Article | Published:

Scaffolds that mimic antigen-presenting cells enable ex vivo expansion of primary T cells

Nature Biotechnology volume 36, pages 160169 (2018) | Download Citation

Abstract

Therapeutic ex vivo T-cell expansion is limited by low rates and T-cell products of limited functionality. Here we describe a system that mimics natural antigen-presenting cells (APCs) and consists of a fluid lipid bilayer supported by mesoporous silica micro-rods. The lipid bilayer presents membrane-bound cues for T-cell receptor stimulation and costimulation, while the micro-rods enable sustained release of soluble paracrine cues. Using anti-CD3, anti-CD28, and interleukin-2, we show that the APC-mimetic scaffolds (APC-ms) promote two- to tenfold greater polyclonal expansion of primary mouse and human T cells compared with commercial expansion beads (Dynabeads). The efficiency of expansion depends on the density of stimulatory cues and the amount of material in the starting culture. Following a single stimulation, APC-ms enables antigen-specific expansion of rare cytotoxic T-cell subpopulations at a greater magnitude than autologous monocyte-derived dendritic cells after 2 weeks. APC-ms support over fivefold greater expansion of restimulated CD19 CAR-T cells than Dynabeads, with similar efficacy in a xenograft lymphoma model.

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References

  1. 1.

    & Adoptive cell transfer as personalized immunotherapy for human cancer. Science 348, 62–68 (2015).

  2. 2.

    , & Adoptive cellular therapy: a race to the finish line. Sci. Transl. Med. 7, 280ps7 (2015).

  3. 3.

    , & Engineered T cells: the promise and challenges of cancer immunotherapy. Nat. Rev. Cancer 16, 566–581 (2016).

  4. 4.

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

  5. 5.

    et al. T cells expressing CD19 chimeric antigen receptors for acute lymphoblastic leukaemia in children and young adults: a phase 1 dose-escalation trial. Lancet 385, 517–528 (2015).

  6. 6.

    et al. Safety and persistence of adoptively transferred autologous CD19-targeted T cells in patients with relapsed or chemotherapy refractory B-cell leukemias. Blood 118, 4817–4828 (2011).

  7. 7.

    et al. CD19 CAR-T cells of defined CD4+:CD8+ composition in adult B cell ALL patients. J. Clin. Invest. 126, 2123–2138 (2016).

  8. 8.

    et al. Kte-C19 (anti-CD19 CAR T Cells) induces complete remissions in patients with refractory diffuse large B-cell lymphoma (DLBCL): results from the pivotal phase 2 Zuma-1. Blood 128, LBA-6 (2016).

  9. 9.

    & T-cell-antigen recognition and the immunological synapse. Nat. Rev. Immunol. 3, 973–983 (2003).

  10. 10.

    et al. The immunological synapse balances T cell receptor signaling and degradation. Science 302, 1218–1222 (2003).

  11. 11.

    , & The immunological synapse: a cause or consequence of T-cell receptor triggering? Immunology 133, 420–425 (2011).

  12. 12.

    , , , & Full activation of the T cell receptor requires both clustering and conformational changes at CD3. Immunity 26, 43–54 (2007).

  13. 13.

    , & Artificial antigen presenting cells: an off the shelf approach for generation of desirable T-cell populations for broad application of adoptive immunotherapy. Adv. Genet. Eng. 4, 130 (2015).

  14. 14.

    et al. Manufacturing validation of biologically functional T cells targeted to CD19 antigen for autologous adoptive cell therapy. J. Immunother. 32, 169–180 (2009).

  15. 15.

    et al. Ex vivo expansion of polyclonal and antigen-specific cytotoxic T lymphocytes by artificial APCs expressing ligands for the T-cell receptor, CD28 and 4-1BB. Nat. Biotechnol. 20, 143–148 (2002).

  16. 16.

    et al. The effect of artificial antigen-presenting cells with preclustered anti-CD28/-CD3/-LFA-1 monoclonal antibodies on the induction of ex vivo expansion of functional human antitumor T cells. Haematologica 93, 1523–1534 (2008).

  17. 17.

    et al. Enrichment and expansion with nanoscale artificial antigen presenting cells for adoptive immunotherapy. ACS Nano 9, 6861–6871 (2015).

  18. 18.

    et al. Polymer-based synthetic dendritic cells for tailoring robust and multifunctional T cell responses. ACS Chem. Biol. 10, 485–492 (2015).

  19. 19.

    & A comprehensive platform for ex vivo T-cell expansion based on biodegradable polymeric artificial antigen-presenting cells. Mol. Ther. 16, 765–772 (2008).

  20. 20.

    et al. A carbon nanotube-polymer composite for T-cell therapy. Nat. Nanotechnol. 9, 639–647 (2014).

