Microenvironmental Hypoxia Induces Dynamic Changes in Lung Cancer Synthesis and Secretion of Into Extracellular Vesicles

Background: Extracellular vesicles (EVs) mediate critical intercellular communication within healthy tissues, but are also exploited by tumour cells to promote angiogenesis, metastasis, and host immunosuppression under hypoxic stress. We hypothesize that oxygen starvation in developing tumours induces specific hypoxia-sensitive proteins for packing into small EVs to modulate its microenvironment for cancer progression and enhance malignancy. Methods: We employed a heavy isotope pulse/trace quantitative proteomic approach to study hypoxia-sensitive EVs proteins (HSEPs) in hypoxic A549 lung adenocarcinoma cells derived small EVs (<200 nm). Proteomics data mining and pathway analysis were used to reveal potential roles of the HSEPs in enhancing tumour cell progression and in modulating host immunity. Functional clustering was applied to study enhanced EVs biogenesis and secretion in hypoxic cancer cells. Subsequent biochemical functional assays were performed in A549 and H1299 lung cancer cells to validate the hypoxic cancer-derived EVs in promoting cancer progression.Results: Results revealed that hypoxia stimulated cancer cells to synthesize EVs proteins involved in enhancing tumour cell proliferation (NRSN2, WISP2, SPRX1, LCK), metastasis (GOLM1, STC1, MGAT5B), stemness (STC1, TMEM59), angiogenesis (ANGPTL4), and suppressing host immunity (CD70). In addition, functional clustering analyses revealed that tumour hypoxia was strongly associated with rapid synthesis and EV loading of lysosome-related hydrolases and membrane-trafficking proteins to enhance EVs secretion. Moreover, lung cancer-derived EVs were also enriched in signalling molecules capable of inducing epithelial-mesenchymal transition in recipient cancer cells to promote their migration and invasion. Conclusion: Together, these data indicate that lung cancer-derived EVs can act as paracrine/autocrine mediators of tumorigenesis and metastasis in hypoxic microenvironments. Tumour EVs may therefore offer novel opportunities for useful biomarkers discovery and therapeutic targeting of different cancer types and at different stages according to microenvironmental conditions.