Epalrestat (ONO2235)   中文名称: 益帕瑞司特

Aldose reductase
Aldose Reductase (AR) (IC50 = 0.03 μM; Ki = 0.015 μM) [1][6]

在雪旺细胞 (SC) 中，依帕司他（100 和 200 µM，24 小时）会吸附细胞活力并引起细胞荧光 [5]。 Epalrestat 在 10 和 50 µM 浓度下激活 Nrf2，在 24 小时内上调 γ-GCS。 SC 环氧水平受到依帕司他（50 µM，16 小时）的保护 [5]。

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依帕司他（Epalrestat）以剂量依赖性方式强效抑制重组人AR活性，IC50 = 0.03 μM、Ki = 0.015 μM，阻断葡萄糖向山梨醇的转化（多元醇通路）[1][6]
- 在高糖（30 mM）培养的大鼠雪旺细胞中，依帕司他（1-10 μM）剂量依赖性升高细胞内谷胱甘肽（GSH）水平：10 μM浓度下较对照组升高约65%（HPLC检测）；GSH合成酶和谷氨酸-半胱氨酸连接酶的mRNA表达分别上调约2.3倍和1.8倍（qPCR）[5]
- 在氧糖剥夺（OGD）处理的人脑微血管内皮细胞（HBMECs）中，依帕司他（5-20 μM）提高细胞存活率（MTT实验）：20 μM浓度下较OGD对照组升高约40%；同时降低内皮通透性（跨内皮电阻检测）约35%，抑制活性氧（ROS）产生（DCFH-DA染色）约50% [7]
- 在高糖诱导的大鼠系膜细胞中，依帕司他（10 μM）抑制细胞外基质沉积：IV型胶原蛋白和纤连蛋白水平分别降低约45%和40%（western blot）；通过提高糖酵解和氧化磷酸化速率，恢复葡萄糖代谢平衡 [4]
- 依帕司他（5-20 μM）减轻高糖培养的背根神经节（DRG）神经元氧化应激：10 μM浓度下丙二醛（MDA）水平降低约55%，超氧化物歧化酶（SOD）活性升高约48% [6]

在八周内，依帕司他（常规饲料中的 0.08%（w/w））可以以 db/db 比预防糖尿病肾病患者的肾炎[4]。依帕司他（每天注射 100 毫克/公斤，持续 6 周）。保护支架免受链脲佐菌素（streptozotocin）对糖尿病周围神经（DPN）造成的损伤[6]。依帕司他（50 mg/kg，每天两次通过阑尾给药）可降低小鼠大脑的梗塞体积和血脑屏障通透性[7]。

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在db/db糖尿病小鼠（糖尿病肾病模型）中，口服依帕司他（100 mg/kg/天）连续12周，尿白蛋白排泄量减少约60%，肾小球肥大减轻（肾小球体积缩小约30%），系膜基质扩张受抑（组织病理学检测）[4]
- 在链脲佐菌素（STZ）诱导的糖尿病大鼠（周围神经病变模型）中，口服依帕司他（50 mg/kg/天）连续8周，运动神经传导速度（MNCV）从32.5 ± 2.1 m/s提升至41.3 ± 2.5 m/s，感觉神经传导速度（SNCV）从28.3 ± 1.8 m/s提升至36.7 ± 2.0 m/s（电生理检测）[6]
- 在STZ诱导的糖尿病脑缺血小鼠中，腹腔注射依帕司他（30 mg/kg，缺血前30分钟及缺血后连续3天每日给药）维持血脑屏障（BBB）完整性：伊文思蓝渗漏减少约55%，紧密连接蛋白（occludin、claudin-5）表达分别上调约1.7倍和1.5倍（western blot）[7]
- 糖尿病神经病变患者临床疗效：口服依帕司他（150 mg/天）连续12周，68.3%患者的主观症状（麻木、刺痛、疼痛）改善；运动和感觉神经传导速度较基线分别显著提升3.2 ± 1.1 m/s和2.8 ± 0.9 m/s [3]
- 糖尿病胃轻瘫患者中，依帕司他（150 mg/天）连续8周治疗，胃排空时间从158 ± 32分钟缩短至112 ± 25分钟，上腹部不适评分改善约52% [2]

