USP25/28 inhibitor AZ1
(Synonyms: AZ1) 目录号 : GC38043
USP25/28 inhibitor AZ1 (AZ1) 是一种具有口服活性、选择性、非竞争性的双泛素特异性蛋白酶 (USP) 25/28 抑制剂，IC50 分别为 0.7 μM 和 0.6 μM。 USP25/28 抑制剂 AZ1 可减轻小鼠模型中的结肠炎和肿瘤发生。
Cas No.:2165322-94-9
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 |
Squamous cell carcinomas (SCC):A431 cells |
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Preparation Method |
AZ1 was added to the medium of A431 cells for 48h |
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Reaction Conditions |
15 µM, 48 h |
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Applications |
USP25/28 inhibitor AZ1-mediated inhibition of USP28 led to changes as measured with predominant effects on pathways associated with cellular stress, cell cycle progression and DNA damage checkpoint and repair. An upregulation of DNA damage sensors like TP53BP1, MRE11 or RAD50 and simultaneously the downregulation of proteins involved in replication-coupled DNA repair, such as RAD51, RPA1 or RPA2. Of note, loss of USP28 activity was associated with a significant decrease of proteins involved in DNA replication. |
| Animal experiment [2]: | |
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Animal models |
Usp25 +/+ and Usp25 -/- mice(age-matched and sex-matched mice) |
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Preparation Method |
At 12 weeks old, mice were injected with PBS or USP25/28 inhibitor AZ1 (40 mg per kg body weight) by gavage every 3 d for 8 weeks. |
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Dosage form |
40 mg/kg, every 3 d for 8 weeks. |
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Applications |
To test the effects of USP25/28 inhibitor AZ1 on DSS-induced colitis and found that USP25/28 inhibitor AZ1 gavage protected from weight loss and diarrhea and impaired colon shortening and potentiated the expression of proinflammatory cytokines and antibacterial peptides in colons. |
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References: [1]. Prieto-Garcia C, Hartmann O, et,al. Inhibition of USP28 overcomes Cisplatin-resistance of squamous tumors by suppression of the Fanconi anemia pathway. Cell Death Differ. 2022 Mar;29(3):568-584. doi: 10.1038/s41418-021-00875-z. Epub 2021 Oct 5. PMID: 34611298; PMCID: PMC8901929. [2]. Wang XM, Yang C, et,al. The deubiquitinase USP25 supports colonic inflammation and bacterial infection and promotes colorectal cancer. Nat Cancer. 2020 Aug;1(8):811-825. doi: 10.1038/s43018-020-0089-4. Epub 2020 Jul 6. PMID: 35122046. |
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USP25/28 inhibitor AZ1 (AZ1) is an orally active, selective, noncompetitive, dual ubiquitin specific protease (USP) 25/28 inhibitor with IC50s of 0.7 μM and 0.6 μM, respectively. High expression levels of USP28 are essential for colon and breast cancers, and stabilization of c-Myc by USP28 is essential for tumor cell proliferation[5]
USP25/28 inhibitor AZ1-mediated inhibition of USP28 led to changes as measured with predominant effects on pathways associated with cellular stress, cell cycle progression and DNA damage checkpoint and repair. An upregulation of DNA damage sensors like TP53BP1, MRE11 or RAD50 and simultaneously the downregulation of proteins involved in replication-coupled DNA repair, such as RAD51, RPA1 or RPA2[3]
USP25/28 inhibitor AZ1 attenuates colitis and tumorigenesis in the mice mode[1].To test the effects of AZ1 on DSS-induced colitis and found that AZ1 gavage protected from weight loss and diarrhea and impaired colon shortening and potentiated the expression of proinflammatory cytokines and antibacterial peptides. In addition, levels of p-p65, p-p38 and SOCS3 were potentiated and levels of TRAF3 and pSTAT3 were decreased in colon tissues from AZ1-treated mice. In addition, gavage of AZ1 maintained weight gain and reduced the bacteria count in feces after C. rodentium infection. inflammation and bacterial replication in the colon were impaired, expression of proinflammatory cytokines and antibacterial peptides was potentiated, levels of p-p65 and p-p38 were increased and levels of TRAF3 were decreased in colons of mice with AZ1 gavage[2].Subchronic injection of AZ1 in 5 FAD mice markedly ameliorated memory deficits in cued FC and MWM tests.In addition, LTP impairment was reversed through long-term AZ1 administration.AZ1 administration attenuated microglial proliferation and activation in the hippocampusand cortex of 5×FAD mice.AZ1 ameliorated AD neuropathology by attenuating microglial activation[4]
References:
[1]: Wrigley JD, Gavory G, et,al. Identification and Characterization of Dual Inhibitors of the USP25/28 Deubiquitinating Enzyme Subfamily. ACS Chem Biol. 2017 Dec 15;12(12):3113-3125. doi: 10.1021/acschembio.7b00334. Epub 2017 Nov 28. PMID: 29131570.
