MLN4924
(Synonyms: 氨基磺酸[(1S,2S,4R)-4-[4-[[(1S)-2,3-二氢-1H-茚-1-基]氨基]-7H-吡咯并[2,3-D]嘧啶-7-基]-2-羟基环戊基]甲基酯,MLN4924) 目录号 : GC15086
MLN4924 是一种有效的选择性 NEDD8 激活酶 (NAE) 小分子抑制剂 (IC50 = 4 nM) ，并且相对于密切相关的酶 UAE、SAE、UBA6 和 ATG7 具有选择性（ IC50 = 1.5、8.2、1.8 和 >；分别为 10 μM）。
Cas No.:905579-51-3
Sample solution is provided at 25 µL, 10mM.
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- Purity: >98.00%
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- Datasheet
| Cell experiment [1]: | |
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Cell lines |
HCT-116 cells |
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Preparation Method |
HCT-116 cells grown in 6-well cell-culture dishes were treated with 0.1% DMSO (control) or MLN4924 for 24 h. Whole cell extracts were prepared and analysed by immunoblotting. For analysis of the E2-UBL thioester levels, lysates were fractionated by non-reducing SDS-PAGE and immunoblotted with polyclonal antibodies to Ubc12, Ubc9 and Ubc10. |
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Reaction Conditions |
0-3µM for 24 hours |
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Applications |
Treatment of HCT-116 cells with MLN4924 for 24 h resulted in a dose-dependent decrease of Ubc12-NEDD8 thioester and NEDD8-cullin conjugates, with an IC50 < 0.1 µM |
| Animal experiment [1]: | |
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Animal models |
Female athymic NCR mice |
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Preparation Method |
All animals were housed and handled in accordance with the Guide for the Care and Use of Laboratory Animals. Mice were inoculated with 2 × 106 HCT-116 cells (or 30-40 mg H522 tumour fragments) subcutaneously in the right flank, and tumour growth was monitored with caliper measurements. When the mean tumour volume reached approximately 200 mm3, animals were dosed subcutaneously with vehicle (10% cyclodextrin) or MLN4924. Inhibition of tumour growth (T/C) was calculated on the last day of treatment. |
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Dosage form |
subcutaneous injection, once (QD) or twice (BID) daily,30 and 60 mg kg-1 |
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Applications |
MLN4924 administered on a BID schedule at 30 and 60 mg kg-1 inhibited tumour growth with T/C values of 0.36 and 0.15, respectively |
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References: [1]: Soucy T A, Smith P G, Milhollen M A, et al. An inhibitor of NEDD8-activating enzyme as a new approach to treat cancer[J]. Nature, 2009, 458(7239): 732-736. |
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MLN4924 is a potent and selective small-molecule inhibitor of NEDD8-activating enzyme (NAE) (IC50 = 4 nM) [1,2], and is selective relative to the closely related enzymes UAE, SAE, UBA6 and ATG7 (IC50 = 1.5, 8.2, 1.8 and >10 μM, respectively) [2]. MLN4924 has a multifaceted mechanism of action that is characterized by the induction of DNA re-replication and DNA damage, increased oxidative stress, inhibition of NF-B activity, apoptotic cell death, and cellular senescence [3].
MLN4924 Treatment of HCT-116 cells for 24 h resulted in a dose-dependent decrease of Ubc12-NEDD8 thioester and NEDD8-cullin conjugates, with an IC50 < 0.1 μM [2], resulting in a reciprocal increase in the abundance of the known CRL substrates CDT1, p27 and NRF2 (also known as NFE2L2), c-Jun27, HIF1α, cyclin E29, CDC25A, EMI1 (also known as FBXO5) and phosphorylated IκBα, but not non-CRL substrates [2]. MLN4924 treatment of human-tumour-derived cell lines, including HCT-116(colon), Calu-6 (lung), SKOV-3 (ovarian), H460 (lung), DLD-1 (colon), CWR22 (prostate) and OCI-LY19 (lymphoma), resulted in S-phase-defective phenotypes [2]. Potent inhibition of MLN4924 of cell viability was observed across 17 lymphoma cell lines tested in an ATPlite viability assay, with EC50 values of 10 to 244nM [1].
