PT  - JOURNAL ARTICLE
AU  - Dimitrios L. Wagner
AU  - Leila Amini
AU  - Desiree J. Wendering
AU  - Petra Reinke
AU  - Hans-Dieter Volk
AU  - Michael Schmueck-Henneresse
TI  - High prevalence of S. pyogenes Cas9-specific T cell sensitization within the adult human population – A balanced effector/regulatory T cell response
AID  - 10.1101/295139
DP  - 2018 Jan 01
TA  - bioRxiv
PG  - 295139
4099  - http://biorxiv.org/content/early/2018/04/04/295139.short
4100  - http://biorxiv.org/content/early/2018/04/04/295139.full
AB  - The field of gene therapy has been galvanized by the discovery of the highly efficient and adaptable site-specific nuclease system CRISPR/Cas9 from bacteria.1,2 Immunity against therapeutic gene vectors or gene-modifying cargo nullifies the effect of a possible curative treatment and may pose significant safety issues.3-5 Immunocompetent mice treated with CRISPR/Cas9-encoding vectors exhibit humoral and cellular immune responses against the Cas9 protein, that impact the efficacy of treatment and can cause tissue damage.5,6 Most applications aim to temporarily express the Cas9 nuclease in or deliver the protein directly into the target cell. Thus, a putative humoral antibody response may be negligible.5 However, intracellular protein degradation processes lead to peptide presentation of Cas9 fragments on the cellular surface of gene-edited cells that may be recognized by T cells. While a primary T cell response could be prevented or delayed, a pre-existing memory would have major impact. Here, we show the presence of a ubiquitous memory/effector T cell response directed towards the most popular Cas9 homolog from Streptococcus pyogenes (SpCas9) within healthy human subjects. We have characterized SpCas9-reactive memory/effector T cells (TEFF) within the CD4/CD8 compartments for multi-effector potency and lineage determination. Intriguingly, SpCas9-specific regulatory T cells (TREG) profoundly contribute to the pre-existing SpCas9-directed T cell immunity. The frequency of SpCas9-reactive TREG cells inversely correlates with the magnitude of the respective TEFF response. SpCas9-specific TREG may be harnessed to ensure the success of SpCas9-mediated gene therapy by combating undesired TEFF response in vivo. Furthermore, the equilibrium of Cas9-specific TEFF and TREG cells may have greater importance in Streptococcus pyogenes-associated diseases. Our results shed light on the T cell mediated immunity towards the much-praised gene scissor SpCas9 and offer a possible solution to overcome the problem of pre-existing immunity.