A biomime c model combining 3D muscle-like ssue, stromal and immune cells for recapitulating the rapid immune signature predic ve of mRNA vaccine immunogenicity

Protec ve responses raised by vaccina on result from the coordinated ac on of immune cells in mul ple loca ons across the body. In this context, system vaccinology studies provide an integrated picture of vaccine-driven molecular networks that are associated with later adap ve vaccine protec ve responses. Blood modules, elicited early a er vaccina on, belong to innate immunity and include pro-inflammatory cytokines, interferon (IFN)-smulated genes and transcripts associated with metabolic altera on cri cal for an viral responses, an gen presenta on and B cell ac va on. In par cular, the cytokines IL-15, IL-6, TNF-α, IFN-γ, the chemokines IP-10 and MCP-1 and the IFN inducible genes MX1 and IRF1 were induced early in subjects that get vaccinated with BNT162b2 mRNA COVID-19 vaccine and were found to correlate with vaccine-specific humoral protec ve responses. In the a empt to reproduce in vitro an immune-relevant scenario, here we simulated the mRNA vaccine intramuscular administra on in IMMUSC model, a heterologous system composed of vaccine-injectable 3D muscle-like ssue (3D-MT) and peripheral blood mononuclear cells (PBMC), represen ng the immune compartment. Moreover, to dissect stroma contribu on in shaping early immune response to BNT162b2 vaccine, vaccine exposed human primary fibroblasts and PBMC were also used. BNT162b2 vaccine was recognized by all cell types, although only fibroblasts translate the spike an gen at early me point a er smula on. Vaccine-injec on in the 3D-MT smulates the recruitment and phenotypical modula on of major an gen presen ng cells, par cularly monocytes and macrophages while factors from stromal compartment mainly account for dendri c cell expansion and ac va on. By integra ng these in vitro systems, we found that ssue resident muscle cells prompt PBMC to generate an immunosmulatory soluble milieu while stromal compartment specifically equipes PBMC to express IFN-induced innate immune mediators. Our results suggest that cell-based systems composed of PBMC, stromal and ssue specific cells can be exploited to recapitulate the early cytokine and chemokine signature previously iden fied ex vivo in sera of subjects vaccinated with BNT162b2 vaccine. Based on that, an advanced version of IMMUSC model combining muscle cells and fibroblasts in a 3D structure and PBMC is under development with the aim to provide a unique valuable tool that could be employed for in vitro valida on of system vaccinology findings as well as for non-animal down-selec on and pre-clinical tes ng of vaccines following intra-muscular administra on route.