Heng Jianfu, Guo Xinwu, Wu Wenhan, Wang Yue, Li Guoli, Chen Ming, Peng Limin, Wang Shouman, Dai Lizhong, Tang Lili, Wang Jun
The State Key Laboratory of Medical Genetics & School of Life Sciences, Central South University, Changsha, Hunan, China.
Sanway Gene Technology Inc., Changsha, Hunan, China.
PLoS One. 2017 Mar 16;12(3):e0174022. doi: 10.1371/journal.pone.0174022. eCollection 2017.
As downstream mediators of PI3K /PTEN /AKT /mTORC1 pathway, the AKT isoforms play critical roles in tumorgenesis. Although the pleiotropic effects of AKT1 in breast cancer have been reported, the genetic and epigenetic characteristics of AKT1 promoter region in breast cancer remains to be identified. In this study we aimed to investigate the promoter mutation spectrum, methylation and gene expression pattern of AKT1 and their relationship with breast cancer.
By using PCR target sequence enrichment and next-generation sequencing technology, we sequenced AKT1 promoter region in pairs of breast tumor and normal tissues from 95 unselected Chinese breast cancer patients. The methylation of the promoter region and the expression profile of AKT1 in the same cohort were detected with bisulfite next-generation sequencing and qPCR, respectively.
We identified 28 somatic mutations in 23 of the 95 (24.2%) breast cancer samples. And 19 of the 28 mutations were located in transcription factor (TF) binding sites. In the 23 patients with somatic mutations, no significant change of methylation or expression was found comparing with other patients. AKT1 promoter region was significantly hypo-methylated in tumor compared with matched normal tissue (P = 0.0014) in the 95 patients. The expression of AKT1 was significantly suppressed in tumor tissue (P = 0.0375). In clinicopathological factor analysis, AKT1 showed significant hypo-methylation (P = 0.0249) and suppressed expression (P = 0.0375) in HER2 negative subtype. And a trend of decrease in expression level (P = 0.0624) of AKT1 in the ER negative subtype was observed, which is significantly decreased in basal-like breast tumor (P = 0.0328).
Hypo-methylation and suppressed expression of AKT1 was observed to be associated with breast cancer in our cohort. The methylation and expression of AKT1 were both significantly associated with HER2 status. The promoter mutation of AKT1 did not show significant association with its methylation and expression status. These results suggested that the promoter mutation, methylation and gene expression of AKT1 may play distinct roles in tumorgenesis of breast cancer and the integrated analysis of methylation and expression of AKT1 might serve as potential biomarkers for diagnosis and classification of breast cancer.
作为PI3K/PTEN/AKT/mTORC1信号通路的下游介质，AKT亚型在肿瘤发生中起关键作用。尽管已有报道AKT1在乳腺癌中具有多效性作用，但其在乳腺癌中的启动子区域的遗传和表观遗传特征仍有待确定。本研究旨在探讨AKT1的启动子突变谱、甲基化和基因表达模式及其与乳腺癌的关系。
采用PCR靶向序列富集和二代测序技术，对95例未经选择的中国乳腺癌患者的肿瘤组织和正常组织配对样本中的AKT1启动子区域进行测序。分别采用亚硫酸氢盐二代测序和qPCR检测同一队列中启动子区域的甲基化和AKT1的表达谱。
在95例（24.2%）乳腺癌样本中的23例中鉴定出28个体细胞突变。28个突变中有19个位于转录因子（TF）结合位点。在23例发生体细胞突变的患者中，与其他患者相比，甲基化或表达无显著变化。在95例患者中，与配对的正常组织相比，肿瘤组织中AKT1启动子区域显著低甲基化（P = 0.0014）。肿瘤组织中AKT1的表达显著受抑制（P = 0.0375）。在临床病理因素分析中，HER2阴性亚型中AKT1显示出显著的低甲基化（P = 0.0249）和表达受抑制（P = 0.0375）。在ER阴性亚型中观察到AKT1表达水平有下降趋势（P = 0.0624），在基底样乳腺癌中显著下降（P = 0.0328）。
在我们的队列中观察到AKT1的低甲基化和表达受抑制与乳腺癌相关。AKT1的甲基化和表达均与HER2状态显著相关。AKT1的启动子突变与其甲基化和表达状态无显著关联。这些结果表明，AKT1的启动子突变、甲基化和基因表达可能在乳腺癌的肿瘤发生中发挥不同作用，对AKT1甲基化和表达的综合分析可能作为乳腺癌诊断和分类的潜在生物标志物。
