FIG. 3.

D6R inhibits anthrax toxemia in Fisher 344 rats. (A) Twenty Fisher 344 male rats (weight, 250 to 300 g) were anesthetized by intraperitoneal injection of ketamine-xylazine and injected intravenously with a mixture of 40 μg of PA and 8 μg of LF per rat (which represents 10 times the minimal lethal dose) in a total volume of 200 μl of phosphate-buffered saline. With this dose of toxin, animals become moribund after approximately 90 to 120 min. D6R (1 mg/100 μl in physiological saline) was given intravenously to 10 rats immediately after administration of anthrax toxins; 100 μl of phosphate-buffered saline was given to the other group. The time until moribund, defined as the point at which severe symptoms of intoxication (highly labored breathing and prostration) appear, was scored at specific time intervals (every 30 min for 2 h and every hour thereafter). When moribund, animals were sacrificed by CO₂ inhalation for humane reasons (6, 17, 19). Rats that did not show these symptoms survived toxin administration, recovered from anesthesia, and were sacrificed after 2 weeks of observation. The first group of 10 rats was treated with anthrax toxin alone (open squares), and the second group was treated with anthrax toxin and 1 mg of D6R (filled circles). A preliminary experiment was performed in a nonblind fashion, while the experiment shown was performed in a double-blind fashion. (B) D6R protects against anthrax toxin-induced intoxication in mice. Groups of FVB mice (10 animals per group, 6 weeks old, both genders) were also given anthrax toxin (10 μg of PA and 0.5 μg of FP59 per mouse), but intraperitoneally (open squares). One group of mice was also administered 100 μg of D6R intraperitoneally immediately after anthrax toxin (filled circles). Mice were observed for 24 h after injection. With the first signs of impending morbidity (intensive and labored breathing accompanied by prostration), mice were sacrificed by CO₂ inhalation. Survival rates were assessed every half hour for the first 2 h, every 2 h thereafter for 12 h, and intermittently thereafter for 48 h. Surviving mice were sacrificed 2 weeks later. Results are presented as percent survival at the times indicated. Surviving mice were observed for 48 h after injection. Per the Fisher exact test, differences between both experimental groups achieved significance at the 6-h time point and afterward (P < 0.05).