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Masitinib抑制Kit功能获得性突变体,包括V559D突变体和Δ27小鼠突变体,在Ba\u002FF3细胞中IC50为3和5nM。 Masitinib抑制包括HMC-1α155和FMA3在内的肥大细胞瘤细胞的增殖,IC50分别为10和30 nM [1]。 Masitinib在两种新型ISS细胞系中抑制细胞生长和PDGFR磷酸化,这表明Masitinib显示出对原发性和转移性ISS细胞系的活性,并可能有助于ISS的临床管理[2]。",{"key":214,"value":215},"体内研究","Masitinib在30 mg\u002Fkg时抑制肿瘤生长并增加表达Δ27的Ba\u002FF3肿瘤模型的中位存活时间,无心脏毒性或遗传毒性[1]。与狗中的安慰剂相比,Masitinib(12.5 mg\u002Fkg\u002Fd,po)可增加总体TTP(肿瘤扩展时间)[3]。 Masitinib \u002F吉西他滨的组合在体外对吉西他滨难治性细胞系Mia Paca2和Panc1的增殖具有协同作用,并且在较小程度上对小鼠的Mia Paca-2胰腺肿瘤具有协同作用[4]。",{"key":217,"value":218},"激酶实验","将96孔微量滴定板用0.25mg \u002F mL聚（Glu，Tyr 4：1）包被过夜，用250μL洗涤缓冲液（10mM磷酸盐缓冲盐水[pH 7.4]和0.05％吐温20）漂洗两次并干燥在室温下保持2小时。在室温下进行测定，最终体积为50μL，在含有ATP的激酶缓冲液（10mM MgCl 2,1mM MnCl 2,1mM原钒酸钠，20mM HEPES，pH 7.8）中，每种浓度至少为Km的两倍。酶和适量的重组酶确保线性反应速率。在引入酶时开始反应，并且每5mol \u002F Lurea混合物加入一个反应体积（50μL）的100mM EDTA终止反应。将板洗涤三次并与1：30,000辣根过氧化物酶缀合的抗磷酸酪氨酸单克隆抗体一起温育，然后洗涤三次并与四甲基联苯胺一起温育。通过分光光度法在450nm处定量最终反应产物。",{"key":220,"value":221},"细胞实验","对于Ba \u002F F3细胞增殖的测定，将微量滴定板以总共104个细胞\u002F孔接种于100μL含有10％胎牛血清的RPMI 1640培养基中，37℃。这些补充或不补充来自X63-IL-3细胞的0.1％条件培养基或250ng \u002F mL鼠SCF。激活Kit的鼠SCF从产生SCF的CHO细胞的条件培养基中纯化。细胞在37℃下与马西替尼一起生长48小时，然后与10μL\u002F孔的WST-1试剂在37℃下孵育3小时。形成的甲dye染料的量通过使用扫描多孔分光光度计在450nm处的吸光度来量化。没有细胞的空白孔用作分光光度计的背景对照。",{"key":223,"value":224},"动物实验","雄性Nog-SCID小鼠（7周龄）在20±1℃下在12小时光照\u002F 12小时黑暗循环中处于无特定病原体的条件下，随意获取食物和过滤水。如上所述培养Mia Paca-2细胞。在第0天（D0），将小鼠在200μLPBS中注射107个Mia Paca-2细胞到右侧腹中。允许肿瘤生长1.5至4周，直至达到所需的肿瘤大小（约200mm 3）。在第28天，将动物分配到四个治疗组（每组n = 7至8），确保每组的平均体重和肿瘤体积匹配良好。然后给予治疗长达4周，之后处死动物。治疗方法包括：a）对照组每日无菌水，b）腹腔注射（ip）50 mg \u002F kg吉西他滨，每周两次，c）每日灌胃100 mg \u002F kg masitinib，或d）腹腔注射联合注射每周两次50mg \u002F kg吉西他滨，每日灌胃100mg \u002F kg马西替尼。用卡尺测量肿瘤大小，并使用以下公式估计肿瘤体积：体积=（长度×宽度2）\u002F 2。肿瘤生长抑制率计算为（100）×（治疗组的中位肿瘤体积）\u002F（对照组的中位肿瘤体积）。",{"key":226,"value":227},"参考文献","[1].\nDubreuil P, et al. Masitinib (AB1010), a Potent and Selective Tyrosine Kinase Inhibitor Targeting KIT. PLoS One, 2009, 4(9), e7258.\n[2].\nLawrence J, et al. Masitinib demonstrates anti-proliferative and pro-apoptotic activity in primary and metastatic feline injection-site sarcoma cells. Vet Comp Oncol, 2011, doi: 10.1111\u002Fj.1476-5829.2011.00291.x.\n[3].\nHahn KA, et al. Masitinib is safe and effective for the treatment of canine mast cell tumors. J Vet Intern Med, 2008, 22(6), 1301-1309.\n[4].\nHumbert M, et al. Masitinib combined with standard gemcitabine chemotherapy: in vitro and in vivo studies in human pancreatic tumour cell lines and ectopic mouse model. 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