The selective 5-HT 1A receptor agonist, NLX-112, overcomes tetrabenazine-induced catalepsy and depression-like behavior in the rat.

Tetrabenazine, a preferential inhibitor of the vesicular monoamine transporter type 2, depletes the brain monoamines dopamine, serotonin and norepinephrine. Tetrabenazine and deutetrabenazine (Austedo ®) are used to treat chorea associated with Huntington's disease. However, both compounds are known to aggravate Parkinsonism and depression observed in Huntington's disease patients. NLX-112 (a.k.a. befiradol/F13640) is a highly selective, potent and efficacious serotonin 5-HT 1A agonist. In animal models, it has robust efficacy in combating other iatrogenic motor disorders such as L-DOPA-induced dyskinesia and has marked antidepressant-like activity in rodent tests. In the present study, we investigated, in rats, the efficacy of NLX-112 to counteract tetrabenazine-induced catalepsy (a model of Parkinsonism) and tetrabenazine-induced potentiation of immobility in the forced swim test (FST, a model to detect antidepressant-like activity). The prototypical 5-HT 1A agonist, (±)8-OH-DPAT, and the 5-HT 1A partial agonist/dopamine D2 receptor blocker, buspirone, were used as comparators. Both NLX-112 and (±)8-OH-DPAT (0.16-2.5 mg/kg p.o. or s.c., respectively) abolished catalepsy induced by tetrabenazine (2 mg/kg i.p.). In comparison, buspirone (0.63-5.0 mg/kg p.o.) was ineffective and even tended to potentiate tetrabenazine-induced catalepsy at 0.63 mg/kg. In the FST, NLX-112 and (±)8-OH-DPAT (0.63 mg/kg) strongly reduced immobility when administered alone but also significantly opposed potentiation of immobility induced by tetrabenazine (1.5 mg/kg i.p.). Buspirone (0.63 and 2.5 mg/kg p.o.) had no effect by itself or against tetrabenazine. These results strongly suggest that selective and highly efficacious 5-HT 1A agonists, such as NLX-112, may be useful in combating tetrabenazine-induced Parkinsonism and/or depression in Huntington's disease patients.

Identification of the 5-HT1A serotonin receptor as a novel therapeutic target in a C. elegans model of Machado-Joseph disease.

Machado-Joseph disease (MJD) or Spinocerebellar ataxia type 3 (SCA3) is a progressive neurodegenerative disorder that affects movement coordination leading to a premature death. Despite several efforts, no disease-modifying treatment is yet available for this disease. Previous studies pinpointed the modulation of serotonergic signaling, through pharmacological inhibition of the serotonin transporter SERT, as a promising therapeutic approach for MJD/SCA3. Here, we describe the 5-HT1A receptor as a novel therapeutic target in MJD, using a C. elegans model of ATXN3 proteotoxicity. Chronic and acute administration of befiradol (also known as NLX-112), a highly specific 5-HT1A agonist, rescued motor function and suppressed mutant ATXN3 aggregation. This action required the 5-HT1A receptor orthologue in the nematode, SER-4. Tandospirone, a clinically tested 5-HT1A receptor partial agonist, showed a limited impact on animals' motor dysfunction on acute administration and a broader receptor activation profile upon chronic treatment, its effect depending on 5-HT1A but also on the 5-HT6/SER-5 and 5-HT7/SER-7 receptors. Our results support high potency and specificity of befiradol for activation of 5-HT1A/SER-4 receptors and highlight the contribution of the auto- and hetero-receptor function to the therapeutic outcome in this MJD model. Our study deepens the understanding of serotonergic signaling modulation in the suppression of ATXN3 proteotoxicity and suggests that a potent and selective 5-HT1A receptor agonist such as befiradol could constitute a promising therapeutic agent for MJD.

Effects of the Serotonin 5-HT1A Receptor Biased Agonists, F13714 and F15599, on Striatal Neurotransmitter Levels Following L-DOPA Administration in Hemi-Parkinsonian Rats.

