{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,24]],"date-time":"2026-09-24T07:08:47Z","timestamp":1790233727367,"version":"4.1.0"},"reference-count":43,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2026,6,15]],"date-time":"2026-06-15T00:00:00Z","timestamp":1781481600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pharmaceutics"],"abstract":"<jats:p>Background\/Objectives: Colorectal cancer (CRC) remains a leading cause of cancer-related mortality, with poor outcomes in advanced stages and significant limitations in current chemotherapy regimens due to systemic toxicity. Extracellular vesicles (EVs) have emerged as promising natural drug delivery vehicles, offering the potential for targeted, less toxic therapies. This study investigates the feasibility of using autologous, patient-derived EVs as a delivery system for the chemotherapeutic agent doxorubicin, focusing on how disease stage and the EV protein corona influence loading and delivery efficiency. Methods: EVs were isolated from plasma and tissue samples of CRC patients at different disease stages, as well as from healthy controls, demonstrating successful isolation and characterisation of EVs, with distinct profiles across different sources. Results: Doxorubicin loading into EVs was significantly higher in CRC patient-derived EVs compared to healthy controls, and tissue-derived EVs yielded higher quantities of drug-loaded particles. Delivery of doxorubicin-loaded EVs to recipient CRC cell lines (SW480 and SW620) revealed that disease stage impacts both EV uptake and drug delivery, with late-stage EVs showing reduced uptake and delivery efficiency. The protein corona, known to coat circulating EVs, was found to influence drug loading and delivery. Pre-treatment of cell line-derived EVs with plasma proteins enhanced EV uptake but reduced doxorubicin loading and subsequent delivery, particularly when using plasma from healthy volunteers. Conclusions: These findings underscore the importance of EV source and protein corona composition in optimising drug delivery strategies. Our results suggest that autologous, patient-derived EVs hold potential as a targeted drug delivery system for CRC, but highlight the need for further optimisation of EV isolation, loading methods, and understanding of how disease progression affects EV functionality. This approach could ultimately reduce systemic toxicity and improve therapeutic outcomes for CRC patients.<\/jats:p>","DOI":"10.3390\/pharmaceutics18060740","type":"journal-article","created":{"date-parts":[[2026,6,17]],"date-time":"2026-06-17T05:56:31Z","timestamp":1781675791000},"page":"740","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Chemotherapeutic Loading and Delivery of Patient-Derived Extracellular Vesicles Are Influenced by Colorectal Cancer Disease Stage and Protein Corona"],"prefix":"10.3390","volume":"18","author":[{"given":"Otman","family":"Saud","sequence":"first","affiliation":[{"name":"Biomolecular Sciences Research Centre, Sheffield Hallam University, Sheffield S1 1WB, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dallal","family":"Blidi","sequence":"additional","affiliation":[{"name":"Biomolecular Sciences Research Centre, Sheffield Hallam University, Sheffield S1 1WB, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Emily","family":"Hayes","sequence":"additional","affiliation":[{"name":"Biomolecular Sciences Research Centre, Sheffield Hallam University, Sheffield S1 1WB, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Celine","family":"Souilhol","sequence":"additional","affiliation":[{"name":"Biomolecular Sciences Research Centre, Sheffield Hallam University, Sheffield S1 1WB, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rawan","family":"Maani","sequence":"additional","affiliation":[{"name":"Biomolecular Sciences Research Centre, Sheffield Hallam University, Sheffield S1 1WB, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7676-2464","authenticated-orcid":false,"given":"Alice","family":"Johnson","sequence":"additional","affiliation":[{"name":"Biomolecular Sciences Research Centre, Sheffield Hallam University, Sheffield S1 1WB, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6523-2230","authenticated-orcid":false,"given":"Keith","family":"Chapple","sequence":"additional","affiliation":[{"name":"Colorectal Surgical Unit, Sheffield Teaching Hospitals NHS Foundation Trust, Sheffield S10 2JF, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9755-3175","authenticated-orcid":false,"given":"Nick","family":"Peake","sequence":"additional","affiliation":[{"name":"Biomolecular Sciences Research Centre, Sheffield Hallam University, Sheffield S1 1WB, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,15]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"2534098","DOI":"10.1080\/07853890.2025.2534098","article-title":"Global, regional, and national trends in colorectal cancer from 2010 to 2021: An analysis of the global burden of disease study 2021","volume":"57","author":"Chen","year":"2025","journal-title":"Ann. Med."