Guidance for Industry 

Considerations for Developmental 

Toxicity Studies for Preventive and 

Therapeutic Vaccines for Infectious 

Disease Indications 

Additional copies of this guidance are available from the Office of Communication, 
Training and Manufacturers Assistance (HFM-40), 1401 Rockville Pike, suite 200N, 
Rockville, MD 20852-1448 or by calling 1-800-835-4709 or 301-827-1800, or from the 
Internet at http://www.fda.gov/cber/guidelines.htm. 
For questions on the content of this guidance, contact CBER Division of Vaccines and 
Related Products Applications at (301) 827-3070. 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Biologics Evaluation and Research 
February 2006  Contains Nonbinding Recommendations
TABLE OF CONTENTS 

I.	 INTRODUCTION............................................................................................................. 1

II.	 BACKGROUND ............................................................................................................... 2

III.	 VACCINE TARGET POPULATION AND TIMING OF PRE-CLINICAL 

DEVELOPMENTAL TOXICITY STUDIES ................................................................ 3

IV.	 DESIGN OF DEVELOPMENTAL TOXICITY STUDIES..........................................4

A.	 General Considerations and Recommendations ................................................ 4

B.	 Specific Considerations ........................................................................................ 6

V.	 REFERENCES................................................................................................................ 12

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1 
Guidance for Industry
Considerations for Developmental Toxicity Studies for Preventive 
and Therapeutic Vaccines for Infectious Disease Indications 
This guidance represents the Food and Drug Administrations (FDAs) current thinking on this 
topic.  It does not create or confer any rights for or on any person and does not operate to bind 
FDA or the public.  You can use an alternative approach if the approach satisfies the 
requirements of the applicable statutes and regulations.  If you want to discuss an alternative 
approach, contact the appropriate FDA staff. If you cannot identify the appropriate FDA staff, 
call the appropriate number listed on the title page of this guidance. 
I.  INTRODUCTION 
This guidance is intended to provide you, sponsors, with recommendations for the conduct of 
developmental toxicity2 studies for investigational preventive and therapeutic vaccines for 
infectious disease indications. The recommendations set forth in this document pertain to the 
assessment of the developmental toxicity potential of preventive and therapeutic vaccines for 
infectious diseases indicated for females of childbearing potential and pregnant individuals.3 
This guidance applies prospectively to investigational vaccines, i.e., vaccines under 
investigational new drug applications (IND) and vaccines the subject of a new biologics license 
application (BLA). These recommendations do not apply retrospectively to already licensed 
vaccines except those the subject of additional INDs. This guidance document finalizes the draft 
guidance entitled Guidance for Industry: Considerations for Reproductive Toxicity Studies for 
Preventive Vaccines for Infectious Disease Indications dated August 2000 (65 FR 54534; 
September 8, 2000). 
FDAs guidance documents, including this guidance, do not establish legally enforceable 
responsibilities. Instead, guidances describe the FDAs 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 FDAs guidances means that something is suggested or 
1 The Maternal Immunization Working Group in the Center for Biologics Evaluation and Research at the Food and 
Drug Administration prepared this guidance.  The document was revised based on public comments submitted to 
the Division of Dockets Management on the draft guidance dated August 2000, and on recommendations made by 
an expert panel convened at a Workshop entitled Non-clinical Safety Evaluation of Preventive Vaccines: Recent 
Advances and Regulatory Considerations held December 2 & 3, 2002, Arlington, VA, discussing approaches for 
developmental toxicity assessments for vaccines. 
2 Developmental toxicity is any adverse effect induced prior to attainment of adult life.  This includes effects 
induced or manifested in the embryonic or fetal period and those induced or manifested postnatally (Ref. 1). 
3 This document does not address concerns regarding male reproductive toxicity and male and female fertility 
studies. 
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recommended, but not required. 
II.  BACKGROUND 
The Center for Biologics Evaluation and Research (CBER) reviews a broad spectrum of 
applications for investigational vaccines intended for the prevention and treatment of infectious 
diseases and indicated for immunization of adolescents and adults.  Thus, the target population 
for vaccines often includes females of childbearing potential who may become pregnant during 
the vaccination period. A number of vaccines are in clinical development specifically for 
maternal immunization indications with the goal of preventing infectious disease in the pregnant 
mother and/or neonate through passive antibody transfer from mother to fetus.  Unless the 
vaccine is specifically indicated for maternal immunization, no studies are conducted prior to 
product licensure to determine the vaccines safety in pregnant women.  During clinical 
development of most vaccines not specifically intended for use during pregnancy, pregnant 