Peak-dose dyskinesia is associated with the dramatic increase in striatal dopamine levels that follows L-DOPA administration. The 'false neurotransmitter' hypothesis postulates that the latter is likely due to an aberrant processing of L-DOPA by serotonergic neurons. In keeping with this hypothesis, two highly selective 'biased agonists' of 5-HT1A receptors-namely F13714 and F15599 (NLX-101)-were recently shown to exhibit exceptionally potent anti-dyskinetic activity without impairing L-DOPA therapeutic properties despite their differential targeting of 5-HT1A receptor sub-populations. In this study, we investigated whether these two compounds dampened peak L-DOPA-induced dopamine microdialysate levels in the striatum of hemi-parkinsonian rats. Acute administration of either F13714 (0.04 and 0.16 mg/kg i.p.) or F15599 (0.16 and 0.64 mg/kg, i.p.) blunted L-DOPA (2 mg/kg)-induced increases in dopamine microdialysate levels in the denervated striatum (following unilateral injection of 6-OHDA into the medial forebrain bundle). No significant changes were observed on the intact side of the brain. Concurrently, both drugs profoundly reduced striatal serotonin levels on both sides of the brain. In addition, F13714 and F15599, in the presence of L-DOPA, produced a dose-dependent increase in glutamate levels, but this effect was restricted to later time points. These finding support the interpretation that F13714 and F15599 mediate their anti-dyskinetic effects by blunting of the peak in dopamine levels via activation of somatodendritic serotonin 5-HT1A receptors and the consequent inhibition of serotonergic neurons. This study adds to the growing body of evidence supporting the development of a potent 5-HT1A receptor agonist for treatment of peak-dose dyskinesia.

Pharmacodynamic, pharmacokinetic and rat brain receptor occupancy profile of NLX-112, a highly selective 5-HT1A receptor biased agonist.

NLX-112 (i.e., F13640, befiradol) exhibits nanomolar affinity, exceptional selectivity and full agonist efficacy at serotonin 5-HT1A receptors. NLX-112 shows efficacy in rat, marmoset and macaque models of L-DOPA induced dyskinesia (LID) in Parkinson's disease and has shown clinical efficacy in a Phase 2a proof-of-concept study for this indication. Here we investigated, in rats, its pharmacodynamic, pharmacokinetic (PK) and brain 5-HT1A receptor occupancy profiles, and its PK properties in the absence and presence of L-DOPA. Total and free NLX-112 exposure in plasma, CSF and striatal ECF was dose-proportional over the range tested (0.04, 0.16 and 0.63 mg/kg i.p.). NLX-112 exposure increased rapidly (Tmax 0.25-0.5h) and exhibited approximately threefold longer half-life in brain than in plasma (1.1 and 3.6h, respectively). At a pharmacologically relevant dose of 0.16 mg/kg i.p., previously shown to elicit anti-LID activity in parkinsonian rats, brain concentration of NLX-112 was 51-63 ng/g from 0.15 to 1h. In microPET imaging experiments, NLX-112 showed dose-dependent reduction of 18F-F13640 (i.e., 18F-NLX-112) brain 5-HT1A receptor labeling in cingulate cortex and striatum, regions associated with motor control and mood, with almost complete inhibition of labeling at the dose of 0.63 mg/kg i.p.. Co-administration of L-DOPA (6 mg/kg s.c., a dose used to elicit LID in parkinsonian rats) together with NLX-112 (0.16 mg/kg i.p.) did not modify PK parameters in rat plasma and brain of either NLX-112 or L-DOPA. Here, we demonstrate that NLX-112's profile is compatible with 'druggable' parameters for CNS indications, and the results provide measures of brain concentrations and 5-HT1A receptor binding parameters relevant to the anti-dyskinetic activity of the compound.

Pharmacodynamic effects of a D-amino acid oxidase inhibitor indicate a spinal site of action in rat models of neuropathic pain.

Inhibition of d-amino acid oxidase (DAAO) activity is a potential target for the treatment of chronic pain. Here we characterized the effects of systemic administration of the DAAO inhibitor 4H-furo[3,2-b]pyrrole-5-carboxylic acid (SUN) in rat models of neuropathic and inflammatory pain. Oral administration of SUN dose dependently attenuated tactile allodynia induced by ligation of the L5 spinal nerve (SNL) and similarly reversed thermal hyperalgesia produced by chronic constriction injury. In addition, SUN was efficacious against complete Freund's adjuvant-induced thermal hyperalgesia. In these models, maximal reversal of pain-related behaviors corresponded with maximum rates of increase in brain and plasma d-serine concentrations, indicative of full inhibition of DAAO activity. To investigate the possible site(s) of action, we recorded spontaneous nerve activity and mechanically evoked responses of central spinal cord dorsal horn neurons and compared these with spontaneous activity of peripheral dorsal root filaments in anesthetized SNL model animals. Oral SUN reduced spontaneous activity in both central and peripheral recordings at doses and pretreatment times that corresponded to reduced mechanical allodynia in behavioral experiments. After intravenous administration of SUN, the onset of action for this central effect was rapid (maximal effects within 30 minutes), but was abolished by severing afferent inputs to the dorsal horn. Overall, these results indicate that inhibition of DAAO in peripheral afferent spinal circuits reduced spontaneous neuronal activity to attenuate pain-related behaviors in rat models of neuropathic and inflammatory pain.

