Advances in Molecular OncologyAdvances in Molecular OncologyУспехи молекулярной онкологии2313-805X2413-3787Publishing House ABV Press54610.17650/2313-805X-2023-10-2-108-116EXPERIMENTAL REPORTЭКСПЕРИМЕНТАЛЬНАЯ СТАТЬЯComparative assessment of the exosomal tumor-associated microRNA levels in blood plasma and ascitic fluid in ovarian cancer patientsСравнительная оценка уровней опухолеассоциированных микроРНК экзосом плазмы крови и асцитической жидкости пациентов с раком яичниковhttps://orcid.org/0000-0003-0465-3124DzhugashviliE. I.ДжугашвилиЕ. И.
Russian Federation

2 Pirogova St., Novosibirsk 630090

630090 Новосибирск, ул. Пирогова, 2

https://orcid.org/0000-0003-4595-4177YunusovaN. V.ЮнусоваН. В.
Russian Federation

5 Cooperative Lane, Tomsk 634009

634009 Томск, пер. Кооперативный, 5

YalovayaA. I.ЯловаяА. И.
Russian Federation

8 Akademika Lavrent’eva Prospekt, Novosibirsk 630090

630090 Новосибирск, проспект Академика Лаврентьева, 8

https://orcid.org/0000-0001-9853-223XGrigorievaA. E.ГригорьеваА. Е.
Russian Federation

8 Akademika Lavrent’eva Prospekt, Novosibirsk 630090

630090 Новосибирск, проспект Академика Лаврентьева, 8

https://orcid.org/0000-0002-7752-9346SeredaE. E.СередаЕ. Е.
Russian Federation

5 Cooperative Lane, Tomsk 634009

634009 Томск, пер. Кооперативный, 5

https://orcid.org/0000-0002-6854-8940KolomietsL. A.КоломиецЛ. А.
Russian Federation

5 Cooperative Lane, Tomsk 634009

634009 Томск, пер. Кооперативный, 5

https://orcid.org/0000-0001-7774-943XTamkovichS. N.ТамковичС. Н.
Russian Federation

Svetlana N. Tamkovich.

2 Pirogova St., Novosibirsk 630090; 8 Akademika Lavrent’eva Prospekt, Novosibirsk 630090

Тамкович Светлана Николаевна.

630090 Новосибирск, ул. Пирогова, 2; 630090 Новосибирск, проспект Академика Лаврентьева, 8

s.tamkovich@g.nsu.ruNovosibirsk State UniversityФГАОУ ВО «Новосибирский национальный исследовательский государственный университет»Scientific Research Institute of Oncology of the Tomsk National Medical Research Center of the Russian Academy of SciencesНаучно-исследовательский институт онкологии ФГБНУ «Томский национальный медицинский исследовательский центр Российской академии наук»Institute of Chemical Biology and Fundamental Medicine of the Siberian Branch of the Russian Academy of SciencesФГБУН «Институт химической биологии и фундаментальной медицины Сибирского отделения Российской академии наук»150520231021081161612202210072023Copyright ©; 2023, Dzhugashvili E.I., Yunusova N.V., Yalovaya A.I., Grigorieva A.E., Sereda E.E., Kolomiets L.A., Tamkovich S.N.Copyright ©; 2023, Джугашвили Е.И., Юнусова Н.В., Яловая А.И., Григорьева А.Е., Середа Е.Е., Коломиец Л.А., Тамкович С.Н.2023Dzhugashvili E.I., Yunusova N.V., Yalovaya A.I., Grigorieva A.E., Sereda E.E., Kolomiets L.A., Tamkovich S.N.Джугашвили Е.И., Юнусова Н.В., Яловая А.И., Григорьева А.Е., Середа Е.Е., Коломиец Л.А., Тамкович С.Н.https://creativecommons.org/licenses/by/4.0https://umo.abvpress.ru/jour/article/view/546

Introduction. Ovarian cancer (OC) is one of the malignant neoplasms of the female reproductive system with a high mortality rate. Currently used tumor markers of this pathology do not have high sensitivity and specificity. In this regard, promising areas of molecular oncology are the study of the mechanisms of carcinogenesis of OC and the search for new biomarkers of liquid biopsy for early non-invasive diagnosis of neoplasms. It is known that tumor cells actively secrete exosomes into the extracellular space, which include biologically active molecules involved in carcinogenesis and claiming to be diagnostic markers. It was previously shown that microRNA-24 (miR-24) and microRNA-101 (miR-101) are transported as part of exosomes in OC and are involved in the degradation of the extracellular matrix, stromal remodeling, angiogenesis, and cancer cell motility.

