CAR T‐cells' Interaction with Artificial Antigen Presenting Cell Surface

Abstract

Adoptive Cell Transfer (ACT) using Chimeric Antigen Receptor (CAR) T-cells shows promising results in treating metastatic melanoma and B cell malignancies. However, the detailed mechanism of CAR T-cell's interaction with tumors is still not well-understood. In this work, we created an artificial antigen presenting cell (APC) surface for 2nd generation (CD3 and CD28 signaling domains) CD19 and GD2 CAR T-cells and studied the interaction of CAR T-cells in terms of synapse formation. We created circular patterns of antibodies against CAR T-cells using micro-contact printing and measured the level of activation and degranulation in both CAR T-cells by quantifying the intensity of the phosphorylated CD3ζ chain (p-CD3ζ) and the Lysosome-Associated Membrane Protein 1 (LAMP-1) respectively. The results show that we can study the interaction of CAR T cell with APC surface in a reproducible manner and that different CAR T cells have a different level of activation and degranulation. Our findings suggest that the artificial APC allows for quantitative measurement of effectiveness for different CAR T cells with controlled APC density. Support or Funding Information This work was supported by the National Science Foundation through the Early-concept Grants for Exploratory Research (EAGER) (Award number: 1649243) and was performed in part at the Joint School of Nanoscience and Nanoengineering, a member of the Southeastern Nanotechnology Infrastructure Corridor (SENIC) and National Nanotechnology Coordinated Infrastructure (NNCI), which is supported by the National Science Foundation (Grant ECCS-1542174). The figure shows the interaction of GD2 CAR T-cell with 5 μm micropatterns of 1A7 (target). The aggregation of the phosphorylated CD3 zeta chain (p-CD3ζ) at the contact with the micropattern is clear. Micropatterns of 1A7 are labeled with Alexa Fluor 568 Goat anti-mouse secondary antibody and p-CD3ζ is labeled with Alexa Fluor 488 primary antibody. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.