CRISPR/Cas9-mediated exploration and disruption of herpes simplex virus type 1

Abstract

Herpesviruses can cause lifelong infections in almost 100% of the population. During infection, the virus can enter a "dormant" phase, also known as latency, in which it produces viral proteins to a very limited extent. This latent virus can therefore not be properly recognized by our immune system and as a result we are unable to clear these viruses from our body. There are no drugs available that can recognize and eliminate these dormant viruses. In this thesis, we try to eliminate herpesvirus infections with new techniques. We usurped an anti-viral system from bacteria: CRISPR/Cas9 to specifically inactivate genomes of human herpes viruses. This approach has proven to be highly effective at eliminating virus infections from various herpes viruses, including Epstein-Barr virus (EBV), human Herpes Simplex virus (HSV-1) and cytomegalovirus (CMV). When using multiple anti-herpesvirus CRISPR we were capable of fully blocking replication of HSV-1. In follow-up experiments we have compared the power of this new technique with the treatment method as it is used in the clinic for treatment against HSV-1. This showed that CRISPR/Cas9 was better able to combat HSV-1 infections and to reduce the formation of medication-avoiding mutants. These findings are important because it offers the possibility to further develop anti-HSV-1 CRISPR/Cas9 into a future treatment against HSV-1. Since the virus, in addition to cold sores, can cause recurring eye infections resulting in vision loss, a healing treatment could have a major medical and societal impact. In addition to directly inactivating viral genomes through CRISPR/Cas9, the technique can also be used to identify, host factors involved in the infection of a virus. Through large genome-wide screens, we found human genes that viruses use in the infection process. In addition to HSV-1, we performed a similar screen for the "MVA" smallpox virus and identified human genes used by the viruses in the infection process. The identification of such genes provides more insight into the interaction between host and virus, and can also provide tools for the targeted development of new treatment methods to prevent or treat virus infections in the future.