ac9b03064_si_001.pdf (1005.28 kB)
Download file

Isolation and Retrieval of Extracellular Vesicles for Liquid Biopsy of Malignant Ground-Glass Opacity

Download (1005.28 kB)
journal contribution
posted on 15.10.2019, 13:33 by Wenjun Mao, Yi Wen, Haozhi Lei, Rongguo Lu, Shengfei Wang, Yuheng Wang, Ruo Chen, Yuanyuan Gu, Lin Zhu, Komal K. Abhange, Zachary J. Quinn, Yundi Chen, Fei Xue, Mingfeng Zheng, Yuan Wan
Extracellular vesicles (EVs) are cell-released vesicles of submicrometer size. EVs contain a tissue-specific signature wherein a variety of proteins and nucleic acids are selectively packaged. Recent studies validate that EVs can be used for cancer diagnostics, staging, and treatment monitoring. EV-related clinical translation requires effective EV isolation as a prerequisite. However, lengthy procedures, low yield, low throughput, and high levels of contaminants disqualify the existing isolation approaches for large-scale clinical use. Hence, new approaches for rapid, efficient, and low-cost isolation of EVs in high purity for flexible analyses of the diverse contents in real-world clinical settings are highly desired yet are currently unavailable. Here, we report the effective use of heparin/polymer-coated microspheres (HPM) for EV isolation and retrieval. Approximately 81% of EVs can be isolated from plasma in 1 h with depletion of ∼99.5% plasma protein and nucleic acid contaminants, and 72% of isolated EVs can be retrieved with saline in 5 min for various cargo analyses. This approach was further validated with clinical samples derived from patients with malignant ground-glass opacity (GGO). In eight patients, the mutation concordance between EV DNA and tissue DNA is 39.8%. The prevalence and mutation count of EGFR, TP53, and NF1 are higher than those of other oncogenes and antioncogenes that are intensely associated with lung adenocarcinoma. Moreover, different mutation prevalence and patterns between smokers and nonsmokers can be observed. Our findings suggest that the combination of HPM assay and targeted sequencing of EV DNA could be translated in the differential diagnosis of malignant GGO with short turnaround time.

History