Abstract:
Malignant cell transformation is accompanied by two processes of DNA methylation changes: promoter hypermethylation of specific genes and hypomethylation of retrotransposons. The composition of circulating DNA (cirDNA) from plasma and cell-surface-bound circulating DNA (csb- cirDNA) was shown earlier to be altered in the blood of cancer patients due to accumulation of tumor- specific aberrantly methylated DNA fragments, which are currently considered valuable cancer markers. The present study compares LINE-1 retrotransposon methylation patterns in plasma cirDNA and csb- cirDNA from 21 untreated lung cancer patients (LC) and 23 healthy donors. Concentrations of methylated LINE-1 region 1 copies (LINE-1met) were assayed by 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 in comparison to healthy donors (Mann- Whitney test, P=0.16). The LINE-1Ind concentration in csb-cirDNA (methylation-independent) was found to be threefold lower in LC patients and fourfold lower in patients with adenocarcinoma than in healthy donors. That is why, along with the expected decrease in LINE-1met concentration in csb-cirDNA, we recorded an unexpected statistically significant increase of the LINE-1 methylation index determined as (LINE-1met/LINE-1Ind) due to the profound LINE-1Ind decrease. Plasma cirDNA demonstrated no difference in the LINE-1 methylation index (LINE-1met/LINE-1Ind) between LC patients and healthy donors (Mann-Whitney test, P = 0.40). The data obtained agree with our earlier results, which showed that csb-cirDNA was a highly informative material for lung cancer diagnostics.
About The Authors:
A. A. Ponomaryova. Tomsk Cancer Research Institute, Tomsk, Russia The National Research Tomsk Polytechnic University, Tomsk, Russia, Russian Federation
N. V. Cherdyntseva. Tomsk Cancer Research Institute, Tomsk, Russia National Research Tomsk State University, Tomsk, Russia, Russian Federation
A. Y. Dobrodeev. Tomsk Cancer Research Institute, Tomsk, Russia, Russian Federation
S. A. Tuzikov. Tomsk Cancer Research Institute, Tomsk, Russia, Russian Federation
T. I. Merkulova. Institute of Cytology and Genetics SB RAS, Novosibirsk, Russian Federation
P. P. Laktionov. Institute of Chemical Biology and Fundamental Medicine SB RAS, Novosibirsk, Russia, Russian Federation
References:
1. Anker P., Lyautey J., Lederrey C, Stroun M. Circulating nucleic acids in plasma or serum. Clin. Chim. Acta. 2001;313:143-146.
2. Aparicio A., North B., Barske L., Wang X., Bollati V., Weisenberger D., Yoo C., Tannir N., Horne E., Groshen S., Jones P., Yang A., Issa J.P. LINE-1 methylation in plasma DNA as a biomarker of activity of DNA methylation inhibitors in patients with solid tumors. Epigenetics. 2009;4:176-184.
3. Beck J., Urnovitz H.B., Mitchell W.M., Schütz E. Next generation sequencing of serum circulating nucleic reveals differences to healthy and nonmalignant controls acids from patients with invasive ductal breast cancer. Mol. Cancer Res. 2010;8(3):335-342. DOI: 10.1158/1541-7786.MCR-09-0314
4. Beck J., Urnovitz H.B., Riggert J., Clerici M., Schütz E. Profile of the circulating DNA in apparently healthy individuals. Clin. Chem. 2009;55(4):730-738. DOI: 10.1373/clinchem.2008.113597
5. Bourc’his D., Bestor T.H. Meiotic catastrophe and retrotransposon reactivation in male germ cells lacking Dnmt3L. Nature. 2004;431(7004):96-99.
6. Bryzgunova O., Laktionov P., Skvortsova T., Bondar A., Morozkin E., Lebedeva A., Krause H., Miller K., Vlassov V. Efficacy of bisulfite modification and recovery of human of genomic and circulating DNA using commercial kits. Eur. J. Mol. Biol. 2013;1(1):1-8. DOI: 10.11648/j.ejmb.20130101.11
7. Carethers J.M. Proteomics, genomics, and molecular biology in the personalized treatment of colorectal cancer. J. Gastrointest Surg. 2012;16(9):1648-1650. DOI: 10.1007/s11605-012-1942-2
8. Dunican D.S., Cruickshanks H.A., Suzuki M., Semple C.A., Davey T., Arceci R.J., Greally J., Adams I.R., Meehan R.R. Lsh regulates LTR retrotransposon repression independently of Dnmt3b function. Genome Biol. 2013;14(12):R146. DOI: 10.1186/gb-2013-1412-r146
9. Ehrlich M. DNA hypomethylation, cancer, the immunodeficiency, centromeric region instability, facial anomalies syndrome and chromosomal rearrangements. J. Nutr. 2002;132. Suppl. 8:2424S2429S.
