RESUMEN

We identified (5'S)-10'-fluoro-6'-methyl-5',6'-dihydro-3'H-spiro[cyclopropane-1,4'-[2,6]diaza[2,5]methano[2,6]benzodiazonin]-7'(1'H)-one, 22b (DS34942424) with a unique and original bicyclic skeleton. 22b showed an orally potent analgesic in the acetic acid-induced writhing test and formalin test in ddY mice without sedation. Moreover, 22b did not exhibit mu opioid receptor agonist activity.

Asunto(s)

Analgésicos Opioides/química , Receptores Opioides mu/agonistas , Compuestos de Espiro/química , Administración Oral , Analgésicos Opioides/metabolismo , Analgésicos Opioides/farmacología , Analgésicos Opioides/uso terapéutico , Animales , Conducta Animal/efectos de los fármacos , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos , Estabilidad de Medicamentos , Masculino , Ratones , Microsomas Hepáticos/metabolismo , Dolor/inducido químicamente , Dolor/tratamiento farmacológico , Dolor/patología , Receptores Opioides mu/metabolismo , Compuestos de Espiro/metabolismo , Compuestos de Espiro/farmacología , Compuestos de Espiro/uso terapéutico , Relación Estructura-Actividad

RESUMEN

We synthesized derivatives of a natural alkaloid, conolidine, and evaluated these derivatives in the acetic acid-induced writhing test and formalin test in ddY mice after oral administration. As a result, we identified (5S)-6-methyl-1,3,4,5,6,8-hexahydro-7H-2,5-methano[1,5]diazonino[7,8-b]indol-7-one sulfate salt, 15a (DS54360155), with a unique and original bicyclic skeleton, as an analgesic more potent than conolidine. Moreover, 15a did not exhibit mu-opioid receptor agonist activity.

Asunto(s)

Analgésicos/uso terapéutico , Receptores Opioides mu/agonistas , Analgésicos/farmacología , Animales , Modelos Animales de Enfermedad , Humanos , Ratones

RESUMEN

We discovered a novel compound, 5-methyl-1,4,5,7-tetrahydro-2,5-ethanoazocino[4,3-b]indol-6(3H)-one sulfuric acid salt (DS39201083), which was formed by derivatization of a natural product, conolidine. DS39201083 had a unique bicyclic skeleton and was a more potent analgesic than conolidine, as revealed in the acetic acid-induced writhing test and formalin test in ddY mice. The compound showed no agonist activity at the mu opioid receptor.

Asunto(s)

Analgésicos Opioides/uso terapéutico , Alcaloides Indólicos/uso terapéutico , Receptores Opioides mu/uso terapéutico , Analgésicos Opioides/farmacología , Animales , Alcaloides Indólicos/farmacología , Ratones , Receptores Opioides mu/agonistas

RESUMEN

PURPOSE: The short half lives of small molecules in the vitreous requires frequent repeated intravitreal injections that are impractical for treatment of chronic eye diseases. We sought to develop a method for increasing the intravitreal half-life of small-molecule drugs. METHODS: We adapted a technology for controlled release of drugs from macromolecular carriers for use as a long-acting intravitreal delivery system for small molecules. As a prototype, a small molecule complement factor D inhibitor with an intravitreal half-life of 7 hours was covalently attached to a 4-arm PEG40kDa by a self-cleaving ß-eliminative linker with a cleavage half-life of approximately 1 week. RESULTS: After intravitreal injection in rabbits, the drug was slowly released in the vitreous, and equilibrated with the retina and choroid. The intravitreal half-life of the intact PEG-drug conjugate in the rabbit was 7 days, and that of the released drug was 3.6 days. We simulated the anticipated pharmacokinetics of the delivery system in human vitreous, and estimated that the half-life of a 4-arm PEG40kDa conjugate would be approximately 2 weeks, and that of the released drug would be approximately 5 days. CONCLUSIONS: We posit that a linker with a cleavage half life of 2 weeks would confer a half life of approximately 7 days to a released small molecule drug in humans, comparable to the half life of approved intravitreal injected macromolecular drugs. TRANSLATIONAL RELEVANCE: With this technology, a potent small molecule with an appropriate therapeutic window should be administrable by intravitreal injections in the human at once-monthly intervals.

