Parallel lipid-carried epitope pillars enhance stimulation of chimeric antigen receptor T cells

Chimeric antigen receptor T cells are genetically engineered to express a specific T cell receptor of interest, such as to target a cancer-specific antigen. Signaling events in chimeric antigen receptor T cells are essential for their proliferation, survival, and function. To achieve optimal antitumor efficacy, signaling motifs as well as the structure of the transmembrane domain of the chimeric antigen receptor have to be carefully designed. However, it remains unclear whether the arrangement, and therefore the movement and elasticity, of tumor antigens influence their stimulation of chimeric antigen receptor T cells. Here we show that MUC1 molecules tethered to a glycosylphosphatidylinositol anchor drastically increased its stimulation of chimeric antigen receptor T cells. In particular, MUC1 molecules containing one tandem repeat sequence showed significantly greater stimulatory activity than five tandem repeat sequences. Thus glycosylphosphatidylinositol-anchored, tumor antigen epitope pillars in parallel are attractive targets for the design of chimeric antigen receptor T cells. These novel findings, which we propose as the parallel lipid-carried epitope model, emphasize the importance of epitope arrangement in selecting highly effective chimeric antigen receptor T cells, potentially revolutionizing the applicability of this therapy. Furthermore, our data have implications for the necessity of methodologies to measure the elasticity, movement, and density of antigen pillars in parallel, as key tools to guide future chimeric antigen receptor T cell therapy.