Human single-chain antibodies inhibit replication of human immunodeficiency virus type 1 (HIV-1).
The F240 human monoclonal antibody specifically recognizes the disulfide loop-bonded immunodominant epitope of gp41 spanning residues 592-606 and expressed broadly on HIV-1 primary isolates. Despite broad reactivity with native virions and HIV-infected cells, the antibody fails to neutralize infection. However, cytoplasmic expression of single-chain antibody (scFv) directed against gp41 of HIV-1 provides a rationale means to inhibit the maturation of envelope protein. The variable regions of the heavy chain and light chain of human monoclonal antibody were amplified by PCR and linked by a 15 amino acid (GGSGS)3 linker in an orientation of VL-linker-VH and retroviral expression vectors were constructed to simultaneously express F240 scFv and eGFP to facilitate selection of scFv-producing cells. Incorporation of a human immunoglobulin signal sequence directed secretion of the F240 scFv (s-scFv) while an otherwise identical vector lacked this sequence (scFv) resulting in intracellular expression of scFv. Transduced human CD4+ H9 T cells were challenged with HIV. While both secreted and nonsecreted F240 scFv inhibited viral production, secretory F240 scFv was more potent. Thus, this novel approach to direct expression of a nonneutralizing scFv using the Ig signal sequence suggests that targeted therapy using antibodies to conserved, highly expressed epitopes may result in a decrease in viral production due to a reduction of viral assembly and/or transport and expression.
Liu
Fangbing
F
Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA 02215, USA.
Kumar
Mukesh
M
Ma
Qiangzhong
Q
Duval
Mark
M
Kuhrt
David
D
Junghans
Richard
R
Posner
Marshall
M
Cavacini
Lisa
L
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001483:Base Sequence; D017931:DNA Primers; D015700:HIV Envelope Protein gp41 / Q000276:immunology; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000276:immunology*; D051379:Mice; D041681:NIH 3T3 Cells; D009500:Neutralization Tests; D014779:Virus Replication / Q000276:immunology*
D000911:Antibodies, Monoclonal; D017931:DNA Primers; D015700:HIV Envelope Protein gp41; D007135:Immunoglobulin Variable Region