An 116-amino acid deletion in the third intracellular loop of a naturally occurring human histamine H3 receptor isoform confers inverse agonist pharmacological differences

Natasha Lethbridge1, Andrew Medhurst2, Paul Chazot1. 1Durham University, Durham, United Kingdom, 2GSK Neuroscience CEDD, Harlow, United Kingdom.

Brain histamine is involved in the regulation of numerous functions of the central nervous system (CNS), with growing evidence for the involvement of the histamine H3 receptor in arousal, cognition, locomotor activity, autonomic and vestibular functions, feeding and drinking, sexual behaviour, and analgesia. Moreover, H3R specific ligands display beneficial effects in pre-clinical animal models of obesity, epilepsy, and cognitive diseases such as Alzheimer’s disease and attention deficit hyperactivity disorder. As a result, H3R antagonists are currently undergoing clinical trials (reviewed in The Third Histamine Receptor Ed. D Vohora).

The aim of the present study is to pharmacologically characterize three common naturally occurring human histamine H3 receptor (hH3R) isoforms, hH3R (445), hH3R (365) and hH3R (329) transiently expressed in HEK 293 cells as previously described (van Rijn et al., 2006) using [3H] GSK189254, a new high affinity and selective H3 receptor inverse agonist (Medhurst et al., 2007). Expression of each isoform was confirmed immunologically. These abundantly expressed human splice variants differ by a deletion of 80 and 116 amino acids in the intracellular loop 3, respectively and display a differential expression pattern in human CNS.

In this report, we show the hH3R (329) to have a 5-fold lower affinity 0.98 +/- 0.4nM (n = 4) for [3H] GSK189254 than either the hH3R (445) 0.16 +/- 0.04nM (n = 4) or hH3R (365) 0.24 +/- 0.07nM (n = 4). Interestingly, co-transfection (cDNA ratio 1:1) of the hH3R (329) isoform had no effect on hH3R (445) affinity 0.27 +/- 0.07nM (n = 4), but reduced by approximately 10-fold, [3H] GSK189254 affinity for hH3R (365) 2.00 +/- 1.1nM (n = 5). Previous reports have shown an 80 amino acid deletion (hH3R (365)) in the intracellular loop 3 to confer higher affinity and potency for H3R agonists and conversely lower affinity and potency for H3R inverse agonists, when compared to the full length hH3R (445) isoform (Bonger et al., (2007).

These differences in H3R pharmacology displayed by GSK189254 for the major human isoforms may be of importance for a detailed understanding of the clinical efficacy of H3R ligands.

The Third Histamine Receptor (2009) (Ed. D Vohora) CRC Press.
Bongers, G. et al., (2007). J. Pharmacol. Exptl. Ther. 323: 888-98.
Medhurst , A. D. et al., (2007). J. Pharmacol. Exptl. Ther. 321: 1032-1045.
Van Rijn, R. M. et al., (2006) . Mol Pharmacol. 70: 604-15.
This work is supported by ESF COST BMO806 (HARR4-EuCOST). NL is a BBSRC/GSK CASE Postgraduate student.