Chordoma is a slow-growing but malignant subtype of bone sarcoma with relatively high recurrence rates and high resistance to chemotherapy. It is urgent to understand the underlying regulatory networks to determine more effective potential targets. Phosphorylative regulation is currently regarded as playing a significant role in tumorigenesis, and the use of tyrosine kinase inhibitors in clinical practice has yielded new promise for the treatment of a variety of sarcoma types.

We performed comprehensive proteomic and phosphoproteomic analyses of chordoma using four-dimensional label-free liquid chromatography–tandem mass spectrometry (LC-MS/MS) and bioinformatics analysis. The potential aberrantly expressed kinases and their functions were validated using western blotting and CCK-8 assays.

Compared with paired normal muscle tissues, 1,139 differentially expressed proteins (DEPs) and 776 differentially phosphorylated proteins (DPPs) were identified in chordoma tumor tissues. The developmentally significant Wnt-signaling pathway and oxidative phosphorylation were aberrant in chordoma. Moreover, we predicted three kinases (AURA, CDK9, and MOK) with elevated activity by kinase-pathway network analysis (KiPNA) and verified their increased expression levels. The knockdown of these kinases markedly suppressed chordoma cell growth, and this was also the case for cells treated with the CDK9 inhibitor AZD4573. We additionally examined 208 proteins whose expression and phosphorylation levels were synergetically altered.

We herein depicted the collective protein profiles of chordomas, providing insight into chordomagenesis and the potential development of new therapeutic targets.

脊索瘤是一种生长缓慢但恶性的骨肉瘤亚型，具有相对较高的复发率和对化疗的高耐药性。迫切需要了解潜在的监管网络以确定更有效的潜在目标。磷酸化调节目前被认为在肿瘤发生中起重要作用，酪氨酸激酶抑制剂在临床实践中的应用为治疗多种肉瘤类型带来了新的希望。

我们使用四维无标记液相色谱-串联质谱 (LC-MS/MS) 和生物信息学分析对脊索瘤进行了全面的蛋白质组学和磷酸化蛋白质组学分析。使用蛋白质印迹和 CCK-8 分析验证了潜在的异常表达激酶及其功能。

与配对的正常肌肉组织相比，在脊索瘤肿瘤组织中鉴定出 1,139 种差异表达蛋白 (DEP) 和 776 种差异磷酸化蛋白 (DPP)。具有发育意义的 Wnt 信号通路和氧化磷酸化在脊索瘤中异常。此外，我们通过激酶通路网络分析 (KiPNA) 预测了三种激酶（AURA、CDK9 和 MOK）具有升高的活性，并验证了它们增加的表达水平。这些激酶的敲低显着抑制了脊索瘤细胞的生长，用 CDK9 抑制剂 AZD4573 处理的细胞也是如此。我们还检查了 208 种蛋白质，它们的表达和磷酸化水平被协同改变。

我们在此描述了脊索瘤的集体蛋白质谱，提供了对脊索瘤形成和新治疗靶点的潜在发展的洞察。