In OCI-Ly10 xenografts, a dose- and time-dependent increase in pIκBα levels was observed after MLN4924 treatment (10, 30, or 60 mg/kg, subcutaneous), with peak elevation occurring 2 hours after dose and levels returning to baseline at 8 hours after dose. A consequence of inhibiting NAE and NF-κB signaling in OCI-Ly10 xenografts was the induction of apoptosis 8 to 12 hours after a single dose of 60 mg/kg MLN4924, as evidenced by detection of cleaved caspase-3 [1]. A single dose of MLN4924 resulted in a dose- and time-dependent decrease of NEDD8-cullin levels as early as 30 min after administration of MLN4924 (10, 30 or 60 mg/kg, subcutaneous), with maximal effect 1-2 h post-dose on HCT-116 tumour-bearing mice [2].
References:
[1]. Milhollen M A, Traore T, Adams-Duffy J, et al. MLN4924, a NEDD8-activating enzyme inhibitor, is active in diffuse large B-cell lymphoma models: rationale for treatment of NF-κB-dependent lymphoma[J]. Blood, The Journal of the American Society of Hematology, 2010, 116(9): 1515-1523.
[2]. Soucy T A, Smith P G, Milhollen M A, et al. An inhibitor of NEDD8-activating enzyme as a new approach to treat cancer[J]. Nature, 2009, 458(7239): 732-736.
[3]. Nawrocki S T, Griffin P, Kelly K R, et al. MLN4924: a novel first-in-class inhibitor of NEDD8-activating enzyme for cancer therapy[J]. Expert opinion on investigational drugs, 2012, 21(10): 1563-1573.
MLN4924 是一种有效的选择性 NEDD8 激活酶 (NAE) 小分子抑制剂 (IC50 = 4 nM) [1,2]，并且相对于密切相关的酶 UAE、SAE、UBA6 和 ATG7 具有选择性（ IC50 = 1.5、8.2、1.8 和 >；分别为 10 μM）[2]。 MLN4924 具有多方面的作用机制，其特征在于诱导 DNA 再复制和 DNA 损伤、增加氧化应激、抑制 NF-B 活性、凋亡细胞死亡和细胞衰老[3] .
MLN4924 处理 HCT-116 细胞 24 小时导致 Ubc12-NEDD8 硫酯和 NEDD8-cullin 结合物呈剂量依赖性降低，IC50 为 <； 0.1 μM [2]，导致已知 CRL 底物 CDT1、p27 和 NRF2（也称为 NFE2L2）、c-Jun27、HIF1α、细胞周期蛋白 E29、CDC25A、EMI1 的丰度相互增加（也称为 FBXO5）和磷酸化 IκBα，但不是非 CRL 底物[2]。 MLN4924 治疗人肿瘤来源的细胞系，包括 HCT-116（结肠）、Calu-6（肺）、SKOV-3（卵巢）、H460（肺）、DLD-1（结肠）、CWR22（前列腺）和OCI-LY19（淋巴瘤）导致 S 期缺陷表型 [2]。在 ATPlite 活力测定中测试的 17 种淋巴瘤细胞系中观察到 MLN4924 对细胞活力的有效抑制，EC50 值为 10 至 244nM [1]。
在 OCI-Ly10 异种移植物中，在 MLN4924 治疗（10、30 或 60 毫克/千克，皮下注射）后观察到 pIκBα 水平呈剂量和时间依赖性升高，给药后 2 小时出现峰值升高，水平恢复给药后 8 小时达到基线。在 OCI-Ly10 异种移植物中抑制 NAE 和 NF-κB 信号传导的结果是在单剂量 60 mg/kg MLN4924 后 8 至 12 小时诱导细胞凋亡，检测到裂解的 caspase-3 [1] 。早在施用 MLN4924（10、30 或 60 mg/kg，皮下）后 30 分钟，单剂量 MLN4924 就会导致 NEDD8-cullin 水平的剂量和时间依赖性降低，在 1-2 小时后产生最大效果-对携带 HCT-116 肿瘤的小鼠的剂量[2]。
| Cas No. | 905579-51-3 | SDF | |
