Pre-existing adaptive immunity to the RNA-editing enzyme Cas13d in humans

RNA-guided RNA-targeting nucleases, such as CRISPR-Cas13 proteins, have therapeutic potential for gene editing. Among Cas13d enzymes, Cas13d from the bacteria Ruminococcus flavefaciens (RfxCas13d) is of particular interest owing to its small size and high specificity. However, the existence of pre-existing immunity against RfxCas13d is unclear. In this study, we evaluated antibody and T cell responses to RfxCas13d in healthy donors using ELISA and T cell culture assays. We found RfxCas13d-reactive antibodies and CD4 and CD8 T cell responses in most donors, comparable to responses against Cas9 proteins from Staphylococcus aureus (SaCas9) and Streptococcus pyogenes (SpCas9). RfxCas13d-responding T cells could produce the inflammatory cytokines IFN-gamma, TNF-alpha and IL-17. These findings should be taken into consideration in the development of RfxCas13d for therapy. Healthy individuals have antibodies and T cells that are reactive to the Cas13d protein from the bovine bacteria Ruminococcus flavefaciens, which may have implications for clinical testing of CRISPR-Cas13 gene editing approaches.

Automated summary

This study evaluated antibody and T cell responses to the Cas13d protein (RfxCas13d) from Ruminococcus flavefaciens in healthy donors. It found that most donors have antibodies and CD4 and CD8 T cell responses to RfxCas13d, similar to other Cas9 proteins. In addition, RfxCas13d-responsive T cells could produce inflammatory cytokines. These findings are important for the development of RfxCas13d for therapy.