Extracellular vesicles released during hypoxia transport heparanase and enhance macrophage migration, endothelial tube formation and cancer cell stemness.

Heparanase is upregulated during the progression of most cancers and via its enzyme activity promotes extracellular matrix degradation, angiogenesis and cell migration. Heparanase expression is often associated with enhanced tumor aggressiveness and chemoresistance. We previously demonstrated that increased heparanase expression in tumor cells enhances secretion and alters the composition of tumor-released exosomes. In the present study, we discovered that extracellular vesicles (EVs) secreted by human multiple myeloma cells growing in hypoxic conditions exhibited elevated levels of heparanase cargo compared to EVs from cells growing in normoxic conditions. When macrophages (RAW 264.7 monocyte/macrophage-like cells) were exposed to EVs released by tumor cells growing in either hypoxic or normoxic conditions, macrophage migration and invasion was elevated by EVs from hypoxic conditions. The elevated invasion of macrophages was blocked by a monoclonal antibody that inhibits heparanase enzyme activity. Moreover, the heparanase-bearing EVs from hypoxic cells greatly enhanced endothelial cell tube formation consistent with the known role of heparanase in promoting angiogenesis. EVs from hypoxic tumor cells when compared with EVs from normoxic cells also enhanced cancer stemness properties of both CAG and RPMI 8226 human myeloma cells. Together these data indicate that under hypoxic conditions, tumor cells secrete EVs having an elevated level of heparanase as cargo. These EVs can act on both tumor and non-tumor cells, enhancing tumor progression and tumor cell stemness that likely supports chemoresistance and relapse of tumor.