Phenamil
目录号 : GC13994A TRPP3 inhibitor
Cas No.:1161-94-0
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >98.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
| Cell experiment [1]: | |
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Cell lines |
Osteoblast-like MC3T3-E1 cells |
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Preparation Method |
The cells were maintained in regular growth medium, i.e. α-minimal essential medium supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin G and 100 mg/ml streptomycin. The culture medium, with or without 10 µM phenamil, was replaced every 3-4 days. |
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Reaction Conditions |
10 µM for 14 days |
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Applications |
Phenamil-treated and -untreated control cells showed a statistically significant increase in cell proliferation from day 7 to day 14. |
| Animal experiment [2]: | |
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Animal models |
Male Sprague-Dawley rats |
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Preparation Method |
Miniature osmotic infusion pumps which deliver saline-DMSO (50% saline-50% DMSO) solution or phenamil in saline-DMSO were implanted subcutaneously in male Sprague-Dawley rats (age, 6 to 7 weeks; weight, 250 to 300 g) between the scapulae and connected via a catheter advanced into the left carotid artery. Four rats were used for each condition. Phenamil (15 or 30 mg/kg of body weight/day) was administered continuously for 21 days at an infusion rate of 1 ml/h. The rats were subjected to hypobaric hypoxia (10.5% oxygen) or normoxia (room air) for 21 days. |
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Dosage form |
15 or 30 mg/kg, Miniature osmotic infusion pump |
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Applications |
Phenamil reduces chronic-hypoxia-induced pulmonary artery hypertension (PAH). |
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References: [1] : Lo K W H, Ulery B D, Kan H M, et al. Evaluating the feasibility of utilizing the small molecule phenamil as a novel biofactor for bone regenerative engineering[J]. Journal of tissue engineering and regenerative medicine, 2014, 8(9): 728-736. [2] :Chan M C, Weisman A S, Kang H, et al. The amiloride derivative phenamil attenuates pulmonary vascular remodeling by activating NFAT and the bone morphogenetic protein signaling pathway[J]. Molecular and cellular biology, 2011, 31(3): 517-530. |
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Phenamil is a selective inhibitor of transient receptor potentials (TRPs) and acid-sensing ion channels (ASICs) [1]. Phenamil is an amiloride analogs and a more potent and less reversible epithelial sodium channel (ENaC) blocker with an IC50 of 400 nM [2]. Phenamil inhibited TRPP3 -mediated Ca2+ transport with an IC50 of 140 nM in a Ca2+ uptake assay [3].
Phenamil (10 μM) Treatment of M2-10B4 (M2) mouse (mesenchymal stem cells) MSCs for 6 days with 10 μM phenamil markedly stimulated expression of the genes encoding ALP, Runx2, OCN, and osterix. Phenamil induces osteogenic gene expression and mineralization in both MSC and primary calvariae organ cultures [4]. Phenamil (10 μM) induced the expression of PPARγ within 24 h in 3T3-F442A preadipocyte cell lines. Phenamil act as a new proadipogenic compound [5].
Phenamil (15 or 30 mg/kg; 21 days) reduced chronic-hypoxia-induced pulmonary artery hypertension (PAH). Infusion of phenamil during hypoxia treatment inhibited pulmonary vascular remodeling [6].
References:
[1]. Chan M C, Weisman A S, Kang H, et al. The amiloride derivative phenamil attenuates pulmonary vascular remodeling by activating NFAT and the bone morphogenetic protein signaling pathway[J]. Molecular and cellular biology, 2011, 31(3): 517-530.
[2]. Hirsh A J, Molino B F, Zhang J, et al. Design, synthesis, and structure? activity relationships of novel 2-substituted pyrazinoylguanidine epithelial sodium channel blockers: drugs for cystic fibrosis and chronic bronchitis[J]. Journal of medicinal chemistry, 2006, 49(14): 4098-4115.
[3]. Dai X Q, Ramji A, Liu Y, et al. Inhibition of TRPP3 channel by amiloride and analogs[J]. Molecular pharmacology, 2007, 72(6): 1576-1585.
[4]. Park K W, Waki H, Kim W K, et al. The small molecule phenamil induces osteoblast differentiation and mineralization[J]. Molecular and cellular biology, 2009, 29(14): 3905-3914.
[5]. Park K W, Waki H, Choi S P, et al. The small molecule phenamil is a modulator of adipocyte differentiation and PPARγ expression [S][J]. Journal of lipid research, 2010, 51(9): 2775-2784.
