Functionalized Exosomes Reprogram the Targeted Recognition Between Immune Cells and Tumor Cells

doi:10.13523/j.cb.2305021

China Biotechnology

2023, Vol. 43

Issue (10): 1-9 DOI: 10.13523/j.cb.2305021

Functionalized Exosomes Reprogram the Targeted Recognition Between Immune Cells and Tumor Cells

XIANG Jian¹,YE Bang-ce¹,YIN Bin-cheng^1,^2,^**(

)

1 State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China
2 School of Chemistry and Chemical Engineering, Shihezi University, Shihezi 832003, China

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Abstract

Objective: To prepare the exosomes that display the vesicular stomatitis virus glycoprotein (VSVG) by utilizing genetic engineering technology, which promotes membrane fusion. Meanwhile, the exosomes were further modified with aptamers that target dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) receptors to prepare functionalized exosomes, which is achieved through DNA hybridization chain reaction mediated by nucleic acid probes. This approach reprograms the targeted recognition process between tumor cells and dendritic cells, ultimately enhancing their interaction. Methods: The paper focuses on mouse mammary tumor cells (4T1) and mouse dendritic cells (DC2.4) as research subjects, which demonstrates through confocal imaging and flow cytometry that VSVG-Exos can specifically bind to 4T1 cells via membrane fusion, and that DC-SIGN aptamers can specifically target DC2.4 cells. Results: Functionalized exosomes can reprogram and modify 4T1 cells to enhance their targeted recognition effect with DC2.4 cells. Conclusion: Functionalized exosomes can effectively deliver molecules with specific functions to the surface of tumor cells and reprogram and modify them, thus improving the ability of immune cells to accurately locate and efficiently attack tumor cells. This provides a new approach and strategy for targeted elimination of tumor cells.

Key words： Functionalized exosomes Targeted recognition Aptamers Membrane fusion Vesicular stomatitis virus glycoprotein

Received: 15 May 2023 Published: 02 November 2023

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XIANG Jian, YE Bang-ce, YIN Bin-cheng. Functionalized Exosomes Reprogram the Targeted Recognition Between Immune Cells and Tumor Cells. China Biotechnology, 2023, 43(10): 1-9.

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https://manu60.magtech.com.cn/biotech/10.13523/j.cb.2305021 OR https://manu60.magtech.com.cn/biotech/Y2023/V43/I10/1

Fig.1 Preparation of VSVG-Exos-Apt and its enhancement of interactions between DC2.4 and 4T1 cells The VSVG plasmid was transfected into HEK 293T cells; anchor DNA probe P on the surface of the isolated and obtained VSVG-Exos membrane; H1 and H2 mediate the HCR and ligate the DC-SIGN aptamer to obtain VSVG-Exos-Apt; VSVG-Exos-Apt binds 4T1 cells by membrane fusion and transfers VSVG protein and DC-SIGN aptamers to 4T1 cell membranes; 4T1 cells specifically target DC2.4 cells via DC-SIGN aptamers

Table 1 Oligonucleotide sequences used in this study

Fig.2 Preparation and characterization of VSVG-Exos (a) Confocal fluorescence microscopy images indicating the expression of VSVG in HEK 293T cells (b) Western blot analysis of the expression of VSVG in both the parental cells and derived exosomes indicated treatments (c) Expression levels of VSVG in HEK 293T cells detected by RT-qPCR. The data are expressed as mean ± SD (n = 3)

Fig.3 Functionalized and characterization of exosomes (a) The relationship between the incubation time of P-Chol and VSVG-Exos (b) The fluorescence intensity of VSVG-Exos corresponding to different amounts of added P-Chol (c) Feasibility verification of HCR cascade amplification reaction (d) Representative graphic and morphological structure chart of the particle number distribution (y-axis) and diameter (nm) (x-axis) of VSVG-Exos-Apt as revealed by the analysis of NTA and TEM images (e) Zeta potential measurement of VSVG-Exos-Apt, VSVG-Exos or Exos (f) The fluorescence intensity and number of FAM-P-Chol tethers on VSVG-Exos-Apt. The data are expressed as mean ± SD (n = 3). Statistical analysis between two sample groups was performed using student’s t-test

Fig.4 Verification of exosome fusion with 4T1 cell membrane (a) Flow cytometry and (b) confocal fluorescence microscopy images of membrane fusion verification with VSVG-Exos or Exos

Fig.5 In vitro targeting validation of aptamers (a, c) Flow cytometry and (b) confocal fluorescence microscopy images of functional verification of DC-SIGN aptamers (d) Flow cytometry of corresponding to different amounts of DC-SIGN aptamer additions

Fig.6 Reprogramming of 4T1 cells by functionalized exosomes (a) Schematic diagram of the HCR reaction (b) Flow cytometry and (c) confocal fluores cence microscopy images were used to validate VSVG-Exo-Apt and Exos-Apt mediated editing of 4T1 cells