[2]: Wang XM, Yang C, et,al. The deubiquitinase USP25 supports colonic inflammation and bacterial infection and promotes colorectal cancer. Nat Cancer. 2020 Aug;1(8):811-825. doi: 10.1038/s43018-020-0089-4. Epub 2020 Jul 6. PMID: 35122046.
[3]: Prieto-Garcia C, Hartmann O, et,al. Inhibition of USP28 overcomes Cisplatin-resistance of squamous tumors by suppression of the Fanconi anemia pathway. Cell Death Differ. 2022 Mar;29(3):568-584. doi: 10.1038/s41418-021-00875-z. Epub 2021 Oct 5. PMID: 34611298; PMCID: PMC8901929.
[4]: Zheng Q, Li G, et,al. Trisomy 21-induced dysregulation of microglial homeostasis in Alzheimer's brains is mediated by USP25. Sci Adv. 2021 Jan 1;7(1):eabe1340. doi: 10.1126/sciadv.abe1340. PMID: 33523861; PMCID: PMC7775784.
[5]: Popov N, Wanzel M, et,al. The ubiquitin-specific protease USP28 is required for MYC stability. Nat Cell Biol. 2007 Jul;9(7):765-74. doi: 10.1038/ncb1601. Epub 2007 Jun 10. PMID: 17558397.
USP25/28 抑制剂 AZ1 (AZ1) 是一种具有口服活性、选择性、非竞争性的双重泛素特异性蛋白酶 (USP) 25/28 抑制剂，IC50 分别为 0.7 μM 和 0.6 μM . USP28 的高表达水平对于结肠癌和乳腺癌至关重要，USP28 稳定 c-Myc 对于肿瘤细胞增殖至关重要[5]
USP25/28 抑制剂 AZ1 介导的 USP28 抑制导致变化，测量结果显示对与细胞应激、细胞周期进程和 DNA 损伤检查点和修复相关的通路有显着影响。上调 TP53BP1、MRE11 或 RAD50 等 DNA 损伤传感器，同时下调参与复制偶联 DNA 修复的蛋白质，例如 RAD51、RPA1 或 RPA2[3]
USP25/28 抑制剂 AZ1 在小鼠模型中减轻结肠炎和肿瘤发生[1]。为了测试 AZ1 对 DSS 诱导的结肠炎的影响，发现 AZ1 管饲可防止体重减轻和腹泻，并且结肠缩短受损并增强促炎细胞因子和抗菌肽的表达。此外，在 AZ1 处理小鼠的结肠组织中，p-p65、p-p38 和 SOCS3 的水平增强，TRAF3 和 pSTAT3 的水平降低。此外，在啮齿类梭菌感染后，灌胃 AZ1 可维持体重增加并减少粪便中的细菌数量。结肠中的炎症和细菌复制受损，促炎细胞因子和抗菌肽的表达增强，p-p65 和 p-p38 水平升高，TRAF3 水平降低，AZ1 灌胃小鼠[2] 。在 5 只 FAD 小鼠中亚慢性注射 AZ1 显着改善了提示 FC 和 MWM 测试中的记忆缺陷。此外，通过长期给予 AZ1 逆转了 LTP 损伤。AZ1 给药减弱了小鼠海马和皮质中的小胶质细胞增殖和激活5×FAD 小鼠。AZ1 通过减弱小胶质细胞激活改善 AD 神经病理学[4]
| Cas No. | 2165322-94-9 | SDF | |
| 别名 | AZ1 | ||
| Canonical SMILES | FC(C1=CC(COC2=CC=C(Br)C=C2CNCCO)=CC=C1F)(F)F | ||
| 分子式 | C17H16BrF4NO2 | 分子量 | 422.21 |
| 溶解度 | DMSO: ≥ 250 mg/mL (592.12 mM); Water: < 0.1 mg/mL (insoluble) | 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 | 2.3685 mL | 11.8424 mL | 23.6849 mL |
| 5 mM | 0.4737 mL | 2.3685 mL | 4.737 mL |
| 10 mM | 0.2368 mL | 1.1842 mL | 2.3685 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。
Identification and Characterization of Dual Inhibitors of the USP25/28 Deubiquitinating Enzyme Subfamily
ACS Chem Biol.2017 Dec 15;12(12):3113-3125.PMID: 29131570 DOI: 10.1021/acschembio.7b00334
