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
Affiliations
Mikołaj Piotr Zaborowski
Department of Neurology, Massachusetts General Hospital, Charlestown, MA 02129, USA; Program in Neuroscience, Harvard Medical School, Boston, MA 02115, USA; Department of Gynecology, Obstetrics and Gynecologic Oncology, Division of Gynecologic Oncology, Poznań University of Medical Sciences, 60-535 Poznań, Poland; Corresponding author
Kyungheon Lee
Center for Systems Biology, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Radiology, Massachusetts General Hospital, Boston, MA 02114, USA
Young Jeong Na
Cancer Center, Massachusetts General Hospital, Boston, MA 02114, USA
Alessandro Sammarco
Department of Neurology, Massachusetts General Hospital, Charlestown, MA 02129, USA; Program in Neuroscience, Harvard Medical School, Boston, MA 02115, USA; Department of Comparative Biomedicine and Food Science, University of Padua, 35020 Padua, Italy
Xuan Zhang
Department of Neurology, Massachusetts General Hospital, Charlestown, MA 02129, USA; Program in Neuroscience, Harvard Medical School, Boston, MA 02115, USA
Marcin Iwanicki
Department of Chemistry and Chemical Biology, Stevens Institute of Technology, Hoboken, NJ 07030, USA
Pike See Cheah
Department of Neurology, Massachusetts General Hospital, Charlestown, MA 02129, USA; Program in Neuroscience, Harvard Medical School, Boston, MA 02115, USA; Department of Human Anatomy, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, 43400 Seri Kembangan, Malaysia
Hsing-Ying Lin
Center for Systems Biology, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Radiology, Massachusetts General Hospital, Boston, MA 02114, USA
Max Zinter
Department of Neurology, Massachusetts General Hospital, Charlestown, MA 02129, USA; Department of Radiology, Massachusetts General Hospital, Boston, MA 02114, USA
Chung-Yu Chou
Center for Systems Biology, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Biomedical Sciences and Engineering, National Central University, Taoyuan City 320, Taiwan
Giulia Fulci
Cancer Center, Massachusetts General Hospital, Boston, MA 02114, USA
Bakhos A. Tannous
Department of Neurology, Massachusetts General Hospital, Charlestown, MA 02129, USA; Program in Neuroscience, Harvard Medical School, Boston, MA 02115, USA; Department of Radiology, Massachusetts General Hospital, Boston, MA 02114, USA
Charles Pin-Kuang Lai
Department of Neurology, Massachusetts General Hospital, Charlestown, MA 02129, USA; Program in Neuroscience, Harvard Medical School, Boston, MA 02115, USA
Michael J. Birrer
Cancer Center, Massachusetts General Hospital, Boston, MA 02114, USA
Ralph Weissleder
Center for Systems Biology, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Radiology, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA
Hakho Lee
Center for Systems Biology, Massachusetts General Hospital, Boston, MA 02114, USA; Department of Radiology, Massachusetts General Hospital, Boston, MA 02114, USA
Xandra O. Breakefield
Department of Neurology, Massachusetts General Hospital, Charlestown, MA 02129, USA; Program in Neuroscience, Harvard Medical School, Boston, MA 02115, USA; Department of Radiology, Massachusetts General Hospital, Boston, MA 02114, USA; Corresponding author
Summary: 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. Keywords: extracellular vesicles, membrane proteins, biomarker, The Cancer Genome Atlas, Genotype-Tissue Expression Project, Human Protein Atlas, palmitoylated fluorescent protein, membrane-bound Gaussia luciferase
