Induction of human immunodeficiency virus neutralizing antibodies using fusion complexes.
Human immunodeficiency virus-1 (HIV-1) infects cells by membrane fusion that is mediated by the envelope proteins gp120/gp41 and the cellular receptors CD4 and CCR5. During this process, some conserved viral epitopes are temporarily exposed and may induce a neutralizing antibody response when fixed in the fusogenic conformation. These transient structures are conserved and may be effective antigens for use in an anti-HIV-1 vaccine. In this study we tested different conditions of preparation of fusion complexes inducing neutralizing antibodies against both R5 and X4 tropic HIV-1 strains. Cell lines expressing HIV-1 gp120/gp41 and CD4-CCR5 were prepared and conditions for producing fusion complexes were tested. Complexes produced at different temperature and fixative combinations were used to immunize mice. Results indicated that (a) fusion complexes prepared at either 21 degrees C, 30 degrees C or 37 degrees C were immunogenic and induced neutralizing antibodies against both R5 and X4 HIV-1 heterologous isolates; (b) after extensive purification of antibodies there was no cytotoxic effect; (c) complexes prepared at 37 degrees C were more immunogenic and induced higher titers of neutralizing antibodies than complexes prepared at either 21 degrees C or 30 degrees C; (d) the fixative used did not affect the titer of neutralizing antibodies except for glutaraldehyde which was ineffective; (e) the neutralizing activity was retained after CD4-CCR5 antibody removal. The production of higher titers of neutralizing antibody with fusion complexes prepared at 37 degrees C, as compared to lower temperatures, may be related to the induction of antibodies against many different conformation intermediates that subsequently act synergistically at different steps in the fusion process.
Zipeto
Donato
D
Laboratory of Molecular Virology, Department of Mother and Child, Biology and Genetics, Section of Biology and Genetics, University of Verona, Strada le Grazie n. 8, 37134 Verona, Italy. donato.zipeto@univr.it
Matucci
Andrea
A
Ripamonti
Chiara
C
Scarlatti
Gabriella
G
Rossolillo
Paola
P
Turci
Marco
M
Sartoris
Silvia
S
Tridente
Giuseppe
G
Bertazzoni
Umberto
U
D000818:Animals; D015704:CD4 Antigens / Q000276:immunology; D016466:CHO Cells; D002459:Cell Fusion / Q000379:methods; D006224:Cricetinae; D004797:Enzyme-Linked Immunosorbent Assay; D015483:HIV Antibodies / Q000096:biosynthesis / Q000097:blood / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000276:immunology; D015700:HIV Envelope Protein gp41 / Q000276:immunology; D015658:HIV Infections / Q000276:immunology*; D015497:HIV-1 / Q000276:immunology*; D007114:Immunization / Q000379:methods; D051379:Mice; D008807:Mice, Inbred BALB C; D041681:NIH 3T3 Cells; D009500:Neutralization Tests; D019713:Receptors, CCR5 / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology
D015704:CD4 Antigens; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; D015700:HIV Envelope Protein gp41; D019713:Receptors, CCR5; D011993:Recombinant Fusion Proteins