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Activation and expansion of human T cells using artificial antigen-presenting cell scaffolds

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Abstract

Synthetic antigen-presenting cells (APCs) are used to mediate scalable ex vivo T-cell expansion for adoptive cell therapy. Recently, we developed APC-mimetic scaffolds (APC-ms), which present signals to T cells in a physiological manner to mediate rapid and controlled T-cell expansion. APC-ms are composed of individual high-aspect-ratio silica microrods loaded with soluble mitogenic cues and coated with liposomes of defined compositions, to form supported lipid bilayers. Membrane-bound ligands for stimulation and co-stimulation of T-cell receptors are presented via the fluid, synthetic membranes, while mitogenic cues are released slowly from the microrods. In culture, interacting T cells assemble the individual APC-ms microrods into a biodegradable 3D matrix. Compared to conventional methods, APC-ms facilitates several-fold greater polyclonal T-cell expansion and improved antigen-specific enrichment of rare T-cell subpopulations. Here we provide a detailed protocol for APC-ms synthesis and use for human T-cell activation, and discuss important considerations for material design and T-cell co-culture. This protocol describes the facile assembly of APC-ms in ~4 h and rapid expansion or enrichment of relevant T-cell clones in <2 weeks, and is applicable for T-cell manufacturing and assay development.

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Fig. 1: Ex vivo manufacture of ACTs requires T-cell activation and expansion.
Fig. 2: Assembly of APC-ms and use for T-cell activation and expansion.
Fig. 3: Relevant APC-ms design considerations.
Fig. 4: Representative data for polyclonal expansion of primary human T cells using APC-ms.

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Data availability

Data for this study are available from the corresponding author upon reasonable request.

Change history

  • 24 January 2020

    Fig. 1 as initially published had labels that were too small to be legible. The figure was replaced, and the legend text adapted accordingly, on 24 January 2020.

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Acknowledgements

We thank O. Ali, A. Najibi, M. Sobral, and H. Wang for critical reading of the protocol. We are grateful to G. Cuneo, T. Ferrante, H. Ijaz, and C. Johnson for their technical expertise and help. This work was partially supported by the Wyss Institute at Harvard University (D.J.M.), the National Institutes of Health (NIH 1R01EB015498 and U01CA214369 to D.J.M.), the Food and Drug Administration (FDA, RFA-FD-18-023), and a fellowship from the Canadian Institutes of Health Research (CIHR, D.K.Y.Z.). Part of this work was performed at the Center for Nanoscale Systems (CNS) at Harvard University, which is supported by the National Science Foundation (NSF; 1541979).

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Contributions

A.C.S. conceived the protocol. D.K.Y.Z. and A.S.C. collected and analyzed the experimental data. D.K.Y.Z. designed the figures. D.K.Y.Z., A.S.C., and D.J.M. wrote the paper.

Corresponding author

Correspondence to David J. Mooney.

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Competing interests

An international patent application (WO2018013797A1) describes APC-ms compositions. A.S.C. is a co-founder of Immulus, Inc. D.J.M. is a co-founder, advisor, and chair of Immulus’ scientific advisory board. D.K.Y.Z. declares no competing interests.

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Cheung, A. S. et al. Nat. Biotechnol. 36, 160–169 (2018): https://www.nature.com/articles/nbt.4047

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Supplementary Figure 1 Flow cytometry analysis of various T cell subpopulations.

Cell populations should be gated using appropriate fluorescence-minus-one controls. Gating strategy for live CD3+CD8+ T-cell subpopulations is shown; the same approach may be applied for CD3+CD4+ T cells. N: naïve T cell; SCM: stem memory T cell; CM: central memory T cell; EM: effector memory T cell; E: effector T cell.

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Zhang, D.K.Y., Cheung, A.S. & Mooney, D.J. Activation and expansion of human T cells using artificial antigen-presenting cell scaffolds. Nat Protoc 15, 773–798 (2020). https://doi.org/10.1038/s41596-019-0249-0

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