Functional and binding kinetic studies make a distinction between OX1 and OX2 orexin receptor antagonists.

In this work the pharmacology and the receptor kinetics of the following orexin receptor antagonists SB-649868, ACT-078573, JNJ-10397049, MK-6096 and Roche-Cp were evaluated at human OX(1) and OX(2) orexin receptors by using functional and receptor binding assays. Kinetic analysis of the unlabeled ligands was carried out by indirect measurement according to the Motulski and Mahan's method as opposed to the direct measure by using labeled test compounds. All compounds antagonized orexin-A-induced inositol 1 phosphate (IP1) accumulation with the following pK(B) values: SB-649868 (OX(1)=9.67; OX(2)=9.64), ACT-078573 (OX(1)=8.44; OX(2)=9.02), JNJ-10397049 (OX(1)=5.97; OX(2)=8.35), MK-6096 (OX(1)=9.13; OX(2)=9.79) and Roche-Cp (OX(1)=7.18; OX(2)=8.83). They displaced the [(3)H]ACT-078573 receptor binding with the following pK(i) values: SB-649868 (OX(1)=9.27; OX(2)=8.91), ACT-078573 (OX(1)=7.80; OX(2)=9.12), JNJ-10397049 (OX(1)=5.18; OX(2)=8.10), MK-6096 (OX(1)=8.39; OX(2)=8.90) and Roche-Cp (OX(1)=6.65; OX(2)=8.54). From dissociation kinetic studies using [(3)H]ACT-078573, the calculated long half-life, (t(½)) supported the non-surmountability profile of SB-649868 (t(½)=35.91min) at OX(1) orexin receptor. Similarly, the long or moderately long t(½) values for ACT-078573 at OX(2) orexin receptor (t(½)=69.71min), MK-6096 (t(½)=17.70min), SB-649868 (t(½)=8.09min) and Roche-Cp (t(½)=5.79min) sustained their non-surmountable profile. JNJ-10397049 showed short t(½) values at both receptor subtypes (OX(1)t(½)=0.19min; OX(2)t(½)=0.60min) with surmountable antagonism.