Epitope determinants of a chimpanzee dengue virus type 4 (DENV-4)-neutralizing antibody and protection against DENV-4 challenge in mice and rhesus monkeys by passively transferred humanized antibody - PubMed (original) (raw)

Epitope determinants of a chimpanzee dengue virus type 4 (DENV-4)-neutralizing antibody and protection against DENV-4 challenge in mice and rhesus monkeys by passively transferred humanized antibody

Ching-Juh Lai et al. J Virol. 2007 Dec.

Abstract

The chimpanzee monoclonal antibody (MAb) 5H2 is specific for dengue virus type 4 (DENV-4) and neutralizes the virus at a high titer in vitro. The epitope detected by the antibody was mapped by sequencing neutralization escape variants of the virus. One variant contained a Lys174-Glu substitution and another contained a Pro176-Leu substitution in domain I of the DENV-4 envelope protein (E). These mutations reduced binding affinity for the antibody 18- to >100-fold. Humanized immunoglobulin G (IgG) 5H2, originally produced from an expression vector, has been shown to be a variant containing a nine-amino-acid deletion in the Fc region which completely ablates antibody-dependent enhancement of DENV replication in vitro. The variant MAb, termed IgG 5H2 deltaD, is particularly attractive for exploring its protective capacity in vivo. Passive transfer of IgG 5H2 deltaD at 20 microg/mouse afforded 50% protection of suckling mice against challenge with 25 50% lethal doses of mouse neurovirulent DENV-4 strain H241. Passive transfer of antibody to monkeys was conducted to demonstrate proof of concept for protection against DENV challenge. Monkeys that received 2 mg/kg of body weight of IgG 5H2 deltaD were completely protected against 100 50% monkey infectious doses (MID50) of DENV-4, as indicated by the absence of viremia and seroconversion. A DENV-4 escape mutant that contained a Lys174-Glu substitution identical to that found in vitro was isolated from monkeys challenged with 10(6) MID50 of DENV-4. This substitution was also present in all naturally occurring isolates belonging to DENV-4 genotype III. These studies have important implications for possible antibody-mediated prevention of DENV infection.

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Figures

FIG. 1.

(A) In vitro neutralization assay of IgG 5H2 ΔD against DENV-4 parental viruses and antigenic variants. (B) Growth curves for DENV-4 antigenic variants and parental viruses in C6/36 cells. The multiplicity of infection was 0.1.

FIG. 2.

(A) Comparison of amino acid sequences of E proteins of wild-type DENV-4, antigenic variants, and other flaviviruses. In the sequence alignment, a dot indicates an identical amino acid compared with DENV-4. The localization of amino acid substitutions at positions 174 and 176 in the 3-D structure of monomeric DENV-4 E, based on the DENV-2 E model, is shown from the top (B) and from the side (C). Domain I is in red, domain II is in yellow, and domain III is in blue. Structural modeling of DENV-4 E was performed using SwissModel and the published crystal coordinates of DENV-2 E (1OAN.pdb) as the template (12, 37). Molecular graphic images were produced using the UCSF Chimera package from the Resource for Bio-Computing, Visualization and Informatics at the University of California, San Francisco (44).

FIG. 3.

ADE of DENV-4-specific epitope and neutralization of DENV-4 by IgG 5H2 ΔD before and after adsorption to Vero cells. A comparison of ADE of replication of parental DENV-4 and its derived antigenic variants in K562 cells exposed to full-length IgG 5H2 was performed. (A) Percentage of cells infected with DENV-4, determined by flow cytometry. (B) In pre- and postadsorption antibody neutralization assays, a constant amount of DENV-4 (70 FFU) was tested against various dilutions of IgG 5H2 ΔD.

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Epitope determinants of a chimpanzee dengue virus type 4 (DENV-4)-neutralizing antibody and protection against DENV-4 challenge in mice and rhesus monkeys by passively transferred humanized antibody - PubMed (original) (raw)