Azamulin
(Synonyms: 阿扎莫林,Antibiotic TDM 85-530) 目录号 : GC42888A selective CYP3A inhibitor
Cas No.:76530-44-4
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
Azamulin is a selective, irreversible inhibitor of cytochrome P450 (CYP) 3A isoforms (IC50 values range from 26 to 240 nM for CYP3A4 and CYP3A4/5 from different sources). [1] It is at least 50-fold less potent against CYP2J2 and 100-fold less effective against all other CYP isoforms. [1] Azamulin potently blocks the hydroxylation of testosterone and midazolam by CYP3A4.[2]
Reference:
[1]. Stresser, D.M., Broudy, M.I., Ho, T., et al. Highly selective inhibition of human CYP3Aa in vitro by azamulin and evidence that inhibition is irreversible. Drug Metabolism and Disposition 32(1), 105-112 (2004).
[2]. Lim, H.K., Duczak, N., Jr., Brougham, K., et al. Automated screening with confirmation of mechanism-based inactivation of CYP3A4, CYP2C9, CYP2C19, CYP2D6, and CYP1A2 in pooled human liver microsomes. Drug Metabolism and Disposition 33(8), 1211-1219 (2005).
| Cas No. | 76530-44-4 | SDF | |
| 别名 | 阿扎莫林,Antibiotic TDM 85-530 | ||
| 化学名 | (3aS,4R,5S,6R,8R,9R,9aR,10R)-6-ethyldecahydro-5-hydroxy-4,6,9,10-tetramethyl-1-oxo-3a,9-propano-3aH-cyclopentacycloocten-8-yl ester 2-[(3-amino-1H-1,2,4-triazol-5-yl)thio]-acetic acid | ||
| Canonical SMILES | O=C1CC[C@@]23[C@@H](C)[C@H](O)[C@@](CC)(C)C[C@@H](OC(CSC4=NN=C(N)N4)=O)[C@](C)([C@H](C)CC3)[C@@]21[H] | ||
| 分子式 | C24H38N4O4S | 分子量 | 478.7 |
| 溶解度 | 20mg/mL in ethanol, or in DMSO, or in DMF | 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 | 2.089 mL | 10.445 mL | 20.8899 mL |
| 5 mM | 0.4178 mL | 2.089 mL | 4.178 mL |
| 10 mM | 0.2089 mL | 1.0445 mL | 2.089 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。
Application of Azamulin to Determine the Contribution of CYP3A4/5 to Drug Metabolic Clearance Using Human Hepatocytes
Drug Metab Dispos 2020 Sep;48(9):778-787.PMID:32532738DOI:10.1124/dmd.120.000017.
Early determination of CYP3A4/5 contribution to the clearance of new chemical entities is critical to inform on the risk of drug-drug interactions with CYP3A inhibitors and inducers. Several in vitro approaches (recombinant P450 enzymes, correlation analysis, chemical and antibody inhibition in human liver microsomes) are available, but they are usually labor-intensive and/or suffer from specific limitations. In the present study, we have validated the use of Azamulin as a specific CYP3A inhibitor in human hepatocytes. Azamulin (3 µM) was found to significantly inhibit CYP3A4/5 (>90%), whereas other P450 enzymes were not affected (less than 20% inhibition). Because human hepatocytes were used as a test system, the effect of Azamulin on other key drug-metabolizing enzymes (aldehyde oxidase, carboxylesterase, UGT, flavin monooxygenase, and sulfotransferase) was also investigated. Apart from some UGTs showing minor inhibition (∼20%-30%), none of these non-P450 enzymes were inhibited by Azamulin. Use of CYP3A5-genotyped human hepatocyte batches in combination with CYP3cide demonstrated that Azamulin (at 3 µM) inhibits both CYP3A4 and CYP3A5 enzymes. Finally, 11 compounds with known in vivo CYP3A4/5 contribution have been evaluated in this human hepatocyte assay. Results showed that the effect of Azamulin on the in vitro intrinsic clearance of these known CYP3A4/5 substrates was predictive of the in vivo CYP3A4/5 contribution. Overall, the study showed that human hepatocytes treated with Azamulin provide a fast and accurate estimation of CYP3A4/5 contribution in metabolic clearance of new chemical entities. SIGNIFICANCE STATEMENT: Accurate estimation of CYP3A4/5 contribution in drug clearance is essential to anticipate risk of drug-drug interactions and select the appropriate candidate for clinical development. The present study validated the use of Azamulin as selective CYP3A4/5 inhibitor in suspended human hepatocytes and demonstrated that this novel approach provides a direct and accurate determination of the contribution of CYP3A4/5 (fraction metabolized by CYP3A4/5) in the metabolic clearance of new chemical entities.
Structural characterization of the homotropic cooperative binding of Azamulin to human cytochrome P450 3A5
J Biol Chem 2022 May;298(5):101909.PMID:35398097DOI:10.1016/j.jbc.2022.101909.
