Targeting mTORC2-Dependent AKT/FOXO1/RNF125 Signaling Exploits a Therapeutic Vulnerability in c-MET-Activated and ß-catenin-Mutated Hepatocellular Carcinoma

Background and Aims: Approximately 10% of human hepatocellular carcinomas (HCC) exhibit concurrent c-MET activation and ß-catenin gain-of-function mutations, representing a clinically relevant HCC subtype. This study aimed to investigate the role of mTORC2/AKT signaling in this subtype and identify potential therapeutic targets. Approach and Results: The mTORC2/AKT cascade was activated in c-Met/ß-catenin?90 HCC lesions. Genetic ablation of Rictor, the essential mTORC2 subunit, strongly suppressed c-Met/ß-catenin?90-dependent hepatocarcinogenesis. Mechanistically, both the TSC2/mTORC1 axis and FOXO1 transcription factors functioned as critical downstream effectors of mTORC2/AKT in this model. We further identified RNF125 as a direct transcriptional target of FOXO1. RNF125 overexpression significantly inhibited tumorigenesis in the c-Met/ß-catenin?90 model and suppressed liver cancer cell growth in vitro. Notably, using an in vivo doxycycline-inducible system, we found that inducing RNF125 expression in established c-Met/ß-catenin?90 HCC suppressed tumor progression, suggesting that activation of RNF125 may have translational implications for HCC treatment. Conclusions: Our study, for the first time, established the mTORC2/AKT/FOXO1/RNF125 axis as a critical driver and therapeutic vulnerability in c-MET-activated/ß-catenin-mutated HCC. Our study filled a critical gap by defining the tumor-suppressive role of FOXO1 specifically in this HCC subtype. Furthermore, our results positioned RNF125 as a promising therapeutic target for this aggressive HCC subtype. Overall design: RNA-seq profiling was performed on pInducer20-RNF125 HepG2 cells treated with or without 2 µM doxycycline hydrochloride (Dox) for 48 hours. The experiment aimed to assess gene expression changes upon RNF125 induction.

研究背景与目的：约10%的人类肝细胞癌（hepatocellular carcinoma, HCC）同时存在c-MET激活与β-连环蛋白（β-catenin）功能获得性突变，属于具有临床意义的HCC亚型。本研究旨在探究哺乳动物雷帕霉素靶蛋白复合物2（mTORC2）/AKT信号通路在该亚型中的作用，并筛选潜在治疗靶点。

研究方法与结果：mTORC2/AKT级联反应在c-Met/β-连环蛋白Δ90 HCC病灶中被激活。遗传敲除作为mTORC2必需亚基的Rictor，可显著抑制c-Met/β-连环蛋白Δ90依赖性肝细胞癌变过程。机制层面，TSC2/mTORC1信号轴与叉头框蛋白O1（FOXO1）转录因子均为该模型中mTORC2/AKT通路的关键下游效应分子。本研究进一步鉴定出环指蛋白125（RNF125）是FOXO1的直接转录靶标。过表达RNF125可显著抑制c-Met/β-连环蛋白Δ90模型中的肿瘤发生，并在体外抑制肝癌细胞增殖。值得注意的是，通过体内多西环素（doxycycline）诱导表达系统，我们发现于已形成的c-Met/β-连环蛋白Δ90 HCC病灶中诱导RNF125表达可抑制肿瘤进展，提示激活RNF125或可为HCC治疗带来转化应用价值。

研究结论：本研究首次确立了mTORC2/AKT/FOXO1/RNF125信号轴作为c-MET激活/β-连环蛋白突变型HCC的关键驱动通路与治疗易感靶点。本研究填补了关键研究空白，明确了FOXO1在该特定HCC亚型中的抑癌作用。此外，本研究结果将RNF125定位为该侵袭性HCC亚型的潜在治疗靶点。

整体实验设计：对经2 μM盐酸多西环素（doxycycline hydrochloride, Dox）处理或未处理的pInducer20-RNF125转染HepG2细胞进行转录组测序（RNA-seq）分析，处理时长为48小时。本实验旨在评估RNF125诱导后细胞的基因表达变化。