RESUMEN

Compounds that modulate the activity of sphingosine 1-phosphate (S1P)-metabolizing enzymes are expected to be potential therapeutic agents for various diseases. Investigation of their potencies requires not only cell-free but also cell-based assays in which intracellular accumulation/depletion of S1P could be monitored. However, conventional methods have limitations to their simplicity, mainly due to the necessity of a separation process that separates S1P from its related substances. Here, we describe a method utilizing a scintillation proximity assay (SPA) for semi-quantifying intracellular [(3)H]-labeled dihydroS1P ([(3)H]dhS1P), which is also a substrate for S1P-metabolizing enzymes. We found that uncoated yttrium silicate SPA beads could selectively bind to and detect [(3)H]dhS1P rather than [(3)H]dihydrosphingosine (the non-phosphorylated form of [(3)H]dhS1P). Based on this, we developed a novel cell-based assay system which does not require any organic solvent extraction or chromatographic separation, and confirmed its practicality by using siRNA targeting S1P lyase (S1PL) and known S1PL inhibitors as models. Our results demonstrated that this assay is useful for rapid and easy evaluation of S1PL inhibitors, and could be potentially applicable for all compounds that modulate the activity of S1P-metabolizing enzymes.

Asunto(s)

Aldehído-Liasas/antagonistas & inhibidores , Técnicas de Química Analítica/métodos , Esfingosina/análogos & derivados , Animales , Células Cultivadas , Inhibidores Enzimáticos , Ratones , ARN Interferente Pequeño/farmacología , Transducción de Señal , Esfingosina/análisis , Esfingosina/química , Itrio/química

RESUMEN

Caramel color is widely used in the food industry, and its many variations are generally considered to be safe. It has been known for a long time that THI (2-acetyl-4-(tetrahydroxybutyl)imidazole), a component of caramel color III, causes lymphopenia in animals through sphingosine 1-phosphate (S1P) lyase (S1PL) inhibition. However, this mechanism of action has not been fully validated because THI does not inhibit S1PL in vitro. To reconcile this situation, we examined molecular details of THI mechanism of action using "smaller" THI derivatives. We identified a bioactive derivative, A6770, which has the same lymphopenic effect as THI via S1PL inhibition. In the case of A6770 we observe this effect both in vitro and in vivo, and demonstrate that A6770 is phosphorylated and inhibits S1PL in the same way as 4-deoxypyridoxine. In addition, A6770 was detected in rat plasma following oral administration of THI, suggesting that A6770 is a key metabolic intermediate of THI.

Asunto(s)

Colorantes de Alimentos/farmacología , Imidazoles/farmacología , Linfopenia/inducido químicamente , Linfopenia/metabolismo , Lisofosfolípidos/antagonistas & inhibidores , Esfingosina/análogos & derivados , Animales , Línea Celular , Relación Dosis-Respuesta a Droga , Colorantes de Alimentos/administración & dosificación , Colorantes de Alimentos/química , Imidazoles/administración & dosificación , Imidazoles/química , Lisofosfolípidos/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratas , Ratas Endogámicas Lew , Esfingosina/antagonistas & inhibidores , Esfingosina/metabolismo , Relación Estructura-Actividad

RESUMEN

Caramel food colorant 2-acetyl-4-(tetrahydroxybutyl)imidazole (THI) causes lymphopenia in animals through sphingosine 1-phosphate lyase (SPL) inhibition. However, this mechanism of action is partly still controversial because THI did not inhibit SPL in vitro either in cell-free or in cell-based systems. It is thought that the in vitro experimental conditions which have been used so far were not suitable for the evaluation of SPL inhibition, especially in case of cell-based experiments. We speculated that the key factor might be the coenzyme pyridoxal 5'-phosphate (PLP), an active form of vitamin B6 (VB6), because media used in cell-based assays usually contain an excess amount of VB6 which leads to the activation of SPL. By the use of VB6-deficient culture medium, we could regulate apo- (without PLP) and holo- (with PLP) SPL enzyme in cultured cells, resulting in the successful detection of SPL inhibition by THI. Although the observed inhibitory effect was not as strong as that of 4-deoxypyridoxine (a VB6 analog SPL inhibitor), these findings may be useful for further understanding the mechanism of action of THI.

