G. SMAGIN1, D. P. BUDAC1, M. CAJINA1, A. LEE1, B. CAMPBELL1, G. LI1, C. SANCHEZ2, C. FORRAY1, V. S. PALAMARCHOUK3, D. SONG1
1Lundbeck Res., Paramus, NJ; 2Lundbeck Res USA, Inc., Paramus, NJ; 3PsychoGenics Inc, Tarrytown, NY
Degradation of the amino acid tryptophan (TRP) along the kynurenine pathway (KP) yields several neuroactive intermediates regulated by enzymes localized in astrocytes and microglial cells. Impairment of KP metabolism is functionally significant in a variety of diseases that affect the brain. It is hypothesized that CNS penetrant small molecules which inhibit kynurenine 3-monooxygenase (KMO) normalize central kynurenine dysregulation and will be beneficial for CNS disorders. We identified a CNS penetrant KMO inhibitor Compound A and characterized its effect on central and peripheral KP metabolites after systemic administration. We demonstrated that by extending the analyte capacity of an LC/MS/MS method, dedicated to KP metabolite, to include drug molecules we could simultaneously monitor KP metabolites and drug compounds in one injection. Utilizing this technique alongside in vivo microdialysis allows us to perform direct comparisons of drug levels and their impact on KP metabolites over a time course.
Mice were implanted with the guide cannula placed into the striatum. One week after surgery, a microdialysis probe was inserted for the microdialysis experiment. Microdialysis samples were collected for 6 hrs after 30 and 100 mg/kg administration of Compound A and analyzed for Compound A, L-kynurenine (KYN), kynurenic acid (KYNA), 3-hydrohykynurenine (3-HK) and quinolinic acid (QUIN), using LC/MS/MS Terminal brain and plasma samples were taken for analysis of KP metabolites and compound exposure.
Administration of Compound A decreased concentrations 3-HK to 60% of baseline which co-incided with the increased concentration of Compound A in dialysis samples. Initial decrease in 3-HK concentrations was followed by a two-fold increase after 6 hrs. Concentrations of KYN, KYNA and QUIN in microdialysates collected from the brain increased 75-fold, 350-fold and 2-fold respectively, compared to vehicle control. The increase lasted through the six hour observation period. KYN and KYNA levels were also elevated in terminal brain and plasma samples collected at the 6 hour mark. The results demonstrate that CNS penetrant KMO inhibitors while affecting both central and peripheral KP metabolites provide an avenue to understanding the biological role of these metabolites in brain disorders. Further studies of brain-penetrating modulators of KP may provide treatment options for such disorders.