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Biodegradable silicon nanoneedles delivering nucleic acids intracellularly induce localized in vivo neovascularization

Nature Materials volume 14, pages 532–539 (2015)Cite this article

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Abstract

The controlled delivery of nucleic acids to selected tissues remains an inefficient process mired by low transfection efficacy, poor scalability because of varying efficiency with cell type and location, and questionable safety as a result of toxicity issues arising from the typical materials and procedures employed. High efficiency and minimal toxicity in vitro has been shown for intracellular delivery of nuclei acids by using nanoneedles, yet extending these characteristics to in vivo delivery has been difficult, as current interfacing strategies rely on complex equipment or active cell internalization through prolonged interfacing. Here, we show that a tunable array of biodegradable nanoneedles fabricated by metal-assisted chemical etching of silicon can access the cytosol to co-deliver DNA and siRNA with an efficiency greater than 90%, and that in vivo the nanoneedles transfect the VEGF-165 gene, inducing sustained neovascularization and a localized sixfold increase in blood perfusion in a target region of the muscle.

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Figure 1: Porous silicon nanoneedles.
Figure 2: Cell interfacing, cytocompatibility and biodegradation.
Figure 3: Intracellular co-delivery of nucleic acids.
Figure 4: Nanoneedles mediate in vivo delivery.
Figure 5: Safety profile of nanoinjection.
Figure 6: Nanoneedles mediate neovascularization.

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References

  1. Leader, B., Baca, Q. J. & Golan, D. E. Protein therapeutics: A summary and pharmacological classification. Nature Rev. Drug Discov. 7, 21–39 (2008).

    Article  CAS  Google Scholar 

  2. Vlieghe, P., Lisowski, V., Martinez, J. & Khrestchatisky, M. Synthetic therapeutic peptides: Science and market. Drug Discov. Today 15, 40–56 (2010).

    Article  CAS  Google Scholar 

  3. Kole, R., Krainer, A. R. & Altman, S. RNA therapeutics: Beyond RNA interference and antisense oligonucleotides. Nature Rev. Drug Discov. 11, 125–140 (2012).

    Article  CAS  Google Scholar 

  4. Khademhosseini, A. Microscale technologies for tissue engineering and biology. Proc. Natl Acad. Sci. USA 103, 2480–2487 (2006).

    Article  CAS  Google Scholar 

  5. Whitehead, K. A., Langer, R. & Anderson, D. G. Knocking down barriers: Advances in siRNA delivery. Nature Rev. Drug Discov. 8, 129–138 (2009).

    Article  CAS  Google Scholar 

  6. Saltzman, W. M. & Luo, D. Synthetic DNA delivery systems. Nature Biotechnol. 18, 33–37 (2000).

    Article  Google Scholar 

  7. Thomas, C. E., Ehrhardt, A. & Kay, M. A. Progress and problems with the use of viral vectors for gene therapy. Nature Rev. Genet. 4, 346–358 (2003).

    Article  CAS  Google Scholar 

  8. Mingozzi, F. & High, K. A. Therapeutic in vivo gene transfer for genetic disease using AAV: Progress and challenges. Nature Rev. Genet. 12, 341–355 (2011).

    Article  CAS  Google Scholar 

  9. Lefesvre, P., Attema, J. & van Bekkum, D. A comparison of efficacy and toxicity between electroporation and adenoviral gene transfer. BMC Mol. Biol. 3, 12 (2002).

    Article  Google Scholar 

  10. Aihara, H. & Miyazaki, J-I. Gene transfer into muscle by electroporation in vivo. Nature Biotechnol. 16, 867–870 (1998).

    Article  CAS  Google Scholar 

  11. Nishikawa, M. & Huang, L. Nonviral vectors in the new millennium: Delivery barriers in gene transfer. Hum. Gene Ther. 12, 861–870 (2001).

    Article  CAS  Google Scholar 

  12. Shalek, A. K. et al. Vertical silicon nanowires as a universal platform for delivering biomolecules into living cells. Proc. Natl Acad. Sci. USA 107, 1870–1875 (2010).

    Article  CAS  Google Scholar 

  13. Robinson, J. T. et al. Vertical nanowire electrode arrays as a scalable platform for intracellular interfacing to neuronal circuits. Nature Nanotech. 7, 180–184 (2012).

    Article  CAS  Google Scholar 

  14. Yan, R. et al. Nanowire-based single-cell endoscopy. Nature Nanotech. 7, 191–196 (2011).

    Article  Google Scholar 

  15. Han, S. W. et al. High-efficiency DNA injection into a single human mesenchymal stem cell using a nanoneedle and atomic force microscopy. Nanomedicine: Nanotechnol. Biol. Med. 4, 215–225 (2008).

    Article  CAS  Google Scholar 

  16. Shalek, A. K. et al. Nanowire-mediated delivery enables functional interrogation of primary immune cells: Application to the analysis of chronic lymphocytic leukemia. Nano Lett. 12, 6498–6504 (2012).

