Thrombopoietin receptor (TpoR)

当荧光素酶报告基因转染小鼠 BAF3 细胞时，艾曲波帕（0.002-50 μM；4 小时）表现出活性[1]。在 N2C-Tpo 细胞中，艾曲波帕（30 μM；120 分钟）对 p-STAT5 激活有影响[1]。在巨核细胞中，艾曲波帕（30 μM；120 分钟）刺激 p-STAT5[1]。艾曲波帕（0.1 nM-10 μM；30 分钟）刺激 BAF3/hTpoR 细胞的增殖[1]。当用elotrombopag (0.03-3 μM)处理十天时，骨髓CD34+细胞更有可能分化为CD41+巨核细胞[1]。 N2C-Tpo 细胞凋亡受艾曲波帕（0-3 μM；72 小时）影响[1]。 eloxacoum 的 MIC50 为 0.3 mg/L，可有效抑制肺炎球菌的生长，而对革兰氏阴性菌没有影响[3]。艾曲波帕（0-200 mg/L；24 小时）的 MIC50 为 1.5 mg/L，可抑制金黄色葡萄球菌的生长。当与 MIC50 为 1.2 mg/L 的万古霉素联合使用时，艾曲波帕的效力会增加[3]。艾曲波帕强烈促进 Huh7 细胞 G0/G1 期阻滞（0 或 10 μg/mL；72 小时）[5]。艾曲波帕在浓度为 0.1-100 μg/mL 时对 HCC 细胞系具有抗增殖作用，作用持续 72 小时[5]。

在黑猩猩中，艾曲波帕乙醇胺（10 mg/kg；每天口服一次，持续五天）表现出良好的耐受性[1]。埃洛曲波帕奥拉明（17.6 mg/kg；IP；每天一次，持续两天）使平均 S 大大降低。小鼠鼻腔感染有大量金黄色葡萄球菌[3]。

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当黑猩猩口服（每天10mg/kg，持续5天）时，艾曲波帕的体内活性表现为血小板数量增加了100%。总之，艾曲波帕选择性地与TpoR相互作用，而不与Tpo竞争，导致人骨髓祖细胞向巨核细胞的增殖和分化增加，血小板产量增加。这些结果表明，艾曲波帕和Tpo可能能够相加作用以增加血小板产量。1.

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黑猩猩重复剂量研究[1]
鉴于艾曲波帕在人类和黑猩猩血小板中表现出STAT激活的独特物种特异性，对五只雌性黑猩猩进行了为期5天的重复剂量安全性和药理学研究。黑猩猩通过口服强饲法单独服用赋形剂（n=2）或在赋形剂中服用艾曲波帕（n=3；每天10mg/kg）。在重复口服剂量后，艾曲波帕的给药耐受性良好。对血液学、凝血或临床化学参数没有不良影响。在接受艾曲波帕治疗的黑猩猩中，一只黑猩猩的血小板计数在最后一次给药后约1周增加了一倍多，另外两只黑猩猩的血小板数量增加了约1.5倍。血小板计数在达到峰值约2周后恢复到接近基线值（图6）。

萤光素酶报告基因检测[1]
洗涤BAF3/hTpoR或32D-mpl细胞，并在处理前用rmIL-3或rhTpo饥饿过夜。在Iscove改良的Dulbecco's培养基（IMDM）/0.5%胎牛血清（FBS）和30μM ZnCl2中，饥饿的亲本BAF3细胞（1×105个细胞/ml）在37°C、5%CO2下用Eltrombopag（0.002-50μM）或rhTpo（100ng/ml）处理4小时。将细胞在100μl裂解缓冲液（25mM tris、15%甘油、2%3-[（3-氨基丙基）二甲基铵]-1-丙磺酸、1%卵磷脂、1%牛血清白蛋白、4mM EGTA、8mM MgCl2、10mM二硫苏糖醇和0.4mM苯甲基磺酰氟，pH 7.8）中裂解15分钟，并加入96孔板（每孔30μl）。在使用化学光度计读取平板读数之前，立即向每个孔中加入Promega Steady Glow（100μl）。萤光素酶测定数据以四个重复孔的平均值和标准误差表示。

