Mutational Pathways, Resistance Profile, and Side Effects of Cyanovirin Relative to Human Immunodeficiency Virus Type 1 Strains with N-Glycan Deletions in Their gp120 Envelopes

Abstract

Limited data are available on the genotypic and phenotypic resistance profile of the ␣-(1-2)mannose oligomer-specific prokaryotic lectin cyanovirin (CV-N). Therefore, a more systematic investigation was carried out to obtain a better view of the interaction between CV-N and human immunodeficiency virus type 1 (HIV-1) gp120. When HIV-1-infected CEM cell cultures were exposed to CV-N in a dose-escalating manner, a total of eight different amino acid mutations exclusively located at N-glycosylation sites in the envelope surface gp120 were observed. Six of the eight mutations resulted in the deletion of high-mannose type N-glycans (i.e., at amino acid positions 230, 332, 339, 386, 392, and 448). Two mutations (i.e., at position 136 and 160) deleted a complex type N-glycan in the variable V1/V2 domain of gp120. The level of phenotypic resistance of the mutated virus strains against CV-N generally correlated with the number of glycan deletions in gp120, although deletion of the glycans at N-230, N-392, and N-448 generally afforded a more pronounced CV-N resistance than other N-glycan deletions. However, the extent of the decrease of antiviral activity of CV-N against the mutated virus strains was markedly less pronounced than observed for ␣(1-3)-and ␣(1-6)-mannose-specific plant lectins Hippeastrum hybrid agglutinin (HHA) and Galanthus nivalis agglutinin (GNA), which points to the existence of a higher genetic barrier for CV-N. This is in agreement with a more consistent suppression of a wider variety of HIV-1 clades by CV-N than by HHA and GNA. Whereas the antiviral and in vitro antiproliferative activity of CV-N can be efficiently reversed by mannan, the pronounced mitogenic activity of CV-N on peripheral blood mononuclear cells was unaffected by mannan, indicating that some of the observed side effects of CV-N are unrelated to its carbohydrate specificity/activity.

Cyanovirin-N potently inhibits human immunodeficiency virus type 1 infection in cellular and cervical explant models

“…The activity of CV-N against a broad range of primary isolates from different clades confirms previous studies and validates the activity of the CV-N used in this study (Balzarini et al, 2006;Boyd et al, 1997). Interestingly, activity seemed higher against primary strains than against laboratory isolates, a positive trait for a potential microbicide candidate.…”

“…The anti-HIV activity of CV-N was then assayed against different HIV-1 primary isolates, representative of clades A, B, C, D, E and O, in PBMC and monitored by detection of p24 antigen release. CV-N demonstrated potent activity against all isolates, with IC 50 values ranging from 0.002 to 0.031 mM (Table 1), in agreement with previous studies (Balzarini et al, 2006;Boyd et al, 1997).…”

“…The compound was previously tested on PBMCs for its cytotoxic effects by a MTT dye reduction assay (CC 50 value 1.63 mM, data not shown) and used at non-toxic concentrations. As previously demonstrated (Balzarini et al, 2006), CV-N has some mitogenic activity after 3 days of incubation with PBMCs, with a maximal stimulation index (SI) of 5 (Fig. 6a).…”

“…Interestingly, activity seemed higher against primary strains than against laboratory isolates, a positive trait for a potential microbicide candidate. The ability of this candidate to inactivate a wide spectrum of laboratory strains and clinical primary isolates underlines the conserved nature of CV-N binding sites on gp120 and the high genetic barrier of this compound, already demonstrated with drug-resistance selection studies (Balzarini et al, 2006;Hu et al, 2007;Witvrouw et al, 2005).…”

“…Thus potent antiviral activity, lack of toxicity, resistance to physico-chemical denaturation as well as the unusual high genetic barrier to resistance (Balzarini et al, 2006;Hu et al, 2007;Witvrouw et al, 2005) suggest that CV-N is a promising anti-HIV molecule for use as a topical microbicide (Boyd et al, 1997;Esser et al, 1999).…”