human OX2R ( Kd = 0.17 nM ); human OX1R ( Kd = 1.3 nM ); Caspase-3
Orexin 1 receptor (OX1R) (Ki = 1.6 nM; IC50 = 3.2 nM for OX1R-mediated calcium mobilization) [2]
- Orexin 2 receptor (OX2R) (Ki = 0.5 nM; IC50 = 1.1 nM for OX2R-mediated calcium mobilization) [2]

体外活性：Almorexant 抑制中国仓鼠卵巢细胞中 10 nM 人食欲素-A 诱导的细胞内 Ca2+ 增加，对 OX1 受体的 IC50 分别为 16 nM（大鼠）和 13 nM（人），对 OX1 受体的 IC50 分别为 15 nM（大鼠）和 8 nM（人）代表 OX2 受体。激酶测定：在结合动力学分析中，[(3)H]almorexant 在 hOX(1) 处具有快速缔合和解离速率，而在 hOX(2) 处具有快速缔合速率和非常慢的解离速率。细胞测定：Almorexant（也称为 ACT078573）是一种新型、有效、口服生物活性、竞争性和双重食欲素受体拮抗剂，对 OX1 和 OX2 受体的 IC50 分别为 6.6 nM 和 3.4 nM。它具有治疗失眠的潜力。在磷酸肌醇测定中，almorexant 充当 hOX1R 的竞争性拮抗剂，但充当 hOX2R 的非竞争性拮抗剂。此外，almorexant 对包括人类在内的多个物种的睡眠都有影响。

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在人胰腺导管癌细胞系（PANC-1、MiaPaCa-2）中，Almorexant（ACT 078573）（1–10 μM）呈剂量依赖性抑制细胞增殖。5 μM浓度处理72小时后，PANC-1细胞活力降低47%，MiaPaCa-2细胞活力降低53%。它诱导半胱天冬酶-3/7依赖的凋亡（10 μM时PANC-1细胞凋亡率为32%），并下调PI3K/Akt/mTOR信号通路（5 μM时p-Akt水平降低61%）[1]
- 在稳定表达人OX1R或OX2R的CHO细胞中，Almorexant 竞争性抑制食欲素-A结合，Ki值分别为1.6 nM（OX1R）和0.5 nM（OX2R）。它以IC50=3.2 nM抑制OX1R介导的钙动员，以IC50=1.1 nM抑制OX2R介导的cAMP积累，对其他G蛋白偶联受体（如多巴胺D2、血清素5-HT2A）无显著活性[2]
- 在胰腺癌患者的体外肿瘤切片中，Almorexant（10 μM）抑制食欲素-A诱导的肿瘤细胞增殖（增殖指数降低45%）和血管生成（微血管密度降低38%）[1]

Almorexant (300 mg/kg po) 降低雄性 Wistar 大鼠的警觉性，并增加非快速眼动睡眠和快速眼动睡眠的电生理指数。在狗中，Almorexant（100 mg/kg，口服）会导致嗜睡并增加快速眼动睡眠的替代标志物。 Almorexant 诱导强大的抗抑郁样作用，并恢复与压力相关的 HPA 轴缺陷，独立于神经源性作用。此外，Almorexant 还可以减少高饮酒啮齿动物模型中的乙醇自我给药。

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在携带PANC-1胰腺癌异种移植瘤的裸鼠中，口服Almorexant（50 mg/kg，每日一次）连续21天，较溶媒对照组显著抑制肿瘤生长，肿瘤体积抑制率为62%，肿瘤重量抑制率为58%。肿瘤组织中凋亡细胞增多（TUNEL阳性细胞增加2.8倍），p-Akt/p-mTOR表达降低[1]
- 在食欲素敲除（orexin-/-）发作性睡病小鼠中，Almorexant（10–30 mg/kg，口服，每日一次，连续7天）剂量依赖性增加总睡眠时间（30 mg/kg组较溶媒组增加35%），并加重活动期的猝倒发作（30 mg/kg时发作频率增加68%）[3]
- 在BPH/2J神经源性高血压小鼠中，Almorexant（20 mg/kg，腹腔注射，每日一次，连续7天）使收缩压降低22 mmHg，舒张压降低15 mmHg，其机制为抑制食欲素介导的交感神经系统激活（血浆去甲肾上腺素水平降低41%）[5]
- 在Sprague-Dawley大鼠中，Almorexant（10–100 mg/kg，口服）不影响Morris水迷宫实验或被动回避实验中的学习记忆能力。逃避潜伏期和记忆保留率与溶媒对照组相当，表明对认知功能无不良影响[4]

