Antitumor effects of PRMT5 inhibition in sarcomas [RNA-seq]

Patients with advanced soft-tissue sarcomas (STSs) have few therapeutic options. Protein arginine methyltransferase 5 (PRMT5), an anticancer target, has been extensively investigated in recent years in epithelial tumors. To date, no data related to the biological role of PRMT5 inhibition and its potential effect as a treatment in STS have been reported. To investigate the therapeutic potential of PRMT5 targeting in STS, we first evaluated the prognostic value of PRMT5 expression in 2 different cohorts of patients with STS. We then used the potent and selective GSK3326595 (GSK595) compound to investigate the antitumor effect of the pharmacological inhibition of PRMT5 in vitro via MTT, apoptosis, cell cycle, clonogenicity and proliferation assays. In vivo studies were performed with two animal models to evaluate the effects of GSK595 on tumor growth. The mechanisms of action were investigated by RNA sequencing, metabolic pathway analysis, Western blotting and glucose uptake/lactate production assays. High PRMT5 gene expression levels were significantly associated with worsened metastasis-free survival of STS patients. GSK595 decreased the global symmetric dimethylarginine level, the proliferation rate and clonogenicity of STS cell lines in vitro and tumor growth in vivo. Moreover, PRMT5 inhibition regulated aerobic glycolysis through downregulation of key enzymes of glycolysis as well as glucose uptake and lactate production. The present study demonstrated that PRMT5 regulates STS cell metabolism and thus represents a potential therapeutic target for STS. Additional studies in diverse sarcoma subtypes will be essential to confirm and expand upon these findings. To decipher the mechanism of action of PRMT5 inhibition in STS cell lines, we explored the transcriptomic changes in IB111 cells after 10 days of treatment by RNA sequencing. IB111 cells were untreated or treated with GSK595 at the IC50 every 3 days for 10 days in triplicate. RNA was then extracted and sequenced on a NovaSeq6000 S2 platform.

晚期软组织肉瘤（soft-tissue sarcomas，STSs）患者的治疗选择极为有限。蛋白精氨酸甲基转移酶5（Protein arginine methyltransferase 5，PRMT5）作为一种抗癌靶点，近年来在上皮性肿瘤中得到了广泛研究。截至目前，尚未有关于PRMT5抑制的生物学作用及其在STSs治疗中的潜在疗效的相关研究数据发表。为探究靶向PRMT5在STSs治疗中的潜力，本研究首先在两个不同队列的STSs患者中评估了PRMT5表达的预后价值。随后，我们使用强效且具有选择性的GSK3326595（GSK595）化合物，通过MTT实验、细胞凋亡实验、细胞周期实验、克隆形成实验及细胞增殖实验，在体外探究了PRMT5药理学抑制的抗肿瘤效应。本研究通过两种动物模型开展体内实验，以评估GSK595对肿瘤生长的影响。本研究通过RNA测序、代谢通路分析、蛋白质印迹（Western blotting）实验及葡萄糖摄取/乳酸生成实验，探究了其作用机制。PRMT5基因高表达与STSs患者的无转移生存期缩短显著相关。GSK595可在体外降低STSs细胞系的整体对称二甲基精氨酸水平、增殖速率及克隆形成能力，并在体内抑制肿瘤生长。此外，PRMT5抑制可通过下调糖酵解关键酶的表达，以及降低葡萄糖摄取和乳酸生成，调控有氧糖酵解过程。本研究证实，PRMT5可调控STSs细胞代谢，因此有望成为STSs的潜在治疗靶点。未来需针对不同肉瘤亚型开展更多研究，以验证并拓展本研究的发现。为阐明PRMT5抑制在STSs细胞系中的作用机制，本研究通过RNA测序探究了IB111细胞经10天处理后的转录组变化。实验设置三组重复：IB111细胞不予处理，或以IC50浓度的GSK595每3天给药一次，持续处理10天。随后提取RNA并在NovaSeq6000 S2测序平台上进行测序。