A cocktail of humanized anti–pertussis toxin antibodies limits disease in murine and baboon models of whooping cough

Abstract

In spite of wide-spread vaccination, pertussis rates are rising in industrialized countries and remain high world-wide. With no specific therapeutics to treat disease, pertussis continues to cause considerable infant morbidity and mortality. The pertussis toxin is a major contributor to disease, responsible for local and systemic effects including leukocytosis and immunosuppression. Here, we humanized two murine monoclonal antibodies that neutralize pertussis toxin and expressed them as human IgG1 molecules with no loss of affinity or in vitro neutralization activity. When administered prophylactically to mice as a binary cocktail, antibody treatment completely mitigated the B. pertussis-induced rise in white blood cell count and decreased bacterial colonization. When administered therapeutically to baboons, antibody-treated but not control animals experienced a blunted rise in white blood cell count and accelerated bacterial clearance rates. These preliminary findings support further investigation into the use of these antibodies to treat human neonatal pertussis in conjunction with antibiotics and supportive care.

Synergistic Neutralization of Pertussis Toxin by a Bispecific Antibody In Vitro and In Vivo

“…The full-length formats showed an approximately 3-fold improvement in effective affinity compared to the Fabs, likely due to avidity effects. The K D values calculated for the full-length hu1B7 and hu11E6 antibodies were 0.8 Ϯ 0.1 nM and 6 Ϯ 2 nM, respectively, and are comparable to our previous competition ELISA and surface plasmon resonance (SPR) measurements (9).…”

“…The hu11E6 antibody binds two homologous sites on the B subunit and thus prevents the initial interaction between the toxin and glycosylated receptors on host cells (33), while the hu1B7 antibody binds the A subunit, preventing ADP-glycosylation of cytoplasmic G i/o receptors (21). We thus hypothesized that the synergy previously observed between hu1B7 and hu11E6 was due to a more complete neutralization of the toxin function when both the binding and catalytic activities of the toxin were blocked (9). This effect would be maximal if the hu1B7 and hu11E6 epitopes on a single PTx molecule could be bound simultaneously by their corresponding antibodies, creating a hu1B7-PTx-hu11E6 complex.…”

“…To provide a point of reference in our functional assays and to assess valency effects, we also produced monoclonal hu1B7 and hu11E6 and their Fab fragments. The monoclonal antibodies were expressed in transient CHO cell cultures and purified by protein A and anion exchange chromatography as previously described (9). The Fab fragments were generated by subsequent digestion of purified antibody with immobilized pa-…”

“…We previously observed that an equimolar mixture of the hu1B7 and hu11E6 anti-PTx antibodies was more effective in vitro than either antibody alone (9). This led us to hypothesize that a bispecific antibody containing these two binding sites would capture the therapeutic potential of the mixture in a single molecule.…”

“…In addition, low titers of PTx-neutralizing antibodies correlate with susceptibility to clinical infection (8). We previously developed a binary mixture of two humanized anti-pertussis toxin antibodies which was able to mitigate whooping cough in mouse and baboon models of infection (9). The antibody hu11E6 blocks binding of the toxin to host cells, while the antibody hu1B7 interferes with the catalytic pathway.…”