N 1 -(3-Cyclohexylbutanoyl)-N 2 -[3-(1H-imidazol-4- yl)propyl]guanidine (UR-AK57), a Potent Partial Agonist for the Human Histamine H 1 - and H 2 - Receptors Sheng-Xue Xie, Anja Kraus, Prasanta Ghorai, Qi-Zhuang Ye, Sigurd Elz, Armin Buschauer, and Roland Seifert High-Throughput Screening Laboratory (S.-X.X., Q.-Z.Y.) and Department of Pharmacology and Toxicology (R.S.), The University of Kansas, Lawrence, Kansas; and Department of Medicinal Chemistry II (A.K., P.G., A.B.), Department of Medicinal Chemistry I (S.E.), and Department of Pharmacology and Toxicology (R.S.), University of Regensburg, Regensburg, Germany Received February 13, 2006; accepted March 17, 2006 ABSTRACT Both the histamine H 1 -receptor (H 1 R) and H 2 -receptor (H 2 R) exhibit pronounced species selectivity in their pharmacological properties; i.e., bulky agonists possess higher potencies and efficacies at guinea pig (gp) than at the corresponding human (h) receptor isoforms. In this study, we examined the effects of N G -acylated imidazolylpropylguanidines substituted with a sin- gle phenyl or cyclohexyl substituent on H 1 R and H 2 R spe- cies isoforms expressed in Sf9 insect cells. N 1 -(3-Cyclohexyl- butanoyl)-N 2 -[3-(1H-imidazol-4-yl)propyl]guanidine (UR-AK57) turned out to be the most potent hH 2 R agonist identified so far (EC 50 of 23 nM in the GTPase assay at the hH 2 R-G s fusion protein expressed in Sf9 insect cells). UR-AK57 was almost a full-hH 2 R agonist and only slightly less potent and efficacious than at gpH 2 R-G s . Several N G -acylated imidazolylpropylgua- nidines showed similar potency at hH 2 R and gpH 2 R. Most unexpectedly, UR-AK57 exhibited moderately strong partial hH 1 R agonism with a potency similar to that of histamine, whereas at gpH 1 R, UR-AK57 was only a very weak partial agonist. Structure/activity relationship studies revealed that both the alkanoyl chain connecting the aromatic or alicyclic substituent with the guanidine moiety and the nature of the carbocycle (cyclohexyl versus phenyl ring) critically determine the pharmacological properties of this class of compounds. Collectively, our data show that gpH 1 R and gpH 2 R do not necessarily exhibit preference for bulky agonists compared with hH 1 R and hH 2 R, respectively, and that UR-AK57 is a promising starting point for the development of both potent and efficacious hH 1 R and hH 2 R agonists. Histamine (HIS) (1) (see Fig. 1) is a neurotransmitter and autacoid and acts through H 1 -, H 2 -, H 3 -, and H 4 -receptors (H 1 R–H 4 R) (Hill et al., 1997; Hough, 2001; Bakker et al., 2002). The H 1 R couples to G q -proteins to mediate phospho- lipase C activation and plays a role in the regulation of alertness and as mediator of type 1 allergic reactions (Hill et al., 1997; Bakker et al., 2002). The H 2 R couples to G s -pro- teins to mediate adenylyl cyclase activation and regulates H + secretion in gastric parietal cells, cardiac contractility, and various myeloid cell functions (Klinker et al., 1996; Hill et al., 1997; Bakker et al., 2002). It has been difficult to establish relevant native test sys- tems for the analysis of the human H 1 R (hH 1 R) and human H 2 R (hH 2 R), because there are unexplained pharmacological differences in the properties of hH 1 R and hH 2 R in native cells relative to standard guinea pig test organs (Burde et al., 1990; Seifert et al., 1994; Klinker et al., 1996). To facilitate the comparison of histamine receptors under identical exper- imental conditions, we established expression systems for the H 1 R and H 2 R in Sf9 insect cells (Kelley et al., 2001; Houston et al., 2002). Sf9 cells express the H 1 R and H 2 R at high levels and can be cultured in large quantities. GPCR/G- protein coupling in Sf9 membranes is monitored with high sensitivity using the steady-state GTPase assay. This assay This work was supported by the National Institutes of Health COBRE Award 1 P20 RR15563 and matching support from the State of Kansas and the University of Kansas (to R.S. and Q.-Z.Y.) and the Graduate Training Program (Graduiertenkolleg GRK 760, “Medicinal Chemistry: Molecular Recognition— Ligand-Receptor Interactions”) of the Deutsche Forschungsgemeinschaft (to R.S., S.E., and A.B.). Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. doi:10.1124/jpet.106.102897. ABBREVIATIONS: HIS, histamine; UR-AK57, N 1 -(3-cyclohexylbutanoyl)-N 2 -[3-(1H-imidazol-4-yl)propyl]guanidine; AIPG, N G -acylated imidazolyl- propylguanidine; GPCR, G-protein-coupled receptor; gpH 1 R, guinea pig histamine H 1 -receptor; gpH 2 R, guinea pig histamine H 2 -receptor; gpH 2 R-G sS , fusion protein of the guinea pig histamine H 2 -receptor and the short splice variant of G s ; hH 1 R, human histamine H 1 -receptor; hH 2 R, human histamine H 2 -receptor; hH 2 R-G sS , fusion protein of the human histamine H 2 -receptor and the short splice variant of G s ; HIS, histamine; RGS, regulator of G-protein signaling; TM, transmembrane domain. 0022-3565/06/3173-1262–1268$20.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 317, No. 3 Copyright © 2006 by The American Society for Pharmacology and Experimental Therapeutics 102897/3117058 JPET 317:1262–1268, 2006 Printed in U.S.A. 1262 at ASPET Journals on February 2, 2016 jpet.aspetjournals.org Downloaded from