The effect of low-profile serine substitutions in the V3 loop of HIV-1 gp120 IIIB/LAI on the immunogenicity of the envelope protein.

Many microbial antigens contain powerful hypervariable epitopes that fail to induce broadly protective immunity because they dominate the immune response at the expense of more conserved but weaker epitopes. If the undesired B cell epitopes are eliminated, the immune system could be focused on the conserved epitopes and produce a stronger antibody response to conserved parts of the protein and thus become a more efficacious immunogen for a vaccine. We examined this possibility using the human immunodeficiency virus envelope glycoprotein (gp)120 IIIB/LAI and selectively replaced the amino acids from the V3 region and analyzed the overall immunogenicity of the mutant proteins after nucleic acid immunization in mice. The most variable residues of the human immunodeficiency virus type 1 gp120 V3 loop sequence were replaced with serine, which has a small uncharged hydrophilic side chain and therefore is likely to be less immunogenic than amino acids found in wildtype V3 sequences. The serine substitutions did not affect the ability of soluble CD4 to bind the mutant molecules compared with wildtype gp120 and monoclonal antibodies against both linear and discontinuous epitopes located in the V1/V2, C1, and C4 regions of the molecule. These data suggest that the V3 loop substitutions did not grossly affect the overall conformation of the envelope molecule. Immunization of CBA x BALB/c F1 mice with DNA expression plasmids for the wild-type gp120 sequence induced a predominantly IgGI antibody response with end point titers of 10(4)-5 x 10(4). The antibodies reacted only with conformationally intact gp120. Serine replacements targeted to both sides of the V3 loop had a major impact on gp120 immunogenicity, with a markedly reduced response in the majority of animals tested. Analysis of the epitope specificity of the responses suggests that N-terminal amino acids in the V3 loop contribute to the major immunodominant epitope and provides no evidence that their removal enhances immunogenicity of the conserved regions.