AAV-CRISPR Gene Editing Is Negated by Pre-existing Immunity to Cas9

Adeno-associated viral (AAV) vectors are a leading candidate for the delivery of CRISPR-Cas9 for therapeutic genome editing in vivo. However, AAV-based delivery involves persistent expression of the Cas9 nuclease, a bacterial protein. Recent studies indicate a high prevalence of neutralizing antibod...

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Bibliographic Details
Published in:Molecular therapy 2020-06, Vol.28 (6), p.1432-1441
Main Authors: Li, Ang, Tanner, Mark R., Lee, Ciaran M., Hurley, Ayrea E., De Giorgi, Marco, Jarrett, Kelsey E., Davis, Timothy H., Doerfler, Alexandria M., Bao, Gang, Beeton, Christine, Lagor, William R.
Format: Article
Language:English
Subjects:
AAV-CRISPR
adeno-associated virus
Animals
Biomarkers
CD8+ T cell
Clustered Regularly Interspaced Short Palindromic Repeats
CRISPR-Associated Protein 9 - adverse effects
CRISPR-Associated Protein 9 - immunology
Dependovirus - genetics
Gene Editing - methods
Gene Expression
Gene Order
gene therapy
Genetic Vectors - genetics
hepatocytes
Hepatocytes - metabolism
Humans
immune response
Immunization
Immunologic Memory
Immunophenotyping
liver
Mice
Original
pre-existing immunity
RNA, Guide, CRISPR-Cas Systems
SaCas9
somatic genome editing
T-Lymphocyte Subsets - immunology
T-Lymphocyte Subsets - metabolism
Transgenes
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Summary:Adeno-associated viral (AAV) vectors are a leading candidate for the delivery of CRISPR-Cas9 for therapeutic genome editing in vivo. However, AAV-based delivery involves persistent expression of the Cas9 nuclease, a bacterial protein. Recent studies indicate a high prevalence of neutralizing antibodies and T cells specific to the commonly used Cas9 orthologs from Streptococcus pyogenes (SpCas9) and Staphylococcus aureus (SaCas9) in humans. We tested in a mouse model whether pre-existing immunity to SaCas9 would pose a barrier to liver genome editing with AAV packaging CRISPR-Cas9. Although efficient genome editing occurred in mouse liver with pre-existing SaCas9 immunity, this was accompanied by an increased proportion of CD8+ T cells in the liver. This cytotoxic T cell response was characterized by hepatocyte apoptosis, loss of recombinant AAV genomes, and complete elimination of genome-edited cells, and was followed by compensatory liver regeneration. Our results raise important efficacy and safety concerns for CRISPR-Cas9-based in vivo genome editing in the liver. [Display omitted] Li and colleagues demonstrated that pre-existing immunity to Cas9 poses a significant barrier to liver-directed genome editing with AAV-CRISPR. They found that AAV expression of Cas9 elicited a robust CD8+ T cell response, resulting in elimination of edited hepatocytes. This raises important safety and efficacy concerns for in vivo editing with CRISPR-Cas9.
ISSN:1525-0016
1525-0024
DOI:10.1016/j.ymthe.2020.04.017