Unexpected relationship between plasma protein binding and the pharmacodynamics of 2‐NAP, a CCK1‐receptor antagonist

What is already known about this subject? • Two chemically diverse CCK1 receptor antagonists have been shown clinically to inhibit CCK‐evoked contraction of human gallbladder [2, 3]. These studies have not examined the relationship between plasma concentration and effect, the latter usually considered to be predictive from the free drug concentration [8]. • We wanted to examine our novel CCK1 receptor antagonist in this validated model and also to explore its PK‐PD relationship. What this study adds • 2‐NAP inhibited CCK‐evoked human gallbladder contraction in vivo but at a plasma free concentration that was, in theory, too low to have achieved adequate CCK1 receptor occupancy. • The study serves as a caveat to the assumption that free plasma concentration can be used to predict pharmacological effect. Aims To study the pharmacokinetics and pharmacodynamics of 2‐NAP (2‐naphthalenesulfonyl‐L‐aspartyl‐(2‐phenethyl)amide), a selective CCK1 receptor antagonist in healthy volunteers. Methods 2‐NAP was given to 12 healthy male volunteers in an ascending dose, safety and PK phase 1a study by 1 h i.v. infusion (0.6–9.6 mg kg−1 h−1). A further 12 healthy male volunteers received i.v. CCK‐8S (6.25 pmol kg−1 h−1) to produce gallbladder contraction, measured by ultrasound recordings of gallbladder volume, and the effect of concurrent i.v. 2‐NAP administration was studied. Plasma protein binding in vitro and ex vivo was measured by ultrafiltration and by equilibrium dialysis. Results 2‐NAP was generally well tolerated, displayed linear pharmacokinetics and a very high degree of plasma protein binding (99.9%). A 105 min i.v. CCK‐8S infusion induced a reduction in gallbladder volume of 14.9 (±7.0) ml during placebo co‐infusion and this was reduced to 2.4 (±5.9) ml when 2‐NAP was co‐infused with CCK‐8S (P = 0.00024, paired t‐test, mean change 12.5 ml; 95% CI For mean 7.4, 18.3 ml). This extent of inhibition was consistent with a 2‐NAP total plasma concentration of 36 µm, but when protein binding corrections were made, the ‘free concentration’ of 2‐NAP was only 0.04 µm, a value much less than the average equilibrium dissociation constant of 2‐NAP for human CCK1 receptors (∼0.7 µm). Conclusions The pharmacological effect of a drug is usually considered to be determined by its free concentration. However, the complete inhibition of CCK‐8S‐evoked gallbladder contraction by a free plasma concentration of 0.04 µm 2‐NAP was much greater than would have been predicted from simple