Abstract
To increase the half-life of a cytokine and target its activation specifically to disease sites, we have engineered a latent cytokine using the latency-associated protein (LAP) of transforming growth factor-β1 (TGF-β1) fused via a matrix metalloproteinase (MMP) cleavage site to interferon (IFN)-β at either its N or C terminus. The configuration LAP-MMP-IFN-β resembles native TGF-β and lacks biological activity until cleaved by MMPs, whereas the configuration IFN-β-MMP-LAP is active. LAP provides for a disulfide-linked shell hindering interaction of the cytokine with its cellular receptors, conferring a very long half-life of 55 h in vivo. Mutations of the disulfide bonds in LAP abolish this latency. Samples of cerebrospinal fluid (CSF) or synovial fluid from patients with inflammatory diseases specifically activate the latent cytokine, whereas serum samples do not. Intramuscular injection in arthritic mice of plasmid DNA encoding these constructs demonstrated a greater therapeutic effect of the latent as compared to the active forms.
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Notes
*Note: In Figure 3b, the zero time point for IFN-LAP + RASF was omitted. This mistake has been corrected in the HTML version and will appear correctly in print. The correction has been appended to the PDF version available online.
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Acknowledgements
This work was funded by the Multiple Sclerosis Society of Great Britain and Northern Ireland, the Arthritis Research Campaign (UK) and Kinetique Biomedical Research Seed Fund UK. We thank Irene Theoharidou for her expert secretarial assistance, G. Scott and G. Giovannoni for clinical samples, S. Amor for samples from rhesus monkeys, S. Mather and D. Ellison for help with protein iodination, J. Wilson and A. Mustafa for technical help, A. Johnston for the analysis of pharmacokinetic data, L. Layward, D. Willoughby, O.L. Podhajcer and P. Colville-Nash for reviewing the manuscript and H. Nagase for helpful and encouraging discussions.
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G.A., H.D. and Y.C. are shareholders in Stealtnyx Therapeutics, Ltd., which holds patents based on this technology.
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Adams, G., Vessillier, S., Dreja, H. et al. Targeting cytokines to inflammation sites. Nat Biotechnol 21, 1314–1320 (2003). https://doi.org/10.1038/nbt888
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DOI: https://doi.org/10.1038/nbt888
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