Dissecting the contribution of 5-HT1A auto- and heteroreceptors in sucrose overconsumption in mice.

The rise in obesity prevalence has been linked to overconsumption of high-sugar containing food and beverages. Recent evidence suggests that chronic sucrose consumption leads to changes in serotonergic neuroplasticity within the neural circuits involved in feeding control. Although there is a relationship between serotonin signalling in the brain and diet-induced obesity, the specific serotonin (5-HT) receptors or pathways involved remain unknown. The 5-HT1A receptor subtype plays a role in regulating mood, anxiety, and appetite, and has been associated with reversing addiction to substances of abuse. However, the respective role of 5-HT1A auto- vs heteroreceptors in sucrose consumption has not been examined. Mice were given controlled access to either 5%, 10% or 25% w/v sucrose, or water as a control, for 12 weeks using the well-established "drinking in the dark" protocol (n = 6-8 mice per group). Ligands selectively targeting 5-HT1A auto- and/or heteroreceptors (NLX-112, unbiased 5-HT1A receptor agonist; NLX-101, preferential heteroreceptor agonist; F13714, preferential autoreceptor agonist) were administered i.p. acutely after 6 and 12 weeks of sucrose consumption. The specific involvement of 5-HT1A receptors in these effects was verified by blockade with the selective 5-HT1A receptors antagonist WAY-100,635. The specific subpopulation of 5-HT1A receptors involved in sucrose consumption was dependent on the concentration of sucrose solution and the duration of exposure to sucrose (6 weeks vs 12 weeks). Long-term sucrose consumption leads to accentuated 5-HT1A autoreceptor function. Thus, targeting 5-HT1A autoreceptors might represent an effective therapeutic strategy to combat the rise in obesity resulting from the overconsumption of high-sugar diet.

Discovery of 1-(3,4-dichlorophenyl)-N,N-dimethyl-1,2,3,4-tetrahydroquinolin-4-amine, a dual serotonin and dopamine reuptake inhibitor.

The present work describes a series of novel tetrahydroquinoline amines that potently inhibit the in vitro reuptake of serotonin and dopamine (dual reuptake inhibitors). The compounds are structurally related to a series we disclosed previously, but are improved with respect to cytochrome P-450 enzyme (CYP) and potassium ion channel Kv11.1 (hERG) inhibition and synthetic accessibility. The detailed synthesis and in vitro activity and ADME profile of the compounds is described, which represent a previously undisclosed dual reuptake inhibitor chemotype.

Benzobistriazinones and related heterocyclic ring systems as potent, orally bioavailable positive allosteric AMPA receptor modulators.

AMPA receptors (AMPARs) are an important therapeutic target in the CNS. A series of substituted benzobistriazinone, benzobispyrimidinone and related derivatives have been prepared with high potency and selectivity for the allosteric binding site of AMPARs. Further improvements have been made to previously reported series of positive AMPAR modulators and these compounds exhibit excellent in vivo activity and improved in vivo metabolic stability with up to 100% oral bioavailability in rat.

Discovery of N-methyl-1-(1-phenylcyclohexyl)methanamine, a novel triple serotonin, norepinephrine, and dopamine reuptake inhibitor.

The current work discloses a novel cyclohexylarylamine chemotype with potent inhibition of the serotonin, norepinephrine, and dopamine transporters and potential for treatment of major depressive disorder. Optimized compounds 1 (SERT, NET, DAT, IC(50)=169, 85, 21 nM) and 42 (SERT, NET, DAT IC(50)=34, 295, 90 nM) were highly brain penetrant, active in vivo in the mouse tail suspension test at 30 mpk po and were not general motor stimulants.

