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

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 Lys 174 -Glu substitution and another contained a Pro 176 -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 ⌬D, is particularly attractive for exploring its protective capacity in vivo. Passive transfer of IgG 5H2 ⌬D at 20 g/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 ⌬D were completely protected against 100 50% monkey infectious doses (MID 50 ) of DENV-4, as indicated by the absence of viremia and seroconversion. A DENV-4 escape mutant that contained a Lys 174 -Glu substitution identical to that found in vitro was isolated from monkeys challenged with 10 6 MID 50 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.The four dengue virus serotypes (dengue virus types 1 to 4 [DENV-1 to DENV-4]) cause more morbidity in humans than any other arthropod-borne flaviviruses (40). Up to 100 million DENV infections occur every year, mostly in tropical and subtropical areas where the vector mosquitoes, principally Aedes aegypti and Aedes albopictus, are present. Dengue illnesses range from mild fever to severe dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS), which has fatality rates ranging from Ͻ1% to 5% in children. The most severe dengue (Ͼ90% fatality rate) occurs in patients reinfected with DENV of a serotype different from that in the primary infection (15, 50). Antibody-dependent enhancement (ADE) of DENV replication has been proposed as an underlying pathogenic mechanism of severe DHF/DSS (17). A safe and effective vaccine against dengue is still not available.Early studies of DENV infections in human volunteers showed that homotypic immunity against the same serotype is life-long but that heterotypic immunity against other serotypes lasts only months (49). Since antibodies provide the important component of acquired immunity against infection, type-specific immunity afforded by antibody may contribute significantly to long-term protection. Antigenic differences exist among strains of the same serotype (21). DENV variants that form ...

Chimpanzees as an animal model for human norovirus infection and vaccine development

“…The chimpanzee animal model has played a key role in the study of several viral pathogens (22)(23)(24)(25)(26)(27)(28)(29)(30). During early HAV challenge experiments in nonhuman primates, it was determined that 10 4.5 -fold more virus was required to infect animals by the oral route compared with the i.v.…”

Functional Analysis of Antibodies against Dengue Virus Type 4 Reveals Strain-Dependent Epitope Exposure That Impacts Neutralization and Protection

“…Two typespecific anti-DENV-4 chimpanzee MAbs (5D9 and 5H2) were generated after immunization with DENV-4 (strain 814669; Caribbean); these MAbs inhibited infection of the homologous strain (PRNT 50 of 580 and 240 ng/ml, respectively) (42). The 5H2 MAb, which localized to an epitope in domain I (amino acid residues K174 and P176) of the E protein (43), neutralized two other DENV-4 isolates (H-241 [Philippines] and 341750 [Caribbean]) from similar and distinct geographic regions (42) and also protected suckling mice from lethal DENV-4 H-241 infection when administered as prophylaxis. A recombinant variant of 5H2 lacking the ability to bind Fc␥ receptors protected monkeys from viremia after challenge with DENV-4 814669.…”

“…While our laboratory and others have generated panels of inhibitory monoclonal antibodies (MAbs) that react specifically with DENV-1, DENV-2, or DENV-3, to date, remarkably few strongly neutralizing type-specific anti-DENV-4 MAbs have been reported: these include three unmapped mouse MAbs (UH-6C3, D4-II-12B2, and D4-I-11D11) (40,41), two chimpanzee MAbs (5D9 and 5H2), one of which is localized to an epitope in domain I of the E protein (42,43), and one human MAb DV22.3, which recognizes an epitope in DI-DII (44). In the current study, we developed a panel of 81 new DENV-4 mouse MAbs and examined their neutralization potentials initially against a genotype II strain (DENV-4 1036; Indonesia 1976) that was used for immunization.…”

Deconstructing the Antiviral Neutralizing-Antibody Response: Implications for Vaccine Development and Immunity

“…Blocking of cell surface attachment or receptor engagement through steric hindrance may be a common mechanism and has been suggested to explain the activity of DENVimmune sera (176). However, many antibodies are also capable of blocking infection at a postattachment step (51,59,167,(177)(178)(179). These NAbs may inhibit conformational changes of a viral protein required to mediate virus entry.…”