Oncogenic mutation of RAS results in aberrant cellular signaling and is responsible for more than 30% of all human tumors. Therefore, pharmacologic modulation of RAS has attracted great interest as a therapeutic strategy. Our laboratory has recently discovered small molecules that activate Son of Sevenless (SOS)–catalyzed nucleotide exchange on RAS and inhibit downstream signaling. Here, we describe how pharmacologically targeting SOS1 induced biphasic modulation of RAS-GTP and ERK phosphorylation levels, which we observed in a variety of cell lines expressing different RAS-mutant isoforms. We show that compound treatment caused an increase in phosphorylation at ERK consensus motifs on SOS1 that was not observed with the expression of a non-phosphorylatable S1178A SOS1 mutant or after pretreatment with an ERK inhibitor. Phosphorylation at S1178 on SOS1 is known to inhibit the association between SOS1 and GRB2 and disrupt SOS1 membrane localization. Consistent with this, we show that wild-type SOS1 and GRB2 dissociated in a time-dependent fashion in response to compound treatment, and conversely, this interaction was enhanced with the expression of an S1178A SOS1 mutant. Furthermore, in cells expressing either S1178A SOS1 or a constitutively membrane-bound CAAX box tagged SOS1 mutant, we observed elevated RAS-GTP levels over time in response to compound, as compared with the biphasic changes in RAS-GTP exhibited in cells expressing wild-type SOS1. These results suggest that small molecule targeting of SOS1 can elicit a biphasic modulation of RAS-GTP and phospho-ERK levels through negative feedback on SOS1 that regulates the interaction between SOS1 and GRB2. Mol Cancer Ther; 17(5); 1051–60. ©2018 AACR.

中文翻译：

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小分子介导的 RAS 激活引起磷酸化 ERK 水平的双相调节，磷酸化 ERK 水平通过对 SOS1 的负反馈进行调节

RAS 的致癌突变导致异常的细胞信号传导，并导致超过 30% 的人类肿瘤。因此，RAS的药理学调节作为一种治疗策略引起了极大的兴趣。我们的实验室最近发现了小分子，这些小分子可以激活 Son of Sevenless (SOS) 催化的 RAS 上的核苷酸交换并抑制下游信号传导。在这里，我们描述了靶向 SOS1 的药理学如何诱导 RAS-GTP 和 ERK 磷酸化水平的双相调节，我们在表达不同 RAS 突变亚型的各种细胞系中观察到了这种调节。我们表明，复合处理导致 SOS1 上 ERK 共有基序的磷酸化增加，这在非磷酸化 S1178A SOS1 突变体的表达或用 ERK 抑制剂预处理后未观察到。已知 SOS1 上 S1178 的磷酸化会抑制 SOS1 和 GRB2 之间的关联并破坏 SOS1 膜定位。与此一致，我们表明野生型 SOS1 和 GRB2 响应于复合处理以时间依赖性方式解离，相反，这种相互作用随着 S1178A SOS1 突变体的表达而增强。此外，在表达 S1178A SOS1 或标记为 SOS1 突变体的组成型膜结合 CAAX 盒的细胞中，我们观察到 RAS-GTP 水平随着时间的推移响应化合物而升高，与表达野生型细胞中 RAS-GTP 的双相变化相比键入 SOS1。这些结果表明，靶向 SOS1 的小分子可以通过调节 SOS1 和 GRB2 之间相互作用的 SOS1 上的负反馈引起 RAS-GTP 和磷酸化 ERK 水平的双相调节。摩尔癌症治疗；17（5）；1051-60。©2018 AACR。