Abstract
Cancer cells release extracellular vesicles known as extracellular vesicles (EVs), containing tumor-derived DNA, RNA and proteins within their cargo, into the circulation. Circulating tumor-derived extracellular vesicles (TEV) can be used in the context of serial “liquid biopsies” for early detection of cancer, for monitoring disease burden in patients, and for assessing recurrence in the post-resection setting. Nonetheless, isolating sufficient TEV by ultracentrifugation-based approaches, in order to enable molecular assessment of EVs cargo, can be an arduous, time-consuming process and is inconsistent in the context of yield and purity among institutions. Herein, we describe a microfluidic platform, which we have named MITEV (Microfluidic Isolation of Tumor-derived Extracellular Vesicles) for the rapid isolation of TEV from the plasma of pancreatic cancer patients. The device, which has ~100,000 pillars placed in a zigzag pattern and is coated with antibodies against generic EV surface proteins (anti-CD63, -CD9, and -CD81 antibodies) or the TEV specific anti-Epithelial Cell Adhesion Molecule (EpCAM) antibody, is capable of high-throughput EVs isolation and yields sufficient DNA (total of ~2–14 ng from 2-ml plasma) for downstream genomic analysis. Using two independent quantitative platforms, droplet digital polymerase chain reaction (ddPCR) and molecular barcoding using nanoString nCounter® technology, we can reliably identify KRAS mutations within isolated TEV of treatment-naïve metastatic pancreatic cancer patients. Our study suggests that the MITEV device can be used for point-of-care applications, such as in the context of monitoring residual or recurrent tumor presence in pancreatic cancer patients undergoing therapy.
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Abbreviations
- TEV:
-
Tumor associated Extracellular Vesicles
- NTEV:
-
Non-Tumor associated Extracellular Vesicles
- MITEV:
-
Microfluidic Isolation of Tumor-derived EVs
- PDAC:
-
Pancreatic Ductal Adenocarcinoma
- EV:
-
Extracellular Vesicles
- EpCAM:
-
Epithelial Cell Adhesion Molecule
- ddPCR:
-
Droplet digital Polymerase Chain Reaction
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Acknowledgements
Scanning electron microscopy was performed at the High-Resolution Electron Microscopy Facility, while nanoString analysis was performed at the Sequencing and Microarray facility (SMF) in MD Anderson Cancer Center (Houston, TX), both of which are supported by the NCI Cancer Center Support Grant (P30CA016672). N.K. was supported by the CPRIT Research Training Program (RP170067).
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A.M. discloses receiving royalties from Hangzhou Guangkeande (Cosmos) Biotechnology Company LTD for being a co-inventor on a license related to pancreatic cancer early detection, and this financial relationship is managed by the MD Anderson Conflict of Interest Committee. There are no other conflicts of interests to declare.
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Kamyabi, N., Abbasgholizadeh, R., Maitra, A. et al. Isolation and mutational assessment of pancreatic cancer extracellular vesicles using a microfluidic platform. Biomed Microdevices 22, 23 (2020). https://doi.org/10.1007/s10544-020-00483-7
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DOI: https://doi.org/10.1007/s10544-020-00483-7