Reversible switching of primary cells between normal and malignant state by oncogenic virus KSHV and CRISPR/Cas9-mediated targeting of a major viral latent protein.

Viral infection has been implicated in the pathogenesis of a plethora of human diseases. Although antiviral therapies effectively confront the viral spread and infection, how to completely eradicate the viral genome from infected cells remains a challenge. In this study, we demonstrated the reversible switching of primary cells between normal and malignant states by an oncogenic virus Kaposi's sarcoma-associated herpesvirus (KSHV) and CRISPR/Cas9-mediated targeting of a major viral latent protein. Primary cells can be transformed into malignant status by infection of KSHV, while elimination of the KSHV genome from latent KSHV-infected cells reverses KSHV-transformed primary cells back to a "normal state" by CRISPR/Cas-mediated knockout of viral major latent gene LANA. As a proof of concept, we demonstrated efficient elimination of KSHV episome in KSHV-associated primary effusion lymphoma cells resulting in the induction of apoptosis by liposome-encapsulated CRISPR/Cas9 ribonucleoprotein complexes (Lipo/Cas9-LANAsgRNA). Our work illustrates CRISPR/Cas as a promising technology for eliminating oncogenic viruses from persistently infected cells by taking advantage of the genetic differences between viral and cellular genomes. Compared to traditional antiviral therapy, our study offer an approach for antagonizing human oncogenic virus-related cancers by directly targeting as well as clearing viral genomes.