An immunoglobulin fusion protein based on the gp120-CD4 receptor complex potently inhibits human immunodeficiency virus type 1 in vitro.
Fusion proteins containing immunoglobulin Fc domains attached to bioactive moieties have been developed as therapeutic agents against several diseases. Here, we describe the development and characteristics of a novel fusion protein (FLSC R/T-IgG1) that targets CCR5, the major coreceptor for HIV-1 during primary infection. FLSC R/T-IgG1 was expressed from a synthetic gene that linked a single chain gp120-CD4 complex containing an R5 gp120 sequence with the hinge-CH2-CH3 portion of human immunoglobulin gamma subtype 1. Purified FLSC R/T-IgG1 exhibited a molecular mass of 189 kDa under reducing conditions, which matched the expected size of one polypeptide chain. Chemically crosslinked or untreated FLSC R/T-IgG1 exhibited a mass of a 360-kDa polypeptide under reducing and nonreducing conditions, which indicated that the molecule adopts a disulfide-linked bivalent structure. The chimeric molecule bound specifically to CCR5-expressing cells and to peptides derived from the CCR5 N-terminus. Such binding was more efficient than what was obtained with a monomeric single chain gp120-CD4 complex. FLSC R/T-IgG1 binding to CCR5 was blocked by preincubation of coreceptor-expressing cells with CCR5 ligands and by antibody to the coreceptor binding domain of gp120. Conversely, FLSC R/T-IgG1 blocked the binding of chemokine to CCR5. However, FLSC R/T-IgG1 did not trigger intracellular Ca2+ mobilization in peripheral blood mononuclear cells. FLSC R/T-IgG1 potently neutralized primary R5 HIV-1 in both a PBMC-based assay and cell line-based assays but did not affect the replication of X4 viruses. These findings suggest that FLSC R/T-IgG1 might be used as a possible therapeutic agent against HIV.
Vu
John R
JR
Department of Microbiology and Immunology, School of Medicine, University of Maryland, Baltimore, Maryland 21201, USA.
Fouts
Timothy
T
Bobb
Katherine
K
Burns
Jennifer
J
McDermott
Brenda
B
Israel
David I
DI
Godfrey
Karla
K
DeVico
Anthony
A
D016915:AIDS Vaccines; D015704:CD4 Antigens / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D015699:HIV Envelope Protein gp120 / Q000235:genetics / Q000378:metabolism*; D015497:HIV-1 / Q000187:drug effects*; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D007963:Leukocytes, Mononuclear / Q000821:virology; D008969:Molecular Sequence Data; D010455:Peptides; D011485:Protein Binding; D019713:Receptors, CCR5 / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D017422:Sequence Analysis, DNA
D016915:AIDS Vaccines; D015704:CD4 Antigens; D015699:HIV Envelope Protein gp120; D007074:Immunoglobulin G; D010455:Peptides; D019713:Receptors, CCR5; D011993:Recombinant Fusion Proteins