Asunto(s)

Aldehído-Liasas/metabolismo , Imidazoles/farmacología , Deficiencia de Vitamina B 6/enzimología , Aldehído-Liasas/antagonistas & inhibidores , Animales , Carbohidratos/farmacología , Ratones , Fosfato de Piridoxal/metabolismo , Deficiencia de Vitamina B 6/metabolismo , Deficiencia de Vitamina B 6/patología

RESUMEN

We have carried out the optimization of substituents at the C-3 or the C-5 position on the pyrrolidine ring of VLA-4 antagonist 3 with 2-(phenylamino)-7-fluorobenzoxazolyl moiety for the purpose of improving in vivo efficacy while maintaining good aqueous solubility. As a result, we successfully increased in vitro activity in the presence of 3% human serum albumin and achieved an exquisite lipophilic and hydrophilic balance of compounds suitable for oral administrative regimen. The modification resulted in the identification of zwitterionic compound 7n with (5S)-[methoxy(methyl)amino]methylpyrrolidine, which significantly alleviated bronchial hyper-responsiveness to acetylcholine chloride at 12.5mg/kg, p.o. in a murine asthma model and showed favorable aqueous solubility (JP1, 89 µg/mL; JP2, 462 µg/mL). Furthermore, this compound showed good oral bioavailability (F=54%) in monkeys.

Asunto(s)

Antiinflamatorios/química , Antiinflamatorios/uso terapéutico , Asma/tratamiento farmacológico , Ácidos Ciclohexanocarboxílicos/química , Ácidos Ciclohexanocarboxílicos/uso terapéutico , Integrina alfa4beta1/antagonistas & inhibidores , Administración Oral , Animales , Antiinflamatorios/administración & dosificación , Antiinflamatorios/farmacocinética , Asma/inmunología , Disponibilidad Biológica , Bronquios/efectos de los fármacos , Bronquios/inmunología , Línea Celular , Ácidos Ciclohexanocarboxílicos/administración & dosificación , Ácidos Ciclohexanocarboxílicos/farmacocinética , Eosinófilos/efectos de los fármacos , Eosinófilos/inmunología , Femenino , Haplorrinos , Humanos , Integrina alfa4beta1/inmunología , Ratones , Ratones Endogámicos BALB C , Pirrolidinas/administración & dosificación , Pirrolidinas/química , Pirrolidinas/farmacocinética , Pirrolidinas/uso terapéutico , Solubilidad , Agua/química

RESUMEN

This contribution describes a concise synthesis to ethyl trans-[(4S)-methoxy-(2S)-pyrrolidinylmethoxy]cyclohexanecarboxylate (2b) as a key intermediate of very late antigen-4 (VLA-4) antagonist trans-4-[1-[[2,5-dichloro-4-(1-methyl-3-indolylcarboxyamide)phenyl]acetyl]-(4S)-methoxy-(2S)-pyrrolidinylmethoxy]cyclohexanecarboxylic acid (1). The synthesis employs a reductive etherification as a key reaction using (2S,4S)-1-benzyloxycarbonyl-4-methoxypyrrolidine-2-carboxyaldehyde (12) and trans-4-triethylsilyloxycyclohexanecarboxilic acid ethyl ester (13b). This synthesis provides 2b in 6 steps with 38% overall yield from commercially available starting material.