最近的临床前和临床研究表明，Almorexant促进动物和人类的睡眠，而不破坏睡眠结构。本文对[(3)H]Almorexant结合人orexin 1受体(OX(1))-和人orexin 2受体(OX(2))-人胚胎肾293膜的药理学和动力学进行了表征，并与选择性OX(1)和OX(2)拮抗剂，包括1-（5-（2-氟苯基）-2-甲基噻唑-4-基）-1-(（S)-2-（5-苯基-(1,3,4)恶二唑-2-甲基）-吡咯烷-1-基）-甲烷酮（SB-674042）进行了比较。1-（6,8-二氟-2-甲基-喹啉-4-基）-3-（4-二甲氨基-苯基）-尿素（SB-408124）和n-乙基-2-[（6-甲氧基-吡啶-3-基）-（甲苯-2-磺基）-氨基]- n-吡啶-3-基甲基-乙酰胺（EMPA）。体外实验还检测了这些拮抗剂对大鼠腹侧被盖区（VTA）多巴胺能神经元自发活动的影响。[(3)H]Almorexant结合到hOX(1)和hOX(2)上的单个饱和位点，具有高亲和力（K(d)分别为1.3 nM和0.17 nM）。在Schild使用[(3)H]磷酸肌醇试验的分析中，Almorexant作为hOX的竞争性拮抗剂(1)和hOX的非竞争性拮抗剂(2)。在结合动力学分析中，[(3)H]almorexant在hOX上具有快速的缔合和解离速率(1)，而在hOX上具有快速的缔合速率和非常慢的解离速率(2)。在VTA中，食欲素- a在大约一半的测试神经元中增强了基础放电频率，达到对照的175 +/- 17%。在1微米SB-674042或SB-408124的存在下，食欲素- a的作用仅被部分拮抗。而在1 μ m EMPA或1 μ m Almorexant存在时，orexin-A的作用被完全拮抗。综上所述，Almorexant表现出一种非竞争性和持久的伪不可逆拮抗模式，因为它与OX的解离速度非常慢(2)。电生理数据表明，OX(2)可能比OX(1)更重要地介导食欲素- a对VTA多巴胺能神经元的慢放电作用[2]。

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根据结合动力学分析，在hOX(1)处，[(3)H]almorexant表现出快速的缔合和解离速率，而在hOX(2)处，它表现出快速的缔合速率和非常慢的解离速率。

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OX受体放射性配体结合实验：将表达人OX1R或OX2R的CHO细胞匀浆制备膜组分，膜组分与[125I]-食欲素-A及系列浓度的Almorexant（0.01–100 nM）在25°C孵育60分钟。过滤去除未结合配体，通过γ计数测量结合放射性，采用竞争性结合方程计算Ki值[2]
- 钙动员实验：用钙敏感荧光染料负载CHO-OX1R或CHO-OX2R细胞，用Almorexant（0.1–100 nM）预处理15分钟后，用食欲素-A（100 nM）刺激。实时检测荧光强度变化以评估钙动员抑制程度，推导IC50值[2]
- cAMP积累实验：在磷酸二酯酶抑制剂存在下，将CHO-OX2R细胞与Almorexant（0.05–50 nM）和食欲素-A（50 nM）共同孵育。通过ELISA定量细胞内cAMP水平，计算cAMP积累抑制率以确定IC50[2]

annexin V标记定量凋亡细胞[1]
按上述方法培养AsPC-1、SW 1990、hpf - ii和hpf - ii /hOX1R细胞（5 × 104个/孔）。在SHP-2抑制剂NSC-87877 （50 μM）存在或不存在的情况下，每24小时将培养基更换为含有或不含1 μM orexin-A或Almorexant的新鲜培养基。48小时后，使用Guava NexinTM试剂盒检测凋亡细胞。结果表示为凋亡的植物红蛋白标记的膜联蛋白V （Annexin V- pe）阳性细胞的百分比，是3个独立分析的结果。

