LINE-1 and Alu Methylation Patterns in Lymph Node Metastases of Head and Neck Cancers

Abstract

<b>Background:</b> The potential use of hypomethylation of Long INterspersed Element 1 (LINE-1) and Alu elements(Alu) as a biomarker has been comprehensively assessed in several cancers, including head and neck squamouscell carcinoma (HNSCC). Failure to detect occult metastatic head and neck tumors on radical neck lymph nodedissection can affect the therapeutic measures taken. <br/><b>Objective</b>: The aim of this study was to investigate theLINE-1 and Alu methylation status and determine whether it can be applied for detection of occult metastatictumors in HNSCC cases. <br/><b>Methods</b>: We used the Combine Bisulfite Restriction Analysis (COBRA) techniqueto analyse LINE-1 and Alu methylation status. In addition to the methylation level, LINE-1 and Alu loci wereclassified based on the methylation statuses of two CpG dinucleotides in each allele as follows: hypermethylation(mCmC), hypomethylation (uCuC), and 2 forms of partial methylation (mCuC and uCmC). Sixty-one lymph nodeswere divided into 3 groups: 1) non-metastatic head and neck cancer (NM), 2) histologically negative for tumor cellsof cases with metastatic head and neck cancer (LN), and 3) histologically positive for tumor cells (LP). <br/><b>Results</b>:Alu methylation change was not significant. However, LINE-1 methylation of both LN and LP was altered, asdemonstrated by the lower LINE-1 methylation levels (p<0.001), higher percentage of mCuC (p<0.01), lowerpercentage of uCmC (p<0.001) and higher percentage of uCuC (p<0.001). Using receiver operating characteristic(ROC) curve analysis, %uCmC and %mCuC values revealed a high level of AUC at 0.806 and 0.716, respectively,in distinguishing LN from NM. <br/><b>Conclusion</b>: The LINE-1 methylation changes in LN have the same pattern asthat in LP. This epigenomic change may be due to the presence of occult metastatic tumor in LN cases.