CD44v3 as a Biomarker of Tumor-derived Exosomes in Plasma from Head and Neck Squamous Cell Carcinoma Patients

Background Circulating exosomes play a key role in immune suppression and disease progression and reflect the cargo of their cell origin. By isolating CD3(-) and CD3(+) exosomes from patients’ plasma, the cellular origins of immunoregulatory proteins they carry were identified. However, a tumor-specific marker for separation of tumor-derived exosomes (TEX) is still needed. Here we present a method to capture TEX from HNSCC patient’s plasma and to use these exosomes as potential biomarkers for disease stage and metastasis.

Methods Exosomes were isolated from the plasma of 25 HNSCC patients and 7 healthy donors. Exosomes were separated by immunoaffinity capture using either CD3 or CD44v3. On-bead flow cytometry was used for detection of FAS-L, CSGP4, PD-L1, TGFϐ or EGFr on exosomes. Results were correlated to clinicopathological parameters.

Results CD44v3 levels were significantly higher on TEX from HNSCC cell lines as well as in the CD3(-) fraction (enriched in TEX) of plasma derived exosomes. CD44v3 on CD3(-) exosomes correlated significantly with UICC stage and lymph node metastasis. Even more, levels of CD44v3 on CD3(-) exosomes were higher in patients than in healthy donors. Separating exosomes via CD44v3 and staining them for various inhibitory markers showed that CD44v3(+) exosomes had significantly higher levels of PD-L1, CSGP4, TGFϐ, EGFr and FAS-L compared to CD44v3(-) exosomes. Even more, levels of these markers on CD44v3(+) exosomes correlated with clinicopathological parameters.

Conclusions Separation of exosomes via CD44v3 presents a way of TEX enrichment and may serve as a liquid biomarker for tumor load and disease stage.

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Publication History

Article published online:
10 June 2020