Sobecka Agnieszka, Blaszczak Wiktoria, Barczak Wojciech, Golusinski Pawel, Rubis Blazej, Masternak Michal M, Suchorska Wiktoria M, Golusinski Wojciech
Department of Head and Neck Surgery, The Greater Poland Cancer Centre, Poznan University of Medical Sciences, Garbary 15 Str, 61-866, Poznan, Poland.
Radiobiology Laboratory, The Greater Poland Cancer Centre, Garbary 15 Str, 61-866, Poznan, Poland.
J Appl Genet. 2018 Nov;59(4):453-461. doi: 10.1007/s13353-018-0458-1. Epub 2018 Aug 7.
Cancer cells, including head and neck cancer cell carcinoma (HNSCC), are characterized by an increased telomerase activity. This enzymatic complex is active in approximately 80-90% of all malignancies, and is regulated by various factors, including methylation status of hTERT gene promoter. hTERT methylation pattern has been thoroughly studied so far. It was proved that hTERT is aberrantly methylated in tumor tissue versus healthy counterparts. However, such effect has not yet been investigated in PBLs (peripheral blood leukocytes) of cancer patients. The aim of this study was to analyze the hTERT gene promoter methylation status in blood leukocytes. DNA was extracted from PBL of 92 patients with histologically diagnosed HNSCC and 53 healthy controls. Methylation status of whole hTERT promoter fragment with independent analysis of each 19 CpG sites was performed using bisulfide conversion technique followed by sequencing of PCR products. Not significant (p = 0.0532) differences in the general frequency of hTERT CpG sites methylation were detected between patients and healthy controls. However, it was discovered that some of analyzed positions (CpG islands: 1 [p = 0.0235], 5 [p = 0.0462], 8 [p = 0.0343]) are significantly more often methylated in HNSCC patients than in controls. The opposite finding was observed in case of CpG position 2 (p = 0.0210). Furthermore, closer analysis of single CpG positions revealed differences in methylation status dependent on anatomical site and TNM classification. To conclude, hTERT promoter methylation status (general or single CpG sites) would be considered as a molecular markers of HNSCC diagnostics.
癌细胞，包括头颈癌细胞癌（HNSCC），其特征在于端粒酶活性增加。这种酶复合物在所有恶性肿瘤中约80-90%具有活性，并受多种因素调节，包括hTERT基因启动子的甲基化状态。到目前为止，hTERT甲基化模式已得到充分研究。事实证明，与健康对照相比，肿瘤组织中hTERT存在异常甲基化。然而，尚未在癌症患者的外周血白细胞（PBLs）中研究这种效应。本研究的目的是分析血液白细胞中hTERT基因启动子的甲基化状态。从92例经组织学诊断为HNSCC的患者和53例健康对照的外周血白细胞中提取DNA。使用亚硫酸氢盐转化技术对整个hTERT启动子片段进行甲基化状态分析，并对每个19个CpG位点进行独立分析，随后对PCR产物进行测序。在患者和健康对照之间未检测到hTERT CpG位点甲基化总体频率的显著差异（p = 0.0532）。然而，发现一些分析位点（CpG岛：1 [p = 0.0235]、5 [p = 0.0462]、8 [p = 0.0343]）在HNSCC患者中的甲基化频率明显高于对照组。在CpG位置2的情况下观察到相反的结果（p = 0.0210）。此外，对单个CpG位置的进一步分析揭示了甲基化状态因解剖部位和TNM分类而异。总之，hTERT启动子甲基化状态（总体或单个CpG位点）可被视为HNSCC诊断的分子标志物。
