NVP-DPP728 - (1-[[[2-[(5-cyanopyridin-2-yl) amino]ethyl] amino] acetyl]-2-cyano-(S)-pyrrolidine), a slow-binding inhibitor of dipeptidyl peptidase IV

Inhibition of dipeptidyl peptidase IV (DPP-IV) has been proposed recently as a therapeutic approach to the treatment of type 2 diabetes. N-Substituted-glycyl-2-cyanopyrrolidide compounds, typified by NVP-DPP728 (1-[[[2-[(5-cyanopyridin-2-yl)amino]ethyl]amino]acetyl]-2-cyano-(S)-pyrrolidine), inhibit degradation of glucagon-like peptide-1 (GLP-1) and thereby potentiate insulin release in response to glucose-containing meals. In the present study NVP-DPP728 was found to inhibit human DPP-IV amidolytic activity with a K-i of 11 nM, a k(on) value of 1.3 x 10(5) M-1 s(-1), and a k(off) of 1.3 x 10(-3) s(-1). Purified bovine kidney DPP-IV bound 1 mol/mol [C-14]-NVP-DPP728 with high affinity (12 nM K-d) The dissociation constant, k(off), was 1.0 x 10(-3) and 1.6 x 10(-3) s(-1) in the presence of 0 and 200 mu M H-Gly-Pro-AMC, respectively (dissociation t(1/2) similar to 10 min). Through kinetic evaluation of DPP-IV inhibition by the D-antipode, des-cyano, and amide analogues of NVP-DPP728, it was determined that the nitrile functionality at the 2-pyrrolidine position is required, in the L-configuration, for maximal activity (K-i of 11 nM vs K-i values of 5.6 to >300 mu M for the other analogues tested). Surprisingly, it was found that the D-antipode, despite being similar to 500-fold less potent than NVP-DPP728, displayed identical dissociation kinetics (k(off) of 1.5 x 10(-3) s(-1)). NVP-DPP728 inhibited DPP-IV in a manner consistent with a two-step inhibition mechanism. Taken together, these data suggest that NVP-DPP728 inhibits DPP-IV through formation of a novel, reversible, nitrile-dependent complex with transition state characteristics.