Efficiency and immunogenicity of lipid nanoparticle-mediated cardiac mRNA delivery are lipid composition-dependent.

The efficiency and safety of modified messenger RNA (modRNA) delivery into the heart using lipid nanoparticles (LNPs) remain undetermined. We previously demonstrated that modRNA encapsulated in C12-200 LNPs outperforms current state-of-the-art intramyocardial modRNA delivery methods. Surprisingly, C12-200 LNPs triggered robust local immune cell activation 5 days post-injection, which was not evident on day 1. To investigate whether this immune response is driven by LNP composition or modRNA transfection efficiency, we systematically compared cardiac transfection efficiency, off-target biodistribution, and immunogenicity of modRNA formulated with clinically validated LNPs from Onpattro, BNT162b2/Comirnaty, and mRNA-1273/Spikevax. All tested formulations outperformed C12-200 in cardiac delivery, with mRNA-1273 and Onpattro showing markedly reduced off-target accumulation in the liver and spleen. Histopathological analysis revealed formulation-dependent immune cell infiltration, most pronounced with C12-200. C12-200 significantly elevated cytokine levels in both serum and heart tissue, whereas Onpattro increased cytokine levels mainly locally. In contrast, cytokine levels in animals treated with BNT162b2 and mRNA-1273 were comparable to those in PBS-treated controls. Importantly, cardiac transfection efficiency did not correlate with cytokine induction or histopathological changes, indicating that LNP-driven immunogenicity is independent of transfection efficacy. These findings provide a foundation for refining LNP formulations to optimize cardiac modRNA delivery while minimizing immune-related adverse effects.