Asunto(s)

Ácidos Ciclohexanocarboxílicos/química , Ácidos Ciclohexanocarboxílicos/síntesis química , Integrina alfa4beta1/antagonistas & inhibidores , Pirrolidinas/química , Pirrolidinas/síntesis química , Integrina alfa4beta1/metabolismo , Cetonas/química , Oxidación-Reducción , Estereoisomerismo

RESUMEN

For the purpose of obtaining orally potent VLA-4 inhibitors, we have carried out structural modification of the (N'-phenylureido)phenyl group in compound 1, where the group was found to be attributed to poor pharmacokinetic profile in our previous research. Through modification, we have identified several compounds with both potent in vitro activity and improved oral exposure. In particular, compound 7e with 7-fluoro-2-(1-methyl-1H-indol-3-yl)-1,3-benzoxazolyl group as a novel replacement of the (N'-phenylureido)phenyl group significantly inhibited eosinophil infiltration into bronchoalveolar lavage fluid at 15mg/kg in an Ascaris-antigen-induced murine bronchial inflammatory model, and its efficacy was comparable to that of the anti-mouse α(4) antibody (R1-2).

Asunto(s)

Asma/tratamiento farmacológico , Ácidos Ciclohexanocarboxílicos/química , Ácidos Ciclohexanocarboxílicos/uso terapéutico , Integrina alfa4beta1/antagonistas & inhibidores , Administración Oral , Animales , Asma/inmunología , Bronquios/efectos de los fármacos , Bronquios/inmunología , Lavado Broncoalveolar , Línea Celular , Ácidos Ciclohexanocarboxílicos/farmacocinética , Ácidos Ciclohexanocarboxílicos/farmacología , Eosinófilos/efectos de los fármacos , Eosinófilos/inmunología , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Relación Estructura-Actividad

RESUMEN

This contribution describes a novel synthetic approach to very late antigen-4 (VLA-4) antagonist trans-4-[1-[[2,5-dichloro-4-(1-methyl-3-indolylcarboxyamide)phenyl]acetyl]-(4S)-methoxy-(2S)-pyrrolidinylmethoxy]cyclohexanecarboxylic acid (1) via tert-butyl trans-[(4S)-methoxy-(2S)-pyrrolidinylmethoxy]cyclohexanecarboxylate (2b) as a key intermediate. The synthesis, which includes n-Bu4NSO3H that catalyzed basic etherification of 12 and iodine-mediated cyclization to provide the 2,4-disubstituted pyrrolidine frame of 2b, is designed to utilize trans-4-hydroxycyclohexanecarboxylic acid (9) as a commercially available starting material.

Asunto(s)

Antiasmáticos/farmacología , Ácidos Ciclohexanocarboxílicos/farmacología , Integrina alfa4beta1/antagonistas & inhibidores , Amidas/química , Antiasmáticos/síntesis química , Antiasmáticos/química , Ciclización , Ácidos Ciclohexanocarboxílicos/síntesis química , Ácidos Ciclohexanocarboxílicos/química , Éteres/química , Espectroscopía de Resonancia Magnética , Modelos Químicos , Pirrolidinas/química , Espectrofotometría Infrarroja , Estereoisomerismo

RESUMEN

We have focused on optimization of the inadequate pharmacokinetic profile of trans-4-substituted cyclohexanecarboxylic acid 5, which is commonly observed in many small molecule very late antigen-4 (VLA-4) antagonists. We modified the lipophilic moiety in 5 and found that reducing the polar surface area of this moiety results in improvement of the PK profile. Consequently, our efforts have led to the discovery of trans-4-[1-[[2,5-dichloro-4-(1-methyl-3-indolylcarboxamido)phenyl]acetyl]-(4S)-methoxy-(2S)-pyrrolidinylmethoxy]cyclohexanecarboxylic acid (14e) with potent activity (IC(50) = 5.4 nM) and significantly improved bioavailability in rats, dogs, and monkeys (100%, 91%, 68%), which demonstrated excellent oral efficacy in murine and guinea pig models of asthma. Based on its overall profile, compound 14e was progressed into clinical trails. In a single ascending-dose phase I clinical study, compound 14e exhibited favorable oral exposure as expected and had no serious adverse events.

