Anthrax:  Developing 
Drugs for Prophylaxis 
of Inhalational 
Anthrax 
Guidance for Industry 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Drug Evaluation and Research (CDER) 
 
May 2018 
Clinical/Antimicrobial 
   
Anthrax:  Developing 
Drugs for Prophylaxis 
of Inhalational 
Anthrax 
Guidance for Industry 
 
Additional copies are available from: 
 
Office of Communications, Division of Drug Information 
Center for Drug Evaluation and Research 
Food and Drug Administration 
10001 New Hampshire Ave., Hillandale Bldg., 4th Floor 
Silver Spring, MD  20993-0002 
Phone: 855-543-3784 or 301-796-3400; Fax: 301-431-6353; Email: druginfo@fda.hhs.gov 
https://www.fda.gov/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/default.htm 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Drug Evaluation and Research (CDER) 
 
May 2018 
Clinical/Antimicrobial 
   
TABLE OF CONTENTS 
 
I.  INTRODUCTION............................................................................................................. 1 
II.  BACKGROUND ............................................................................................................... 2 
A.  Historical Background................................................................................................................... 2 
B.  Indication for Prophylaxis of Inhalational Anthrax ................................................................... 2 
III.  DEVELOPMENT PROGRAM ....................................................................................... 3 
A.  General Considerations ................................................................................................................. 3 
1. Efficacy Considerations ................................................................................................................... 3 
2. Human Safety Considerations.......................................................................................................... 4 
3. Nonclinical Safety Considerations ................................................................................................... 5 
4. Clinical Pharmacology Considerations ........................................................................................... 5 
5. Microbiology Considerations .......................................................................................................... 5 
B.  Considerations for the Adequate and Well-Controlled Animal Efficacy Studies .................... 7 
1. Animal Models ................................................................................................................................. 7 
2. Study Conduct .................................................................................................................................. 7 
3. Bacterial Challenge ......................................................................................................................... 7 
4. Selection of the Dose for the Investigational Drug .......................................................................... 8 
5. Choice of Comparators .................................................................................................................... 9 
6. Efficacy Endpoints ........................................................................................................................... 9 
7. Study Pharmacokinetic Assessments .............................................................................................. 10 
8. Statistical Considerations .............................................................................................................. 10 
C.  Other Considerations................................................................................................................... 10 
1. Pediatrics ....................................................................................................................................... 10 
2. Postapproval Studies ..................................................................................................................... 11 
3. Labeling ......................................................................................................................................... 11 
REFERENCES ............................................................................................................................ 12 
 
 
  Contains Nonbinding Recommendations 
Anthrax:  Developing Drugs for 
Prophylaxis of Inhalational Anthrax 
Guidance for Industry1 
 
 
 
 
This guidance represents the current thinking of the Food and Drug Administration (FDA or Agency) on 
this topic.  It does not establish any rights for any person and is not binding on FDA or the public.  You 
can use an alternative approach if it satisfies the requirements of the applicable statutes and regulations.  
To discuss an alternative approach, contact the FDA office responsible for this guidance as listed on the 
title page. 
 
 
 
 
I.  INTRODUCTION  
 
The purpose of this guidance is to assist sponsors in the development of drugs for the indication 
of prophylaxis of inhalational anthrax in persons who have or may have inhaled aerosolized 
Bacillus anthracis spores but who have not yet manifested clinical evidence of disease.2  The 
indication also applies to persons with anticipated exposure to B. anthracis spores (e.g., first 
responders for anthrax incidents); in such cases, initiation of antibacterial therapy would begin 
immediately before entering the B. anthracis-contaminated environment.  For more information 
regarding the indication, see section II.B., Indication for Prophylaxis of Inhalational Anthrax.  
 
This guidance clarifies that drugs developed for the prophylaxis of inhalational anthrax are to be 
considered for approval under the animal rule.3  
 
This guidance does not contain discussion of the general issues of statistical analysis or clinical 
trial design.  Those topics are addressed in the ICH guidances for industry E9 Statistical 
                                                 
1 This guidance has been prepared by the Division of Anti-Infective Products in the Center for Drug Evaluation and 
Research at the Food and Drug Administration.  
 
2 For the purposes of this guidance, all references to drugs include both human drugs and therapeutic biological 
products such as therapeutic proteins and monoclonal antibodies, unless otherwise specified, and references to 
approval include new drug application approval for drugs or biologics license application licensure for therapeutic 
proteins and monoclonal antibodies.  Sponsors interested in developing other types of biological products, such as 
vaccines and immunoglobulin preparations, should contact the appropriate review division in the Center for 
Biologics Evaluation and Research. 
 