Discovery of N-methyl-1-(1-phenylcyclohexyl)ethanamine, a novel triple serotonin, norepinephrine and dopamine reuptake inhibitor.

Novel chiral cyclohexylaryl amines were developed with potent reuptake inhibition against the serotonin, norepinephrine and dopamine transporters and activity at 10 and 30 mpk PO in the mouse tail suspension test. Prototype compound 31 (SERT, NET, DAT IC(50) ≤ 1, 21, 28 nM) was highly brain penetrant, had minimal CYP and hERG inhibition, and represents a previously undisclosed architecture with potential for treatment of major depressive disorder.

Synthesis and pharmacological evaluation of 4-(3,4-dichlorophenyl)-N-methyl-1,2,3,4-tetrahydronaphthalenyl amines as triple reuptake inhibitors.

The present work describes a series of novel chiral amines that potently inhibit the in vitro reuptake of serotonin, norepinephrine and dopamine (triple reuptake inhibitors) and were active in vivo in a mouse model predictive of antidepressant like activity. The detailed synthesis and in vitro activity and ADME profile of compounds is described, which represent a previously undisclosed triple reuptake inhibitor chemotype.

The selective 5-HT1A receptor agonist, NLX-112, exerts anti-dyskinetic and anti-parkinsonian-like effects in MPTP-treated marmosets.

l-DOPA is the gold-standard pharmacotherapy for treatment of Parkinson's disease (PD) but can lead to the appearance of troubling dyskinesia which are attributable to 'false neurotransmitter' release of dopamine by serotonergic neurons. Reducing the activity of these neurons diminishes l-DOPA-induced dyskinesia (LID), but there are currently no clinically approved selective, high efficacy 5-HT1A receptor agonists. Here we describe the effects of NLX-112, a highly selective and efficacious 5-HT1A receptor agonist, on LID in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated marmosets, a non-human primate model of PD. NLX-112 exhibited modest plasma half-life (~2h) and marked plasma protein binding (96%). When administered to parkinsonian marmosets with l-DOPA (7 mg/kg p.o.), NLX-112 (0.025, 0.1 and 0.4 mg/kg p.o.) reduced LID scores at early time-points after administration, whilst only minimally interfering with the l-DOPA-induced reversal of motor disability. In contrast, the prototypical 5-HT1A receptor agonist, (+)8-OH-DPAT (0.6 and 2 mg/kg p. o.), reduced LID but also abolished l-DOPA's anti-disability activity. Administered by itself, NLX-112 (0.1, 0.2 mg/kg p.o.) produced very little dyskinesia or locomotor activity, but reduced motor disability scores by about half the extent elicited by l-DOPA, suggesting that it may have motor facilitation effects of its own. Both NLX-112 and (+)8-OH-DPAT induced unusual and dose-limiting behaviors in marmoset that resembled 'serotonin behavioral syndrome' observed previously in rat. Overall, the present study showed that NLX-112 has anti-LID activity at the doses tested as well as reducing motor disability. The data suggest that additional investigation of NLX-112 is desirable to explore its potential as a treatment for PD and PD-LID.

In vivo saturation binding of GABA-A receptor ligands to estimate receptor occupancy using liquid chromatography/tandem mass spectrometry.

Typically, the dose-occupancy curves for GABA-A receptor ligands are determined using in vivo binding of [3H]flumazenil. This study describes in vivo binding experiments without the use of tracer ligands. Bound and free fractions were measured directly using a highly sensitive LC/MS/MS detection method after in vivo administration of the GABA-A ligands zolpidem, (RS)-zopiclone, L-838417 and flumazenil, to demonstrate affinity and saturation of the filter-retained, membrane-bound fraction. The in vivo binding of flumazenil and L-838417 both saturated around 200 nM, at a similar level to the specific binding of (S)-zopiclone after doses of the racemic zopiclone, using (R)-zopiclone to estimate non-specific binding. This saturable component represented an estimate of benzodiazepine binding sites available on GABA-A receptors in vivo (200 nM). Dose-occupancy curves were constructed to estimate the dose required to achieve 50% occupancy and matched estimates obtained with tracer methods. In contrast to tracer methods, this method is uniquely suitable to the demonstration of stereoselective binding of the (S)-isomer in vivo after doses of racemic zopiclone. These results demonstrate that the LC/MS/MS measurements of total drug concentrations typically used in early drug development can be adapted to provide information about receptor occupancy in vivo.

