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Preclinical in vivo characterization of lemborexant (E2006), a novel dual orexin receptor antagonist for sleep/wake regulation - PubMed

️Tue Jan 01 2019

Preclinical in vivo characterization of lemborexant (E2006), a novel dual orexin receptor antagonist for sleep/wake regulation

Carsten Theodor Beuckmann et al. Sleep. 2019.

Abstract

Study objectives: To present results from in vivo studies underlying the preclinical development of lemborexant (E2006), a novel dual orexin (hypocretin) receptor antagonist for sleep/wake regulation.

Methods: Rodent (wild-type rats and wild-type and orexin neuron-deficient [orexin/ataxin-3 Tg/+] mice) studies were performed to evaluate the effects of single-dose oral lemborexant (1-300 mg/kg) on orexin-induced increases in plasma adrenocorticotropic hormone (ACTH), locomotor activity, vigilance state measures (wakefulness, nonrapid eye movement [non-REM] sleep, rapid eye movement [REM] sleep), ethanol-induced anesthesia, and motor coordination, and the effects of multiple-dose oral lemborexant (30 mg/kg) on vigilance state measures. Active comparators were almorexant and zolpidem. Pharmacokinetics were assessed after single-dose lemborexant in mice and rats.

Results: Lemborexant prevented the orexin-promoted increase in ACTH in rats, therefore demonstrating inhibition of the orexin signaling pathway. Furthermore, lemborexant promoted sleep in wild-type mice and rats. Lemborexant promoted REM and non-REM sleep at an equal rate (there was no change in the REM sleep ratio). In contrast, zolpidem reduced REM sleep. The sleep-promoting effect of lemborexant was mediated via the orexin-peptide signaling pathway as demonstrated by a lack of sleep promotion in orexin neuron-deficient mice. Chronic dosing was not associated with a change in effect size or sleep architecture immediately postdosing. Lemborexant did not increase the sedative effects of ethanol or impair motor coordination, showing good safety margin in animals. Pharmacokinetic/pharmacodynamic data for mice and rats were well aligned.

Conclusions: These findings supported further clinical evaluation (ongoing at this time) of lemborexant as a potential candidate for treating insomnia and other sleep disorders.

Keywords: E2006; antagonist; dual orexin receptor antagonist; in vivo; insomnia; lemborexant; mouse; orexin; rat; sleep.

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Figures

**Figure 1.**
Inhibitory effect of single-dose lemborexant on the plasma ACTH increase induced by centrally applied [Ala¹¹, D-Leu¹⁵]-orexin B in rats. Rats received single-dose oral doses of vehicle or lemborexant (1, 3, 10, or 30 mg/kg) (all n = 5, except for 30 mg/kg n = 4) during the light phase. ^#p < 0.05 versus i.c.v. PBS control (t-test); *p < 0.05 versus vehicle/[Ala¹¹, D-Leu¹⁵]-orexin B (one-way analysis of variance followed by Dunnett multiple comparison test).

**Figure 2.**
Relationship between unbound plasma concentrations of lemborexant and corresponding inhibition of [Ala¹¹, D-Leu¹⁵]-orexin B-triggered increases in plasma ACTH concentrations in rats. Data for individual animals are depicted as white open symbols; cohort means are depicted as gray open symbols (mean ± standard error of the mean). Circles = 1 mg/kg lemborexant; squares = 3 mg/kg lemborexant; triangles = 10 mg/kg lemborexant; diamonds = 30 mg/kg lemborexant.

**Figure 3.**
Effect of single-dose lemborexant on spontaneous locomotor activity in wild-type (A) and orexin neuron-deficient (B) mice. Wild-type mice received single oral doses of vehicle (n = 16), lemborexant (30 mg/kg [n = 8]), or lemborexant (100 mg/kg [n = 7]). Orexin neuron-deficient *orexin/ataxin-3* Tg/+ mice received single oral doses of vehicle (n = 8) or lemborexant (100 mg/kg [n = 8]) at Zeitgeber time 3:40 or 5:30 (Zeitgeber time 0:00 = lights on; Zeitgeber time 12:00 = lights off). Data are mean ± standard error of the mean and represent all horizontal and vertical infrared light beam breaks within 1 h after introduction of the mouse to the open field arena. **p < 0.01 versus vehicle (wild-type mice: one-way analysis of variance followed by Dunnett multiple comparison test; orexin neuron-deficient mice: two-tailed unpaired t test).

