Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

In situ programming of leukaemia-specific T cells using synthetic DNA nanocarriers

Abstract

An emerging approach for treating cancer involves programming patient-derived T cells with genes encoding disease-specific chimeric antigen receptors (CARs), so that they can combat tumour cells once they are reinfused. Although trials of this therapy have produced impressive results, the in vitro methods they require to generate large numbers of tumour-specific T cells are too elaborate for widespread application to treat cancer patients. Here, we describe a method to quickly program circulating T cells with tumour-recognizing capabilities, thus avoiding these complications. Specifically, we demonstrate that DNA-carrying nanoparticles can efficiently introduce leukaemia-targeting CAR genes into T-cell nuclei, thereby bringing about long-term disease remission. These polymer nanoparticles are easy to manufacture in a stable form, which simplifies storage and reduces cost. Our technology may therefore provide a practical, broadly applicable treatment that can generate anti-tumour immunity ‘on demand’ for oncologists in a variety of settings.

Your institute does not have access to this article

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: Design and manufacture of lymphocyte-programming nanoparticles.
Figure 2: DNA nanocarriers choreograph robust and persistent CAR production by lymphocytes in vitro.
Figure 3: CD3-targeted nanoparticles bind to circulating T cells in mice.
Figure 4: Reprogramming host T cells with leukaemia-specific CAR genes.
Figure 5: Nanoparticle-programmed CAR lymphocytes can cause tumour regression with efficacies similar to adoptive T-cell therapy.

References

  1. Rosenberg, S. A. & Restifo, N. P. Adoptive cell transfer as personalized immunotherapy for human cancer. Science 348, 62–68 (2015).

    CAS  Article  Google Scholar 

  2. Plumridge, H. New costly cancer treatments face hurdles getting to patients. The Wall Street Journal (6 October 2014); https://www.wsj.com/articles/new-costly-cancer-treatments-face-hurdles-getting-to-patients-1412627150

  3. Mangraviti, A. et al. Polymeric nanoparticles for nonviral gene therapy extend brain tumor survival in vivo. ACS Nano 9, 1236–1249 (2015).

    CAS  Article  Google Scholar 

  4. Narayanan, K. et al. Mimicking cellular transport mechanism in stem cells through endosomal escape of new peptide-coated quantum dots. Sci. Rep. 3, 2184 (2013).

    Article  Google Scholar 

  5. Maude, S. L. et al. Chimeric antigen receptor T cells for sustained remissions in leukemia. N. Engl. J. Med. 371, 1507–1517 (2014).

    Article  Google Scholar 

  6. Davila, M. L., Kloss, C. C., Gunset, G. & Sadelain, M. CD19 CAR-targeted T cells induce long-term remission and B cell aplasia in an immunocompetent mouse model of B cell acute lymphoblastic leukemia. PLoS ONE 8, e61338 (2013).

    CAS  Article  Google Scholar 

  7. Nakazawa, Y. et al. Evaluation of long-term transgene expression in piggyBac-modified human T lymphocytes. J. Immunother. 36, 3–10 (2013).

    CAS  Article  Google Scholar 

  8. Burnight, E. R. et al. A hyperactive transposase promotes persistent gene transfer of a piggyBac DNA transposon. Mol. Ther. Nucleic Acids 1, e50 (2012).

    Article  Google Scholar 

  9. Gade, T. P. et al. Targeted elimination of prostate cancer by genetically directed human T lymphocytes. Cancer Res. 65, 9080–9088 (2005).

    CAS  Article  Google Scholar 

  10. Maude, S. L., Teachey, D. T., Porter, D. & Grupp, S. A. CD19-targeted chimeric antigen receptor T-cell therapy for acute lymphoblastic leukemia. Blood 125, 4017–4023 (2015).

    CAS  Article  Google Scholar 

  11. Grupp, S. A. et al. Chimeric antigen receptor-modified T cells for acute lymphoid leukemia. N. Engl. J. Med. 368, 1509–1518 (2013).

    CAS  Article  Google Scholar 

  12. Rosenberg, S. A. Cell transfer immunotherapy for metastatic solid cancer—what clinicians need to know. Nat. Rev. Clin. Oncol. 8, 577–585 (2011).

    CAS  Article  Google Scholar 

  13. Wang, X. et al. Large-scale clinical-grade retroviral vector production in a fixed-bed bioreactor. J. Immunother. 38, 127–135 (2015).

    Article  Google Scholar 

  14. Kuchenbaecker, K. B. et al. Identification of six new susceptibility loci for invasive epithelial ovarian cancer. Nat. Genet. 47, 164–171 (2015).

    CAS  Article  Google Scholar 

  15. Johnson, L. A. et al. Rational development and characterization of humanized anti-EGFR variant III chimeric antigen receptor T cells for glioblastoma. Sci. Transl. Med. 7, 275ra222 (2015).

    Article  Google Scholar 

  16. Magnani, C. F. et al. Immunotherapy of acute leukemia by chimeric antigen receptor-modified lymphocytes using an improved sleeping beauty transposon platform. Oncotarget 7, 51581–51597 (2016).

    Article  Google Scholar 

  17. Abate-Daga, D. et al. A novel chimeric antigen receptor against prostate stem cell antigen mediates tumor destruction in a humanized mouse model of pancreatic cancer. Hum. Gene Ther. 25, 1003–1012 (2014).

