Potent programmable antiviral against dengue virus in primary human cells by Cas13b RNP with short spacer and delivery by virus-like particle

Abstract

SummaryWith sequencing as a standard frontline protocol to identify emerging viruses such zika virus and SARS-CoV2, direct utilization of sequence data to program antivirals against the viruses could accelerate drug development to treat their infections. CRISPR-Cas effectors are promising candidates that could be programmed to inactivate viral genetic material based on sequence data but several challenges such as delivery and design of effective crRNA need to be addressed to realize practical use. Here, we showed that virus-like particle (VLP) could deliver PspCas13b-crRNA ribonucleoprotein (RNP) in nanomolar range to efficiently suppress dengue virus infection in primary human target cells. Shortening spacer length could significantly enhance RNA-targeting efficiency of PspCas13b in mammalian cells compared to the natural length of 30 nucleotides without compromising multiplex targeting by a crRNA array. Our results demonstrate the potentials of applying PspCas13b RNP to suppress RNA virus infection, with implications in targeting host RNA as well.

A comprehensive analysis and resource to use CRISPR-Cas13 for broad-spectrum targeting of RNA viruses

“…2 A few researchers also reported that CRISPR-Cas13a/b systems can be used to inhibit RNA viruses, including lymphocytic choriomeningitis virus (LCMV), vesicular stomatitis virus (VSV), human immunodeficiency virus type 1 (HIV-1), human respiratory syncytial virus (HRSV), Dengue virus, Turnip mosaic virus, and tobacco turnip mosaic RNA virus (TuMV). [3][4][5][6][7][8][9] The CRISPR-Cas13 systems (sub-types a, b, and d) offer attractive prophylactic or therapeutic options due to 2 key features: (1) CRISPR-Cas13 is highly programmable, which allows it to be rapidly adapted against new RNA viruses by designing the CRISPR RNA (crRNA) sequence 5,9 and (2) crRNAs can be designed to target highly conserved viral sequences to cover multiple strains and prevent viral mutational escape. 10 One remaining question is whether the system can be expanded to target a broad spectrum of RNA viruses.…”