Determination of the Postexposure Prophylactic Benefit of Oral Azithromycin and Clarithromycin Against Inhalation Anthrax in Cynomolgus Macaques.

BACKGROUND: Sufficient and diverse medical countermeasures against severe pathogenic infections, such as inhalation anthrax, are a critical need. Azithromycin and clarithromycin are antimicrobials commonly used for both upper and lower respiratory infections. They inhibit protein synthesis by blocking the formation of the 50S ribosomal subunit. To expand the armamentarium, these 2 antibiotics were evaluated in a postexposure prophylactic model of inhalation anthrax in cynomolgus macaques. METHODS: This prophylaxis study had 4 test arms: azithromycin, clarithromycin, a levofloxacin control, and a placebo. Beginning 24 hours after exposure to a target challenge dose of 200 lethal dose 50 (LD50) of Bacillus anthracis Ames spores, animals were treated orally until 30 days postchallenge and then observed until 75 days postchallenge. RESULTS: The test group that received clarithromycin had a survival rate of 67%. The test group that received azithromycin had a survival rate of 50%, but the peak azithromycin plasma levels achieved were <30 ng/mL-much lower than the expected 410 ng/mL. The levofloxacin positive control had a survival rate of 50%; all of the negative controls succumbed to infection. CONCLUSIONS: The efficacy of clarithromycin prophylaxis was statistically significant compared with placebo, while azithromycin prophylaxis was indistinguishable from placebo. Given the low plasma concentrations of azithromycin achieved in the study, it is not surprising that half the animals succumbed to anthrax during the dosing period; the animals that survived beyond the time during which placebo control animals succumbed survived to the end of the observation period.

Evaluation of Amoxicillin and Amoxicillin-Clavulanate (Augmentin) for Antimicrobial Postexposure Prophylaxis Following Bacillus anthracis Inhalational Exposure in Cynomolgus Macaques.

Amoxicillin is a broad-spectrum antibiotic used to treat a variety of gram-positive and gram-negative infections, such as infections of the ear, nose, and throat, genitourinary tract, skin, and lower respiratory tract; gonorrhea; and Helicobacter pylori. The prophylactic benefit of both amoxicillin and Augmentin (amoxicillin-clavulanate for use against ß-lactamase-expressing bacteria) was evaluated for inhalation anthrax in cynomolgus macaques in 2 studies. A pilot study on amoxicillin-clavulanate that used a portion of the study animals demonstrated empirically that dosing twice a day was efficacious. In a subsequent study on both amoxicillin and amoxicillin-clavulanate that used the remaining study animals, the animals were treated orally every 12 hours on days 1-28 postchallenge and followed for an additional 60 days (total of 88 days from day of aerosol challenge to when the animals were culled). The animals from each treatment arm of the 2 studies were completely protected. All untreated animals succumbed to the infection. The degree of protection observed in this study suggests that both amoxicillin and amoxicillin-clavulanate, administered prophylactically over a period of 28 days after a lethal exposure to Bacillus anthracis spores, is sufficient for full protection.

ACTIVating Resources for the COVID-19 Pandemic: In Vivo Models for Vaccines and Therapeutics.

The Preclinical Working Group of Accelerating COVID-19 Therapeutic Interventions and Vaccines (ACTIV), a public-private partnership spearheaded by the National Institutes of Health, has been charged with identifying, prioritizing, and communicating SARS-CoV-2 preclinical resources. Reviewing SARS-CoV-2 animal model data facilitates standardization and harmonization and informs knowledge gaps and prioritization of limited resources. To date, mouse, hamster, ferret, guinea pig, and non-human primates have been investigated. Several species are permissive for SARS-CoV-2 replication, often exhibiting mild disease with resolution, reflecting most human COVID-19 cases. More severe disease develops in a few models, some associated with advanced age, a risk factor for human disease. This review provides a snapshot that recommends the suitability of models for testing vaccines and therapeutics, which may evolve as our understanding of COVID-19 disease biology improves. COVID-19 is a complex disease, and individual models recapitulate certain aspects of disease; therefore, the coordination and assessment of animal models is imperative.

The African Green Monkey Model of Pneumonic Plague and US Food and Drug Administration Approval of Antimicrobials Under the Animal Rule.