Asunto(s)

Ácidos Ciclohexanocarboxílicos/química , Ácidos Ciclohexanocarboxílicos/farmacocinética , Indoles/farmacocinética , Integrina alfa4beta1/antagonistas & inhibidores , Pirrolidinas/farmacocinética , Administración Oral , Animales , Disponibilidad Biológica , Células CHO , Cricetinae , Cricetulus , Ácidos Ciclohexanocarboxílicos/síntesis química , Ácidos Ciclohexanocarboxílicos/farmacología , Perros , Cobayas , Haplorrinos , Humanos , Indoles/síntesis química , Indoles/química , Indoles/farmacología , Ratones , Pirrolidinas/síntesis química , Pirrolidinas/química , Pirrolidinas/farmacología , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad

RESUMEN

During the course of our study, it was revealed that the poor pharmacokinetic properties of a series of benzoic acid derivatives such as 1 should be attributed to the diphenylurea moiety. Thus, we replaced the diphenylurea moiety in 1 with a 2-(2-methylphenylamino)benzoxazole moiety which mimics the diphenylurea structure. However, this modification resulted in a significant decrease (3, IC(50)=19 nM) in VLA-4 inhibitory activity compared to 1 (IC(50)=1.6 nM). To address this discrepancy, we worked on optimization of the carboxylic acid moiety in compound 3. As a result, our efforts have led to the discovery of trans-4-substituted cyclohexanecarboxylic acid derivative 11b (IC(50)=2.8 nM) as a novel and potent VLA-4 antagonist. In addition, compound 11b exhibited favorable pharmacokinetic properties (CL=3.3 ml/min/kg, F=51%) in rats.

Asunto(s)

Benzoxazoles/química , Ácidos Ciclohexanocarboxílicos/química , Integrina alfa4beta1/antagonistas & inhibidores , Animales , Benzoxazoles/síntesis química , Benzoxazoles/farmacocinética , Células CHO , Cricetinae , Cricetulus , Cristalografía por Rayos X , Ácidos Ciclohexanocarboxílicos/síntesis química , Ácidos Ciclohexanocarboxílicos/farmacocinética , Concentración 50 Inhibidora , Integrina alfa4beta1/metabolismo , Masculino , Conformación Molecular , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad

RESUMEN

Optimization of benzoic acid derivatives by introducing substituents into the diphenyl urea moiety led to the identification of compound 20l as a potent VLA-4 antagonist. Compound 20l inhibited eosinophil infiltration into bronchial alveolar lavage fluid in a murine asthma model by oral dosing and its efficacy was comparable to anti-mouse alpha4 antibody (R1-2). Furthermore, this compound significantly blocked bronchial hyper-responsiveness in the model.

Asunto(s)

Benzoatos/farmacología , Integrina alfa4beta1/antagonistas & inhibidores , Pirroles/farmacología , Administración Oral , Animales , Antiinflamatorios/farmacología , Asma/tratamiento farmacológico , Benzoatos/administración & dosificación , Benzoatos/síntesis química , Células Cultivadas , Modelos Animales de Enfermedad , Perros , Femenino , Concentración 50 Inhibidora , Ratones , Ratones Endogámicos BALB C , Permeabilidad , Pirroles/administración & dosificación , Pirroles/síntesis química , Ratas

RESUMEN

A series of benzoic acid derivatives was synthesized as VLA-4 antagonists. Introduction of chlorine or bromine into the 3-position on the central benzene of the diphenylurea portion as in lead compound 2 led to improvement in the pharmacokinetic properties. In particular, 12l demonstrated an acceptable plasma clearance and bioavailability in mice and rats as well as dogs (mice, CL=18.5 ml/min/kg,F=28%; rats, CL=5.2 ml/min/kg,F=36%; dogs, CL=3.6 ml/min/kg,F=55%). Additionally, 12l exhibited potent activity with an IC50 value of 0.51 nM and efficacy by oral administration at a dosage of 10 mg/kg in a rat pleurisy model.