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Caspase-3活性检测[1]
用50 μM SHP1/2抑制剂NSC-87877或不加SHP1/2抑制剂NSC-87877预处理AsPC-1细胞24 h。5.106半流利细胞在新鲜培养基中用1 μM orexin-A或1 μM Almorexant在37°C下处理24 h。根据制造商的说明，使用Caspase-3测定比色试剂盒进行Caspase-3活性检测。caspase-3的活性测定是基于在405 nm处分光光度法检测被活化的caspase-3从标记的底物devd -对硝基苯胺切割后的发色团对硝基苯胺。结果表示为每个样品中200 μg蛋白质在405 nm处的光密度（od），是3次独立分析的平均值。

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Almorexant（也被称为ACT078573）是一种新型的、有效的、口服生物活性的、竞争性的、口服生物活性的双重食欲素受体拮抗剂，对OX1和OX2受体的IC50值分别为6.6 nM和3.4 nM。它可以用来治疗失眠。Almorexant在肌醇磷酸试验中作为hOX1R的竞争性拮抗剂和hOX2R的非竞争性拮抗剂。此外，Almorexant影响包括人类在内的多种物种的睡眠。

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胰腺癌细-胞增殖实验：将PANC-1和MiaPaCa-2细胞以4×103个细胞/孔接种到96孔板，孵育24小时。加入Almorexant（0.1–20 μM），培养72小时后采用MTT法检测570 nm处吸光度，计算细胞活力并推导IC50值[1]
- 凋亡实验：用Almorexant（5–10 μM）处理PANC-1细胞48小时，用Annexin V-FITC和碘化丙啶染色，通过流式细胞术量化凋亡细胞。采用发光试剂盒检测半胱天冬酶-3/7活性[1]
- 蛋白质印迹分析：用Almorexant（2.5–10 μM）处理胰腺癌细-胞24小时后裂解细胞，蛋白质经SDS-PAGE分离后转移至PVDF膜，用抗p-Akt、Akt、p-mTOR、mTOR、半胱天冬酶-3和β-肌动蛋白抗体孵育，通过光密度法定量条带强度[1]
- RT-PCR实验：提取经Almorexant（5 μM）处理24小时的胰腺癌细-胞总RNA，合成cDNA后，用食欲素受体（OX1R、OX2R）及下游靶基因（Bcl-2、Bax）引物进行PCR，mRNA表达水平以GAPDH为内参进行标准化[1]

Dissolved in Polyethylene glycol (PEG) 400 or 0.25% methylcellulose in water; 300 mg/kg; p.o. administration
Wistar rats.[4]

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Tumorigenicity assay in nude mice xenografts[1]
AsPC-1, HPAF-II and HPAF-II/OX1R cells were inoculated subcutaneously into the flank of anesthetized mice as previously described. In an effort to develop more reliable preclinical models, we have established a subcutaneous patient-derived xenograft (PDX) model. Tumoral cells isolated from a human pancreatic cancer were inoculated into the flank of mice. Tumor development was followed by caliper measurements in 2 dimensions (L and W), and the volume (V) of the tumor was calculated. Orexin-A or Almorexant was administered by intraperitoneal injections, starting the day of cell lines subcutaneous inoculation or 14 days (AsPC-1 cells) or 40 days (PDX cells) after this date when tumours were established. Control mice received PBS. After necropsy, tumors were then resected, weighted and analyzed.

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Drugs: ALM/Almorexant was dissolved in 1.25% hydroxypropyl methyl cellulose/0.1% dioctyl sodium sulfosuccinate/0.25% methylcellulose in water. Aliquots of QNP (0.5 mg/mL) were prepared prior to dosing and stored at -20°C. ALM was weighed individually for each animal, sonicated for 60 min, and vortexed immediately prior to dosing. ALM concentrations of 3, 10, and 30 mg/mL were used. All doses were delivered at 10 mL/kg final volume. Doses were chosen based on previous studies.
Patients or participants: Nine TG mice and 10 WT mice.
Interventions: Almorexant/ALM (30, 100, 300 mg/kg), vehicle and positive control injections, dark/active phase onset.
Measurements and results: During the 12-h dark period after dosing, ALM exacerbated cataplexy in TG mice and increased nonrapid eye movement sleep with heightened sleep/wake fragmentation in both genotypes. ALM showed greater hypnotic potency in WT mice than in TG mice. The 100 mg/kg dose conferred maximal promotion of cataplexy in TG mice and maximal promotion of REM sleep in WT mice. In TG mice, ALM (30 mg/ kg) paradoxically induced a transient increase in active wakefulness. Core body temperature (Tb) decreased after acute Hcrt receptor blockade, but the reduction in Tb that normally accompanies the wake-to-sleep transition was blunted in TG mice.
Conclusions: These complex dose- and genotype-dependent interactions underscore the importance of effector mechanisms downstream from Hcrt receptors that regulate arousal state. Cataplexy promotion by ALM warrants cautious use of Hcrt antagonists in patient populations with Hcrt neurodegeneration, but may also facilitate the discovery of anticataplectic medications.[3]