women are ineligible to participate in clinical trials.  If pregnancy occurs during a study, 
treatment is usually discontinued and the woman does not receive additional immunizations.  
Consequently, there are few clinical data to address developmental toxicity of the vaccine in 
pregnant women or females of childbearing potential at the time of product licensure.   
As more women of childbearing potential participate in clinical trials and as more preventive and 
therapeutic vaccines are developed for adolescents and adults, there is increasing concern about 
the unintentional exposure of an embryo/fetus before information is available about the risk 
versus benefit of the vaccine. Also, following approval, vaccines not indicated for use during 
pregnancy may be recommended by health policy makers for use in pregnant women (Ref. 2).  In 
addition, use of licensed vaccines in females of childbearing potential will likely result in 
inadvertent exposure of the pregnant woman and her fetus to the vaccine.  Considering that more 
than half of pregnancies in the U.S. are unintended, it is unlikely that vaccine exposure would be 
avoided in these pregnancies prior to their clinical recognition (Ref. 3). In these situations, in the 
absence of clinical data, it is difficult for the practitioner to make an informed risk assessment, 
even in situations where immunization of pregnant women may be appropriate.  
Until recently, few licensed vaccines have been tested for developmental toxicity in animals 
prior to their use in humans.  However, for the reasons listed above, we, FDA, recommend that 
you address during the IND studies the risks versus the benefits of immunization programs for 
pregnant women and/or females of childbearing potential by performing developmental toxicity 
studies in animal models.  Potential risks involved in prenatal immunization programs include 
developmental adverse effects caused by the inherent biological activity of the vaccine antigen 
and constituents of the vaccine product (e.g., adjuvants, preservatives, and stabilizers). In 
addition, potential adverse effects on the pregnancy status and or the developing embryo/fetus 
may be the result of maternal immune modulation (Refs. 4 and 5). 
Because pregnant women are usually excluded from clinical trials, data from developmental 
toxicity studies in animal models offer one approach to screen for potential developmental 
hazards. Studies in animal models may frequently present the only information available to draw 
conclusions regarding developmental risk to be included in the product labeling required under 
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section 201.57(f)(6) in Title 21 Code of Federal Regulations ( 201.57(f)(6)). As there is 
virtually no scientific literature on animal developmental toxicity testing for vaccine products, 
this guidance will outline general and specific recommendations that should be taken into 
account in the assessment of developmental toxicity for preventive and therapeutic vaccines for 
infectious disease indications. 
III. 	 VACCINE TARGET POPULATION AND TIMING OF PRE-CLINICAL 
DEVELOPMENTAL TOXICITY STUDIES 
For the purpose of this document, vaccines are a heterogeneous class of preventive, in some 
cases therapeutic, medicinal products, the administration of which is intended to elicit an 
immune response(s) that can prevent and/or lessen the severity of one or more infectious 
diseases.  A vaccine may be a live attenuated preparation of bacteria, viruses, or parasites, 
inactivated (killed) whole organisms, living irradiated cells, crude fractions or purified 
immunogens, including those derived from recombinant deoxyribonucleic acid (DNA) in a host 
cell, conjugates formed by covalent linkage of components, synthetic antigens, polynucleotides 
(such as plasmid DNA vaccines), living vectored cells expressing specific heterologous 
immunogens, or cells pulsed with immunogen.  It may also be a combination of vaccines listed 
above. Antigens may be presented plain or delivered in combination with other antigens, 
adjuvants, additives and other excipients. Therapeutic vaccines for non-infectious diseases and 
monoclonal antibodies used as immunogens are not considered in this guidance (Ref. 6).  
Developmental toxicity studies are usually not necessary for vaccines indicated for 
immunization during childhood.  However, for vaccines indicated for adolescents and adults and 
for vaccines that are indicated or may have the potential to be indicated for immunization of 
pregnant women, we recommend that you consider developmental toxicity studies. 
There are currently differences in the timing of these developmental toxicity studies, as follows, 
to support inclusion of either pregnant individuals or females of childbearing potential in clinical 
trials. 
Maternal immunization: For products indicated specifically for immunization of pregnant 
women, we recommend that you have the data from non-clinical developmental toxicity studies 
available prior to the initiation of any clinical trial enrolling pregnant women.  
Females of childbearing potential: For vaccines indicated for females of childbearing potential, 
you may include subjects in clinical trials without non-clinical developmental toxicity studies, 
provided that appropriate precautions are taken by subjects enrolled in these trials to avoid 
vaccination during pregnancy, such as pregnancy testing and use of birth control. For these 
products, we recommend that you include data from developmental toxicity studies with the 
initial BLA submission ( 601.2), regardless of whether such studies have been submitted earlier 