3 The animal rule sets forth a pathway for approval of drug or biological products when human efficacy studies are 
not ethical or feasible.  See 21 CFR part 314, subpart I, for drugs and 21 CFR part 601, subpart H, for biological 
products.  See also the guidance for industry Product Development Under the Animal Rule.  We update guidances 
periodically.  To make sure you have the most recent version of a guidance, check the FDA guidance web page at 
https://www.fda.gov/RegulatoryInformation/Guidances/default.htm.  
1  Contains Nonbinding Recommendations 
Principles for Clinical Trials and E10 Choice of Control Group and Related Issues in Clinical 
Trials, respectively.   
 
In general, FDAs guidance documents do not establish legally enforceable responsibilities.  
Instead, guidances describe the Agencys current thinking on a topic and should be viewed only 
as recommendations, unless specific regulatory or statutory requirements are cited.  The use of 
the word should in Agency guidances means that something is suggested or recommended, but 
not required.  
 
 
II.  BACKGROUND 
 
A.  Historical Background 
 
In the fall of 2001, B. anthracis spores (Ames strain) were used as an agent of bioterrorism and 
sent through the U.S. mail, resulting in cases of inhalational and cutaneous anthrax.  Post-
exposure prophylaxis of inhalational anthrax was administered to thousands of persons, most of 
whom received ciprofloxacin or doxycycline (Jernigan, Stephens, et al. 2002; Martin, Tierney, et 
al. 2005; Doolan, Freilich, et al. 2007; Inglesby, OToole, et al. 2002).  
 
At the time of the anthrax attacks, ciprofloxacin was already approved (in August 2000) for post-
exposure prophylaxis of inhalational anthrax under FDAs accelerated approval regulations.  In 
November 2001, a notice in the Federal Register clarified that penicillin G procaine and 
doxycycline, both of which included anthrax or B. anthracis in their previously approved 
labelings, are indicated for prophylaxis of inhalational anthrax.4  Levofloxacin also was 
approved (in November 2004) for this indication under the accelerated approval regulations.  
Subsequently, some drugs approved under the animal rule regulations received an indication for 
prophylaxis of inhalational anthrax but with a more limited indication as follows: 
 
for prophylaxis of inhalational anthrax when alternative therapies are not available or are 
not appropriate.5   
 
B.  Indication for Prophylaxis of Inhalational Anthrax 
 
A window of opportunity for preventing illness and reducing mortality exists between the time of 
inhalation of aerosolized B. anthracis spores and the development of signs and symptoms of 
inhalational anthrax.  The indication of prophylaxis of inhalational anthrax was previously 
known as inhalational anthrax (post-exposure)  to reduce the incidence or progression of 
disease following exposure to aerosolized B. anthracis.  However, situations can arise in which 
persons (e.g., first responders) anticipate an imminent risk of exposure to aerosolized B. 
                                                 
4 See Prescription Drug Products; Doxycycline and Penicillin G Procaine Administration for Inhalational Anthrax 
(Post-Exposure) (66 FR 55679, November 2, 2001). 
 
5 In 2012 and 2016, respectively, raxibacumab and obiltoxaximab were approved for treatment of inhalational 
anthrax caused by B. anthracis in combination with appropriate antibacterial drugs and for prophylaxis of 
inhalational anthrax when alternative therapies are not available or are not appropriate.  See section III.A., General 
Considerations.   
2  Contains Nonbinding Recommendations 
anthracis spores.  Starting therapy immediately before the anticipated or potential exposure can 
reduce the risk of illness and reduce mortality from inhalational anthrax.  Therefore, the 
indication has been revised.  The indication for drugs to reduce the risk of disease in persons who 
have inhaled, or are likely to imminently inhale, aerosolized B. anthracis spores, but who do not 
yet have the established disease, is now referred to as prophylaxis of inhalational anthrax. 
 
 
III.  DEVELOPMENT PROGRAM 
 
A.  General Considerations 
 
The antibacterial drugs recently approved for prophylaxis of inhalational anthrax were found to 
be safe and effective in a number of indications, marketed for many years, and prescribed to 
millions of patients before being approved for prophylaxis of inhalational anthrax.  Therefore, 
the level of experience with these drugs was quite extensive, and the safety profiles were well-
characterized beforehand.  Because a large number of people are expected to possibly receive 
these drugs as prophylaxis, FDA recommends that antibacterial drugs being developed for this 
indication have sufficient safety experience to assess the risk and benefit among people who are 
determined to be at risk for inhalational exposure to B. anthracis spores.  Sufficient human safety 
experience is unlikely to be obtained for an investigational antibacterial drug for which 
prophylaxis of inhalational anthrax is the only indication under development.  Thus, an 
indication for prophylaxis of inhalational anthrax is likely to be reserved almost exclusively for 
antibacterial drugs that have established uses and safety data in other infectious diseases.6  
 
Safety and efficacy information derived from the development and use of a drug for other 
indications may provide additional information for developing a drug for the prophylaxis of 
inhalational anthrax indication.  If there is substantially limited human safety and efficacy 
information available for evaluation of an investigational drug, sponsors should provide a 
proposed justification for the anticipated benefit that will offset the risk of the investigational 
drug for prophylaxis of inhalational anthrax.   
 