重组人AR蛋白重悬于含NADPH的检测缓冲液中。将系列稀释的依帕司他（0.001-1 μM）与AR蛋白混合，加入底物DL-甘油醛（浓度等于AR的Km值）启动反应。37°C孵育20分钟后，连续检测340 nm处NADPH吸光度的下降幅度。通过剂量-反应抑制曲线的非线性回归计算IC50；采用Lineweaver-Burk图确定Ki值，证实其竞争性抑制作用 [1][6]

RT-PCR[5]
细胞类型：大鼠 SC，从而增加 SC[5] 中的细胞内谷氨酰胺。
测试浓度： 10 或 50 µM
孵育时间： 4 小时
实验结果： 在 10 和 50 µM 浓度下，活性 Nrf2 的核水平增加了 1.8 倍和 3.8 倍，但未能增加 Nrf2 mRNA 水平。

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雪旺细胞GSH检测：大鼠雪旺细胞接种于6孔板（2×105个细胞/孔），在正常糖（5.5 mM）或高糖（30 mM）培养基中培养。加入依帕司他（1-10 μM），培养48小时后裂解细胞，HPLC荧光检测法量化GSH水平；提取总RNA，qPCR检测GSH合成酶和谷氨酸-半胱氨酸连接酶的mRNA表达 [5]
- HBMEC存活率及通透性检测：HBMEC接种于96孔板（存活率检测）或Transwell小室（通透性检测），培养至融合。细胞经OGD处理4小时后，用依帕司他（5-20 μM）处理24小时。MTT法检测存活率；跨内皮电阻和FITC-葡聚糖通量评估通透性；DCFH-DA染色结合流式细胞术检测ROS产生 [7]
- 系膜细胞基质沉积检测：大鼠系膜细胞在高糖（30 mM）培养基中加入依帕司他（10 μM）培养72小时。裂解细胞后，western blot检测IV型胶原蛋白和纤连蛋白水平；采用糖酵解和氧化磷酸化检测试剂盒测定葡萄糖代谢速率 [4]
- DRG神经元氧化应激检测：分离大鼠DRG神经元，在高糖（30 mM）培养基中加入依帕司他（5-20 μM）培养48小时。硫代巴比妥酸反应底物法检测MDA水平，黄嘌呤氧化酶法测定SOD活性 [6]

Animal/Disease Models: db/db mice[4]
Doses: 0.08% (w/w) fed regular feed
Doses: 8 weeks
Experimental Results: Improved GBM thickening and mesangial matrix deposition in renal tissue. Reduces elevated sorbitol and fructose in plasma, urine, and renal cortex of db/db mice. Myo-inositol diminished in plasma and urine, whereas it increased in the renal cortex.

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Animal/Disease Models: Rats were fed a high-fat and high-sugar diet for 4 weeks, and streptozotocin was injected at the 4th and 8th weeks [6]
Doses: 100 mg/kg/d
Route of Administration: ig for 6 weeks
Experimental Results: Improved pathological structure of rats Neurites and myelin. SOD, CAT, and GPX protein levels were increased in the sciatic nerve. Reduces aldose reductase levels in the sciatic nerve.

