Applicability of conventional genotoxicity test guidelines to mRNA vaccines: a four-battery assessment of a SARS-CoV-2 mRNA vaccine candidate

Abstract mRNA vaccine formulations are generally considered to pose a lower genotoxic risk than conventional small-molecule pharmaceuticals; however, long-term data regarding their potential genotoxicity remain limited. Furthermore, to date, no standardized genotoxicity testing guidelines specifically tailored to the unique characteristics of mRNA vaccines have been formally established. In this study, we conducted a comprehensive genotoxicity assessment of a SARS-CoV-2 Omicron variant mRNA vaccine candidate using a four-test battery, following a modified regimen that accounted for the unique characteristics of mRNA vaccines while adhering to OECD test guidelines: bacterial reverse mutation test (OECD TG 471), in vitro chromosomal aberration test (OECD TG 473), in vitro micronucleus test (OECD TG 487), and in vivo erythrocyte micronucleus test in mice (OECD TG 474). No significant genotoxic effects of the mRNA vaccine candidate were observed in any of the four assays. However, during the design and conduct of this study in accordance with existing genotoxicity guidelines, ambiguity arose as to whether test concentrations should be determined according to the mRNA component or the LNP component. In addition, we observed that, unlike the LNP administered alone, the mRNA vaccine candidate was associated with a reduction in the proportion of PCEs in the in vivo micronucleus assay. Although our mRNA vaccine candidate showed no significant evidence of genotoxicity, the experimental design and dosing regimen highlight the need to refine current regulatory testing guidelines for mRNA vaccines. We anticipate that our findings will contribute to the development of more appropriate genotoxicity guidelines for advanced therapy medicinal products (ATMPs), including mRNA vaccines.