Advances in Molecular OncologyAdvances in Molecular OncologyУспехи молекулярной онкологии2313-805X2413-3787Publishing House ABV Press67910.17650/2313-805X-2024-11-2-85-96RESEARCH ARTICLESЭКСПЕРИМЕНТАЛЬНЫЕ СТАТЬИLINE-1 hypomethylation and HIST1H4F hypermethylation as oncomarkers in liquid biopsy of colorectal cancerГипометилирование LINE-1 и гиперметилирование HIST1H4F как онкомаркеры в жидкостной биопсии колоректального ракаhttps://orcid.org/0000-0003-0614-8789KondratovaV. N.КондратоваВ. Н.
Russian Federation

115522; 24 Kashirskoe Shosse; Moscow

115522; Каширское шоссе, 24; Москва

https://orcid.org/0000-0002-0297-4963BotezatuI. V.БотезатуИ. В.
Russian Federation

115522; 24 Kashirskoe Shosse; Moscow

115522; Каширское шоссе, 24; Москва

https://orcid.org/0000-0002-7297-5240StroganovaA. M.СтрогановаА. М.
Russian Federation

115522; 24 Kashirskoe Shosse; Moscow

115522; Каширское шоссе, 24; Москва

https://orcid.org/0000-0003-3315-0817DrankoS. L.ДранкоС. Л.
Russian Federation

115522; 24 Kashirskoe Shosse; Moscow

115522; Каширское шоссе, 24; Москва

https://orcid.org/0000-0002-0190-5069LichtensteinA. V.ЛихтенштейнA. В.
Russian Federation

Anatoly Vladimirovitch Lichtenstein

115522; 24 Kashirskoe Shosse; Moscow

Анатолий Владимирович Лихтенштейн

115522; Каширское шоссе, 24; Москва

alicht@mail.ruN. N. Blokhin National Medical Research Center of Oncology, Ministry of Health of RussiaФГБУ «Национальный медицинский исследовательский центр онкологии им. Н. Н. Блохина» Минздрава России1505202411285962806202428062024Copyright ©; 2024, Kondratova V.N., Botezatu I.V., Stroganova A.M., Dranko S.L., Lichtenstein A.V.Copyright ©; 2024, Кондратова В.Н., Ботезату И.В., Строганова А.М., Дранко С.Л., Лихтенштейн A.В.2024Kondratova V.N., Botezatu I.V., Stroganova A.M., Dranko S.L., Lichtenstein A.V.Кондратова В.Н., Ботезату И.В., Строганова А.М., Дранко С.Л., Лихтенштейн A.В.https://creativecommons.org/licenses/by/4.0https://umo.abvpress.ru/jour/article/view/679

   Introduction. Local hypermethylation of gene promoters and global genome hypomethylation are well-known manifestations of aberrant methylation associated with carcinogenesis. We investigated this phenomenon as a possible diagnostic marker for liquid biopsy of colorectal cancer using the original quantitative DNA melting analysis with hybridiza-tion probes (qDMA-HP) method.

   Aim. To quantify the methylation of HIST1H4F promoter and LINE-1 transposon in circulating blood plasma DNA of colorectal cancer patients.

   Materials and methods. Bisulfite-treated DNA samples isolated from blood plasma of healthy donors and cancer patients were analyzed. HIST1H4F methylation was assessed by asymmetric polymerase chain reaction with hybridized probe and post-amplification melting of probe / amplicon hybrids. To test for repetitive and highly polymorphic LINE-1 sequences, asymmetric polymerase chain reaction with hybridized probe and SYBR Green intercalating dye was used, followed by melting of hybrids and analysis of multicomponent melt curves.

   Results. High diagnostic efficiency of LINE-1 and HIST1H4F methylation markers in liquid biopsy of colorectal cancer was demonstrated with the area under the ROC curve = 0.92, sensitivity – 100 %, specificity – 84 %. Cross validation supports this result. Hypermethylation of HIST1H4F and hypomethylation of LINE-1 are statistically significantly correlated (Spearman correlation coefficient r = 0.4; p = 0.01).

   Conclusion. The qDMA-HP is suitable for quantitative assessment of aberrant methylation of various clinically significant genes.

   Введение. Локальное гиперметилирование промоторов уникальных генов и глобальное гипометилирование генома – хорошо известные проявления ассоциированного с канцерогенезом аберрантного метилирования. Мы исследовали этот феномен в качестве возможного диагностического маркера для жидкостной биопсии колоректальногорака, используя оригинальный метод количественного анализа плавления гибридов ДНК-зонд (quantitative DNA melting analysis with hybridization probes, qDMA-HP).

   Цель исследования – количественная оценка метилирования гена HIST1H4F и транспозона LINE-1 в циркулирующей ДНК плазмы крови больных колоректальным раком.

   Материалы и методы. Проанализированы обработанные бисульфитом образцы ДНК, выделенные из плазмы крови здоровых доноров и онкологических больных. Метилирование HIST1H4F оценивали с помощью асимметричной полимеразной цепной реакции с гибридизуемым зондом и плавлением гибридов ампликон / зонд после завершения амплификации. Для тестирования многократно повторяющихся и характеризующихся выраженным полиморфизмомпоследовательностей LINE-1 использовали асимметричную полимеразную цепную реакцию с гибридизуемым зондом и интеркалирующим красителем SYBR Green, после чего плавили гибриды и анализировали многокомпонентные профили плавления.

   Результаты. Показана высокая диагностическая эффективность панели маркеров LINE-1 и HIST1H4F в жидкостной биопсии колоректального рака: площадь под ROC-кривой 0,92; чувствительность 100 %; специфичность 84 %. Перекрестная валидизация подтверждает этот результат. Гиперметилирование HIST1H4F и гипометилирование LINE-1статистически значимо коррелируют (коэффициент корреляции Спирмена r = 0,4; p = 0,01).

   Заключение. Метод qDMA-HP можно использовать для количественной оценки аберрантного метилирования клинически значимых генов.

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