An anthrax vaccine may be administered to persons receiving a drug for prophylaxis of 
inhalational anthrax.  Sponsors should discuss with FDA the data that would be collected to 
assess whether there are drug-vaccine interactions.. 
 
1.  Efficacy Considerations 
 
Definitive human efficacy studies cannot be conducted because naturally occurring inhalational 
anthrax is extremely rare and it would be unethical to deliberately expose healthy human 
volunteers to B. anthracis spores; thus, as previously noted, drugs developed for prophylaxis of 
                                                 
6 In addition, in some circumstances when a drug appears to offer potential benefit complementary to drugs already 
approved for the proposed indication but may not be studied for broader indications in other diseases, a more limited 
indication (e.g., reserved for use when an effective treatment regimen cannot be otherwise provided) for prophylaxis 
of inhalational anthrax may be considered (e.g., raxibacumab). 
 
3  Contains Nonbinding Recommendations 
inhalational anthrax should be developed for approval consideration under the animal rule.7  In 
general, FDA relies on evidence from animal studies to provide substantial evidence of 
effectiveness to support approval only when the four criteria listed in the animal rule regulations, 
as follows, are met:8 
 
(1) There is a reasonably well-understood pathophysiological mechanism of the toxicity of 
the substance and its prevention or substantial reduction by the product;  
 
(2) The effect is demonstrated in more than one animal species expected to react with a 
response predictive for humans, unless the effect is demonstrated in a single animal 
species that represents a sufficiently well-characterized animal model for predicting the 
response in humans; 
 
(3) The animal study endpoint is clearly related to the desired benefit in humans, generally 
the enhancement of survival or prevention of major morbidity; and 
 
(4) The data or information on the kinetics and pharmacodynamics of the product or other 
relevant data or information, in animals and humans, allows selection of an effective dose 
in humans. 
 
FDA emphasizes that development proposals for prophylaxis of inhalational anthrax may be 
more convincing if the drug is shown to be safe and effective in the treatment of other relevant 
infectious diseases (e.g., certain types of pneumonia).   
 
2.  Human Safety Considerations 
 
Drugs evaluated for efficacy under the animal rule are evaluated for safety under the existing 
requirements for establishing the safety of new drugs (21 CFR 314.50(d)(5)(vi) and 21 CFR 
314.610(a) for drugs and 21 CFR 601.2(a) and 21 CFR 601.91 for biological products).  The 
risks of the use of any drug are weighed against its benefits in the populations likely to use the 
drug for the stated purpose.  For antibacterial drugs, the anticipated duration of therapy for 
prophylaxis of inhalational anthrax is 60 days.  The drug development plan should address 
assembling a safety database adequate to support the proposed dose and duration for prophylaxis 
of inhalational anthrax.  This is important because shorter-duration or lower-dose uses of 
previously approved or studied antibacterial drugs may demonstrate safety concerns relevant for 
both short- and long-term therapy, but those uses cannot rule out additional safety concerns with 
longer-duration or higher-dose uses of the drug.  Sponsors should discuss with FDA the 
appropriate size and nature of the preapproval safety database.  
 
                                                 
7 See 21 CFR part 314, subpart I, for drugs and 21 CFR part 601, subpart H, for biological products.  See also the 
guidance for industry Product Development Under the Animal Rule.  
 
8 See 21 CFR 314.610 and 601.91.  For this guidance, the term substance refers to B. anthracis, and the term 
product refers to an investigational drug being evaluated for prophylaxis of inhalational anthrax. 
 
4  Contains Nonbinding Recommendations 
3.  Nonclinical Safety Considerations  
 
Guidances for industry are available to provide information for sponsors on general nonclinical 
safety considerations for drug development.9  To support the indication for prophylaxis of 
inhalational anthrax, animal toxicity studies in two or more species (e.g., rat, mouse, dog, or 
monkey) are recommended to characterize nonclinical safety.  For a previously approved 
antibacterial drug for which the nonclinical safety characterization and accumulated clinical data 
on the use of the drug support a 60-day duration of therapy, the available nonclinical safety data 
are usually sufficient.  
   