    Article  CAS  Google Scholar 

  17. Xu, A. M. et al. Quantification of nanowire penetration into living cells. Nature Commun. 5, 3613 (2014).

    Article  Google Scholar 

  18. Sharei, A. et al. A vector-free microfluidic platform for intracellular delivery. Proc. Natl Acad. Sci. USA 110, 2082–2087 (2013).

    Article  CAS  Google Scholar 

  19. Hanson, L., Lin, Z. C., Xie, C., Cui, Y. & Cui, B. Characterization of the cell–nanopillar interface by transmission electron microscopy. Nano Lett. 12, 5815–5820 (2012).

    Article  CAS  Google Scholar 

  20. Xie, X. et al. Mechanical model of vertical nanowire cell penetration. Nano Lett. 13, 6002–6008 (2013).

    Article  CAS  Google Scholar 

  21. Xie, C., Lin, Z., Hanson, L., Cui, Y. & Cui, B. Intracellular recording of action potentials by nanopillar electroporation. Nature Nanotech. 7, 185–190 (2012).

    Article  CAS  Google Scholar 

  22. Wang, Y. et al. Poking cells for efficient vector-free intracellular delivery. Nature Commun. 5, 4466 (2014).

    Article  CAS  Google Scholar 

  23. Anderson, S. H. C., Elliott, H., Wallis, D. J., Canham, L. T. & Powell, J. J. Dissolution of different forms of partially porous silicon wafers under simulated physiological conditions. Phys. Status Solidi A 197, 331–335 (2003).

    Article  CAS  Google Scholar 

  24. Canham, L. T. Bioactive silicon structure fabrication through nanoetching techniques. Adv. Mater. 7, 1033–1037 (1995).

    Article  CAS  Google Scholar 

  25. Chiappini, C. et al. Tailored porous silicon microparticles: Fabrication and properties. ChemPhysChem 11, 1029–1035 (2010).

    Article  CAS  Google Scholar 

  26. Tasciotti, E. et al. Mesoporous silicon particles as a multistage delivery system for imaging and therapeutic applications. Nature Nanotech. 3, 151–157 (2008).

    Article  CAS  Google Scholar 

  27. Tanaka, T. et al. Sustained small interfering RNA delivery by mesoporous silicon particles. Cancer Res. 70, 3687–3696 (2010).

    Article  CAS  Google Scholar 

  28. Park, J-H. et al. Biodegradable luminescent porous silicon nanoparticles for in vivo applications. Nature Mater. 8, 331–336 (2009).

    Article  CAS  Google Scholar 

  29. Goh, A. S-W. et al. A novel approach to brachytherapy in hepatocellular carcinoma using a phosphorous32 (32P) brachytherapy delivery device—a first-in-man study. Int. J. Radiat. Oncol. Biol. Phys. 67, 786–792 (2007).

    Article  CAS  Google Scholar 

  30. Anglin, E., Cheng, L., Freeman, W. & Sailor, M. Porous silicon in drug delivery devices and materials. Adv. Drug Deliv. Rev. 60, 1266–1277 (2008).

    Article  CAS  Google Scholar 

  31. Chiappini, C., Liu, X., Fakhoury, J. R. & Ferrari, M. Biodegradable porous silicon barcode nanowires with defined geometry. Adv. Funct. Mater. 20, 2231–2239 (2010).

    Article  CAS  Google Scholar 

  32. Obataya, I., Nakamura, C., Han, S., Nakamura, N. & Miyake, J. Mechanical sensing of the penetration of various nanoneedles into a living cell using atomic force microscopy. Biosens. Bioelectron. 20, 1652–1655 (2005).

    Article  CAS  Google Scholar 

  33. Na, Y-R. et al. Probing enzymatic activity inside living cells using a nanowire–cell ‘sandwich’ assay. Nano Lett. 13, 153–158 (2013).

    Article  CAS  Google Scholar 

  34. Muralidharan-Chari, V., Clancy, J. W., Sedgwick, A. & D’Souza-Schorey, C. Microvesicles: Mediators of extracellular communication during cancer progression. J. Cell Sci. 123, 1603–1611 (2010).

    Article  CAS  Google Scholar 

  35. Ratajczak, J. et al. Embryonic stem cell-derived microvesicles reprogram hematopoietic progenitors: Evidence for horizontal transfer of mRNA and protein delivery. Leukemia 20, 847–856 (2006).

    Article  CAS  Google Scholar 

  36. Yuan, A. et al. Transfer of microRNAs by embryonic stem cell microvesicles. PLoS ONE 4, e4722 (2009).

    Article  Google Scholar 

  37. Shen, Z. et al. Porous silicon as a versatile platform for laser desorption/ionization mass spectrometry. Anal. Chem. 73, 612–619 (2000).

    Article  Google Scholar 

  38. Lee, J. W., Park, J-H. & Prausnitz, M. R. Dissolving microneedles for transdermal drug delivery. Biomaterials 29, 2113–2124 (2008).