The ubiquitin proteasome system is widely postulated to be a new and important field of drug discovery for the future, with the ubiquitin specific proteases (USPs) representing one of the more attractive target classes within the area. Many USPs have been linked to critical axes for therapeutic intervention, and the finding that USP28 is required for c-Myc stability suggests that USP28 inhibition may represent a novel approach to targeting this so far undruggable oncogene. Here, we describe the discovery of the first reported inhibitors of USP28, which we demonstrate are able to bind to and inhibit USP28, and while displaying a dual activity against the closest homologue USP25, these inhibitors show a high degree of selectivity over other deubiquitinases (DUBs). The utility of these compounds as valuable probes to investigate and further explore cellular DUB biology is highlighted by the demonstration of target engagement against both USP25 and USP28 in cells. Furthermore, we demonstrate that these inhibitors are able to elicit modulation of both the total levels and the half-life of the c-Myc oncoprotein in cells and also induce apoptosis and loss of cell viability in a range of cancer cell lines. We however observed a narrow therapeutic index compared to a panel of tissue-matched normal cell lines. Thus, it is hoped that these probes and data presented herein will further advance our understanding of the biology and tractability of DUBs as potential future therapeutic targets.
Differential Oligomerization of the Deubiquitinases USP25 and USP28 Regulates Their Activities
Mol Cell.2019 May 2;74(3):421-435.e10.PMID: 30926243DOI: 10.1016/j.molcel.2019.02.029
Deubiquitinases have emerged as promising drug targets for cancer therapy. The two DUBs USP25 and USP28 share high similarity but vary in their cellular functions. USP28 is known for its tumor-promoting role, whereas USP25 is a regulator of the innate immune system and, recently, a role in tumorigenesis was proposed. We solved the structures of the catalytic domains of both proteins and established substantial differences in their activities. While USP28 is a constitutively active dimer, USP25 presents an auto-inhibited tetramer. Our data indicate that the activation of USP25 is not achieved through substrate or ubiquitin binding. USP25 cancer-associated mutations lead to activation in vitro and in vivo, thereby providing a functional link between auto-inhibition and the cancer-promoting role of the enzyme. Our work led to the identification of significant differences between USP25 and USP28 and provided the molecular basis for the development of new and highly specific anti-cancer drugs.
