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Membrane radiolabelling of exosomes for comparative biodistribution analysis in immunocompetent and immunodeficient mice – a novel and universal approach

Faruqu, Farid N., Wang, Julie Tzu-Wen, Xu, Lizhou, McNickle, Luke, Chong, Eden Ming-Yiu, Walters, Adam, Gurney, Mark, Clayton, Aled, Smyth, Lesley A., Hider, Robert, Sosabowski, Jane and Al-Jamal, Khuloud 2019. Membrane radiolabelling of exosomes for comparative biodistribution analysis in immunocompetent and immunodeficient mice – a novel and universal approach. Theranostics 9 (6) , pp. 1666-1682. 10.7150/thno.27891

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Abstract

Extracellular vesicles, in particular exosomes, have recently gained interest as novel drug delivery vectors due to their biological origin and inherent intercellular biomolecule delivery capability. An in-depth knowledge of their in vivo biodistribution is therefore essential. This work aimed to develop a novel, reliable and universal method to radiolabel exosomes to study their in vivo biodistribution. Methods: Melanoma (B16F10) cells were cultured in bioreactor flasks to increase exosome yield. B16F10-derived exosomes (ExoB₁₆) were isolated using ultracentrfugation onto a single sucrose cushion, and were characterised for size, yield, purity, exosomal markers and morphology using Nanoparticle Tracking Analysis (NTA), protein measurements, flow cytometry and electron microscopy. ExoB₁₆ were radiolabelled using 2 different approaches – intraluminal labelling (entrapment of ¹¹¹Indium via tropolone shuttling); and membrane labelling (chelation of ¹¹¹Indium via covalently attached bifunctional chelator DTPA-anhydride). Labelling efficiency and stability was assessed using gel filtration and thin layer chromatography. Melanoma-bearing immunocompetent (C57BL/6) and immunodeficient (NSG) mice were injected intravenously with radiolabelled ExoB₁₆ (1x10¹¹ particles/mouse) followed by metabolic cages study, whole body SPECT-CT imaging and ex vivo gamma counting at 1, 4 and 24 h post-injection. Results: Membrane-labelled ExoB₁₆ showed superior radiolabelling efficiency and radiochemical stability (19.2 ± 4.53 % and 80.4 ± 1.6 % respectively) compared to the intraluminal-labelled exosomes (4.73 ± 0.39 % and 14.21 ± 2.76 % respectively). Using the membrane-labelling approach, the in vivo biodistribution of ExoB₁₆ in melanoma-bearing C57Bl/6 mice was carried out, and was found to accumulate primarily in the liver and spleen (~56% and ~38% ID/gT respectively), followed by the kidneys (~3% ID/gT). ExoB₁₆ showed minimal tumour i.e. self-tissue accumulation (~0.7% ID/gT). The membrane-labelling approach was also used to study ExoB₁₆ biodistribution in melanoma-bearing immunocompromised (NSG) mice, to compare with that in the immunocompetent C57Bl/6 mice. Similar biodistribution profile was observed in both C57BL/6 and NSG mice, where prominent accumulation was seen in liver and spleen, apart from the significantly lower tumour accumulation observed in the NSG mice (~0.3% ID/gT). Conclusion: Membrane radiolabelling of exosomes is a reliable approach that allows for accurate live imaging and quantitative biodistribution studies to be performed on potentially all exosome types without engineering parent cells.

Item Type: Article
Date Type: Published Online
Status: Published
Schools: Medicine
Publisher: Ivyspring International Publisher
ISSN: 1838-7640
Date of First Compliant Deposit: 28 January 2019
Date of Acceptance: 8 November 2018
Last Modified: 03 Jul 2019 15:19
URI: http://orca.cf.ac.uk/id/eprint/118875

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