4.  Clinical Pharmacology Considerations 
 
An important component to establishing substantial evidence of effectiveness of a drug approved 
according to the animal rule regulations is the selection of an effective dose in humans based on 
pharmacokinetics and pharmacodynamics of the drug in animals and humans or other relevant 
information (21 CFR 314.610(a)(4) for drugs and 21 CFR 601.91(a)(4) for biological products).  
Because effectiveness of drugs for prophylaxis of inhalational anthrax cannot be tested in 
humans, a comparison between systemic drug exposures achieved in healthy human subjects and 
those observed in animal models of inhalational anthrax obtained in the adequate and well-
controlled animal efficacy studies is used to support the selection of an effective dose in humans.  
This comparison should take into account the variability of exposure parameters in both animals 
and humans, and any outlying values of exposure in humans should be greater than those 
associated with efficacy in animals, to minimize the possibility of subtherapeutic exposures in 
humans.10  Sponsors should discuss with FDA whether information other than pharmacokinetics 
and pharmacodynamics can support the selection of an effective dose and regimen in humans. 
 
The drugs absorption, distribution, metabolism, and excretion should be characterized and 
plasma protein binding determined both in the animal species selected for efficacy testing and in 
humans.  Obtaining pharmacokinetic (PK) data for specific populations (e.g., geriatrics, pregnant 
women, obese/morbidly obese patients, patients with renal or hepatic impairment, and pediatrics, 
if possible (see section III.C.1., Pediatrics)) is recommended, as well as conducting studies to 
investigate the potential for drug-drug interactions with medicinal products likely to be co-
administered in the clinical scenario.   
 
5.  Microbiology Considerations  
 
Sponsors should provide information on the in vitro susceptibility of a spectrum of B. anthracis 
isolates to the investigational drug.  This testing should be performed on approximately 50 
isolates, provided there is evidence that all isolates have uniformly low variability in minimum 
inhibitory concentrations (MICs) to the investigational drug.  If there is multiple-fold variability 
                                                 
9 See the Pharmacology/Toxicology guidance web page at 
https://www.fda.gov/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/ucm065014.htm; for example, 
see the guidance for industry Content and Format of Investigational New Drug Applications (INDs) for Phase 1 
Studies of Drugs, Including Well-Characterized, Therapeutic, Biotechnology-Derived Products and the ICH 
guidance for industry S6 Preclinical Safety Evaluation of Biotechnology-Derived Pharmaceuticals. 
 
10 See the guidance for industry Product Development Under the Animal Rule.   
5  Contains Nonbinding Recommendations 
in the MICs, data on a larger number of isolates should be submitted (up to 100 isolates).  A 
variety of isolates should be selected to represent geographic diversity, human and animal 
isolates, and naturally occurring antibacterial resistance. In addition, susceptibility testing using 
several well-characterized strains (including Vollum, Ames, and Sterne) and genetically diverse 
strains from an established repository should be performed.  Sponsors should discuss with the 
FDA if they believe that the breadth of genetic variation can be captured using a different or 
smaller number of strains/isolates or if there are difficulties obtaining an adequate number of 
isolates for testing. Reproducibility of the results should be demonstrated at different 
laboratories.  For example, testing should be performed in at least two laboratories with three or 
more of the same isolates tested in each laboratory.11   
 
Initial susceptibility testing of genetically diverse repository isolates might provide some initial 
insight into potential mechanisms of resistance to the investigational drug if not already known.  
Sponsors should discuss with FDA their proposed approaches for further assessment of the 
potential for resistance emergence and should obtain appropriate review (including through their 
institutions and funding agencies) of any elements that might raise dual-use issues of concern.12  
This discussion should include plans for susceptibility assessment of any bacteria cultured from 
animals that develop anthrax disease during prophylaxis or in the follow-up period.  Post-
treatment MIC values should be compared to the baseline MIC values.  This information may be 
important for understanding the mechanism of prophylaxis failures and its relevance to dosing 
considerations.  If antibacterial resistance develops spontaneously following exposure to the 
investigational drug, the mechanism of resistance should be characterized, when possible.   
 
Drugs that have an FDA-approved indication for prophylaxis of inhalational anthrax (such as 
ciprofloxacin, levofloxacin, doxycycline, or penicillin G procaine) should be included in 
susceptibility tests as control drugs.   
 
The details of the procedure and methods for susceptibility testing should be provided.  
Susceptibility testing should be performed using a standardized method, such as that 
recommended by the Clinical and Laboratory Standards Institute (CLSI).13  If an alternative or 
experimental testing method is used, then details of the method and performance characteristics 
should be provided.  The range of concentrations to be tested should be sufficiently broad to 
ensure that MICs are reported as a specific value.  
 
Laboratory work with B. anthracis must be conducted in compliance with the requirements of 
the select agent regulations (42 CFR part 73) and should incorporate relevant biosafety 
procedures.  For more information regarding biosafety procedures, sponsors should contact the 
                                                 
11 For more information about microbiology considerations, see the guidance for industry Microbiology Data for 
Systemic Antibacterial Drugs  Development, Analysis, and Presentation. 
 
