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

A. A. Ponomaryova^{a, c}, E. Y. Rykova^b, N. V. Cherdyntseva^{a, d}, A. A. Bondar^b,
A. Y. Dobrodeev^a, A. A. Zavyalov^a, S. A. Tuzikov^a, L. O. Bryzgalov^e,
T. I. Merkulova^e, V. V. Vlassov^b, and P. P. Laktionov^b

^aFederal Government Budget Institute Tomsk Cancer Research Institute, Tomsk, Russia

^bFederal Government Budget Institute of Science Institute of Chemical Biology and Fundamental Medicine,
Russian Academy of Sciences, Siberian Brunch, Novosibirsk, Russia

^cFederal Government Autonomous Educational Institute of Higher Professional Education the National Research
Tomsk Polytechnic University, Tomsk, Russia

^dFederal Government Autonomous Educational Institute of Higher Education National Research
Tomsk State University, Tomsk, Russia

^eFederal Government Budget Institute of Science Institute of Cytology and Genetics,
Russian Academy of Sciences, Siberian Branch, Novosibirsk, Russia

e-mail: rykova@niboch.nsc.ru

Received October 20, 2014; in final form, December 15, 2014

Abstract—The main components of DNA aberrant methylation in malignant cells are promoter hypermethy-
lation 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 cir-
DNA and csb-cirDNA plasma from 21 untreated LC patients and 23 healthy donors. Concentrations of methy-
lated 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 LINE-1met 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 LINE-1met 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 differ-
ences 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.

Keywords: blood-circulating DNA, aberrant methylation, LINE-1 retrotransposons, diagnostics, lung cancer

DOI: 10.1134/S2079059716010111

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