Serotonin 5-HT1A Receptor Biased Agonists Display Differential Anxiolytic Activity in a Rat Social Interaction Model.

When placed in an unfamiliar and brightly lit open-field, two adult male rats that have not previously interacted display a low level of social interaction (SI) attributed to an anxiety-like state. The SI test has therefore been used to explore anxiolytic/antistress activity. Here, we investigated the effects of serotonin 5-HT1A receptor agonists displaying various activity profiles, i.e. partial vs full agonist efficacy and pre- versus postsynaptic 5-HT1A receptor preferential activation by "biased agonists". Adult male Sprague-Dawley rats were housed singly before starting the social interaction session. At 30 min before being placed in an open-field, both rats of the dyad were injected (i.p or s.c.) with either vehicle, diazepam (as a reference compound), or one of six 5-HT1A receptor agonists: NLX-101 (a.k.a. F15599), F13714, S15535, flesinoxan, 8-OH-DPAT, and buspirone. Time spent in SI (following, sniffing, playing) was recorded for 10 min. Time spent in SI was inversely correlated with light intensity, with values dropping nearly by half (212.6 ± 18.8 vs 113.7 ± 7.0 s) between 10 and 300 lx (measured at floor level). Under the high light intensity conditions (300 lx), diazepam showed a bell-shaped curve, significantly increasing SI (78% increase in interaction time above control) at 1 mg/kg i.p. only. In the case of 5-HT1A receptor ligands, full agonists, whether nonpreferential (flesinoxan, (±)8-OH-DPAT) or preferential for presynaptic receptors (F13714), showed the strongest activity in this model. The preferential presynaptic receptor partial agonist, S15535, was also active over a wide dose-range, although with lower efficacy than F13714. In contrast, NLX-101, a high-efficacy biased agonist that preferentially activates postsynaptic 5-HT1A receptors, exhibited little activity. The clinical anxiolytic, buspirone, showed a marked effect likely due to its partial agonist activity at 5-HT1A presynaptic receptors. These data support the hypothesis that enhancement of SI in this model is mediated by preferential agonist activation of presynaptic 5-HT1A receptors, and confirm previous studies using local microinjections of (±)8-OH-DPAT. They further support the utility of noninvasive administration of biased agonists for exploring the activity of 5-HT1A receptor subpopulations.

Cortical 5-hydroxytryptamine 1A receptor biased agonist, NLX-101, displays rapid-acting antidepressant-like properties in the rat chronic mild stress model.

BACKGROUND: NLX-101 (also known as F15599) is a highly selective and efficacious 'biased' agonist at cortical 5-hydroxytryptamine 1A (5-HT1A) heteroreceptors. In rodents, it possesses marked antidepressant-like activity, potently and completely abolishing immobility in the forced swim test (FST) with extended duration of action. METHODS: We investigated the antidepressant-like activity of NLX-101 using the rat chronic mild stress (CMS) model of depression, considered to have a higher translational potential than the FST, as it possesses construct, face and predictive validity. The effects of CMS and repeated NLX-101 treatment were tested using sucrose consumption (a measure of anhedonia), novel object recognition (NOR; a measure of working memory) and elevated plus maze (EPM; a measure of anxiety) tests. RESULTS: NLX-101 reversed the CMS-induced decrease of sucrose intake on day 1 of testing, with full reversal observed at the dose of 0.16 mg/kg and a less pronounced but still significant effect at 0.04 mg/kg, both given twice a day intraperitoneally. The effects of NLX-101 were maintained over the 2 week treatment period and persisted for four weeks following cessation of treatment. In the NOR test, both doses of NLX-101 rescued the deficit in discrimination index caused by CMS, without any effect on locomotor activity. However, NLX-101 had no effect on the reduction of open-arms entries produced by CMS in the EPM model. In control, non-stressed rats, NLX-101 produced non-significant effects in all three models. CONCLUSIONS: NLX-101 displayed efficacious activity in the CMS test, with more rapid (1 day) antidepressant-like effects than pharmacological compounds tested previously under the same experimental conditions. These observations suggest that biased agonist targeting of cortical 5-HT1A receptors constitutes a promising strategy to achieve rapid-acting and sustained antidepressant effects.

Enhanced aggressive phenotype of Tph2 knockout rats is associated with diminished 5-HT1A receptor sensitivity.

