Functionalizing hydrogel nanovials with vesicles mimicking antigen-presenting vesicles and cancer exosomes improves T cell capture and activation

Recent advances have demonstrated the application of microcavity-containing hydrogel microparticles, known as nanovials, for the massively parallel and high-throughput screening of therapeutic T cell populations for adoptive cell therapies. Nanovial cavities coated with peptide-MHC (pMHC) or antigen tetramers selectively bind to their cognate T cell receptor (TCR) or chimeric antigen receptor (CAR) to activate T cells and capture secreted cytokines. However, binding of tetramers or recombinantly expressed antigen by T cells is not always correlated with T cell activation or cytotoxicity as the binding interface is not fully representative of the natural immunological synapse formed between T cells and professional antigen-presenting cells (APCs). Here, we leverage the recent discovery of an ESCRT- and ALIX-binding region (EABR) sequence to generate antigen-presenting vesicles and cancer-mimicking exosomes from standard HEK293T and Expi293F cell cultures. EABR-mediated vesicles present natural, full-length oncologically-relevant membrane proteins embedded in lipid bilayers to functionalize the nanovial cavity with cell-like membranes. These hydrogel nanovials functionalized with the EABR-mediated vesicles show improved T cell capture of 1G4 T cells and enhanced activation of HER2 CAR-T cells compared to hydrogel surfaces functionalized with recombinantly-expressed soluble proteins.