Identification of Small Molecules with Type I Interferon Inducing Properties by High-Throughput Screening

Abstract

The continuous emergence of virus that are resistant to current anti-viral drugs, combined with the introduction of new viral pathogens for which no therapeutics are available, creates an urgent need for the development of novel broad spectrum antivirals. Type I interferon (IFN) can, by modulating the cellular expression profile, stimulate a non-specific antiviral state. The antiviral and adjuvant properties of IFN have been extensively demonstrated; however, its clinical application has been so far limited. We have developed a human cell-based assay that monitors IFN-β production for use in a high throughput screen. Using this assay we screened 94,398 small molecules and identified 18 compounds with IFN-inducing properties. Among these, 3 small molecules (C3, E51 and L56) showed activity not only in human but also in murine and canine derived cells. We further characterized C3 and showed that this molecule is capable of stimulating an anti-viral state in human-derived lung epithelial cells. Furthermore, the IFN-induction by C3 is not diminished by the presence of influenza A virus NS1 protein or hepatitis C virus NS3/4A protease, which make this molecule an interesting candidate for the development of a new type of broad-spectrum antiviral. In addition, the IFN-inducing properties of C3 also suggest its potential use as vaccine adjuvant.

Dual Myxovirus Screen Identifies a Small-Molecule Agonist of the Host Antiviral Response

“…However, specific activation of innate immune signaling pathways for an antiviral endpoint has experienced a renaissance in recent drug screening campaigns (68)(69)(70). Several of these activities have identified fused heterocyclic compounds with DNA-binding (68) and/or DNA-intercalation activity (70,71), which is inherently associated with high mutagenic potential (72), creating a developmental liability.…”

“…However, specific activation of innate immune signaling pathways for an antiviral endpoint has experienced a renaissance in recent drug screening campaigns (68)(69)(70). Several of these activities have identified fused heterocyclic compounds with DNA-binding (68) and/or DNA-intercalation activity (70,71), which is inherently associated with high mutagenic potential (72), creating a developmental liability. Compound 09167 is structurally distinct from this set of DNA-active small molecules, but it features a thiophene substructure which is potentially reactive (61) and has appeared in low-potency hits (i.e., hits with EC 50 s of 13 to 300 M) in previous screening campaigns (73,74).…”

Genome rearrangement of influenza virus for anti-viral drug screening

“…Given that most methods of anti-viral screening use cell-based assays (Dai et al, 2012; Hossain et al, 2010; Martinez-Gil et al, 2012; Ozawa et al, 2013; Zhang et al, 2011), and that amantadine treatment decreased both luciferase expression and viral titers of Sens/GlucCa04, Sens and Res/GlucCa04 were evaluated in the context of an anti-viral screening assay. Thus, in vitro screening assays were performed in which Sens and Res/GlucCa04 (MOI 0.01 and 0.001) were pre-incubated with increasing concentrations of amantadine and then overlaid on MDCK cells.…”

“…These assays are cell-based and include the generation of stable cell lines (MDCK, Hela, or 293T) expressing influenza driven Renilla luciferase (RLuc) or Firefly luciferase (FLuc) reporter constructs (Hossain et al, 2010; Martinez-Gil et al, 2012; Zhang et al, 2011), and a 293T cell line expressing the viral ribonucleoprotein genes (Ozawa et al, 2013). Importantly, these assays take advantage of luciferase expression that can be quickly assayed from cell lysates.…”

The future of antivirals

“…Nevertheless, 5 0 pppRNA still requires parenteral administration. The use of small molecules that trigger an interferon-response, for example, compound C3 [73], provides an interesting possibility to overcome parenteral administration.…”