Gao Aidi, He Chao, Chen Hengrui, Liu Qianlin, Chen Yin, Sun Jianying, Wu Chuanfeng, Pan Ya, Rocha Sonia, Wang Mu, Zhou Jundong
Suzhou Cancer Center Core Laboratory, The Affiliated Suzhou Hospital of Nanjing Medical University, Suzhou, Jiangsu, P.R. China.
Wisdom Lake Academy of Pharmacy, Xi'an Jiaotong-Liverpool University, Suzhou, Jiangsu, P.R. China.
Proteomics Clin Appl. 2025 Jan;19(1):e202400010. doi: 10.1002/prca.202400010. Epub 2024 Oct 7.
The recurrence of esophageal squamous cell carcinoma (ESCC) in radiation therapy treatment presents a complex challenge due to its resistance to radiation. However, the mechanism underlying the development of radioresistance in ESCC remains unclear. In this study, we aim to uncover the mechanisms underlying radioresistance in ESCC cells and identify potential targets for radiosensitization.
We established two radio-resistant cell lines, TE-1R and KYSE-150R, from the parental ESCC cell lines TE-1 and KYSE-150 through fractionated irradiation. A TMT-based quantitative proteomic profiling approach was applied to identify changes in protein expression patterns. Cell Counting Kit-8, colony formation, γH2AX foci immunofluorescence and comet assays were utilized to validate our findings. The downstream effectors of the DNA repair pathway were confirmed using an HR/NHEJ reporter assay and Western blot analysis. Furthermore, we evaluated the expression of potential targets in ESCC tissues through immunohistochemistry combined with mass spectrometry.
Over 2,000 proteins were quantitatively identified in the ESCC cell lysates. A comparison with radio-sensitive cells revealed 61 up-regulated and 14 down-regulated proteins in the radio-resistant cells. Additionally, radiation treatment induced 24 up-regulated and 12 down-regulated proteins in the radio-sensitive ESCC cells. Among the differentially expressed proteins, S100 calcium binding protein A6 (S100A6), glutamine gamma-glutamyltransferase 2 (TGM2), glycogen phosphorylase, brain form (PYGB), and Thymosin Beta 10 (TMSB10) were selected for further validation studies as they were found to be over-expressed in the accumulated radio-resistant ESCC cells and radio-resistant cells. Importantly, high S100A6 expression showed a positive correlation with cancer recurrence in ESCC patients. Our results suggest that several key proteins, including S100A6, TGM2, and PYGB, play a role in the development of radioresistance in ESCC.
Our results revealed that several proteins including Protein S100-A6 (S100A6), Protein-glutamine gamma-glutamyltransferase 2 (TGM2), Glycogen phosphorylase, brain form (PYGB) were involved in radio-resistance development. These proteins could potentially serve as biomarkers for ESCC radio-resistance and as therapeutic targets to treat radio-resistant ESCC cells.
食管鳞状细胞癌（ESCC）在放射治疗中出现复发是一个复杂的挑战，因为其对放疗具有抗性。然而，ESCC中放射抗性产生的潜在机制仍不清楚。在本研究中，我们旨在揭示ESCC细胞中放射抗性的潜在机制，并确定放射增敏的潜在靶点。
我们通过分次照射，从亲本ESCC细胞系TE-1和KYSE-150建立了两个放射抗性细胞系TE-1R和KYSE-150R。采用基于TMT的定量蛋白质组学分析方法来鉴定蛋白质表达模式的变化。利用细胞计数试剂盒-8、集落形成、γH2AX焦点免疫荧光和彗星试验来验证我们的发现。使用HR/NHEJ报告基因检测和蛋白质印迹分析来确认DNA修复途径的下游效应器。此外，我们通过免疫组织化学结合质谱法评估ESCC组织中潜在靶点的表达。
在ESCC细胞裂解物中定量鉴定出2000多种蛋白质。与放射敏感细胞相比，放射抗性细胞中有61种蛋白质上调，14种蛋白质下调。此外，放射治疗在放射敏感的ESCC细胞中诱导了24种蛋白质上调和12种蛋白质下调。在差异表达的蛋白质中，选择了S100钙结合蛋白A6（S100A6）、谷氨酰胺γ-谷氨酰转移酶2（TGM2）、脑型糖原磷酸化酶（PYGB）和胸腺素β10（TMSB10）进行进一步的验证研究，因为它们在累积的放射抗性ESCC细胞和放射抗性细胞中被发现过表达。重要的是，高S100A6表达与ESCC患者的癌症复发呈正相关。我们的结果表明，包括S100A6、TGM2和PYGB在内的几种关键蛋白质在ESCC放射抗性的发展中起作用。
我们的结果表明，包括蛋白质S100-A6（S100A6）、蛋白质-谷氨酰胺γ-谷氨酰转移酶2（TGM2）、脑型糖原磷酸化酶（PYGB）在内的几种蛋白质参与了放射抗性的发展。这些蛋白质可能作为ESCC放射抗性的生物标志物，并作为治疗放射抗性ESCC细胞的治疗靶点。