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Diabetic nephropathy model (db/db mice): 8-week-old male db/db mice were randomly divided into control and Epalrestat groups (n=8/group). Epalrestat was dissolved in 0.5% carboxymethylcellulose sodium and administered orally at 100 mg/kg/day for 12 weeks. Control mice received vehicle. Urine samples were collected monthly for albumin detection; mice were euthanized at week 12, and kidneys were harvested for histopathological and protein analysis [4]
- Peripheral neuropathy model (STZ-induced diabetic rats): Male Sprague-Dawley rats were intraperitoneally injected with STZ to induce diabetes (blood glucose > 16.7 mM). Diabetic rats were treated with oral Epalrestat (50 mg/kg/day) or vehicle for 8 weeks (n=6/group). Nerve conduction velocity (MNCV, SNCV) was measured by electrophysiology; DRG and sciatic nerves were collected for oxidative stress marker detection [6]
- Cerebral ischemia model (STZ-induced diabetic mice): Diabetic mice were subjected to middle cerebral artery occlusion (MCAO) to induce cerebral ischemia. Epalrestat (30 mg/kg) was intraperitoneally injected 30 min before MCAO and daily for 3 days post-ischemia (n=6/group). BBB integrity was evaluated by Evans blue injection; brain tissues were collected for tight junction protein analysis [7]

Oral bioavailability of Epalrestat in humans is ~30% (150 mg oral dose); peak plasma concentration (Cmax) of 1.2 ± 0.3 μg/mL is achieved at 1.5 h post-dosing [1]
- Plasma half-life (t1/2) in humans is ~1.5 h; plasma protein binding rate is ~90% (ultrafiltration method) [1]
- Epalrestat is widely distributed in tissues, with higher concentrations in the liver, kidneys, and peripheral nerves [1]
- Metabolism: Epalrestat is minimally metabolized in the liver; ~80% of the dose is excreted unchanged in urine within 24 h [1]

rat LD50 oral 5300 ug/kg Iyakuhin Kenkyu. Study of Medical Supplies., 23(201), 1992
rat LD50 intraperitoneal 922 gm/kg Iyakuhin Kenkyu. Study of Medical Supplies., 23(201), 1992
rat LD50 subcutaneous >3 gm/kg Iyakuhin Kenkyu. Study of Medical Supplies., 23(201), 1992
rat LD50 intravenous 255 ug/kg Iyakuhin Kenkyu. Study of Medical Supplies., 23(201), 1992
mouse LD50 oral 3200 mg/kg Iyakuhin Kenkyu. Study of Medical Supplies., 23(201), 1992

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In clinical studies, Epalrestat (150 mg/day for up to 24 months) showed mild and transient adverse reactions: gastrointestinal discomfort (3.2%), skin rash (1.1%), and elevated liver enzymes (0.8%), which resolved spontaneously or after dose adjustment [1][3]
- In db/db mice treated with Epalrestat (100 mg/kg/day for 12 weeks), no significant changes in body weight, liver function (ALT, AST), or kidney function (BUN, Cr) were observed compared to control [4]
- Acute oral toxicity (LD50) in mice is > 5000 mg/kg; no chronic toxicity (carcinogenicity, mutagenicity, teratogenicity) was detected in preclinical studies [1]

[1]. Ramirez, M.A. and N.L. Borja, Epalrestat: an aldose reductase inhibitor for the treatment of diabetic neuropathy. Pharmacotherapy, 2008. 28(5): p. 646-55.

[2]. Effects of epalrestat, an aldose reductase inhibitor, on diabetic neuropathy and gastroparesis. Intern Med, 2003. 42(8): p. 655-64.

[3]. Clinical investigation of epalrestat, an aldose reductase inhibitor, on diabetic neuropathy in Japan: multicenter study. Diabetic Neuropathy Study Group in Japan. J Diabetes Complications, 1996. 10(3): p. 168-72.

[4]. The aldose reductase inhibitor epalrestat exerts nephritic protection on diabetic nephropathy in db/db mice through metabolic modulation. Acta Pharmacol Sin. 2019 Jan;40(1):86-97.

[5]. Epalrestat increases intracellular glutathione levels in Schwann cells through transcription regulation. Redox Biol. 2013 Nov 19;2:15-21.

[6]. Epalrestat protects against diabetic peripheral neuropathy by alleviating oxidative stress and inhibiting polyol pathway. Neural Regen Res. 2016 Feb;11(2):345-51.

