Modulation of the c-Met/hepatocyte growth factor pathway in small cell lung cancer.
Modulation of the c-Met/hepatocyte growth factor pathway in small cell lung cancer.
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发表时间：
2002-02
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通讯作者：
G. Maulik;T. Kijima;P. Ma;S. Ghosh;Jeffrey Lin;G. Shapiro;E. Schaefer;E. Tibaldi;B. Johnson;R. Salgia
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作者：
G. Maulik;T. Kijima;P. Ma;S. Ghosh;Jeffrey Lin;G. Shapiro;E. Schaefer;E. Tibaldi;B. Johnson;R. Salgia
c-Met 受体酪氨酸激酶及其配体 HGF（肝细胞生长因子）已被证明参与血管生成、细胞运动、生长、侵袭和分化。 c-Met/HGF 轴在小细胞肺癌 (SCLC) 中的作用此前尚未有报道。我们通过免疫印迹法测定了七种 SCLC 细胞系中 p170(c-Met) 前体和 p140(c-Met) β 链的表达。我们使用表达丰富 c-Met 的 SCLC 细胞系 H69 来研究 c-Met 激活的功能和下游影响。用 HGF 刺激 H69 细胞（40 ng/ml，6 小时刺激）显着改变了 SCLC 细胞的细胞运动性，丝状伪足和膜褶皱（表现为膜起泡）的形成增加，以及细胞簇迁移增加。我们进一步研究了HGF/c-Met在H69细胞系中的信号转导通路。用 HGF（40 ng/ml，>24 小时，1 小时时最大）刺激 H69 使形成的活性氧数量增加 34%。 H69 细胞的 HGF 刺激（40 ng/ml，7.5 分钟刺激）显示在 M(r) 68,000、120,000-140,000 和 200,000 处鉴定的酪氨酸磷酸化条带增加。其中一些酪氨酸磷酸化条带被鉴定为粘着斑蛋白桩蛋白、FAK、PYK2 和 c-Met 受体本身。磷酸化特异性抗体显示桩蛋白第 31 位氨基酸 (a.a.) 处的酪氨酸以及 a.a. 处的自磷酸化位点。 p125FAK 的 397 和 a.a. PYK2 的 402 位响应 HGF/c-Met 信号传导而被磷酸化。我们还证明，Hsp90 抑制剂格尔德霉素（也影响 c-Met）在 72 小时内使四种 SCLC 细胞系中的四种的生长和活力降低了 25% 至 85%。格尔德霉素引起 SCLC 细胞凋亡，并导致 Hsp70 水平增加，但不增加 Hsp90 水平。这些结果表明 c-Met/HGF 通路在 SCLC 中发挥作用，针对该通路进行新的治疗将是有用的。
The c-Met receptor tyrosine kinase and its ligand HGF (hepatocyte growth factor) have been shown to be involved in angiogenesis, cellular motility, growth, invasion, and differentiation. The role of c-Met/HGF axis in small cell lung cancer (SCLC) has not been reported previously. We have determined the expression of p170(c-Met) precursor and p140(c-Met) beta-chain in seven SCLC cell lines by immunoblotting. We used the SCLC cell line H69, which expressed an abundant amount of c-Met to study the function and downstream effects of c-Met activation. Stimulation of H69 cells with HGF (40 ng/ml, 6-h stimulation) significantly altered cell motility of the SCLC cells with increased formation of filopodia and membrane ruffling, characterized as membrane blebbing, as well as increased migration of the cellular clusters were seen. We have further studied the signal transduction pathways of HGF/c-Met in the H69 cell line. The stimulation of H69 with HGF (40 ng/ml, >24 h, maximal at 1 h) increased the amount of reactive oxygen species formed by 34%. HGF stimulation (40 ng/ml, 7.5-min stimulation) of H69 cells showed increased tyrosine phosphorylated bands identified at M(r) 68,000, 120,000-140,000, and 200,000. Some of these tyrosine-phosphorylated bands were identified as the focal adhesion proteins paxillin, FAK, PYK2, and the c-Met receptor itself. Phospho-specific antibodies show that tyrosines at amino acid (a.a.) 31 of paxillin, and autophosphorylation sites at a.a. 397 of p125FAK, and a.a. 402 of PYK2 are phosphorylated in response to HGF/c-Met signaling. We also demonstrate that the Hsp90 inhibitor geldanamycin, which also affects c-Met, reduced the growth and viability of four of four SCLC cell lines by 25% to 85%, over a 72-h time period. Geldanamycin caused apoptosis of SCLC cells, as well as led to increased levels of Hsp70 but not Hsp90. These results demonstrate that c-Met/HGF pathway is functional in SCLC, and it would be useful to target this pathway toward novel therapy.