title: Development of an edema factor-mediated cAMP-induction bioassay for detecting antibody-mediated neutralization of anthrax protective antigen.
abstract: Intoxication of mammalian cells by Bacillus anthracis requires the coordinate activity of three distinct bacterial proteins: protective antigen (PA), edema factor (EF), and lethal factor (LF). Among these proteins, PA has become the major focus of work on monoclonal antibodies and vaccines designed to treat or prevent anthrax infection since neither EF nor LF is capable of inducing cellular toxicity in its absence. Here, we present the development of a sensitive, precise, and biologically relevant bioassay platform capable of quantifying antibody-mediated PA neutralization. This bioassay is based on the ability of PA to bind and shuttle EF, a bacterial adenylate cyclase, into mammalian cells leading to an increase in cAMP that can be quantified using a sensitive chemiluminescent ELISA. The results of this study indicate that the cAMP-induction assay possesses the necessary performance characteristics for use as both a potency-indicating release assay in a quality control setting and as a surrogate pharmacodynamic marker for ensuring the continued bioactivity of therapeutic antibodies against PA during clinical trials.
authors.affiliation: Human Genome Sciences, Inc., 14200 Shady Grove Road, Rockville, MD 20850, USA. Jonathan_Zmuda@HGSI.com
authors.forename: Jonathan F
authors.initials: JF
authors.lastname: Zmuda
authors.forename: Linyi
authors.initials: L
authors.lastname: Zhang
authors.forename: Terri
authors.initials: T
authors.lastname: Richards
authors.forename: Quyen
authors.initials: Q
authors.lastname: Pham
authors.forename: David
authors.initials: D
authors.lastname: Zukauskas
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Pierre
authors.forename: Michael W
authors.initials: MW
authors.lastname: Laird
authors.forename: Janine
authors.initials: J
authors.lastname: Askins
authors.forename: Gil H
authors.initials: GH
authors.lastname: Choi
mesh_terms: D000818:Animals; D000942:Antigens, Bacterial / Q000276:immunology*; D001408:Bacillus anthracis / Q000276:immunology; D001427:Bacterial Toxins / Q000276:immunology*; D001681:Biological Assay / Q000379:methods*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014757:Viper Venoms / Q000276:immunology / Q000378:metabolism*
chemical_list: D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D014757:Viper Venoms; C030325:anthrax toxin; C416405:edema factor; D000242:Cyclic AMP