    Article  CAS  Google Scholar 

  39. Gross, S. et al. Bioluminescence imaging of myeloperoxidase activity in vivo. Nature Med. 15, 455–461 (2009).

    Article  CAS  Google Scholar 

  40. Luxembourg, A., Evans, C. F. & Hannaman, D. Electroporation-based DNA immunisation: Translation to the clinic. Expert Opin. Biol. Ther. 7, 1647–1664 (2007).

    Article  CAS  Google Scholar 

  41. Yancopoulos, G. D. et al. Vascular-specific growth factors and blood vessel formation. Nature 407, 242–248 (2000).

    Article  CAS  Google Scholar 

  42. Tammela, T., Enholm, B., Alitalo, K. & Paavonen, K. The biology of vascular endothelial growth factors. Cardiovasc. Res. 65, 550–563 (2005).

    Article  CAS  Google Scholar 

  43. DeMuth, P. C. et al. Polymer multilayer tattooing for enhanced DNA vaccination. Nature Mater. 12, 367–376 (2013).

    Article  CAS  Google Scholar 

  44. Kim, Y-C., Park, J-H. & Prausnitz, M. R. Microneedles for drug and vaccine delivery. Adv. Drug Deliv. Rev. 64, 1547–1568 (2012).

    Article  CAS  Google Scholar 

  45. Gershonowitz, A. & Gat, A. VoluDerm Microneedle Technology for Skin Treatments—In Vivo Histological Evidence. J. Cosmetic Laser Surg. 17, 9–14 (2015).

    Article  Google Scholar 

  46. Barrientos, S., Stojadinovic, O., Golinko, M. S., Brem, H. & Tomic-Canic, M. Growth factors and cytokines in wound healing. Wound Repair Regen. 16, 585–601 (2008).

    Article  Google Scholar 

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Acknowledgements

The authors would like to thank M. Giacca at the International Centre for Genetic Engineering and Biotechnology (ICGEB in Trieste, Italy) for kindly donating the pDNA-expressing VEGF-165, and I. K. Yazdi and R. Palomba for the amplification and purification of the plasmids used in the study. We are grateful to S. Amra for histology-slide preparation, D. Tinkey for surgical procedures, S. Zacchigna for suggestions on PCR protocols, P. Campagnolo for advice on SMA and isolectin staining, and S. T. Gammon for advice on bioluminescence imaging with luminol. This work was financially supported by: the US Department of Defense (W81XWH-12-10414) and the NIH (1R21CA173579-01A1 and 5U54CA143837); C.C. was supported by a Newton International Fellowship and a Marie Curie International Incoming Fellowship; J.O.M. was supported by a NIH pre-doctoral fellowship 5F31CA154119-02. M.M.S. holds an ERC grant ‘Naturale-CG’ and is supported by a Wellcome Trust Senior Investigator Award. E.T. holds grants from the Hearst Foundation and the Cullen Trust Foundation. C.C. and M.M.S. thank the Rosetrees Trust and the Stoneygate Trust for funding.

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Author notes

  1. C. Chiappini and E. De Rosa: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Materials, Imperial College London, London, SW6 7PB, UK

    C. Chiappini, J. Steele & M. M. Stevens

  2. Department of Bioengineering and Institute of Biomedical Engineering, Imperial College London, London, SW6 7PB, UK

    C. Chiappini, J. Steele & M. M. Stevens

  3. Department of Nanomedicine, Houston Methodist Research Institute, Houston, Texas 77030, USA

    E. De Rosa, J. O. Martinez, X. Liu & E. Tasciotti

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  1. C. Chiappini
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Contributions

C.C., E.T. and M.M.S. designed the research. C.C. and E.T. conceived the nanoneedles; C.C. developed the nanoneedles with contributions from X.L.; C.C. evaluated loading, release, delivery and efficacy with contributions from E.D.R.; C.C. imaged biodegradation and cell interaction with contributions from E.D.R.; C.C. wrote the initial manuscript with contributions from E.D.R. and J.O.M. C.C. performed all electron microscopy analysis. E.D.R. and J.O.M. assessed cytocompatibility; E.D.R. designed and performed animal surgeries, intravital imaging and quantification of vascularization, vasculature pattern, and extravasation and flow rate over time on VEGF treatment, with contributions from J.O.M. J.O.M. evaluated degradation by ICP-AES, fluorescent and bioluminescent imaging and analysis, real-time PCR, and histological evaluation and staining of tissues with contributions from E.D.R. J.S. performed compressive mechanical testing. E.T. and M.M.S. contributed equally to the work, with M.M.S. supervising the in vitro studies and E.T. the in vivo work. All authors discussed and commented on the manuscript.

Corresponding authors

Correspondence to M. M. Stevens or E. Tasciotti.

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Chiappini, C., De Rosa, E., Martinez, J. et al. Biodegradable silicon nanoneedles delivering nucleic acids intracellularly induce localized in vivo neovascularization. Nature Mater 14, 532–539 (2015). https://doi.org/10.1038/nmat4249

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