Phosphorothioate Oligonucleotides Inhibit Human Immunodeficiency Virus Type 1 Fusion by Blocking gp41 Core Formation

Abstract

Several studies have shown that phosphorothioate oligodeoxynucleotides (PS-ONs) have a sequence-independent antiviral activity against human immunodeficiency virus type 1 (HIV-1). It has also been suggested that PS-ONs inhibit HIV-1 by acting as attachment inhibitors that bind to the V3 loop of gp120 and prevent the gp120-CD4 interaction. Here we show that PS-ONs (and their fully 2-O-methylated derivatives) are potent inhibitors of HIV-1-mediated membrane fusion and HIV-1 replication in a size-dependent, phosphorothioation-dependent manner. PS-ONs interact with a peptide derived from the N-terminal heptad repeat region of gp41, and the HIV-1 fusion-inhibitory activity of PS-ONs is closely correlated with their ability to block gp41 six-helix bundle formation, a critical step during the process of HIV-1 fusion with the target cell. These results suggest that the increased hydrophobicity of PS-ONs may contribute to their inhibitory activity against HIV-1 fusion and entry, because longer PS-ONs (>30 bases) which have a greater hydrophobicity are more potent in blocking the hydrophobic interactions involved in the gp41 six-helix bundle formation and inhibiting the HIV-1-mediated cell-cell fusion than shorter PS-ONs (<30 bases). This novel antiviral mechanism of action of long PS-ONs has implications for therapy against infection by HIV-1 and other enveloped viruses with type I fusion proteins.Several studies have suggested that the antiviral activity of phosphorothioated oligonucleotides (PS-ONs) with different sequences or different intended mechanisms of action (i.e., antisense or sequence-specific aptamers) all work as inhibitors of human immunodeficiency virus (HIV) viral entry by binding to the V3 loop of gp120 and preventing CD4 interaction and attachment to the host cell (13, 43). The general conservation of this mechanism of action with different oligonucleotides having different sequences suggested that the antiviral activity of oligonucleotides against HIV was sequence independent.The sequence-independent nature of the antiviral activity of oligonucleotides (ONs) was more clearly demonstrated in several studies using degenerate or homopolymeric ONs (24,30,37). In these studies, the anti-HIV activities of 28-mer phosphorothioated polycytidylic, polyadenylic, and polythimidylic or fully degenerate ONs were studied in vitro, and in the case of polycytidylic ONs, it was also discovered that for ONs up to 28 bases in length increasingly larger ONs displayed more potent antiviral activity. The binding of 28-mer polycytidylic ONs to the V3 loop of gp120 was also described (38).PS-ONs are also polyanionic, and the antiviral activities of a variety of other classes of polyanions, such as sulfated saccharides, polysulfonates, polyhydroxycarboxylates, and polyoxometalates, have also been well established (11,28,31,34,35).There is good evidence to suggest that these compounds also interfere with viral attachment by binding to the V3 loop of gp120 (2,12,33,40).To further explore the sequence-independent mechanisms of t...

Nucleic Acid Polymers Prevent the Establishment of Duck Hepatitis B Virus Infection In Vivo

“…In the case of viral entry, NAP-protein interactions destroy the ability of amphipathic protein domains present in viral fusion glycoproteins to interact with each other, a process known to be important in catalyzing the entry of many enveloped viruses into their host cells (15). In HIV-1 and LCMV, the interaction of NAPs with viral fusion proteins is consistent with their ability to block viral entry, and they have little or no postentry antiviral activity (13,14).…”

“…For example, NAPs have been shown to interact with prion proteins (12) and type 1 viral fusion glycoproteins in both human immunodeficiency virus type 1 (HIV-1) and lymphocytic choriomeningitis virus (LCMV) (13,14). In the case of viral entry, NAP-protein interactions destroy the ability of amphipathic protein domains present in viral fusion glycoproteins to interact with each other, a process known to be important in catalyzing the entry of many enveloped viruses into their host cells (15).…”

Phosphorothioate-Modified TLR9 Ligands Protect Cancer Cells against TRAIL-Induced Apoptosis

“…binding inhibition. This phenomenon could be extended to the inhibition of virus entry, as it has been recently shown that human cytomegalovirus, hepatitis C virus, and HIV entry are blocked by phosphorothioate-modified ODNs, regardless of CpG motifs (33)(34)(35)(36). One could therefore take advantage of this to design antiviral vaccines that target virus entry.…”

Azasugar-Containing Phosphorothioate Oligonucleotide (AZPSON) DBM-2198 Inhibits Human Immunodeficiency Virus Type 1 (HIV-1) Replication by Blocking HIV-1 gp120 without Affecting the V3 Region

“…In both cases, the tetrameric G-quartet structure and the sulfur-containing backbone are believed to be involved in binding to V3, followed by the inhibition of virus-to-cell and cell-to-cell infections. Another study showed that P = S ONs interact with a peptide derived from the N-terminal heptad repeat region of gp41, blocking gp41 sixhelix bundle formation (Vaillant et al, 2006). Recently, a short 14-mer P = S 2′ deoxyribose backbone was demonstrated to enhance anti-HIV-1 activity by blocking Toll-like receptor 7 (TLR7) and TLR9, thus inhibiting the initial spread of HIV-1 after infection (Fraietta et al, 2010).…”

“…These suggested mechanisms include blocking of viral adsorption (Luganini et al, 2008;Wyatt et al, 1994) and/or inhibition of HIV-1-specific enzymes, such as reverse transcriptase (Marshall and Caruthers, 1993;Marshall et al, 1992) or integrase (Jing and Hogan, 1998;Jing et al, 2000). P = S oligodeoxycytidine [poly (SdC) 28 ] interacts specifically with the positively-charged V3 loop of HIV-1 gp120 (Vaillant et al, 2006), possibly resulting in the inhibition of HIV-1 replication. It was also reported that T2G4T2 (5-TTGGGGTT-3; ISIS-5320), a P = S ON identified as an HIV-1 inhibitor, forms a tetrameric G-quartet structure and binds to the cationic V3 loop region of the gp120 viral envelope glycoprotein , as does poly(SdC) 28 .…”