A Chemical Switch for the Modulation of the Functional Activity of Higher Homologues of Histamine on the Human Histamine H 3 Receptor: Effect of Various Substitutions at the Primary Amino Function Marinella Govoni, Herman D. Lim, Dris El-Atmioui, Wiro M. P. B. Menge, Henk Timmerman, Remko A. Bakker, Rob Leurs, and Iwan J. P. De Esch* Leiden/Amsterdam Center for Drug Research, Department of Pharmacochemistry, Vrije UniVersiteit Amsterdam, De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands ReceiVed May 8, 2005 In an effort to establish the structural requirements for agonism, neutral antagonism, and inverse agonism at the human histamine H 3 receptor (H 3 R) we have prepared a series of higher homologues of histamine in which the terminal nitrogen of the side chain has been either mono- or disubstituted with several aliphatic, alicyclic, and aromatic moieties or incorporated in cyclic systems. The novel ligands have been pharmacologically investigated in vitro for their affinities on the human H 3 R and H 4 R subtypes by radioligand displacement experiments and for their intrinsic H 3 R activities via a CRE-mediated -galactosidase reporter gene assay. Subtle changes of the substitution pattern at the side chain nitrogen alter enormously the pharmacological activity of the ligands, resulting in a series of compounds with a wide spectrum of pharmacological activities. Among the several neutral H 3 R antagonists identified within this series, compounds 2b and 2h display an H 3 R affinity in the low nanomolar concentration range (pK i values of 8.1 and 8.4, respectively). A very potent and selective H 3 R agonist (1l, pEC 50 ) 8.9, R) 0.94) and a very potent, though not highly selective, H 3 R inverse agonist (2k, pIC 50 ) 8.9, R)-0.97) have been identified as well. Introduction The biogenic amine histamine exerts its multiple biological activities through the activation of at least four distinct histamine receptors, i.e., H 1 ,H 2 ,H 3 , and H 4 receptors, 1-4 all belonging to the superfamily of heptahelical (7TM) G-protein coupled receptors. The histamine H 3 receptor (H 3 R) was discovered in 1983 by Arrang and co-workers as a presynaptic autoreceptor, 5 and the gene was successively cloned in 1999 by Lovenberg and colleagues. 3 Tissue distribution analysis indicated that the expression of the receptor is predominantly restricted to the brain. 6 The H 3 R does not only mediate the inhibition of synthesis and release of histamine from histaminergic neurons via a negative feedback loop, 7,8 but also exerts modulatory effects on other neurotransmitter systems, e.g. the cholinergic, 9,10 dopaminergic, 11 noradrenergic, 12 and serotoninergic 13 systems, in both the central and peripheral nervous systems. It has been recognized that the H 3 R is a potential therapeutic target. 14-16 H 3 R antagonists have been proposed to be potential drugs for the treatment of several CNS disorders, such as attention-deficit hyperactivity disorder (ADHD), 17,18 Alzheimer’s disease, 19 epilepsy, 20-22 schizophrenia, 22,23 and obesity, 24-26 whereas the therapeutic potential of H 3 R agonists has been shown for myocardial ischemia, 27 inflammatory 28 and gastric acid related diseases, 29 migraine, and sleep disorders. 30 The constitutive activity of several 7TM receptor systems has been investigated within the past decade. 31-36 For a long time it was not known whether this phenomenon existed only in cells overexpressing (mutant) GPCRs or also occurred in vivo. Constitutive activity has been shown recently for both the histamine H 1 and H 2 receptors. 37,38 Our group reported that the therapeutically important H 1 and H 2 receptor antagonists act, in fact, as inverse agonists, but we also identified neutral antagonists for both receptors. 37,39 Histaminergic neurotrans- mission in rodent brain was shown to be regulated by consti- tutively active H 3 Rs in vitro as well as in vivo, 40 and our group also demonstrated that the human and rat H 3 Rs stably expressed in SK-N-MC cells 3,6 show a high level of constitutive activ- ity. 41,42 This led to the identification of several standard H 3 R antagonists (thioperamide and clobenpropit) as inverse agonists in this cell system. Moreover, burimamide and impentamine (Figure 1), previously identified as H 3 R antagonists, 5,43,44 behave as H 3 R agonists at both recombinant H 3 Rs (human and rat). We also showed that in a small series of impentamine analogues we were able to manipulate the intrinsic activity (R) by substitution of the amino group. 41 In an attempt to establish a correlation between the structure of the ligands and their functional activity at the human H 3 R, * Author for correspondence. Tel: 31-20-597600. Fax: 31-20- 5987610. E-mail: i.de.esch@few.vu.nl. Figure 1. Structures of 4-[1H-imidazol-4-yl]butylamine (imbutamine, 1a), 5-[1H-imidazol-4-yl]pentylamine (impentamine, 2a), [5-(1H- imidazol-4-yl)pentyl]isopropylamine (2d), clobenpropit, GT-2331 (cipral- isant, Perceptin), and proxyfan. 2549 J. Med. Chem. 2006, 49, 2549-2557 10.1021/jm0504353 CCC: $33.50 © 2006 American Chemical Society Published on Web 03/22/2006