A Monoclonal Immunoglobulin G Antibody Directed against an Immunodominant Linear Epitope on the Ricin A Chain Confers Systemic and Mucosal Immunity to Ricin

Abstract

ABSTRACT Due to the potential use of ricin and other fast-acting toxins as agents of bioterrorism, there is an urgent need for the development of safe and effective antitoxin vaccines. A candidate ricin subunit vaccine (RiVax) consisting of a recombinant attenuated enzymatic A chain (RTA) has been shown to elicit protective antitoxin antibodies in mice and rabbits and is currently being tested in phase I human clinical trials. However, evaluation of the efficacy of this vaccine for humans is difficult for a number of reasons, including the fact that the key neutralizing B-cell epitopes on RTA have not been fully defined. Castelletti and colleagues (Clin. Exp. Immunol. 136: 365-372, 2004) recently identified a linear epitope on RTA, spanning residues L161 to I175, as a primary target of serum antibodies derived from humans who had been treated with ricin immunotoxin. While affinity-purified polyclonal IgG antibodies against this region of RTA were capable of neutralizing ricin in vitro , their capacity to confer protection against ricin challenge in vivo was not determined. In this report, we describe the production and characterization of GD12, a murine monoclonal IgG1 antibody specifically directed against residues 163 to 174 (TLARSFIICIQM) of RTA. GD12 bound ricin holotoxin with high affinity ( K D [dissociation constant], 2.9 × 10 −9 M) and neutralized it with a 50% inhibitory concentration of ∼0.25 μg/ml, as determined by a Vero cell-based cytotoxicity assay. Passive administration of GD12 was sufficient to protect BALB/c mice against intraperitoneal and intragastric ricin challenges. These data are important in terms of vaccine development, since they firmly establish that preexisting serum antibodies directed against residues 161 to 175 on RTA are sufficient to confer both systemic and mucosal immunity to ricin. The potential of GD12 to serve as a therapeutic following ricin challenge was not explored in this study.