Methods for Systematic Identification of Membrane Proteins for Specific Capture of Cancer-Derived Extracellular Vesicles

Mikołaj Piotr Zaborowski, Kyungheon Lee, Young Jeong Na, Alessandro Sammarco, Xuan Zhang, Marcin Iwanicki, Pike See Cheah, Hsing Ying Lin, Max Zinter, Chung Yu Chou, Giulia Fulci, Bakhos A. Tannous, Charles Pin Kuang Lai, Michael J. Birrer, Ralph Weissleder, Hakho Lee, Xandra O. Breakefield

Research output: Contribution to journalArticlepeer-review

34 Scopus citations

Abstract

Analysis of cancer-derived extracellular vesicles (EVs) in biofluids potentially provides a source of disease biomarkers. At present there is no procedure to systematically identify which antigens should be targeted to differentiate cancer-derived from normal host cell-derived EVs. Here, we propose a computational framework that integrates information about membrane proteins in tumors and normal tissues from databases: UniProt, The Cancer Genome Atlas, the Genotype-Tissue Expression Project, and the Human Protein Atlas. We developed two methods to assess capture of EVs from specific cell types. (1) We used palmitoylated fluorescent protein (palmtdTomato) to label tumor-derived EVs. Beads displaying antibodies of interest were incubated with conditioned medium from palmtdTomato-expressing cells. Bound EVs were quantified using flow cytometry. (2) We also showed that membrane-bound Gaussia luciferase allows the detection of cancer-derived EVs in blood of tumor-bearing animals. Our analytical and validation platform should be applicable to identify antigens on EVs from any tumor type. Cancer cell-derived extracellular vesicles (EVs) can be used in diagnostics, but their enrichment remains challenging. Zaborowski et al. identify membrane proteins enriched on the surface of cancer cells compared with normal tissues using TCGA, the Human Protein Atlas, and GTEx and present methods to measure immunocapture of cancer EVs in vitro and in animal models.

Original languageEnglish
Pages (from-to)255-268.e6
JournalCell Reports
Volume27
Issue number1
DOIs
StatePublished - 2 Apr 2019

Keywords

  • Genotype-Tissue Expression Project
  • Human Protein Atlas
  • The Cancer Genome Atlas
  • biomarker
  • extracellular vesicles
  • membrane proteins
  • membrane-bound Gaussia luciferase
  • palmitoylated fluorescent protein

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