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Caspase-3和Caspase-7检测[1]
Caspase-Glo 3/7测定法（Promega）是一种发光测定法，用于测量半胱氨酸天冬氨酸蛋白酶-3和胱天蛋白酶-7的活性。添加胱天蛋白酶-Glo试剂导致细胞裂解，随后胱天蛋白酶切割底物并产生发光信号；发光量与存在的胱天蛋白酶的量成比例。细胞因子饥饿的N2C-Tpo细胞（最终浓度为1.4×105个细胞/ml）在白色观察板上生长，并在37°C下暴露于Eltrombopag（0.003-3μM）和/或rhTpo（1-100ng/ml）72小时。加入Caspase-Glo（100μl），在室温下孵育细胞90分钟。

细胞活力测定[1]
细胞类型： 鼠 BAF3 细胞
测试浓度： 0.002-50 μM
孵育时间：4小时
实验结果：用人TpoR有效抑制小鼠BAF3细胞，EC50值为0.27 μM。

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蛋白质印迹分析[1]
细胞类型： N2C-Tpo 细胞和 CD34+
测试浓度： N2C-Tpo 30 μM细胞; CD34+ 为 0、1、3 和 10 μM
孵育时间：N2C- Tpo 细胞为 120 分钟； CD34+ 30 分钟
实验结果：在 N2C-Tpo 细胞中处理后 60 分钟显示出激活的磷酸-STAT5 和最大信号强度。 CD34+ 治疗后 30 分钟，剂量依赖性激活 STAT5 磷酸化。

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细胞增殖检测[1]
细胞类型： BAF3/hTpoR 细胞
测试浓度： 0.1 nM-10 μM
孵育时间： 2 天
实验结果： 孵育 2 天后，EC50 为 0.03 μM，促进 BAF3/hTpoR 细胞增殖。

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细胞分化测定[1]
细胞类型： CD34+
测试浓度： 0.003, 0.01, 0.03, 0.1, 0.3, 1和 3 μM
孵育时间： 10 天
实验结果： 剂量依赖性刺激骨 m 分化

Animal/Disease Models: Female chimpanzees[1]
Doses: 10 mg/kg
Route of Administration: po (oral gavage); 10 mg/kg one time/day; for 5 days
Experimental Results: Appeared a goes up and then goes back tendency of platelet counts after treatment, and demonstrated no bad effects of hematology, coagulation, or clinical chemistry parameters on animal.

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Animal/Disease Models: C57BL/6 male mice (7 weeks, 20-22 g; injected S. aureus (5 × 108 CFU suspended in 40 µL PBS) into the nasal cavities)[3]
Doses: 17.6 mg/kg
Route of Administration: IP; one time/day for 2 days
Experimental Results: Dramatically decreased mean bacterial counts (5.0 × 106 CFU/lung) in the nasal infection model compared with control PBS (5.2 × 107 CFU/lung) mice.

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Female chimpanzees (approximately 7–8 years of age) were given either Eltrombopag (10 mg/kg) in aqueous 2% hydroxypropyl methylcellulose with 0.2% sodium lauryl sulfate vehicle or vehicle alone by oral gavage at a dose volume of 1 ml/kg. Chimpanzees were given five daily doses of vehicle alone (n = 2) or Eltrombopag (n = 3). Platelet counts and reticulated platelet counts were performed prior to, during, and following the treatment regimen. At the end of the study, all chimpanzees were returned to the stock colony.[1]

Although only limited pharmacokinetic sampling in three chimpanzees was performed, the data suggest that the pharmacodynamic signal of a change in platelet count from baseline for eltrombopag in the chimpanzees was detected at minimum concentration (Cmin), Cmax, and area under the curve (AUC) values of approximately 0.107 μg/ml, 0.525 μg/ml, and 12.1 μg h/ml, respectively. [1]

Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
Minimal information is available on the use of eltrombopag during breastfeeding. One breastfed infant with thrombocytosis at birth that was possibly prolonged by eltrombopag in milk. Until more data become available, romiplostim should be used with careful infant monitoring of infant blood parameters during breastfeeding, especially while nursing a newborn or preterm infant. The manufacturer recommends avoiding breastfeeding during the use of eltrombopag. Based on the drug’s half-life, the drug should be eliminated by the mother 8 days after the last dose.
◉ Effects in Breastfed Infants
An infant was born to a mother taking eltrombopag in a maximum dose of 100 mg during pregnancy. At birth, the infant had thrombocytosis, which persisted for a few weeks while the mother was breastfeeding. The extent of breastfeeding and the maternal dose were not stated. The authors felt that the persistence of thrombocytosis in the infant was possibly caused by eltrombopag in milk.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

[1]. Preclinical activity of eltrombopag (SB-497115), an oral, nonpeptide thrombopoietin receptor agonist. Stem Cells. 2009 Feb;27(2):424-30.

[2]. Discovery and characterization of a selective, nonpeptidyl thrombopoietin receptor agonist. Exp Hematol. 2005 Jan;33(1):85-93.

[3]. Repurposing Eltrombopag for Multidrug Resistant Staphylococcus aureus Infections. Antibiotics (Basel). 2021 Nov 9;10(11):1372.

[4]. Identification of Eltrombopag as a Repurposing Drug Against Staphylococcus epidermidis and its Biofilms. Curr Microbiol. 2021 Feb 21.

[5]. The Eltrombopag antitumor effect on hepatocellular carcinoma. Int J Oncol. 2015 Nov;47(5):1696-702.

See also: Eltrombopag Olamine (annotation moved to).

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Drug Indication
Revolade is indicated for the treatment of adult patients with primary immune thrombocytopenia (ITP) who are refractory to other treatments (e. g. corticosteroids, immunoglobulins) (see sections 4. 2 and 5. 1). Revolade is indicated for the treatment of paediatric patients aged 1 year and above with primary immune thrombocytopenia (ITP) lasting 6 months or longer from diagnosis and who are refractory to other treatments (e. g. corticosteroids, immunoglobulins) (see sections 4. 2 and 5. 1). Revolade is indicated in adult patients with chronic hepatitis C virus (HCV) infection for the treatment of thrombocytopenia, where the degree of thrombocytopenia is the main factor preventing the initiation or limiting the ability to maintain optimal interferon-based therapy (see sections 4. 4 and 5. 1). Revolade is indicated in adult patients with acquired severe aplastic anaemia (SAA) who were either refractory to prior immunosuppressive therapy or heavily pretreated and are unsuitable for haematopoietic stem cell transplantation (see section 5. 1). either refractory to prior immunosuppressive therapy or heavily pretreated and are unsuitable for haematopoietic stem cell transplantation (see section 5. 1).

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Eltrombopag is a hydrazine in which each nitrogen atom is substituted, one by a 3'-carboxy-2-hydroxy[1,1'-biphenyl]-3-yl group and the other by a 1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydro-4H-pyrazol-4-ylidene group. A small molecule agonist of the c-mpl (TpoR) receptor (the physiological target of the hormone thrombopoietin), it has been developed as a medication for conditions that lead to thrombocytopenia (abnormally low platelet counts). It has a role as a thrombopoietin receptor agonist and a xenobiotic. It is a member of hydrazines, a member of pyrazoles and a member of benzoic acids.
Eltrombopag is a Thrombopoietin Receptor Agonist. The mechanism of action of eltrombopag is as a Thrombopoietin Receptor Agonist, and Organic Anion Transporting Polypeptide 1B1 Inhibitor, and Breast Cancer Resistance Protein Inhibitor, and UGT1A1 Inhibitor, and UGT1A3 Inhibitor, and UGT1A4 Inhibitor, and UGT1A6 Inhibitor, and UGT1A9 Inhibitor, and UGT2B7 Inhibitor, and UGT2B15 Inhibitor. The physiologic effect of eltrombopag is by means of Increased Megakaryocyte Maturation, and Increased Platelet Production.
See also: Romiplostim (annotation moved to); Eltrombopag (annotation moved to).

