CRAF inhibition induces apoptosis in melanoma cells with non-V600E BRAF mutations.
CRAF inhibition induces apoptosis in melanoma cells with non-V600E BRAF mutations.
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DOI：
10.1038/onc.2008.362
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发表时间：
2009-01-08
期刊：
影响因子：
8
通讯作者：
Herlyn, M.
中科院分区：
文献类型：
--
作者：
Smalley, K. S. M.;Xiao, M.;Villanueva, J.;Nguyen, T. K.;Flaherty, K. T.;Letrero, R.;Van Belle, P.;Elder, D. E.;Wang, Y.;Nathanson, K. L.;Herlyn, M.
在这里，我们确定了一组黑色素瘤细胞系与非V600 E突变的BRAF。发现这些G469 E和D594 G突变的黑素瘤表现出组成性水平的pERK、低水平的pMEK，并且对MEK抑制具有抗性。在使用CRAF抑制剂索拉非尼治疗后，这些细胞系发生了细胞凋亡，与线粒体去极化和AIF重新定位相关，而BRAF-V600 E突变的黑色素瘤则没有。研究表明BRAF的低活性突变体（G469 E/D594 G）通过CRAF发出信号。与BRAF不同，CRAF通过线粒体定位直接调节细胞凋亡，在线粒体定位中CRAF结合Bcl-2并磷酸化BAD。发现CRAF抑制剂索拉非尼仅在D594 G/G469 E系中诱导BAD磷酸化和Bcl-2表达的时间依赖性降低。使用慢病毒shRNA敲低CRAF抑制Bcl-2表达并诱导D594 G黑色素瘤细胞系的细胞凋亡，但在V600 E突变细胞系中没有。最后，我们在一系列异种移植研究中发现，索拉非尼在减少D594 G突变肿瘤生长方面比V600 E突变肿瘤更有效。总之，我们已经确定了一组黑色素瘤与低活性BRAF突变依赖于CRAF介导的生存活动。
Here we identify a panel of melanoma lines with non-V600E mutations in BRAF. These G469E and D594G-mutated melanomas were found to exhibit constitutive levels of pERK, low levels of pMEK and were resistant to MEK inhibition. Upon treatment with the CRAF inhibitor sorafenib, these lines underwent apoptosis, associated with mitochondrial depolarization and relocalization of AIF, whereas the BRAF-V600E mutated melanomas did not. Studies have shown low-activity mutants of BRAF (G469E/D594G) instead signal via CRAF. Unlike BRAF, CRAF directly regulates apoptosis through mitochondrial localization where it binds to Bcl-2, and phosphorylates BAD. The CRAF inhibitor sorafenib was found to induce a time-dependent reduction in both BAD phosphorylation and Bcl-2 expression in the D594G/G469E lines only. Knockdown of CRAF using a lentiviral shRNA suppressed both Bcl-2 expression and induced apoptosis in the D594G melanoma line but not in a V600E mutated line. Finally, we showed in a series of xenograft studies that sorafenib was more potent at reducing the growth of tumors with the D594G mutation than those with the V600E mutation. In summary, we have identified a group of melanomas with low-activity BRAF mutations that are reliant upon CRAF-mediated survival activity.
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