Sphingolipids (SLs), glycosphingolipids (GSLs), and eicosanoids are bioactive lipids, which play important roles in the etiology of various diseases, including cancer. However, their content and roles in cancer cells, and in particular in the exosomes derived from tumor cells, remain insufficiently characterized. In this study, we evaluated alterations of SL and GSL levels in transformed cells and their exosomes, using comparative HPLC-MS/MS analysis of parental human bronchial epithelial cells HBEC-12KT and their derivative, benzo[a]pyrene-transformed HBEC-12KT-B1 cells with the acquired mesenchymal phenotype. We examined in parallel SL/GSL contents in the exosomes released from both cell lines. We found significant alterations of the SL/GSL profile in the transformed cell line, which corresponded well with alterations of the SL/GSL profile in exosomes derived from these cells. This suggested that a majority of SLs and GSLs were transported by exosomes in the same relative pattern as in the cells of origin. The only exceptions included decreased contents of sphingosin, sphingosin-1-phosphate, and lactosylceramide in exosomes derived from the transformed cells, as compared with the exosomes derived from the parental cell line. Importantly, we found increased levels of ceramide phosphate, globoside Gb3, and ganglioside GD3 in the exosomes derived from the transformed cells. These positive modulators of epithelial-mesenchymal transition and other pro-carcinogenic processes might thus also contribute to cancer progression in recipient cells. In addition, the transformed HBEC-12KT-B1 cells also produced increased amounts of eicosanoids, in particular prostaglandin E2. Taken together, the exosomes derived from the transformed cells with specifically upregulated SL and GSL species, and increased levels of eicosanoids, might contribute to changes within the cancer microenvironment and in recipient cells, which could in turn participate in cancer development. Future studies should address specific roles of individual SL and GSL species identified in the present study.

Department of Cytokinetics Institute of Biophysics of the Czech Academy of Sciences Kralovopolska 135 61265 Brno Czech Republic

Department of Pharmacology and Toxicology Veterinary Research Institute Hudcova 296 70 62100 Brno Czech Republic

Section for Occupational Toxicology National Institute of Occupational Health 0304 Oslo Norway

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Kalra H., Drummen G.P., Mathivanan S. Focus on extracellular vesicles: Introducing the next small big thing. Int. J. Mol. Sci. 2016;17:170. doi: 10.3390/ijms17020170. PubMed DOI PMC

Gurung S., Perocheau D., Touramanidou L., Baruteau J. The exosome journey: From biogenesis to uptake and intracellular signalling. Cell Commun. Signal. 2021;19:47. doi: 10.1186/s12964-021-00730-1. PubMed DOI PMC

Gurunathan S., Kang M.-H., Qasim M., Khan K., Kim J.-H. Biogenesis, membrane trafficking, functions, and next generation nanotherapeutics medicine of extracellular vesicles. Int. J. Nanomed. 2021;16:3357–3383. doi: 10.2147/IJN.S310357. PubMed DOI PMC

Al-Nedawi K., Meehan B., Micallef J., Lhotak V., May L., Guha A., Rak J. Intercellular transfer of the oncogenic receptor EGFRvIII by microvesicles derived from tumour cells. Nat. Cell Biol. 2008;10:619–624. doi: 10.1038/ncb1725. PubMed DOI

Skog J., Würdinger T., van Rijn S., Meijer D.H., Gainche L., Sena-Esteves M., Curry W.T., Jr., Carter B.S., Krichevsky A.M., Breakefield X.O. Glioblastoma microvesicles transport RNA and proteins that promote tumour growth and provide diagnostic biomarkers. Nat. Cell Biol. 2008;10:1470–1476. doi: 10.1038/ncb1800. PubMed DOI PMC

Keller S., König A.K., Marmé F., Runz S., Wolterink S., Koensgen D., Mustea A., Sehouli J., Altevogt P. Systemic presence and tumor-growth promoting effect of ovarian carcinoma released exosomes. Cancer Lett. 2009;278:73–81. doi: 10.1016/j.canlet.2008.12.028. PubMed DOI

Di Vizio D., Morello M., Dudley A.C., Schow P.W., Adam R.M., Morley S., Mulholland D., Rotinen M., Hager M.H., Insabato L., et al. Large oncosomes in human prostate cancer tissues and in the circulation of mice with metastatic disease. Am. J. Pathol. 2012;181:1573–1584. doi: 10.1016/j.ajpath.2012.07.030. PubMed DOI PMC