12 See the Dual Use of Concern web page on the Department of Health and Human Services Public Health 
Emergency website available at https://www.phe.gov/s3/dualuse/Pages/default.aspx.  
 
13 CLSI publishes documents, which are updated periodically, that describe standardized susceptibility testing.  
These documents can be found at http://clsi.org.  
 
6  Contains Nonbinding Recommendations 
Centers for Disease Control and Prevention (CDC) (https://www.CDC.gov) and the National 
Institutes of Health (https://www.nih.gov/research-training) (CDC 2009). 
 
B.  Considerations for the Adequate and Well-Controlled Animal Efficacy 
Studies 
 
1.  Animal Models 
 
Before initiating the animal studies, sponsors should discuss with FDA the proposed animal 
models (in which efficacy will be tested) and the study designs for the adequate and well-
controlled animal efficacy studies to obtain concurrence on the details of the models and the 
design of the studies.14  Nonhuman primate study data together with extensive human experience 
in other infections contributed to the approvals of ciprofloxacin and levofloxacin discussed 
above, while combinations of nonhuman primate and rabbit data contributed to more recent 
development plans for anthrax treatment and prophylaxis under the animal rule.  Sponsors should 
discuss with FDA the appropriateness of specific animal models and combinations of models for 
proposed drug development programs, including the time of initiation of the investigational 
therapy for the prophylaxis of inhalational anthrax.  
 
2.  Study Conduct 
 
FDA considers the good laboratory practice (GLP) for nonclinical laboratory studies regulations 
to be an established and relevant system for ensuring data quality and integrity.  FDA 
recommends the use of GLP, to the extent practicable, for these studies.15  Before initiating the 
studies, sponsors should identify exceptions to GLP regulations, if any, and seek concurrence 
from FDA on alternative methods to ensure data quality and integrity in the event of such 
exceptions.   
 
Animal studies must comply with the applicable laws and regulations as outlined in the Animal 
Welfare Act (7 U.S.C. 2131 et seq.) and the U.S. Public Health Service Policy on Humane Care 
and Use of Laboratory Animals.16  FDA recommends following the principles of replacement, 
reduction, and refinement of the use of animals in biomedical research (Russell and Burch 1959). 
 
3.  Bacterial Challenge 
 
Well-characterized strains of B. anthracis with known virulence in humans and the chosen 
animal species should be used for challenge.  Mortality is expected to be greater than or equal to 
90 percent in the infected, untreated control group.   
 
                                                 
14 See the guidance for industry Product Development Under the Animal Rule.   
 
15 See 21 CFR part 58 and the guidance for industry Product Development Under the Animal Rule. 
 
16 The policy document is accessible at https://grants.nih.gov/grants/olaw/references/PHSPolicyLabAnimals.pdf.  
See also Appendix A in the guidance for industry Product Development Under the Animal Rule. 
7  Contains Nonbinding Recommendations 
The route of B. anthracis exposure in the animal efficacy studies should be aerosol inhalation, as 
is anticipated in humans from an intentional release.  Sponsors wishing to explore the possibility 
of administering spores via direct placement into the trachea or nasal passages should discuss 
these plans with FDA early in the development program. 
 
The preparation of the inoculum of spores to be used for the inhalational challenge should be 
standardized and well characterized.  Standardization and optimization of the inoculum 
concentration are important because the animal survival time may vary with the experimental 
conditions (e.g., animal species, strain of B. anthracis, method of inoculum preparation, 
inoculum size, and exposure time).17 
 
4.  Selection of the Dose for the Investigational Drug  
 
The selection of the dose for the investigational drug to be studied in the adequate and well-
controlled animal studies should be based on an understanding of the exposure-response 
relationship in the proposed animal model and an understanding of any differences in absorption, 
distribution, metabolism, and excretion of the drug between humans and the proposed animal 
species.18  Sponsors should provide evidence to support a conclusion that humans receiving the 
proposed dose would safely and reliably have exposures to the drug greater than therapeutic 
exposures observed in the animals in the efficacy studies used to support approval.  Achieving a 
higher exposure in humans is important to minimize the possibility of subtherapeutic exposures.  
The human dose selected for this indication should also be adequately supported by human 
safety data.  In general for antibacterial drugs, the duration of administration in the adequate and 
well-controlled animal studies has been approximately 30 days; this has allowed for 
demonstration of robust protective effects in the nonhuman primate model, whereas the 60-day 
recommended human antibacterial drug regimen addresses the possibility of occasional later 
spore germination.19   
 
