Resistance of Human Immunodeficiency Virus Type 1 to the High-Mannose Binding Agents Cyanovirin N and Concanavalin A

ABSTRACT Due to the biological significance of the carbohydrate component of the human immunodeficiency virus type 1 (HIV-1) glycoproteins in viral pathogenesis, the glycosylation step constitutes an attractive target for anti-HIV therapy. Cyanovirin N (CV-N), which specifically targets the high-mannose (HM) glycans on gp120, has been identified as a potent HIV-1 entry inhibitor. Concanavalin A (ConA) represents another mannose-binding lectin, although it has a lower specificity for HM glycans than that of CV-N. For the present study, we selected CV-N- and ConA-resistant HIV-1 strains in the presence of CV-N and ConA, respectively. Both resistant strains exhibited a variety of mutations eliminating N-linked glycans within gp120. Strains resistant to CV-N or ConA displayed high levels of cross-resistance towards one another. The N-glycan at position 302 was eliminated in both of the lectin-resistant strains. However, the elimination of this glycan alone by site-directed mutagenesis was not sufficient to render HIV-1 resistant to CV-N or ConA, suggesting that HIV-1 needs to mutate several N-glycans to become resistant to these lectins. Both strains also demonstrated clear cross-resistance towards the carbohydrate-dependent monoclonal antibody 2G12. In contrast, the selected strains did not show a reduced susceptibility towards the nonlectin entry inhibitors AMD3100 and enfuvirtide or towards reverse transcriptase or protease inhibitors. Recombination of the mutated gp160 genes of the strains resistant to CV-N or ConA into a wild-type background fully reproduced the (cross-)resistance profiles of the originally selected strains, pointing to the impact of the N-glycan mutations on the phenotypic resistance profiles of both selected strains.

[1]  E. De Clercq,et al.  Profile of Resistance of Human Immunodeficiency Virus to Mannose-Specific Plant Lectins , 2004, Journal of Virology.

[2]  K. Gustafson,et al.  Cyanovirin-N inhibits AIDS virus infections in vaginal transmission models. , 2004, AIDS research and human retroviruses.

[3]  Ping Zhu,et al.  Antibody Domain Exchange Is an Immunological Solution to Carbohydrate Cluster Recognition , 2003, Science.

[4]  K. Tashima,et al.  Fusion inhibition--a major but costly step forward in the treatment of HIV-1. , 2003, The New England journal of medicine.

[5]  Benhur Lee,et al.  Human Immunodeficiency Virus Envelope (gp120) Binding to DC-SIGN and Primary Dendritic Cells Is Carbohydrate Dependent but Does Not Involve 2G12 or Cyanovirin Binding Sites: Implications for Structural Analyses of gp120-DC-SIGN Binding , 2002, Journal of Virology.

[6]  E. De Clercq,et al.  env Chimeric Virus Technology for Evaluating Human Immunodeficiency Virus Susceptibility to Entry Inhibitors , 2002, Antimicrobial Agents and Chemotherapy.

[7]  Alexander Wlodawer,et al.  Structures of the Complexes of a Potent Anti-HIV Protein Cyanovirin-N and High Mannose Oligosaccharides* , 2002, The Journal of Biological Chemistry.

[8]  H. Katinger,et al.  The Broadly Neutralizing Anti-Human Immunodeficiency Virus Type 1 Antibody 2G12 Recognizes a Cluster of α1→2 Mannose Residues on the Outer Face of gp120 , 2002, Journal of Virology.

[9]  Peter D. Kwong,et al.  The Mannose-Dependent Epitope for Neutralizing Antibody 2G12 on Human Immunodeficiency Virus Type 1 Glycoprotein gp120 , 2002, Journal of Virology.

[10]  C. Bewley,et al.  Solution Structure of a Cyanovirin-N:Manα1-2Manα Complex , 2001 .

[11]  M. Boyd,et al.  Selective interactions of the human immunodeficiency virus-inactivating protein cyanovirin-N with high-mannose oligosaccharides on gp120 and other glycoproteins. , 2001, The Journal of pharmacology and experimental therapeutics.

[12]  M. Boyd,et al.  Cyanovirin-N defines a new class of antiviral agent targeting N-linked, high-mannose glycans in an oligosaccharide-specific manner. , 2001, Molecular pharmacology.