| 别名 | 氨基磺酸[(1S,2S,4R)-4-[4-[[(1S)-2,3-二氢-1H-茚-1-基]氨基]-7H-吡咯并[2,3-D]嘧啶-7-基]-2-羟基环戊基]甲基酯,MLN4924 | ||
| 化学名 | [(1S,2S,4R)-4-[4-[[(1S)-2,3-dihydro-1H-inden-1-yl]amino]pyrrolo[2,3-d]pyrimidin-7-yl]-2-hydroxycyclopentyl]methyl sulfamate | ||
| Canonical SMILES | C1CC2=CC=CC=C2C1NC3=NC=NC4=C3C=CN4C5CC(C(C5)O)COS(=O)(=O)N | ||
| 分子式 | C21H25N5O4S | 分子量 | 443.53 |
| 溶解度 | ≥ 22.2mg/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 | 2.2546 mL | 11.2732 mL | 22.5464 mL |
| 5 mM | 0.4509 mL | 2.2546 mL | 4.5093 mL |
| 10 mM | 0.2255 mL | 1.1273 mL | 2.2546 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。
Neddylation-Independent Activities of MLN4924
Adv Exp Med Biol2020;1217:363-372.PMID: 31898238DOI: 10.1007/978-981-15-1025-0_21
MLN4924, also known as pevonedistat, is a highly selective small-molecule inhibitor of NEDD8 (neuronal precursor cell-expressed developmentally downregulated protein 8)-activating enzyme (NAE) to block the entire neddylation modification cascade, leading to inactivation of cullin-RING ligases (CRLs), since activation of CRLs requires cullin neddylation. MLN4924 showed impressive anticancer activity in many preclinical studies and is currently in several Phase I/II clinical trials for anticancer therapy as a single agent or in combination with chemotherapeutic drugs.In addition to well-characterized anti-neddylation activity, recent studies showed that MLN4924 has several neddylation-independent activities. First, MLN4924 triggers EGFR dimerization to activate EGFR and its downstream RAS/MAPK and PI3K/AKT1 signals, leading to enhanced tumor sphere formation, accelerated EGF-mediated wound healing, and inhibited ciliogenesis. Second, MLN4924 induces PKM2 tetramerization to promote glycolysis, thus affecting energy metabolism. Third, MLN4924 inhibits the interaction between ACT1 (NF-百B activator 1) and TRAF6 (tumor necrosis factor receptor-associated factor 6) and attenuates IL-17A-mediated activation of NF-百B to reduce pulmonary inflammation. Fourth, MLN4924 inhibits IRF3 binding to the IFN-汿promoter to inhibit IFN-汿production. And finally, MLN4924 activates the JNK signaling pathway to reduce c-FLIP levels, thus enhancing TRAIL-induced apoptosis. This chapter will summarize these neddylation-independent activities of MLN4924 and discuss the underlying mechanisms and potential therapeutic applications.