BACKGROUND: Additional treatment options for pneumonic plague, the most severe form of infection by Yersinia pestis, are needed, as past US Food and Drug Administration (FDA) approvals were not based on clinical trials that meet today's standards, and multiple drugs are sought to counter resistance or use in special populations. Due to the sporadic nature of outbreaks and the low number of pneumonic cases of disease, we sought FDA approval of antimicrobials for treatment under the Animal Efficacy Rule, where efficacy can be demonstrated in 1 or more well-characterized animal models that sufficiently represent human disease. METHODS: A model was developed in African green monkeys (AGMs) after challenge with a lethal dose of Y. pestis delivered as an aerosol, in 4 independent studies in 3 laboratories. The primary data points were bacteremia (daily), body temperature and heart rate (continuously monitored by telemetry), and survival. In antimicrobial efficacy studies, human-equivalent doses of gentamicin, ciprofloxacin, levofloxacin, and doxycycline were administered upon fever onset for 10 days. RESULTS: Disease in AGMs was similar to case reports of human disease. Fever was determined to be a reliable sign of disease and selected as a treatment trigger. Gentamicin was 60%-80% effective depending on the dose given to animals. Ciprofloxacin and levofloxacin were found to be >90% efficacious. These data were submitted to FDA and plague indications were approved. Doxycycline was less effective. CONCLUSIONS: The AGM model of pneumonic plague is reproducible, well-characterized, and mimics human disease. It has been used to support plague indications for fluoroquinolones and to test the efficacy of additional antimicrobials.

Effect of Delaying Treatment on Efficacy of Ciprofloxacin and Levofloxacin in the African Green Monkey Model of Pneumonic Plague.

BACKGROUND: Ciprofloxacin and levofloxacin, 2 fluoroquinolone antimicrobials, are ≥90% effective for the treatment of inhalational plague when administered within 2-6 hours of fever onset in African green monkeys (AGM). Based on data in the AGM model, these antimicrobials were approved under the Food and Drug Administration's Animal Efficacy Rule. However, that data did not address the issue of how long treatment with these antimicrobials would remain effective after fever onset. METHODS: The AGM model of pneumonic plague was used to explore the effect of delaying treatment with ciprofloxacin and levofloxacin on efficacy. In 2 studies, AGMs were challenged with inhaled lethal doses of Yersinia pestis. Treatment with ciprofloxacin and levofloxacin was initiated from 0 to up to 30 hours after fever onset. RESULTS: Challenged animals all developed fever within 78 hours and were treated with ciprofloxacin (n = 27) or levofloxacin (n = 29) at various predetermined time points postfever. When administered 10 hours after fever onset, 10 days of ciprofloxacin and levofloxacin treatment remained very effective (90 or 100%, respectively). The efficacy of both antimicrobials declined as treatment initiation was further delayed. Statistical analyses estimated the treatment delay times at which half of the AGMs were no longer expected to survive as 19.7 hours for ciprofloxacin and 26.5 hours for levofloxacin. CONCLUSIONS: This study demonstrates that there is a narrow window following fever onset during which ciprofloxacin and levofloxacin are fully effective treatment options for pneumonic plague in AGMs. Since the timing of disease is similar in humans and AGMs, these AGM data are reasonably likely to predict response times for treatment in humans.

Retrospective Analysis of Pneumonic Tularemia in Operation Whitecoat Human Subjects: Disease Progression and Tetracycline Efficacy.

Francisella tularensis is a highly infectious Gram-negative bacterium that is the etiologic agent of tularemia in animals and humans. The incidence of tularemia is very low with a lack of comprehensive data that describe disease in humans due to difficulty in understanding time and routes of exposure. Under the title Operation Whitecoat, researchers at Ft. Detrick, MD conducted 40 clinical studies of tularemia from 1958 to 1968. In these studies, one of the objectives was to evaluate candidate countermeasures for treatment or prophylaxis of disease after exposure to Francisella tularensis strain Schu S4 by inhalation. These studies were reviewed retrospectively to delineate the early signs and symptoms or natural history of pneumonic tularemia and examine the efficacy of tetracycline in controlled human clinical studies. Using vital signs, onset of fever was objectively defined and calculated for each subject, while Adverse Events reported after exposure were also used to define the timing of disease onset and symptoms of early disease. There was a dose response relationship between time to fever onset and exposed dose at 200 cfu (172.8 h), 700 cfu (163.2 h), 2,500 cfu (105.3 h), and 25,000 cfu (75.5 h). Onset of fever was typically the earliest sign of disease at all doses but was often accompanied by symptoms such as headache, myalgia, chest pain, and nausea, irrespective of dose except at 200 cfu where only 50% of subjects exhibited fever onset or symptoms. Examining the efficacy of different treatment regimens of tetracycline, ineffective treatments were indicated by relapse of disease (fever and Adverse Events) after cessation of antibiotic treatment. Stratification of the data suggested that treatment for <14 days or doses <2g/day was associated with increased percentage of subjects with relapse of disease symptoms. Although these types of human challenge studies would not be ethically possible now, the climate post-World War II supported human testing under rigorous conditions with informed consent. Thus, going back and analyzing these unique clinical human challenge studies has helped describe the course of infection and disease induced by a biothreat pathogen and possible countermeasures for treatment under controlled conditions.