Kim J., Kim T.Y., Lee M.S., Mun J.Y., Ihm C., Kim S.A. Exosome cargo reflects TGF-β1-mediated epithelial-to-mesenchymal transition (EMT) status in A549 human lung adenocarcinoma cells. Biochem. Biophys. Res. Commun. 2016;478:643–648. doi: 10.1016/j.bbrc.2016.07.124. PubMed DOI

El-Sayed I.Y., Daher A., Destouches D., Firlej V., Kostallari E., Maillé P., Huet E., Haidar-Ahmad N., Jenster G., de la Taille A., et al. Extracellular vesicles released by mesenchymal-like prostate carcinoma cells modulate EMT state of recipient epithelial-like carcinoma cells through regulation of AR signaling. Cancer Lett. 2017;410:100–111. doi: 10.1016/j.canlet.2017.09.010. PubMed DOI

Bebelman M.P., Smit M.J., Pegtel D.M., Baglio S.R. Biogenesis and function of extracellular vesicles in cancer. Pharmacol. Ther. 2018;188:1–11. doi: 10.1016/j.pharmthera.2018.02.013. PubMed DOI

Latifkar A., Cerione R.A., Antonyak M.A. Probing the mechanisms of extracellular vesicle biogenesis and function in cancer. Biochem. Soc. Trans. 2018;46:1137–1146. doi: 10.1042/BST20180523. PubMed DOI PMC

Jabalee J., Towle R., Garnis C. The role of extracellular vesicles in cancer: Cargo, function, and therapeutic implications. Cells. 2018;7:93. doi: 10.3390/cells7080093. PubMed DOI PMC

Subra C., Laulagnier K., Perret B., Record M. Exosome lipidomics unravels lipid sorting at the level of multivesicular bodies. Biochimie. 2007;89:205–212. doi: 10.1016/j.biochi.2006.10.014. PubMed DOI

Record M., Carayon K., Poirot M., Silvente-Poirot S. Exosomes as new vesicular lipid transporters involved in cell-cell communication and various pathophysiologies. Biochim. Biophys. Acta. 2014;1841:108–120. doi: 10.1016/j.bbalip.2013.10.004. PubMed DOI

Record M., Silvente-Poirot S., Poirot M., Wakelam M.J.O. Extracellular vesicles: Lipids as key components of their biogenesis and functions. J. Lipid Res. 2018;59:1316–1324. doi: 10.1194/jlr.E086173. PubMed DOI PMC

Llorente A., Skotland T., Sylvänne T., Kauhanen D., Róg T., Orłowski A., Vattulainen I., Ekroos K., Sandvig K. Molecular lipidomics of exosomes released by PC-3 prostate cancer cells. Biochim. Biophys. Acta. 2013;1831:1302–1309. doi: 10.1016/j.bbalip.2013.04.011. PubMed DOI

Lydic T.A., Townsend S., Adda C.G., Collins C., Mathivanan S., Reid G.E. Rapid and comprehensive ‘shotgun’ lipidome profiling of colorectal cancer cell derived exosomes. Methods. 2015;87:83–95. doi: 10.1016/j.ymeth.2015.04.014. PubMed DOI PMC

Haraszti R.A., Didiot M.C., Sapp E., Leszyk J., Shaffer S.A., Rockwell H.E., Gao F., Narain N.R., DiFiglia M., Kiebish M.A., et al. High-resolution proteomic and lipidomic analysis of exosomes and microvesicles from different cell sources. J. Extracell. Vesicles. 2016;5:32570. doi: 10.3402/jev.v5.32570. PubMed DOI PMC

Skotland T., Sagini K., Sandvig K., Llorente A. An emerging focus on lipids in extracellular vesicles. Adv. Drug Deliv. Rev. 2020;159:308–321. doi: 10.1016/j.addr.2020.03.002. PubMed DOI

Hannun Y.A., Obeid L.M. Sphingolipids and their metabolism in physiology and disease. Nat. Rev. Mol. Cell. Biol. 2018;19:175–191. doi: 10.1038/nrm.2017.107. PubMed DOI PMC

