Abstract 459: Lipidomic analysis of extracellular vesicles and its potential for the identification of body fluid-based biomarkers for breast cancer diagnosis
Authors: Dorado Erika , Doria M Luisa , Nagelkerke Anika , McKenzie James S , Maneta-Stavrakaki Stefania , Ion Charlotte , Whittaker Thomas , Nicholson Jeremy , Stevens Molly M , Coombes R Charles , Takats Zoltan 
Published: July 2021

Abstract
Background: Tumor-derived Extracellular Vesicles (EVs) are found in body fluids of cancer patients. These EVs include exosomes and microvesicles, which play an important role in tumor development and metastasis. Although EVs have great potential as liquid biopsies for early detection of breast cancer, the lipid composition of breast cancer-derived EVs is unknown. Here, we studied the lipid composition of EVs and its potential for the identification of body fluid-based biomarkers for breast cancer diagnosis.
Methods: The lipid composition of EVs and their parental cells were evaluated for nine breast cancer and two non-cancerous mammary cell lines. EVs were isolated by size-exclusion chromatography (SEC). Furthermore, ten blood plasma samples were studied from women diagnosed with primary breast cancer before surgery, nine samples from patients with progressive metastatic breast cancer and ten samples from healthy women who neither had pre-existing medical conditions nor infections at the time the samples were collected or in the previous weeks. EVs were obtained from blood plasma samples by a combination of density gradient ultracentrifugation and SEC. Lipids were extracted from EVs and analyzed by reverse-phase liquid chromatography mass-spectrometry (LC-MS).
Results: We found that EVs produced by breast cancer cell lines were enriched in sphingomyelins and ceramides when compared to their parental cells. Furthermore, phospholipids such as phosphatidylcholine and phosphatidylethanolamine were found to be abundant in these EVs. Multivariate analyses showed that EVs released by breast cancer cells can be distinguished from those released by non-cancerous cells based on their phospholipid and sphingolipid composition (AUCROC=0.99, p&lt;0.001). A key finding of this study is that we also found significant differences in the phospholipid and sphingolipid composition between EVs found in blood plasma from breast cancer patients and healthy volunteers (AUCROC=0.99, p&lt;0.001). Importantly with respect to the practical applications of this study for early diagnosis of breast cancer, these differences in EV's lipid content were also significant between EVs from primary breast cancer patients and healthy volunteers (AUCROC=0.99, p&lt;0.001). Annotation of specific lipid species are currently being carried out by MS/MS analysis.
Conclusion: Our findings demonstrate the potential of the lipid content of cancer-derived EVs for the identification of lipid biomarkers for breast cancer diagnosis and its use for the development of non-invasive (or minimally invasive) diagnostic methods based on blood plasma analysis.
Citation Format: Erika Dorado, M Luisa Doria, Anika Nagelkerke, James S McKenzie, Stefania Maneta-Stavrakaki, Charlotte Ion, Thomas Whittaker, Jeremy Nicholson, Molly M Stevens, R Charles Coombes, Zoltan Takats. Lipidomic analysis of extracellular vesicles and its potential for the identification of body fluid-based biomarkers for breast cancer diagnosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 459.