Pharmacodynamic effects of BIA 10-2474, a novel FAAH inhibitor and evidence of target engagement in the mouse

M. J. Bonifácio1, C. Lopes1, N. Pires1, A. I. Loureiro1, P. Moser1, P. Soares-da-Silva1,2. 1Dept. Research & Development, BIAL – Portela & Cª, S.A., 4745-457 Coronado (S. Romão e S. Mamede), PORTUGAL, 2Dept. Pharmacology & Therapeutics, Faculty of Medicine, University Porto, Porto, PORTUGAL.

Introduction: BIA 10-2474 (3-(1-(cyclohexyl (methyl)carbamoyl)-1H-imidazol-4-yl)pyridine 1-oxide) is a novel fatty acid amide hydrolase (FAAH) inhibitor. This study reports on the PK/PD relationship for BIA 10-2474 and FAAH inhibition to assist in defining its clinical development.

Methods: Animal experiments were conducted in strict adherence to the 2010/63/EU and Portuguese law on animal welfare. Used 48 CD1 and138 NMRI mice. BIA 10-2474, anandamide (AEA), palmitoylethanolamide (PEA) and oleoylethanolamide (OEA) were quantified in plasma from CD1 mice orally given BIA 10-2474, by LC/MS. FAAH activity (using [3H]-AEA as substrate) was assayed in NMRI liver and brain homogenates at 1 and 4h after BIA 10-2474 administration (1). A functional interaction with FAAH was further demonstrated by co-administering BIA 10-2474 with AEA using the formalin paw test (2). 25µl of 5% formalin was injected into the plantar region of NMRI mouse hind-paw and paw licking behaviour was evaluated between 15 and 50 minute post-administration.

Results: Oral administration of BIA 10-2474 to mice (0.001-3 mg/kg, n=4) resulted in dose and time-dependent inhibition of FAAH activity in the brain with ED50 values of approximately 94 and 27 µg/kg at 1h and 8h post-dosing and in the liver (11 and 7 µg/kg respectively). Administration of BIA 10-2474 to mice resulted in significant increase in AEA brain levels after 4h (vehicle BLQ, 1 mg/kg: 98±4 pmol/g; 3 mg/kg: 92±5 pmol/g). A PK/PD study using 10 mg/kg (po) showed that BIA 10-2474 had a Cmax in plasma at 1h (3616±437 ng/ml, n=4) and was almost undetectable at 8h (83±14 ng/ml, n=4). In contrast, brain AEA levels (which were very low in control animals, 0-13 pmol/g) peaked at 4h (113.7±8.8 pmol/g, n=4), were still elevated at 8h (100.4±9.3 pmol/g, n=3) but returned to control levels by 24h. Brain concentrations of PEA and OEA, showed similar time-courses. In the formalin paw test, AEA (5 mg/kg ip), 10 min after formalin slightly but significantly reduced licking behaviour (table 1). Administration of BIA 10-2474 markedly facilitated the analgesic effects of exogenously applied AEA (table 1, n=10 for all groups).

Conclusion: These data show that BIA 10-2474 is a potent FAAH inhibitor in vivo with sustained inhibition of FAAH for up to 8h in the mouse. BIA 10-2474 increases brain levels of AEA and other fatty acid amides and is able to potentiate the analgesic effects of AEA.

1- Boldrup L. et al (2004) JBiochemBiophysMeth 60:171-177; 2- Wheeler-Aceto H. & Cowan A. (1991) Psychopharmacology 104:35-44