(-)-Stepholidine is a potent pan-dopamine receptor antagonist of both G protein- and ß-arrestin-mediated signaling.

RATIONALE: (-)-Stepholidine is a tetrahydroberberine alkaloid that is known to interact with dopamine receptors and has also been proposed as a novel antipsychotic agent. Its suggested novelty lies in the fact that it has been proposed to have D1-like receptor agonist and D2-like receptor antagonist properties. Thus, it might be effective in treating both positive and negative (cognition) symptoms of schizophrenia. However, its activity on specific dopamine receptor subtypes has not been clarified, especially with respect to its ability to activate D1-like receptors. OBJECTIVES: We wished to examine the affinity and functional activity of (-)-stepholidine at each of the human dopamine receptor subtypes expressed in a defined cellular environment. METHODS: D1-D5 dopamine receptors were stably expressed in cell lines and their interactions with (-)-stepholidine were examined using radioligand binding and various functional signaling assays. Radioligand binding assays were also performed using bovine striatal membranes. RESULTS: (-)-Stepholidine exhibited high (nM) affinity for D1 and D5 receptors, somewhat lower (two- to four-fold) affinity for D2 and D3 receptors, and low micromolar affinity for D4 receptors. Functionally, (-)-stepholidine was ineffective in activating G protein-mediated signaling of D1-like and D2 receptors and was also ineffective in stimulating ß-arrestin recruitment to any dopamine receptor subtype. It did, however, antagonize all of these responses. It also antagonized D1-D2 heteromer-mediated Ca(2+) mobilization. Radioligand binding assays of D1-like receptors in brain membranes also indicated that (-)-stepholidine binds to the D1 receptor with antagonist-like properties. CONCLUSIONS: (-)-Stepholidine is a pan-dopamine receptor antagonist and its in vivo effects are largely mediated through dopamine receptor blockade with potential cross-talk to other receptors or signaling proteins.

Biphasic effect of melanocortin agonists on metabolic rate and body temperature.

The melanocortin system regulates metabolic homeostasis and inflammation. Melanocortin agonists have contradictorily been reported to both increase and decrease metabolic rate and body temperature. We find two distinct physiologic responses occurring at similar doses. Intraperitoneal administration of the nonselective melanocortin agonist MTII causes a melanocortin-4 receptor (Mc4r)-mediated hypermetabolism/hyperthermia. This is preceded by a profound, transient hypometabolism/hypothermia that is preserved in mice lacking any one of Mc1r, Mc3r, Mc4r, or Mc5r. Three other melanocortin agonists also caused hypothermia, which is actively achieved via seeking a cool environment, vasodilation, and inhibition of brown adipose tissue thermogenesis. These results suggest that the hypometabolic/hypothermic effect of MTII is not due to a failure of thermoregulation. The hypometabolism/hypothermia was prevented by dopamine antagonists, and MTII selectively activated arcuate nucleus dopaminergic neurons, suggesting that these neurons may contribute to the hypometabolism/hypothermia. We propose that the hypometabolism/hypothermia is a regulated response, potentially beneficial during extreme physiologic stress.

Development of a highly selective, orally bioavailable and CNS penetrant M1 agonist derived from the MLPCN probe ML071.

Herein we report the discovery and SAR of a novel series of M(1) agonists based on the MLPCN probe, ML071. From this, VU0364572 emerged as a potent, orally bioavailable and CNS penetrant M(1) agonist with high selectivity, clean ancillary pharmacology and enantiospecific activity.

Differential effects of allosteric M(1) muscarinic acetylcholine receptor agonists on receptor activation, arrestin 3 recruitment, and receptor downregulation.

Muscarinic acetylcholine receptors (mAChRs) are drug targets for multiple neurodegenerative and neuropsychiatric disorders, but the full therapeutic potential of mAChR-targeted drugs has not been realized, mainly because of a lack of subtype-selective agonists. Recent advances have allowed the development of highly selective agonists that bind to an allosteric site on the M(1) mAChR that is spatially distinct from the orthosteric acetylcholine binding site, but less is known about the profile of intracellular signals activated by orthosteric versus allosteric M(1) mAChR agonists. We investigated the activation and regulatory mechanisms of two structurally distinct allosteric M(1) mAChR agonists, AC260584 and TBPB. We show that allosteric agonists potently activate multiple signal transduction pathways linked to the M(1) mAChR receptor but, compared to orthosteric agonists, much less efficiently recruit arrestin 3, a protein involved in regulation of G-protein coupled receptor signaling. Consistent with decreased arrestin recruitment, both allosteric agonists showed blunted responses in measurements of receptor desensitization, internalization, and downregulation. These results advance the understanding of mAChR biology and may shed light on unanticipated differences in the pharmacology of orthosteric vs. allosteric agonists that might be capitalized upon for drug development for the treatment of CNS diseases.

