Biomolecule Conjugation Strategy for HAGM Cryogels to Create 3D Immune Niches that Induce Multifunctional T Cells

Recently, biomaterials have emerged as tools to activate and expand T cells in the context of cancer immunotherapy. Most designs accommodate T cells with a stimulatory two-dimensional (2D) environment. In contrast, three-dimensional (3D) scaffolds, mimicking the complex architecture of the lymph node, have been shown to outperform 2D synthetic constructs, resulting in a more optimal T-cell expansion and phenotype. Here, we used injectable glycidyl methacrylated hyaluronic acid (HAGM)-based cryogel scaffolds to create a modular biodegradable 3D stimulatory immune niche. We developed a strategy to achieve highly specific and efficient covalent linking of immune-activating biomolecules, such as T-cell-activating peptide MHC complexes and antibodies, to HAGM scaffolds without compromising the injectable properties of the cryogels. Importantly, because our conjugation strategy is carried out postcryogelation, biomolecules are not exposed to free radicals and freeze–thawing cycles, facilitating highly reproducible covalent attachment. Our scaffold potently activates human- and murine-T cells, inducing higher levels of multifunctional T cells with a less exhausted phenotype compared to 2D cultures. Following injections, HAGM scaffolds retain up to 60% of highly proliferative T cells. In conclusion, our HAGM scaffolds are an easily adaptable tool for robust T-cell activation, thus further expanding the biomaterial-based immunotherapy toolbox.