Isolation and Characterization of Cross-Reactive Human Monoclonal Antibodies That Potently Neutralize Australian Bat Lyssavirus Variants and Other Phylogroup 1 Lyssaviruses

Abstract

Australian bat lyssavirus (ABLV) is a rhabdovirus that circulates in four species of pteropid bats (ABLVp) and the yellow-bellied sheath-tailed bat (ABLVs) in mainland Australia. In the three confirmed human cases of ABLV, rabies illness preceded fatality. As with rabies virus (RABV), post-exposure prophylaxis (PEP) for potential ABLV infections consists of wound cleansing, administration of the rabies vaccine and injection of rabies immunoglobulin (RIG) proximal to the wound. Despite the efficacy of PEP, the inaccessibility of human RIG (HRIG) in the developing world and the high immunogenicity of equine RIG (ERIG) has led to consideration of human monoclonal antibodies (hmAbs) as a passive immunization option that offers enhanced safety and specificity. Using a recombinant vesicular stomatitis virus (rVSV) expressing the glycoprotein (G) protein of ABLVs and phage display, we identified two hmAbs, A6 and F11, which completely neutralize ABLVs/ABLVp, and RABV at concentrations ranging from 0.39 and 6.25 µg/mL and 0.19 and 0.39 µg/mL respectively. A6 and F11 recognize overlapping epitopes in the lyssavirus G protein, effectively neutralizing phylogroup 1 lyssaviruses, while having little effect on phylogroup 2 and non-grouped diverse lyssaviruses. These results suggest that A6 and F11 could be effective therapeutic and diagnostic tools for phylogroup 1 lyssavirus infections.

mAb therapy controls CNS‐resident lyssavirus infection via a CD4 T cell‐dependent mechanism

“…F11 is a recombinant human IgG1 mAb that targets the ABLV envelope glycoprotein (G) (see Materials and Methods and Weir et al , 2021 ). As shown in Fig EV1A , F11 demonstrates potent in vitro neutralization of ABLV‐luc with a neutralization titer of 0.267 μg/ml.…”

“…Although normally excluded from the brain by the blood–brain barrier (BBB), cells of the immune system can be efficiently recruited to the brain in specific circumstances, including viral infection (Engelhardt, 2006 ; Arima et al , 2013 ). To determine whether F11 therapy changes the spectrum of leukocytes within the infected brain, we used flow cytometry to compare leukocyte subtypes in infected animals treated with either F11 or with the anti‐henipavirus G glycoprotein huIgG1 mAb, m102.4 (Zhu et al , 2008 ) (which has an identical constant region to F11, but has no measurable interaction with ABLV G (Weir et al , 2021 )). Animals were infected with 2 × 10 5 FFU of ABLV‐luc and treated with either F11 or with m102.4 at day 5 post‐infection.…”

“…With a half-life of approximately 13 days, 140 days is more than ten half-lives, corresponding to greater than a 1,000fold reduction from the starting in vivo concentration. We recently reported that the IC 50 for mAb F11 is 0.6 ng/ml for ABLV and several other phylogroup I lyssaviruses (Weir et al, 2021). While we measured the initial in vivo concentration of F11 at 154 ng/ml, a value well above the IC 50 , the concentration is reduced to 0.15 ng/ml after 10 half-lives, which is below the IC 50 , strongly suggesting that meaningful neutralization no longer occurs at such late post-therapy time points.…”

“…More specifically, we used longitudinal imaging of lethal-dose ABLV infection in a mouse model to trace virus location and relative viral load in the presence and absence of therapeutic intervention. We demonstrate that a single dose of human anti-ABLV/RABV mAb F11 (Weir et al, 2021) can limit lyssavirus replication in the CNS, resolve morbidity and promote survival for more than 4 months post-treatment, even when administered after demonstrable viral replication in the CNS and onset of disease signs. Surprisingly, we found that F11 efficacy requires an intact host adaptive immune response, particularly CD4 T cells, and that immune cell infiltrates in the brains of F11-treated animals differ significantly from untreated animals.…”

Structural Determination of the Australian Bat Lyssavirus Nucleoprotein and Phosphoprotein Complex

“…Screening of Australian bat populations has shown ABLV in five out of six bat families present, and widespread prevalence [ 9 ]. As such, all contact with bats should be considered a potential transmission event, and exposed individuals should receive post-exposure prophylaxis in the form of a rabies vaccine and hIG as these species are in the same lyssavirus phylogroup, and cross-reactivity is present [ 10 , 11 ].…”