Expression of kinase-defective mutants of c-Src in human metastatic colon cancer cells decreases Bcl-xL and increases oxaliplatin- and Fas-induced apoptosis
Expression of kinase-defective mutants of c-Src in human metastatic colon cancer cells decreases Bcl-xL and increases oxaliplatin- and Fas-induced apoptosis
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DOI：
10.1074/jbc.m408550200
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发表时间：
2004-10-29
影响因子：
4.8
通讯作者：
Dive, C
中科院分区：
文献类型：
--
作者：
Griffiths, GJ;Koh, MY;Dive, C
肿瘤对现有药物的抗药性阻碍了人类结肠癌的根治治疗。存在对有效的、肿瘤特异性化疗方法的迫切需求。非受体酪氨酸激酶c-Src在70%的人类结肠癌中过度表达，是一种易于处理的药物靶点。KM12L4A人转移性结肠癌细胞被稳定地导入两个不同的c-src激酶缺陷突变体。比较了它们对奥沙利铂、伊立替康的活性代谢物SN38以及死亡受体Fas的激活在细胞周期停滞和细胞凋亡方面的反应。这两种激酶缺陷形式的c-Src共同敏化了奥沙利铂和Fas激活诱导的细胞凋亡，但不是SN38。在SH3或SH2结构域中携带功能阻断点突变的c-Src激酶缺陷形式的细胞对奥沙利铂也同样敏感，这表明，观察到的敏感性改变是因为激酶活性的降低，而不是Src SH2-SH3支架功能的降低。奥沙利铂诱导的细胞凋亡依赖于caspase8的激活，并与Bid的切割有关，而c-Src突变体可增强这种作用。在每个稳定的细胞系中，表达c-Src的激酶缺陷突变体都降低了Bclx的水平(L)。然而，在载体对照细胞中，PP2抑制c-Src激酶活性并没有改变72 h内奥沙利铂的反应，也没有降低Bc l-x的水平(L)。这些数据表明，可能需要长期抑制Src激酶的活性，以降低Bclx(L)的水平，并使结肠癌细胞对奥沙利铂诱导的细胞凋亡敏感。
Tumor resistance to current drugs prevents curative treatment of human colon cancer. A pressing need for effective, tumor-specific chemotherapies exists. The non-receptor-tyrosine kinase c-Src is overexpressed in >70% of human colon cancers and represents a tractable drug target. KM12L4A human metastatic colon cancer cells were stably transfected with two distinct kinase-defective mutants of c-src. Their response to oxaliplatin, to SN38, the active metabolite of irinotecan ( drugs active in colon cancer), and to activation of the death receptor Fas was compared with vector control cells in terms of cell cycle arrest and apoptosis. Both kinase-defective forms of c-Src co-sensitized cells to apoptosis induced by oxaliplatin and Fas activation but not by SN38. Cells harboring kinase-defective forms of c-Src carrying function blocking point mutations in SH3 or SH2 domains were similarly sensitive to oxaliplatin, suggesting that reduction in kinase activity and not a Src SH2-SH3 scaffold function was responsible for the observed altered sensitivity. Oxaliplatin-induced apoptosis, potentiated by kinase-defective c-Src mutants, was dependent on activation of caspase 8 and associated with Bid cleavage. Each of the stable cell lines in which kinase-defective mutants of c-Src were expressed had reduced levels of Bcl-x(L). However, inhibition of c-Src kinase activity by PP2 in vector control cells did not alter the oxaliplatin response over 72 h nor did it reduce Bcl-x(L) levels. The data suggest that longer term suppression of Src kinase activity may be required to lower Bcl-x(L) levels and sensitize colon cancer cells to oxaliplatin-induced apoptosis.