Proteomics profiling identifies extracellular vesicles’ cargo associated with tumour cell induced platelet aggregation

Abstract

Background Cancer patients have an increased risk of developing venous thromboembolism, with up to 30% dying within a month of their development. Some cancer cells are known to induce platelet aggregation, and this interaction is understood to contribute to thrombosis and haematogenous metastasis. Many researchers have reported on extracellular vesicles (EVs) released from platelets. However, less is known about how cancer cells’ EVs may affect platelet function. Here EVs released by triple-negative breast cancer (TNBC) cell line variants were extensively investigated in this regard. Methods EVs were separated from conditioned media of TNBC Hs578T and Hs578Ts(i)8 cells using filtration and ultracentrifugation and were characterised by nanoparticle tracking analysis, immunoblots, and transmission electron microscopy. Blood samples from consenting donors were procured, and their platelets collected by differential centrifugation. Light transmission aggregometry and optical microscopy evaluated the potential interaction of TNBC cells and their EVs with platelets. Global proteomic analysis was performed on the EVs, by in-solution digestion and mass spectrometry. Data analysis included the use of Perseus, FunRich, and Vesiclepedia. Immunoblotting was used as a secondary method to investigate some key EV cargo proteins identified by the global proteomics approach. Results Both TNBC cell variants induced platelet aggregation. Increasing cell numbers significantly reduced the time taken for platelet aggregation to occur. EVs released by the cells also resulted in platelet aggregation. The time to induce platelet aggregation was EV dose-dependent. Proteomics profiling and immunoblotting of the EVs’ cargo identified candidate proteins (including uPAR and PDGFRβ) that may be involved during this process. Conclusions TNBC cells induce platelet aggregation. Furthermore, the cell-free EVs induced this undesirable effect. A number of EV cargo proteins were identified that may be relevant as therapeutic targets.

Reprogramming the immunosuppressive tumor microenvironment: exploiting angiogenesis and thrombosis to enhance immunotherapy

“…Similarly, TF can directly activate platelets or facilitate tumor cell-platelet interactions ( 116 , 117 ). EVs released by triple-negative breast cancer cells contain uPAR and PDGFRβ that can induce platelet aggregation ( 118 ). Platelet activation leads to degranulation and release of ADP and thromboxane A2, which can function in an autocrine manner to amplify platelet activation and aggregation ( 119 ).…”

Design, Synthesis, and Antitumor Evaluation of an Opioid Growth Factor Bioconjugate Targeting Pancreatic Ductal Adenocarcinoma

“…Unfortunately, pancreatic cancer is diagnosed in patients at a late stage, when there are already metastases to other tissues, liver, peritoneum, lungs, and bones. The process of cancer metastasis is also associated with the formation of embolisms in the circulatory system caused by the appearance of aggregates between the CTCs and the platelets [ 16 , 19 , 117 , 118 ]. Thrombocytes play a significant role in cancer progression, contributing to the free circulation of CTCs within the blood.…”

“…Platelets also release metalloproteins, which contribute to the digestion of the extracellular matrix and create a new environment for the tumor. Thus, the development of effective drugs that inhibit the TICPA process would help to reduce the pancreatic cancer progression [ 16 , 17 , 18 , 19 , 20 , 21 ].…”

Extracellular vesicles: Function, resilience, biomarker, bioengineering, and clinical implications

“…Accordingly, the cargo carried by EVs encompasses a diverse array of biomolecules, including proteins, nucleic acids, lipids, and metabolites, which reflect the physiological state and pathological alterations of the parent cells. By analyzing the intracellular cargo of EVs, researchers can elucidate their roles in various biological processes and disease pathogenesis [ 28 29 30 32 50 51 52 53 ].…”