Before conducting the adequate and well-controlled animal efficacy studies, the 
PK/pharmacodynamic (PD) parameters that correlate with activity and efficacy of the drug 
                                                 
17 For example, in the Friedlander and colleagues study (Friedlander, Welkos, et al. 1993), rhesus monkeys were 
infected with approximately 5.5 x 105 spores, a mean of 11 times the amount that kills 50 percent of the test animals 
(LD ), with a range of 5 to 30 times the LD  of Vollum 1B strain of B. anthracis.  In another animal study included 
50 50
in labeling for levofloxacin, rhesus monkeys were infected with approximately 2.7 x 106 spores (a mean of 49 times 
the LD ), with a range of 17 to 118 times the LD  of Ames strain of B. anthracis.   
50 50
 
18 See 21 CFR 314.610(a)(4) and 21 CFR 601.91(a)(4). 
 
19 Discussions of the duration of dosing of antibacterial drugs after inhalational exposure to anthrax spores have 
focused on the objective of preventing disease arising from germination of inhaled spores until sufficient spore 
clearance occurs to substantially reduce the risk of disease developing after drug cessation.  For example, during the 
Anti-Infective Drugs Advisory Committee meeting on July 28, 2000 (transcript available at https://wayback.archive-
it.org/7993/20170403222328/https://www.fda.gov/ohrms/dockets/ac/cder00.htm#Anti-Infective), discussions 
suggested that post-exposure prophylaxis durations up to 20 days might delay but not prevent disease and death in 
nonhuman primates, while 30 days might provide statistically significant but not complete protection.  A total of 60 
days was agreed upon as a reasonable duration to recommend for humans based on the combination of animal 
survival and spore clearance data.  Drugs with major differences in mechanism of action, dosing feasibility, or risk-
benefit assessment might raise additional considerations that sponsors should discuss with FDA on a case-by-case 
basis (Bell, Kozarsky, et al. 2002; Inglesby, Henderson, et al. 1999).  
8  Contains Nonbinding Recommendations 
should be identified via in vitro PK/PD approaches and in animal models.  This information can 
guide the selection of doses to be tested in adequate and well-controlled animal studies to 
evaluate efficacy.  In vitro studies can be used to determine potentially relevant PK/PD 
relationships between PK parameters, for example the maximal concentration (C ) and the area 
max
under the curve (AUC) and susceptibility based on the MIC.  Nonclinical studies then can be 
used to identify potentially relevant PK/PD parameters (e.g., C /MIC ratio, AUC/MIC ratio, 
max
the time the concentration remains above the MIC) that could correlate with an effective 
response.  Similar PK/PD parameters should be identified for humans to support human dose 
selection.  
 
5.  Choice of Comparators 
 
A control group should be included.  This can serve as an untreated control, or placebo/vehicle 
control, to verify aspects of study conduct and bacterial inoculum preparation by comparing the 
progression of disease in the absence of treatment to what is anticipated based on previous 
experience with that animal model.  A randomized, masked (blinded) design is particularly 
important for this type of animal study to minimize the risk that comparisons between treatment 
and control groups could be affected by differences in baseline characteristics, supportive care, 
clinical evaluation, or use of euthanasia criteria based on treatment group designation.  Sponsors 
may wish to consider and discuss with FDA when to include an adequately powered third active 
control arm.   
 
6.  Efficacy Endpoints  
 
The primary endpoint should be survival to the end of the study.  When survival is used as an 
endpoint in animal studies, sponsors should prospectively define and include in the protocol 
euthanasia criteria to reduce or eliminate unnecessary terminal distress and to support study 
goals.  Furthermore, sponsors should consider incorporating telemetry with real-time monitoring 
into euthanasia criteria, if feasible.  Sponsors should increase observation frequency around the 
time of major morbidity or death to prevent unrelieved pain or distress and to minimize a 
potential compromise or loss of data.  The proportion of animals achieving the primary endpoint 
should be compared between the drug group and the control group.  Secondary endpoints should 
include bacteremia at different time intervals during or after treatment and a quantitation of the 
microbial burden in infected organs and/or tissues (e.g., blood, spleen, liver, brain, lymph nodes, 
cerebrospinal fluid) collected at the time of necropsy.  
 