  21. 21.

    , , & Particle shape dependence of CD8+ T cell activation by artificial antigen presenting cells. Biomaterials 35, 269–277 (2014).

  22. 22.

    et al. Enhanced cellular activation with single walled carbon nanotube bundles presenting antibody stimuli. Nano Lett. 8, 2070–2076 (2008).

  23. 23.

    et al. Biodegradable nanoellipsoidal artificial antigen presenting cells for antigen specific T-cell activation. Small 11, 1519–1525 (2015).

  24. 24.

    , , , & An artificial antigen-presenting cell with paracrine delivery of IL-2 impacts the magnitude and direction of the T cell response. J. Biol. Chem. 286, 34883–34892 (2011).

  25. 25.

    Novartis CTL019 Oncologic Drugs Advisory Committee Briefing Document (2017).

  26. 26.

    & Comparison of anti-CD3 and anti-CD28-coated beads with soluble anti-CD3 for expanding human T cells: differing impact on CD8 T cell phenotype and responsiveness to restimulation. J. Transl. Med. 8, 104 2010).

  27. 27.

    et al. Allogeneic lymphocyte-licensed DCs expand T cells with improved antitumor activity and resistance to oxidative stress and immunosuppressive factors. Mol. Ther. Methods Clin. Dev. 1, 14001 (2014).

  28. 28.

    et al. CD27 is required for generation and long-term maintenance of T cell immunity. Nat. Immunol. 1, 433–440 (2000).

  29. 29.

    et al. Treatment of metastatic melanoma with autologous CD4+ T cells against NY-ESO-1. N. Engl. J. Med. 358, 2698–2703 (2008).

  30. 30.

    et al. Phase I study of adoptive T-cell therapy using antigen-specific CD8+ T cells for the treatment of patients with metastatic melanoma. J. Clin. Oncol. 24, 5060–5069 (2006).

  31. 31.

    et al. Infusion of cytotoxic T cells for the prevention and treatment of Epstein-Barr virus-induced lymphoma in allogeneic transplant recipients. Blood 92, 1549–1555 (1998).

  32. 32.

    et al. Dendritic cells are dysfunctional in patients with operable breast cancer. Cancer Immunol. Immunother. 53, 510–518 (2004).

  33. 33.

    & Antigen-specific activation and cytokine-facilitated expansion of naive, human CD8+ T cells. Nat. Protoc. 9, 950–966 (2014).

  34. 34.

    et al. The immunological synapse: a molecular machine controlling T cell activation. Science 285, 221–227 (1999).

  35. 35.

    et al. Injectable, spontaneously assembling, inorganic scaffolds modulate immune cells in vivo and increase vaccine efficacy. Nat. Biotechnol. 33, 64–72 (2015).

  36. 36.

    et al. The effect of surface modification of mesoporous silica micro-rod scaffold on immune cell activation and infiltration. Biomaterials 83, 249–256 (2016).

  37. 37.

    , , & Membrane-mediated effect on ion channels induced by the anesthetic drug ketamine. J. Am. Chem. Soc. 132, 7990–7997 (2010).

  38. 38.

    , , , & Functional single-cell analysis of T-cell activation by supported lipid bilayer-tethered ligands on arrays of nanowells. Lab Chip 13, 90–99 (2013).

  39. 39.

    et al. Acquisition of full effector function in vitro paradoxically impairs the in vivo antitumor efficacy of adoptively transferred CD8+ T cells. J. Clin. Invest. 115, 1616–1626 (2005).

  40. 40.

    et al. Isolation of neoantigen-specific T cells from tumor and peripheral lymphocytes. J. Clin. Invest. 125, 3981–3991 (2015).

  41. 41.

    et al. Targeting of cancer neoantigens with donor-derived T cell receptor repertoires. Science 352, 1337–1341 (2016).

  42. 42.

    HLA anchor optimization of the melan-A-HLA-A2 epitope within a long peptide is required for efficient cross-priming of human tumor-reactive T cells. J. Immunol. 188, 2102–2110 (2012).

  43. 43.

    & Artificial antigen-presenting cells for use in adoptive immunotherapy. Cancer J. 16, 374–381 (2010).

  44. 44.

    , , & Towards efficient cancer immunotherapy: advances in developing artificial antigen-presenting cells. Trends Biotechnol. 32, 456–465 (2014).

  45. 45.

    & Planar lipid bilayers on solid supports from liposomes--factors of importance for kinetics and stability. Biochim. Biophys. Acta 1327, 149–161 (1997).

  46. 46.

    , , & Characterization and control of lipid layer fluidity in hybrid bilayer membranes. J. Am. Chem. Soc. 129, 2094–2100 (2007).