Aim. To evaluate the representation and diagnostic significance of miR-24 and miR-101 in plasma exosomes and ascitic fluid of OC patients.

Materials and methods. The study included blood and ascitic fluid samples from OC patients (n = 20) and blood samples from healthy women (n = 19). The exosomal nature of the vesicles was confirmed by transmission electron microscopy, nanotracing analysis, and flow cytometry. After isolation of exosomal RNA, the relative level of miRNA was determined using reverse transcription and real-time polymerase chain reaction.

Results. The highest concentration of exosomes was found in the ascitic fluid of OC patients, while the concentration of exosomes in the blood plasma of these patients was significantly higher than in healthy women. Relative levels of miR-24 and miR-101 in exosomes of blood plasma of healthy women were significantly higher than in exosomes of blood plasma and ascitic fluid of OC patients; at the same time, the levels of these miRNAs in exosomes of plasma and ascitic fluid of patients did not differ significantly.

Conclusion. The results obtained confirm the promise of exosomal miR-101 and miR-24 for the diagnosis of OC by liquid biopsy.

Введение. Рак яичников (РЯ) входит в число злокачественных новообразований женской репродуктивной системы с высокой летальностью. применяемые в настоящие время онкомаркеры данной патологии не обладают высокими чувствительностью и специфичностью. в связи с этим перспективными направлениями молекулярной онкологии являются исследование механизмов канцерогенеза РЯ и поиск новых биомаркеров жидкостной биопсии для ранней неинвазивной диагностики новообразований. известно, что опухолевые клетки активно секретируют во внеклеточное пространство экзосомы, в состав которых входят биологически активные молекулы, участвующие в канцерогенезе и претендующие на роль диагностических маркеров. Ранее было показано, что микроРНК-24 (miR-24) и микро-РНК-101 (miR-101) переносятся в составе экзосом при РЯ и участвуют в процессе деградации внеклеточного матрикса, ремоделировании стромы, ангиогенезе и подвижности раковых клеток.

Цель исследования – оценка представленности и диагностической значимости miR-24 и miR-101 в экзосомах плазмы и асцитической жидкости больных РЯ.

Материалы и методы. В исследование включены образцы крови и асцитической жидкости больных РЯ (n = 20) и образцы крови здоровых женщин (n = 19). экзосомальную природу везикул подтверждали с помощью трансмиссионной электронной микроскопии, трекового анализа и проточной цитофлуориметрии. после выделения экзосомальной РНК определяли относительный уровень микроРНК с использованием обратной транскрипции и полимеразной цепной реакции в реальном времени.

Результаты. Наибольшая концентрация экзосом выявлена в асцитической жидкости больных РЯ, при этом концентрация экзосом в плазме крови этих пациенток оказалась достоверно выше, чем у здоровых женщин. Относительные уровни miR-24 и miR-101 в экзосомах плазмы крови здоровых женщин были достоверно выше, чем в экзосомах плазмы крови и асцитической жидкости больных РЯ; при этом уровни этих микроРНК в экзосомах плазмы и асцитической жидкости пациентов достоверно не различались.

Заключение. Полученные результаты подтверждают перспективность экзосомальных miR-101 и miR-24 для диагностики РЯ методом жидкостной биопсии.