García-González V., Díaz-Villanueva J.F., Galindo-Hernández O., Martínez-Navarro I., Hurtado-Ureta G., Pérez-Arias A.A. Ceramide metabolism balance, a multifaceted factor in critical steps of breast cancer development. Int. J. Mol. Sci. 2018;19:2527. doi: 10.3390/ijms19092527. PubMed DOI PMC

Furukawa K., Ohmi Y., Ohkawa Y., Bhuiyan R.H., Zhang P., Tajima O., Hashimoto N., Hamamura K., Furukawa K. New era of research on cancer-associated glycosphingolipids. Cancer Sci. 2019;10:1544–1551. doi: 10.1111/cas.14005. PubMed DOI PMC

Cumin C., Huang Y.L., Everest-Dass A., Jacob F. Deciphering the importance of glycosphingolipids on cellular and molecular mechanisms associated with epithelial-to-mesenchymal transition in cancer. Biomolecules. 2021;11:62. doi: 10.3390/biom11010062. PubMed DOI PMC

Furukawa K., Ohkawa Y., Yamauchi Y., Hamamura K., Ohmi Y., Furukawa K. Fine tuning of cell signals by glycosylation. J. Biochem. 2012;151:573–578. doi: 10.1093/jb/mvs043. PubMed DOI

Machala M., Procházková J., Hofmanová J., Králiková L., Slavík J., Tylichová Z., Ovesná P., Kozubík A., Vondráček J. Colon cancer and perturbations of the sphingolipid metabolism. Int. J. Mol. Sci. 2019;20:6051. doi: 10.3390/ijms20236051. PubMed DOI PMC

Russo D., Capolupo L., Loomba J.S., Sticco L., D’Angelo G. Glycosphingolipid metabolism in cell fate specification. J. Cell Sci. 2018;131:jcs219204. doi: 10.1242/jcs.219204. PubMed DOI

Finetti F., Travelli C., Ercoli J., Colombo G., Buoso E., Trabalzini L. Prostaglandin E2 and cancer: Insight into tumor progression and immunity. Biology. 2020;9:434. doi: 10.3390/biology9120434. PubMed DOI PMC

Pettus B.J., Chalfant C.E., Hannun Y.A. Sphingolipids in inflammation: Roles and implications. Curr. Mol. Med. 2004;4:405–418. doi: 10.2174/1566524043360573. PubMed DOI

Kawamori T., Kaneshiro T., Okumura M., Maalouf S., Uflacker A., Bielawski J., Hannun Y.A., Obeid L.M. Role for sphingosine kinase 1 in colon carcinogenesis. FASEB J. 2009;23:405–414. doi: 10.1096/fj.08-117572. PubMed DOI PMC

Bersaas A., Arnoldussen Y.J., Sjøberg M., Haugen A., Mollerup S. Epithelial-mesenchymal transition and FOXA genes during tobacco smoke carcinogen induced transformation of human bronchial epithelial cells. Toxicol. In Vitro. 2016;35:55–65. doi: 10.1016/j.tiv.2016.04.012. PubMed DOI

Menter D.G., Dubois R.N. Prostaglandins in cancer cell adhesion, migration and invasion. Int. J. Cell Biol. 2012;2012:723419. doi: 10.1155/2012/723419. PubMed DOI PMC

Biringer R.G. A review of prostanoid receptors: Expression, characterization, regulation and mechanism of action. J. Cell Commun. Signal. 2021;15:155–184. doi: 10.1007/s12079-020-00585-0. PubMed DOI PMC

Vangaveti V., Baune B.T., Kennedy R.L. Hydroxyoctadecadienoic acids: Novel regulators of macrophage differentiation and atherogenesis. Ther. Adv. Endocrinol. Metab. 2010;1:51–60. doi: 10.1177/2042018810375656. PubMed DOI PMC

Van’t Erve T.J., Lih F.B., Jelsema C., Deterding L.J., Eling T.E., Mason R.P., Kadiiska M.B. Reinterpreting the best biomarker of oxidative stress: The 8-iso-prostaglandin F2α/prostaglandin F2α ratio shows complex origins of lipid peroxidation biomarkers in animal models. Free Radic. Biol. Med. 2016;95:65–73. doi: 10.1016/j.freeradbiomed.2016.03.001. PubMed DOI PMC

Patwardhan G.A., Liu Y.Y. Sphingolipids and expression regulation of genes in cancer. Prog. Lipid Res. 2011;50:104–114. doi: 10.1016/j.plipres.2010.10.003. PubMed DOI PMC

