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EBioMedicine. 2019 Oct;48:23-35. doi: 10.1016/j.ebiom.2019.09.025. Epub 2019 Oct 15.

Characterization of plasma-derived protoporphyrin-IX-positive extracellular vesicles following 5-ALA use in patients with malignant glioma.

Author information

1
Department of Neurosurgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, United States.
2
Department of Pathology, Flow Cytometry Core, Massachusetts General Hospital, Boston, MA, United States.
3
Scintillon Institute, La Jolla, CA, United States.
4
Flow Cytometry Core, Beth Israel Deaconess Medical Center, Boston, MA, United States.
5
Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, United States.
6
Department of Neurosurgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, United States. Electronic address: balaj.leonora@mgh.harvard.edu.
7
Department of Neurosurgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, United States. Electronic address: bcarter@mgh.harvard.edu.

Abstract

BACKGROUND:

Malignant gliomas are rapidly progressive brain tumors with high mortality. Fluorescence guided surgery (FGS) with 5-aminolevulinic acid (5-ALA) provides fluorescent delineation of malignant tissue, which helps achieve maximum safe resection. 5-ALA-based fluorescence is due to preferential accumulation of the fluorophore protoporphyrin-IX (PpIX) in malignant glioma tissue. Additionally, gliomas cells release extracellular vesicles (EVs) which carry biomarkers of disease. Herein, we performed animal and human studies to investigate whether 5-ALA dosed glioma cells, in vitro and in vivo, release PpIX positive EVs in circulation which can be captured and analyzed.

METHODS:

We used imaging flow cytometry (IFC) to characterize PpIX-positive EVs released from 5-ALA-dosed glioma cells, glioma-bearing xenograft models, as well as patients with malignant glioma undergoing FGS.

FINDINGS:

We first show that glioma cells dosed with 5-ALA release 247-fold higher PpIX positive EVs compared to mock dosed glioma cells. Second, we demonstrate that the plasma of glioma-bearing mice (n = 2) dosed with 5-ALA contain significantly higher levels of circulating PpIX-positive EVs than their pre-dosing background (p = 0.004). Lastly, we also show that the plasma of patients with avidly fluorescent tumors (n = 4) undergoing FGS contain circulating PpIX-positive EVs at levels significantly higher than their pre-dosing background (p = 0.00009) and this rise in signal correlates with enhancing tumor volumes (r 2  = 0.888).

INTERPRETATION:

Our findings highlight the potential of plasma-derived PpIX-positive EV-based diagnostics for malignant gliomas, offering a novel liquid biopsy platform for confirming and monitoring tumor status.

KEYWORDS:

5-ALA; Extracellular vesicles; Fluorescence guided surgery; Imaging flow cytometry; Liquid biopsy; Malignant gliomas

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