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Despite promising progress in cancer vaccination, therapeutic effectiveness is often insufficient. Cancer vaccine effectiveness could be enhanced by targeting vaccine antigens to antigen-presenting cells, thereby increasing T-cell activation. CD169-expressing splenic macrophages efficiently capture particulate antigens from the blood and transfer these antigens to dendritic cells for the activation of CD8+ T cells. In this study, we incorporated a physiological ligand for CD169, the ganglioside GM3, into liposomes to enhance liposome uptake by CD169+ macrophages. We assessed how variation in the amount of GM3, surface-attached PEG and liposomal size affected the binding to, and uptake by, CD169+ macrophages in vitro and in vivo. As a proof of concept, we prepared GM3-targeted liposomes containing a long synthetic ovalbumin peptide and tested the capacity of these liposomes to induce CD8+ and CD4+ T-cell responses compared to control liposomes or soluble peptide. The data indicate that the delivery of liposomes to splenic CD169+ macrophages can be optimized by the selection of liposomal constituents and liposomal size. Moreover, optimized GM3-mediated liposomal targeting to CD169+ macrophages induces potent immune responses and therefore presents as an interesting delivery strategy for cancer vaccination.

Self-assembling peptides are an exemplary class of supramolecular biomaterials of broad biomedical utility. Mechanistic studies on the peptide self-assembly demonstrated the importance of the oligomeric intermediates towards the properties of the supramolecular biomaterials being formed. In this study, we demonstrate how the overall yield of the supramolecular assemblies are moderated through subtle molecular changes in the peptide monomers. This strategy is exemplified with a set of surfactant-like peptides (SLPs) with different β-sheet propensities and charged residues flanking the aggregation domains. By integrating different techniques, we show that these molecular changes can alter both the nucleation propensity of the oligomeric intermediates and the thermodynamic stability of the fibril structures. We demonstrate that the amount of assembled nanofibers are critically defined by the oligomeric nucleation propensities. Our findings offer guidance on designing self-assembling peptides for different biomedical applications, as well as insights into the role of protein gatekeeper sequences in preventing amyloidosis.

Recent studies have shown a high potency of protein-based vaccines for cell-mediated cancer immunotherapy. However, due to their poor cellular uptake, efficient immune responses with soluble protein antigens are often not observed. As a result of superior cellular uptake, nanogels loaded with antigenic peptides were investigated in this study as carrier systems for cancer immunotherapy. Different synthetic long peptides (SLPs) containing the CTL and CD4+ T-helper (Help) epitopes were synthesized and covalently conjugated via disulfide bonds to the polymeric network of cationic dextran nanogels. Cationic nanogels with a size of 210 nm, positive zeta potential (+24 mV) and high peptide loading content (15%) showed triggered release of the loaded peptides under reducing conditions. An in vitro study demonstrated the capability of cationic nanogels to maturate dendritic cells (DCs). Importantly, covalently SLP-loaded nanogels adjuvanted with poly(I:C) showed superior CD8+ T cell responses compared to soluble peptides and nanogel formulations with physically loaded peptides both in vitro and in vivo. In conclusion, covalently SLPs-loaded cationic nanogels are a promising system to provoke immune responses for therapeutic cancer vaccination.

The current treatment of autoimmune and chronic inflammatory diseases entails systemic immune suppression, which is associated with increased susceptibility to infections. To restore immune tolerance and reduce systemic side effects, a targeted approach using tolerogenic dendritic cells (tolDCs) is being explored. tolDCs are characterized by the expression of CD11c, the major histocompatibility complex (MHC)II and low levels of co-stimulatory molecules CD40 and CD86. In this study, tolDCs were generated using a human-proteoglycan-derived peptide (hPG) and all-trans retinoic acid (RA). RA-tolDCs not only display a tolerogenic phenotype but also can induce an antigen-specific regulatory T cell (Treg) response in vitro. However, further analysis showed that RA-tolDCs make up a heterogeneous population of DCs, with only a small proportion being antigen-associated tolDCs. To increase the homogeneity of this population, 1,2-distearoyl-sn-glycero-3-phosphoglycerol (DSPG)-containing liposomes were used to encapsulate the relevant antigen together with RA. These liposomes greatly enhanced the proportion of antigen-associated tolDCs in culture. In addition, in mice, we showed that the liposomal co-delivery of antigen and RA can be a more targeted approach to induce antigen-specific tolerance compared to the injection of RA-tolDCs, and that these liposomes can stimulate the generation of antigen-specific Tregs. This work highlights the importance of the co-delivery of an antigen and immunomodulator to minimize off-target effects and systemic side effects and provides new insights in the use of RA for antigen-specific immunotherapy for autoimmune and chronic inflammatory diseases.

Successful delivery of mRNA into the cytosol of professional antigen-presenting cells (APCs) poses one of the biggest challenges in developing effective mRNA vaccines to treat various cancers and viral infectious diseases. However, most polymeric mRNA delivery systems fail to transfect APCs. We have discovered that decoration of pH-sensitive endosome-disruptive GALA peptides on the surface of mRNA polyplexes leads to efficient targeting and transfection of APCs. GALA peptides not only enhance specific uptake in APCs through binding to sialic acid moieties, they also facilitate the endosomal escape of mRNA especially in dendritic cells (DCs). Here, we describe in detail the production of stabilized mRNA polyplexes post-conjugated with GALA peptides via copper-free click chemistry. Methods described here include the synthesis and purification of GALA peptides and its conjugation to mRNA polyplexes.