A cocktail of human monoclonal antibodies broadly neutralizes North American rabies virus variants as a promising candidate for rabies post-exposure prophylaxis

Abstract

Human rabies remains a globally significant public health problem. Replacement of polyclonal anti-rabies immunoglobulin (RIG), a passive component of rabies post-exposure prophylaxis (PEP), with a monoclonal antibody (MAb), would eliminate the cost and availability constraints associated with RIG. Our team has developed and licensed a human monoclonal antibody RAB1 (Rabishield^©), as the replacement for RIG where canine rabies is enzootic. However, for the highly diverse rabies viruses of North America, a cocktail containing two or more MAbs targeting different antigenic sites of the rabies glycoprotein should be included to ensure neutralization of all variants of the virus. In this study, two MAb cocktails, R172 (RAB1-RAB2) and R173 (RAB1-CR57), were identified and evaluated against a broad range of rabies variants from North America. R173 was found to be the most potent cocktail, as it neutralized all the tested North American RABV isolates and demonstrated broad coverage of isolates from both terrestrial and bat species. R173 could be a promising candidate as an alternative or replacement for RIG PEP in North America.

Figure 1

RAB2 and R172 binding and neutralization of rabies glycoproteins. (a) Schematic of rabies glycoprotein showing non-overlapping antigenic site locations of RAB1 (antigenic site III) and RAB2 (antigenic site II), figure was created with BioRender.com. (b) RAB2 recognition of antigenic site II alanine mutants. Each amino acid in the antigenic site II of ERA full length surface glycoprotein (a.a. 1–524) was substituted with alanine by site directed mutagenesis. (c) ELISA binding of RAB2 to the wild-type ERA439 or RAB1 escape N336D R346K and RAB2 escape E33K mutants. (d) Affinity measurement of RAB2 antibody against wild type ERA439, conducted by Bio-layer interferometry. (e) ELISA binding of R172 to the wild-type ERA439 or RAB1 escape N336D R346K and RAB2 escape E33K mutants. (f–h) R172 pseudovirus neutralization against lentivirus bearing the wild-type ERA glycoprotein (f), E33K (g) and N336D R346K (h) mutants. ERA glycoprotein was used as backbone to introduce all mutants. EC50 values were calculated by nonlinear regression analysis using Prism version 8.1.1. Data is plotted as the mean ± s.d. from n = 4 independent experiments.

Figure 2

CR57 and R173 binding and neutralization of rabies glycoproteins. (a) Schematic of rabies glycoprotein showing non-overlapping antigenic sites locations of RAB1 (antigenic site III) and CR57 (antigenic site I), figure was created with BioRender.com. (b, c) ELISA binding of CR57 (b) and R173 (c) to the wild type ERA₄₃₉ or RAB1 escape mutant N336D R346K. (d, e) Pseudovirus neutralization of RAB1, CR57, and R173 against lentivirus bearing the wild type ERA glycoprotein (d) and N336 R346K mutant (e). EC₅₀ values were calculated by nonlinear regression analysis using Prism version 8.1.1. Data is plotted as the mean ± s.d. from n = 4 independent experiments.