[6]. Chan M C, Weisman A S, Kang H, et al. The amiloride derivative phenamil attenuates pulmonary vascular remodeling by activating NFAT and the bone morphogenetic protein signaling pathway[J]. Molecular and cellular biology, 2011, 31(3): 517-530.
Phenamil 是瞬时受体电位 (TRP) 和酸敏感离子通道 (ASIC) 的选择性抑制剂[1]。 Phenamil 是一种阿米洛利类似物，是一种更有效且可逆性更差的上皮钠通道 (ENaC) 阻断剂，IC50 为 400 nM [2]。在 Ca2+ 摄取试验中，Phenamil 抑制 TRPP3 介导的 Ca2+ 转运，IC50 为 140 nM [3]。
Phenamil (10 μM) 用 10 μM phenamil 处理 M2-10B4 (M2) 小鼠（间充质干细胞）MSCs 6 天，可显着刺激编码 ALP、Runx2、OCN 和 osterix 的基因的表达。 Phenamil 在 MSC 和原代颅骨器官培养物中诱导成骨基因表达和矿化[4]。 Phenamil (10 μM) 在 24 小时内诱导 3T3-F442A 前脂肪细胞系中 PPARγ 的表达。 Phenamil 作为一种新的促脂肪形成化合物[5]。
Phenamil（15 或 30 mg/kg；21 天）可减少慢性缺氧引起的肺动脉高压 (PAH)。缺氧治疗期间输注非那米抑制肺血管重构[6]。
| Cas No. | 1161-94-0 | SDF | |
| 化学名 | (E)-3,5-diamino-N-(amino(phenylamino)methylene)-6-chloropyrazine-2-carboxamide methanesulfonate | ||
| Canonical SMILES | ClC(N=C1C(/N=C(N)/NC2=CC=CC=C2)=O)=C(N)N=C1N.OS(=O)(C)=O | ||
| 分子式 | C12H12ClN7O.CH3SO3H | 分子量 | 401.83 |
| 溶解度 | DMF: 0.1 mg/ml,DMSO: 1 mg/ml,DMSO:PBS (pH 7.2) (1:1): 0.5 mg/ml | 储存条件 | Store at -20°C |
| General tips | 请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。 储备液的保存方式和期限：-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。 为了提高溶解度，请将管子加热至37℃，然后在超声波浴中震荡一段时间。 |
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| Shipping Condition | 评估样品解决方案：配备蓝冰进行发货。所有其他可用尺寸：配备RT，或根据请求配备蓝冰。 | ||
| 制备储备液 | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 2.4886 mL | 12.4431 mL | 24.8861 mL |
| 5 mM | 0.4977 mL | 2.4886 mL | 4.9772 mL |
| 10 mM | 0.2489 mL | 1.2443 mL | 2.4886 mL |
| 第一步：请输入基本实验信息（考虑到实验过程中的损耗，建议多配一只动物的药量） | ||||||||||
| 给药剂量 | mg/kg |  |
动物平均体重 | g | |
每只动物给药体积 | ul | |
动物数量 | 只 |
| 第二步：请输入动物体内配方组成（配方适用于不溶于水的药物；不同批次药物配方比例不同，请联系GLPBIO为您提供正确的澄清溶液配方） | ||||||||||
| % DMSO % % Tween 80 % saline | ||||||||||
| 计算重置 | ||||||||||
计算结果:
工作液浓度： mg/ml；
DMSO母液配制方法： mg 药物溶于 μL DMSO溶液（母液浓度 mg/mL，
体内配方配制方法：取 μL DMSO母液，加入 μL PEG300，混匀澄清后加入μL Tween 80，混匀澄清后加入 μL saline，混匀澄清。
1. 首先保证母液是澄清的；
2.