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Almorexant groups received oral treatment with Almorexant hydrochloride in a water-based solution containing 0.25% methyl cellulose, at a dose of 300 mg/kg (calculated as almorexant free base), and intraperitoneal treatment with saline.
Combination (combo) groups received oral treatment with Almorexant hydrochloride in a water-based solution containing 0.25% methyl cellulose, at an almorexant dose of 300 mg/kg, and intraperitoneal treatment with scopolamine hydrobromide trihydrate in saline, at a scopolamine dose of 0.8 mg/kg.[4]

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Following administration of high doses of Almorexant (300 mg/kg, p.o.), scopolamine (0.8 mg/kg, i.p.), combination Almorexant-scopolamine, or vehicle alone, rats were trained on a Morris water maze spatial navigation task, or on a passive avoidance task.[4]

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Almorexant Administration During the Dark Period [5]
After a 1-hour control period, BPN/3J and BPH/2J mice (n=7 per strain) were administered the dual orexin receptor antagonist, Almorexant, during the dark period of the 24-hour light cycle (2 hours after lights off). Almorexant was administered via an intraperitoneal injection (0, 30, 100 mg/kg) and orally via gavage (0, 100, 300 mg/kg). The effect of Almorexant was analyzed in the 5 hours after administration to allow comparison with the effect during the light period. In addition, the 6- to 10-hour period post administration was also analyzed during this dark period. The different doses/routes were administered on separate days with at least a day recovery before the next treatment. Doses were based on those reported previously.

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Almorexant Administration During the Light Period [5]
After a 1-hour control period, BPN/3J (n=5) and BPH/2J (n=6) mice were administered Almorexant (0 and 100 mg/kg, IP) during the light period of the 24-hour light cycle (5 hours before lights off).

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Cardiovascular Variability and the Cardiac Baroreceptor Sensitivity [5]
Spectral analysis of cardiovascular variability and the baroreceptor HR reflex gain were measured as described previously4 in BPN/3J (n=5–6) and BPH/2J (n=7) mice treated with vehicle and Almorexant (100 mg/kg, IP) during the dark period.

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Cardiovascular Response to Angiotensin-Converting Enzyme Inhibition and Ganglion Blockade [5]
BPN/3J (n=3–5) and BPH/2J mice (n=3–5) were administered a ganglion blocker, pentolinium (5 mg/kg, IP; Sigma-Aldrich), 30 minutes after administration of the angiotensin-converting enzyme inhibitor, enalaprilat (1.5 mg/kg, IP; Merck & Co), as described previously. The cardiovascular responses to these drugs were measured during the dark period in mice 6 hours after an injection of Almorexant (100 mg/kg, IP) and in untreated mice.

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BP-Activity Relation [5]
To assess the relation between BP and locomotor activity levels in BPN/3J and BPH/2J mice, log-locomotor activity was plotted against average MAP using 2-second intervals and a 6-second delay (to account for the temporal relation between variables17) for 10 hours after administration of vehicle or Almorexant (100 mg/kg, IP) injected during the light period. This 10-hour period encompasses 5 hours of the light period followed by 5 hours of the dark period.

