Discovery of a small molecule RXFP3/4 agonist that increases food intake in rats upon acute central administration.

The relaxin family peptide receptors have been implicated in numerous physiological processes including energy homeostasis, cardiac function, wound healing, and reproductive function. Two family members, RXFP3 and RXFP4, are class A GPCRs with endogenous peptide ligands (relaxin-3 and insulin-like peptide 5 (INSL5), respectively). Polymorphisms in relaxin-3 and RXFP3 have been associated with obesity, diabetes, and hypercholesterolemia. Moreover, central administration of relaxin-3 in rats has been shown to increase food intake, leading to body weight gain. Reported RXFP3 and RXFP4 ligands have been restricted to peptides (both endogenous and synthetic) as well as a low molecular weight positive allosteric modulator requiring a non-endogenous orthosteric ligand. Described here is the discovery of the first potent low molecular weight dual agonists of RXFP3/4. The scaffold identified is competitive with a chimeric relaxin-3/INSL5 peptide for RXFP3 binding, elicits similar downstream signaling as relaxin-3, and increases food intake in rats following acute central administration. This is the first report of small molecule RXFP3/4 agonism.

Synthesis and biological evaluation of novel hybrids of highly potent and selective α4ß2-Nicotinic acetylcholine receptor (nAChR) partial agonists.

We previously reported the cyclopropylpyridine and isoxazolylpyridine ether scaffolds to be versatile building blocks for creating potent α4ß2 nicotinic acetylcholine receptor (nAChR) partial agonists with excellent selectivity over the α3ß4 subtype. In our continued efforts to develop therapeutic nicotinic ligands, seven novel hybrid compounds were rationally designed, synthesized, and evaluated in [3H]epibatidine binding competition studies. Incorporation of a cyclopropane- or isoxazole-containing side chain onto the 5-position of 1-(pyridin-3-yl)-1,4-diazepane or 2-(pyridin-3-yl)-2,5-diazabicyclo[2.2.1]heptane led to highly potent and selective α4ß2* nAChR partial agonists with Ki values of 0.5-51.4 nM for α4ß2 and negligible affinities for α3ß4 and α7. Moreover, compounds 21, 25, and 30 maintained the functional profiles (EC50 and IC50 values of 15-50 nM) of the parent azetidine-containing compounds 3 and 4 in the 86Rb+ ion flux assays. In vivo efficacy of the most promising compound 21 was confirmed in the mouse SmartCube® platform and classical forced swim tests, supporting the potential use of α4ß2 partial agonists for treatment of depression.

Characterization of maleimide-based glycogen synthase kinase-3 (GSK-3) inhibitors as stimulators of steroidogenesis.

Inhibition of GSK-3ß has been well documented to account for the behavioral actions of the mood stabilizer lithium in various animal models of mood disorders. Recent studies have showed that genetic or pharmacological inhibition of GSK-3ß resulted in anxiolytic-like and pro-social behavior. In our ongoing efforts to develop GSK-3ß inhibitors for the treatment of mood disorders, SAR studies on maleimide-based compounds were undertaken. We present herein for the first time that some of these GSK-3ß inhibitors, in particular analogues 1 and 9, were able to stimulate progesterone production in the MA-10 mouse tumor Leydig cell model of steroidogenesis without any significant toxicity. These two compounds were tested in the SmartCube behavioral assay and showed anxiolytic-like signatures following daily dose administration (50 mg/kg, ip) for 13 days. Taken together, these results support the hypothesis that GSK-3ß inhibition could influence neuroactive steroid production thereby mediating the modulation of anxiety-like behavior in vivo.

Discovery of highly potent and selective α4ß2-nicotinic acetylcholine receptor (nAChR) partial agonists containing an isoxazolylpyridine ether scaffold that demonstrate antidepressant-like activity. Part II.

In our continued efforts to develop α4ß2-nicotinic acetylcholine receptor (nAChR) partial agonists as novel antidepressants having a unique mechanism of action, structure-activity relationship (SAR) exploration of certain isoxazolylpyridine ethers is presented. In particular, modifications to both the azetidine ring present in the starting structure 4 and its metabolically liable hydroxyl side chain substituent have been explored to improve compound druggability. The pharmacological characterization of all new compounds has been carried out using [(3)H]epibatidine binding studies together with functional assays based on (86)Rb(+) ion flux measurements. We found that the deletion of the metabolically liable hydroxyl group or its replacement by a fluoromethyl group not only maintained potency and selectivity but also resulted in compounds showing antidepressant-like properties in the mouse forced swim test. These isoxazolylpyridine ethers appear to represent promising lead candidates in the design of innovative chemical tools containing reporter groups for imaging purposes and of possible therapeutics.

