Using extracellular vesicles for earlier detection of bile duct cancer

Hyungsoon Im, PhD, an investigator in the Center for Systems Biology at Massachusetts General Hospital and an assistant professor of Radiology at Harvard Medical School, is the senior author of a new study in Advanced Sciences, Plasmon-Enhanced Single Extracellular Vesicle Analysis for Cholangiocarcinoma Diagnosis.  

What Question Were You Investigating?

Cholangiocarcinoma (CCA), cancer that forms in the bile ducts, is a fatal disease that is often detected after it has spread too far to be surgically removed. There are currently no effective biomarkers or diagnostic tools to detect CCA with high confidence.

Extracellular vesicles (EVs) are small particles released from cells that are used to transport different proteins, nucleic acids, lipids and other materials between cells.

We wanted to see if molecular analysis of extracellular vesicles (EVs) in human bile samples could accurately detect patients with cholangiocarcinoma from patients with other benign or inflammatory conditions.

What Was Your Approach?

We first developed a nanoplasmonic sensing technology, named FLEX (fluorescence-amplified extracellular vesicle sensing technology). FLEX enables multiplexed single EV analysis through a simple and rapid detection assay.

After validation in cell line-derived EVs, we applied the technology to human bile samples collected through an endoscopic retrograde cholangiopancreatography (ERCP) procedure.

Fluorescence-amplified extracellular vesicle (FLEX) sensing technology

A) FLEX gold nanowell chips fabricated on a 4 inch Si wafer. The fabricated wafer is diced into 1 cm2 chips for EV assays. B) A scanning electron micrograph of the FLEX chip shows the periodic gold nanowell structures. C) Finite-difference time-domain (FDTD) simulation shows the enhanced electromagnetic fields confined near the gold nanowell surface. The enhanced fields are responsible for plasmon-enhanced fluorescence amplification. D) Reflectance spectra of the gold nanowell arrays with 200 nm diameter and 500 nm periodicity from FDTD simulation (blue dashed line) and experimental measurement (solid red line). E) Fluorescence signal enhancement of AF488, AF555, and AF647 fluorophores on the FLEX chips compared to a plain glass substrate. We formed a thin polyvinyl alcohol (PVA) layer containing the fluorescence dyes spun-coated on the gold nanowell and glass surfaces, as shown in the schematic in the inset. The data are displayed as mean ± standard deviation from triplicate measurements. F) A representative schematic of an EV captured on the gold nanowell surface. The gold surface is functionalized by a polyethylene glycol (PEG) layer to capture EVs. The captured EVs are then labeled by primary (1°) antibodies followed by fluorophore-conjugated secondary antibodies (2°). G) Representative fluorescence image of fluorescently labeled EVs captured on FLEX and plain gold substrates. For the same sample, EVs captured on the FLEX chip generate stronger fluorescence intensities.

What Did You Find?

  1. The FLEX chips could sensitively detect small tumor-derived EVs that were missed by conventional EV methods.
  2. The molecular analysis of tumor-derived EVs could accurately detect CCA patients with significantly better accuracy (93%) than the current ERCP-based tissue biopsies or serum biomarkers.

What Are the Clinical Implications?

  1. The new method has the potential to detect CCA at earlier stages than is currently possible, which could significantly improve the five-year survival rate for the disease, which remains under 20%.
  2. The molecular analysis of a bile sample could further improve the diagnostic accuracy of the current gold standard pathology diagnosis through ERCP.
  3. With ERCP, up to 20% of brush biopsies are found inconclusive, requiring repeated procedures and delaying treatment.

SourceMassachusetts General Hospital 

Jeong MH, Son T, Tae YK, Park CH, Lee HS, Chung MJ, Park JY, Castro CM, Weissleder R, Jo JH, Bang S, Im H. (2023) Plasmon-Enhanced Single Extracellular Vesicle Analysis for Cholangiocarcinoma Diagnosis. Adv Sci (Weinh) [Epub ahead of print]. [article]

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