Defining Optimal Conditions for Tumor Extracellular Vesicle DNA Extraction for Mutation Profiling

(1) Background: Extracellular vesicles (EVs) have emerged as crucial players in the communication between cells in both physiological and pathological scenarios. The functions of EVs are strongly determined by their molecular content, which includes all bioactive molecules, such as proteins, lipids,...

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Published in:Cancers 2022-07, Vol.14 (13), p.3258
Main Authors: Elzanowska, Julia, Berrocal, Laura, García-Peláez, Beatriz, Vives-Usano, Marta, Sebo, Beatriz Passos, Maia, Joana, Batista, Silvia, Teppo, Jaakko, Varjosalo, Markku, Moraes, Maria Carolina Strano, Molina-Vila, Miguel Ángel, Costa-Silva, Bruno
Format: Article
Language:English
Subjects:
Antibodies
Biomarkers
Biopsy
Cell culture
Cell interactions
Colorectal cancer
DNA fingerprinting
Genomes
Infrared imaging systems
Lipids
Mutation
Next-generation sequencing
Pancreatic cancer
Plasma
Proteins
Quality control
Standardization
Sucrose
Tumors
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Summary:(1) Background: Extracellular vesicles (EVs) have emerged as crucial players in the communication between cells in both physiological and pathological scenarios. The functions of EVs are strongly determined by their molecular content, which includes all bioactive molecules, such as proteins, lipids, RNA, and, as more recently described, double-stranded DNA. It has been shown that in oncological settings DNA associated with EVs (EV-DNA) is representative of the genome of parental cells and that it reflects the mutational status of the tumor, gaining much attention as a promising source of biomarker mutant DNA. However, one of the challenges in studies of EV-DNA is the lack of standardization of protocols for the DNA extraction from EVs, as well as ways to assess quality control, which hinders its future implementation in clinics. (2) Methods: We performed a comprehensive comparison of commonly used approaches for EV-DNA extraction by assessing DNA quantity, quality, and suitability for downstream analyses. (3) Results: We here established strategic points to consider for EV-DNA preparation for mutational analyses, including qPCR and NGS. (4) Conclusions: We put in place a workflow that can be applied for the detection of clinically relevant mutations in the EV-DNA of cancer patients.
ISSN:2072-6694
2072-6694
DOI:10.3390/cancers14133258