Epoxomicin
(Synonyms: 环氧酶素; BU-4061T) 目录号 : GC12494
环氧霉素（Epoxomicin）是一种选择性蛋白酶体抑制剂，有效地抑制20S蛋白酶体的胰凝乳蛋白酶样（CH-L）活性，IC50约为40-80nM。
Cas No.:134381-21-8
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >98.50%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
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Cell experiment [1]: |
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Cell lines |
HEK293T cells |
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Preparation method |
HEK293T cells were grown to 90% confluence. At the start of the experiment, media were removed from all plates and media with or without epoxomicin were placed into the plates incubated at 37°C for 1h. Two concentrations of epoxomicin were used, 0.2 and 2μM, prepared by a 1:2500 dilution of 0.5 and 5 mM solutions of epoxomicin in DMSO, respectively. |
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Reaction Conditions |
0.2 and 2μM; 1 h |
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Applications |
Peptides are degraded from cytoplasmic, mitochondrial and nuclear proteins by proteasomes. Epoxomicin inhibits proteasome beta-2 and beta-5 subunits in HEK293T cells. |
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Animal experiment [2]: |
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Animal models |
C57BL/6 mice |
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Preparation method |
mice were weighed and injected intraperitoneally with vehicle or epoxomicin , 0.5mg/kg. One hour after the first injection, animals were injected with vehicle or LPS, 7.5 mg/kg. Mice were anesthetized eight hours after LPS administration with a lethal injection of a mix of ketamine and xylazine. Measuring all three β1, β2 and β5 activites of the 20S and 26S proteasomes, the levels of steady state polyubiquitinated proteins. |
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Dosage form |
0.5mg/kg; i.p. |
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Applications |
Epoxomicin decreased polyubiquitination by 23% in GAS muscle and conversely increased it by 41% in liver, the latter being consistent with proteasome β5 inhibition. |
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References: [1] Fricker L D, Gelman J S, Castro L M, et al. Peptidomic analysis of HEK293T cells: effect of the proteasome inhibitor epoxomicin on intracellular peptides[J]. Journal of proteome research, 2012, 11(3): 1981-1990. [2] Jamart, Cécile, Gomes A V , Dewey S ,et al. Regulation of ubiquitin-proteasome and autophagy pathways after acute LPS and epoxomicin administration in mice[J].Bmc Musculoskeletal Disorders, 2014, 15(1):166. |
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Epoxomicin is a selective proteasome inhibitor that effectively inhibits the chymotrypsin-like (CH-L) activity of the 20S proteasome, with an IC50 of approximately 40-80nM[1]. Epoxomicin covalently binds to the catalytic β subunits LMP 7, X, Z, and MECL 1 of the proteasome, leading to inhibition of the proteasome subunits' CH-L, T-L, and PGPH catalytic activities[2].
In vitro, Epoxomicin shows considerable cytotoxicity against B16-F10, HCT116, Moser, P388, and K562 cells, with IC50 values of 0.002μg/mL, 0.005μg/mL, 0.044μg/mL, 0.002μg/mL, and 0.037μg/mL, respectively[3]. Epoxymycin (0.2 and 2 μM) treated HEK293T cells for 1 hour, inhibiting the expression of proteasome beta-2 and beta-5 subunits in the cells[4]. Epoxomicin (100 nM) for 24 hours in Plasmodium falciparum cells results in a 77% reduction in gametocytes[5].
In vivo, Epoxymycin (0.5mg/kg), administered intraperitoneally, reduced polyubiquitination in GAS muscle of C57BL/6 mice by 23% and increased polyubiquitination in liver by 41%, the latter associated with The inhibitory effect of proteasome β5 is consistent [6]. Epoxymycin (0.5mg/kg/day) was administered to KI and WT mice for 1 week using a subcutaneously implanted osmotic minipump, which reduced the chymotrypsin-like activity of the mice by approximately 50%[7] .
References:
[1] Meng L , Mohan R , Kwok B H B ,et al.Epoxomicin, a potent and selective proteasome inhibitor, exhibits in vivo antiinflammatory activity[J]. Proceedings of the National Academy of Sciences, 1999, 96(18):10403-10408.
[2] Aboulaila M , Nakamura K , Govind Y ,et al.Evaluation of the in vitro growth-inhibitory effect of epoxomicin on Babesia parasites.[J].Veterinary Parasitology, 2010, 167(1):19-27.
[3] Hanada M, Sugawara K, Kaneta K, et al. Epoxomicin, a new antitumor agent of microbial origin.[J] Antibiot (Tokyo). 1992 Nov;45(11):1746-52.