一定要按照顺序依次将溶剂加入，进行下一步操作之前必须保证上一步操作得到的是澄清的溶液，可采用涡旋、超声或水浴加热等物理方法助溶。
3. 以上所有助溶剂都可在 GlpBio 网站选购。
Mineralized nanofibrous scaffold promotes phenamil-induced osteoblastic differentiation while mitigating adipogenic differentiation
J Tissue Eng Regen Med2020 Mar;14(3):464-474.PMID: 31840422DOI: 10.1002/term.3007
Large bone defects represent a significant unmet medical challenge. Cost effectiveness and better stability make small molecule organic compounds a more promising alternative compared with biomacromolecules, for example, growth factors/hormones, in regenerative medicine. However, one common challenge for the application of these small compounds is their side-effect issue. Phenamil is emerging as an intriguing small molecule to promote bone repair by strongly activating bone morphogenetic protein signaling pathway. In addition to osteogenesis, phenamil also induces significant adipogenesis based on some in vitro studies, which is a concern that impedes it from potential clinical applications. Besides the soluble chemical signals, cellular differentiation is heavily dependent on the microenvironments provided by the 3D scaffolds. Therefore, we developed a 3D nanofibrous biomimetic scaffold-based strategy to harness the phenamil-induced stem cell lineage differentiation. Based on the gene expression, alkaline phosphatase activity, and mineralization data, we indicated that bone-matrix mimicking mineralized-gelatin nanofibrous scaffold effectively improved phenamil-induced osteoblastic differentiation, while mitigating the adipogenic differentiation in vitro. In addition to normal culture conditions, we also indicated that mineralized matrix can significantly improve phenamil-induced osteoblastic differentiation in simulated inflammatory condition. In viewing of the crucial role of mineralized matrix, we developed an innovative and facile mineral deposition-based strategy to sustain release of phenamil from 3D scaffolds for efficient local bone regeneration. Overall, our study demonstrated that biomaterials played a crucial role in modulating small molecule drug phenamil-induced osteoblastic differentiation by providing a bone-matrix mimicking mineralized gelatin nanofibrous scaffolds.
Phenamil enhances the adipogenic differentiation of hen preadipocytes
Cell Biol Int2016 Oct;40(10):1123-8.PMID: 27460177DOI: 10.1002/cbin.10651
A study was conducted to examine the effect of phenamil on adipogenic differentiation and expression of key adipogenic transcripts in hen preadipocytes. Preadipocytes were isolated from 20-week old Single Comb White Leghorn hens (Gallas gallus, Lohman strain). The experiment lasted for 48 h and had six treatments. Non-treated control (C) cells, cells treated with dexamethasone, 3-isobutyl-1-methylxanthine, insulin, and oleic acid (DMIOA) (T1), DMIOA + 15 米M phenamil (T2), DMIOA + 30 米M phenamil (T3), 15 米M phenamil alone (T4), and 30 米M phenamil alone (T5). Neutral lipid accumulation and the mRNA expression of key adipogenic transcripts were measured in all treatments and compared. Lipid accumulation was detected in T1, T2, and T3 only. Expression of peroxisome proliferator receptor-activator gamma 2 (PPAR汿), the core enhancer binding protein 汿(C/EBP汿, C/EBP汿 fatty acid binding protein 4 (FABP4), and lipoprotein lipase (LPL) as well as ETS variant 4 (ETV4) and 5 was higher (P < 0.05) in T2, T3, T4, and T5 compared to C. Expression of these transcripts was higher (P < 0.05) in T2 and T3 compared to T4 and T5. The core enhancer binding protein 汿 C/EBP汿 and FABP4 were highly expressed (P < 0.05) in T1 compared to C. However, the expression of PPAR汿, LPL, and ETV4 and ETV5 was not significantly different. Expression of C/EBP汿 C/EBP汿 and FABP4 was higher (P < 0.05) in T2 and T3 compared to T1. Expression of sterol regulatory element binding protein 1 (SREBP1) and leptin receptor (LEPR) was not significantly different among the treatments. In conclusion, phenamil enhances DMIOA-induced adipogenic differentiation of hen preadipocytes but does not induce adipogenesis by itself.
Osteogenic Differentiation Effect of BMP-9 with Phenamil and Simvastatin on Intact Human Amniotic Epithelial Stem Cells
Iran Biomed J2022 Nov 1;26(6):463-74.PMID: 36437797DOI: 10.52547/ibj.3748
Background: Background: Bone tissue engineering has shown to be a promising strategy for repairing bone defects without causing harmful side effects to the patient. Three main building blocks of tissue engineering, including seeding cells, scaffold, and signaling molecules, are required for adequate bone regeneration. The human amniotic membrane (hAM) is the innermost of the placental membranes. In addition to providing a source of stem cells and growth factors, hAM has several features that make it an appropriate scaffold containing stem cells for use in tissue engineering purposes. The present investigation aimed to assess the effect of bone morphogenetic protein-9 (BMP-9) combined with phenamil and simvastatin on osteogenic induction of hAM with its human amniotic membrane epithelial cells (hAECs).
