Ectosomes are small heterogeneous membrane vesicles generated by budding from the plasma membrane in a variety of cell types and, more frequently, in tumor cells. They are shed into the extracellular space and are proposed as a novel form of intracellular communication in which information is transmitted from the originating cell to recipient cells without direct cell-to-cell contact. This review focuses on a single population of extracellular vesicles—ectosomes. We summarize recent studies of tumor-derived ectosomes which examine their biogenesis and protein cargo, and their influence on different aspects of cancer progression. We discuss possible clinical implications involving ectosomes as potential biomarkers, diagnostic tools and treatment targets in oncology. The unique composition of the molecules (cargo) that ectosomes carry, and their functional role, depends largely on the state of their originating cell. Through horizontal transfer of a variety of biologically active molecules (including proteins, lipids and nucleic acids) between donor and recipient cells, tumor-derived ectosomes may play functional roles in oncogenic transformation, tumor progression, invasion, metastasis, angiogenesis promotion, escape from immune surveillance, and drug resistance, thereby facilitating disease progression. The presence of tumor-derived ectosomes in body fluids such as the blood and urine of cancer patients makes them potentially useful prognostic and predictive biomarkers. Tumor-derived ectosomes also offer possible targets for multiple therapeutic strategies.

Dramatic changes in glycan biosynthesis during oncogenic transformation result in the emergence of marker glycans on the cell surface. We analysed the N-linked glycans of L1CAM from different stages of melanoma progression, using high-performance liquid chromatography combined with exoglycosidase sequencing, matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry, and lectin probes. L1CAM oligosaccharides are heavily sialylated, mainly digalactosylated, biantennary complex-type structures with galactose β1-4/3-linked to GlcNAc and with or without fucose α1-3/6-linked to GlcNAc. Hybrid, bisected hybrid, bisected triantennary and tetraantennary complex oligosaccharides, and β1-6-branched complex-type glycans with or without lactosamine extensions are expresses at lower abundance. We found that metastatic L1CAM possesses only α2-6-linked sialic acid and the loss of α2-3-linked sialic acid in L1CAM is a phenomenon observed during the transition of melanoma cells from VGP to a metastatic stage. Unexpectedly, we found a novel monoantennary complex-type oligosaccharide with a Galβ1-4Galβ1- epitope capped with sialic acid residues A1[3]G(4)2S_2-3. To our knowledge this is the first report documenting the presence of this oligosaccharide in human cancer. The novel and unique N-glycan should be recognised as a new class of human melanoma marker. In functional tests we demonstrated that the presence of cell surface α2-3-linked sialic acid facilitates the migratory behaviour and increases the invasiveness of primary melanoma cells, and it enhances the motility of metastatic cells. The presence of cell surface α2-6-linked sialic acid enhances the invasive potential of both primary and metastatic melanoma cells. Complex-type oligosaccharides in L1CAM enhance the invasiveness of metastatic melanoma cells.

The aim of the present study was to determine whether stage of invasiveness of bladder cancer cell lines contributes to alterations in glycan pattern of their cadherins. Human non-malignant epithelial cell of ureter HCV29, v-raf transfected HCV29 line (BC3726) and transitional cell cancers of urine bladder Hu456 and T24 were grown in cell culture. Equal amounts of protein from each cell extracts were separated by SDS-PAGE electrophoresis and were blotted on an Immobilon P membrane. Cadherins were immunodetected using anti-pan cadherin mAb and lectin blotting assays were performed, in parallel. N-oligosaccharides were analysed by specific reaction with Galanthus nivalis agglutinin (GNA), Sambucus nigra agglutinin (SNA), Maackia amurensis agglutinin (MAA), Datura stramonium agglutinin (DSA), Aleuria aurantia agglutinin (AAA), Phaseolus vulgaris agglutinin (PHA-L) and wheat germ agglutinin (WGA). The cadherin from HCV29 cell line possessed bi- and/or 2,4-branched triantennary complex type glycans, some of which were alpha2,6-sialylated. The cadherin from BC3726 cell line exhibited exclusively high mannose type glycans. Cadherins from Hu456 and T24 cell lines expressed high mannose type glycans as well as beta1,6-branched oligosaccharides with poly-N-acetyllactosamine structures and alpha2,3-linked sialic acid residues. Additionally, the presence of fucose and alpha2,6-sialic acid residues on the cadherin from T24 cell line was detected. These results indicate that N-glycosylation pattern of cadherin from bladder cancer cell line undergoes modification during carcinogenesis.

Tumour-associated alterations of cell surface glycosylation play a crucial role in the adhesion and metastasis of cancer cells. It is well known that the metastatic potential is associated with increased GlcNAc β1–6 branching in N-glycans of tumour cells specifically recognized by a lectin from Phaseolus vulgaris leukoagglutinin (PHA-L). We identified proteins bearing GlcNAc β1–6 branched N-glycans in the A375 human melanoma cell line by affinity chromatography separation on a PHA-L agarose column, followed by immunoidentification and tandem mass spectrometry (MS/MS) analysis. Amongst the proteins identified were integrin subunits α₂, α₃, α₅ and β₁, as well as N-cadherin and lysosome-associated membrane proteins (LAMP-1 and LAMP-2). In addition, L1, Mac-2 binding protein (Mac-2-BP), activated leukocyte cell adhesion molecule/CD166 (ALCAM) and melanotransferrin were shown to react with PHA-L. Some of these proteins are connected mainly with nervous tissues or the immune system and play a crucial role in cell adhesion processes. The presence of GlcNAc β1–6 branched oligosaccharides in these proteins may influence their adhesion properties, reducing adhesion of the cells to the extracellular matrix (ECM) and thus facilitating tumour cell invasion.