Epigenetic Probes for Lung Cancer Monitoring: Line-1 Methylation Pattern in Blood-Circulating DNA

The main components of DNA aberrant methylation in malignant cells are promoter hypermethylation of specific genes and hypomethylation of the main part of DNA, in particular, the repeated sequences of retrotransposons. The hypomethylation of retrotransposons from the LINE-1 family was revealed in the lung cancer (LC) tissue. The composition of circulating DNA (cirDNA) from the plasma and blood cell surface bound circulating DNA (csb-cirDNA) was shown earlier to be altered in the blood of cancer patients due to the accumulation of tumor-specific aberrantly methylated DNA fragments, which are currently considered to be valuable cancer markers. The present study compares LINE-1 retrotransposon methylation patterns in the cirDNA and csb-cirDNA plasma from 21 untreated LC patients and 23 healthy donors. Concentrations of methylated LINE-1 region 1 copies (LINE-1met) were assessed by quantitative real-time methylation-specific PCR. In order to normalize the LINE-1 methylation level, the LINE-1 region 2 concentration was evaluated, which was independent of the methylation status (LINE-1Ind). The LINE1met concentration in csb-cirDNA tended to decrease (by a factor of 1.4) in blood from LC patients compared to healthy donors (Mann–Whitney test, P = 0.16). The LINE-1Ind (methylation-independent) concentration in csb-cirDNA was found to be lower by a factor of three in LC patients and by a factor of four in patients with adenocarcinoma than in healthy donors. Thus, along with the expected decrease in the LINE1met concentration in csb-cirDNA from LC patients’ blood, we recorded an unexpected statistically significant increase of the LINE-1 methylation index determined as (LINE-1met/LINE-1Ind) due to the profound decrease of the total LINE-1Ind number. There were no differences between the plasma cirDNA LINE-1 methylation indexes in the LC patients and healthy donors (Mann–Whitney test, P = 0.40). The data obtained agreed with our earlier results, which showed that csb-cirDNA was highly informative material for lung cancer diagnostics.