RESUMEN
A strategy to conformationally restrain a series of GlyT1 inhibitors identified potent analogs that exhibited slowly interconverting rotational isomers. Further studies to address this concern led to a series of azetidine-based inhibitors. Compound 26 was able to elevate CSF glycine levels in vivo and demonstrated potency comparable to Bitopertin in an in vivo rat receptor occupancy study. Compound 26 was subsequently shown to enhance memory in a Novel Object Recognition (NOR) behavioral study after a single dose of 0.03 mg/kg, and in a contextual fear conditioning (cFC) study after four QD doses of 0.01-0.03 mg/kg.
Asunto(s)
Azetidinas/farmacología , Proteínas de Transporte de Glicina en la Membrana Plasmática/antagonistas & inhibidores , Memoria/efectos de los fármacos , Azetidinas/síntesis química , Azetidinas/química , Relación Dosis-Respuesta a Droga , Células HEK293 , Humanos , Estructura Molecular , Relación Estructura-ActividadRESUMEN
We report here the identification and optimization of a novel series of potent GlyT1 inhibitors. A ligand design campaign that utilized known GlyT1 inhibitors as starting points led to the identification of a novel series of pyrrolo[3,4- c]pyrazoles amides (21-50) with good in vitro potency. Subsequent optimization of physicochemical and in vitro ADME properties produced several compounds with promising pharmacokinetic profiles. In vivo inhibition of GlyT1 was demonstrated for select compounds within this series by measuring the elevation of glycine in the cerebrospinal fluid (CSF) of rats after a single oral dose of 10 mg/kg. Ultimately, an optimized lead, compound 46, demonstrated in vivo efficacy in a rat novel object recognition (NOR) assay after oral dosing at 0.1, 1, and 3 mg/kg.
Asunto(s)
Diseño de Fármacos , Proteínas de Transporte de Glicina en la Membrana Plasmática/antagonistas & inhibidores , Memoria/efectos de los fármacos , Pirazoles/síntesis química , Pirazoles/farmacología , Animales , Técnicas de Química Sintética , Proteínas de Transporte de Glicina en la Membrana Plasmática/química , Proteínas de Transporte de Glicina en la Membrana Plasmática/metabolismo , Células HEK293 , Humanos , Modelos Moleculares , Conformación Molecular , Permeabilidad , Pirazoles/química , Pirazoles/metabolismo , RatasRESUMEN
BACKGROUND: Davalintide, an investigational therapeutic peptide for the treatment of obesity, is rapidly metabolized by enzymatic cleavage of its N-terminal lysine residue to produce an active des-Lys metabolite in vivo. While a sensitive ELISA assay is available, it is unable to distinguish davalintide from its metabolite. Consequently, we developed an online SPE-LC-MS/MS method for simultaneous quantification of the drug and its active metabolite in beagle and rat plasma samples and compared the resulting pharmacokinetic profiles with those determined by ELISA. RESULTS: The total concentration of active drug measured by ELISA correlated well with the total concentration of davalintide and its metabolite using online SPE-LC-MS/MS. CONCLUSION: The technique is a viable alternative to immunochemistry-based methods for peptide quantitation in terms of sensitivity, reproducibility and specificity, and importantly, does not require developing antibody-based reagents.
