Supramolecular hydrogels are used in the 3D printing of high-resolution, multi-material structures. The non-covalent bonds allow the extrusion of the inks into support gels to directly write structures continuously in 3D space. This material system supports the patterning of multiple inks, cells, and void spaces.

Tumour-associated macrophages (TAMs) are abundant in many cancers, and often display an immune-suppressive M2-like phenotype that fosters tumour growth and promotes resistance to therapy. Yet macrophages are highly plastic and can also acquire an anti-tumourigenic M1-like phenotype. Here, we show that R848, an agonist of the toll-like receptors (TLRs) TLR7 and TLR8 identified in a morphometric-based screen, is a potent driver of the M1 phenotype in vitro and that R848-loaded β-cyclodextrin nanoparticles (CDNPs) lead to efficient drug delivery to TAMs in vivo. As a monotherapy, the administration of CDNP-R848 in multiple tumour models in mice altered the functional orientation of the tumour immune microenvironment towards an M1 phenotype, leading to controlled tumour growth and protecting the animals against tumour rechallenge. When used in combination with the immune checkpoint inhibitor anti-PD-1, we observed improved immunotherapy response rates, also in a tumour model resistant to anti-PD-1 therapy. Our findings demonstrate the ability of rationally engineered drug–nanoparticle combinations to efficiently modulate TAMs for cancer immunotherapy.

RNA-binding proteins (RBPs) with prion-like domains (PrLDs) phase transition to functional liquids, which can mature into aberrant hydrogels composed of pathological fibrils that underpin fatal neurodegenerative disorders. Several nuclear RBPs with PrLDs, including TDP-43, FUS, hnRNPA1, and hnRNPA2, mislocalize to cytoplasmic inclusions in neurodegenerative disorders, and mutations in their PrLDs can accelerate fibrillization and cause disease. Here, we establish that nuclear-import receptors (NIRs) specifically chaperone and potently disaggregate wild-type and disease-linked RBPs bearing a NLS. Karyopherin-β2 (also called Transportin-1) engages PY-NLSs to inhibit and reverse FUS, TAF15, EWSR1, hnRNPA1, and hnRNPA2 fibrillization, whereas Importin-α plus Karyopherin-β1 prevent and reverse TDP-43 fibrillization. Remarkably, Karyopherin-β2 dissolves phase-separated liquids and aberrant fibrillar hydrogels formed by FUS and hnRNPA1. In vivo, Karyopherin-β2 prevents RBPs with PY-NLSs accumulating in stress granules, restores nuclear RBP localization and function, and rescues degeneration caused by disease-linked FUS and hnRNPA2. Thus, NIRs therapeutically restore RBP homeostasis and mitigate neurodegeneration.

The development of printable biomaterial inks is critical to the application of 3D printing in biomedicine. To print high-resolution structures with fidelity to a computer-aided design, materials used in 3D printing must be capable of being deposited on a surface and maintaining a printed structure. A dual-cross-linking hyaluronic acid system was studied here as a printable hydrogel ink, which encompassed both shear-thinning and self-healing behaviors via guest–host bonding, as well as covalent cross-linking for stabilization using photopolymerization. When either guest–host assembly or covalent cross-linking was used alone, long-term stable structures were not formed, because of network relaxation after printing or dispersion of the ink filaments prior to stabilization, respectively. The dual-cross-linking hydrogel filaments formed structures with greater than 16 layers that were stable over a month with no loss in mechanical properties and the printed filament size ranged from 100 to 500 μm, depending on printing parameters (needle size, speed, and extrusion flux). Printed structures were further functionalized (i.e., RGD peptide) to support cell adhesion. This work highlights the importance of ink formulation and cross-linking on the printing of stable hydrogel structures.