10. Esteller M. Epigenetics in cancer. N. Engl. J. Med. 2008;358(11): 1148-1159.
11. Hoshimoto S., Kuo C.T., Chong K.K., Takeshima T.L., Takei Y., Li M.W., Huang S.K., Sim M.S., Morton D.L., Hoon D.S. AIM1 and LINE-1 epigenetic aberrations in tumor and serum relate to melanoma progression and disease outcome. J. Invest. Dermatol. 2012;132(6): 1689-1697. DOI: 10.1038/jid.2012.36
12. Kristensen L.S., Hansen L.L. PCR-based methods for detecting single-locus DNA methylation biomarkers in cancer diagnostics, prognostics, and response to treatment. Clin. Chem. 2009;55(8): 1471-1483. DOI:10.1373/clinchem.2008.121962
13. Lander E.S., Linton L.M., Birren B., Lander E.S., Linton L.M., Birren B., Nusbaum C., Zody M.C., Baldwin J., Devon K., Dewar K., Doyle M., FitzHugh W., Funke R., Gage D., Harris K., Heaford A., Howland J., Kann L., Lehoczky J., LeVine R., McEwan P., McKernan K., Meldrim J., Mesirov J.P., Miranda C., Morris W., Naylor J., Raymond C., Rosetti M., Santos R., SheridanA., Sougnez C., StangeThomann N., Stojanovic N., Subramanian A., Wyman D., Rogers J., Sulston J., Ainscough R., Beck S., Bentley D., Burton J., Clee C., Carter N., Coulson A., Deadman R., Deloukas P., Dunham A., A.A. Ponomaryova, E.Y. Rykova, N.V. Cherdyntseva, … 2015Dunham I., Durbin R., French L., Grafham D., Gregory S., Hubbard T., Humphray S., Hunt A., Jones M., Lloyd C., McMurray A., Matthews L., Mercer S., Milne S., Mullikin J.C., Mungall A., Plumb R., Ross M., Shownkeen R., Sims S., Waterston R.H., Wilson R.K., Hillier L.W., McPherson J.D., Marra M.A., Mardis E.R., Fulton L.A., Chinwalla A.T., Pepin K.H., Gish W.R., Chissoe S.L., Wendl M.C., Delehaunty K.D., Miner T.L., Delehaunty A., Kramer J.B., Cook L.L., Fulton RS, Johnson D.L., Minx P.J., Clifton S.W., Hawkins T., Branscomb E., Predki P., Richardson P., Wenning S., Slezak T., Doggett N., Cheng J.F., Olsen A., Lucas S., Elkin C., Uberbacher E., Frazier M., Gibbs R.A., Muzny D.M., Scherer S.E., Bouck J.B., Sodergren E.J., Worley K.C., Rives C.M., Gorrell J.H., Metzker M.L., Naylor S.L., Kucherlapati R.S., Nelson D.L., Weinstock G.M., Sakaki Y., Fujiyama A., Hattori M., Yada T., Toyoda A., Itoh T., Kawagoe C., Watanabe H., Totoki Y., Taylor T., Weissenbach J., Heilig R., Saurin W., Artiguenave F., Brottier P., Bruls T., Pelletier E., Robert C., Wincker P., Smith D.R., Doucette-Stamm L., Rubenfield M., Weinstock K., Lee H.M., Dubois J., Rosenthal A., Platzer M., Nyakatura G., Taudien S., Rump A., YangH.,YuJ.,WangJ.,HuangG.,GuJ.,HoodL.,RowenL.,MadanA., Qin S, Davis R.W., Federspiel N.A., Abola A.P., Proctor M.J., Myers R.M., Schmutz J., Dickson M., Grimwood J., Cox D.R., Olson M.V., Kaul R., Raymond C., Shimizu N., Kawasaki K., Minoshima S., Evans G.A., Athanasiou M., Schultz R., Roe B.A., Chen F., Pan H., Ramser J., Lehrach H., Reinhardt R., McCombie W.R., de la Bastide M., Dedhia N., Blöcker H., Hornischer K., Nordsiek G., Agarwala R., Aravind L., Bailey J.A., Bateman A., Batzoglou S., Birney E., Bork P., Brown D.G., Burge C.B., Cerutti L., Chen H.C., Church D., Clamp M., Copley R.R., Doerks T., Eddy S.R., Eichler E.E., Furey T.S., Galagan J., Gilbert J.G, Harmon C., Hayashizaki Y., Haussler D., Hermjakob H., Hokamp K., Jang W., Johnson L.S., Jones T.A., Kasif S., Kaspryzk A., Kennedy S., Kent W.J., Kitts P., Koonin E.V., Korf I., Kulp D., Lancet D., Lowe T.M., McLysaght A., Mikkelsen T, Moran J.V., Mulder N., Pollara VJ, Ponting C.P., Schuler G., Schultz J., Slater G., Smit A.F., Stupka E., Szustakowski J., Thierry-Mieg D., ThierryMieg J., Wagner L., Wallis J., Wheeler R., Williams A., Wolf Y.I., Wolfe K.H., Yang SP, Yeh R.F., Collins F, Guyer M.S., Peterson J., Felsenfeld A., Wetterstrand K.A., Patrinos A., Morgan M.J., de Jong P., Catanese J.J., Osoegawa K., Shizuya H., Choi S., Chen Y.J. Initial sequencing and analysis of the human genome. Nature. 2001;409(6822):860-921.