Challenges and Benefits of Repurposing Products for Use during a Radiation Public Health Emergency: Lessons Learned from Biological Threats and other Disease Treatments.

The risk of a radiological or nuclear public health emergency is a major growing concern of the U.S. government. To address a potential incident and ensure that the government is prepared to respond to any subsequent civilian or military casualties, the U.S. Department of Health and Human Services and the Department of Defense have been charged with the development of medical countermeasures (MCMs) to treat the acute and delayed injuries that can result from radiation exposure. Because of the limited budgets in research and development and the high costs associated with bring promising approaches from the bench through advanced product development activities, and ultimately, to regulatory approval, the U.S. government places a priority on repurposing products for which there already exists relevant safety and other important information concerning their use in humans. Generating human data can be a costly and time-consuming process; therefore, the U.S. government has interest in drugs for which such relevant information has been established (e.g., products for another indication), and in determining if they could be repurposed for use as MCMs to treat radiation injuries as well as chemical and biological insults. To explore these possibilities, the National Institute of Allergy and Infectious Diseases (NIAID) convened a workshop including U.S. government, industry and academic subject matter experts, to discuss the challenges and benefits of repurposing products for a radiation indication. Topics covered included a discussion of U.S. government efforts (e.g. funding, stockpiling and making products available for study), as well unique regulatory and other challenges faced when repurposing patent protected or generic drugs. Other discussions involved lessons learned from industry on repurposing pre-license, pipeline products within drug development portfolios. This report reviews the information presented, as well as an overview of discussions from the meeting.

The Cynomolgus Macaque Natural History Model of Pneumonic Tularemia for Predicting Clinical Efficacy Under the Animal Rule.

Francisella tularensis is a highly infectious Gram-negative bacterium that is the etiologic agent of tularemia in animals and humans and a Tier 1 select agent. The natural incidence of pneumonic tularemia worldwide is very low; therefore, it is not feasible to conduct clinical efficacy testing of tularemia medical countermeasures (MCM) in human populations. Development and licensure of tularemia therapeutics and vaccines need to occur under the Food and Drug Administration's (FDA's) Animal Rule under which efficacy studies are conducted in well-characterized animal models that reflect the pathophysiology of human disease. The Tularemia Animal Model Qualification (AMQ) Working Group is seeking qualification of the cynomolgus macaque (Macaca fascicularis) model of pneumonic tularemia under Drug Development Tools Qualification Programs with the FDA based upon the results of studies described in this manuscript. Analysis of data on survival, average time to death, average time to fever onset, average interval between fever and death, and bacteremia; together with summaries of clinical signs, necropsy findings, and histopathology from the animals exposed to aerosolized F. tularensis Schu S4 in five natural history studies and one antibiotic efficacy study form the basis for the proposed cynomolgus macaque model. Results support the conclusion that signs of pneumonic tularemia in cynomolgus macaques exposed to 300-3,000 colony forming units (cfu) aerosolized F. tularensis Schu S4, under the conditions described herein, and human pneumonic tularemia cases are highly similar. Animal age, weight, and sex of animals challenged with 300-3,000 cfu Schu S4 did not impact fever onset in studies described herein. This study summarizes critical parameters and endpoints of a well-characterized cynomolgus macaque model of pneumonic tularemia and demonstrates this model is appropriate for qualification, and for testing efficacy of tularemia therapeutics under Animal Rule.

Accelerating Biomedical Discoveries through Rigor and Transparency.

Difficulties in reproducing published research findings have garnered a lot of press in recent years. As a funder of biomedical research, the National Institutes of Health (NIH) has taken measures to address underlying causes of low reproducibility. Extensive deliberations resulted in a policy, released in 2015, to enhance reproducibility through rigor and transparency. We briefly explain what led to the policy, describe its elements, provide examples and resources for the biomedical research community, and discuss the potential impact of the policy on translatability with a focus on research using animal models. Importantly, while increased attention to rigor and transparency may lead to an increase in the number of laboratory animals used in the near term, it will lead to more efficient and productive use of such resources in the long run. The translational value of animal studies will be improved through more rigorous assessment of experimental variables and data, leading to better assessments of the translational potential of animal models, for the benefit of the research community and society.