Targeting herpes simplex virus with CRISPR–Cas9 cures herpetic stromal keratitis in mice

Abstract

Herpes simplex virus type 1 (HSV-1) is a leading cause of infectious blindness. Current treatments for HSV-1 do not eliminate the virus from the site of infection or latent reservoirs in the trigeminal ganglia. Here, we target HSV-1 genomes directly using mRNA-carrying lentiviral particles that simultaneously deliver SpCas9 mRNA and viral-gene-targeting guide RNAs (designated HSV-1-erasing lentiviral particles, termed HELP). We show that HELP efficiently blocks HSV-1 replication and the occurrence of herpetic stromal keratitis (HSK) in three different infection models. HELP was capable of eliminating the viral reservoir via retrograde transport from corneas to trigeminal ganglia. Additionally, HELP inhibited viral replication in human-derived corneas without causing off-target effects, as determined by whole-genome sequencing. These results support the potential clinical utility of HELP for treating refractory HSK.

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Fig. 1: HELP blocks HSV-1 replication in vitro.
Fig. 2: HELP blocks HSV-1 infection of corneas and neurons in a prevention model.
Fig. 3: HELP suppresses HSV-1-associated disease pathologies in the prevention model.
Fig. 4: Eye health after HELP treatment in the prevention model.
Fig. 5: HELP cures HSK in the therapeutic and recurrent models.
Fig. 6: HELP eliminates HSV-1 in tissue culture of human corneas.

Data availability

Data generated or analyzed during this study are available from the corresponding author on reasonable request. The deep-sequencing and whole-genome sequencing data are available at NCBI BioProject. The BioProject IDs are PRJNA668071 and PRJNA668060, respectively. Source data are provided with this paper.

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Acknowledgements

We thank F. Zhang (MIT, USA) for reading and commenting on our manuscript. The work was supported by grants from the National Natural Science Foundation of China (31971364), the Pujiang Talent Project of Shanghai (18PJ1404500), the Natural Science Foundation of Shanghai (18ZR1419300) and startup funding from the Shanghai Center for Systems Biomedicine, Shanghai Jiao Tong University (WF220441504) to Y.C. and by the National Natural Science Foundation of China (81970766 and 81670818), the Shanghai Rising-Star Program (18QA1401100), the Shanghai Innovation Development Program (2020779) and the Shanghai Key Clinical Research Program (SHDC2020CR3052B) to J.H. S.R.P. is supported by the European Research Council (ERC-AdG ENVISION; 786602).

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D.Y., S.L., J.H. and Y.C. conceptualized the study and designed the experiments; D.Y., S.L., D.W., Y.D., H.J. and X.Z. performed the experiments; S.R.P. provided the HSV-1 strains and facilitated building the mouse HSK model; all the authors analyzed the data; D.Y., S.L. and Y.C. wrote the manuscript with help from all the authors.

Corresponding authors

Correspondence to Jiaxu Hong or Yujia Cai.

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Peer review information Nature Biotechnology thanks Paul R. Kinchington and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Yin, D., Ling, S., Wang, D. et al. Targeting herpes simplex virus with CRISPR–Cas9 cures herpetic stromal keratitis in mice. Nat Biotechnol (2021). https://doi.org/10.1038/s41587-020-00781-8

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