exosomesplasmaascitestumor-associated microRNAsmiR-24-3pmiR-101ovarian cancerэкзосомыплазмаасцитопухолеассоциированные микроРНКmiR-24-3pmiR-101рак яичниковThe work was carried out within the framework of the basic budget financing project of the Ministry of Education and Science of Russia No. 121030200173-6 “Diagnosis and therapy of oncological diseases”.Работа выполнена в рамках проекта базового бюджетного финансирования Минобрнауки России № 121030200173-6 «Диагностика и терапия онкологических заболеваний».1. Ding H., Zhang J., Zhang F. et al. Nanotechnological approaches for diagnosis and treatment of ovarian cancer: a review of recent trends. Drug Deliv 2022;29(1):3218–32. DOI: 10.1080/10717544.2022.2132032Ding H., Zhang J., Zhang F. et al. Nanotechnological approaches for diagnosis and treatment of ovarian cancer: a review of recent trends. Drug Deliv 2022;29(1):3218–32. DOI: 10.1080/10717544.2022.21320322. Bhardwaj B., Thankachan S., Venkatesh T. et al. Liquid biopsy in ovarian cancer. Clinica Chimica Acta 2020;510:28–34. DOI: 10.1016/j.cca.2020.06.047Bhardwaj B., Thankachan S., Venkatesh T. et al. Liquid biopsy in ovarian cancer. Clinica Chimica Acta 2020;510:28–34. DOI: 10.1016/j.cca.2020.06.0473. Guo L., Guo N. Exosomes: potent regulators of tumor malignancy and potential bio-tools in clinical application. Crit Rev Oncol Hematol 2015;95(3):346–58. DOI: 10.1016/j.critrevonc.2015.04.002Guo L., Guo N. Exosomes: potent regulators of tumor malignancy and potential bio-tools in clinical application. Crit Rev Oncol Hematol 2015;95(3):346–58. DOI: 10.1016/j.critrevonc.2015.04.0024. Tenchov R., Sasso J.M., Wang X. et al. Exosomes-Nature’s lipid nanoparticles, a rising star in drug delivery and diagnostics. ACS Nano 2022;16(11):17802–46. DOI: 10.1021/acsnano.2c08774Tenchov R., Sasso J.M., Wang X. et al. Exosomes-Nature’s lipid nanoparticles, a rising star in drug delivery and diagnostics. ACS Nano 2022;16(11):17802–46. DOI: 10.1021/acsnano.2c087745. Yunusova N., Kolegova E., Sereda E. et al. Plasma exosomes of patients with breast and ovarian tumors contain an inactive 20S proteasome. Molecules 2021;26(22):6965. DOI: 10.3390/molecules26226965Yunusova N., Kolegova E., Sereda E. et al. Plasma exosomes of patients with breast and ovarian tumors contain an inactive 20S proteasome. Molecules 2021;26(22):6965. DOI: 10.3390/molecules262269656. Tutanov O., Orlova E., Proskura K. et al. Proteomic analysis of blood exosomes from healthy females and breast cancer patients reveals an association between different exosomal bioactivity on non-tumorigenic epithelial cell and breast cancer cell migration in vitro. Biomolecules 2020;10(4):495. DOI: 10.3390/biom10040495Tutanov O., Orlova E., Proskura K. et al. Proteomic analysis of blood exosomes from healthy females and breast cancer patients reveals an association between different exosomal bioactivity on non-tumorigenic epithelial cell and breast cancer cell migration in vitro. Biomolecules 2020;10(4):495. DOI: 10.3390/biom100404957. Ye S.B., Li Z.L., Luo D.H. et al. Tumor-derived exosomes promote tumor progression and T-cell dysfunction through the regulation of enriched exosomal microRNAs in human nasopharyngeal carcinoma. Oncotarget 2014;5(14):5439–52. DOI: 10.18632/oncotarget.2118Ye S.B., Li Z.L., Luo D.H. et al. Tumor-derived exosomes promote tumor progression and T-cell dysfunction through the regulation of enriched exosomal microRNAs in human nasopharyngeal carcinoma. Oncotarget 2014;5(14):5439–52. DOI: 10.18632/oncotarget.21188. Xia Y., Hu X., Di K. et al. Combined detection of exosome concentration and tumor markers in gastric