FDA recommends the assessment of the primary efficacy endpoint (survival) after a period of 
observation following completion of treatment (e.g., up to and including 30 days following the 
end of treatment).  A complete histopathologic evaluation should be performed on animals that 
die during the study, including animals that met prespecified criteria for euthanasia (Bregman, 
Alder, et al. 2003; Jacobs, El Hage, et al. 2003).  Sponsors should provide a justification for their 
euthanasia criteria and discuss them with FDA before initiating the protocol.20 
 
                                                 
20 See the guidance for industry Product Development Under the Animal Rule. 
9  Contains Nonbinding Recommendations 
7.  Study Pharmacokinetic Assessments 
 
PK data should be collected for the antibacterial drugs tested in the adequate and well-controlled 
animal studies.  Determining the systemic exposure that was achieved in surviving animals 
exposed to B. anthracis spores and that prevented the inhalational anthrax infection and 
consequent death is necessary for comparison to human exposures for the determination of a 
human dose (see 21 CFR 314.610(a)(4) and section III.A.4., Clinical Pharmacology 
Considerations).  Therefore, blood samples for PK analysis should be collected from each 
animal, and sample size (number of animals) and PK sampling strategies should be adequate to 
characterize relevant exposure parameters.  For nonantibacterial drugs, sponsors should discuss 
with FDA whether other important assessments are also needed to allow for the selection of an 
effective dose in humans. 
 
8.  Statistical Considerations 
 
The goal of the adequate and well-controlled animal studies should be to demonstrate that the 
investigational drug is statistically superior to placebo and confers a treatment effect considered 
likely to be clinically meaningful.  Sponsors should ensure that euthanasia criteria and any 
appropriate and needed supportive care are justified, adequately described in the protocol, and 
applied consistently across treatment groups through measures such as investigator blinding.21  
These considerations, power calculations, and proposed statistical analyses should be discussed 
with FDA before studies are initiated. 
 
C.  Other Considerations 
 
1.  Pediatrics 
 
Sponsors are encouraged to begin discussions about their pediatric clinical development plans as 
early as is feasible because pediatric studies are a required part of the overall drug development 
program.  Sponsors are required to submit pediatric study plans no later than 60 days after an 
end-of-phase 2 meeting or such other time as may be agreed upon by FDA and the sponsor.22 
 
Obtaining data to support pediatric dosing is challenging.  A study giving healthy children an 
investigational drug for the purpose of obtaining PK data would not likely meet the criteria for 
institutional review board approval of pediatric studies under 21 CFR part 50, subpart D, for 
several reasons including that the administration of an investigational drug would present more 
than minimal risk, that healthy, unexposed children would have no prospect of direct benefit 
from the administration of the investigational drug, and that naturally occurring exposure to 
anthrax spores would be unlikely for children in the United States. 
                                                 
21 For definitions of supportive care, see the guidance for industry Product Development Under the Animal Rule.   
 
22 See the Pediatric Research Equity Act (Public Law 108-155; section 505B(e)(2)(A) of the Federal Food, Drug, 
and Cosmetic Act; 21 U.S.C. 355B) as amended by the Food and Drug Administration Safety and Innovation Act 
(Public Law 112-144).  See also the draft guidance for industry Pediatric Study Plans:  Content of and Process for 
Submitting Initial Pediatric Study Plans and Amended Initial Pediatric Study Plans.  When final, this guidance will 
represent the FDAs current thinking on this topic.  For the most recent version of a guidance, check the FDA 
guidance web page at https://www.fda.gov/RegulatoryInformation/Guidances/default.htm.  
10  Contains Nonbinding Recommendations 
 
Accordingly, other approaches to pediatric dose selection should be explored, such as applying 
PK data from the use of the investigational drug for other more common diseases, obtaining 
population PK data during use of the investigational drug if specific situations warranting 
pediatric use arise during drug development, modeling pediatric exposure from existing adult 
data from the investigational drug, or using data available from other sufficiently similar drugs.  
For example, PK modeling served as the basis for raxibacumabs pediatric dosing 
recommendations based on existing adult raxibacumab exposure data in combination with PK 
data in adults and pediatric patients from other monoclonal antibodies.  Regardless of the chosen 
approach, the objective is to derive the dose and administration regimens that are predicted to 
provide the pediatric population with adequate drug exposure.23 
 
2.  Postapproval Studies 
 
If a drug is approved under the animal rule regulations, postmarketing studies (e.g., field studies) 
are required to provide evaluation of safety and clinical benefit if circumstances arise in which a 
study would be feasible and ethical (in the case of a drug for prophylaxis of inhalational anthrax 
if the drug is used in the event of an accidental or intentional exposure to aerosolized B. 
anthracis).24  A plan for a postmarketing study is required as part of a new drug application or 
biologics license application under the animal rule.25  
 
3.  Labeling 
 
The indication granted will be prophylaxis of inhalational anthrax.  In addition to providing the 
appropriate dosing regimen in the DOSAGE AND ADMINISTRATION section, the prescribing 
information should list the organism Bacillus anthracis under the Antimicrobial Activity heading 
in the Microbiology subsection and provide a summary of the efficacy data that served as the 
basis of approval in the CLINICAL STUDIES section.  Patient labeling (e.g., Medication Guide, 
patient information) should also be drafted and discussed with FDA, including, but not 
necessarily limited to, the explanation that, for ethical and feasibility reasons, the drugs approval 
is based on efficacy studies conducted in animals alone (21 CFR 314.610(b)(3) for drugs and 21 
CFR 601.91(b)(3) for biological products).   
 
