Quantitative Proteome and Phosphoproteome Profiling across Three Cell Lines Revealed Potential Proteins Relevant to Nasopharyngeal Carcinoma Metastasis

Despite the substantial reduction in the mortality rates
of nasopharyngeal
carcinoma (NPC), metastasis remains the primary cause of death in
NPC cases. To explore metastasis-related proteins, we conducted proteomic
and phosphoproteomic analyses of three NPC cell lines: SUNE1 and its
subclones, 5–8F (high metastatic potential) and 6–10B
(low metastatic potential). Using TMT-based quantification, we identified
1231, 1524, and 166 differentially regulated proteins (DRPs), as well
as 177, 270, and 20 differentially regulated phosphoproteins (DRpPs)
in 5–8F/SUNE1, 6–10B/SUNE1 and 5–8F/6–10B,
respectively. These were enriched in cancer metastasis-related pathways,
including cell migration and PPAR and PI3K pathways. Notably, 5–8F
and 6–10B showed greater proteomic and phosphoproteomic similarity.
To identify key proteins involved in NPC metastasis, we focused on
the top 10 DRPs in 5–8F/6–10B. Knockdown experiments
revealed that eight of these proteins, CRABP2, DNAJC15, NACAD, MYL9,
DPYSL3, MAOA, MCAM, and S100A2, significantly influenced cell migration
or invasion, with CRABP2, NACAD, and DPYSL3 dramatically enhancing
these processes. Notably, DNAJC15 and NACAD are identified for the
first time as novel metastasis-related proteins. Our findings demonstrate
the effectiveness of this approach in identifying NPC metastasis biomarker
candidates and offer new insights into underlying metastasis mechanisms,
thus laying the groundwork for future research endeavors.

尽管鼻咽癌（nasopharyngeal carcinoma, NPC）的死亡率已显著降低，肿瘤转移仍是此类患者的首要致死原因。为探索与鼻咽癌转移相关的蛋白，我们对三株鼻咽癌细胞系开展了蛋白质组学与磷酸化蛋白质组学分析：SUNE1及其亚克隆5–8F（高转移潜能）与6–10B（低转移潜能）。基于TMT定量技术，我们分别在5–8F/SUNE1、6–10B/SUNE1及5–8F/6–10B三组比较中，鉴定出1231、1524和166个差异调控蛋白（differentially regulated proteins, DRPs），以及177、270和20个差异调控磷酸化蛋白（differentially regulated phosphoproteins, DRpPs）。这些分子显著富集于癌症转移相关通路，包括细胞迁移、过氧化物酶体增殖物激活受体（PPAR）与磷脂酰肌醇3-激酶（PI3K）信号通路。值得注意的是，5–8F与6–10B两株细胞的蛋白质组与磷酸化蛋白质组特征更为相似。为筛选参与鼻咽癌转移的关键蛋白，我们聚焦于5–8F/6–10B组中排名前10的差异调控蛋白。敲低实验结果显示，其中8种蛋白——CRABP2、DNAJC15、NACAD、MYL9、DPYSL3、MAOA、MCAM及S100A2——可显著影响细胞迁移与侵袭能力，其中CRABP2、NACAD与DPYSL3可显著增强上述过程。尤为关键的是，DNAJC15与NACAD首次被鉴定为新型转移相关蛋白。本研究证实了该策略在筛选鼻咽癌转移生物标志物候选物方面的有效性，为阐明转移潜在机制提供了新视角，亦为后续研究奠定了坚实基础。