[13]  C. Bewley,et al.  The potent anti-HIV protein cyanovirin-N contains two novel carbohydrate binding sites that selectively bind to Man(8) D1D3 and Man(9) with nanomolar affinity: implications for binding to the HIV envelope protein gp120. , 2001, Journal of the American Chemical Society.

[14]  David Yang,et al.  The N-Terminal V3 Loop Glycan Modulates the Interaction of Clade A and B Human Immunodeficiency Virus Type 1 Envelopes with CD4 and Chemokine Receptors , 2000, Journal of Virology.

[15]  E. De Clercq,et al.  Polyanionic (i.e., polysulfonate) dendrimers can inhibit the replication of human immunodeficiency virus by interfering with both virus adsorption and later steps (reverse transcriptase/integrase) in the virus replicative cycle. , 2000, Molecular pharmacology.

[16]  L. Pannell,et al.  Isolation and characterization of Myrianthus holstii lectin, a potent HIV-1 inhibitory protein from the plant Myrianthus holstii. , 2000 .

[17]  D. Burton,et al.  Antibody and virus: binding and neutralization. , 2000, Virology.

[18]  D. Burton,et al.  Vaccines and the induction of functional antibodies: Time to look beyond the molecules of natural infection? , 2000, Nature Medicine.

[19]  R. Doms,et al.  Determinants of CD4 Independence for a Human Immunodeficiency Virus Type 1 Variant Map outside Regions Required for Coreceptor Specificity , 1999, Journal of Virology.

[20]  Q. Sattentau,et al.  Cyanovirin-N Binds to gp120 To Interfere with CD4-Dependent Human Immunodeficiency Virus Type 1 Virion Binding, Fusion, and Infectivity but Does Not Affect the CD4 Binding Site on gp120 or Soluble CD4-Induced Conformational Changes in gp120 , 1999, Journal of Virology.

[21]  J. Sodroski,et al.  Probability Analysis of Variational Crystallization and Its Application to gp120, The Exterior Envelope Glycoprotein of Type 1 Human Immunodeficiency Virus (HIV-1)* , 1999, The Journal of Biological Chemistry.

[22]  J. Hansen,et al.  Induction of antibodies against epitopes inaccessible on the HIV type 1 envelope oligomer by immunization with recombinant monomeric glycoprotein 120. , 1998, AIDS research and human retroviruses.

[23]  Nicole Zitzmann,et al.  α‐Glucosidase inhibitors as potential broad based anti‐viral agents , 1998, FEBS letters.

[24]  Peter D. Kwong,et al.  The antigenic structure of the HIV gp120 envelope glycoprotein , 1998, Nature.

[25]  J. Sodroski,et al.  Structure of an HIV gp120 envelope glycoprotein in complex with the CD4 receptor and a neutralizing human antibody , 1998, Nature.

[26]  P. S. Kim,et al.  HIV Entry and Its Inhibition , 1998, Cell.

[27]  L. Pannell,et al.  Recombinant production of cyanovirin-N, a potent human immunodeficiency virus-inactivating protein derived from a cultured cyanobacterium. , 1998, Protein expression and purification.

[28]  D. Burton.  A vaccine for HIV type 1: the antibody perspective. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[29]  L K Pannell,et al.  Discovery of cyanovirin-N, a novel human immunodeficiency virus-inactivating protein that binds viral surface envelope glycoprotein gp120: potential applications to microbicide development , 1997, Antimicrobial agents and chemotherapy.

[30]  E. De Clercq,et al.  Development of resistance of human immunodeficiency virus type 1 to dextran sulfate associated with the emergence of specific mutations in the envelope gp120 glycoprotein. , 1997, Molecular pharmacology.

[31]  J. Hansen,et al.  Influence of N-linked glycans in V4-V5 region of human immunodeficiency virus type 1 glycoprotein gp160 on induction of a virus-neutralizing humoral response. , 1996, Journal of acquired immune deficiency syndromes and human retrovirology : official publication of the International Retrovirology Association.

[32]  E. De Clercq,et al.  The molecular target of bicyclams, potent inhibitors of human immunodeficiency virus replication , 1996, Journal of virology.

[33]  A. Sjölander,et al.  N-linked glycans in the CD4-binding domain of human immunodeficiency virus type 1 envelope glycoprotein gp160 are essential for the in vivo priming of T cells recognizing an epitope located in their vicinity. , 1996, Virology.