Insights into the structural determinants required for high-affinity binding of chiral cyclopropane-containing ligands to α4ß2-nicotinic acetylcholine receptors: an integrated approach to behaviorally active nicotinic ligands.

Structure-based drug design can potentially accelerate the development of new therapeutics. In this study, a cocrystal structure of the acetylcholine binding protein (AChBP) from Capitella teleta (Ct) in complex with a cyclopropane-containing selective α4ß2-nicotinic acetylcholine receptor (nAChR) partial agonist (compound 5) was acquired. The structural determinants required for ligand binding obtained from this AChBP X-ray structure were used to refine a previous model of the human α4ß2-nAChR, thus possibly providing a better understanding of the structure of the human receptor. To validate the potential application of the structure of the Ct-AChBP in the engineering of new α4ß2-nAChR ligands, homology modeling methods, combined with in silico ADME calculations, were used to design analogues of compound 5. The most promising compound, 12, exhibited an improved metabolic stability in comparison to the parent compound 5 while retaining favorable pharmacological parameters together with appropriate behavioral end points in the rodent studies.

Chemistry and behavioral studies identify chiral cyclopropanes as selective α4ß2-nicotinic acetylcholine receptor partial agonists exhibiting an antidepressant profile.

Despite their discovery in the early 20th century and intensive study over the last 20 years, nicotinic acetylcholine receptors (nAChRs) are still far from being well understood. Only a few chemical entities targeting nAChRs are currently undergoing clinical trials, and even fewer have reached the marketplace. In our efforts to discover novel and truly selective nAChR ligands, we designed and synthesized a series of chiral cyclopropane-containing α4ß2-specific ligands that display low nanomolar binding affinities and excellent subtype selectivity while acting as partial agonists at α4ß2-nAChRs. Their favorable antidepressant-like properties were demonstrated in the classical mouse forced swim test. Preliminary ADMET studies and broad screening toward other common neurotransmitter receptors were also carried out to further evaluate their safety profile and eliminate their potential off-target activity. These highly potent cyclopropane ligands possess superior subtype selectivity compared to other α4ß2-nAChR agonists reported to date, including the marketed drug varenicline, and therefore may fully satisfy the crucial prerequisite for avoiding adverse side effects. These novel chemical entities could potentially be advanced to the clinic as new drug candidates for treating depression.

Identification of novel α4ß2-nicotinic acetylcholine receptor (nAChR) agonists based on an isoxazole ether scaffold that demonstrate antidepressant-like activity.

There is considerable evidence to support the hypothesis that the blockade of nAChR is responsible for the antidepressant action of nicotinic ligands. The nicotinic acetylcholine receptor (nAChR) antagonist, mecamylamine, has been shown to be an effective add-on in patients that do not respond to selective serotonin reuptake inhibitors. This suggests that nAChR ligands may address an unmet clinical need by providing relief from depressive symptoms in refractory patients. In this study, a new series of nAChR ligands based on an isoxazole-ether scaffold have been designed and synthesized for binding and functional assays. Preliminary structure-activity relationship (SAR) efforts identified a lead compound 43, which possesses potent antidepressant-like activity (1 mg/kg, IP; 5 mg/kg, PO) in the classical mouse forced swim test. Early stage absorption, distribution, metabolism, excretion, and toxicity (ADME-Tox) studies also suggested favorable drug-like properties, and broad screening toward other common neurotransmitter receptors indicated that compound 43 is highly selective for nAChRs over the other 45 neurotransmitter receptors and transporters tested.

Discovery of isoxazole analogues of sazetidine-A as selective α4ß2-nicotinic acetylcholine receptor partial agonists for the treatment of depression.

Depression, a common neurological condition, is one of the leading causes of disability and suicide worldwide. Standard treatment, targeting monoamine transporters selective for the neurotransmitters serotonin and noradrenaline, is not able to help many patients that are poor responders. This study advances the development of sazetidine-A analogues that interact with α4ß2 nicotinic acetylcholine receptors (nAChRs) as partial agonists and that possess favorable antidepressant profiles. The resulting compounds that are highly selective for the α4ß2 subtype of nAChR over α3ß4-nAChRs are partial agonists at the α4ß2 subtype and have excellent antidepressant behavioral profiles as measured by the mouse forced swim test. Preliminary absorption, distribution, metabolism, excretion, and toxicity (ADMET) studies for one promising ligand revealed an excellent plasma protein binding (PPB) profile, low CYP450-related metabolism, and low cardiovascular toxicity, suggesting it is a promising lead as well as a drug candidate to be advanced through the drug discovery pipeline.

Dissociation between duration of action in the forced swim test in mice and nicotinic acetylcholine receptor occupancy with sazetidine, varenicline, and 5-I-A85380.

