title: Biochemically defined HIV-1 envelope glycoprotein variant immunogens display differential binding and neutralizing specificities to the CD4-binding site.
abstract: HIV-1 gp120 binds the primary receptor CD4. Recently, a plethora of broadly neutralizing antibodies to the gp120 CD4-binding site (CD4bs) validated this region as a target for immunogen design. Here, we asked if modified HIV-1 envelope glycoproteins (Env) designed to increase CD4 recognition might improve recognition by CD4bs neutralizing antibodies and more efficiently elicit such reactivities. We also asked if CD4bs stabilization, coupled with altering the Env format (monomer to trimer or cross-clade), might better elicit neutralizing antibodies by focusing the immune response on the functionally conserved CD4bs. We produced monomeric and trimeric Envs stabilized by mutations within the gp120 CD4bs cavity (pocket-filling; PF2) or by appending heterologous trimerization motifs to soluble Env ectodomains (gp120/gp140). Recombinant glycoproteins were purified to relative homogeneity, and ligand binding properties were analyzed by ELISA, surface plasmon resonance, and isothermal titration microcalorimetry. In some formats, the PF2 substitutions increased CD4 affinity, and importantly, PF2-containing proteins were better recognized by the broadly neutralizing CD4bs mAbs, VRC01 and VRC-PG04. Based on this analysis, we immunized selected Env variants into rabbits using heterologous or homologous regimens. Analysis of the sera revealed that homologous inoculation of the PF2-containing, variable region-deleted YU2 gp120 trimers (ΔV123/PF2-GCN4) more rapidly elicited CD4bs-directed neutralizing antibodies compared with other regimens, whereas homologous trimers elicited increased neutralization potency, mapping predominantly to the gp120 third major variable region (V3). These results suggest that some engineered Env proteins may more efficiently direct responses toward the conserved CD4bs and be valuable to elicit antibodies of greater neutralizing capacity.
authors.affiliation: IAVI Neutralizing Antibody Center, Department of Immunology and Microbial Science, The Scripps Research Institute, La Jolla, California 92037, USA.
authors.forename: Yu
authors.initials: Y
authors.lastname: Feng
authors.forename: Krisha
authors.initials: K
authors.lastname: McKee
authors.forename: Karen
authors.initials: K
authors.lastname: Tran
authors.forename: Sijy
authors.initials: S
authors.lastname: O'Dell
authors.forename: Stephen D
authors.initials: SD
authors.lastname: Schmidt
authors.forename: Adhuna
authors.initials: A
authors.lastname: Phogat
authors.forename: Mattias N
authors.initials: MN
authors.lastname: Forsell
authors.forename: Gunilla B
authors.initials: GB
authors.lastname: Karlsson Hedestam
authors.forename: John R
authors.initials: JR
authors.lastname: Mascola
authors.forename: Richard T
authors.initials: RT
authors.lastname: Wyatt
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology*; D000918:Antibody Specificity; D001665:Binding Sites; D015704:CD4 Antigens / Q000378:metabolism*; D005260:Female; D015699:HIV Envelope Protein gp120 / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D015497:HIV-1; D007700:Kinetics; D015202:Protein Engineering; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D011817:Rabbits; D013816:Thermodynamics; D054299:env Gene Products, Human Immunodeficiency Virus / Q000276:immunology
chemical_list: D057134:Antibodies, Neutralizing; D015704:CD4 Antigens; D015699:HIV Envelope Protein gp120; D054299:env Gene Products, Human Immunodeficiency Virus; C514793:gp120 protein, Human immunodeficiency virus 1; C097729:gp140 envelope protein, Human immunodeficiency virus 1