Role of mitophagy in head and neck squamous cell carcinoma:Prognosis and immune insights

Objective:To explore the correlation between mitophagy and the tumor microenvironment(TME)in patients with head and neck squamous cell carcinoma(HNSCC),with an aim to enhance therapeutic efficacy for HNSCC.Methods:A machine learning-based multigene prognostic signature was developed based on mitophagy-related differentially expressed genes(MRGs)identified in The Cancer Genome Atlas cohort.This signature was correlated with the TME using gene set enrichment analysis.The association between this prognostic signature and various immunological features of the TME was explored,including status of tumor-infiltrating immune cells,expression of immune checkpoint molecules,and the immunoscore.Immunohistochemistry validated the expression of hub gene CSNK2A2 and assessed its relationship with immunomarker expression.Quantitative PCR validated CSNK2A2 knockdown in HNSCC cell lines.Functional experiments including Transwell assays to determine cell migration and invasion,Cell Counting Kit 8 assay,and 5-ethynyl-2-deoxyuridine assay were performed to confirm the role of CSNK2A2 in HNSCC.Finally,a subcutaneous xenograft model was generated in C3H mice to validate our findings.Results:The MRG-based prognostic signature showed excellent predictive performance.High-risk patients had significantly shorter progression-free and overall survival(P<0.0001)than low-risk patients.CD8+T cell infiltration was lower in high-risk groups,whereas low-risk groups showed higher immunological marker expression.Thus,the low-risk HNSCC subtype may benefit from immune therapy,while high-risk subtypes may benefit from chemotherapy(P<0.001).CSNK2A2 was highly expressed and strongly correlated with CD8 and PD-L1 based on immunohistochemistry of the HNSCC tissue microarray.CSNK2A2 knockdown reduced cell migration,invasion,and proliferation,and arrested cells in G1 phase.In vivo,it led to slower tumor growth and smaller tumor volumes.Conclusion:We established a potential prognostic signature that could improve HNSCC management in the future.CSNK2A2 may be a new biomarker to predict immunotherapy efficacy in HNSCC.

人源酪蛋白激酶Ⅱ催化亚基的构建及表达纯化

目的:构建人源CK2催化亚基pET-28a(+)-hcsnk2a1和pET-28a(+)-hcsnk2a2重组质粒,并进行原核表达纯化得到高纯度融合蛋白,为进一步开展CK2生理病理机制研究以及CK2作为肿瘤抑制靶点相关抑制剂的筛选和评价提供实验基础。方法:利用合成的人源csnk2a1和csnk2a2目的基因片段,将酶切连接得到的重组质粒经测序验证后,进行大肠杆菌感受态BL21 (DE3)/Transetta (DE3)转化。使用合适浓度IPTG诱导融合蛋白的表达以得到可溶性的融合蛋白,并应用AKTA avant蛋白纯化仪和Ni2+-NTA预装柱进行纯化,蛋白纯度最后经SDS-PAGE胶分离后考马斯亮蓝染色和Western Blot检测鉴定。结果:测序结果表明pET-28a(+)-hcsnk2a1和pET-28a(+)-hcsnk2a2质粒均成功被构建;经转化诱导表达后,成功纯化得到相对分子量为42KD的his-hcsnk2a1和38KD的his-hcsnk2a2目的融合蛋白。结论:首次成功构建得到pET-28a(+)-hcsnk2a1和pET-28a(+)-hcsnk2a2质粒;并表达纯化得到高浓度和高纯度的his-hcsnk2a1和his-hcsnk2a2融合蛋白,为后期的蛋白功能研究和抑制剂筛选评价提供了基础。