[34]  J. Naismith,et al.  Structural Basis of Trimannoside Recognition by Concanavalin A (*) , 1996, The Journal of Biological Chemistry.

[35]  J. McKeating,et al.  Characterization of neutralization epitopes in the V2 region of human immunodeficiency virus type 1 gp120: role of glycosylation in the correct folding of the V1/V2 domain , 1995, Journal of virology.

[36]  R. Pauwels,et al.  Synthesis and structure-activity relationships of phenylenebis(methylene)-linked bis-tetraazamacrocycles that inhibit HIV replication. Effects of macrocyclic ring size and substituents on the aromatic linker. , 1995, Journal of medicinal chemistry.

[37]  Lizhong,et al.  Glycosylation is necessary for the correct folding of human immunodeficiency virus gp120 in CD4 binding , 1993, Journal of virology.

[38]  J. Hansen,et al.  Carbohydrate determinant NeuAc-Gal beta (1-4) of N-linked glycans modulates the antigenic activity of human immunodeficiency virus type 1 glycoprotein gp120. , 1992, The Journal of general virology.

[39]  E. De Clercq,et al.  The mannose-specific plant lectins from Cymbidium hybrid and Epipactis helleborine and the (N-acetylglucosamine)n-specific plant lectin from Urtica dioica are potent and selective inhibitors of human immunodeficiency virus and cytomegalovirus replication in vitro. , 1992, Antiviral research.

[40]  E. De Clercq,et al.  Alpha-(1-3)- and alpha-(1-6)-D-mannose-specific plant lectins are markedly inhibitory to human immunodeficiency virus and cytomegalovirus infections in vitro , 1991, Antimicrobial Agents and Chemotherapy.

[41]  Reed J. Harris,et al.  Assignment of intrachain disulfide bonds and characterization of potential glycosylation sites of the type 1 recombinant human immunodeficiency virus envelope glycoprotein (gp120) expressed in Chinese hamster ovary cells. , 1990, The Journal of biological chemistry.

[42]  J. Hansen,et al.  Correlation between carbohydrate structures on the envelope glycoprotein gp120 of HIV-1 and HIV-2 and syncytium inhibition with lectins. , 1989, AIDS.

[43]  J. Schneider,et al.  Carbohydrates of human immunodeficiency virus. Structures of oligosaccharides linked to the envelope glycoprotein 120. , 1988, The Journal of biological chemistry.

[44]  J Desmyter,et al.  Rapid and automated tetrazolium-based colorimetric assay for the detection of anti-HIV compounds. , 1988, Journal of virological methods.

[45]  H. Gendelman,et al.  Production of acquired immunodeficiency syndrome-associated retrovirus in human and nonhuman cells transfected with an infectious molecular clone , 1986, Journal of virology.

[46]  J U Baenziger,et al.  Structural determinants of concanavalin A specificity for oligosaccharides. , 1979, The Journal of biological chemistry.

[47]  J. Horwitz,et al.  Nucleosides. V. The Monomesylates of 1-(2'-Deoxy-β-D-lyxofuranosyl)thymine1,2 , 1964 .

[48]  K. Schønning.  Antibody neutralization of human immunodeficiency virus type 1 (HIV-1). , 2003, APMIS. Supplementum.

[49]  Raymond A. Dwek,et al.  Targeting glycosylation as a therapeutic approach , 2002, Nature Reviews Drug Discovery.

[50]  E. De Clercq.  Highlights in the development of new antiviral agents. , 2002, Mini reviews in medicinal chemistry.

[51]  C. Bewley.  Solution structure of a cyanovirin-N:Man alpha 1-2Man alpha complex: structural basis for high-affinity carbohydrate-mediated binding to gp120. , 2001, Structure.

[52]  L. Pannell,et al.  Isolation and characterization of Myrianthus holstii lectin, a potent HIV-1 inhibitory protein from the plant Myrianthus holstii(1). , 2000, Journal of natural products.

[53]  R. Dwek,et al.  The role of N-linked glycosylation in the secretion of hepatitis B virus. , 1998, Advances in experimental medicine and biology.

[54]  J. Hansen,et al.  Carbohydrates of human immunodeficiency virus. , 1992, APMIS. Supplementum.

[55]  M. Haine,et al.  Van Damme A. , 1986 .

[56]  Y. Ohtsuki,et al.  Type C Virus-Producing Cell Lines Derived from Adult T Cell Leukemia , 1982 .