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Wu, S., Zhang, Y., Lin, Z., and Wei, M. (2025). Global burden of colorectal cancer in 2022 and projections to 2050: Incidence and mortality estimates from GLOBOCAN. BMC Cancer, 25.","DOI":"10.1186\/s12885-025-15138-0"},{"key":"ref_3","unstructured":"(2026, March 06). CRUK Statistics. Available online: https:\/\/www.cancerresearchuk.org\/about-cancer\/bowel-cancer\/survival."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1136","DOI":"10.5306\/wjco.v15.i9.1136","article-title":"Colorectal cancer: Recent advances in management and treatment","volume":"15","author":"Fadlallah","year":"2024","journal-title":"World J. Clin. Oncol."},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Han, C.J., Ning, X., Burd, C.E., Spakowicz, D.J., Tounkara, F., Kalady, M.F., Noonan, A.M., McCabe, S., and Von Ah, D. (2024). Chemotoxicity and Associated Risk Factors in Colorectal Cancer: A Systematic Review and Meta-Analysis. Cancers, 16.","DOI":"10.20944\/preprints202405.0890.v1"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"e12144","DOI":"10.1002\/jev2.12144","article-title":"A brief history of nearly EV-erything\u2014The rise and rise of extracellular vesicles","volume":"10","author":"Couch","year":"2021","journal-title":"J. Extracell. Vesicles"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"110651","DOI":"10.1016\/j.celrep.2022.110651","article-title":"Uptake, functionality, and re-release of extracellular vesicle-encapsulated cargo","volume":"39","author":"Ughetto","year":"2022","journal-title":"Cell Rep."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1864","DOI":"10.1038\/s41467-021-22126-y","article-title":"Quantitative characterization of extracellular vesicle uptake and content delivery within mammalian cells","volume":"12","author":"Bonsergent","year":"2021","journal-title":"Nat. Commun."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"28","DOI":"10.1186\/s12575-023-00220-3","article-title":"Extracellular Vesicles for Drug Delivery in Cancer Treatment","volume":"25","author":"Wang","year":"2023","journal-title":"Biol. Proced. Online"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Wang, J., Yin, B., Lian, J., and Wang, X. (2024). Extracellular Vesicles as Drug Delivery System for Cancer Therapy. Pharmaceutics, 16.","DOI":"10.3390\/pharmaceutics16081029"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"329","DOI":"10.1038\/nature15756","article-title":"Tumour exosome integrins determine organotropic metastasis","volume":"527","author":"Hoshino","year":"2015","journal-title":"Nature"},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Linxweiler, J., Kolbinger, A., Himbert, D., Zeuschner, P., Saar, M., St\u00f6ckle, M., and Junker, K. (2021). Organ-Specific Uptake of Extracellular Vesicles Secreted by Urological Cancer Cells. Cancers, 13.","DOI":"10.3390\/cancers13194937"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"27","DOI":"10.1016\/j.ejpb.2019.10.005","article-title":"Cancer cell-type tropism is one of crucial determinants for the efficient systemic delivery of cancer cell-derived exosomes to tumor tissues","volume":"145","author":"Emam","year":"2019","journal-title":"Eur. J. Pharm. Biopharm."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"101259","DOI":"10.1016\/j.omtm.2024.101259","article-title":"Bioengineering extracellular vesicle cargo for optimal therapeutic efficiency","volume":"32","author":"Parks","year":"2024","journal-title":"Mol. Ther. Methods Clin. Dev."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"35","DOI":"10.1016\/j.jconrel.2014.11.029","article-title":"Active loading into extracellular vesicles significantly improves the cellular uptake and photodynamic effect of porphyrins","volume":"205","author":"Fuhrmann","year":"2015","journal-title":"J. Control. Release"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"102096","DOI":"10.1016\/j.jsps.2024.102096","article-title":"Human plasma derived exosomes: Impact of active and passive drug loading approaches on drug delivery","volume":"32","author":"Gul","year":"2024","journal-title":"Saudi Pharm. J."