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Drug Indication
Revolade is indicated for the treatment of adult patients with primary immune thrombocytopenia (ITP) who are refractory to other treatments (e. g. corticosteroids, immunoglobulins) (see sections 4. 2 and 5. 1). Revolade is indicated for the treatment of paediatric patients aged 1 year and above with primary immune thrombocytopenia (ITP) lasting 6 months or longer from diagnosis and who are refractory to other treatments (e. g. corticosteroids, immunoglobulins) (see sections 4. 2 and 5. 1). Revolade is indicated in adult patients with chronic hepatitis C virus (HCV) infection for the treatment of thrombocytopenia, where the degree of thrombocytopenia is the main factor preventing the initiation or limiting the ability to maintain optimal interferon-based therapy (see sections 4. 4 and 5. 1). Revolade is indicated in adult patients with acquired severe aplastic anaemia (SAA) who were either refractory to prior immunosuppressive therapy or heavily pretreated and are unsuitable for haematopoietic stem cell transplantation (see section 5. 1).

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Eltrombopag is a first-in-class, orally bioavailable, small-molecule, nonpeptide agonist of the thrombopoietin receptor (TpoR), which is being developed as a treatment for thrombocytopenia of various etiologies. In vitro studies have demonstrated that the activity of eltrombopag is dependent on expression of TpoR, which activates the signaling transducers and activators of transcription (STAT) and mitogen-activated protein kinase signal transduction pathways. The objective of this preclinical study is to determine if eltrombopag interacts selectively with the TpoR to facilitate megakaryocyte differentiation in platelets. Functional thrombopoietic activity was demonstrated by the proliferation and differentiation of primary human CD34(+) bone marrow cells into CD41(+) megakaryocytes. Measurements in platelets in several species indicated that eltrombopag specifically activates only the human and chimpanzee STAT pathways. [1]

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The continuous rise of antimicrobial resistance urgently demands new therapeutic agents for human health. Drug repurposing is an attractive strategy that could significantly save time delivering new antibiotics to clinics. We screened 182 US Food and Drug Administration (FDA)-approved drugs to identify potential antibiotic candidates against Staphylococcus aureus, a major pathogenic bacterium. This screening revealed the significant antibacterial activity of three small molecule drugs against S. aureus: (1) LDK378 (Ceritinib), an anaplastic lymphoma kinase (ALK) inhibitor for the treatment of lung cancer, (2) dronedarone HCl, an antiarrhythmic drug for the treatment of atrial fibrillation, and (3) eltrombopag, a thrombopoietin receptor agonist for the treatment of thrombocytopenia. Among these, eltrombopag showed the highest potency against not only a drug-sensitive S. aureus strain but also 55 clinical isolates including 35 methicillin-resistant S. aureus (Minimum inhibitory concentration, MIC, to inhibit 50% growth [MIC50] = 1.4-3.2 mg/L). Furthermore, we showed that eltrombopag inhibited bacterial growth in a cell infection model and reduced bacterial loads in infected mice, demonstrating its potential as a new antibiotic agent against S. aureus that can overcome current antibiotic resistance.[3]

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Staphylococcus epidermidis is a common cause of nosocomial infections, and readily adheres to medical apparatus to form biofilms consisting of highly resistant persister cells. Owing to the refractory infections caused by S. epidermidis biofilms and persisters in immunosuppressed patients, it is crucial to develop new antimicrobials. In the present study, we analyzed the antimicrobial effects of the thrombopoietin receptor agonist eltrombopag (EP) against S. epidermidis planktonic cells, biofilms, and persister cells. EP was significantly toxic to S. epidermidis with the minimal inhibitory concentration of 8 μg/ml, and effectively inhibited the biofilms and persisters in a strain-dependent manner. In addition, EP was only mildly toxic to mammalian cells after 12 to 24 h treatment. It also partially synergized with vancomycin against S. epidermidis, which enhanced its antimicrobial effects and reduced its toxicity to mammalian cells. Taken together, EP is a potential antibiotic for treating refractory infections caused by S. epidermidis.[4]

