Alda 1 synthesis Compared to the other histamine receptor su

Compared to the other Alda 1 synthesis receptor subtypes, the H3 receptor exhibits rather complex pharmacology, being subject to extensive splicing, leading to two major (H3-445 and H3-365) and possibly more functional human receptor isoforms; for a comprehensive review please see Leurs et al. (2005) and Bongers et al. (2007). The receptor is also reportedly to be constitutively active to various degrees, rendering mechanistic understanding of the pharmacology of “antagonists” as partial agonists, neutral antagonists and inverse agonists essential, as differential activity can yield different functional or “protean” outcomes (Gbahou et al., 2003; Krueger et al., 2005). Despite these challenges, multiple compounds have progressed into clinical development (exemplified in Fig. 1) for a range of target indications, including Alzheimer's disease, neuropathic pain, narcolepsy, and excessive daytime sleepiness. Pitolisant (also known as tiprolisant, ciproxidine or BF 2649) was the first H3 receptor antagonist to progress into clinical development, ultimately being approved for the treatment of narcolepsy (with further Phase III trial results pending; Abad and Guilleminault, 2017; Wozniak and Quinnell, 2015). Other agents, including ABT-239, GSK189254, S 38093-2, JNJ-5207852 and PF-3654746, progressed into either Phase I or Phase II trials but were discontinued due to a lack of efficacy, insufficient safety or a strategic withdrawal from neuroscience R&D (Kubo et al., 2015; Lazewska and Kiec-Kononowicz, 2018). Although each and every one of the agents listed above has been independently described in the literature by the originating laboratories, the extent to which their comparative receptor pharmacology might underlie their varied efficacy profile across different pre-clinical models and clinical populations remains relatively unexplored. Accordingly, we examined the pharmacology of the aforementioned investigational and approved histamine H3 receptor antagonists to compare their affinities, inverse agonist efficacies and any differential isoform activity at H3-445 and H3-365 utilising cAMP accumulation, GTPγ35S binding and intracellular Ca2+ mobilization in recombinant cells and/or rat striatal tissue. We show that all H3 receptor antagonists exhibit inverse agonist activity in membranes prepared from recombinant H3 receptor expressing CHO cells, and use functional kinetic assays (Riddy et al., 2015; Christopoulos et al., 1999; Mathiesen et al., 2006) to demonstrate that these compounds fall broadly into two distinct kinetic classes: “slow-off rate” antagonists (GSK189254, JNJ-5207852 and PF-3654746) and “fast-off rate” antagonists (ABT-239, pitolisant and S 38093-2), properties that do not consistently correlate with their equilibrium affinity. Histamine H3 antagonists including thioperamide and iodophenpropit display sub-micromolar affinity for other receptors, including the sigma-1 receptor (Leurs et al., 1995). Although not a G protein-coupled receptor, sigma-1 receptors are known to regulate ion channel function, including NMDA/GluN receptors and K+/Ca2+ channels (Cobos et al., 2008). Antagonists of this receptor are effective in models of neuropathic pain (Abadias et al., 2013; Zamanillo et al., 2013) and off target profiling of S 38093-2 revealed modest activity at the peripheral sigma-1 receptor. Due to this activity we hypothesized that part of the observed in vivo activity of H3 antagonists, including S 38093-2, may be due to a combination of both H3 and sigma-1 activities. Using in vitro [3H]-(+)-pentazocine binding and ex vivo receptor occupancy studies we demonstrate that S 38093-2 preferentially occupies sigma-1, but not histamine H3 receptors, at cognition-enhancing doses. Indeed, mouse brain histamine H3 versus sigma-1 affinity ratios also differentiated the other agents tested; GSK189254 and JNJ-5207852 display high histamine H3 receptor selectivity, whereas PF-3654746, ABT-239 and pitolisant have similar histamine H3 and sigma-1 receptor affinity, suggesting that the latter property could contribute to their in vivo efficacy.