                                                 
23 See, for example, documents for the Anti-Infective Drugs Advisory Committee meeting of November 2, 2012 
(available at https://wayback.archive-
it.org/7993/20170403223651/https://www.fda.gov/AdvisoryCommittees/CommitteesMeetingMaterials/Drugs/Anti-
InfectiveDrugsAdvisoryCommittee/ucm293600.htm); and the biologics license application review documents of 
raxibacumab found on the Drugs@FDA web page (available at 
https://www.accessdata.fda.gov/scripts/cder/daf/index.cfm?event=overview.process&ApplNo=125349).   
 
24 21 CFR 314.610(b)(1) for drugs and 21 CFR 601.91(b)(1) for biological products. 
 
25 Ibid. 
11  Contains Nonbinding Recommendations 
REFERENCES 
 
Bell DM, Kozarsky PE, and Stephens DS, 2002, Clinical Issues in the Prophylaxis, Diagnosis, 
and Treatment of Anthrax, Emerg Infect Dis, 8(2):222225. 
 
Brachman PS, 1980, Inhalational Anthrax, Ann NY Acad Sci, 353:8393. 
 
Bregman CL, Alder RR, Morton DG, et al., 2003, Recommended Tissue List for Histopathologic 
Examination in Repeat-dose Toxicity and Carcinogenicity Studies:  A Proposal of the Society of 
Toxicologic Pathology (STP), Toxicol Pathol, 31:252253. 
 
CDC, 2006, Inhalation Anthrax Associated with Dried Animal Hides  Pennsylvania and New 
York City, 2006, MMWR Morb Mortal Wkly Rep, 55(10):280282. 
 
CDC/NIH, 2009, Biosafety in Microbiological and Biomedical Laboratories (BMBL) 5th 
Edition. 
 
Doolan DL, Freilich DA, Brice GT, et al., 2007, The U.S. Capital Bioterrorism Anthrax 
Exposures:  Clinical Epidemiological and Immunological Characteristics, J Infect Dis, 
195(2):174184. 
 
Friedlander AM, Welkos SL, Pitt MLM, et al., 1993, Post-Exposure Prophylaxis Against 
Experimental Inhalation Anthrax, J Infect Dis, 167:12391243. 
 
Henderson DW, Peacock S, and Belton FC, 1956, Observations on the Prophylaxis of 
Experimental Pulmonary Anthrax in the Monkey, J Hyg, 54:2836. 
 
Inglesby TV, Henderson DA, Bartlett JG, et al., 1999, Anthrax as a Biological Weapon, JAMA, 
281(18):17351745. 
 
Inglesby TV, OToole T, Henderson DA, et al., 2002, Anthrax as a Biological Weapon; Updated 
Recommendations for Management, JAMA, 288:22362252. 
 
Jacobs A, El Hage J, Hastings K, et al., 2003, Recommended Tissue List for Histopathologic 
Examination:  Letters to the Editor, Toxicol Pathol, 31:571. 
 
Jernigan DB, Stephens DS, Ashford DA, et al., 2002, Investigation of Bioterrorism-related 
Anthrax, United States, 2001:  Epidemiologic Findings, Emerg Infect Dis, 8(10):10191028. 
 
Martin SW, Tierney BC, Aranas A, et.al., 2005, An Overview of Adverse Events Reported by 
Participants in CDCs Anthrax Vaccine and Antimicrobial Availability Program, 
Pharmacoepidemiol Drug Saf, 14(6):393401. 
 
Meselson M, Guillemin J, Hugh-Jones M, et al., 1994, The Sverdlovsk Anthrax Outbreak of 
1979, Science, 266:12021208. 
 
12  Contains Nonbinding Recommendations 
Plotkin SA, Brachman PS, Utell M, Bumford FH, and Atchison MM, 2002, An Epidemic of 
Inhalation Anthrax, The First in the Twentieth Century:  I.  Clinical Features, Am J Med, 112;4
12 (originally published as Am J Med 1960, 29:9921001). 
 
Russell, WMS and RL Burch, 1959, The Principles of Humane Experimental Techniques, 
London: Methuen and Co. Ltd.  Reissued: 1992, Herts, UK: Universities Federation for Animal 
Welfare. 
 
Suffin SC, Carnes WH, Kaufmann AF, et al., 1978, Inhalational Anthrax in a Home Craftsman, 
Hum Pathol, 9(5):594597. 
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