Cytochrome P450 3A4 and 3A5 catalyze the metabolic clearance of a large portion of therapeutic drugs. Azamulin is used as a selective inhibitor for 3A4 and 3A5 to define their roles in metabolism of new chemical entities during drug development. In contrast to 3A4, 3A5 exhibits homotropic cooperativity for the sequential binding of two Azamulin molecules at concentrations used for inhibition. To define the underlying sites and mechanisms for cooperativity, an X-ray crystal structure of 3A5 was determined with two Azamulin molecules in the active site that are stacked in an antiparallel orientation. One Azamulin resides proximal to the heme in a pose similar to the 3A4-azamulin complex. Comparison to the 3A5 apo structure indicates that the distal Azamulin in 3A5 ternary complex causes a significant induced fit that excludes water from the hydrophobic surfaces of binding cavity and the distal Azamulin, which is augmented by the stacking interaction with the proximal Azamulin. Homotropic cooperativity was not observed for the binding of related pleuromutilin antibiotics, tiamulin, retapamulin, and lefamulin, to 3A5, which are larger and unlikely to bind in the distal site in a stacked orientation. Formation of the 3A5 complex with two Azamulin molecules may prevent time-dependent inhibition that is seen for 3A4 by restricting alternate product formation and/or access of reactive intermediates to vulnerable protein sites. These results also contribute to a better understanding of sites for cooperative binding and the differential structural plasticity of 3A5 and 3A4 that contribute to differential substrate and inhibitor binding.
Structural Insights into the Interaction of Cytochrome P450 3A4 with Suicide Substrates: Mibefradil, Azamulin and 6',7'-Dihydroxybergamottin
Int J Mol Sci 2019 Aug 30;20(17):4245.PMID:31480231DOI:10.3390/ijms20174245.
Human cytochrome P450 3A4 (CYP3A4) is the most important drug-metabolizing enzyme. Some drugs and natural compounds can act as suicide (mechanism-based) inactivators of CYP3A4, leading to unanticipated drug-drug interactions, toxicity and therapeutic failures. Despite significant clinical and toxicological implications, the mechanism-based inactivation remains incompletely understood. This study provides the first direct insights into the interaction of CYP3A4 with three suicide substrates: mibefradil, an antihypertensive drug quickly withdrawn from the market; a semi-synthetic antibiotic Azamulin; and a natural furanocoumarin, 6',7'-dihydroxybergamottin. Novel structural findings help better understand the suicide substrate binding and inhibitory mechanism, and can be used to improve the predictability of the binding ability, metabolic sites and inhibitory/inactivation potential of newly developed drugs and other chemicals relevant to public health.
Highly selective inhibition of human CYP3Aa in vitro by Azamulin and evidence that inhibition is irreversible
Drug Metab Dispos 2004 Jan;32(1):105-12.PMID:14709627DOI:10.1124/dmd.32.1.105.
Azamulin [14-O-(5-(2-amino-1,3,4-triazolyl)thioacetyl)-dihydromutilin] is an azole derivative of the pleuromutilin class of antiinfectives. We tested the inhibition potency of Azamulin toward 18 cytochromes P450 using human liver microsomes or microsomes from insect cells expressing single isoforms. In a competitive inhibition model, IC(50) values for CYP3A (0.03-0.24 microM) were at least 100-fold lower than all other non-CYP3A enzymes except CYP2J2 ( approximately 50-fold lower). The IC(50) value with heterologously expressed CYP3A4 was 15-fold and 13-fold less than those of CYP3A5 and CYP3A7, respectively. The reference inhibitor ketoconazole was less selective and exhibited potent inhibition (IC(50) values <10 microM) for CYP1A1, CYP1B1, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP4F2, and CYP4F12. Inhibition of CYP3A by Azamulin appeared sigmoidal and well behaved with the substrates 7-benzyloxy-4-trifluoromethylcoumarin, testosterone, and midazolam. Preincubation of 4.8 microM Azamulin in the presence of NADPH for 10 min inhibited approximately 95% of testosterone 6beta-hydroxylase activity compared with preincubation in the absence of NADPH. Catalytic activities of CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP2E1 were unaffected by similar experiments. Incubation of Azamulin with heterologously expressed CYP3A4 yielded a type I binding spectrum with a spectral dissociation constant of 3.5 microM, whereas no interaction was found with CYP2D6. Azamulin exhibited good chemical stability when stored in acetonitrile for up to 12 days. Aqueous solubility was found to be >300 microM. Azamulin represents an important new chemical tool for use in characterizing the contribution of CYP3A to the metabolism of xenobiotics.
Roles of human cytochrome P450 3A4/5 in dexamethasone 6β-hydroxylation mediated by liver microsomes and humanized liver in chimeric mice metabolically suppressed with Azamulin
Drug Metab Pharmacokinet 2023 Mar 6;50:100504.PMID:37031476DOI:10.1016/j.dmpk.2023.100504.
The urinary metabolic ratio of 6β-hydroxydexamethasone to dexamethasone reportedly acts as a noninvasive marker for human cytochrome P450 (P450) 3A4/5, which is induced by rifampicin in humanized-liver mice. In the current study, the pharmacokinetics of dexamethasone in humanized-liver mice after intravenous administration (10 mg/kg) were investigated using Azamulin (a time-dependent P450 3A4/5 inhibitor). After intravenous dexamethasone administration, significant differences were observed in the time-dependent plasma and 24-h urinary concentrations of 6β-hydroxydexamethasone between untreated humanized-liver mice and humanized-liver mice treated with Azamulin (daily oral doses of 15 mg/kg for 3 days). The mean ratios of 6β-hydroxydexamethasone to dexamethasone for the maximum concentrations, the areas under the plasma concentration-versus-time curves, and urinary concentrations were significantly lower in the azamulin-treated group (59%, 58%, and 41% of the untreated values, respectively). 6β-Hydroxydexamethasone formation was suppressed by 93% by replacing control human liver microsomes with P450 3A4/5-inactivated liver microsomes. These results suggest that the oxidation of dexamethasone in humans is mediated mainly by P450 3A4/5 (which is suppressed by Azamulin), and that humanized-liver mice orally treated with Azamulin may constitute an in vivo model for metabolically inactivated P450 3A4/5 in human hepatocytes transplanted into chimeric mice.