Passive Immunotherapy for Anthrax Mediated by an Adenovirus Expressing Anti-Protective Antigen Single Chain Antibody
Authors: Kazuhiko Kasuya, Julie L. Boyer, Yadi Tan, D. Olivier Alipui, Neil R. Hackett, Ronald G. Crystal
Journal: Molecular Therapy (2004)

Bacillus anthracis was responsible for 45% of deaths in the 2001 US bioterrorism attacks, despite optimal medical therapy. Any delay in treatment of anthrax significantly affects the chances of survival, therefore, rapid therapy is required. Several vaccines have been developed against B. anthracis, but all require multiple administrations to elicit protective immunity. An anthrax vaccine composed of an adenovirus vector expressing the protective antigen (PA) component of anthrax lethal toxin only requires a single administration to be effective in protecting mice against the anthrax lethal toxin, but it takes >2 wk for anti-anthrax immunity to develop. To address the need for rapid protection, this study describes an in vivo gene transfer-based therapy with a human adenovirus (Ad)-based vector (AdαPAscAb) encoding a single chain antibody that neutralizes protective antigen (PA), a critical component of B. anthracis lethal toxin. To examine the expression and secretion of the anti-PA single chain antibody by AdαPAscAb, A549 cells were infected with AdαPAscAb and, as controls, cells were infected with AdαCTXscAb (an Ad vector encoding an irrelevant single chain antibody) or AdNull (an Ad vector with no transgene). Supernatants from cells infected with AdαPAscAb and AdαCTXscAb both contained antibody reactive with the common human Igκ light chain region. Western analysis of purified protective-antigen or lethal factor proteins with supernatants from AdαPAscAb-infected cells showed that the anti-PA single chain antibody specifically recognized PA. The in vivo expression profile of AdαPAscAb was assessed in serum 0 to 14 days after intravenous administration to BALB/c mice by quantifying anti-PA single chain antibody levels by ELISA and anti-PA neutralizing antibody levels. When measured with a PA-specific ELISA, the anti-PA serum antibody titer in mice receiving AdαPAscAb reached significant levels over a 2 wk period (peak reciprocal titer at day 5 = 12,400 ± 8800). Serum from AdαPAscAb-immunized mice, but not from AdαCTXscAb- or AdNull-immunized mice, had neutralizing activity that paralleled the ELISA titers (peak 50% neutralizing titer at day 5 = 260). Mice that had been injected with AdαPAscAb or AdNull were challenged with lethal toxin at 72 hr post-administration, using naive mice as an additional control. Over a seven day observation period, mice that received AdαPAscAb exhibited a significant survival advantage (100%) relative to naive mice (0%) or mice that received AdαCTXscAb (0%) or AdNull (0%). To determine the timing of protection by AdαPAscAb, mice were given a single intravenous administration with AdαPAscAb, AdαCTXscAb or AdNull at -14 to -1 day before, at the time of, or 0.5 days after challenge with lethal toxin. Although mice immunized at the time of challenge or post toxin administration did not survive, mice that were immunized as early as up to 14 days pre-challenge had a survival advantage relative to control animals (90% protection at day 1, 100% protection at day 2 to day 5, 10% protection at day 14). Passive immunotherapy with AdαPAscAb is a rapid, convenient approach to protect against anthrax or to use in conjunction with antibiotic treatment.