to the IND. 
Males: Currently, males may be included in Phase I, II, and III clinical trials in the absence of 
nonclinical male fertility studies, although such studies may be recommended for certain 
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products in the future. 
IV.  DESIGN OF DEVELOPMENTAL TOXICITY STUDIES 
A.  General Considerations and Recommendations 
The decision whether a developmental toxicity study needs to be performed should be 
made on a case-by-case basis taking into consideration historical use, product features, 
intended target population and intended clinical use. We recommend the developmental 
toxicity study be designed to detect potential developmental adverse effects induced by 
components present in the vaccine formulation.  However, despite efforts to maximize 
the predictive value of developmental toxicity studies, there may always be limitations in 
evaluating or screening for potential risks and thus, limitations in reducing the 
uncertainties of risk. Also, lack of adverse effects on embryo/fetal development in an 
animal study does not necessarily imply absence of risk for humans.  Factors that may 
limit risk prediction include, but are not limited to, species-specific differences in the 
immune response, different developmental time lines, and differences in placentation.  
Nevertheless, developmental toxicity studies in animal models are the best currently 
available non-clinical tools to screen for adverse developmental effects of the product in 
humans.  Information about developmental risk from animal data is frequently the only 
information available at the time of product licensure.  The good laboratory practice 
regulations in 21 CFR Part 58 apply to nonclinical laboratory studies that support or are 
intended to support applications. 
1.  Previous clinical experience 
All available clinical experience in pregnant women should be considered in any 
potential application with respect to the design of developmental toxicity studies 
in animals.  Clinical experience derived from immunization of pregnant women 
may be helpful in the evaluation of the potential for any adverse outcome on the 
viability and development of offspring.  Such information may also aid in the 
design/monitoring of appropriate non-clinical studies, and for product labeling.   
However, clinical data that may have been obtained from small numbers of 
pregnant women enrolled in non-IND studies, e.g., immunized with the vaccine or 
a related product, will generally not replace the need for animal developmental 
toxicity studies. 
2.  Previous non-clinical experience 
We recommend that you review all data generated from prior acute or repeat dose 
non-clinical toxicity studies for their possible contribution to the interpretation of 
any adverse developmental effects that appear in the developmental toxicology 
studies. In addition, data from prior non-clinical studies do frequently form the 
basis for the choice of the animal model and vaccine dose used in the 
developmental toxicity study. 
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3.  Vaccine formulation 
We recommend that you perform the non-clinical developmental toxicity study on 
the same lot as proposed for the clinical trial.  If this is not feasible, we 
recommend that non-clinical lots be comparable to clinical lots with respect to 
physico-chemical data, stability, and formulation and be manufactured in 
accordance with applicable cGMPstandards. 
In addition, even though pivotal clinical studies are frequently conducted with an 
intended final formulation, optimizations of formulations are frequently made 
prior to product marketing.  In these cases, we will assess, on a case-by-case 
basis, the applicability of non-clinical studies conducted with earlier clinical 
formulations of the vaccine to the commercial formulation of the vaccine.  For a 
product specifically intended for maternal immunization, we recommend that you 
perform non-clinical developmental toxicity studies in advance of clinical studies 
that enroll pregnant women.  In these cases, to avoid performing multiple 
developmental toxicology studies during development, you may find it 
advantageous to conduct Phase I and Phase II studies in non-pregnant subjects. 
Results from these studies can be used as the basis for advancing the most 
promising product formulation(s) to studies that enroll pregnant women.  
4.  Vaccine product class 
There are a number of vaccines in clinical development that may be similar to or 
of the same product class as either investigational or already licensed products, 
for which developmental toxicity studies may have been performed.  In these 
cases, we will examine the need for additional development toxicity studies for 
the product under investigation on a case-by case basis. Regarding combination 
vaccines in clinical development, for which the individual components are 
licensed and on which developmental toxicity assessments have been performed, 
we may not recommend further developmental toxicity assessments.  However, if 
the combination vaccine is formulated with new adjuvant, new preservative or if 
significant changes to the individual products or their manufacture were made 
and/or concerns exist that combining the individual licensed products may 
increase their toxic potential, we recommend additional developmental toxicity 
studies. Similarly, if no developmental toxicity studies have been conducted for 
the individual licensed or unlicensed components, we recommend that you 
conduct developmental toxicity studies.  In some instances, documentation on 
