A cell-based, infectious-free, platform to identify inhibitors of lassa virus ribonucleoprotein (vRNP) activity

Abstract

A B S T R A C TThe mammarenavirus Lassa (LASV) is highly prevalent in West Africa where it infects several hundred thousand individuals annually resulting in a high number of Lassa fever (LF) cases, a febrile disease associated with high morbidity and significant mortality. Mounting evidence indicates that the worldwide-distributed prototypic mammarenavirus lymphocytic choriomeningitis virus (LCMV) is a neglected human pathogen of clinical significance. There are not Food and Drug Administration (FDA) licensed vaccines and current anti-mammarenavirus therapy is limited to an off-label use of ribavirin that is only partially effective and can cause significant side effects. Therefore, there is an unmet need for novel antiviral drugs to combat LASV. This task would be facilitated by the implementation of high throughput screens (HTS) to identify inhibitors of the activity of the virus ribonucleoprotein (vRNP) responsible for directing virus RNA genome replication and gene transcription. The use of live LASV for this purpose is jeopardized by the requirement of biosafety level 4 (BSL4) containment. We have developed a virus-free cell platform, where expression levels of reporter genes serve as accurate surrogates of vRNP activity, to develop cell-based assays compatible with HTS to identify inhibitors of LASV and LCMV mammarenavirus vRNP activities.

RNA-Dependent RNA Polymerase as a Target for COVID-19 Drug Discovery

“… 56 Similarly, for Lassa virus, the minimal viral trans-acting factors required for genome replication are the L protein, which contains the RdRP activity, and the NP. 58 These proteins were stably expressed in a cell line along with Gaussia luciferase and green fluorescent protein (GFP) reporters. 58 …”

“… 58 These proteins were stably expressed in a cell line along with Gaussia luciferase and green fluorescent protein (GFP) reporters. 58 …”

Drug Discovery Strategies for SARS-CoV-2

“…These nonviral assays used for viral entry or replication are not infectious and can be used in a BSL-2 facility for screening large compound collections. This strategy has been used to screen compounds for BSL-3/4 viruses such as EBOV ( Tscherne et al, 2010 ; Kouznetsova et al, 2014 ), Lassa virus ( Cubitt et al, 2020 ), SARS, and MERS-CoV ( de Wilde et al, 2014 ; Dyall et al, 2014 ). Recently, Letko et al (2020) showed chimeric S proteins containing RBD of SARS-CoV-2 can confer receptor specificity to the full S protein sequence.…”

The Nonribosomal Peptide Valinomycin: From Discovery to Bioactivity and Biosynthesis

“…As a result, most of the oligosaccharides in VSV G protein were not converted to their mature forms, leading to the failure of transport of G protein to the cell surface and its further incorporation into budding viral particles [ 42 ]. In another way, valinomycin inhibited the activity of the viral ribonucleoprotein (vRNP) that is responsible for directing viral RNA genome replication and gene transcription [ 62 ]. In addition, Sandler et al [ 57 ] also reported that valinomycin significantly blocked virus replication and reduced the number of viral genomes by >90%.…”