Passacantilli Ilaria, Frisone Paola, De Paola Elisa, Fidaleo Marco, Paronetto Maria Paola
Laboratory of Cellular and Molecular Neurobiology, Fondazione Santa Lucia, Via del Fosso di Fiorano, 64, 00143 Rome, Italy.
University of Rome 'Foro Italico', Piazza Lauro de Bosis 6, 00135 Rome, Italy.
Nucleic Acids Res. 2017 Dec 1;45(21):12270-12284. doi: 10.1093/nar/gkx831.
Ewing sarcomas (ES) are biologically aggressive tumors of bone and soft tissues for which no cure is currently available. Most ES patients do not respond to chemotherapeutic treatments or acquire resistance. Since the PI3K/AKT/mTOR axis is often deregulated in ES, its inhibition offers therapeutic perspective for these aggressive tumors. Herein, by using splicing sensitive arrays, we have uncovered an extensive splicing program activated upon inhibition of the PI3K/AKT/mTOR signaling pathway by BEZ235. Bioinformatics analyses identified hnRNPM as a key factor in this response. HnRNPM motifs were significantly enriched in introns flanking the regulated exons and proximity of binding represented a key determinant for hnRNPM-dependent splicing regulation. Knockdown of hnRNPM expression abolished a subset of BEZ235-induced splicing changes that contained hnRNPM binding sites, enhanced BEZ235 cytotoxicity and limited the clonogenicity of ES cells. Importantly, hnRNPM up-regulation correlates with poor outcome in sarcoma patients. These findings uncover an hnRNPM-dependent alternative splicing program set in motion by inhibition of the mTOR/AKT/PI3K pathway in ES cells that limits therapeutic efficacy of pharmacologic inhibitors, suggesting that combined inhibition of the PI3K/AKT/mTOR pathway and hnRNPM activity may represent a novel approach for ES treatment.
尤因肉瘤（ES）是一种具有生物学侵袭性的骨和软组织肿瘤，目前尚无治愈方法。大多数ES患者对化疗无反应或产生耐药性。由于PI3K/AKT/mTOR轴在ES中常失调，对其抑制为这些侵袭性肿瘤提供了治疗前景。在此，通过使用剪接敏感阵列，我们发现了一个广泛的剪接程序，该程序在BEZ235抑制PI3K/AKT/mTOR信号通路后被激活。生物信息学分析确定hnRNPM是这一反应中的关键因子。hnRNPM基序在受调控外显子侧翼的内含子中显著富集，结合的接近度是hnRNPM依赖性剪接调控的关键决定因素。敲低hnRNPM表达消除了一部分包含hnRNPM结合位点的BEZ235诱导的剪接变化，增强了BEZ235的细胞毒性并限制了ES细胞的克隆形成能力。重要的是，hnRNPM上调与肉瘤患者的不良预后相关。这些发现揭示了ES细胞中通过抑制mTOR/AKT/PI3K途径启动的一个依赖hnRNPM的可变剪接程序，该程序限制了药物抑制剂的治疗效果，表明联合抑制PI3K/AKT/mTOR途径和hnRNPM活性可能是ES治疗的一种新方法。
