Effects of UVA Irradiation, ARYl Azides, and Reactive Oxygen Species on the Viral Membrane: Making Inactivated HIV Detergent Resistant

Abstract

Previously we reported that UV-activatable hydrophobic aryl azides that partition into the hydrophobic regions of the viral membrane of enveloped viruses, inactivate the virus upon UVA irradiation with preservation of surface epitopes (Raviv et al, J. Virol. 2005; Belanger et al. Photochem. Photobiol. 2010). Treatment of HIV-1 with 1,5-diazidonapthalene, 1-iodo, 5-azidonaphthalene, 1-azidonaphthalene or 4,4’-diazidobiphenyl followed by UVA irradiation for 2 minutes resulted in at least 2 logs of viral inactivation, whereas treatment using non-azido controls had no effect. Prolonged UVA irradiation (15 minutes) of the virus with the azido compounds resulted in viral protein aggregation as visualized via Western blot analysis, due to reactive oxygen species (ROS) formation. Treated HIV maintained key surface antigenic structures (recognized by neutralizing antibodies), critical for the creation of efficient vaccines from these inactivated virus preparations. Subsequent treatment of the ROS-modified virus preparation with detergent indicates that the virions are partially detergent resistant. When ROS-modified HIV virus preparations were treated with 1% Triton X-100, there was an increase in the percent of viral proteins (gp41, p24) in the viral pellet after ultracentrifugation through sucrose, as characterized using Western blot, compared to controls. Studies are ongoing to further characterize these preparations using electron microscopy and to extend this method to other enveloped viruses for its use as a novel orthogonally inactivated virus vaccine strategy. Funded in part by NCI Contract HHSN26120080001E.