Synthesis and SAR of N-(4-(4-alklylpiperazin-1-yl)phenyl)benzamides as muscarinic acetylcholine receptor subtype 1 (M1) anatgonists.

This Letter describes the synthesis and SAR, developed through an iterative analog library approach, of a novel series of selective M(1) mAChR antagonists, based on an N-(4-(4-alkylpiperazin-1-yl)phenyl)benzamide scaffold for the potential treatment of Parkinson's disease, dystonia and other movement disorders. Compounds in this series possess M(1) antagonist IC(50)s in the 350 nM to >10 microM range with varying degrees of functional selectivity versus M(2)-M(5).

Context-dependent pharmacology exhibited by negative allosteric modulators of metabotropic glutamate receptor 7.

Phenotypic studies of mice lacking metabotropic glutamate receptor subtype 7 (mGluR7) suggest that antagonists of this receptor may be promising for the treatment of central nervous system disorders such as anxiety and depression. Suzuki et al. (J Pharmacol Exp Ther 323:147-156, 2007) recently reported the in vitro characterization of a novel mGluR7 antagonist called 6-(4-methoxyphenyl)-5-methyl-3-(4-pyridinyl)-isoxazolo[ 4,5-c]pyridin-4(5H)-one (MMPIP), which noncompetitively inhibited the activity of orthosteric and allosteric agonists at mGluR7. We describe that MMPIP acts as a noncompetitive antagonist in calcium mobilization assays in cells coexpressing mGluR7 and the promiscuous G protein G alpha(15). Assessment of the activity of a small library of MMPIP-derived compounds using this assay reveals that, despite similar potencies, compounds exhibit differences in negative cooperativity for agonist-mediated calcium mobilization. Examination of the inhibitory activity of MMPIP and analogs using endogenous G(i/o)-coupled assay readouts indicates that the pharmacology of these ligands seems to be context-dependent, and MMPIP exhibits differences in negative cooperativity in certain cellular backgrounds. Electrophysiological studies reveal that, in contrast to the orthosteric antagonist (2S)-2-amino-2-[(1S,2S)-2-carboxyclycloprop-1-yl]-3-(xanth-9-yl) propanoic acid (LY341495), MMPIP is unable to block agonist-mediated responses at the Schaffer collateral-CA1 synapse, a location at which neurotransmission has been shown to be modulated by mGluR7 activity. Thus, MMPIP and related compounds differentially inhibit coupling of mGluR7 in different cellular backgrounds and may not antagonize the coupling of this receptor to native G(i/o) signaling pathways in all cellular contexts. The pharmacology of this compound represents a striking example of the potential for context-dependent blockade of receptor responses by negative allosteric modulators.

Synthesis and SAR of analogs of the M1 allosteric agonist TBPB. Part II: Amides, sulfonamides and ureas--the effect of capping the distal basic piperidine nitrogen.

This letter describes the further synthesis and SAR, developed through an iterative analog library approach, of analogs of the highly selective M1 allosteric agonist TBPB by deletion of the distal basic piperidine nitrogen by the formation of amides, sulfonamides and ureas. Despite the large change in basicity and topology, M1 selectivity was maintained.

Synthesis and SAR of analogues of the M1 allosteric agonist TBPB. Part I: Exploration of alternative benzyl and privileged structure moieties.

This Letter describes the first account of the synthesis and SAR, developed through an iterative analogue library approach, of analogues of the highly selective M1 allosteric agonist TBPB. With slight structural changes, mAChR selectivity was maintained, but the degree of partial M1 agonism varied considerably.

A new multi-gram synthetic route to labeling precursors for the D(2/3) PET agent 18F-fallypride.

This Letter describes a new multi-gram synthetic protocol for the preparation of the classic tosylate labeling precursor for the D(2/3) PET agent [(18)F]fallypride. In the course of our studies, we also discovered two novel labeling precusors, the previously undescribed mesylate and chloro congeners of fallypride.