In vitro modeling for the in-depth characterization of mRNA vaccines

Introduction/Rationale: Pre-clinical evaluation of mRNA vaccine formulations requires ranking and de-risking diverse lipid nanoparticles (LNPs). The Modular Immune In vitro Construct Peripheral Tissue Equivalent (MIMIC PTE) system is a state-of-the-art, fully human platform that enables high-throughput evaluation of vaccine reactogenicity within a physiologically relevant environment.

Methods: The MIMIC PTE system is a three-dimensional construct composed of endothelial cells layered on a collagen matrix. Peripheral blood mononuclear cells (PBMCs) from individual human donors are introduced into the system, allowing antigen presenting cells (APCs) to migrate through the endothelial layer. Following vaccine treatment, the construct is incubated to enable cellular uptake, PBMC maturation and differentiation, and cytokine expression. Subsequent analyses include bead-based cytokine assays, flow cytometry, and transcriptomic profiling.

Results: This enhanced system successfully ranks mRNA-LNPs formulations across a variety of ionizable and cationic lipid formulations. Using this platform, we have deepened our understanding of LNP mechanisms of action, identifying a substantial population of cells that produce pro-inflammatory cytokines without expressing the vaccine antigen, indicative of pleiotropic activation. Cell depletion experiments demonstrated that CD14+ and/or CD11b+ myeloid cells, despite representing < 25% of all PBMCs, are the primary drivers of the global cytokine profile elicited by mRNA vaccines. In addition to inflammasome-mediated IL-1 production, we observed robust activation of TLR3 and TLR9 pathways, suggesting potential DAMP signaling via self-nucleic acids.

Conclusion: The fully human MIMIC PTE platform substantially improves our capacity to investigate mechanisms of action and predict vaccine safety profiles within physiologically relevant immune cell populations, thereby accelerating the development of safer, more effective vaccines and mitigating clinical trial risks.