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Identification of Pre-Existing Adaptive Immunity to Cas9 Proteins in Humans

Charlesworth, Carsten T., Deshpande, Priyanka S., Dever, Daniel P., Dejene, Beruh, Gomez-Ospina, Natalia, Mantri, Sruthi, Pavel-Dinu, Mara, Camarena, Joab, Weinberg, Kenneth I., Porteus, Matthew H.·2018
DOI 10.1101/243345

Preprint — published as 10.1038/s41591-018-0326-x · Paper page

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Abstract

The CRISPR-Cas9 system has proven to be a powerful tool for genome editing, allowing for the precise modification of specific DNA sequences within a cell. Many efforts are currently underway to use the CRISPR-Cas9 system for the therapeutic correction of human genetic diseases. The most widely used homologs of the Cas9 protein are derived from the bacteria Staphylococcus aureus ( S. aureus ) and Streptococcus pyogenes ( S. pyogenes ). Based on the fact that these two bacterial species cause infections in the human population at high frequencies, we looked for the presence of pre-existing adaptive immune responses to their respective Cas9 homologs, SaCas9 ( S. aureus homolog of Cas9) and SpCas9 ( S. pyogenes homolog of Cas9). To determine the presence of anti-Cas9 antibodies, we probed for the two homologs using human serum and were able to detect antibodies against both, with 79% of donors staining against SaCas9 and 65% of donors staining against SpCas9. Upon investigating the presence of antigen-specific T-cells against the two homologs in human peripheral blood, we found anti-SaCas9 T-cells in 46% of donors. Upon isolating, expanding, and conducting antigen re-stimulation experiments on several of these donors’ anti-SaCas9 T-cells, we observed an SaCas9-specific response confirming that these T-cells were antigen-specific. We were unable to detect antigen-specific T-cells against SpCas9, although the sensitivity of the assay precludes us from concluding that such T-cells do not exist. Together, this data demonstrates that there are pre-existing humoral and cell-mediated adaptive immune responses to Cas9 in humans, a factor which must be taken into account as the CRISPR-Cas9 system moves forward into clinical trials.

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Metadata

Authors
Charlesworth, C. T., Deshpande, P. S., Dever, D. P., Dejene, B., Gomez-Ospina, N., Mantri, S., Pavel-Dinu, M., Camarena, J., Weinberg, K. I., & Porteus, M. H.
Publisher
openRxiv
Published
2018-01-05
DOI
10.1101/243345
Cited by
71
References
0
Type
Preprint

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CoChat. (2026). Citation check: Identification of Pre-Existing Adaptive Immunity to Cas9 Proteins in Humans. https://app.cochat.ai/papers/10.1101/243345
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Charlesworth, C. T., Deshpande, P. S., Dever, D. P., Dejene, B., Gomez-Ospina, N., Mantri, S., Pavel-Dinu, M., Camarena, J., Weinberg, K. I., & Porteus, M. H. (2018). Identification of Pre-Existing Adaptive Immunity to Cas9 Proteins in Humans. https://doi.org/10.1101/243345