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BMC Cancer
Published in: BMC Cancer 1/2021

Open Access 01-12-2021 | Research article

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

Authors: Silvia R. Vitale, Jean A. Helmijr, Marjolein Gerritsen, Hicret Coban, Lisanne F. van Dessel, Nick Beije, Michelle van der Vlugt-Daane, Paolo Vigneri, Anieta M. Sieuwerts, Natasja Dits, Martin E. van Royen, Guido Jenster, Stefan Sleijfer, Martijn Lolkema, John W. M. Martens, Maurice P. H. M. Jansen

Published in: BMC Cancer | Issue 1/2021

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Abstract

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.
Appendix
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Metadata
Title
Detection of tumor-derived extracellular vesicles in plasma from patients with solid cancer
Authors
Silvia R. Vitale
Jean A. Helmijr
Marjolein Gerritsen
Hicret Coban
Lisanne F. van Dessel
Nick Beije
Michelle van der Vlugt-Daane
Paolo Vigneri
Anieta M. Sieuwerts
Natasja Dits
Martin E. van Royen
Guido Jenster
Stefan Sleijfer
Martijn Lolkema
John W. M. Martens
Maurice P. H. M. Jansen
Publication date
01-12-2021
Publisher
BioMed Central
Published in
BMC Cancer / Issue 1/2021
Electronic ISSN: 1471-2407
DOI
https://doi.org/10.1186/s12885-021-08007-z

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