PIK3CA基因突变及分子动力学模拟分析在子宫颈鳞状细胞癌中的意义研究

Mutation analysis of PIK3CA gene and molecular dynamics simulation analysis in cervical squamous cell carcinoma

    摘要
  • 摘要:
    目的 探讨磷脂酰肌醇-4,5-二磷酸3-激酶催化亚基α（phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha，PIK3CA）基因在子宫颈鳞癌（cervical squamous cell carcinoma，CSCC）发生发展中的意义及与临床病理特征的关系。方法：
    贵州医科大学附属医院2013年1月至2019年5月期间资料完善且诊断明确的112例CSCC组织样本，并以癌旁组织作为对照，通过NGS检测PIK3CA基因的突变，并用IHC方法进行检测蛋白表达，并通过癌症基因组图谱（the Cancer Genome Atlas，TCGA）数据库进行基因本体（gene ontology，GO）和京都基因与基因组百科全书（Kyoto encyclopedia of genes and genomes，KEGG）富集，同时利用分子动力学模拟研究E542K、E545K突变对PIK3CA蛋白的结构、功能和稳定性的影响。
    结果 在112例CSCC中，PIK3CA突变频率为27.68%（31/112），热点突变位点为E545K和E542K，明显高于癌旁组织。IHC检测发现PIK3CA蛋白在CSCC中表达明显强于癌旁组织，差异具有统计学意义（χ2=28.52，P<0.0001），且PIK3CA蛋白表达水平与Ki-67呈正相关（χ2=5.684，P=0.0171），E545K和E542K突变组PIK3CA蛋白表达明显高于野生型组（P<0.01），其蛋白表达与HPV16感染有统计学意义（χ2=9.584，P=0.002）；分子动力学模拟发现E545K和E542K突变导致PIK3CA蛋白复合物（p110α-p85α）结构、功能和稳定性均发生改变。
    结论 PIK3CA基因热点突变位点E545K和E542K在宫颈鳞癌中突变频率较高，其突变使蛋白表达增加，并协同HPV感染促进CSCC细胞增殖和侵袭，可能机制是其破坏了PIK3CA蛋白复合物中关键亚基的相互作用稳定性，导致PI3K信号通路失调，进而促进肿瘤细胞增殖，从而导致CSCC的发生发展，提示PIK3CA可能作为CSCC发生发展评估的生学物标志物或分子靶点。

     

    Abstract:
    Objective To investigate the role and clinical significance of the phosphatidylinositol-4, 5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) gene in the occurrence and development of cervical squamous cell carcinoma（CSCC）and its relationship with clinicopathological features.
    Methods A total of 112 CSCC tissue samples with complete clinicopathologicaldata and confirmed diagnoses collected between January 2013 and May 2019 at the Affiliated Hospital of Guizhou Medical University were analyzed, with adjacent tissues serving as controls. The mutations of the PIK3CA gene were detected by next-generation sequencing (NGS), and the protein expression was detected by immunohistochemistry (IHC). Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analyses were performed through the Cancer Genome Atlas (TCGA) database. Molecular dynamics simulations were used to study the effects of E542K and E545K mutations on the structure, function, and stability of the PIK3CA protein.
    Results Among the 112 CSCC tissue samples, the mutation frequency of PIK3CA was 27.68% (31/112), with E545K and E542K identified as hotspot mutations, which occurred at significantly higher frequencies than those in adjacent tissues. IHC detection found that the expression of PIK3CA protein in CSCC was significantly stronger than that in adjacent tissues, and the difference was statistically significant (χ2=28.52, P<0.0001). The expression level of PIK3CA protein was positively correlated with Ki-67 (χ2= 5.684, P=0.0171), and its expression levels were significantly higher in the E545K and E542K mutant groups than in the wild-type group (P<0.01). A significant association was also observed between PIK3CA expression and HPV16 infection (χ2=9.584, P=0.002). Molecular dynamics simulations found that E545K and E542K mutations induced structural, functional, and stability alterations in the PIK3CA protein complex (p110α-p85α).
    Conclusions The mutation frequency of the hot spot mutation sites E545K and E542K of PIK3CA gene in CSCC is relatively high. PIK3CA mutations increase protein expression and promote the proliferation and invasion of CSCC cells in collaboration with HPV infection. The underlying mechanism is that E542K and E545K mutations disrupt the stability of the interactions between key subunits in the PIK3CA protein complex, leading to dysregulation of the PI3K signaling pathway and promoting tumor cell proliferation, thereby causing the occurrence and development of CSCC. This suggests that PIK3CA may serve as a potential biomarker or molecular target for the diagnosis and treatment of CSCC.

     

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