Self‐Assembly of Immune Signals Improves Codelivery to Antigen Presenting Cells and Accelerates Signal Internalization, Processing Kinetics, and Immune Activation

Abstract Vaccines and immunotherapies that elicit specific types of immune responses offer transformative potential to tackle disease. The mechanisms governing the processing of immune signals—events that determine the type of response generated—are incredibly complex. Understanding these processes would inform more rational vaccine design by linking carrier properties, processing mechanisms, and relevant timescales to specific impacts on immune response. This goal is pursued using nanostructured materials—termed immune polyelectrolyte multilayers—built entirely from antigens and stimulatory toll‐like receptors agonists (TLRas). This simplicity allows isolation and quantification of the rates and mechanisms of intracellular signal processing, and the link to activation of distinct immune pathways. Each vaccine component is internalized in a colocalized manner through energy‐dependent caveolae‐mediated endocytosis. This process results in trafficking through endosome/lysosome pathways and stimulation of TLRs expressed on endosomes/lysosomes. The maximum rates for these events occur within 4 h, but are detectable in minutes, ultimately driving downstream proimmune functions. Interestingly, these uptake, processing, and activation kinetics are significantly faster for TLRas in particulate form compared with free TLRa. Our findings provide insight into specific mechanisms by which particulate vaccines enhance initiation of immune response, and highlight quantitative strategies to assess other carrier technologies.