Editors' ChoiceGenome Editing

Immune-orthogonal orthologs: The solution for genome editing?

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Science Translational Medicine  14 Aug 2019:
Vol. 11, Issue 505, eaay7701
DOI: 10.1126/scitranslmed.aay7701

Abstract

Immune-orthogonal Cas9 allows for genome editing in pre-immunized mice.

In recent years, the number of protein therapeutics reaching the clinic increased dramatically. One important limitation of this class of therapeutics is that they are a possible target for the host adaptive immune system. Both pre-existing immunity and treatment-induced immune responses can potentially reduce the efficacy of the treatment.

CRISPR-CAS9 (clustered regularly interspaced short palindromic repeats–CRISPR-associated protein 9) revolutionized the genome-editing field due to its efficacy and the high-throughput capacity of this technology. A clinical trial using CRISPR-Cas9–mediated genome editing started early this year, and more are coming. However, concerns remain for the potential immunogenicity of the Cas9 protein in humans—in particular, for in vivo applications in which adeno-associated virus (AAV) vectors mediate the delivery of the nuclease. Recently, pre-existing immunity to Cas9 proteins was reported in humans by independent groups. Fortunately, several variants (orthologs) of Cas9 derived from different bacterial species are found in nature.

With these considerations in mind, Moreno and colleagues analyzed the immune-orthogonality of variants of Cas9 and AAV by predicting the binding of their respective amino acid sequences to major histocompatibility complex (MHC) class I and II. By using both Cas9 and AAV immune-orthogonal orthologs, they demonstrated efficient genome editing in animals pre-immunized for the Cas9 protein. Interestingly, the pre-immunization resulted only in a partial reduction of the editing rate, suggesting a limited impact of the anti-Cas9 adaptive immune response in vivo.

One important limitation of the study, however, is the use of inbred mice with a limited MHC repertoire compared with humans. As admitted by the authors, the use of immune-orthogonal orthologs is likely to be a poor solution to reduce anti-capsid immunogenicity in humans because of the high degree of conservation between the different AAV capsids and the presence of numerous MHC-encoding genes in the human populations. The use of Cas9 in vivo may present similar limitations. However, the evolution of Cas9 in bacteria has a longer history, and new Cas9 variants are isolated every month from different bacterial strains. Another possible solution to reduce the risk of developing an adaptive immune response against Cas9 is transient expression of the nuclease achieved by mRNA or protein administration.

The immune orthogonality of the Cas9 proteins existing in nature seems to provide a possible solution to reduce the impact of both pre-existing and post-treatment immunity on genome editing efficacy. If confirmed in animal models with a larger MHC repertoire, this approach may allow for the application of genome editing for the modification of somatic cells and in vivo correction of genetic mutations in humans.

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