[3H]BMS-599240--a novel tritiated ligand for the characterization of BACE1 inhibitors.

In this report we describe a novel radioligand, [(3)H](S)-2-((S)-3-Acetylamino-3-sec-butyl-2-oxo-pyrrolidin-1-yl)-N-[(1S,2R)-1-benzyl-2-hydroxy-3-(3-methoxy-benzylamino)-propyl]-4-phenyl-butyramide ([(3)H]BMS-599240), that exhibits robust specific binding in homogenates from cell cultures overexpressing beta-site amyloid precursor protein cleaving enzyme-1 (BACE1). Radioligand binding exhibited high affinity, K(d)=2 nM, commensurate with its inhibitory potency against BACE1. Inhibition of radioligand binding in the presence of a range of different BACE1 inhibitors exhibited the same rank order of potency as for inhibition of BACE1 enzymatic activity. BACE1-dependent binding of the radioligand was also demonstrated in mouse brain homogenates, where genetic ablation of BACE1 eliminated high affinity binding. Thus, the radioligand [(3)H]BMS-599240 is a novel tool potentially useful for evaluation of BACE1 enzyme in biological samples, and for evaluation of inhibitor binding to BACE1.