RATIONALE: Nicotinic acetylcholine receptor (nAChR) agonists, partial agonists, and antagonists have antidepressant-like effects in rodents and reduce symptoms of depression in humans. OBJECTIVES: The study determined whether the antidepressant-like effect of the nAChR ß2* partial agonist sazetidine-A (sazetidine) in the forced swim test was due to activation or desensitization of ß2* nAChRs. The study also determined if sazetidine's behavioral responses in the forced swim test corresponded to ß2* nAChRs receptor occupancy and drug bioavailability. RESULTS: Acute antidepressant-like effects in the forced swim test were seen with sazetidine and the full ß2* agonist 5-I-A8350 (BALB/cJ mice) and the less selective ß2* partial agonist varenicline in C57BL/6J but not BALB/cJ mice. The role of ß2* nAChRs was confirmed by results showing: (1) reversal of sazetidine's antidepressant-like effects in the forced swim test by nAChR antagonists mecamylamine and dihydro-ß-erythroidine; (2) absence of sazetidine's effect in mice lacking the ß2 subunit of the nAChR; and (3) a high correspondence between behaviorally active doses of sazetidine and ß2* receptor occupancy. ß2* receptor occupancy following acute sazetidine, varenicline, and 5-I-A8350 lasted beyond the duration of action in the forced swim test. Sazetidine's long lasting receptor occupancy did not diminish behavioral efficacy in the forced swim test following repeated dosing. CONCLUSIONS: Results demonstrate that activation of a small population of ß2* nAChRs (10-40%) is sufficient to elicit sazetidine's antidepressant-like actions without producing tolerance and suggest that ligands that activate ß2* nAChRs would be promising targets for the development of a new class of antidepressant.

Psychostimulant-like discriminative stimulus and locomotor sensitization properties of the wake-promoting agent modafinil in rodents.

UNLABELLED: The present studies assessed the potential abuse liability and likely mechanism(s) of action of the wake-promoting agent modafinil. METHODS: Experiments assessed the locomotor sensitization (LS) and discriminative stimulus (DS) properties of modafinil in mouse and rat, respectively. Comparative data were generated with a range of psychostimulants and monoamine reuptake inhibitors. RESULTS: Repeated administration of d-amphetamine and cocaine, psychostimulants with high abuse liability, resulted in the induction and expression of LS in mice. Bupropion and caffeine, two psychostimulants not abused in humans, were not associated with LS. GBR12909 induced LS during repeated exposure, but there was no evidence of expression of LS after acute challenge following withdrawal. In contrast, repeated administration of modafinil resulted in the expression, but not induction, of LS. d-amphetamine, but not the mu-opioid agonist morphine or the nAChR agonist nicotine, fully substituted for the cocaine DS in rats. The selective dopamine transporter (DAT) inhibitor GBR12909 fully substituted, the preferential norepinephrine transporter (NET) inhibitor desipramine partially substituted, and the selective serotonin reuptake inhibitor citalopram failed to substitute for cocaine. Modafinil fully substituted for cocaine, similar to the mixed DAT/NET inhibitor bupropion. CONCLUSIONS: Two preclinical assays indicated potential abuse liability of modafinil; drug discrimination studies suggest DAT blockade by modafinil is a likely mechanism of action in vivo.

Structure-based design leads to the identification of lithium mimetics that block mania-like effects in rodents. possible new GSK-3beta therapies for bipolar disorders.

More than two million American adults, or approximately one percent of the population 18 years or older, suffer from bipolar disorder. Current treatments include the so-called "mood stabilizers," lithium and valproic acid. Both are relatively dated drugs that are only partially effective and produce various undesirable side effects including weight gain. Based upon continued efforts to understand the molecular target for lithium, it now appears that specific inhibitors of the enzyme glycogen synthase kinase-3beta (GSK-3beta) may mimic the therapeutic action of mood stabilizers and might therefore allow for the design of improved drugs for treating patients with bipolar disorder as well as certain neurodegenerative disorders. Furthermore, the pro-apoptotic properties of the GSK-3 enzyme suggest the possible use of such inhibitors as neuroprotective agents. In fact, neuroprotection may contribute to the treatment of mood disorders. The present chemistry, modeling, and biology efforts have identified 3-benzofuranyl-4-indolylmaleimides as potent and relatively selective GSK-3beta inhibitors. The best ligand in this series (having a Ki value of 4.6 nM against GSK-3beta) was studied in a novel mouse model of mania that has recently been validated with several clinically effective mood stabilizers. This study presents the first demonstration of the efficacy of a GSK-3beta inhibitor in this mouse model of mania. Selective brain penetrable GSK-3 ligands like those described herein become valuable research tools in better defining the role of this multifaceted kinase in both physiological and pathophysiological events.