**Figure 4.**
Effect of single-dose lemborexant on cumulative wakefulness time (A), non-REM sleep time (B), REM sleep time (C), and REM sleep ratio (D) over 3 h after dosing in wild-type mice. Mice received single oral doses of vehicle (n = 5), lemborexant (1 mg/kg, n = 5; 10 mg/kg, n = 4), almorexant (10 mg/kg, n = 5; 100 mg/kg, n = 4), or zolpidem (3 mg/kg, n = 5; 30 mg/kg, n = 4) at Zeitgeber time 3:30–4:10 (Zeitgeber time 0:00 = lights on; Zeitgeber time 12:00 = lights off). Data are mean ± standard error of the mean. *p < 0.05; **p < .01; ***p < 0.001 versus vehicle (one-way analysis followed by Dunnett multiple comparison test).

**Figure 5.**
Effect of single-dose lemborexant on cumulative total sleep time (A) and REM sleep ratio (B) over 3 h after dosing in wild-type and orexin neuron-deficient *orexin/ataxin-3* Tg/+ mice. Mice received single oral doses of vehicle or lemborexant 30 mg/kg (both n = 7 per genotype) at Zeitgeber time 3:30–3:48 (Zeitgeber time 0:00 = lights on; Zeitgeber time 12:00 = lights off). Data are mean ± standard error of the mean. **p < 0.01 versus vehicle (t test).

**Figure 6.**
Effect of single-dose lemborexant on total sleep time (A) and REM sleep ratio (B) in rats. Rats received single oral doses of vehicle, lemborexant (3, 10, 30, 100, or 300 mg/kg), or zolpidem (3, 10, 30, or 100 mg/kg) (all n = 6) at Zeitgeber time 2:00–3:00 (Zeitgeber time 0:00 = lights on; Zeitgeber time 12:00 = lights off). Data are mean ± standard error of the mean for the 2 h after dosing. *p < 0.05 versus vehicle (mixed effect model followed by Dunnett multiple comparison test).

**Figure 7.**
Effect of chronic-dose lemborexant on total sleep time (A), non-REM sleep time (B), REM sleep time (C), and sleep latency (D) in rats. Rats received once-daily oral doses of vehicle (n = 2 per time point), lemborexant 30 mg/kg (n = 5 per time point), or zolpidem 100 mg/kg (n = 4–5 per time point) at Zeitgeber time 2:00–3:00 (Zeitgeber time 0:00 = lights on; Zeitgeber time 12:00 = lights off). Data are mean ± standard error of the mean. *p < 0.05 versus pretreatment value for lemborexant group; ^#p < 0.05 versus pretreatment value for zolpidem group (both repeated measures analysis of variance followed by Dunnett type multiple comparison test).

**Figure 8.**
Effect of single-dose lemborexant on ethanol-induced anesthesia duration in wild-type mice. (A) Mice received single oral doses of vehicle, lemborexant (1, 3, or 10 mg/kg), almorexant (30, 100, or 300 mg/kg), or zolpidem (3, 10, or 30 mg/kg) (all n = 6) during the light phase. (B) Mice received single oral doses of vehicle or lemborexant (30, 100, or 300 mg/kg) (all n = 6) during the light phase. Data are mean ± standard error of the mean. *p < 0.05 versus vehicle (one-way analysis of variance followed by Dunnett multiple comparison test).

**Figure 9.**
Effect of single-dose lemborexant on the latency to fall in wild-type mice. Mice received single oral doses of vehicle, lemborexant (30, 100, or 300 mg/kg), or zolpidem 100 mg/kg (all n = 11 per time point) at Zeitgeber time 4:00 (Zeitgeber time 0:00 = lights on; Zeitgeber time 12:00 = lights off) and had to balance on a rotating axle for a maximum of 180 s at Zeitgeber times 4:30, 6:00, 7.30, and 9:00. Data are mean ± standard error of the mean. There were no significant differences between the vehicle and lemborexant groups. *p < 0.05 versus vehicle (repeated measures analysis of covariance using Zeitgeber time 2:00 predosing values as a covariate).

**Figure 10.**
Plasma concentration profile of lemborexant over time in mice. Mice received single oral doses of lemborexant (10 and 300 mg/kg; both n = 3 per time point) at Zeitgeber time 3:00–9:15 (Zeitgeber time 0:00 = lights on; Zeitgeber time 12:00 = lights off) and plasma samples were taken over 24 h. Data are mean ± standard error of the mean.

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Preclinical in vivo characterization of lemborexant (E2006), a novel dual orexin receptor antagonist for sleep/wake regulation - PubMed