Balancing the Nanoscale Organization in Multivalent Materials for Functional Inhibition of the Programmed Death-1 Immune Checkpoint.

Dendritic cells (DCs) regulate immune priming by expressing programmed death ligand 1 (PD-L1) and PD-L2, which interact with the inhibitory receptor PD-1 on activated T cells. PD-1 signaling regulates T cell effector functions and limits autoimmunity. Tumor cells can hijack this pathway by overexpressing PD-L1 to suppress antitumor T cell responses. Blocking this inhibitory pathway has been beneficial for the treatment of various cancer types, although only a subset of patients responds. A deepened understanding of the spatial organization and molecular interplay between PD-1 and its ligands may inform the design of more efficacious nanotherapeutics. We visualized the natural molecular PD-L1 organization on DCs by DNA-PAINT microscopy and created a template to engineer DNA-based nanoclusters presenting PD-1 at defined valencies, distances, and patterns. These multivalent nanomaterials were examined for their cellular binding and blocking ability. Our data show that PD-1 nano-organization has profound effects on ligand interaction and that the valency of PD-1 molecules modulates the effectiveness in restoring T cell function. This work highlights the power of spatially controlled functional materials to unravel the importance of multivalent patterns in the PD-1 pathway and presents alternative design strategies for immune-engineering.

Multivalent Pattern Recognition through Control of Nano-Spacing in Low-Valency Super-Selective Materials.

Super-selective multivalent ligand-receptor interactions display a signature step-like onset in binding when meeting a characteristic density of target receptors. Materials engineered for super-selective binding generally display a high number of flexible ligands to enhance the systems' avidity. In many biological processes, however, ligands are present in moderate copy numbers and arranged in spatio-temporal patterns. In this low-valency regime, the rigidity of the ligand-presenting architecture plays a critical role in the selectivity of the multivalent complex through decrease of the entropic penalty of binding. Exploiting the precision in spatial design inherent to the DNA nanotechnology, we engineered a library of rigid architectures to explore how valency, affinity, and nano-spacing control the presence of super-selectivity in multivalent binding. A micromolar monovalent affinity was required for super-selective binding to be observed within low-valency systems, and the transition point for stable interactions was measured at hexavalent ligand presentation, setting the limits of the low-valency regime. Super-selective binding was observed for all hexavalent architectures, and, more strikingly, the ligand pattern determined the selectivity onset. Hereby, we demonstrate for the first time that nano-control of geometric patterns can be used to discriminate between receptor densities in a super-selective manner. Materials that were indistinguishable in their molecular composition and ligand valency bound with various efficacies on surfaces with constant receptor densities. We define this new phenomenon in super-selective binding as multivalent pattern recognition.

Nucleic acids presenting polymer nanomaterials as vaccine adjuvants.

Most vaccines developed today include only the antigens that best stimulate the immune system rather than the entire virus or microbe, which makes vaccine production and use safer and easier, though they lack potency to induce acceptable immunity and long-term protection. The incorporation of additional immune stimulating components, named adjuvants, is required to generate a strong protective immune response. Nucleic acids (DNA and RNA) and their synthetic analogs are promising candidates as vaccine adjuvants activating Toll-like receptors (TLRs). Additionally, in the last few years several nanocarriers have emerged as platforms for targeted co-delivery of antigens and adjuvants. In this review, we focus on the recent developments in polymer nanomaterials presenting nucleic acids as vaccine adjuvants. We aim to compare the effectiveness of the various classes of polymers in immune modulating materials (nanoparticles, dendrimers, single-chain particles, nanogels, polymersomes and DNA-based architectures). In particular, we address the critical role of parameters such as size, shape, complexation and release of TLR ligands, cellular uptake, stability, toxicity and potential importance of spatial control in ligand presentation.