The global outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) necessitates the rapid development of new therapies against coronavirus disease 2019 (COVID-19) infection. Here, we present the identification of 200 approved drugs, appropriate for repurposing against COVID-19. We constructed a SARS-CoV-2–induced protein network, based on disease signatures defined by COVID-19 multiomics datasets, and cross-examined these pathways against approved drugs. This analysis identified 200 drugs predicted to target SARS-CoV-2–induced pathways, 40 of which are already in COVID-19 clinical trials, testifying to the validity of the approach. Using artificial neural network analysis, we classified these 200 drugs into nine distinct pathways, within two overarching mechanisms of action (MoAs): viral replication (126) and immune response (74). Two drugs (proguanil and sulfasalazine) implicated in viral replication were shown to inhibit replication in cell assays. This unbiased and validated analysis opens new avenues for the rapid repurposing of approved drugs into clinical trials.

Helium ion microscopy (HIM) offers the opportunity to obtain direct views of biological samples such as cellular structures, virus particles, and microbial interactions. Imaging with the HIM combines sub-nanometer resolution, large depth of field, and high surface sensitivity. Due to its charge compensation capability, the HIM can image insulating biological samples without additional conductive coatings. Here, we present an exploratory HIM study of SARS-CoV-2 infected Vero E6 cells, in which several areas of interaction between cells and virus particles, as well as among virus particles, were imaged. The HIM pictures show the three-dimensional appearance of SARS-CoV-2 and the surface of Vero E6 cells at a multiplicity of infection of approximately 1 with great morphological detail. The absence of a conductive coating allows for a distinction between virus particles bound to the cell membrane and virus particles lying on top of the membrane. After prolonged imaging, it was found that ion-induced deposition of hydrocarbons from the vacuum renders the sample sufficiently conductive to allow for imaging even without charge compensation. The presented images demonstrate the potential of the HIM in bioimaging, especially for the imaging of interactions between viruses and their host organisms.

Viruses with an RNA genome are often the cause of zoonotic infections. In order to identify novel pro-viral host cell factors, we screened a haploid insertion–mutagenized mouse embryonic cell library for clones that are resistant to Rift Valley fever virus (RVFV). This screen returned the low-density lipoprotein receptor–related protein 1 (LRP1) as a top hit, a plasma membrane protein involved in a wide variety of cell activities. Inactivation ofLRP1in human cells reduced RVFV RNA levels already at the attachment and entry stages of infection. Moreover, the role of LRP1 in promoting RVFV infection was dependent on physiological levels of cholesterol and on endocytosis. In the human cell line HuH-7, LRP1 also promoted early infection stages of sandfly fever Sicilian virus and La Crosse virus, but had a minor effect on late infection by vesicular stomatitis virus, whereas encephalomyocarditis virus was entirely LRP1-independent. Moreover, siRNA experiments in human Calu-3 cells demonstrated that also SARS-CoV-2 infection benefitted from LRP1. Thus, we identified LRP1 as a host factor that supports infection by a spectrum of RNA viruses.

Viruses with an RNA genome are the main causes of zoonotic infections. In order to identify novel pro-viral host cell factors, we screened a haploid insertion-mutagenized mouse embryonic cell library for clones that rendered them resistant to the zoonotic Rift Valley fever virus (RVFV; family Phleboviridae, order Bunyavirales). This screen returned the Low Density Lipoprotein Receptor-Related protein 1 (LRP1, or CD91) as top hit, a 600 kDa plasma membrane protein known to be involved in a wide variety of cell activities. Inactivation of LRP1 expression in human cells reduced RVFV infection at the early stages of infection, including the particle attachment to the cell. In the highly LRP1-positive human HuH-7 cell line, LRP1 was required for the early infection stages also of Sandfly fever Sicilian virus (SFSV; family Phleboviridae, order Bunyavirales), vesicular stomatitis (VSV; family Rhabdoviridae, order Mononegavirales), Encephalomyocarditis virus (EMCV, family Picornaviridae), and the coronaviruses MERS-CoV, SARS-CoV-1, and SARS-CoV-2. While for RVFV, EMCV, and MERS-CoV the replication cycle could eventually catch up, LRP1 requirement for the late infection stage in HuH-7 cells was observed for SFSV, La Crosse virus (LACV; family Peribunyaviridae, order Bunyavirales), VSV, SARS-CoV-1, and SARS-CoV-2. For SARS-CoV-2, the absence of LRP1 stably reduced viral RNA levels in human lung Calu-3 cells, and both RNA levels and particle production in the hepatic HuH-7 cells. Thus, we identified LRP1 as a host factor that supports various infection cycle stages of a broad spectrum of RNA viruses.

The global outbreak of SARS-CoV-2 necessitates the rapid development of new therapies against COVID-19 infection. Here, we present the identification of 200 approved drugs, appropriate for repurposing against COVID-19. We constructed a SARS-CoV-2-induced protein (SIP) network, based on disease signatures defined by COVID-19 multi-omic datasets(Bojkova et al., 2020; Gordon et al., 2020), and cross-examined these pathways against approved drugs. This analysis identified 200 drugs predicted to target SARS-CoV-2-induced pathways, 40 of which are already in COVID-19 clinical trials(Clinicaltrials.gov, 2020) testifying to the validity of the approach. Using artificial neural network analysis we classified these 200 drugs into 9 distinct pathways, within two overarching mechanisms of action (MoAs): viral replication (130) and immune response (70). A subset of drugs implicated in viral replication were tested in cellular assays and two (proguanil and sulfasalazine) were shown to inhibit replication. This unbiased and validated analysis opens new avenues for the rapid repurposing of approved drugs into clinical trials.