Brain serotonin (5-HT) plays a key role in aggressive behaviours and related psychopathologies, but its precise mechanism of action remains elusive. Genetic animal models may provide a tool to elucidate the relationship between aggression and serotonin. The present study showed that tryptophan hydroxylase 2 (Tph2) knockout (KO) rats, which exhibit profoundly diminished extracellular serotonin levels, display increased aggressiveness compared to their Tph2 wildtype (WT) counterparts. However, the level of aggression in Tph2 KO rats did not equal that of feral wild type Groningen (WTG) rats. To investigate whether enhanced 5-HT1A receptor functionality may be present in Tph2 KO rats, we tested the acute anti-aggressive potency of the highly selective 5-HT1A receptor full agonist NLX-112 (a.k.a. befiradol or F13640). Data show that compared to Tph2 WT and WTG rats, the NLX-112 dose-effect curve was shifted to the right in Tph2 KO animals. These results suggest that, unlike previous reports in Tph2 KO mice, Tph2 KO rats have a decreased 5-HT1A receptor sensitivity compared to both Tph2 WT and WTG animals.

Novel Aryloxyethyl Derivatives of 1-(1-Benzoylpiperidin-4-yl)methanamine as the Extracellular Regulated Kinases 1/2 (ERK1/2) Phosphorylation-Preferring Serotonin 5-HT1A Receptor-Biased Agonists with Robust Antidepressant-like Activity.

Novel 1-(1-benzoylpiperidin-4-yl)methanamine derivatives were designed as "biased agonists" of serotonin 5-HT1A receptors. The compounds were tested in signal transduction assays (ERK1/2 phosphorylation, cAMP inhibition, Ca2+ mobilization, and ß-arrestin recruitment) which identified ERK1/2 phosphorylation-preferring aryloxyethyl derivatives. The novel series showed high 5-HT1A receptor affinity, >1000-fold selectivity versus noradrenergic α1, dopamine D2, serotonin 5-HT2A, histamine H1, and muscarinic M1 receptors, and favorable druglike properties (CNS-MPO, Fsp3, LELP). The lead structure, (3-chloro-4-fluorophenyl)(4-fluoro-4-(((2-(pyridin-2-yloxy)ethyl)amino)methyl)piperidin-1-yl)methanone (17, NLX-204), displayed high selectivity in the SafetyScreen44 panel (including hERG channel), high solubility, metabolic stability, and Caco-2 penetration and did not block CYP3A4, CYP2D6 isoenzymes, or P-glycoprotein. Preliminary in vivo studies confirmed its promising pharmacokinetic profile. 17 also robustly stimulated ERK1/2 phosphorylation in rat cortex and showed highly potent (MED = 0.16 mg/kg) and efficacious antidepressant-like activity, totally eliminating immobility in the rat Porsolt test. These data suggest that the present 5-HT1A receptor-biased agonists could constitute promising antidepressant drug candidates.

Activity of Serotonin 5-HT1A Receptor Biased Agonists in Rat: Anxiolytic and Antidepressant-like properties.

Although serotonin 5-HT1A receptors constitute attractive therapeutic targets, there is a lack of potential clinical candidates that have a high degree of selectivity and full agonist efficacy. Recently, novel 5-HT1A receptor "biased agonists" F15599 (also known as NLX-101) and F13714 have been reported that exhibit distinctive properties for in vitro signaling, neurochemical, electrophysiological effects, and in brain imaging. The present study characterized their effects in rat models of anxiety (elevated plus-maze, EPM, and Vogel tests), in depressive-like behavior (forced swim test), and on the induction of the three serotonergic behaviors (forepaw treading, flat body posture, and lower lip retraction). The prototypical 5-HT1A receptor ligands (±)8-OH-DPAT and buspirone were tested as comparators. In the elevated plus-maze, F15599, F13714, and (±)8-OH-DPAT dose-dependently increased the amount and percentage of time spent in the open arms with minimal effective doses (MED) of 5 mg/kg p.o., 2.5 mg/kg p.o. and 1.25 mg/kg s.c., respectively. The effects of the three agonists were abolished by pretreatment with the selective 5-HT1A receptor antagonist, WAY100635 (0.63 mg/kg s.c.). Buspirone did not show significant activity in the EPM. In contrast, in the Vogel test only buspirone was active, significantly increasing the number of licks and shocks accepted (active dose: 1.25 mg/kg s.c.). However, WAY100635 failed to reverse the effects of buspirone in this test, suggesting that they were not 5-HT1A receptor-mediated. In the forced swim test, F15599, F13714, and (±)8-OH-DPAT were potently active, abolishing immobility (MED: 0.63 mg/kg p.o., 0.63 mg/kg p.o. and 0.16 mg/kg s.c., respectively). Buspirone was not active. In measures of serotonergic behavior, F13714 and (±)8-OH-DPAT robustly elicited all three signs of serotonergic behaviors, whereas F15599 and buspirone elicited only lower-lip retraction. Taken together, these observations highlight the distinct profiles of activity of 5-HT1A agonists and suggest that the novel biased agonist F15599 combines pronounced activity in a test of anxiety (elevated plus-maze) with potent antidepressant-like effects and low propensity to induce serotonergic behaviors. These data suggest that selective biased agonists could constitute promising pharmacotherapeutics for mood disorders.