cancer. J Biomed Nanotechnol 2020;16(2):252–8. DOI: 10.1166/jbn.2020.2887Xia Y., Hu X., Di K. et al. Combined detection of exosome concentration and tumor markers in gastric cancer. J Biomed Nanotechnol 2020;16(2):252–8. DOI: 10.1166/jbn.2020.28879. Yunusova N., Patysheva M., Molchanov S. et al. Metalloproteinases at the surface of small extrcellular vesicles in advanced ovarian cancer: Relationships with ascites volume and peritoneal canceromatosis index. Clin Chim Acta 2019;494:116–22. DOI: 10.1016/j.cca.2019.03.1621Yunusova N., Patysheva M., Molchanov S. et al. Metalloproteinases at the surface of small extrcellular vesicles in advanced ovarian cancer: Relationships with ascites volume and peritoneal canceromatosis index. Clin Chim Acta 2019;494:116–22. DOI: 10.1016/j.cca.2019.03.162110. Joyce D.P., Kerin M.J., Dwyer R.M. Exosome-encapsulated microRNAs as circulating biomarkers for breast cancer. Int J Cancer 2016;139(7):1443–8. DOI: 10.1002/ijc.30179Joyce D.P., Kerin M.J., Dwyer R.M. Exosome-encapsulated microRNAs as circulating biomarkers for breast cancer. Int J Cancer 2016;139(7):1443–8. DOI: 10.1002/ijc.3017911. Kalra H., Drummen G.P., Mathivanan S. Focus on extracellular vesicles: introducing the next small big thing. Int J Mol Sci 2016;17(2):170. DOI: 10.3390/ijms17020170Kalra H., Drummen G.P., Mathivanan S. Focus on extracellular vesicles: introducing the next small big thing. Int J Mol Sci 2016;17(2):170. DOI: 10.3390/ijms1702017012. Yang D., Sun Y., Hu L. et al. Integrated analyses identify a master microRNA regulatory network for the mesenchymal subtype in serous ovarian cancer. Cancer Cell 2013;23(2):186–99. DOI: 10.1016/j.ccr.2012.12.020Yang D., Sun Y., Hu L. et al. Integrated analyses identify a master microRNA regulatory network for the mesenchymal subtype in serous ovarian cancer. Cancer Cell 2013;23(2):186–99. DOI: 10.1016/j.ccr.2012.12.02013. Braga E.A., Fridman M.V., Kushlinskii N.E. Molecular mechanisms of ovarian carcinoma metastasis: key genes and regulatory microRNAs. Biochemistry (Mosc) 2017;82(5):529–41. DOI: 10.1134/S0006297917050017Braga E.A., Fridman M.V., Kushlinskii N.E. Molecular mechanisms of ovarian carcinoma metastasis: key genes and regulatory microRNAs. Biochemistry (Mosc) 2017;82(5):529–41. DOI: 10.1134/S000629791705001714. Liu W., Wang S., Zhou S. et al. A systems biology approach to identify microRNAs contributing to cisplatin resistance in human ovarian cancer cells. Mol Biosyst 2017;13(11):2268–76. DOI: 10.1039/c7mb00362eLiu W., Wang S., Zhou S. et al. A systems biology approach to identify microRNAs contributing to cisplatin resistance in human ovarian cancer cells. Mol Biosyst 2017;13(11):2268–76. DOI: 10.1039/c7mb00362e15. Konoshenko M., Sagaradze G., Orlova E. et al. Total blood exosomes in breast cancer: potential role in crucial steps of tumorigenesis. Int J Mol Sci 2020;21(19):7341. DOI: 10.3390/ijms21197341Konoshenko M., Sagaradze G., Orlova E. et al. Total blood exosomes in breast cancer: potential role in crucial steps of tumorigenesis. Int J Mol Sci 2020;21(19):7341. DOI: 10.3390/ijms2119734116. Tutanov O., Shtam T., Grigor’eva A. et al. Blood plasma exosomes contain circulating DNA in their crown. Diagnostics (Basel) 2022;12(4):854. DOI: 10.3390/diagnostics12040854Tutanov O., Shtam T., Grigor’eva A. et al. Blood plasma exosomes contain circulating DNA in their crown. Diagnostics (Basel) 2022;12(4):854. DOI: 10.3390/diagnostics1204085417. Konoshenko M.Y., Bryzgunova O.E., Lekchnov E.A. et al. The influence of radical prostatectomy on the expression of cell-free MiRNA. Diagnostics (Basel) 2020;10(8):600. DOI: 