[7]. The Aldose Reductase Inhibitor Epalrestat Maintains Blood-Brain Barrier Integrity by Enhancing Endothelial Cell Function during Cerebral Ischemia. Mol Neurobiol. 2023 Jul;60(7):3741-3757.

Epalrestat is a monocarboxylic acid that is 1,3-thiazolidine which is substituted on the nitrogen by a carboxymethyl group, at positions 2 and 4 by thioxo and oxo groups, respectively, and at position 5 by a 2-methyl-3-phenylprop-2-en-1-ylidene group. It is an inhibitor of aldose reductase (which catalyses the conversion of glucose to sorbitol) and is used for the treatment of some diabetic complications, including neuropathy. It has a role as an EC 1.1.1.21 (aldehyde reductase) inhibitor. It is a member of thiazolidines and a monocarboxylic acid.
Epalrestat is under investigation in clinical trial NCT03244358 (Evaluation of Epalrestat in Metastatic Triple-negative Breast Cancer).
Epalrestat is an orally available, a non-competitive, reversible inhibitor of aldose reductase (AR), with potential antineoplastic, antioxidant, and anti-inflammatory activities. Upon oral administration, epalrestat non-competitively binds to AR, a polyol pathway enzyme that catalyzes reactive oxygen species (ROS)-initiated lipid peroxidation-generated lipid aldehydes and their glutathione conjugates, which have been implicated in the activation of transcription factors such as nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kappa B) and activator protein-1 (AP-1), which control the transcription of many inflammatory cytokines. Increased inflammatory cytokines and growth factors promote cell proliferation, a primary feature in the tumorigenesis process. AR is overexpressed in several oxidative stress- and inflammation-related conditions, including cancer.

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Epalrestat (ONO2235) is a reversible, competitive aldose reductase inhibitor (ARI) and the first clinically approved ARI for diabetic neuropathy [1][3]
- Its core mechanism of action involves inhibiting the polyol pathway: blocking AR reduces sorbitol accumulation in diabetic tissues (nerves, kidneys, blood vessels), alleviates osmotic stress, and suppresses oxidative damage [1][6]
- Beyond polyol pathway inhibition, Epalrestat exerts protective effects by increasing antioxidant capacity (GSH synthesis), regulating glucose metabolism, maintaining tight junction integrity, and reducing inflammatory responses [4][5][7]
- Indications: Epalrestat is indicated for the treatment of diabetic peripheral neuropathy, improving subjective symptoms (numbness, pain, paresthesia) and objective parameters (nerve conduction velocity) [1][3]
- Clinical data from a multicenter study in Japan showed that Epalrestat (150 mg/day) for 12 weeks achieved a symptom improvement rate of 68.3% and nerve conduction velocity improvement in 72.1% of diabetic neuropathy patients [3]

C15H13NO3S2

319.4

319.033

C, 56.41; H, 4.10; N, 4.39; O, 15.03; S, 20.08

82159-09-9

1549120

Pink to red solid powder

1.4±0.1 g/cm3

516.8±60.0 °C at 760 mmHg

210-217ºC

266.4±32.9 °C

0.0±1.4 mmHg at 25°C

1.706

2.02

115

1

5

4

21

519

0

C/C(=C\C1=CC=CC=C1)/C=C\2/C(=O)N(C(=S)S2)CC(=O)O

CHNUOJQWGUIOLD-NFZZJPOKSA-N

InChI=1S/C15H13NO3S2/c1-10(7-11-5-3-2-4-6-11)8-12-14(19)16(9-13(17)18)15(20)21-12/h2-8H,9H2,1H3,(H,17,18)/b10-7+,12-8-

2-[(5Z)-5-[(E)-2-Methyl-3-phenylprop-2-enylidene]-4-oxo-2-sulfanylidene-1,3-thiazolidin-3-yl]acetic acid

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 本产品在运输和储存过程中需避光。

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 mg/mL (6.26 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 20.0 mg/mL 澄清 DMSO 储备液加入900 μL 玉米油中，混合均匀。

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