Gomez-Muñoz A., Presa N., Gomez-Larrauri A., Rivera I.G., Trueba M., Ordoñez M. Control of inflammatory responses by ceramide, sphingosine 1-phosphate and ceramide 1-phosphate. Prog. Lipid Res. 2016;61:51–62. doi: 10.1016/j.plipres.2015.09.002. PubMed DOI

Basu S., Rossary A., Vasson M. Role of inflammation and eicosanoids in breast cancer. Lipid Technol. 2016;28:60–64. doi: 10.1002/lite.201600017. DOI

Moreno J.J. New aspects of the role of hydroxyeicosatetraenoic acids in cell growth and cancer development. Biochem. Pharmacol. 2009;77:1–10. doi: 10.1016/j.bcp.2008.07.033. PubMed DOI

Nakamura H., Murayama T. The role of sphingolipids in arachidonic acid metabolism. J. Pharmacol. Sci. 2014;124:307–312. doi: 10.1254/jphs.13R18CP. PubMed DOI

Sampaio J.L., Gerl M.J., Klose C., Ejsing C.S., Beug H., Simons K., Shevchenko A. Membrane lipidome of an epithelial cell line. Proc. Natl. Acad. Sci. USA. 2011;108:1903–1907. doi: 10.1073/pnas.1019267108. PubMed DOI PMC

Peng J., Chen B., Shen Z., Deng H., Liu D., Xie X., Gan X., Xu X., Huang Z., Chen J. DNA promoter hypermethylation contributes to down-regulation of galactocerebrosidase gene in lung and head and neck cancers. Int. J. Clin. Exp. Pathol. 2015;8:11042–11050. PubMed PMC

Kovbasnjuk O., Mourtazina R., Baibakov B., Wang T., Elowsky C., Choti M.A., Kane A., Donowitz M. The glycosphingolipid globotriaosylceramide in the metastatic transformation of colon cancer. Proc. Natl. Acad. Sci. USA. 2005;102:19087–19092. doi: 10.1073/pnas.0506474102. PubMed DOI PMC

Tyler A., Johansson A., Karlsson T., Gudey S.K., Brännström T., Grankvist K., Behnam-Motlagh P. Targeting glucosylceramide synthase induction of cell surface globotriaosylceramide (Gb3) in acquired cisplatin-resistance of lung cancer and malignant pleural mesothelioma cells. Exp. Cell Res. 2015;336:23–32. doi: 10.1016/j.yexcr.2015.05.012. PubMed DOI

Ramirez R.D., Sheridan S., Girard L., Sato M., Kim Y., Pollack J., Peyton M., Zou Y., Kurie J.M., Dimaio J.M., et al. Immortalization of human bronchial epithelial cells in the absence of viral oncoproteins. Cancer Res. 2004;64:9027–9034. doi: 10.1158/0008-5472.CAN-04-3703. PubMed DOI

Théry C., Amigorena S., Raposo G., Clayton A. Isolation and characterization of exosomes from cell culture supernatants and biological fluids. Curr. Protoc. Cell Biol. 2006;30:3–22. doi: 10.1002/0471143030.cb0322s30. PubMed DOI

Pospichalova V., Svoboda J., Dave Z., Kotrbova A., Kaiser K., Klemova D., Ilkovics L., Hampl A., Crha I., Jandakova E., et al. Simplified protocol for flow cytometry analysis of fluorescently labeled exosomes and microvesicles using dedicated flow cytometer. J. Extracell. Vesicles. 2015;4:25530. doi: 10.3402/jev.v4.25530. PubMed DOI PMC

Prochazkova J., Slavik J., Bouchal J., Levkova M., Huskova Z., Ehrmann J., Ovesna P., Kolar Z., Skalicky P., Strakova N., et al. Specific alterations of sphingolipid metabolism identified in EpCAM-positive cells isolated from human colon tumors. Biochim. Biophys. Acta Mol. Cell Biol. Lipids. 2020;1865:158742. doi: 10.1016/j.bbalip.2020.158742. PubMed DOI

Schmittgen T.D., Livak K.J. Analyzing real-time PCR data by the comparative C(T) method. Nat. Protoc. 2008;3:1101–1108. doi: 10.1038/nprot.2008.73. PubMed DOI

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