Anthrax immunoglobulin

The polyclonal immune globulin G is a passive immunizing agent that neutralizes anthrax toxin. Anthrax immunoglobulin binds to protective antigen (PA) to prevent PA mediated cellular entry of anthrax edema factor and lethal factor. Anthrax immunoglobulin is administered in combination with appropriate antibiotic therapy as the product itself is not known to have direct antibacterial activity against anthrax bacteria, which otherwise may continue to grow and product anthrax toxins.

The mean TNA activities for three doses of anthrax immunoglobulin (210, 420 and 840 units TNA) in the clinical trial in healthy volunteers are plotted on a semi-log scale in Figure 1. The pharmacokinetics of anthrax immunoglobulin after intravenous infusion of the three dose levels were characterized; the peak levels of anthrax immunoglobulin were reached immediately after infusion and then declined over the duration of study (84 days). The mean TNA activity remained above the lower limit of quantitation (5 milliunits per mL) over the entire 84-day post-dose period for the three doses studied.

Figure 1. Mean TNA Activities for Three Doses of anthrax immunoglobulin:

A summary of the mean pharmacokinetic results for the TNA data collected in the healthy volunteer study is presented in Table 3.

Table 3. Summary of Mean PK Results by Treatment (TNA Data):

In comparison to healthy subjects, patients with inhalational anthrax are expected to initially have greater clearance of anti-PA antibodies and lower AUC from anthrax immunoglobulin administration due to the presence of PA antigen.

Mean PK results (TNA data) were evaluated by sex and revealed no sex-related differences over the dose range studied. Systemic exposure of anthrax immunoglobulin increased in a dose-proportional manner over the dose range studied. Anthrax immunoglobulin has a serum elimination half-life of 24 to 28 days in healthy humans.

Inhalational anthrax patients, concomitantly treated with antibiotics and a single 420 unit TNA dose of anthrax immunoglobulin, exhibited increases in serum and pleural anti-PA levels; these levels remained at >50% of the peak anti-PA levels over the next five days. The peak serum anti-PA levels in these patients following anthrax immunoglobulin administration (132 to 160 mcg/mL, mean 145 mcg/mL) overlapped with those obtained with the 420 unit dose in healthy volunteers (135 to 250 mcg/mL, mean 190 mcg/mL, median 192 mcg/mL), although mean levels were approximately 25% lower in the inhalational anthrax patients. In the three inhalational anthrax patients, serum and pleural levels of lethal factor declined after initiation of antibiotics and further decreased over the period of five days following anthrax immunoglobulin administration; however, due at least in part to anthrax immunoglobulin targeting the PA component of lethal toxin, plasma and pleural fluid lethal factor levels remained detectable when measured two to five days following anthrax immunoglobulin administration.

Because the effectiveness of anthrax immunoglobulin cannot ethically be tested in placebo-controlled trials in humans, a comparison of anthrax immunoglobulin exposures achieved in healthy human subjects to those observed in animal models of inhalational anthrax in therapeutic efficacy studies was necessary to support the dosage regimen. A dose of 420 units has a similar exposure to the efficacious dose of 15 U/kg administered to New Zealand white rabbits and cynomolgus macaques. In cynomolgus macaques treated with anthrax immunoglobulin monotherapy, a higher dose of 30 U/kg, with a similar exposure to a human dose of 840 units, may result in improved survival. As a result, the initial dosing regimen is given as a range of 420 to 840 units, and the recommended regimen includes the potential for repeat dosing.

Anthrax infected New Zealand white rabbits and cynomolgus macaques administered an intravenous injection of anthrax immunoglobulin (15 units TNA per kg) that did not survive their infection showed an increase in the severity and/or incidence of central nervous system lesions (bacteria, hemorrhage and necrosis) as compared to immune globulin (“placebo”) treated animals who also did not survive the infection. The mean time to death between non-surviving anthrax immunoglobulin and placebo treated animals was comparable. Surviving rabbits had no evidence of central nervous system lesions at the end of the study. No surviving cynomolgus macaques in monotherapeutic studies were tested for central nervous system lesions.

Monotherapeutic Studies in Animal Models

In a monotherapeutic efficacy study, rabbits were exposed to a target dose of 200 x LD₅₀ aerosolized anthrax spores and then administered 15 units per kg of anthrax immunoglobulin at the onset of toxemia, as determined by the presence of PA in serum samples. Detection of PA was used as the trigger for initiation of treatment, while bacteremia status provided a retrospective confirmation of disease. Ninety-eight (98) percent of the treated animals were bacteremic prior to treatment. Of the animals that were toxemic and bacteremic prior to treatment, anthrax immunoglobulin treatment resulted in a 26% survival in comparison to a 2% survival with IGIV placebo treatment (Table 4) over the 36 day duration of the study. Anthrax immunoglobulin treatment resulted in a significant decrease in the proportion of rabbits that were toxemic or bacteremic. The time to resolution of toxemia (p=0.0006) or bacteremia (p=0.0074) was also significantly reduced in rabbits that received anthrax immunoglobulin.

