| 名稱 | Torin 1 |
| 描述 | Torin 1 is an mTOR inhibitor that inhibits mTORC1 and mTORC2 (IC50=2/10 nM) selectively and ATP-competitively. Torin 1 induces cellular autophagy and possesses anti-tumor, anti-inflammatory, and anti-aging activities. |
| 細(xì)胞實(shí)驗(yàn) | Cell viability is assessed with the CellTiter-Glo Luminescent Cell Viability Assay. On Day 0, 96-well plates are seeded with 500 cells per well and grown overnight. On Day 1, cells are treated with the appropriate compounds and subsequently analyzed on Days 3-5. For analysis, plates are incubated for 60 min at room temperature; 50 μL of CellTiter-Glo reagent is added to each well, and plates are mixed on an orbital shaker for 12 min. Luminescence is quantified on a standard plate luminometer. (Only for Reference) |
| 激酶實(shí)驗(yàn) | mTORC1 and mTORC2 in Vitro Kinase Assays: To produce soluble mTORC1, HEK-293T cell lines that stably express N-terminally FLAG-tagged Raptor are generated using vesicular stomatitis virus G-pseudotyped MSCV retrovirus. For mTORC2, similar HeLa cells that stably express N-terminally FLAG-tagged Protor-1 are generated. Both complexes are purified by lysing cells in 50 mm HEPES, pH 7.4, 10 mm sodium pyrophosphate, 10 mm sodium β-glycerophosphate, 100 mm NaCl, 2 mm EDTA, 0.3% CHAPS. Cells are lysed at 4 °C for 30 min, and the insoluble fraction is removed by microcentrifugation at 13,000 rpm for 10 min. Supernatants are incubated with FLAG-M2 monoclonal antibody-agarose for 1 h and then washed three times with lysis buffer and once with lysis buffer containing a final concentration of 0.5 M NaCl. Purified mTORC1 is eluted with 100 μg/mL 3×FLAG peptide in 50 mm HEPES, pH 7.4, 100 mm NaCl. Eluate can be aliquoted and stored at -80 °C. Substrates S6K1 and Akt1 are purified. Kinase assays are performed for 20 min at 30 °C in a final volume of 20 μL consisting of the kinase buffer (25 mm HEPES, pH 7.4, 50 mm KCl, 10 mm MgCl2, 500 μm ATP) and 150 ng of inactive S6K1 or Akt1 as substrates. Reactions are stopped by the addition of 80 μL of sample buffer and boiled for 5 min. Samples are subsequently analyzed by SDS-PAGE and immunoblotting. |
| 體外活性 | 方法：野生型 MEFs 細(xì)胞用 Torin 1 (250 nm) 處理 4 天，使用 CellTiterGlo viability assay 檢測(cè)細(xì)胞增殖。
結(jié)果：250 nm Torin1 完全抑制細(xì)胞增殖。[1]
方法：人類內(nèi)分泌細(xì)胞系 BON 用 Torin 1 (62.5-150 μM) 處理 3-24 h，使用 Western Blot 檢測(cè)靶點(diǎn)蛋白表達(dá)水平。
結(jié)果：Torin1 以劑量依賴的方式增加 NT 分泌；在 Torin1 處理后 3-24 h，NT 分泌增加。Torin1 處理降低了 Akt (S473) 磷酸化，p70S6K (T389) 以及 4E-BP1 (T37/46) 的磷酸化也受到抑制。[2] |
| 體內(nèi)活性 | 方法：為檢測(cè)體內(nèi)抗腫瘤活性，將 Torin 1 (20 mg/kg，20% N-mentyl-2-pyrrolidone, 40% PEG400 and 40% water) 腹腔注射給攜帶人膠質(zhì)母細(xì)胞瘤 U87MG 的 NCR nude 小鼠，每天一次，持續(xù)十天。
結(jié)果：連續(xù)給藥十天導(dǎo)致大于 99% 的腫瘤生長抑制。停止藥物治療后，腫瘤繼續(xù)生長，這表明 Torin 1 治療主要是細(xì)胞抑制性的，并且大量腫瘤細(xì)胞在治療過程中仍然存活。[3] |
| 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 | 10% DMSO+90% Saline : 0.15 mg/mL (0.25 mM), Solution.
DMSO : 1 mg/mL (1.65 mM), Sonication is recommended.
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| 關(guān)鍵字 | Torin-1 | Torin1 | Torin 1 | p110γ | mTORC2 | mTORC1 | mTOR | Mammalian target of Rapamycin | Inhibitor | inhibit | DNA-PK | DNAPK | Autophagy |
| 相關(guān)產(chǎn)品 | Oxyresveratrol | Guanidine hydrochloride | Naringin | Valproic Acid | Taurine | Gefitinib | Aceglutamide | Hydroxychloroquine | Curcumin | Stavudine | Paeonol | Sodium 4-phenylbutyrate |
| 相關(guān)庫 | 抑制劑庫 | 細(xì)胞焦亡化合物庫 | 抗乳腺癌化合物庫 | 經(jīng)典已知活性庫 | 已知活性化合物庫 | 自噬庫 | 激酶抑制劑庫 | 高選擇性抑制劑庫 | 抗衰老化合物庫 | 糖代謝化合物庫 | 免疫/炎癥分子化合物庫 | 細(xì)胞重編程化合物庫 |