Laboratory of Molecular Tumor Pathology and Cancer Systems Biology, Institute of Pathology, Charité Universitätsmedizin Berlin, Charitéplatz 1, D-10117, Berlin, Germany.
Laboratory of Molecular Tumor Pathology and Cancer Systems Biology, Institute of Pathology, Charité Universitätsmedizin Berlin, Charitéplatz 1, D-10117, Berlin, Germany; Integrative Research Institute Life Sciences, Humboldt University Berlin, Philippstraße 13, Building 18, D-10115, Berlin, Germany.
Adv Biol Regul. 2021 Jan;79:100778. doi: 10.1016/j.jbior.2020.100778. Epub 2020 Dec 31.
In colorectal cancer (CRC), the prevalence of NRAS mutations (5-9%) is inferior to that of KRAS mutations (40-50%). NRAS mutations feature lately during tumour progression and drive resistance to anti-EGFR therapy in KRAS wild-type tumours. To elucidate specific functions of NRAS mutations in CRC, we expressed doxycycline-inducible G12D and Q61K mutations in the CRC cell line Caco-2. A focused phospho-proteome analysis based on the Bio-Plex platform, which interrogated the activity of MAPK, PI3K, mTOR, STAT, p38, JNK and ATF2, did not reveal significant differences between Caco-2 cells expressing NRAS, NRAS and KRAS. However, phenotypic read-outs were different. The NRAS Q61K mutation promoted anchorage-independent proliferation and tumorigenicity, similar to features driven by canonical KRAS mutations. In contrast, expression of NRAS resulted in reduced proliferation and apoptosis. At the transcriptome level, we saw upregulation of cytokines and chemokines. IL1A, IL11, CXCL8 (IL-8) and CCL20 exhibited enhanced secretion into the culture medium. In addition, RNA sequencing results indicated activation of the IL1-, JAK/STAT-, NFκB- and TNFα signalling pathways. These results form the basis for an NRAS-driven inflammatory phenotype in CRC.
在结直肠癌（CRC）中，NRAS 突变的发生率（5-9%）低于 KRAS 突变（40-50%）。NRAS 突变在肿瘤进展过程中较晚出现，并导致 KRAS 野生型肿瘤对抗 EGFR 治疗产生耐药性。为了阐明 NRAS 突变在 CRC 中的特定功能，我们在 CRC 细胞系 Caco-2 中表达了可诱导的 G12D 和 Q61K 突变。基于 Bio-Plex 平台的靶向磷酸化蛋白质组分析，该分析检测了 MAPK、PI3K、mTOR、STAT、p38、JNK 和 ATF2 的活性，并未显示表达 NRAS、NRAS 和 KRAS 的 Caco-2 细胞之间存在显著差异。然而，表型读数有所不同。NRAS Q61K 突变促进了非锚定依赖性增殖和致瘤性，类似于经典 KRAS 突变驱动的特征。相比之下，表达 NRAS 会导致增殖减少和细胞凋亡。在转录组水平上，我们观察到细胞因子和趋化因子的上调。IL1A、IL11、CXCL8（IL-8）和 CCL20 的分泌到培养基中增强。此外，RNA 测序结果表明 IL1-、JAK/STAT-、NFκB-和 TNFα 信号通路被激活。这些结果为 CRC 中的 NRAS 驱动的炎症表型奠定了基础。
