of unwanted side effects, parallel structure activity studies for competition with the [3H]-rolipram binding site in rat brain were performed. In this fashion 5-[4-(3-cyclopentyloxy-4-methoxyphenyl)-2-oxo-pyrrolidin-1-yl]-3-(3-methoxybenzyloxy)benzoic acid N',N'-dimethylhydrazide (22) was identified as a potent inhibitor of PDE4 which exhibits >1000 fold selectivity versus PDE3, and is a nanomolar inhibitor in all the cellular assays tested. Studies on the stereoselectivity of PDE4 inhibition of this class of rolipram based compounds revealed, that for example (S)-11 is a more potent inhibitor than (R)-11. This effect can also be observed in primary human cells where the (S)-enantiomer is about 10 fold more potent than the corresponding (R)-enantiomer.
Assuntos
3',5'-AMP Cíclico Fosfodiesterases/antagonistas & inibidores , Inibidores de Fosfodiesterase/farmacologia , Rolipram/análogos & derivados , Rolipram/farmacologia , 3',5'-AMP Cíclico Fosfodiesterases/metabolismo , Animais , Encéfalo/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4 , Eosinófilos/efeitos dos fármacos , Eosinófilos/enzimologia , Eosinófilos/metabolismo , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Inibidores de Fosfodiesterase/química , Ratos , Explosão Respiratória/efeitos dos fármacos , Explosão Respiratória/fisiologia , Rolipram/química , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
The synthesis and pharmacological profile of a novel series of 7-methoxybenzofuran-4-carboxamides is described. Some of these compounds were found to be potent inhibitors of phosphodiesterase type 4 (PDE4).