New Broad-Spectrum Viral Fusion Inhibitors Act by Deleterious Effect on the Viral Membrane through the Production Singlet Oxygen Molecules

Abstract

Targeting membranes of enveloped viruses represents an exciting new paradigm to explore regarding the development of broad-spectrum antivirals. Recently, broad-spectrum small-molecule antiviral drugs were described, which prevent enveloped virus entry at an intermediate step, after virus binding but before virus-cell fusion [1, 2]. These compounds, an aryl methyldiene rhodanine derivate, named LJ001, and oxazolidine-2,4-dithione derivates named JL103 and JL 118, act deleteriously on the virus envelope but not at the cell membrane level. The aim of the present work was to study the interactions of active compounds (LJ001, JL103 and JL118) and LJ025 (an inactive analog used as negative control) with biological membrane models, in order to clarify the mechanism of action of these new enveloped virus entry inhibitors. Fluorescence spectroscopy was used to quantify the partition and determine the location of the molecules on membranes. The ability of the compounds to produce reactive oxygen molecules in the membrane was tested by using 9,10-dimethylanthracene, which reacts selectively with singlet oxygen. Changes on the lipid packing and fluidity of membranes were assessed by fluorescence anisotropy and generalized polarization measurements. Finally, the ability to inhibit membrane fusion was evaluated using FRET. Our results indicate that singlet oxygen production by LJ001, JL103 and JL118 is able to induce several changes on membrane properties, specially related with a decrease on its fluidity, concomitant with an increase on the order on the polar head groups region, resulting in an inhibition of the membrane fusion necessary for cell infection.1. Wolf et al. (2010) PNAS 107, 3157-62.2. Vigant et al. (2013) PLoS Pathog 9: e1003297.