Signal Transduction in Tumour Development and Drug Resistance Group, Institute of Molecular Medicine and Cell Research (IMMZ), Faculty of Medicine, University of Freiburg, Freiburg, Germany.
Faculty of Biology, University of Freiburg, Freiburg, Germany.
Oncogene. 2018 Mar;37(12):1576-1593. doi: 10.1038/s41388-017-0063-5. Epub 2018 Jan 12.
BRAF mutations occur in ~10% of colorectal cancer (CRC) and are associated with poor prognosis. Inhibitors selective for the BRAF oncoprotein, the most common BRAF mutant, elicit only poor response rates in BRAF-mutant CRC as single agents. This unresponsiveness was mechanistically attributed to the loss of negative feedbacks on the epidermal growth factor receptor (EGFR) and initiated clinical trials that combine BRAF (and MEK) inhibitors, either singly or in combination, with the anti-EGFR antibodies cetuximab or panitumumab. First results of these combinatorial studies demonstrated improved efficacy, however, the response rates still were heterogeneous. Here, we show that BRAF inhibition leads to the upregulation of a variety of receptor tyrosine kinases (RTKs) in CRC cell lines, including not only the EGFR, but also human epidermal growth factor receptor (HER) 2 and HER3. Importantly, combination of the BRAF inhibitors (BRAFi) vemurafenib (PLX4032), dabrafenib, or encorafenib with inhibitors dually targeting the EGFR and HER2 (such as lapatinib, canertinib, and afatinib) significantly reduced the metabolic activity and proliferative potential of CRC cells. This re-sensitization was also observed after genetic depletion of HER2 or HER3. Interestingly, BRAF inhibitors did not only upregulate RTKs, but also increased the abundance of the GRB2-associated binders (Gab) 1 and Gab2, two important amplifiers of RTK signaling. An allele-specific shRNA-mediated knockdown of BRAF revealed that Gab2 upregulation was directly dependent on the loss of the oncoprotein and was not caused by an "off-target" effect of these kinase inhibitors. Furthermore, Gab2 and Gab2-mediated Shp2 signaling were shown to be functionally important in BRAFi resistance. These findings highlight potential new escape mechanisms to these targeted therapies and indicate that a broad suppression of RTK signaling might be beneficial and should be taken into account in future research addressing targeted therapy in BRAF-mutant CRC.
BRAF 突变发生在约 10%的结直肠癌（CRC）中，与预后不良相关。针对 BRAF 癌蛋白的抑制剂，即最常见的 BRAF 突变体，作为单一药物在 BRAF 突变型 CRC 中的反应率很差。这种无反应性在机制上归因于对表皮生长因子受体（EGFR）的负反馈的丧失，并启动了临床试验，将 BRAF（和 MEK）抑制剂单独或联合与抗 EGFR 抗体西妥昔单抗或帕尼单抗联合使用。这些组合研究的初步结果表明疗效有所提高，但反应率仍然存在异质性。在这里，我们表明 BRAF 抑制导致 CRC 细胞系中多种受体酪氨酸激酶（RTK）的上调，不仅包括 EGFR，还包括人表皮生长因子受体（HER）2 和 HER3。重要的是，BRAF 抑制剂（BRAFi）vemurafenib（PLX4032）、dabrafenib 或 encorafenib 与双重靶向 EGFR 和 HER2 的抑制剂（如 lapatinib、canertinib 和 afatinib）联合使用可显著降低 CRC 细胞的代谢活性和增殖潜力。在 HER2 或 HER3 基因缺失后也观察到这种重新敏感化。有趣的是，BRAF 抑制剂不仅上调了 RTKs，还增加了 GRB2 相关结合物（Gab）1 和 Gab2 的丰度，这两种是 RTK 信号的重要放大器。通过等位基因特异性 shRNA 介导的 BRAF 敲低揭示了 Gab2 的上调直接依赖于癌蛋白的丢失，而不是这些激酶抑制剂的“脱靶”效应引起的。此外，Gab2 和 Gab2 介导的 Shp2 信号在 BRAFi 耐药中具有功能重要性。这些发现强调了这些靶向治疗的潜在新逃逸机制，并表明广泛抑制 RTK 信号可能是有益的，并应在未来研究中考虑针对 BRAF 突变型 CRC 的靶向治疗。
