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Adaptor-associated kinase 1 (AAK1), a member of the Ark1/Prk1 family of serine/threonine kinases, plays a role in modulating clatherin coated endocytosis of specific surface receptors. We have demonstrated that AAK1 inhibitors are efficacious in rodent models of neuropathic pain (Kostich et al., 2016). Here we have characterized the binding properties and distribution pattern of the tritiated AAK1 radioligand, [ 3 H]BMT-046091, in rodents and cynomolgus monkeys, and used the radioligand to measure the brain target occupancy following systemic administration of AAK1 inhibitors. We have found that [ 3 H]BMT-046091 is potent and selective AAK1 inhibitor. It inhibits AAK1 phosphorylation of a peptide derived from a physiologic substrate, the μ2 subunit of the adaptor protein complex, with an IC 50 value of 2.8 nM, and is inactive at >5 μM in a panel of functional or binding assays for receptors, transporters and enzymes. [ 3 H]BMT-046091 binding in the brain is absent in the AAK1 knockout mouse, and is displaceable with a high concentration of AAK1 inhibitors in wild type mice. Specific [ 3 H]BMT-046091 binding is widespread in the brain and spinal cord with the highest density in the cortex, hippocampus, amygdala, striatum and thalamus. In the spinal cord, [ 3 H]BMT-046091 binding appears enriched in the dorsal horn superficial layers. Oral administration of LP-935509, an AAK1 inhibitor, results in a dose-dependent occupation of AAK1 binding sites in the brain and spinal cord. The increase in AAK1 binding site occupancy by LP-935509 correlates with the decrease in antinociceptive responses in the rat chronic constriction injury model of neuropathic pain. [ABSTRACT FROM AUTHOR]

Abstract: BMS-763534 is a potent (CRF1 IC50 = 0.4 nM) and selective (>1000-fold selectivity vs. all other sites tested) CRF1 receptor antagonist (pA2 = 9.47 vs. CRF1-mediated cAMP production in Y79 cells). BMS-763534 accelerated the dissociation of 125I-o-CRF from rat frontal cortex membrane CRF1 receptors consistent with a negative allosteric modulation of CRF binding. BMS-763534 produced dose-dependent increases in CRF1 receptor occupancy and anxiolytic efficacy; lowest effective anxiolytic dose = 0.56 mg/kg, PO, which was associated with 71 ± 5% CRF1 receptor occupancy of frontoparietal CRF1 receptors. Sedative/ataxic effects of BMS-763534 were only observed at high dose multiples (54–179×) relative to the lowest dose required for anxiolytic efficacy. At doses of 5- to 18-fold higher than the lowest efficacious dose in the anxiety assay, BMS-763534 shared subjective effects with the benzodiazepine chlordiazepoxide. Interestingly BMS-790318, the O-demethylated metabolite of BMS-763534, showed weak affinity for the TBOB site of the GABAA receptor (67% inhibition at 10 μM) and augmented GABA evoked currents (EC50 = 1.6 μM). Thus, the unanticipated signal in the drug discrimination assay may have resulted from an interaction of the metabolite BMS-790318 with the TBOB site on the GABAA channel where it appears to behave as an allosteric potentiator of GABA evoked currents. [Copyright &y& Elsevier]