14. Morozkin E.S., Loseva E.M., Morozov I.V., Kurilshikov A.M., Bondar A.A., Rykova E.Y., Rubtsov N.B., Vlassov V.V., Laktionov P.P. A comparative study of cell-free apoptotic and genomic DNA using FISH and massive parallel sequencing. Expert Opin. Biol. Ther. 2012;12(1):S141-S153. DOI:10.1517/14712598.2012.670631
15. Peters D.L., Pretorius P.J. Origin, translocation and destination of extracellular occurring DNA-a new paradigm in genetic behaviour. Clin. Chim. Acta. 2011;412(11/12):806-811. DOI: 10.1016/j.cca.2011.01.026
16. Ponomaryova A., Rykova E., Cherdyntseva N., Skvortsova T.E., Dobrodeev A.Y., Zav’yalov A.A., Bryzgalov L.O., Tuzikov S.A., Vlassov V.V., Laktionov P.P. Potentialities of aberrantly methylated circulating DNA for diagnostics and post-treatment follow-up of lung cancer patients. Lung Cancer. 2013;81(3):397-403. DOI: 10.1016/j.lungcan.2013.05.016
17. Radpour R., Barekati Z., Kohler C., Lv Q., Bürki N., Diesch C., Bitzer J., Zheng H., Schmid S., Zhong X.Y. Hypermethylation of tumor suppressor genes involved in critical regulatory pathways for developing a blood-based test in breast cancer. PLoS ONE. 2011;6(1):e16080. DOI: 10.1371/journal.pone.0016080
18. Ramzy I.I., Omran D.A., Hamad O., Shaker O., Abboud A. Evaluation of serum LINE-1 hypomethylation as a prognostic marker for hepatocellular carcinoma. Arab J. Gastroenterol. 2011;12(3):139142. DOI:10.1016/j.ajg.2011.07.002
19. Rykova E.Y., Morozkin E.S., Ponomaryova A.A., Loseva E.M., Zaporozhchenko I.A., Cherdyntseva N.V., Vlassov V.V., Laktionov P.P. Cell-free and cell-bound circulating nucleic acid complexes: mechanisms of generation, concentration and content. Expert Opin. Biol. Th. 2012;12. Suppl. 1:S141-S153. DOI:10.1517/14712598.2012.673577
20. Saito K., Kawakami K., Matsumoto I., Oda M., Watanabe G., Minamoto T. Long interspersed nuclear element 1 hypomethylation is a marker of poor prognosis in stage IA non-small cell lung cancer. Clin. Cancer Res. 2010;16(8):2418-2426. DOI: 10.1158/10780432.CCR-09-2819
21. Schwarzenbach H., Nishida N., Calin G.A., Pantel K. Clinical relevance of circulating cell-free microRNAs in cancer. Nature Rev. Clin. Oncol. 2014;11(3):145-156. DOI: 10.1038/nrclinonc.2014.5
22. Skvortsova T.E., Bryzgunova O.E., Lebedeva A.O., Mak V.V., Vlassov V.V., Laktionov P.P. Methylated cell-free DNA in vitro and in vivo. Circulating nucleic acids in plasma and serum (Ed. P.B .Gahan). United Kingdom: Springer, 2011:85-194. DOI: 10.1007/97890-481-9382-0_25
23. Suzuki M., Shiraishi K., Eguchi A., Ikeda K., Mori T., Yoshimoto K., Ohba Y., Yamada T., Ito T., Baba Y., Baba H. Aberrant methylation of LINE-1, SLIT2, MAL and IGFBP7 in non-small cell lung cancer. Oncol. Rep. 2013;29(4):1308-1314. DOI: 10.3892/or.2013.2266
24. Van der Vaart M., Semenov D.V., Kuligina E.V., Richter V.A., Pretorius P.J. Characterisation of circulating DNA by parallel tagged sequencing on the 454 platform. Clin. Chim. Acta. 2009;409(1/2): 21-27. DOI:10.1016/j.cca.2009.08.011