Efficacy of anthrax immunoglobulin was also assessed in cynomolgus macaques exposed to a target dose of 200 x LD₅₀ aerosolized anthrax spores. Treatment with placebo or one of three dose levels of anthrax immunoglobulin was initiated after animals became toxemic (positive for PA detection in serum samples), and bacteremia status provided a retrospective confirmation of disease. Survival was assessed over a period of 88 days in toxemic animals that were confirmed to be bacteremic at the time of treatment. Survival was 0% in placebo treated animals. Animals treated with 7.5 units per kg exhibited 36% survival, those treated with 15 units per kg exhibited 43% survival, and those treated with 30 units per kg exhibited 70% survival (Table 4). Compared to placebo, these survival rates were statistically significant at p=0.0451, 0.0339, and 0.0031, respectively. The differences in survival between the 7.5, 15, and 30 unit per kg doses of anthrax immunoglobulin were not statistically significant. Anthrax immunoglobulin treated animals showed a statistically significant reduction in anthrax toxin when compared to placebo treated animals.

Table 4. Survival Rates in NZW Rabbits and Cynomolgus Macaques Treated with anthrax immunoglobulin:

^a Survival among animals that were bacteremic and toxemic prior to treatment
^b Two-sided Fisher’s exact test
^c Bonferroni-Holm adjusted one-sided Fisher’s exact test
^d Dose not evaluated in rabbits in this study
PI = Post-infection

Anthrax immunoglobulin Efficacy in Combination with Antibiotics

The efficacy of anthrax immunoglobulin administered with levofloxacin was determined in New Zealand white rabbits with systemic disease. No significant difference between the control (normal immune globulin [IGIV] plus levofloxacin) and treatment groups (anthrax immunoglobulin plus levofloxacin) was seen when combination treatment was delayed up to 60 hours post-challenge. There was no observed antagonism between levofloxacin and anthrax immunoglobulin in this study. This study also supported that anthrax immunoglobulin effectively cleared toxemia when administered with antibiotics. In anthrax immunoglobulin treated groups, all animals cleared PA toxemia post-anthrax immunoglobulin administration and only 4/31 (13%) of anthrax immunoglobulin treated animals exhibited a single transient positive PA result for toxemia at the 12 or 18 hour time point post-dosing. Placebo control animals exhibited more persistent toxemia, with 26/32 (81%) having positive PA results for 18 to 90 hours post-treatment.

In a second study, treatment was delayed beyond 60 hours to simulate a clinical scenario. When combination treatment was initiated at 60, 72, 84 or 96 hours post anthrax exposure, differences in survival were seen, but no statistically significant added survival benefit was observed between groups that received placebo (IGIV plus levofloxacin) or anthrax immunoglobulin (15 units per kg plus levofloxacin). An increase in survival was observed with anthrax immunoglobulin when treatment was delayed to 96 hours post exposure, but was not statistically significant. When treatment was delayed to 96 hours, survival was 25% (2/8) in the antibiotic plus IGIV control group and 71% (5/7) in the anthrax immunoglobulin plus levofloxacin group. A marginal improvement of 10 to 15% was observed at other time points, suggesting a trend in added benefit with anthrax immunoglobulin. This study also demonstrated a significant effect of anthrax immunoglobulin on toxemia. The majority of anthrax immunoglobulin treated animals became negative for PA (toxemia) within one hour post-infusion of anthrax immunoglobulin and remained negative, even with the delayed treatment from 60 to 96 hours post-anthrax challenge and high levels of toxemia pretreatment. In contrast, placebo treated animals remained toxemic up to three days after initiating antibiotic treatment.

The efficacy of anthrax immunoglobulin co-administered with levofloxacin was evaluated in New Zealand white rabbits when treatment was delayed to 96 hours after anthrax spore inhalation. The dose of levofloxacin was chosen to yield a comparable exposure to that achieved by the recommended dose in humans. Of the animals that survived to be treated (19% of those challenged), antibacterial drug plus anthrax immunoglobulin (15 units per kg) resulted in 58% (18/31) survival compared to 39% (13/33) survival in rabbits treated with antibacterial drug and IGIV placebo (p=0.14, Z-test).

When animals were stratified by pre-treatment toxemia (PA) in a post hoc analysis, added benefit was observed in animals treated with anthrax immunoglobulin and levofloxacin when they had pre-treatment PA levels between 200 and 800 ng/mL (p=0.02, Fisher’s exact test). When pre-treatment toxemia was low (PA <200 ng/mL), survival was greater than 90% in all animals, regardless of treatment (Table 5). Animals with very high levels of toxemia (>800 ng/mL) did not survive irrespective of the treatment administered.

Table 5. Survival Rates in New Zealand White Rabbits Stratified by Pre-treatment PA Levels:

Anthrax immunoglobulin and antibiotic combination treatment was also studied in the cynomolgus macaque model of inhalational anthrax. In this study, delay of initiation of treatment to 64 hours post anthrax exposure resulted in 75% (9/12) survival in the placebo plus ciprofloxacin treatment group versus 83% (10/12) survival in the anthrax immunoglobulin (15 units per kg) plus ciprofloxacin group (p=1).

No antagonism of anthrax immunoglobulin when administered with antibiotic as a concomitant therapy was observed.

Anthrax immunoglobulin in Post-exposure Prophylaxis

A post exposure prophylactic study assessed the survival following aerosol exposure to a lethal dose of anthrax spores (200 x LD₅₀) in New Zealand white rabbits administered anthrax immunoglobulin (7.5, 15 or 30 units TNA per kg) at 30 hours post-anthrax challenge compared to placebo controls. All three doses of anthrax immunoglobulin improved survival when given 30 hours post-anthrax challenge. When animals that were both bacteremic and toxemic were treated at 30 hours following challenge, there was a 22% (2/9) survival with a dose of 15 units TNA per kg and a 33% (4/12) survival with a dose of 30 units TNA per kg. All rabbits in the placebo arm died.