The NEDD8-activating enzyme inhibitor MLN4924 reduces ischemic brain injury in mice
Proc Natl Acad Sci U S A2022 Feb 8;119(6):e2111896119.PMID: 35101976DOI: 10.1073/pnas.2111896119
Blood-brain barrier (BBB) breakdown and inflammation occurring at the BBB have a key, mainly a deleterious role in the pathophysiology of ischemic stroke. Neddylation is a ubiquitylation-like pathway that is critical in various cellular functions by conjugating neuronal precursor cell-expressed developmentally down-regulated protein 8 (NEDD8) to target proteins. However, the roles of neddylation pathway in ischemic stroke remain elusive. Here, we report that NEDD8 conjugation increased during acute phase after ischemic stroke and was present in intravascular and intraparenchymal neutrophils. Inhibition of neddylation by MLN4924, also known as pevonedistat, inactivated cullin-RING E3 ligase (CRL), and reduced brain infarction and improved functional outcomes. MLN4924 treatment induced the accumulation of the CRL substrate neurofibromatosis 1 (NF1). By using virus-mediated NF1 silencing, we show that NF1 knockdown abolished MLN4924-dependent inhibition of neutrophil trafficking. These effects were mediated through activation of endothelial P-selectin and intercellular adhesion molecule-1 (ICAM-1), and blocking antibodies against P-selectin or anti-ICAM-1 antibodies reversed NF1 silencing-induced increase in neutrophil infiltration in MLN4924-treated mice. Furthermore, we found that NF1 silencing blocked MLN4924-afforded BBB protection and neuroprotection through activation of protein kinase C 汿(PKC汿, myristoylated alanine-rich C-kinase substrate (MARCKS), and myosin light chain (MLC) in cerebral microvessels after ischemic stroke, and treatment of mice with the PKC汿inhibitor rottlerin reduced this increased BBB permeability. Our study demonstrated that increased neddylation promoted neutrophil trafficking and thus exacerbated injury of the BBB and stroke outcomes. We suggest that the neddylation inhibition may be beneficial in ischemic stroke.
MLN4924 therapy as a novel approach in cancer treatment modalities
J Chemother2016 Apr;28(2):74-82.PMID: 26292710DOI: 10.1179/1973947815Y.0000000066
MLN4924 is an investigational and a newly discovered small molecule that is a potent and selective inhibitor of the NEDD8 (Neural precursor cell-Expressed Developmentally down-regulated 8) Activating Enzyme (NAE), a pivotal regulator of the Cullin Ring Ligases E3 (CRL), which has been implicated recently in DNA damage. MLN4924 effectively inhibits tumour cell growth by inducing all three common types of death, namely apoptosis, autophagy and senescence and it was also reported that the formation of capillary vessels was significantly suppressed by MLN4924.In this review, we are going to highlight the molecular mechanism of MLN4924 in cancer therapy and its pros and cons in cancer therapy.
Cardioprotective effect of MLN4924 on ameliorating autophagic flux impairment in myocardial ischemia-reperfusion injury by Sirt1
Redox Biol2021 Oct;46:102114.PMID: 34454165DOI: 10.1016/j.redox.2021.102114
Neddylation is essential for cardiomyocyte survival in the presence of oxidative stress, and it participates in autophagy regulation. However, whether MLN4924-an inhibitor of neddylation-exerts cardioprotective effects against myocardial ischemia/reperfusion (MI/R) remains unknown. In the present study, MLN4924 exerted strong cardioprotective effects, demonstrated by significantly elevated cell viability, a decreased LDH leakage rate, and improved cell morphology following H2O2-induced injury in vitro. MLN4924 also markedly decreased the serum myocardial zymogram level, ameliorated cardiac histopathological alterations, and alleviated left ventricular contractile dysfunction, thus limiting the cardiac infarct size in vivo compared with those in MI/R mice. Amazingly, such action of MLN4924 was abrogated by a combined treatment with the autophagic flux inhibitor, chloroquine. The mRFP-GFP-LC3 assay illustrated that MLN4924 restored the defective autophagic flux via enhancing the autolysosome formation. Notably, the expression levels of Rab7 and Atg5 were markedly up-regulated in MLN4924 treated cells and mice subjected to H2O2 or MI/R, respectively, while knockdown of Sirt1 in cells and heart tissue largely blocked such effect and induced autophagosome accumulation by inhibiting its fusion with lysosomes. Transmission electron microscopic analysis, histopathological assay and TUNEL detection of the heart tissues showed that the absence of Sirt1 blocked the cardioprotective effect of MLN4924 by further exacerbating the impaired autophagic flux during MI/R injury in vivo. Taken together, MLN4924 exhibited the strong cardioprotective action via restoring the impaired autophagic flux in H2O2-induced injury in vitro and in MI/R mice. Our work implicated that Sirt1 played a critical role in autophagosome clearance, likely through up-regulating Rab7 in MI/R.