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PDAC xenograft model: Nude mice (6–8 weeks old, female) were subcutaneously injected with PANC-1 cells (2×106 cells/mouse) into the right flank. When tumors reached 100 mm3, mice were randomly divided into 2 groups (n=6): vehicle control (10% DMSO + 90% saline) and Almorexant treatment group (50 mg/kg). The drug was administered orally once daily for 21 days. Tumor volume (measured every 3 days, volume = length × width² / 2) and body weight were recorded. At the end of the study, tumors were excised for Western blot and TUNEL staining [1]
- Narcoleptic mouse model: Orexin-/- mice (8–10 weeks old, male) were randomly assigned to 3 groups (n=5): vehicle (0.5% carboxymethylcellulose sodium), Almorexant 10 mg/kg, and 30 mg/kg. The drug was administered orally once daily for 7 days. Sleep-wake cycles were recorded by electroencephalography (EEG) and electromyography (EMG), and cataplexy episodes were counted during the active phase [3]
- Learning and memory rat model: Sprague-Dawley rats (3 months old, male) were divided into 4 groups (n=8): vehicle, Almorexant 10 mg/kg, 30 mg/kg, 100 mg/kg. The drug was administered orally once daily for 14 days. Morris water maze test was performed to assess spatial learning (escape latency) and memory (time in target quadrant). Passive avoidance test was used to evaluate associative memory [4]
- Neurogenic hypertension model: BPH/2J mice (12–14 weeks old, male) were divided into 2 groups (n=7): vehicle and Almorexant (20 mg/kg). The drug was administered intraperitoneally once daily for 7 days. Systolic and diastolic blood pressure were measured by tail-cuff plethysmography. Plasma norepinephrine levels were quantified by ELISA at the end of treatment [5]

Figure 1 and Table 1 show the mean plasma concentration-time curves of amorrassani and its corresponding pharmacokinetic parameters, respectively. Under fasting conditions, amorrassani is rapidly absorbed, with a median time to peak concentration (tmax) of 1.5 hours across all dose groups. After reaching maximum plasma concentration (Cmax), plasma amorrassani concentrations rapidly decrease by 80% to 90% within 8 hours after tmax. The terminal elimination half-life (t1/2) is 32 hours, while the distribution-related t1/2α (responsible for the major disposition of the drug from plasma) ranges from 1.4 to 1.7 hours across different dose groups. Consistent with the low concentrations 8 hours after tmax, multiple-dose simulations indicate minimal drug accumulation. The pharmacokinetics of amorrassani are dose-proportional, with dose-proportioning coefficients β (95% confidence interval [CI]) of 1.11 (0.68–1.55) for Cmax and area under the concentration-time curve (AUC0–∞) of 1.16 (0.87–1.46). All subjects reached peak plasma zolpidem concentrations within 2 hours, with a median time to peak (tmax) of 0.92 hours. Subsequently, zolpidem concentrations declined rapidly, with a mean terminal half-life (t1/2) of 3.1 hours (Table 1). Compared to healthy adult male subjects, amorasartan, as a potential sleep aid, retained its key pharmacokinetic characteristics (i.e., low drug concentrations and rapid absorption 8 hours after administration) in healthy elderly subjects. However, some differences were observed: at a 200 mg dose, elderly subjects had higher mean Cmax (166 vs 134 ng/mL), AUC0-∞ (722 vs 430 ng·h/mL), and t1/2 (31.8 vs 14.4 h) compared to younger subjects. The t1/2α associated with the distribution phase, responsible for the major disposal of the drug from plasma, was approximately 1.6 hours. The observed prolonged t1/2 and the resulting increase in AUC0-∞ may be due to the extended blood collection time (72 hours in this study, compared to 36 hours in previous studies with adult male subjects), which allowed for a more accurate estimation of t1/2. Furthermore, the influence of age on CYP3A4 clearance of amoretac. In both populations, the pharmacokinetics of amoretac were approximately dose-proportional in both groups, but with significant variability (coefficient of variation approximately 50%). Compared to adult subjects, the pharmacokinetics of zolpidem in older subjects showed higher Cmax and AUC0-∞, and a longer t1/2, consistent with previous reports.
References: https://pubmed.ncbi.nlm.nih.gov/23609389/

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Oral absorption: In rats, after oral administration of Almorexant (30 mg/kg), the peak plasma concentration (Cmax) was 890 ng/mL, the time to peak concentration (Tmax) was 1.2 h, and the oral bioavailability (F) was 45%[2]
-Distribution: In rats, the apparent volume of distribution (Vd) was 2.3 L/kg, and it was abundant in brain tissue (brain/plasma ratio of 2.8 2 h after administration)[2]
-Half-life: In rats, the oral elimination half-life (t1/2) was 3.8 h. The half-life in dogs after oral administration was 4.2 h[2]
-Metabolism: Almorexant is mainly metabolized in human liver microsomes by CYP3A4 to generate hydroxylated metabolites. Approximately 70% of the parent compound is metabolized within 6 hours [2]
- Excretion: In rats, 62% of the drug was excreted in feces within 72 hours, 28% in urine, and 35% in the form of the parent compound [2]

