Colorectal cancer (CRC) is associated with resistance to anti-epidermal growth factor receptor (EGFR) antibodies (both acquired and intrinsic), owing to the amplification or mutation of the KRAS oncogene. However, the mechanism underlying this resistance is incompletely understood. DLD1 cells with WT (+/−) or KRAS G13D mutant allele were treated with different concentrations of Cetuximab (Cet) or panitumumab (Pab) to study the mechanism underlying the KRAS mutation-induced resistance to anti-EGFR antibodies. The function of AMPK in KRAS mutation-induced resistance to anti-EGFR antibodies in CRC cells, and the regulatory role of Bcl-2 family proteins in DLD1 cells with WT or mutated KRAS upon AMPK activation were investigated. In addition, xenograft tumor models with the nude mouse using DLD1 cells with WT or mutated KRAS were established to examine the effects of AMPK activation on KRAS mutation-mediated anti-EGFR antibody resistance. Higher levels of AMPK activity in CRC cells with wild-type KRAS treated with anti-EGFR antibody resulted in apoptosis induction. In contrast, CRC cells with mutated KRAS showed lower AMP-activated protein kinase (AMPK) activity and decreased sensitivity to the inhibitory effect of anti-EGFR antibody. CRC cells with mutated KRAS showed high levels of glycolysis and produced an excessive amount of ATP, which suppressed AMPK activation. The knockdown of AMPK expression in CRC cells with WT KRAS produced similar effects to those observed in cells with mutated KRAS and decreased their sensitivity to cetuximab. On the contrary, the activation of AMPK by metformin (Met) or 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) could overcome the KRAS-induced resistance to the anti-EGFR antibody in vivo and in vitro. The activation of AMPK resulted in the inhibition of myeloid cell leukemia 1 (Mcl-1) translation through the suppression of the mammalian target of rapamycin (mTOR) pathway. The results established herein indicate that targeting AMPK is a potentially promising and effective CRC treatment strategy.

中文翻译：

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AMPK 激活克服了结直肠癌中 KRAS 突变诱导的抗 EGFR 抗体耐药性。

由于 KRAS 癌基因的扩增或突变，结直肠癌 (CRC) 与抗表皮生长因子受体 (EGFR) 抗体（获得性和内在性）的耐药性相关。然而，这种抵抗背后的机制尚不完全清楚。用不同浓度的西妥昔单抗 (Cet) 或帕尼单抗 (Pab) 处理具有 WT (+/-) 或 KRAS G13D 突变等位基因的 DLD1 细胞，以研究 KRAS 突变诱导抗 EGFR 抗体抗性的潜在机制。研究了 AMPK 在 KRAS 突变诱导的 CRC 细胞抗 EGFR 抗体抗性中的功能，以及 Bcl-2 家族蛋白在 AMPK 激活后具有 WT 或突变 KRAS 的 DLD1 细胞中的调节作用。此外，使用带有 WT 或突变 KRAS 的 DLD1 细胞建立裸鼠异种移植肿瘤模型，以检查 AMPK 激活对 KRAS 突变介导的抗 EGFR 抗体抗性的影响。用抗 EGFR 抗体处理的具有野生型 KRAS 的 CRC 细胞中较高水平的 AMPK 活性导致细胞凋亡诱导。相比之下，具有突变 KRAS 的 CRC 细胞表现出较低的 AMP 活化蛋白激酶 (AMPK) 活性和对抗 EGFR 抗体抑制作用的敏感性降低。具有突变 KRAS 的 CRC 细胞显示出高水平的糖酵解并产生过量的 ATP，从而抑制了 AMPK 的活化。使用 WT KRAS 的 CRC 细胞中 AMPK 表达的敲低产生了与在具有突变 KRAS 的细胞中观察到的相似的效果，并降低了它们对西妥昔单抗的敏感性。相反，通过二甲双胍 (Met) 或 5-氨基咪唑-4-甲酰胺核糖核苷酸 (AICAR) 激活 AMPK 可以在体内和体外克服 KRAS 诱导的抗 EGFR 抗体耐药性。AMPK 的激活通过抑制哺乳动物雷帕霉素靶蛋白 (mTOR) 途径导致髓细胞白血病 1 (Mcl-1) 翻译的抑制。本文建立的结果表明，靶向 AMPK 是一种潜在的有前途且有效的 CRC 治疗策略。