Synthesis and evaluation of the inhibitory activity of the four stereoisomers of the potent and selective human γ-glutamyl transpeptidase inhibitor GGsTop

2-Amino-4-{[3-(carboxymethyl) phenoxy](methoxy) phosphoryl} butanoic acid (GGsTop) is a potent, highly selective, nontoxic, and irreversible inhibitor of c-glutamyl transpeptidase (GGT). GGsTop has been widely used in academic and medicinal research, and also as an active ingredient (Nahlsgen) in commercial anti-aging cosmetics. GGsTop consists of four stereoisomers due to the presence of two stereogenic centers, i.e., the alpha-carbon atom of the glutamate mimic (L/D) and the phosphorus atom (R-P/S-P). In this study, each stereoisomer of GGsTop was synthesized stereoselectively and their inhibitory activity against human GGT was evaluated. The L- and D-configurations of each stereoisomer were determined by a combination of a chiral pool synthesis and chiral HPLC analysis. The synthesis of the four stereoisomers of GGsTop used chiral synthetic precursors that were separated by chiral HPLC on a preparative scale. With respect to the configuration of the alpha-carbon atom of the glutamate mimic, the L-isomer (k(on) = 174 M-1 s(-1)) was ca. 8-fold more potent than the D-isomer (k(on) = 21.5 M-1 s(-1)). In contrast, the configuration of the phosphorus atom is critical for GGT inhibitory activity. Based on a molecular modeling approach, the absolute configuration of the phosphorus atom of the active GGsTop isomers was postulated to be S-P. The S-P-isomers inhibited human GGT (k(on) = 21.5-174 M-1 s(-1)), while the R-P-isomers were inactive even at concentrations of 0.1 mM. (C) 2017 Elsevier Ltd. All rights reserved.