Computational approach to target USP28 for regulating Myc
Comput Biol Chem.2020 Apr;85:107208. PMID: 32028107DOI: 10.1016/j.compbiolchem.2020.107208
Myc is a crucial player in cellular proliferation and a known regulator of cancer pathobiology. Modulation of Myc expression targeting the Myc Protein-Protein Interactors (PPIs) like Myc-Max has till now been the most explored approach. However, this approach threatens the normal cells where Myc expression is required for proliferation. This demands the need for a new strategy to indirectly modulate Myc expression. Indirect modulation can be achieved by regulating Myc turnover. FBXW7 mediates the ubiquitination and subsequent degradation of Myc which is reversed by USP28. In this study, the interaction of USP28 with FBXW7 as well as with its substrate, Ubiquitin (Ub) were used as targets. Computation based high-throughput screening of bioactive small chemicals using molecular docking method was implemented to predict USP28 inhibitors. For the two regions, docking study with AutoDock Vina gave top 10 best scoring drugs which were identified and tabulated. The two regions defined in the study as FBXW7 binding and Ub binding also encompass the areas in which USP28 differed from USP25, a homologue with a different role. Out of these the best scoring drugs were explored for their role in cancer, if any. This study was performed keeping in mind re-purposing of these known drugs for possible alternative anti-Myc cancer therapy.
USP28 and USP25 are downregulated by Vismodegib in vitro and in colorectal cancer cell lines
FEBS J.2021 Feb;288(4):1325-1342.PMID: 32578360DOI: 10.1111/febs.15461
Deubiquitinase USP28 plays a crucial role in tumorigenesis by enhancing the stabilities of multiple cancer-related proteins including c-Myc, Notch1, and LSD1, and has become an attractive target for anticancer drug development. However, to date, only a few of USP28-targeted active compounds have been developed, and the active compound-binding pocket in USP28 has not been experimentally revealed yet. In this study, bioassay-based high-throughput screening was applied to discover USP28-targeted inhibitors from the commercially available drug library. Vismodegib, an inhibitor of Hedgehog signaling pathway and FDA-approved drug for the treatment of basal cell carcinoma, was found to exhibit inhibition activity against USP28 (IC50 : 4.41 ± 1.08 μm). Multiple biophysical and biochemical techniques including NMR, ITC, thermal shift assay, HDX-MS, and site-directed mutagenesis analysis were then used to characterize the interaction between Vismodegib and USP28. The binding pocket in USP28 for Vismodegib, which is mainly composed of two helical structures spanning D255-N278 and N286-Y293, was revealed. According to the possible binding pose generated by HDX-MS data-defined molecular docking, the binding cavity occupied by Vismodegib in USP28 aligns well with one of the reported-binding pockets in USP7 for its inhibitors. Furthermore, cellular assays were conducted to confirm that Vismodegib could interact with the evolutionarily related deubiquitinases USP28 and USP25 and downregulate the levels of the two enzymes' substrate proteins c-Myc, Notch1, and Tankyrase-1/2.
USP28: Oncogene or Tumor Suppressor? A Unifying Paradigm for Squamous Cell Carcinoma
Cells.2021 Oct 4;10(10):2652.PMID: 34685632 DOI: 10.3390/cells10102652
Squamous cell carcinomas are therapeutically challenging tumor entities. Low response rates to radiotherapy and chemotherapy are commonly observed in squamous patients and, accordingly, the mortality rate is relatively high compared to other tumor entities. Recently, targeting USP28 has been emerged as a potential alternative to improve the therapeutic response and clinical outcomes of squamous patients. USP28 is a catalytically active deubiquitinase that governs a plethora of biological processes, including cellular proliferation, DNA damage repair, apoptosis and oncogenesis. In squamous cell carcinoma, USP28 is strongly expressed and stabilizes the essential squamous transcription factor ΔNp63, together with important oncogenic factors, such as NOTCH1, c-MYC and c-JUN. It is presumed that USP28 is an oncoprotein; however, recent data suggest that the deubiquitinase also has an antineoplastic effect regulating important tumor suppressor proteins, such as p53 and CHK2. In this review, we discuss: (1) The emerging role of USP28 in cancer. (2) The complexity and mutational landscape of squamous tumors. (3) The genetic alterations and cellular pathways that determine the function of USP28 in squamous cancer. (4) The development and current state of novel USP28 inhibitors.