FOCiS 2018 Abstract: Single-Cell Highly Multiplexed Proteomics Identifies Novel Polyfunctional Human CD8+ T Cell Signatures Induced by a Nanoparticle-based Melanoma Vaccine in Human Immune System Mice
Polyfunctional T cells (co-secretion of 2+ proteins per single cell) are an important functional attribute of a quality human T-cell immune response to antigen. Assessment of human T-cell polyfunctional responses to tumor vaccines in humanized mice, which are reconstituted with human immune system (HIS), provides a potentially valuable translational tool for tumor vaccine candidate screening and efficacy determination pre-clinically.
We utilized 32-plex single-cell functional proteomics platform to profile human CD8+ T-cells in HIS mice induced by a nanoparticle-based melanoma vaccine (NP). HIS mice were vaccinated with different formulations of NP or vehicle. Splenic human CD8+ T-cells were isolated, stimulated with anti-human CD3/CD28 at 37°C, 5% CO2 for 24 hours and loaded into a single-cell IsoCode Chip, pre-patterned with a 32-plex antibody ELISA array per cellular microchamber. Protein secretions were analyzed from ~1000 single human CD8+ T-cells after 16-hour-on-chip incubation.
The single-cell analysis demonstrates sensitive detection of robust upregulation of polyfunctional human CD8+ T-cell subsets in NP-vaccinated HIS mice compared to controls. The enhanced polyfunctional strength index of the human CD8+ T-cells by the vaccine was driven by antitumor-associated proteins including granzyme B, IFN-γ, MIP-1α, perforin and TNF-α. Additionally, novel single-cell visualizations revealed distinct polyfunctional cell signatures with these combinatorial human cytokine secretions that drove human CD8+ T-cell response by the NP. Polyfunctional human CD8+ T-cell responses sensitively demonstrated successful induction of the tumor vaccine in HIS mice, which may serve as a translational tool for more accurate evaluation of tumor vaccine efficacy in a pre-clinical setting.
Mackay S, Morse K, Paczkowski P, Huang J, Tsuji M, and Zhou J. Single-Cell Highly Multiplexed Proteomics Identifies Novel Polyfunctional Human CD8+ T Cell Signatures Induced by a Nanoparticle-based Melanoma Vaccine in Human Immune System Mice. To be presented at FOCiS Annual Meeting 2018.
Lu Y, Xue Q, Eisele MR, Sulistijo E, Brower K, Han L, Amir ED, Pe’er D, Miller-Jensen K, and Fan R. Highly Multiplexed Profiling of Immune Effector Functions Reveals Deep Functional Heterogeneity in Response to Pathogenic Ligands. Proc. Natl. Acad. Sci., 112 (7), 607-615 (2015).
Lu Y, Chen JJ, Mu L, Xue Q, Wu Y, Wu PH, Li J, Vortmeyer AO, Miller-Jensen K, Wirtz D, and Fan R. High-throughput Secretomic Analysis of Single Cells to Assess Functional Cellular Heterogeneity. Analytical Chemistry, 85 (4), 2548– 2556 (2013).
Ma C, Fan R, Ahmad H, Shi Q, Comin-Anduix B, Chodon T, Koya RC, Liu CC, Kwong GA, Radu CG, Ribas A, and Heath JR. A Clinical Microchip for Evaluation of Single Immune Cells Reveals High Functional Heterogeneity in Phenotypically Similar T Cells. Nature Medicine, 17, 738-743 (2011).