Inhibition of Herpes Simplex Virus Type 1 Infection by Silver Nanoparticles Capped with Mercaptoethane Sulfonate Dana Baram-Pinto, †,‡ Sourabh Shukla, † Nina Perkas, † Aharon Gedanken, † and Ronit Sarid* ,‡ Department of Chemistry and Kanbar Laboratory for Nanomaterials at Bar-Ilan University Center for Advanced Materials and Nanotechnology, Bar-Ilan University, 52900 Ramat-Gan, Israel, and The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, 52900 Ramat-Gan, Israel. Received January 22, 2009; Revised Manuscript Received June 6, 2009 Interactions between biomolecules and nanoparticles suggest the use of nanoparticles for various medical interventions. The attachment and entry of herpes simplex virus type 1 (HSV-1) into cells involve interaction between viral envelope glycoproteins and cell surface heparan sulfate (HS). Based on this mechanism, we designed silver nanoparticles that are capped with mercaptoethane sulfonate (Ag-MES). These nanoparticles are predicted to target the virus and to compete for its binding to cellular HS through their sulfonate end groups, leading to the blockage of viral entry into the cell and to the prevention of subsequent infection. Structurally defined Ag-MES nanoparticles that are readily redispersible in water were sonochemically synthesized. No toxic effects of these nanoparticles on host cells were observed. Effective inhibition of HSV-1 infection in cell culture by the capped nanoparticles was demonstrated. However, application of the soluble surfactant MES failed to inhibit viral infection, implying that the antiviral effect of Ag-MES nanoparticles is imparted by their multivalent nature and spatially directed MES on the surface. Our results suggest that capped nanoparticles may serve as useful topical agents for the prevention of infections with pathogens dependent on HS for entry. INTRODUCTION The application of nanotechnology in therapeutics, biological imaging, drug delivery, biosensors, and cell labeling is being intensively explored (1-6). Biologically important entities, such as proteins, antibodies, antigens, and DNA, which possess nanometer size dimensions, may serve as good candidate targets for interactions with such nanoparticles (1, 7, 8). Viruses are common nanoelement pathogens that can infect and cause diseases in plants, animals, and humans, while viral diseases present challenging problems with worldwide social and economic impli- cations. Development of antiviral drugs able to target the virus while maintaining host cell viability is challenging (9, 10). Herpes simplex virus type 1 (HSV-1) is a common infectious agent that occurs worldwide and infects humans of all ages (11). The outcome of HSV-1 infection includes a wide variety of clinical manifestations, ranging from asymptomatic infection to oral cold sores and severe encephalitis. The HSV-1 virion consists of a 152-kbp double-stranded DNA genome enclosed by an icosahedral capsid, which is surrounded by a lipid bilayer envelope that accommodates 11-12 virally encoded glycopro- teins (11, 12). The envelope diameter ranges from 170 to 200 nm and contains an array of protruding glycoprotein spikes, making the full diameter of the virion about 225 nm on average. Each virion contains 600-750 spikes, with variable packing densities (12). The mechanism of HSV entry into the cell, involving its virus-receptor interaction, is one of the most comprehensively understood routes among members of the Herpesviridae family (13). HSV entry occurs when extracellular virions attach to the cell surface via glycoprotein C (gC) and gB, promoting the binding of gD to one of three alternative cellular receptors. In turn, membrane fusion machinery com- prised of gB, gH, and gL is activated to mediate fusion with plasma or endocytotic membranes (11, 14). During the attach- ment phase, gC and gB interact independently with cellular heparan sulfate (HS). This reversible interaction likely creates multiple points of adhesion and occurs in both wild-type and laboratory viral strains (14). The affinity of the binding of gC to HS is on the order of 10 -8 M and is considered to be the major binding interaction during attachment (13). When gB and gC are absent, viral binding to the cell surface is severely reduced, signifying the important role of this step for viral entry (11, 15). Accordingly, cells that are defective in HS exhibit a dramatic reduction in susceptibility to infection (13). It is worth noting that the interaction with cell surface HS has been found to be a common pathway for attachment by several other human and animal viruses as well (15). HS is a ubiquitous constituent of cell plasma membranes and extracellular matrices, which mediates various physiological processes, such as development, cell adhesion, tumorigenesis, and viral and bacterial infections (16). It is a structurally diverse, highly sulfonated polysaccharide belonging to the family of glycosaminoglycans. HS is composed of alternating sequences of glucosamine and uronic acid. The amino sugars may be N-acetylated or N-sulfated (17-19). The glucosamine residues may, in some cases, carry O-sulfated groups at C6 or C3, and the uronic acid moieties may be O-sulfated at C2 (17). HS is characterized by great structural heterogeneity in terms of chain length and size and the extent of sulfation and epimerization within the modified segments (19). Topical, oral, or intravenous nucleoside derivatives (e.g., acyclovir) have been approved for the treatment of HSV infections and are widely used. However, the emergence of resistant viral strains, mainly after prolonged treatment in immunocompromised patients, is one of the main reasons for the continuous search for new antiviral drugs that can prevent or inhibit infection by both wild-type viruses and drug-resistant strains (13, 20). Compounds that mimic HS, such as sulfated polysaccharide (e.g., dextran sulfate, pentosan polysulfate, a * Corresponding author. Tel: 972-3-5317853. Fax: 972-3-7384058. E-mail: saridr@mail.biu.ac.il. † Department of Chemistry and Kanbar Laboratory for Nanomaterials at Bar-Ilan University Center for Advanced Materials and Nanotechnology. ‡ The Mina and Everard Goodman Faculty of Life Sciences. Bioconjugate Chem. 2009, 20, 1497–1502 1497 10.1021/bc900215b CCC: $40.75  2009 American Chemical Society Published on Web 07/08/2009