[4] Fricker L D, Gelman J S, Castro L M, et al. Peptidomic analysis of HEK293T cells: effect of the proteasome inhibitor epoxomicin on intracellular peptides[J]. Journal of proteome research, 2012, 11(3): 1981-1990.
[5] Czesny B , Goshu S , Cook J L ,et al. The Proteasome Inhibitor Epoxomicin Has Potent Gametocytocidal Activity[J]. 2009.
[6]Jamart, Cécile, Gomes A V , Dewey S ,et al. Regulation of ubiquitin-proteasome and autophagy pathways after acute LPS and epoxomicin administration in mice[J].Bmc Musculoskeletal Disorders, 2014, 15(1):166.
[7]Schlossarek,Saskia,Singh,et al. Proteasome inhibition slightly improves cardiac function in mice with hypertrophic cardiomyopathy[J].Frontiers in Physiology, 2014.
环氧霉素（Epoxomicin）是一种选择性蛋白酶体抑制剂，有效地抑制20S蛋白酶体的胰凝乳蛋白酶样（CH-L）活性，IC50约为40-80nM[1]。环氧霉素共价结合蛋白酶体的LMP 7、X、Z和MECL 1催化β亚基，导致蛋白酶体亚基的CH-L、T-L和PGPH催化活性受到抑制[2]。
在体外，环氧霉素对B16-F10、HCT116、Moser、P388和K562细胞均显示出相当强的细胞毒性，IC50 值分为 0.002 μg/mL、0.005 μg/mL、0.044 μg/mL、0.002 μg/mL和0.037μg/mL[3]。环氧霉素(0.2和2μM)处理HEK293T细胞1小时，抑制了细胞中的蛋白酶体beta-2和beta-5亚基的表达[4]。环氧霉素(100 nM)处理恶性疟原虫细胞24小时，配子细胞减少了77%[5]。
在体内，环氧霉素（0.5mg/kg）通过腹腔注射，使C57BL/6小鼠GAS肌肉中的多泛素化降低 23%，使肝脏中的多泛素化增加 41%，后者与蛋白酶体β5抑制作用一致[6]。环氧霉素（0.5mg/kg/天）采用皮下植入的渗透微型泵持续1周对KI和WT小鼠 给药，使小鼠的胰凝乳蛋白酶样活性降低了约50%[7]。
| Cas No. | 134381-21-8 | SDF | |
| 别名 | 环氧酶素; BU-4061T | ||
| 化学名 | (2S,3S)-2-[[(2S,3S)-2-[acetyl(methyl)amino]-3-methylpentanoyl]amino]-N-[(2S,3R)-3-hydroxy-1-[[(2S)-4-methyl-1-[(2R)-2-methyloxiran-2-yl]-1-oxopentan-2-yl]amino]-1-oxobutan-2-yl]-3-methylpentanamide | ||
| Canonical SMILES | CCC(C)C(C(=O)NC(C(C)O)C(=O)NC(CC(C)C)C(=O)C1(CO1)C)NC(=O)C(C(C)CC)N(C)C(=O)C | ||
| 分子式 | C28H50N4O7 | 分子量 | 554.7 |
| 溶解度 | ≥ 27.74mg/mL in DMSO | 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 | 1.8028 mL | 9.0139 mL | 18.0278 mL |
| 5 mM | 0.3606 mL | 1.8028 mL | 3.6056 mL |
| 10 mM | 0.1803 mL | 0.9014 mL | 1.8028 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。
From epoxomicin to carfilzomib: chemistry, biology, and medical outcomes
The initial enthusiasm following the discovery of a pharmacologically active natural product is often fleeting due to the poor prospects for its ultimate clinical application. Despite this, the ever-changing landscape of modern biology has a constant need for molecular probes that can aid in our understanding of biological processes. After its initial discovery by Bristol-Myers Squibb as a microbial anti-tumor natural product, epoxomicin was deemed unfit for development due to its peptide structure and potentially labile epoxyketone pharmacophore. Despite its drawbacks, epoxomicin's pharmacophore was found to provide unprecedented selectivity for the proteasome. Epoxomicin also served as a scaffold for the generation of a synthetic tetrapeptide epoxyketone with improved activity, YU-101, which became the parent lead compound of carfilzomib (Kyprolis?), the recently approved therapeutic agent for multiple myeloma. In this era of rational drug design and high-throughput screening, the prospects for turning an active natural product into an approved therapy are often slim. However, by understanding the journey that began with the discovery of epoxomicin and ended with the successful use of carfilzomib in the clinic, we may find new insights into the keys for success in natural product-based drug discovery.
