Glycan modification of glioblastoma-derived extracellular vesicles enhances receptor-mediated targeting of dendritic cells

Sophie A Dusoswa; Sophie K Horrevorts; Martino Ambrosini; Hakan Kalay; Nanne J Paauw; Rienk Nieuwland; Michiel D Pegtel; Tom Würdinger; Yvette Van Kooyk; Juan J Garcia-Vallejo

doi:10.1080/20013078.2019.1648995

Glycan modification of glioblastoma-derived extracellular vesicles enhances receptor-mediated targeting of dendritic cells

J Extracell Vesicles. 2019 Aug 9;8(1):1648995. doi: 10.1080/20013078.2019.1648995. eCollection 2019.

Affiliations

¹ Department of Molecular Cell Biology & Immunology, Amsterdam Infection & Immunity Institute and Cancer Center Amsterdam, Amsterdam UMC, Vrije Universiteit Amsterdam, Amsterdam, The Netherlands.
² Laboratory of Experimental Clinical Chemistry, and Vesicle Observation Centre, Amsterdam UMC, University of Amsterdam, Amsterdam, The Netherlands.
³ Department of Pathology, Amsterdam UMC, Vrije Universiteit Amsterdam, Amsterdam, The Netherlands.

Abstract

Glioblastoma is the most prevalent and aggressive primary brain tumour for which total tumour lysate-pulsed dendritic cell vaccination is currently under clinical evaluation. Glioblastoma extracellular vesicles (EVs) may represent an enriched cell-free source of tumour-associated (neo-) antigens to pulse dendritic cells (DCs) for the initiation of an anti-tumour immune response. Capture and uptake of EVs by DCs could occur in a receptor-mediated and presumably glycan-dependent way, yet the glycan composition of glioblastoma EVs is unknown. Here, we set out to characterize the glycocalyx composition of glioblastoma EVs by lectin-binding ELISA and comprehensive immunogold transmission electron microscopy (immuno-TEM). The surface glycan profile of human glioblastoma cell line-derived EVs (50-200 nm) was dominated by α-2,3- and α-2,6 linked sialic acid-capped complex N-glycans and bi-antennary N-glycans. Since sialic acids can trigger immune inhibitory sialic acid-binding Ig-like lectin (Siglec) receptors, we screened for Siglec ligands on the EVs. Glioblastoma EVs showed significant binding to Siglec-9, which is highly expressed on DCs. Surprisingly, however, glioblastoma EVs lack glycans that could bind Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Non-integrin (DC-SIGN, CD209), a receptor that mediates uptake and induction of CD4⁺ and CD8⁺ T cell activation. Therefore, we explored whether modification of the EV glycan surface could reduce immune inhibitory Siglec binding, while enhancing EV internalization by DCs in a DC-SIGN dependent manner. Desialylation with a pan-sialic acid hydrolase led to reduction of sialic acid expression on EVs. Moreover, insertion of a high-affinity ligand (Lewis^Y) for DC-SIGN resulted in a four-fold increase of uptake by monocyte-derived DCs. In conclusion, we show that the glycocalyx composition of EVs is a key factor of efficient DC targeting and that modification of the EV glycocalyx potentiates EVs as anti-cancer vaccine.

Keywords: DC; EV; Glioblastoma; cancer vaccine; dendritic cells; extracellular vesicles; glioma; glycans.

Grants and funding

This work was supported by the European Research Council [ERC Advanced Glycotreat 339977]; Institute for Chemical Immunology [ICI0011]; Cancer Center Amsterdam [CCA2014-5-18].