Exosomes play important roles in cancer progression. Although its contents (e.g., proteins and microRNAs) have been focused on in cancer research, particularly as potential diagnostic markers, the exosome behavior and methods for exosome quantification remain unclear.
In the present study, researchers from the Chiba University Graduate School of Medicine, Japan analyzed the tumor-derived exosome behavior and assessed the quantification of exosomes in patient plasma as a biomarker for esophageal squamous cell carcinoma (ESCC). A CD63-GFP expressing human ESCC cell line (TE2-CD63-GFP) was made by transfection, and mouse subcutaneous tumor models were established. Fluorescence imaging was performed on tumors and plasma exosomes harvested from mice. GFP-positive small vesicles were confirmed in the plasma obtained from TE2-CD63-GFP tumor-bearing mice. Patient plasma was collected in Chiba University Hospital (n=86). Exosomes were extracted from 100 µl of the plasma and quantified by acetylcholinesterase (AChE) activity. The relationship between exosome quantification and the patient clinical characteristics was assessed. The quantification of exosomes isolated from the patient plasma revealed that esophageal cancer patients (n=66) expressed higher exosome levels than non-malignant patients (n=20) (P=0.0002). Although there was no correlation between the tumor progression and the exosome levels, exosome number was the independent prognostic marker and low levels of exosome predicted a poor prognosis (P=0.03).
(A) R epresentative images of subcutaneous tumor composed of TE2-CD63-GFP and TE2 cells [bright-field (BF), GFP fluorescence, H&E stain, immunohistochemistry (IHC) staining of CD63]. Each tumor was established by a subcutaneous injection of 1×107 cells of TE2-CD63-GFP or TE2. Eight weeks after injection, the tumors were harvested and imaged. The tumors had similar appearance in H&E stain and IHC stain of CD63, but only TE2-CD63-GFP tumor had GFP-positive foci in the tumor. (B) The enlarged image of TE2-CD63-GFP tumor. GFP expressing foci were confirmed mainly in cytoplasm as well as the cultured TE2-CD63-GFP cells (bar, 20 μm). (C) A representative GFP image of isolated exosomes from mouse plasma bearing a TE2-CD63-GFP tumor. GFP-positive, but RFP‑negative, small foci were confirmed (bar, 10 μm).