10.3390/diagnostics10080600Konoshenko M.Y., Bryzgunova O.E., Lekchnov E.A. et al. The influence of radical prostatectomy on the expression of cell-free MiRNA. Diagnostics (Basel) 2020;10(8):600. DOI: 10.3390/diagnostics1008060018. Tamkovich S., Tutanov O., Efimenko A. et al. Blood circulating exosomes contain distinguishable fractions of free and cell-surface-associated vesicles. Curr Mol Med 2019;19(4):273–85. DOI: 10.2174/1566524019666190314120532Tamkovich S., Tutanov O., Efimenko A. et al. Blood circulating exosomes contain distinguishable fractions of free and cell-surface-associated vesicles. Curr Mol Med 2019;19(4):273–85. DOI: 10.2174/156652401966619031412053219. Zhang W., Ou X., Wu X. Proteomics profiling of plasma exosomes in epithelial ovarian cancer: a potential role in the coagulation cascade, diagnosis and prognosis. Int J Oncol 2019;54(5):1719–33. DOI: 10.3892/ijo.2019.4742Zhang W., Ou X., Wu X. Proteomics profiling of plasma exosomes in epithelial ovarian cancer: a potential role in the coagulation cascade, diagnosis and prognosis. Int J Oncol 2019;54(5):1719–33. DOI: 10.3892/ijo.2019.474220. Hagrass H.A., Sharaf S., Pasha H.F. et al. Circulating microRNAs – a new horizon in molecular diagnosis of breast cancer. Genes Cancer 2015;6(5–6):281–7. DOI: 10.18632/genesandcancer.66Hagrass H.A., Sharaf S., Pasha H.F. et al. Circulating microRNAs – a new horizon in molecular diagnosis of breast cancer. Genes Cancer 2015;6(5–6):281–7. DOI: 10.18632/genesandcancer.6621. Bryzgunova O.E., Zaripov M.M., Skvortsova T.E. et al. Comparative study of extracellular vesicles from the urine of healthy individuals and prostate cancer patients. PLoS One 2016;11(6):e0157566. DOI: 10.1371/journal.pone.0157566Bryzgunova O.E., Zaripov M.M., Skvortsova T.E. et al. Comparative study of extracellular vesicles from the urine of healthy individuals and prostate cancer patients. PLoS One 2016;11(6):e0157566. DOI: 10.1371/journal.pone.015756622. McDermott A.M., Kerin M.J., Miller N. Identification and validation of miRNAs as endogenous controls for RQ-PCR in blood specimens for breast cancer studies. PLoS One 2013;8(12):e83718. DOI: 10.1371/journal.pone.0083718McDermott A.M., Kerin M.J., Miller N. Identification and validation of miRNAs as endogenous controls for RQ-PCR in blood specimens for breast cancer studies. PLoS One 2013;8(12):e83718. DOI: 10.1371/journal.pone.008371823. Matsumoto Y., Kano M., Murakami K. et al. Tumor-derived exosomes influence the cell cycle and cell migration of human esophageal cancer cell lines. Cancer Sci 2020;111(12):4348–58. DOI: 10.1111/cas.14660Matsumoto Y., Kano M., Murakami K. et al. Tumor-derived exosomes influence the cell cycle and cell migration of human esophageal cancer cell lines. Cancer Sci 2020;111(12):4348–58. DOI: 10.1111/cas.1466024. Pathania A.S., Prathipati P., Challagundla K.B. New insights into exosome mediated tumor-immune escape: Clinical perspectives and therapeutic strategies. Biochim Biophys Acta Rev Cancer 2021;1876(2):188624. DOI: 10.1016/j.bbcan.2021.188624Pathania A.S., Prathipati P., Challagundla K.B. New insights into exosome mediated tumor-immune escape: Clinical perspectives and therapeutic strategies. Biochim Biophys Acta Rev Cancer 2021;1876(2):188624. DOI: 10.1016/j.bbcan.2021.18862425. Paskeh M.D.A., Entezari M., Mirzaei S. et al. Emerging role of exosomes in cancer progression and tumor microenvironment remodeling. J Hematol Oncol 2022;15(1):83. DOI: 10.1186/s13045-022-01305-4Paskeh M.D.A., Entezari M., Mirzaei S. et al. Emerging role of exosomes in cancer progression and tumor microenvironment remodeling. J Hematol Oncol 