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Chandler, K., Millar, J., Ward, G., Boyall, C., White, T., Ready, J.D., Maani, R., Chapple, K., Tempest, R., and Brealey, J. (2023). Imaging of Light-Enhanced Extracellular Vesicle-Mediated Delivery of Oxaliplatin to Colorectal Cancer Cells via Laser Ablation, Inductively Coupled Plasma Mass Spectrometry. Cells, 13.","DOI":"10.3390\/cells13010024"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1256","DOI":"10.1039\/c5mt00028a","article-title":"Tumor microenvironment in focus: LA-ICP-MS bioimaging of a preclinical tumor model upon treatment with platinum(IV)-based anticancer agents","volume":"7","author":"Theiner","year":"2015","journal-title":"Metallomics"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"727","DOI":"10.1080\/10837450.2024.2384448","article-title":"Extracellular vesicle-based formulation of doxorubicin: Drug loading optimization, characterization, and cytotoxicity evaluation in tumor spheroids","volume":"29","author":"Mehryab","year":"2024","journal-title":"Pharm. Dev. Technol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"5","DOI":"10.1186\/s12645-023-00242-2","article-title":"Anticancer platinum-drug delivered by mesenchymal stromal cells improves its activity on glioblastoma","volume":"15","author":"Martegani","year":"2024","journal-title":"Cancer Nano"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"e12433","DOI":"10.1002\/jev2.12433","article-title":"Extracellular vesicles as human therapeutics: A scoping review of the literature","volume":"13","author":"Fusco","year":"2024","journal-title":"J. Extracell. Vesicles"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"1752","DOI":"10.1016\/j.immuni.2024.07.009","article-title":"The biology and function of extracellular vesicles in immune response and immunity","volume":"57","author":"Kalluri","year":"2024","journal-title":"Immunity"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"e2403199","DOI":"10.1002\/adma.202403199","article-title":"Immunogenicity of Extracellular Vesicles","volume":"36","author":"Xia","year":"2024","journal-title":"Adv. Mater."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"10","DOI":"10.1186\/1479-5876-3-10","article-title":"Vaccination of metastatic melanoma patients with autologous dendritic cell (DC) derived-exosomes: Results of thefirst phase I clinical trial","volume":"3","author":"Escudier","year":"2005","journal-title":"J. Transl. Med."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"782","DOI":"10.1038\/mt.2008.1","article-title":"Phase I clinical trial of autologous ascites-derived exosomes combined with GM-CSF for colorectal cancer","volume":"16","author":"Dai","year":"2008","journal-title":"Mol. Ther."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e12140","DOI":"10.1002\/jev2.12140","article-title":"Formation of a protein corona on the surface of extracellular vesicles in blood plasma","volume":"10","author":"Visnovitz","year":"2021","journal-title":"J. Extracell. Vesicles"},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Guarnerio, S., Tempest, R., Maani, R., Hunt, S., Cole, L.M., Le Maitre, C.L., Chapple, K., and Peake, N. (2023). Cellular Responses to Extracellular Vesicles as Potential Markers of Colorectal Cancer Progression. Int. J. Mol. Sci., 24.","DOI":"10.3390\/ijms242316755"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"446","DOI":"10.1002\/1096-9896(2000)9999:9999<::AID-PATH775>3.0.CO;2-K","article-title":"Validation of a model of colon cancer progression","volume":"192","author":"Hewitt","year":"2000","journal-title":"J. Pathol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1032","DOI":"10.1016\/j.jtha.2023.01.007","article-title":"Acidification of blood plasma facilitates the separation and analysis of extracellular vesicles","volume":"21","author":"Khamari","year":"2023","journal-title":"J. Thromb. Haemost."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"e2307240","DOI":"10.1002\/smll.202307240","article-title":"Chemically-Induced Lipoprotein Breakdown for Improved Extracellular Vesicle Purification","volume":"20","author":"Iannotta","year":"2024","journal-title":"Small"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"299","DOI":"10.1002\/hcs2.70028","article-title":"Exploring Regulatory Frameworks for Exosome Therapy: Insights and Perspectives","volume":"4","author":"Li","year":"2025","journal-title":"Health Care Sci."