Fig.7 Analysis of the interaction of 4T1 with DC2.4 cells (a) Flow cytometry and (b) confocal fluorescence microscopy images of interaction of 4T1 with DC2.4 cell


[1]	Zhang H M, Chen J B. Current status and future directions of cancer immunotherapy. Journal of Cancer, 2018, 9(10): 1773-1781. doi: 10.7150/jca.24577 pmid: 29805703

[2]	Hegde P S, Chen D S. Top 10 challenges in cancer immunotherapy. Immunity, 2020, 52(1): 17-35. doi: S1074-7613(19)30530-8 pmid: 31940268

[3]	Coulie P G, Van den Eynde B J, van der Bruggen P, et al. Tumour antigens recognized by T lymphocytes: at the core of cancer immunotherapy. Nature Reviews Cancer, 2014, 14(2): 135-146. doi: 10.1038/nrc3670 pmid: 24457417

[4]	Kim G B, Nam G H, Hong Y, et al. Xenogenization of tumor cells by fusogenic exosomes in tumor microenvironment ignites and propagates antitumor immunity. Science Advances, 2020, 6(27): eaaz2083.

[5]	Sadelain M, Rivière I, Riddell S. Therapeutic T cell engineering. Nature, 2017, 545(7655): 423-431. doi: 10.1038/nature22395

[6]	Maude S L, Frey N, Shaw P A, et al. Chimeric antigen receptor T cells for sustained remissions in leukemia. The New England Journal of Medicine, 2014, 371(16): 1507-1517. doi: 10.1056/NEJMoa1407222 pmid: 25317870

[7]	Jackson H J, Rafiq S, Brentjens R J. Driving CAR T-cells forward. Nature Reviews Clinical Oncology, 2016, 13(6): 370-383. doi: 10.1038/nrclinonc.2016.36 pmid: 27000958

[8]	Bonifant C L, Jackson H J, Brentjens R J, et al. Toxicity and management in CAR T-cell therapy. Molecular Therapy - Oncolytics, 2016, 3: 16011. doi: 10.1038/mto.2016.11

[9]	Zhang Y, Bi J Y, Huang J Y, et al. Exosome: a review of its classification, isolation techniques, storage, diagnostic and targeted therapy applications. International Journal of Nanomedicine, 2020, 15: 6917-6934. doi: 10.2147/IJN.S264498 pmid: 33061359

[10]	Gurung S, Perocheau D, Touramanidou L, et al. The exosome journey: from biogenesis to uptake and intracellular signalling. Cell Communication and Signaling, 2021, 19(1): 47. doi: 10.1186/s12964-021-00730-1 pmid: 33892745

[11]	Xu M, Feng T, Liu B W, et al. Engineered exosomes: desirable target-tracking characteristics for cerebrovascular and neurodegenerative disease therapies. Theranostics, 2021, 11(18): 8926-8944. doi: 10.7150/thno.62330 pmid: 34522219

[12]	Wang X Y, Huang J, Chen W J, et al. The updated role of exosomal proteins in the diagnosis, prognosis, and treatment of cancer. Experimental & Molecular Medicine, 2022, 54(9): 1390-1400.

[13]	Liang Y J, Duan L, Lu J P, et al. Engineering exosomes for targeted drug delivery. Theranostics, 2021, 11(7): 3183-3195. doi: 10.7150/thno.52570 pmid: 33537081

[14]	Yan H, Li S, Zhou L F, et al. Selection of DNA aptamers against DC-SIGN protein. Molecular and Cellular Biochemistry, 2007, 306(1-2): 71-77. pmid: 17660953

[15]	Yang Y, Hong Y, Nam G H, et al. Virus-mimetic fusogenic exosomes for direct delivery of integral membrane proteins to target cell membranes. Advanced Materials, 2017, 29(13): 1605604. doi: 10.1002/adma.v29.13

[16]	Zhao H, Xu J, Li Y, et al. Nanoscale coordination polymer based nanovaccine for tumor immunotherapy. ACS Nano, 2019, 13(11): 13127-13135. doi: 10.1021/acsnano.9b05974 pmid: 31710460

[17]	Valadi H, Ekström K, Bossios A, et al. Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells. Nature Cell Biology, 2007, 9(6): 654-659. doi: 10.1038/ncb1596 pmid: 17486113

[18]	Cho E, Nam G H, Hong Y, et al. Comparison of exosomes and ferritin protein nanocages for the delivery of membrane protein therapeutics. Journal of Controlled Release, 2018, 279: 326-335. doi: S0168-3659(18)30221-9 pmid: 29679665

[19]	Koh E, Lee E J, Nam G H, et al. Exosome-SIRPα, a CD47 blockade increases cancer cell phagocytosis. Biomaterials, 2017, 121: 121-129. doi: S0142-9612(17)30004-2 pmid: 28086180

[20]	Yang Y, Hong Y, Cho E, et al. Extracellular vesicles as a platform for membrane-associated therapeutic protein delivery. Journal of Extracellular Vesicles, 2018, 7(1): 1440131. doi: 10.1080/20013078.2018.1440131

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