T cells have seen it all before

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Science Immunology  01 Mar 2019:
Vol. 4, Issue 33, eaax1019
DOI: 10.1126/sciimmunol.aax1019


Two recent studies demonstrate widespread preexisting immunity to Cas9 in healthy adults.

The therapeutic application of CRISPR-Cas9 systems depends on the efficient delivery of Cas9 proteins into human cells using transient or stable expression systems. Two of the most widely used Cas9 proteins derive from Staphylococcus aureus (Sa) and Streptococcus pyogenes (Sp), which are pervasive commensal bacteria and the causative agents in several common infections. Thus far, it has been uncertain whether preexisting cellular and humoral immunity against these bacteria–which is known to exist in the general population–would also confer immunity to Sa and Sp Cas9 proteins. If present, preexisting immunity to Cas9 would complicate the therapeutic application of CRISPR-Cas9 for gene editing in vivo.

Two recent articles by Wagner et al. and Charlesworth et al. have reported the identification of widespread preexisting immunity in humans against both SaCas9 and SpCas9. Using a combination of techniques—including antigen stimulation of peripheral blood mononuclear cells (PBMC), ELISpot, flow cytometry, and serum enzyme-linked immunosorbent assay (ELISA)—these studies report that specific effector T cell and antibody populations to SaCas9 and SpCas9 exist in the peripheral blood of most healthy adults. Wagner et al. identified small populations of effector-memory T cells that become activated (as measured by CD137 up-regulation) in response to stimulation with recombinant SpCas9 protein. These T cell population were present in greater than 95% of the healthy donors studied, the majority of whom also evidenced positive antibody titers against canonical Sp antigens, indicating prior exposure. Using slightly more stringent criteria, Charlesworth et al. reported similar findings and demonstrated specific T cell responses to SpCas9 as well as SaCas9 in 67% and 78% of healthy donors, respectively, and serum immunoglobulin G antibodies against SpCas9 and SaCas9 in 58% and 78% of donors, respectively.

As both studies point out, the presence of preexisting immunity to Cas9 proteins may not have a significant impact on applications that utilize transient delivery of Cas9 ex vivo. In this setting, Cas9-associated peptide antigens would presumably have time to dissipate prior to in vivo use. However, strategies that depend on the stable expression of Cas9 proteins in vivo are more complicated as immune responses in this setting could render the therapy ineffective or trigger immune-mediated tissue damage. Given the promise and rapid pace of development of CRISPR-Cas9 systems, the authors’ findings suggest that close evaluation of immunogenicity should accompany efforts for their therapeutic development in humans.

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