    CAS  Article  Google Scholar 

  18. Sadelain, M. CAR therapy: the CD19 paradigm. J. Clin. Invest. 125, 3392–3400 (2015).

    Article  Google Scholar 

  19. Meacham, C. E. & Morrison, S. J. Tumour heterogeneity and cancer cell plasticity. Nature 501, 328–337 (2013).

    CAS  Article  Google Scholar 

  20. Li, X. et al. Nanoparticle-mediated transcriptional modification enhances neuronal differentiation of human neural stem cells following transplantation in rat brain. Biomaterials 84, 157–166 (2016).

    CAS  Article  Google Scholar 

  21. Kim, J., Kang, Y., Tzeng, S. Y. & Green, J. J. Synthesis and application of poly(ethylene glycol)-co-poly(beta-amino ester) copolymers for small cell lung cancer gene therapy. Acta Biomater. 41, 293–301 (2016).

    CAS  Article  Google Scholar 

  22. Zhang, X., Edwards, J. P. & Mosser, D. M. The expression of exogenous genes in macrophages: obstacles and opportunities. Methods Mol. Biol. 531, 123–143 (2009).

    CAS  Article  Google Scholar 

  23. Marodon, G. et al. Specific transgene expression in human and mouse CD4+ cells using lentiviral vectors with regulatory sequences from the CD4 gene. Blood 101, 3416–3423 (2003).

    CAS  Article  Google Scholar 

  24. Ellmeier, W., Sunshine, M. J., Losos, K., Hatam, F. & Littman, D. R. An enhancer that directs lineage-specific expression of CD8 in positively selected thymocytes and mature T cells. Immunity 7, 537–547 (1997).

    CAS  Article  Google Scholar 

  25. Wu, C. Y., Rupp, L. J., Roybal, K. T. & Lim, W. A. Synthetic biology approaches to engineer T cells. Curr. Opin. Immunol. 35, 123–130 (2015).

    CAS  Article  Google Scholar 

  26. Kebriaei, P. et al. Phase I trials using sleeping beauty to generate CD19-specific CAR T cells. J. Clin. Invest. 126, 3363–3376 (2016).

    Article  Google Scholar 

  27. Monjezi, R. et al. Enhanced CAR T-cell engineering using non-viral sleeping beauty transposition from minicircle vectors. Leukemia 31, 186–194 (2017).

    CAS  Article  Google Scholar 

  28. Brooks, P. J., Yang, N. N. & Austin, C. P. Gene therapy: the view from NCATS. Hum. Gene Ther. 27, 7–13 (2016).

    CAS  Article  Google Scholar 

  29. Gaspar, V. et al. Minicircle DNA vectors for gene therapy: advances and applications. Expert Opin. Biol. Ther. 15, 353–379 (2015).

    CAS  Article  Google Scholar 

  30. Dobrenkov, K. et al. Monitoring the efficacy of adoptively transferred prostate cancer-targeted human T lymphocytes with PET and bioluminescence imaging. J. Nucl. Med. 49, 1162–1170 (2008).

    Article  Google Scholar 

  31. Fischer, K., Andreesen, R. & Mackensen, A. An improved flow cytometric assay for the determination of cytotoxic T lymphocyte activity. J. Immunol. Methods 259, 159–169 (2002).

    CAS  Article  Google Scholar 

  32. May, C. et al. Therapeutic haemoglobin synthesis in β-thalassaemic mice expressing lentivirus-encoded human β-globin. Nature 406, 82–86 (2000).

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank I. Stanishevskaya and D. Ehlert (cognitionstudio.com) for the design of the illustrations. We also thank M. Sadelain (Memorial Sloan-Kettering Cancer Center, New York, New York) for the Eμ-ALL01 cell line, and for the DNA construct that encodes an all-murine CD19-specific CAR. This work was supported in part by the Fred Hutchinson Cancer Research Center's Immunotherapy Initiative with funds provided by the Bezos Family Foundation, a New Idea Award from the Leukemia & Lymphoma Society, the Phi Beta Psi Sorority, the National Science Foundation (CAREER, award no. 1452492 and EAGER award no. 1644363), and the National Cancer Institute of the National Institutes of Health under award no. R01CA207407. M.T.S. was also supported by a Research Scholar Grant (RSG-16-110-01 – LIB) from the American Cancer Society.

Author information

Authors and Affiliations

Authors

Contributions

T.T.S., S.B.S. and H.F.M. designed and performed experiments and analysed and interpreted data. L.E.M. helped clone plasmid vectors, W.J. synthesized the PBAE polymer, and D.R. and E.B. helped with the large-scale purification of CAR-encoding plasmid DNA. M.E.W. performed the Southern blot analysis. S.P.S.P. performed and analysed in vivo safety/toxicity studies, and M.T.S. designed the study, performed experiments, analysed and interpreted data, and wrote the manuscript.

Corresponding author

Correspondence to Matthias T. Stephan.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary information

Supplementary information (PDF 2240 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Smith, T., Stephan, S., Moffett, H. et al. In situ programming of leukaemia-specific T cells using synthetic DNA nanocarriers. Nature Nanotech 12, 813–820 (2017). https://doi.org/10.1038/nnano.2017.57

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nnano.2017.57

Further reading

Search

Quick links

Find nanotechnology articles, nanomaterial data and patents all in one place. Visit Nano by Nature Research