2022;15(1):83. DOI: 10.1186/s13045-022-01305-426. Sahebi R., Langari H., Fathinezhad Z. et al. Exosomes: new insights into cancer mechanisms. J Cell Biochem 2020;121(1):7–16. DOI: 10.1002/jcb.29120Sahebi R., Langari H., Fathinezhad Z. et al. Exosomes: new insights into cancer mechanisms. J Cell Biochem 2020;121(1):7–16. DOI: 10.1002/jcb.2912027. Yi Y., Wu M., Zeng H. et al. Tumor-derived exosomal non-coding rnas: the emerging mechanisms and potential clinical applications in breast cancer. Front Oncol 2021;11:738945. DOI: 10.3389/fonc.2021.738945Yi Y., Wu M., Zeng H. et al. Tumor-derived exosomal non-coding rnas: the emerging mechanisms and potential clinical applications in breast cancer. Front Oncol 2021;11:738945. DOI: 10.3389/fonc.2021.73894528. Zheng H.B., Zheng X.G., Liu B.P. miRNA-101 inhibits ovarian cancer cells proliferation and invasion by down-regulating expression of SOCS-2. Int J Clin Exp Med 2015;8(11):20263–70.Zheng H.B., Zheng X.G., Liu B.P. miRNA-101 inhibits ovarian cancer cells proliferation and invasion by down-regulating expression of SOCS-2. Int J Clin Exp Med 2015;8(11):20263–70.29. Liang H., Yu T., Han Y. et al. LncRNA PTAR promotes EMT and invasion-metastasis in serous ovarian cancer by competitively binding miR-101-3p to regulate ZEB1 expression. Mol Cancer 2018;17(1):119. DOI: 10.1186/s12943-018-0870-5Liang H., Yu T., Han Y. et al. LncRNA PTAR promotes EMT and invasion-metastasis in serous ovarian cancer by competitively binding miR-101-3p to regulate ZEB1 expression. Mol Cancer 2018;17(1):119. DOI: 10.1186/s12943-018-0870-530. Xu Y., Xu L., Zheng J. et al. MiR-101 inhibits ovarian carcinogenesis by repressing the expression of brain-derived neurotrophic factor. FEBS Open Bio 2017;7(9):1258–66. DOI: 10.1002/2211-5463.12257Xu Y., Xu L., Zheng J. et al. MiR-101 inhibits ovarian carcinogenesis by repressing the expression of brain-derived neurotrophic factor. FEBS Open Bio 2017;7(9):1258–66. DOI: 10.1002/2211-5463.1225731. Liu L., Guo J., Yu L. et al. miR-101 regulates expression of EZH2 and contributes to progression of and cisplatin resistance in epithelial ovarian cancer. Tumour Biol 2014;35(12):12619–26. DOI: 10.1007/s13277-014-2585-6Liu L., Guo J., Yu L. et al. miR-101 regulates expression of EZH2 and contributes to progression of and cisplatin resistance in epithelial ovarian cancer. Tumour Biol 2014;35(12):12619–26. DOI: 10.1007/s13277-014-2585-632. Wang H., Chen C., Ding K. et al. MiR-24-3p as a prognostic indicator for multiple cancers: from a meta-analysis view. Biosci Rep 2020;40(12):BSR20202938. DOI: 10.1042/BSR20202938Wang H., Chen C., Ding K. et al. MiR-24-3p as a prognostic indicator for multiple cancers: from a meta-analysis view. Biosci Rep 2020;40(12):BSR20202938. DOI: 10.1042/BSR2020293833. Yuan D., Cui X., Wang Y. et al. Enrichment analysis identifies functional microRNA-disease associations in humans. PLoS One 2015;10(8):e0136285. DOI: 10.1371/journal.pone.0136285Yuan D., Cui X., Wang Y. et al. Enrichment analysis identifies functional microRNA-disease associations in humans. PLoS One 2015;10(8):e0136285. DOI: 10.1371/journal.pone.013628534. Lin H., Xu X., Chen K. et al. LncRNA CASC15, MiR-23b cluster and SMAD3 form a novel positive feedback loop to promote epithelial-mesenchymal transition and metastasis in ovarian cancer. Int J Biol Sci 2022;18(5):1989–2002. DOI: 10.7150/ijbs.67486Lin H., Xu X., Chen K. et al. LncRNA CASC15, MiR-23b cluster and SMAD3 form a novel positive feedback loop to promote epithelial-mesenchymal transition and metastasis in ovarian cancer. Int J Biol Sci 2022;18(5):1989–2002. DOI: 10.7150/ijbs.67486