Site-Specific Proteasome Inhibitors
Proteasome is a multi-subunit protein degradation machine, which plays a key role in the maintenance of protein homeostasis and, through degradation of regulatory proteins, in the regulation of numerous cell functions. Proteasome inhibitors are essential tools for biomedical research. Three proteasome inhibitors, bortezomib, carfilzomib, and ixazomib are approved by the FDA for the treatment of multiple myeloma; another inhibitor, marizomib, is undergoing clinical trials. The proteolytic core of the proteasome has three pairs of active sites, β5, β2, and β1. All clinical inhibitors and inhibitors that are widely used as research tools (e.g., epoxomicin, MG-132) inhibit multiple active sites and have been extensively reviewed in the past. In the past decade, highly specific inhibitors of individual active sites and the distinct active sites of the lymphoid tissue-specific immunoproteasome have been developed. Here, we provide a comprehensive review of these site-specific inhibitors of mammalian proteasomes and describe their utilization in the studies of the biology of the active sites and their roles as drug targets for the treatment of different diseases.
Epoxomicin, a Selective Proteasome Inhibitor, Activates AIM2 Inflammasome in Human Retinal Pigment Epithelium Cells
Emerging evidence suggests that the intracellular clearance system plays a vital role in maintaining homeostasis and in regulating oxidative stress and inflammation in retinal pigment epithelium (RPE) cells. Dysfunctional proteasomes and autophagy in RPE cells have been associated with the pathogenesis of age-related macular degeneration. We have previously shown that the inhibition of proteasomes using MG-132 activates the NLR family pyrin domain containing 3 (NLRP3) inflammasome in human RPE cells. However, MG-132 is a non-selective proteasome inhibitor. In this study, we used the selective proteasome inhibitor epoxomicin to study the effect of non-functional intracellular clearance systems on inflammasome activation. Our data show that epoxomicin-induced proteasome inhibition promoted both nicotinamide adenine dinucleotide phosphate oxidase and mitochondria-mediated oxidative stress and release of mitochondrial DNA to the cytosol, which resulted in potassium efflux-dependent absence in melanoma 2 (AIM2) inflammasome activation and subsequent interleukin-1β secretion in ARPE-19 cells. The non-specific proteasome inhibitor MG-132 activated both NLRP3 and AIM2 inflammasomes and oxidative stress predominated as the activation mechanism, but modest potassium efflux was also detected. Collectively, our data suggest that a selective proteasome inhibitor is a potent inflammasome activator in human RPE cells and emphasize the role of the AIM2 inflammasome in addition to the more commonly known NLRP3 inflammasome.
Suppression of the TRIF-dependent signaling pathway of TLRs by epoxomicin
Toll-like receptors (TLRs) can recognize specific signatures of invading microbial pathogens and activate a cascade of downstream signals to induce the secretion of inflammatory cytokines, chemokines, and type I interferons. The activation of TLRs triggers two downstream signaling pathways: the MyD88- and the TRIF-dependent pathways. To evaluate the therapeutic potential of epoxomicin, a member of the linear peptide α',β'-epoxyketone first isolated from an actinomycetes strain, we examined its effects on signal transduction via TLR signaling pathways. Epoxomicin inhibited the activation of NF-kB and IRF3 induced by TLR agonists, decreased the expression of interferon-inducible protein-10, and inhibited the activation of NF-kB and IRF3 induced by overexpression of downstream signaling components of TLR signaling pathways. These results suggest that epoxomicin can regulate both the MyD88- and TRIF-dependent signaling pathways of TLRs. Thus, it might have potential as a new therapeutic drug for a variety of inflammatory diseases.
Synthesis and Application of a Clickable Epoxomicin-Based Probe for Proteasome Activity Analysis
The proteasome is a multisubunit protein complex responsible for the degradation of proteins, making it essential in myriad cellular processes. Several reversible and irreversible peptide substrates inspired by known proteasome inhibitors have been developed to visualize it and monitor its activity; however, they have limited commercial availability or possess fluorophores that overlap with other known chemical probes, limiting their simultaneous use. The protocols presented here describe the synthesis of a clickable epoxomicin-based probe followed by the copper-catalyzed installment of an azide-containing fluorophore, and the application of the synthesized peptide in proteasome activity assays by SDS-PAGE and flow cytometry. ? 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Solid-phase synthesis of clickable peptide fragment (2) Basic Protocol 2: In-solution coupling of epoxy-ketone moiety to fragment (2) Basic Protocol 3: Copper-catalyzed click reaction of (3) with fluorophore of choice Basic Protocol 4: Monitoring proteasome activity by SDS-PAGE in HEK-293T cells Alternate Protocol: Monitoring proteasome activity by flow cytometry in HEK-293T cells.