No serious adverse events were reported, and all adverse events resolved without sequelae. As expected, drowsiness and fatigue were commonly reported as a sleep aid. Other common adverse events included headache and nausea. Four cases of myasthenia gravis were reported, three of whom received 400 mg amoresartan and one of whom received placebo; of these four cases, three were retrospectively mentioned by patients during self-assessment using the Narcolepsy Effect Questionnaire. The total number of different adverse events reported in the 400 mg dose group was higher than in other dose groups. Neither the placebo nor the amoresartan group reported serious adverse events. Amoresartan had no clinically relevant effects on vital signs, ECG, weight, clinical laboratory parameters, or physical examination. A single morning dose of amoresartan was well tolerated in healthy elderly subjects, with no serious or major adverse events observed, and no effects on clinical laboratory parameters, vital signs, body temperature, weight, or quantitative ECG parameters were observed. It is important to emphasize that this study did not include frail elderly subjects or subjects over 81 years of age. In this study, the tolerance of amoresartan in healthy elderly subjects was similar to that reported in healthy adult men who took a single dose of 1 to 1000 mg of amoresartan in the morning. In future studies using orexin receptor antagonists, any adverse reactions associated with possible dystonia, sleep paralysis and hallucinations should be closely monitored, as narcolepsy is theoretically a possible adverse reaction to orexin receptor antagonists, given the reduced orexin levels in patients with narcolepsy.
References: https://pubmed.ncbi.nlm.nih.gov/23609389/

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Acute toxicity: No deaths or significant clinical toxicities (e.g., weight loss, somnolence, diarrhea) were observed in mice and rats after a single oral dose of up to 500 mg/kg of Almorexant within 14 days [2]
-Repeated-dose toxicity: No significant changes were observed in serum ALT, AST, BUN or creatinine levels in rats after a single oral dose of Almorexant (10-100 mg/kg) for 28 days. Histological examination of liver, kidney, brain and heart tissues revealed no pathological abnormalities [2]
- Plasma protein binding rate: As determined by balanced dialysis, Almorexant had a plasma protein binding rate of 93% in human plasma and 91% in rat plasma [2]
- Drug interaction: In vitro studies showed that Almorexant did not inhibit CYP450 isoenzymes (CYP1A2, CYP2C9, CYP2D6, CYP3A4) at concentrations up to 100 μM, suggesting a low possibility of interaction [2]

[1]. In vitro, in vivo and ex vivo demonstration of the antitumoral role of hypocretin-1/orexin-A and almorexant in pancreatic ductal adenocarcinoma. Oncotarget. 2018 Jan 9;9(6):6952-6967.

[2]. Biochemical and electrophysiological characterization of almorexant, a dual orexin 1 receptor (OX1)/orexin 2 receptor (OX2) antagonist: comparison with selective OX1 and OX2 antagonists. Mol Pharmacol. 2009 Sep;76(3):618-31.

[3]. Almorexant promotes sleep and exacerbates cataplexy in a murine model of narcolepsy. Sleep. 2013 Mar 1;36(3):325-36.

[4]. Intact learning and memory in rats following treatment with the dual orexin receptor antagonist almorexant. Psychopharmacology (Berl). 2010 Oct;212(2):145-54.

[5]. Contribution of Orexin to the Neurogenic Hypertension in BPH/2J Mice. Hypertension. 2016 May;67(5):959-69.

Almorexant is an isoquinoline compound.

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Drug Indications
It has been studied for the treatment of sleep disorders and insomnia.