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Currently, sorafenib is the only available chemotherapeutic agent for advanced hepatocellular carcinoma (HCC), but it cannot be used in patients with liver cirrhosis (LC) or thrombocytopenia. In these cases, sorafenib is likely effective if given in combination with treatments that increase the number of platelets, such as thrombopoietin (TPO) receptor agonists. Increasing the platelet count via TPO treatment resulted in reduction of LC. Eltrombopag (EP), a TPO receptor agonist, has been reported to have antitumor effects against certain cancers, despite their lack of TPO receptor expression. We hypothesized that EP may possess antitumor activity against HCC in addition to its ability to suppress hepatic fibrosis by increasing the platelet count. In the present study, the antitumor activity of EP was examined by assessing the inhibition of cell proliferation and then ascertaining the ability of iron supplementation to reverse these effects in HepG2, Hep3B and Huh7 cells. In addition, a cell cycle assay was performed using flow cytometry, and signal transduction was evaluated by analyzing cell cycle-related protein expression. The results of EP were compared with those of the most common iron chelator, deferoxamine (DFO). The combined effect of EP and sorafenib was also assessed. The results revealed that EP exerts antitumor activity in HCC that is mediated by the modulation of intracellular iron content. EP suppressed the expression of the cell cycle-related protein cyclin D1 and elicited cell cycle arrest in the G0/G1 phase. The activity of EP was comparable to that of DFO in HCC, and EP did not compete with sorafenib at low concentrations. In conclusion, our findings suggest that EP is a good candidate chemotherapeutic agent for the treatment of HCC in patients with LC and thrombocytopenia.[5]

C29H36N6O6

564.63

564.269

C, 61.69; H, 6.43; N, 14.88; O, 17.00

496775-62-3

Eltrombopag;496775-61-2;(E/Z)-Eltrombopag-13C4;1217230-31-3

135449331

Purple to black solid powder

3.127

209.05

7

11

7

41

822

0

PLILLUUXAVKBPY-SBIAVEDLSA-N

InChI=1S/C25H22N4O4.2C2H7NO/c1-14-10-11-19(12-15(14)2)29-24(31)22(16(3)28-29)27-26-21-9-5-8-20(23(21)30)17-6-4-7-18(13-17)25(32)33;2*3-1-2-4/h4-13,26,30H,1-3H3,(H,32,33);2*4H,1-3H2/b27-22-

(Z)-3-(2-(1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydro-4H-pyrazol-4-ylidene)hydrazinyl)-2-hydroxy-[1,1-biphenyl]-3-carboxyiic acid;2-aminoethan-1-ol (1:2)

SB-497115-GR Ethanolamine, SB497115GR; Eltrombopag ethanolamine; Eltrombopag Mono-Ethanolamine; CHEMBL461806; SCHEMBL2236368; SCHEMBL2236372; SB497115; SB-497115; SB 497115; trade name: PROMACTA

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 请将本产品存放在密封且受保护的环境中，避免吸湿/受潮。

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: ≥ 1.07 mg/mL (1.90 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 10.7 mg/mL澄清的DMSO储备液加入到400 μL PEG300中，混匀；再向上述溶液中加入50 μL Tween-80，混匀；然后加入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: 1.07 mg/mL (1.90 mM) 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), 悬浊液; 超声助溶。
例如，若需制备1 mL的工作液，可将 100 μL 10.7mg/mL澄清的DMSO储备液加入到900μL 20％SBE-β-CD生理盐水中，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: 10 mg/mL (17.71 mM) in 50% PEG300 50% Saline (这些助溶剂从左到右依次添加，逐一添加), 悬浮液； 超声助溶 (<60°C).
*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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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));
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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；
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