Method: Methods: Using six different osteogenic medium (OMs), we cultured hAM for 14 days. The basic OMs were chosen as the first group and other media were made by adding BMP-9, phenamil, simvastatin, BMP-9 alongside phenamil, and BMP-9 alongside simvastatin to the basic OMs. Finally, viability assay, tissue mineralization, calcium and phosphate content determination, and measurement of lactic acid dehydrogenase (LDH), and alkaline phosphatase (ALP) activity were performed.
Results: Results: Among all study groups, groups containing simvastatin showed a significantly lower level of viability. Although all media could induce osteogenic features, the hAECs cultured in media containing BMP-9 and phenamil demonstrated a wider area of mineralization and a significantly higher level of calcium and phosphate content, LDH, and ALP activity.
Conclusion: Conclusion: Our findings indicated that the use of phenamil together with BMP-9 could synergistically show in situ osteogenic induction in hAECs, which could be a new insight into translational medicine.
Phenamil, an amiloride analogue, inhibits differentiation of Friend murine erythroleukemic cells
Am J Physiol1988 Jan;254(1 Pt 1):C122-9.PMID: 3422134DOI: 10.1152/ajpcell.1988.254.1.C122
Amiloride has been reported to inhibit Friend murine erythroleukemic (MEL) cell commitment to differentiate by inhibiting the MEL cell plasma membrane Na+-Ca2+ antiporter (R. L. Smith, I. G. Macara, R. Levenson, D. Housman, and L. Cantley. J. Biol. Chem. 257: 773-780, 1982). We therefore screened a series of amiloride analogues to determine whether a more potent and specific inhibitor of MEL cell differentiation could be found. In our experiments, as in those of Lubin (J. Cell. Physiol. 124: 539-544, 1985), amiloride itself did not inhibit MEL cell differentiation. However, we did find that the amiloride analogue phenamil reversibly inhibits dimethyl sulfoxide (DMSO)-induced MEL cell commitment to differentiate with a K1/2 of 2.5-5.0 microM (in plasma clot assay). At an extracellular concentration of 15 microM, phenamil inhibits commitment to differentiate by approximately 90% in the plasma clot assay while having a minimal effect on growth. Phenamil is not metabolized but is rapidly taken up by MEL cells. Phenamil was most effective as an inhibitor when present during the first 12 h of DMSO treatment, indicating that phenamil affects the early commitment process rather than later steps involved in hemoglobin synthesis. Phenamil does not, however, inhibit the early differentiation-induced decrease in [Na+]i and the concomitant drop in the Na+-K+ pump rate. A specific binding site for phenamil is suggested because some analogues in which the phenamil structure is slightly modified are unable to inhibit differentiation.
Phenamil, an amiloride derivative, restricts long bone growth and alters keeled-sternum bone architecture in growing chickens
Poult Sci2017 Jul 1;96(7):2471-2479.PMID: 28340021DOI: 10.3382/ps/pex034
"Broiler-type" chickens are fast-grow-ing, heavy-bodied birds with high demands on bone quality. Phenamil increased mineralization in cultured murine mesenchymal stem cells. Phenamil effects were tested in 2 groups of weight and gender matched day-old broiler chickens (n = 13). Oral administration of 30 mg phenamil/kg body weight d 1 to 13 reduced growth of chicks d 5 to 14 (P = 0.002); with phenamil-treated (PT) chick body weight being 84% of vehicle-treated (VT) chicks' body weight on d 14. Tissues collected on d 15 showed that femur lengths and widths did not differ, but tibias from PT chicks were 6% shorter (P = 0.002) and 13% narrower (P = 0.012) with 18% thinner tibial cross-sections (P < 0.008) than in VT chicks. Angles of the caudal aspect of the anterior surface of keeled-sternums were 166ｿin PT chicks, flatter than the 148ｿfound in VT chicks (P = 0.000). Total mineral content of both tibia and femur were lower in PT chicks (P = 0.005 for both). Bone Ca, P, and Mg (ppm) in ash were similar, but Ca:P was lower (1.70 vs 1.75) in PT versus VT chicks (P < 0.05). Osteocalcin was ?20% lower (P = 0.020), PINP was ?45% higher (P = 0.000) in PT chicks. Carboxy-terminal telopeptide type I collagen (ICTP) and cross-linked N-telopeptide of type I collagen (NTX1) were similar in the 2 groups. Phenamil had unexpected and detrimental effects on bone formation in growing broiler chicks, reducing linear skeletal growth and markedly changing bone architecture.