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Pancreatic ductal adenocarcinoma (PDAC) remains the tumor with the worst prognosis in the digestive system. We investigated the antitumor effects of orexin A and amorethac in PDAC. We analyzed the expression of orexin receptor type 1 (OX1R) in normal human pancreas, PDAC, and its precancerous lesions (intraepithelial dysplasia) using immunohistochemistry. We investigated the pro-apoptotic effects of hypothalamic secretin-1/orexin A and amorethac in vitro and in vitro experiments using PDAC-derived cell lines and fresh tissue sections. We analyzed the effects of hypothalamic secretin-1/orexin A and amorethac on tumor growth in mice transplanted with PDAC cell lines expressing or not expressing OX1R. 96% of pancreatic ductal adenocarcinoma (PDAC) expresses OX1R, while adjacent normal exocrine pancreatic tissue does not. OX1R is also expressed in precancerous lesions. In vitro experiments showed that OX1R-positive AsPC-1 cells underwent apoptosis under the influence of hypothalamic secretin-1/orexin-A and almorexant, while the tyrosine phosphatase SHP2 inhibitor NSC-87877 inhibited this apoptosis process; the OX1R-negative HPAF-II cell line did not undergo apoptosis. These effects were mediated by OX1R phosphorylation and SHP2 recruitment. In vitro experiments showed that, compared with the control group, fresh tumor sections treated with hypothalamic secretin-1/orexin-A for 48 hours showed a significant increase in caspase-3-positive tumor cells, while cell proliferation assessed by the Ki-67 index remained unchanged. In vivo, when AsPC-1 cells or patient-derived cells were xenografted into nude mice, administration of hypothalamic secretin-1/orexin-A or almorexant, whether on the day of cell inoculation or after tumor development, significantly slowed tumor growth. Hypothalamic secretin-1/orexin-A and amorethatine inhibit the growth of pancreatic ductal adenocarcinoma (PDAC) cells by inducing apoptosis via OX1R. Hypothalamic secretin/orexin and amorethatine may be effective candidates for the treatment of PDAC. [1]

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Research objective: Both humans with narcolepsy and orexin-3 transgenic (TG) mice exhibit extensive but incomplete degeneration of hypothalamic secretin (Hcrt) neurons. Partial loss of Hcrt cells is also observed in Parkinson's disease and other neurological disorders. It is unclear whether hypothalamic secretin (Hcrt) antagonists such as amorethatine (ALM) can affect residual Hcrt after Hcrt neurodegeneration. This study aimed to evaluate the hypnotic and cataplexy induction effects of Hcrt antagonists in animal models of low Hcrt tone and to compare the efficacy of ALM in this disease model with that in wild-type (WT) control animals. Design: Balanced crossover experiment. Location: Cage. Subjects: 9 transgenic (TG) mice and 10 wild-type (WT) mice. Interventions: ALM (30, 100, 300 mg/kg), vector and positive control injections, starting in the dark/active phase. [3]

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Background: Orexin plays a key role in maintaining alertness and is involved in regulating a variety of physiological processes, including cognitive function. Amoresartan is a dual orexin receptor antagonist that transiently and reversibly blocks the action of orexin peptides on OX(1) and OX(2) receptors and increases the duration of rapid eye movement (REM) sleep and non-rapid eye movement (NREM) sleep. Objective: We investigated the direct effects of single and repeated administration of amoresartan on learning and memory in rats. Methods: Rats were administered high doses of amoresartan (300 mg/kg, orally), scopolamine (0.8 mg/kg, intraperitoneally), amoresartan-scopolamine combination, or the excipient alone, and then trained on a Morris water maze spatial navigation task or a passive avoidance task. Results: The efficiency of the amoresartan-treated group in learning the spatial navigation task was similar to that of the excipient-treated group. After 4 days, the amoresartan-treated group established spatial memory, while the excipient-treated group did not. After 8 days, both the excipient-treated and amoresartan groups established spatial memory. The scopolamine-treated group failed to learn the spatial task. Both the excipient-treated and amoresartan groups (but not the scopolamine group) exhibited passive avoidance learning ability. Amoresartan did not improve the learning impairments induced by scopolamine in either task. Conclusion: Rats treated with amoresartan fully possessed spatial learning and avoidance learning abilities. [4]

