Department of Pathology, Moores Cancer Center, University of California San Diego, La Jolla, California, USA.
Nat Med. 2011 Nov 13;17(12):1641-5. doi: 10.1038/nm.2464.
RAF kinases regulate cell proliferation and survival and can be dysregulated in tumors. The role of RAF in cell proliferation has been linked to its ability to activate mitogen-activated protein kinase kinase 1 (MEK) and mitogen-activated protein kinase 1 (ERK). Here we identify a MEK-independent role for RAF in tumor growth. Specifically, in mitotic cells, CRAF becomes phosphorylated on Ser338 and localizes to the mitotic spindle of proliferating tumor cells in vitro as well as in murine tumor models and in biopsies from individuals with cancer. Treatment of tumors with allosteric inhibitors, but not ATP-competitive RAF inhibitors, prevents CRAF phosphorylation on Ser338 and localization to the mitotic spindle and causes cell-cycle arrest at prometaphase. Furthermore, we identify phospho-Ser338 CRAF as a potential biomarker for tumor progression and a surrogate marker for allosteric RAF blockade. Mechanistically, CRAF, but not BRAF, associates with Aurora kinase A (Aurora-A) and Polo-like kinase 1 (Plk1) at the centrosomes and spindle poles during G2/M. Indeed, allosteric or genetic inhibition of phospho-Ser338 CRAF impairs Plk1 activation and accumulation at the kinetochores, causing prometaphase arrest, whereas a phospho-mimetic Ser338D CRAF mutant potentiates Plk1 activation, mitosis and tumor progression in mice. These findings show a previously undefined role for RAF in tumor progression beyond the RAF-MEK-ERK paradigm, opening new avenues for targeting RAF in cancer.
RAF 激酶调节细胞增殖和存活，并且在肿瘤中可能失调。RAF 在细胞增殖中的作用与其激活丝裂原活化蛋白激酶激酶 1（MEK）和丝裂原活化蛋白激酶 1（ERK）的能力有关。在这里，我们确定了 RAF 在肿瘤生长中的 MEK 非依赖性作用。具体而言，在有丝分裂细胞中，CRAF 在丝氨酸 338 上被磷酸化，并在体外增殖肿瘤细胞的有丝分裂纺锤体中以及在鼠肿瘤模型和癌症患者的活检中定位于有丝分裂纺锤体。用别构抑制剂而不是 ATP 竞争性 RAF 抑制剂治疗肿瘤可防止 CRAF 在丝氨酸 338 上的磷酸化和向有丝分裂纺锤体的定位，并导致细胞周期在前期停滞。此外，我们确定磷酸化 Ser338 CRAF 作为肿瘤进展的潜在生物标志物和别构 RAF 阻断的替代标志物。在机制上，CRAF 而不是 BRAF，在 G2/M 期间与中心体和纺锤体极上的 Aurora 激酶 A（Aurora-A）和 Polo 样激酶 1（Plk1）相关。实际上，别构或遗传抑制磷酸化 Ser338 CRAF 会损害 Plk1 在动粒处的激活和积累，导致前期停滞，而磷酸化模拟 Ser338D CRAF 突变体能增强 Plk1 的激活、有丝分裂和在小鼠中的肿瘤进展。这些发现表明 RAF 在 RAF-MEK-ERK 范例之外在肿瘤进展中具有以前未定义的作用，为癌症中靶向 RAF 开辟了新途径。
