Inhibition of Monkeypox virus replication by RNA interference

Abstract

The Orthopoxvirus genus of Poxviridae family is comprised of several human pathogens, including cowpox (CPXV), Vaccinia (VACV), monkeypox (MPV) and Variola (VARV) viruses. Species of this virus genus cause human diseases with various severities and outcome ranging from mild conditions to death in fulminating cases. Currently, vaccination is the only protective measure against infection with these viruses and no licensed antiviral drug therapy is available. In this study, we investigated the potential of RNA interference pathway (RNAi) as a therapeutic approach for orthopox virus infections using MPV as a model. Based on genome-wide expression studies and bioinformatic analysis, we selected 12 viral genes and targeted them by small interference RNA (siRNA). Fortyeight siRNA constructs were developed and evaluated in vitro for their ability to inhibit viral replication. Two genes, each targeted with four different siRNA constructs in one pool, were limiting to viral replication. Seven siRNA constructs from these two pools, targeting either an essential gene for viral replication (A6R) or an important gene in viral entry (E8L), inhibited viral replication in cell culture by 65-95% with no apparent cytotoxicity. Further analysis with wild-type and recombinant MPV expressing green fluorescence protein demonstrated that one of these constructs, siA6-a, was the most potent and inhibited viral replication for up to 7 days at a concentration of 10 nM. These results emphasis the essential role of A6R gene in viral replication, and demonstrate the potential of RNAi as a therapeutic approach for developing oligonucleotidebased drug therapy for MPV and other orthopox viruses.

Control of African swine fever virus replication by small interfering RNA targeting the A151R and VP72 genes

“…At 24 h post-transfection of siRNAs against G7L, the VACV titre produced after infection with 0.1 MOI and incubation over 24 h was reduced by 99% [32], which is 1-2 log 10 values less than the reduction achieved in our work. By contrast, a fourth study targeting the A6R gene of Monkeypox virus showed an almost complete inhibition of virus replication when specific siRNAs were delivered at 40 nM, 18 h before infection at 0.1 MOI [33]. The antiviral therapeutic effect of siRNAs against 0.0001 MOI of VACV at 1, 8 and 24 h post-infection showed a reduction of VACV replication of 92.0%, 72.3% and 74.2% respectively [30].…”

State-of-the-Art Therapeutic Medical Countermeasures for Viral Threat Agents

“…21 RNA interference (RNAi) has been evaluated as a potential therapy for orthopox viruses. 22 Using monkeypox (MPX) as a model system, 48 siRNA constructs were screened in vitro against genes essential for viral replication or entry. These constructs were able to inhibit viral expression by 65% to 95% without apparent signs of cytotoxicity.…”

RNA interference and its therapeutic potential

“…In animals, the first evidence that dsRNA could lead to gene silencing came from work in Caenorhabditis elegans [30]. They found gene silencing in their control set by using sense strand and it was quite an unusual result in comparison to the antisense technique [1,30,59]. In 1998, Fire and Mello recognized the fact that the dsRNA is responsible for endogenous gene silencing.…”

“…Monkeypox or other orthopox viruses poses a serious threat to public health and there are no licensed drugs to treat these infections [18]. The potential role of RNAi as a therapeutic approach for monkeypox virus infections using MPV as a model has been reported [1].…”