| 名稱 | Fedratinib |
| 描述 | Fedratinib (TG-101348) is a JAK2 inhibitor that is selective, ATP-competitive, and orally active, inhibiting JAK2 and JAK2V617F kinases (IC50=3 nM). Fedratinib induces apoptosis in tumor cells and may be used in myeloproliferative diseases. |
| 細(xì)胞實(shí)驗(yàn) | Cells were treated with DMSO and increasing concentrations of inhibitor for 4 hr in RPMI-1640 before collected in 13 Cell Lysis Buffer, containing 1 mM PMSF, and protease inhibitor cocktail tablets. Protein lysates were quantified with BCA assay. Similar protein amounts were mixed with Laemmli sample buffer plus b-mercaptoethanol, boiled for 5 min, and separated on a 4%–15% Tris-HCl gradient electrophoresis gel. Gels were blotted onto a 0.45 mm nitrocellulose membrane, which was blocked in 5% nonfat dry milk and incubated with primary antibodies in either blocking solution or 5% BSA. The membranes were subsequently incubated with a mixture of donkey anti-rabbit IgG conjugated with infrared fluorophore (700 nm emission) and goat anti-mouse IgG conjugated with infrared fluorophore (800 nm emission). Following washing with PBS, the membranes were scanned on an Odyssey scanner to detect total (red) and phospho-STAT5 (green) proteins [1]. |
| 激酶實(shí)驗(yàn) | IC50 values for TG101348 are determined commercially using the InVitrogen kinase profiling service for a 223 kinase screen that included JAK2 and JAK2V617F or Carna Biosciences for the screen of all Janus kinase family members including JAK1 and Tyk2. ATP concentration is set to approximately the Km value for each kinase [1]. |
| 動(dòng)物實(shí)驗(yàn) | The murine BM transplant model was generated and analyzed exactly as previously described. Briefly, C57BL/6 mice were intravenously injected with 1×10^6 whole bone marrow expressing JAK2V617F. Full development of disease was assessed with differential peripheral blood counts at day 26 after bone marrow transplantation. TG101348 was administered by oral gavage twice daily (b.i.d.) at 60 mg/kg, 120 mg/kg, or placebo from day 28 on for 42 days. Differential blood counts were assessed by retro-orbital nonlethal eye bleeds using EDTA glass capillary tubes before study initiation, during the study, and at study endpoints. C57/Bl6 mice were sacrificed at study endpoint or at times indicated based on an IUCAC-approved protocol that includes assessment of morbidity by > 10% loss of weight, scruffy appearance, lethargy, and/or splenomegaly extending across the midline. For histopathology, tissues were fixed in 10% neutral buffered formalin, embedded in paraffin, and stained with hematoxylin and eosin or, to assess for fibrosis, stained with reticulin. Images of histological slides were obtained on a Nikon Eclipse E400 microscope equipped with a SPOT RT color digital camera model 2.1.1. Images were analyzed in Adobe Photoshop 6.0. For flow cytometry, cells were washed in PBS, washed in 2% fetal bovine serum, blocked with Fc-Block for 10 min on ice, and stained with monoclonal antibodies in PBS and 2% FCS for 30 min on ice. Antibodies used were allophycocyanin (APC)-conjugated ter119, Gr-1, CD4, and B220 and phycoerythrin (PE)-conjugated, Mac1, CD8 (all 1:200), and CD71(1:100) rat anti-mouse. After washing, cells were resuspended in PBS and 2% FCS containing 0.5 mg/ml 7-amino-actinomycin D (7-AAD) to allow discrimination of nonviable cells. Flow cytometry was performed on a FACS Calibur cytometer, at least 10,000 events were acquired, and data were analyzed using FloJo software.The results are presented as graphs and representative dot plots of viable cells selected on the basis of scatter and 7-AAD staining [1]. |
| 體外活性 | 方法：人成紅細(xì)胞白血病細(xì)胞 HEL 和 Ba/F3 JAK2V617F 細(xì)胞用 Fedratinib (0-30 μM) 處理 72 h，使用 XTT assay 檢測(cè)細(xì)胞增殖。
結(jié)果：Fedratinib 抑制了攜帶 JAK2V617F 突變的 HEL 細(xì)胞系以及表達(dá)人 JAK2V617F (Ba/F3 JAK2V617F) 的鼠前 B 細(xì)胞系的增殖，兩種細(xì)胞系的 IC50 值分別為 305 nM 和 270 nM。[1]
方法：人骨髓間充質(zhì)干細(xì)胞 hMSC-TERT 用 Fedratinib (3 μM) 處理 10 天，進(jìn)行堿性磷酸酶 (ALP) 染色檢測(cè)成骨分化。
結(jié)果：Fedratinib 處理的 hMSC-TERT 細(xì)胞表現(xiàn)出 ALP 產(chǎn)生的顯著減少。與對(duì)照細(xì)胞相比，成骨細(xì)胞分化誘導(dǎo)后第 10 天的 ALP 活性測(cè)量值降低。Fedratinib 對(duì) hMSC-TERT 細(xì)胞的活力沒(méi)有產(chǎn)生顯著影響。[2] |
| 體內(nèi)活性 | 方法：為研究對(duì)已建立的真性紅細(xì)胞增多癥的療效，將 Fedratinib (60-120 mg/kg) 灌胃給藥給 C57BL/6 小鼠骨髓移植模型，每天兩次，持續(xù) 42 天。
結(jié)果：在接受治療的動(dòng)物中，紅細(xì)胞壓積和白細(xì)胞計(jì)數(shù)在統(tǒng)計(jì)學(xué)上顯著降低，髓外造血的減少/消除呈劑量依賴性，至少在某些情況下，有證據(jù)表明骨髓纖維化減輕。[1] |
| 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+40% PEG300+5% Tween 80+45% Saline : 9.3 mg/mL (17.73 mM), Please add co-solvents sequentially, clarifying the solution as much as possible before adding the next one. Dissolve by heating and/or sonication if necessary. Working solution is recommended to be prepared and used immediately.
DMSO : 50 mg/mL (95.3 mM)
H2O : < 1 mg/mL (insoluble or slightly soluble)
Ethanol : < 1 mg/mL (insoluble or slightly soluble)
|
| 關(guān)鍵字 | Apoptosis | Fedratinib | STAT5 | Inhibitor | Janus kinase | phosphorylation | FLT3 | myeloproliferative | SAR-302503 | JAK2V617F | orally | inhibit | SAR302503 | anti-proliferation | JAK2 | TG 101348 | TG101348 | JAK | RET | anti-cancer |
| 相關(guān)產(chǎn)品 | L-Glutamic acid | Metronidazole | 5-Fluorouracil | Dextran sulfate sodium salt (MW 4500-5500) | Stavudine | Tributyrin | Myricetin | Sorafenib | L-Ascorbic acid | Acetylcysteine | Salicylic acid | Sodium 4-phenylbutyrate |
| 相關(guān)庫(kù) | 抑制劑庫(kù) | 經(jīng)典已知活性庫(kù) | 已知活性化合物庫(kù) | EMA 上市藥物庫(kù) | 激酶抑制劑庫(kù) | FDA 上市藥物庫(kù) | 藥物功能重定位化合物庫(kù) | FDA 上市激酶抑制劑庫(kù) | 抗癌臨床化合物庫(kù) | 抗癌藥物庫(kù) |