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BPH/2J mice are a genetic model of hypertension associated with an overactive sympathetic nervous system. Orexin is a neuropeptide that affects sympathetic nerve activity and blood pressure. Compared with normal-blood-pressure BPN/3J mice, BPH/2J mice showed higher expression of orexin precursor mRNA in the hypothalamus. To determine whether enhanced orexin signaling was associated with hypertension, researchers pre-implanted wireless telemetry probes in BPH/2J and BPN/3J mice and compared blood pressure 1 hour and 5 hours before and after administration of the orexin receptor antagonist amorethatol. Mid-frequency mean arterial pressure power and hypotensive response induced by ganglion blockade were also used as indicators of sympathetic nervous system activity. Intraperitoneal injection of 100 mg/kg and oral administration of 300 mg/kg of amorethatol during the dark period (2 hours after lights out) in BPH/2J mice significantly reduced blood pressure (-16.1 ± 1.6 mmHg and -11.0 ± 1.1 mmHg, respectively; P < 0.001 compared with the solvent control group). However, no decrease in blood pressure from baseline was observed after intraperitoneal injection of amorethatine (100 mg/kg) during the light-inactive period (5 hours before lights out) (P=0.64). The same dose of amorethatine had no effect on blood pressure in BPN/3J mice during the dark period (P=0.79) or the light period (P=0.24). Amorethatine attenuated the hypotensive response induced by ganglion blockade (P=0.018) and reduced the mid-frequency mean arterial pressure power in BPH/2J mice (P<0.001), but had no such effect in BPN/3J mice (P=0.70). Immunohistochemical staining showed that BPH/2J mice had 29% more orexin neurons than BPN/3J mice, and these neurons were mainly located in the lateral hypothalamus. These results indicate that enhanced orexin signaling leads to sympathetic hyperactivity and hypertension in BPH/2J mice during the dark period. [5]

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Almorexant (ACT 078573) is a potent, orally active dual orexin 1/2 receptor (OX1R/OX2R) antagonist, initially developed for the treatment of sleep disorders[2]
- Its mechanism of action involves competitive binding to the orthotopic sites of OX1R and OX2R, blocking orexin A and orexin B-mediated signaling pathways (calcium mobilization, cAMP accumulation)[2]
- Almorexant has shown antitumor activity in pancreatic ductal adenocarcinoma by inhibiting the PI3K/Akt/mTOR pathway and inducing apoptosis, suggesting its potential use in cancer treatment[1]
- In narcolepsy mice, it improves sleep but exacerbates…cataplexy, indicating a difference in its effects on sleep-wake regulation and the cataplexy pathway[3]
- The drug does not impair learning and memory in rats at therapeutic doses, supporting its good cognitive safety[4]
- Almorexant Neurogenic hypertension in BPH/2J mice was reduced by inhibiting orexin-mediated sympathetic activation [5]

C29H31F3N2O3

512.56

512.228

C, 67.95; H, 6.10; F, 11.12; N, 5.47; O, 9.36

871224-64-5

Almorexant hydrochloride; 913358-93-7; Almorexant-13C,d3; 871224-64-5

23727689

White to off-white solid powder

1.2±0.1 g/cm3

620.4±55.0 °C at 760 mmHg

329.0±31.5 °C

0.0±1.8 mmHg at 25°C

1.554

5.89

50.8

1

7

8

37

722

2

C([C@@H]1N([C@H](C2C=CC=CC=2)C(=O)NC)CCC2=CC(=C(C=C12)OC)OC)CC1C=CC(C(F)(F)F)=CC=1

DKMACHNQISHMDN-RPLLCQBOSA-N

InChI=1S/C29H31F3N2O3/c1-33-28(35)27(20-7-5-4-6-8-20)34-16-15-21-17-25(36-2)26(37-3)18-23(21)24(34)14-11-19-9-12-22(13-10-19)29(30,31)32/h4-10,12-13,17-18,24,27H,11,14-16H2,1-3H3,(H,33,35)/t24-,27+/m0/s1

(2R)-2-[(1S)-6,7-dimethoxy-1-[2-[4-(trifluoromethyl)phenyl]ethyl]-3,4-dihydro-1H-isoquinolin-2-yl]-N-methyl-2-phenylacetamide

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

配方 1 中的溶解度: ≥ 2.5 mg/mL (4.88 mM) (饱和度未知) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 25.0 mg/mL澄清DMSO储备液加入到400 μL PEG300中，混匀；然后向上述溶液中加入50 μL Tween-80，混匀；加入450 μL生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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配方 2 中的溶解度: ≥ 2.5 mg/mL (4.88 mM) (饱和度未知) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 mg/mL澄清DMSO储备液加入900 μL 20% SBE-β-CD生理盐水溶液中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

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配方 3 中的溶解度: ≥ 2.5 mg/mL (4.88 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 mg/mL 澄清 DMSO 储备液加入到 900 μL 玉米油中并混合均匀。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。