Extracellular Vesicle Membrane-Associated Proteins: Emerging Roles in Tumor Angiogenesis and Anti-Angiogenesis Therapy Resistance

Abstract

The tumor vasculature is essential for tumor growth and metastasis, and is a prime target of several anti-cancer agents. Increasing evidence indicates that tumor angiogenesis is stimulated by extracellular vesicles (EVs) that are secreted or shed by cancer cells. These EVs encapsulate a variety of biomolecules with angiogenic properties, and have been largely thought to stimulate vessel formation by transferring this luminal cargo into endothelial cells. However, recent studies have revealed that EVs can also signal to recipient cells via proteins on the vesicular surface. This review discusses and integrates emerging insights into the diverse mechanisms by which proteins associate with the EV membrane, the biological functions of EV membrane-associated proteins in tumor angiogenesis, and the clinical significance of these proteins in anti-angiogenic therapy.

Keywords: anti-angiogenic therapy; cytokines; extracellular vesicles; membrane proteins; protein sorting; therapy resistance; tumor angiogenesis.

Figure 1

Schematic representation of protein sorting and secretory pathways. (A) Cell surface receptors and other integral membrane proteins (shown in dark blue) are internalized by endocytosis and routed to early endosomes (green arrow). From there, the proteins are recycled back to the plasma membrane, or are sorted into intraluminal vesicles (ILV) that form through inward budding of endosomes and give rise to multivesicular bodies (MVB). Fusion of MVBs with lysosomes results in degradation of ILVs and their cargo, whereas fusion of MVBs with the plasma membrane results in release of ILVs as exosomes. Unlike exosomes, ectosomes (also known as microvesicles or microparticles) form through outward budding of the plasma membrane. (B) Most receptor ligands (shown in pink) are trafficked to the endoplasmic reticulum (ER) and then to the Golgi complex (red arrow). From there, the proteins are routed to endosomes, or are packaged into secretory granules that then fuse with the plasma membrane, resulting in release of proteins into the extracellular milieu.

Figure 2

Differential sorting of vascular endothelial growth factor (VEGF) isoforms to EVs and impact on anti-angiogenic therapy. Alternative splicing of vascular endothelial growth factor A (VEGFA) mRNA yields several VEGF isoforms of which the 121, 165, and 189 amino acid variants are the most commonly expressed isoforms in tumors. The VEGF₁₈₉ isoform is selectively enriched in cancer cell-derived small EVs (sEV). VEGF₁₈₉ is present on the surface of small EVs as a covalent homodimer and interacts with the EV surface through binding heparan sulfate. By contrast, VEGF_90K is selectively enriched in cancer cell-derived microvesicles (MV). VEGF_90K comprises VEGF₁₆₅ that is crosslinked by tissue transglutaminase (tTG) and interacts with the EV surface through binding heat shock protein 90 (Hsp90). Both small EV-associated VEGF and microvesicle-associated VEGF stimulate VEGF receptor signaling and tumor angiogenesis, but are not neutralized by the therapeutic VEGF antibody bevacizumab.