},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Kong, Y., Li, J., Zhao, X., Wu, Y., and Chen, L. (2025). CAR-T cell therapy: Developments, challenges and expanded applications from cancer to autoimmunity. Front. Immunol., 15.","DOI":"10.3389\/fimmu.2024.1519671"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"452","DOI":"10.1007\/BF01372550","article-title":"Treatment of advanced colorectal cancer with doxorubicin combined with two potential multidrug-resistance-reversing agents: High-dose oral tamoxifen and dexverapamil","volume":"123","author":"Kornek","year":"1997","journal-title":"J. Cancer Res. Clin. Oncol."},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Mas-Bargues, C., and Alique, M. (2023). Extracellular Vesicles as \u201cVery Important Particles\u201d (VIPs) in Aging. Int. J. Mol. Sci., 24.","DOI":"10.3390\/ijms24044250"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"158","DOI":"10.1186\/s12964-020-00630-w","article-title":"Inhibition of \u03b1v\u03b23 integrin impairs adhesion and uptake of tumor-derived small extracellular vesicles","volume":"18","author":"Altei","year":"2020","journal-title":"Cell Commun. Signal"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1274","DOI":"10.1089\/ten.tea.2017.0158","article-title":"A Systematic Evaluation of Factors Affecting Extracellular Vesicle Uptake by Breast Cancer Cells","volume":"23","author":"Stranford","year":"2017","journal-title":"Tissue Eng. Part A"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"e70017","DOI":"10.1002\/jex2.70017","article-title":"Mechanisms of extracellular vesicle uptake and implications for the design of cancer therapeutics","volume":"3","author":"Flemming","year":"2024","journal-title":"J. Extracell. Biol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"e12260","DOI":"10.1002\/jev2.12260","article-title":"Understanding the extracellular vesicle surface for clinical molecular biology","volume":"11","author":"Hallal","year":"2022","journal-title":"J. Extracell. Vesicles"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"563","DOI":"10.1039\/C9NA00537D","article-title":"Isolation methods for particle protein corona complexes from protein-rich matrices","volume":"2","author":"Stock","year":"2020","journal-title":"Nanoscale Adv."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"102121","DOI":"10.1016\/j.tranon.2024.102121","article-title":"A comprehensive review on recent advances in exosome isolation and characterization: Toward clinical applications","volume":"50","author":"Dilsiz","year":"2024","journal-title":"Transl. Oncol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"19861","DOI":"10.3402\/jev.v2i0.19861","article-title":"How pure are your vesicles?","volume":"2","author":"Webber","year":"2013","journal-title":"J. Extracell. Vesicles"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"44386","DOI":"10.1021\/acsami.4c03869","article-title":"Improving the Purity of Extracellular Vesicles by Removal of Lipoproteins from Size Exclusion Chromatography- and Ultracentrifugation-Processed Samples Using Glycosaminoglycan-Functionalized Magnetic Beads","volume":"16","author":"Chou","year":"2024","journal-title":"ACS Appl. Mater. Interfaces"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"1372","DOI":"10.1016\/j.jtha.2024.01.010","article-title":"Density-based lipoprotein depletion improves extracellular vesicle isolation and functional analysis","volume":"22","author":"Merij","year":"2024","journal-title":"J. Thromb. Haemost."}],"container-title":["Pharmaceutics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4923\/18\/6\/740\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,17]],"date-time":"2026-06-17T06:05:37Z","timestamp":1781676337000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4923\/18\/6\/740"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,6,15]]},"references-count":43,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2026,6]]}},"alternative-id":["pharmaceutics18060740"],"URL":"https:\/\/doi.org\/10.3390\/pharmaceutics18060740","relation":{},"ISSN":["1999-4923"],"issn-type":[{"value":"1999-4923","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,6,15]]}}}