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Detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer

BACKGROUND: Extracellular vesicles (EVs) are actively secreted by cells into body fluids and contain nucleic acids of the cells they originate from. The goal of this study was to detect circulating tumor-derived EVs (ctEVs) by mutant mRNA transcripts (EV-RNA) in plasma of patients with solid cancers...

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Autores principales: Vitale, Silvia R., Helmijr, Jean A., Gerritsen, Marjolein, Coban, Hicret, van Dessel, Lisanne F., Beije, Nick, van der Vlugt-Daane, Michelle, Vigneri, Paolo, Sieuwerts, Anieta M., Dits, Natasja, van Royen, Martin E., Jenster, Guido, Sleijfer, Stefan, Lolkema, Martijn, Martens, John W. M., Jansen, Maurice P. H. M.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7992353/
https://www.ncbi.nlm.nih.gov/pubmed/33761899
http://dx.doi.org/10.1186/s12885-021-08007-z
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author Vitale, Silvia R.
Helmijr, Jean A.
Gerritsen, Marjolein
Coban, Hicret
van Dessel, Lisanne F.
Beije, Nick
van der Vlugt-Daane, Michelle
Vigneri, Paolo
Sieuwerts, Anieta M.
Dits, Natasja
van Royen, Martin E.
Jenster, Guido
Sleijfer, Stefan
Lolkema, Martijn
Martens, John W. M.
Jansen, Maurice P. H. M.
author_facet Vitale, Silvia R.
Helmijr, Jean A.
Gerritsen, Marjolein
Coban, Hicret
van Dessel, Lisanne F.
Beije, Nick
van der Vlugt-Daane, Michelle
Vigneri, Paolo
Sieuwerts, Anieta M.
Dits, Natasja
van Royen, Martin E.
Jenster, Guido
Sleijfer, Stefan
Lolkema, Martijn
Martens, John W. M.
Jansen, Maurice P. H. M.
author_sort Vitale, Silvia R.
collection PubMed
description BACKGROUND: Extracellular vesicles (EVs) are actively secreted by cells into body fluids and contain nucleic acids of the cells they originate from. The goal of this study was to detect circulating tumor-derived EVs (ctEVs) by mutant mRNA transcripts (EV-RNA) in plasma of patients with solid cancers and compare the occurrence of ctEVs with circulating tumor DNA (ctDNA) in cell-free DNA (cfDNA). METHODS: For this purpose, blood from 20 patients and 15 healthy blood donors (HBDs) was collected in different preservation tubes (EDTA, BCT, CellSave) and processed into plasma within 24 h from venipuncture. EVs were isolated with the ExoEasy protocol from this plasma and from conditioned medium of 6 cancer cell lines and characterized according to MISEV2018-guidelines. RNA from EVs was isolated with the ExoRNeasy protocol and evaluated for transcript expression levels of 96 genes by RT-qPCR and genotyped by digital PCR. RESULTS: Our workflow applied on cell lines revealed a high concordance between cellular mRNA and EV-RNA in expression levels as well as variant allele frequencies for PIK3CA, KRAS and BRAF. Plasma CD9-positive EV and GAPDH EV-RNA levels were significantly different between the preservation tubes. The workflow detected only ctEVs with mutant transcripts in plasma of patients with high amounts (> 20%) of circulating tumor DNA (ctDNA). Expression profiling showed that the EVs from patients resemble healthy donors more than tumor cell lines supporting that most EVs are derived from healthy tissue. CONCLUSIONS: We provide a workflow for ctEV detection by spin column-based generic isolation of EVs and PCR-based measurement of gene expression and mutant transcripts in EV-RNA derived from cancer patients’ blood plasma. This workflow, however, detected tumor-specific mutations in blood less often in EV-RNA than in cfDNA. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12885-021-08007-z.
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spelling pubmed-79923532021-03-25 Detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer Vitale, Silvia R. Helmijr, Jean A. Gerritsen, Marjolein Coban, Hicret van Dessel, Lisanne F. Beije, Nick van der Vlugt-Daane, Michelle Vigneri, Paolo Sieuwerts, Anieta M. Dits, Natasja van Royen, Martin E. Jenster, Guido Sleijfer, Stefan Lolkema, Martijn Martens, John W. M. Jansen, Maurice P. H. M. BMC Cancer Research Article BACKGROUND: Extracellular vesicles (EVs) are actively secreted by cells into body fluids and contain nucleic acids of the cells they originate from. The goal of this study was to detect circulating tumor-derived EVs (ctEVs) by mutant mRNA transcripts (EV-RNA) in plasma of patients with solid cancers and compare the occurrence of ctEVs with circulating tumor DNA (ctDNA) in cell-free DNA (cfDNA). METHODS: For this purpose, blood from 20 patients and 15 healthy blood donors (HBDs) was collected in different preservation tubes (EDTA, BCT, CellSave) and processed into plasma within 24 h from venipuncture. EVs were isolated with the ExoEasy protocol from this plasma and from conditioned medium of 6 cancer cell lines and characterized according to MISEV2018-guidelines. RNA from EVs was isolated with the ExoRNeasy protocol and evaluated for transcript expression levels of 96 genes by RT-qPCR and genotyped by digital PCR. RESULTS: Our workflow applied on cell lines revealed a high concordance between cellular mRNA and EV-RNA in expression levels as well as variant allele frequencies for PIK3CA, KRAS and BRAF. Plasma CD9-positive EV and GAPDH EV-RNA levels were significantly different between the preservation tubes. The workflow detected only ctEVs with mutant transcripts in plasma of patients with high amounts (> 20%) of circulating tumor DNA (ctDNA). Expression profiling showed that the EVs from patients resemble healthy donors more than tumor cell lines supporting that most EVs are derived from healthy tissue. CONCLUSIONS: We provide a workflow for ctEV detection by spin column-based generic isolation of EVs and PCR-based measurement of gene expression and mutant transcripts in EV-RNA derived from cancer patients’ blood plasma. This workflow, however, detected tumor-specific mutations in blood less often in EV-RNA than in cfDNA. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12885-021-08007-z. BioMed Central 2021-03-24 /pmc/articles/PMC7992353/ /pubmed/33761899 http://dx.doi.org/10.1186/s12885-021-08007-z Text en © The Author(s) 2021 Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
spellingShingle Research Article
Vitale, Silvia R.
Helmijr, Jean A.
Gerritsen, Marjolein
Coban, Hicret
van Dessel, Lisanne F.
Beije, Nick
van der Vlugt-Daane, Michelle
Vigneri, Paolo
Sieuwerts, Anieta M.
Dits, Natasja
van Royen, Martin E.
Jenster, Guido
Sleijfer, Stefan
Lolkema, Martijn
Martens, John W. M.
Jansen, Maurice P. H. M.
Detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer
title Detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer
title_full Detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer
title_fullStr Detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer
title_full_unstemmed Detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer
title_short Detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer
title_sort detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7992353/
https://www.ncbi.nlm.nih.gov/pubmed/33761899
http://dx.doi.org/10.1186/s12885-021-08007-z
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