  47. 47.

    & Tuning lipid mixtures to induce or suppress domain formation across leaflets of unsupported asymmetric bilayers. Proc. Natl. Acad. Sci. USA 105, 124–128 (2008).

  48. 48.

    et al. Asymmetric structural features in single supported lipid bilayers containing cholesterol and GM1 resolved with synchrotron X-Ray reflectivity. Biophys. J. 95, 657–668 (2008).

  49. 49.

    , & Stability and liquid-liquid phase separation in mixed saturated lipid bilayers. Biophys. J. 96, 3977–3986 (2009).

  50. 50.

    , & Induction of potent anti-tumor responses while eliminating systemic side effects via liposome-anchored combinatorial immunotherapy. Biomaterials 32, 5134–5147 (2011).

  51. 51.

    et al. Bioorthogonal copper-free click chemistry in vivo for tumor-targeted delivery of nanoparticles. Angew. Chem. Int. Ed. 51, 11836–11840 (2012).

  52. 52.

    , , , & Versatile click alginate hydrogels crosslinked via tetrazine-norbornene chemistry. Biomaterials 50, 30–37 (2015).

  53. 53.

    et al. Chimeric receptors with 4-1BB signaling capacity provoke potent cytotoxicity against acute lymphoblastic leukemia. Leukemia 18, 676–684 (2004).

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Acknowledgements

This work was supported by the National Institutes of Health (NIH) (1R01EB015498 U01 CA214369) and the Wyss Institute for Biologically Inspired Engineering at Harvard University. D.K.Y.Z. was supported by the Canadian Institutes of Health Research (CIHR-DFSA). S.T.K. was supported by an HHMI ISRF. This work was performed in part at the Center for Nanoscale Systems (CNS), a member of the National Nanotechnology Coordinated Infrastructure Network (NNCI), which is supported by the National Science Foundation under NSF award no. 1541959. CNS is part of Harvard University. We thank the National Institutes of Health (NIH) Tetramer Core Facility for the SIINFEKL/H-2K(b) biotinylated monomer, Alexa Fluor 647-labeled SIINFEKL/H-2K(b) tetramer, CLGGLLTMV/HLA-A*0201 biotinylated monomer, Alexa Fluor 647-labeled CLGGLLTMV/HLA-A*02:01 tetramer, GLCTLVAML/HLA-A*02:01 biotinylated monomer, and Alexa Fluor 647-labeled GLCTLVAML/HLA-A*0201 tetramer, N. Shastri for the B3Z cell line, and G. Freeman for the T2 cell line. We thank Unum Therapeutics for the luciferized Raji cell line and the 19BBz CAR-T cells. We also thank C. Stamoulis from Boston Children′s Hospital and the Harvard Catalyst for her help with statistical analysis; Harvard Catalyst is supported, in part, by the NIH (UL1 TR001102). Lastly, we thank R. Bates, M. Pezone, B. Schultes, L. Edwards, G. Motz, T. Barnitz, T. Hickman, K. McGinness, J. Ritz, M. Maus, T. Snyder, and W.A. Li for valuable scientific discussions.

Author information

Affiliations

  1. John A. Paulson School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts, USA.

    • Alexander S Cheung
    • , David K Y Zhang
    • , Sandeep T Koshy
    •  & David J Mooney
  2. The Wyss Institute for Biologically Inspired Engineering Harvard University, Cambridge, Massachusetts, USA.

    • Alexander S Cheung
    • , David K Y Zhang
    • , Sandeep T Koshy
    •  & David J Mooney
  3. Harvard-MIT Division of Health Sciences and Technology, Cambridge, Massachusetts, USA.

    • Sandeep T Koshy

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Contributions

A.S.C. and D.J.M. conceived and designed the experiments. A.S.C., D.K.Y.Z., and S.T.K. performed the experiments. A.S.C., D.K.Y.Z., and D.J.M. analyzed the data. A.S.C., D.K.Y.Z., and D.J.M. wrote the manuscript. All authors discussed the results and commented on the manuscript. The principal investigator is D.J.M.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to David J Mooney.

Integrated supplementary information

Supplementary information

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  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–9

  2. 2.

    Life Sciences Reporting Summary

  3. 3.

    Supplementary Table 1

    Enrichment of EBV-specific CD8+ 1 T cells following a single stimulation with APC-ms presenting either CLG or GLC.

Videos

  1. 1.

    Formation of APC-ms in culture.

    Individual rods randomlysettled and stacked to form a 3D scaffold in culture.

  2. 2.

    Infiltration of APC-ms by primary T cells.

    The large interparticle spaces formed by 3D scaffold were infiltrated by mouse T cells.

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DOI

https://doi.org/10.1038/nbt.4047

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