| 名稱 | COH000 |
| 描述 | COH000 is a covalent and irreversible inhibitor of small ubiquitin-like modifier (SUMO)-activating enzyme and inhibited SUMOylation (IC50: ~ 0.2 μM in vitro). |
| 細(xì)胞實(shí)驗(yàn) | Cell proliferation was measured using a CellTiter 96 AQueous One Solution Cell Proliferation Assay (MTS-based) after COH000 or its analog (54 or 55) treatment at the indicated concentrations and the time points. Briefly, cells were incubated with 20 mL of CellTiter 96 AQueous reagentafter the treatment and incubated at 37C until color development. Absorbance measurements were performed using a SpectraMax M5 reader. For assays measuring anti-proliferation effects, all values were normalized to the vehicle treatment. All values are represented graphically as mean ± standard deviation (STDEV) from three independent samples (n=3). |
| 激酶實(shí)驗(yàn) | Briefly, SAE, Ubc9, GST-SUMO and His6-RanGap-1 proteins were expressed and purified as described previously.Assay buffer contained 50 mMTris-HCl pH 7.4, 0.3 mM DTT, 10 mM MgCl2, and 0.005% Tween-20. The assays were conducted using 1536-well, white plates. 2 μL of Mixture 1, containing 12.5 nM SAE and 100 nM His6-RanGap-1 in the assay buffer was mixed with 70 nl of 2 mM compounds dissolved in DMSO. Then 2 μL of Mixture 2, containing 20 mM ATP, 12.5 nM E2 and 30 nM GST-SUMO in assay buffer, was added. After incubation for 90 min at room temperature, 1 μl of pre-mixed Ni acceptor beads and Glutathione Donor beads, at 10 μg/ml each, was added. After incubation for 60 min at room temperature, readings were made on a BMG LabtechPheraStar in an AlphaScreen mode (Ex: 680 nm; Em: 570 nm). |
| 動(dòng)物實(shí)驗(yàn) | Mice were housed in a controlled environment (12-h light/12-h dark cycle) with access to waterand fed a standard diet generally from 6 to 8 weeks of age. Body weight was measured weekly and food intake was monitored. For colorectal cancer xenograft with COH000 treatment, HCT116 cells were s.c. injected into a plasma esterase-deficient SCID mouse strain (Es1e/SCID), male, 8-10 weeks. When the tumors became palpable, COH000 was administered subcutaneously peritumoral injection once a day at a dose of 10 mg per kilogram of body weight. Mice in the control group received equal volumes of vehicle (5% DMSO, 30% solutol in PBS). Tumor volume was measured until the endpoint was reached. Mice were euthanized using CO2 inhalation and tumors were excised. |
| 體外活性 | COH000在體外平均IC50約為0.2 μM的條件下抑制了SUMOylation，但在相同條件下，即使?jié)舛雀哌_(dá)100 μM，在以Ubc13介導(dǎo)的多聚泛素化試驗(yàn)中也未能抑制泛素化。COH000處理誘導(dǎo)了HCT-116細(xì)胞中的凋亡。通過轉(zhuǎn)染過表達(dá)SAE2，凋亡水平降低；而當(dāng)在HCT116細(xì)胞中敲除SAE2時(shí)，凋亡水平增加。 |
| 體內(nèi)活性 | 當(dāng)腫瘤變得可觸及時(shí)，攜帶腫瘤的Es1e/SCID小鼠接受了COH000或?qū)φ談┑闹委?，為?4天。COH000顯著抑制了腫瘤生長(zhǎng)，并且顯著降低了腫瘤組織中SAE2的水平。 |
| 存儲(chǔ)條件 | Powder: -20°C for 3 years | In solvent: -80°C for 1 year | Shipping with blue ice. |
| 溶解度 | H2O : < 0.1 mg/mL (insoluble)
DMSO : 35.71 mg/mL (85.13 mM), Sonication is recommended.
|
| 關(guān)鍵字 | Ubiquitin conjugating enzyme | E1 activating enzyme | COH000 | E1/E2/E3 Enzyme | E2 conjugating enzyme | Ubiquitin ligase | Ubiquitin activating enzyme | COH-000 | COH 000 | Inhibitor | E3 ligating enzyme | inhibit |
| 相關(guān)產(chǎn)品 | Nutlin-3 | NAE-IN-M22 | BC-1382 | RG7112 | NSC697923 | Indole-3-carbinol | PYR-41 | Idasanutlin | Navtemadlin | NAcM-OPT | PYZD-4409 | NSC232003 |
| 相關(guān)庫 | 抑制劑庫 | 經(jīng)典已知活性庫 | 已知活性化合物庫 | HIF-1化合物庫 | NO PAINS 化合物庫 | 泛素化化合物庫 | 表型篩選靶點(diǎn)鑒定庫 | 共價(jià)抑制劑庫 |