A modular platform for targeted RNAi therapeutics

Article metrics

Abstract

Previous studies have identified relevant genes and signalling pathways that are hampered in human disorders as potential candidates for therapeutics. Developing nucleic acid-based tools to manipulate gene expression, such as short interfering RNAs1,2,3 (siRNAs), opens up opportunities for personalized medicine. Yet, although major progress has been made in developing siRNA targeted delivery carriers, mainly by utilizing monoclonal antibodies (mAbs) for targeting4,5,6,7,8, their clinical translation has not occurred. This is in part because of the massive development and production requirements and the high batch-to-batch variability of current technologies, which rely on chemical conjugation. Here we present a self-assembled modular platform that enables the construction of a theoretically unlimited repertoire of siRNA targeted carriers. The self-assembly of the platform is based on a membrane-anchored lipoprotein that is incorporated into siRNA-loaded lipid nanoparticles that interact with the antibody crystallizable fragment (Fc) domain. We show that a simple switch of eight different mAbs redirects the specific uptake of siRNAs by diverse leukocyte subsets in vivo. The therapeutic potential of the platform is demonstrated in an inflammatory bowel disease model by targeting colon macrophages to reduce inflammatory symptoms, and in a Mantle Cell Lymphoma xenograft model by targeting cancer cells to induce cell death and improve survival. This modular delivery platform represents a milestone in the development of precision medicine.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: ASSET platform design and construction.
Fig. 2: TsiLNP functionality in vitro and in vivo.
Fig. 3: In vivo targeting of TsiLNPs.
Fig. 4: TsiLNP-mediated therapeutic gene silencing in a murine colitis model.

References

  1. 1.

    Dahlman, J. E. et al. In vivo endothelial siRNA delivery using polymeric nanoparticles with low molecular weight. Nat. Nanotechnol. 9, 648–655 (2014).

  2. 2.

    McNamara, J. O. II et al. Cell type-specific delivery of siRNAs with aptamer-siRNA chimeras. Nat. Biotechnol. 24, 1005–1015 (2006).

  3. 3.

    Wittrup, A. & Lieberman, J. Knocking down disease: a progress report on siRNA therapeutics. Nat. Rev. Genet. 16, 543–552 (2015).

  4. 4.

    Ramishetti, S. et al. Systemic gene silencing in primary T lymphocytes using targeted lipid nanoparticles. ACS Nano 9, 6706–6716 (2015).

  5. 5.

    Weinstein, S. et al. Harnessing RNAi-based nanomedicines for therapeutic gene silencing in B-cell malignancies. Proc. Natl Acad. Sci. USA 113, E16–E22 (2016).

  6. 6.

    Katakowski, J. A. et al. Delivery of siRNAs to dendritic cells using DEC205-targeted lipid nanoparticles to inhibit immune responses. Mol. Ther. 24, 146–155 (2016).

  7. 7.

    Peer, D., Zhu, P., Carman, C. V., Lieberman, J. & Shimaoka, M. Selective gene silencing in activated leukocytes by targeting siRNAs to the integrin lymphocyte function-associated antigen-1. Proc. Natl Acad. Sci. USA 104, 4095–4100 (2007).

  8. 8.

    Song, E. et al. Antibody mediated in vivo delivery of small interfering RNAs via cell-surface receptors. Nat. Biotechnol. 23, 709–717 (2005).

  9. 9.

    Springer, T. A., Bhattacharya, A., Cardoza, J. T. & Sanchez-Madrid, F. Monoclonal antibodies specific for rat IgG1, IgG2a, and IgG2b subclasses, and kappa chain monotypic and allotypic determinants: reagents for use with rat monoclonal antibodies. Hybridoma 1, 257–273 (1982).

  10. 10.

    Laukkanen, M. L., Teeri, T. T. & Keinanen, K. Lipid-tagged antibodies: bacterial expression and characterization of a lipoprotein-single-chain antibody fusion protein. Protein Eng. 6, 449–454 (1993).

  11. 11.

    de Kruif, J., Storm, G., van Bloois, L. & Logtenberg, T. Biosynthetically lipid-modified human scFv fragments from phage display libraries as targeting molecules for immunoliposomes. FEBS Lett. 399, 232–236 (1996).

  12. 12.

    Semple, S. C. et al. Rational design of cationic lipids for siRNA delivery. Nat. Biotechnol. 28, 172–176 (2010).

  13. 13.

    Cohen, Z. R. et al. Localized RNAi therapeutics of chemoresistant grade IV glioma using hyaluronan-grafted lipid-based nanoparticles. ACS Nano. 9, 1581–1591 (2015).

  14. 14.

    Tiisala, S., Paavonen, T. & Renkonen, R. Alpha E beta 7 and alpha 4 beta 7 integrins associated with intraepithelial and mucosal homing, are expressed on macrophages. Eur. J. Immunol. 25, 411–417 (1995).

  15. 15.

    Peer, D., Park, E. J., Morishita, Y., Carman, C. V. & Shimaoka, M. Systemic leukocyte-directed siRNA delivery revealing cyclin D1 as an anti-inflammatory target. Science 319, 627–630 (2008).

  16. 16.

    Zigmond, E. et al. Ly6C hi monocytes in the inflamed colon give rise to proinflammatory effector cells and migratory antigen-presenting cells. Immunity 37, 1076–1090 (2012).

  17. 17.

    Grainger, J. R. et al. Inflammatory monocytes regulate pathologic responses to commensals during acute gastrointestinal infection. Nat. Med. 19, 713–721 (2013).

  18. 18.

    Benhar, I. & Pastan, I. Cloning, expression and characterization of the Fv fragments of the anti-carbohydrate mAbs Bl and B5 as single-chain immunotoxins. Protein Eng. 7, 1509–1515 (1994).

  19. 19.

    Vaks, L. & Benhar, I. Production of stabilized scFv antibody fragments in the E. coli bacterial cytoplasm. Methods Mol. Biol. 1060, 171–184 (2014).

Download references

Acknowledgements

This work was supported in part by grants from the Dotan Hemato-oncology Center at Tel Aviv University, by The Leona M. and Harry B. Helmsley Nanotechnology Research Fund, by the Kenneth Rainin Foundation and by the ERC grant LeukoTheranostics (number 647410) awarded to D.P.

Author information

R.K., N.V. and D.P. conceived and designed the project. R.K., N.V. S.R., M.G., D.R. N.D., L.N., I.H.-H., S.L.-B.-A. and M.H. performed the experimental work. R.K., N.V., M.B., I.B. J.L. and D.P. analysed the data. R.K. N.V. and D.P. wrote the manuscript. All authors discussed the results.

Correspondence to Dan Peer.

Ethics declarations

Competing interests

M.B. is an employee of Integrated DNA Technologies, Inc. J.L. is on the Scientific Advisory Board of Alnylam Pharmaceuticals. D.P. declares financial interests in Quiet Therapeutics. The rest of the authors declare no competing financial interests.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Figures

Supplementary Figures 1–6.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Kedmi, R., Veiga, N., Ramishetti, S. et al. A modular platform for targeted RNAi therapeutics. Nature Nanotech 13, 214–219 (2018) doi:10.1038/s41565-017-0043-5

Download citation

Further reading