Asunto(s)

Benzoatos/síntesis química , Benzoatos/farmacocinética , Integrina alfa4beta1/antagonistas & inhibidores , Administración Oral , Animales , Benzoatos/farmacología , Modelos Animales de Enfermedad , Perros , Inflamación/tratamiento farmacológico , Concentración 50 Inhibidora , Ratones , Farmacocinética , Pleuresia/tratamiento farmacológico , Ratas , Relación Estructura-Actividad

RESUMEN

A novel series of benzoic acid derivatives as VLA-4 antagonists were synthesized. Optimization, focusing on activity and lipophilicity needed for cell permeability, resulted in the identification of 15b and 15e with good activity (IC50 = 1.6 nM each) and moderate lipophilicity (Log D = 2.0, 1.8). Furthermore, 15e demonstrated efficacy in murine asthma model by an oral dose of 30 mg/kg.

Asunto(s)

Asma/tratamiento farmacológico , Benzoatos/administración & dosificación , Benzoatos/farmacocinética , Integrina alfa4beta1/antagonistas & inhibidores , Pirrolidinas/química , Pirrolidinonas/administración & dosificación , Pirrolidinonas/farmacocinética , Administración Oral , Animales , Benzoatos/química , Permeabilidad de la Membrana Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Perros , Evaluación Preclínica de Medicamentos , Activación Enzimática/efectos de los fármacos , Éteres de Hidroxibenzoatos , Riñón/citología , Masculino , Ratones , Ratones Endogámicos BALB C , Conformación Molecular , Pirrolidinonas/química , Ratas , Ratas Sprague-Dawley , Estereoisomerismo , Relación Estructura-Actividad

RESUMEN

A series of prolyl-1-piperazinylacetic acid and prolyl-4-piperidinylacetic acid derivatives were synthesized and evaluated for their activity as VLA-4 antagonists. Of 22 compounds synthesized, 19 compounds showed potent activity with low nanomolar IC50 values. In addition, the representative compounds 11o and 11p with a hydroxy group in the pyrrolidine ring showed moderate plasma clearance in rats (11o, 30 ml/min/kg and 11p, 21 ml/min/kg) and in dogs (11o, 12 ml/min/kg and 11p, 9 ml/min/kg).

Asunto(s)

Acetatos/síntesis química , Integrina alfa4beta1/antagonistas & inhibidores , Piperazinas/síntesis química , Piperidinas/síntesis química , Prolina/análogos & derivados , Acetatos/farmacocinética , Acetatos/farmacología , Animales , Perros , Espectroscopía de Resonancia Magnética , Masculino , Piperazinas/farmacocinética , Piperazinas/farmacología , Piperidinas/farmacocinética , Piperidinas/farmacología , Prolina/síntesis química , Prolina/farmacocinética , Prolina/farmacología , Ratas , Ratas Sprague-Dawley , Espectrometría de Masa Bombardeada por Átomos Veloces

RESUMEN

An investigation into the structure-activity relationship of a lead compound, prolyl-5-aminopentanoic acid 4, led to the identification of a novel series of 4-piperidinylacetic acid, 1-piperazinylacetic acid, and 4-aminobenzoic acid derivatives as potent VLA-4 antagonists with low nanomolar IC(50) values. A representative compound morpholinyl-4-piperidinylacetic acid derivative (13d: IC(50)=4.4 nM) showed efficacy in the Ascaris-antigen sensitized murine airway inflammation model by oral administration.

Asunto(s)

Integrina alfa4beta1/antagonistas & inhibidores , Morfolinas/farmacología , Piperidinas/farmacología , Administración Oral , Animales , Asma/etiología , Asma/metabolismo , Ratones , Morfolinas/administración & dosificación , Morfolinas/química , Piperidinas/administración & dosificación , Piperidinas/química