Abstract
DNA origami is capable of spatially organizing molecules into sophisticated geometric patterns with nanometric precision. Here we describe a reconfigurable, two-dimensional DNA origami with geometrically patterned CD95 ligands that regulates immune cell signalling to alleviate rheumatoid arthritis. In response to pH changes, the device reversibly transforms from a closed to an open configuration, displaying a hexagonal pattern of CD95 ligands with ~10 nm intermolecular spacing, precisely mirroring the spatial arrangement of CD95 receptor clusters on the surface of immune cells. In a collagen-induced arthritis mouse model, DNA origami elicits robust and selective activation of CD95 death-inducing signalling in activated immune cells located in inflamed synovial tissues. Such localized immune tolerance ameliorates joint damage with no noticeable side effects. This device allows for the precise spatial control of cellular signalling, expanding our understanding of ligand–receptor interactions and is a promising platform for the development of pharmacological interventions targeting these interactions.
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Data availability
The data that support the findings of this study are available within the Article and its Supplementary Information files. Source data can be found on figshare at https://doi.org/10.6084/m9.figshare.25111502 (ref. 51). Source data are provided with this paper.
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Acknowledgements
We acknowledge financial support from the National University of Singapore (grant nos. NUHSRO/2020/133/Startup/08, NUHSRO/2023/008/NUSMed/TCE/LOA and NUHSRO/2021/034/TRP/09/Nanomedicine), National Medical Research Council (grant no. MOH-OFIRG23jan-0005), Singapore Ministry of Education (grant no. MOE-000387-00), National Research Foundation (grant no. NRF-000352-00), National Natural Science Foundation of China (grant nos. NSFC 82202311, 62288102, 82302356 and 22274081), Science Foundation of Fujian Normal University (grant no. Y07204080K13) and Natural Science Foundation of Jiangsu Province, Major Project (grant no. BK20212012). We thank H. Liu at the National University of Singapore for providing A20 cells. We thank X. Song at the West China School of Public Health and West China Fourth Hospital, Sichuan University for help with confocal microscopy imaging. We thank B. Zhou at the State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University for help with establishing the CIA model. We thank Guangzhou Sagene Biotech Co., Ltd. for help in making the pattern diagrams.
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X.C., J.C., Z.Y., L.L., J.Y., W.M. and L.T. conceived and designed the experiments. L.L., J.Y., W.M., L.T. and L.Y. performed experiments and wrote the manuscript. J.Z., T.D. and Y.Z. performed data analysis. X.C., J.C. and Z.Y. supervised the entire project. L.W. and C.F. conducted proofreading of the manuscript. All authors discussed the results and commented on the manuscript.
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Extended data
Extended Data Fig. 1 The spatial arrangement of the CD95L array directs the activation of CD95 signaling in Jurkat cells.
a, Clustering of CD95 receptors in Jurkat cells 2 h following treatment with NS-empty (G2), sCD95L (G3), NS-10 (G4), ND-10 in the culture medium (pH 7.4) (G5), ND-10 in the culture medium (pH 6.5) (G6). Untreated (G1); Scale bar, 5 μm. The experiment was performed three times with similar results. (b, c), Colocalization of CD95 with the endosomal marker-early endosome antigen (EEA1) following NS-10 stimulation (b) or ND-10 stimulation in the culture medium (pH 6.5) (c). Scale bar, 5 μm. The experiment was performed three times with similar results. d, Immunocytochemistry analysis of Cleaved Caspase-8 in Jurkat cells 6 h following treatment with NS-empty (G2), sCD95L (G3), NS-10 (G4), ND-10 in the culture medium (pH 7.4) (G5), ND-10 in the culture medium (pH 6.5) (G6). Untreated (G1); Scale bar, 5 μm. The experiment was performed three times with similar results.
Extended Data Fig. 2 DNA origami nanodevice exhibits conformational transition in response to pH trigger in vivo.
Representative in vivo fluorescence images of CIA mice after intravenous injection with pH-sensitive nanodevice with i-motif-based fasteners labeled with both Cy5.5 fluorophore and BBQ650 quencher (w/ Cy5.5 & BBQ650), pH-sensitive nanodevice with i-motif-based fasteners labeled with Cy5.5 fluorophore (w/ Cy5.5), or pH-non-sensitive nanodevice with the pH-non-sensitive fasteners labeled with both Cy5.5 fluorophore and BBQ650 quencher (w/ Cy5.5 & BBQ650).
Extended Data Fig. 3 Designer DNA origami shows minimal immunomodulation potential.
(a, b) Concentration of TNF-α, IL-6, and IL-1β in the supernatants of RAW 264.7 cells (a) and BMDMs (b) following ND-empty treatment at indicated concentration. LPS (lipopolysaccharide, Invitrogen) was used as a control. n = 3 biologically independent samples per group. Note that the cytokine concentrations in some samples were below the threshold of the assay range and they were determined by extrapolating from the standard curve. Data presented as means ± s.d. Statistical significance was calculated via One-way ANOVA with Tukey post-hoc test (a, b).
Extended Data Fig. 4 Designer DNA origami yields long-term therapeutic efficacy in CIA mice.
a, Schematic illustration of the long-term treatment regimen. b, Clinical arthritis scores of the CIA mice following indicated treatments. n = 12 biologically independent mice per group. c, The paw swelling measurement of the CIA mice following indicated treatments. n = 12 biologically independent mice per group. Data presented as means ± s.d. Statistical significance was calculated via One-way ANOVA with Tukey post-hoc test (b, c). The experiment was performed twice with similar results.
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DNA sequences of the staple strand pool and functional staple strands.
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Li, L., Yin, J., Ma, W. et al. A DNA origami device spatially controls CD95 signalling to induce immune tolerance in rheumatoid arthritis. Nat. Mater. (2024). https://doi.org/10.1038/s41563-024-01865-5
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DOI: https://doi.org/10.1038/s41563-024-01865-5
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