Antinociceptive, antiallodynic and antihyperalgesic effects of the 5-HT1A receptor selective agonist, NLX-112 in mouse models of pain.

BACKGROUND AND PURPOSE: NLX-112 (a.k.a. befiradol, F13640) is a drug candidate intended for the treatment of l-DOPA-induced dyskinesia. It is a highly selective serotonin 5-HT1A receptor full agonist which has been previously tested in a variety of models of CNS effects including analgesic activity in rat. Its activity in mouse models of pain has not been previously investigated. EXPERIMENTAL APPROACH: The activity of NLX-112 was tested in mouse models of acute pain (hot plate), tonic pain (intraplantar formalin test), in the oxaliplatin-induced neuropathic pain model of chemotherapy-induced peripheral neuropathy and in the streptozotocin (STZ)-induced model of painful diabetic neuropathy. KEY RESULTS: The main findings indicate that (i) NLX-112 was markedly active in the formalin test with potent reduction of paw licking in both phases of the test (minimal effective dose (MED) 0.5 mg/kg i.p. and p.o. in acute phase, and 0.1 mg/kg i.p. and 1 mg/kg p.o. in late phase). The effects of NLX-112 in this test were completely abolished by the selective 5-HT1A receptor antagonist, WAY100635; (ii) NLX-112 was active in the hot plate test and in the oxaliplatin-induced neuropathic pain model of chemotherapy-induced peripheral neuropathy, but at markedly higher doses (MED 2.5 mg/kg i.p.); (iii) NLX-112 was least active in the STZ-induced model of painful diabetic neuropathy (MED 5 mg/kg i.p.); (iv) NLX-112 did not affect locomotor activity. CONCLUSIONS AND IMPLICATIONS: NLX-112 may have significant potential for treatment of tonic pain but may be less promising as a candidate for treatment of chemotherapy-induced or diabetic neuropathic pain.

Distinctive in vitro signal transduction profile of NLX-112, a potent and efficacious serotonin 5-HT1A receptor agonist.

OBJECTIVES: NLX-112 (befiradol, F13640) is a selective serotonin 5-HT1A receptor agonist. Although it has been tested in vivo, little has been reported on its in vitro signal transduction profile. METHODS: NLX-112 was tested on G-protein activation, inhibition of adenylyl cyclase, ERK1/2 phosphorylation (pERK) and receptor internalization in recombinant cell lines. NLX-112 was also tested on G-protein activation in rat hippocampal membranes. Gα subunit mRNA expression in cell lines and rat brain tissue was quantified by quantitative PCR. KEY FINDINGS: For all signalling measures, NLX-112 exhibited agonist efficacy greater than for reference compounds ((±)8-OH-DPAT or buspirone), but similar to the endogenous agonist, serotonin, and was more potent for pERK than other responses. In rat hippocampal membranes, NLX-112 stimulated 'total G-proteins' but, unlike (±)8-OH-DPAT and buspirone, was more potent for Gαo activation. Cell lines predominantly expressed Gαi1 and Gαi2 mRNA, with low levels of Gαo, whereas in rat brain Gαo subunits showed highest mRNA expression. CONCLUSIONS: Unlike reference compounds, NLX-112 was a highly efficacious agonist in vitro, preferentially activating pERK in cell lines and Gαo proteins in rat hippocampal membranes. However, Gα subunit mRNA levels differ markedly between rat brain and cell lines, warranting caution when extrapolating from recombinant systems to native tissues.