clinical and epidemiological data, e.g., exposure to the infectious agent or use of 
related, licensed vaccines during pregnancy, may be sufficient to evaluate the risk 
of the investigational product and may be provided by you and considered by 
FDA in determining the need for developmental toxicity studies in animal 
models.  In these cases, we recommend that you contact FDA to reach agreement 
regarding the need for additional developmental toxicity studies for that particular 
product. 
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5.  Application of ICH guidance document S5A 
The ICH S5A guidance document entitled "Detection of Toxicity to Reproduction 
for Medicinal Products," addresses the design of animal studies primarily for 
detection of toxicity on reproduction, dividing the reproductive cycle into 
different segments, defined as stages A  F (see Ref. 1).  The ICH S5A guidance 
suggests that different studies can be conducted to address the various segments 
of the reproduction cycle. For preventive and therapeutic vaccines for infectious 
diseases, the primary concern is potential untoward effects of the test article on 
development and growth of the embryo and fetus.  Thus, the primary focus is on 
developmental toxicity studies to detect adverse effects on the pregnant/lactating 
female and development of the embryo/fetus and the offspring following exposure 
of the female to the vaccine from implantation through the end of pregnancy, with 
follow-up of the offspring through weaning. These stages are defined as stages C, 
D, and E in the ICH S5A document.  Depending on the product and on a case-by-
case basis, we might require additional studies to address additional segments of 
the reproductive cycle. 
We recommend use of the ICH S5A guidance as a general point of reference to 
assist you in the general design of developmental toxicity studies and evaluation 
of endpoints. However, we want to emphasize distinguishable factors relevant to 
vaccines. The most important feature distinguishing vaccines from other 
pharmaceutical products is the intended vaccine-induced immune response.  Also, 
vaccines are usually administered in limited, episodic dosages with months or 
even years between doses. Vaccines include a broad range of product categories 
such as live attenuated, inactivated, recombinant, polynucleotide, polysaccharide, 
protein antigens, vectored vaccines, and conjugate vaccines. These may be 
adjuvanted or consist of a combination of different vaccine antigens.  Thus, given 
the complexity of these issues, the non-clinical testing strategies outlined in the 
ICH S5A document may not be directly applicable to vaccines, and study designs 
outlined in the ICH S5A document may need to be tailored to the vaccine product 
under consideration. Outlined below are specific considerations that we 
recommend you take into account when designing developmental toxicity studies 
for vaccines. We also recommend that prior to the conduct of the study, you 
establish an early dialogue with CBER to reach agreement on a specific protocol 
including study endpoints. 
B.  Specific Considerations 
1.  Animal model 
We recommend that you provide in your IND submission a justification for the 
choice of the animal model to be used in the developmental toxicity study.  This 
should include a demonstration that the species is able to develop an immune 
response to the vaccine antigen, even though there may be quantitative and 
qualitative differences in immune responses between species.  The laboratory 
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species most often used for developmental toxicity studies, on the basis of 
availability of background data and historical experience, are rats, rabbits and 
mice.  Most human vaccines are immunogenic in rodents or rabbits.  In some 
cases, only non-human primates may show an adequate immune response.  
However, because of the technical and logistic difficulties involved in using non 
human primates for developmental toxicity studies, we recommend you only 
consider these animals if no alternative models are available. 
In addition to demonstrating an immune response in the pregnant female, we 
recommend that you verify the exposure of the fetus to maternal antibodies.  
Thus, since there are differences between primates, non-rodents, and rodent 
animal species in terms of timing of maternal antibody transfer to the offspring, 
we recommend that you evaluate the pre- and postnatal exposure of the offspring 
to maternal antibody as a criterion for selecting the most appropriate experimental 
model.  In addition, the species selected should be amenable to fetal and postnatal 
examinations.  
In cases where lack of an appropriate animal model hinders the assessment of an 
immune response, developmental toxicity studies may still provide important 
information regarding potential embryo/fetal toxic effects of the vaccine 
components/formulation and safety of the product in the pregnant animal.   
In most cases, it is sufficient to conduct developmental toxicity studies using only 
one species; thus, there is no requirement for the routine use of two species, i.e., 
one rodent and one non-rodent. 
The number of animals per group should be sufficient to allow meaningful 
interpretation of the data. For example, for a developmental toxicity study using 
rats or rabbits, we recommend that you assign an adequate number of animals to 
each group to allow an evaluation of at least 40 animals per group.  These animals 
can be further allocated to the Caesarean and littering subgroup using 20 animals 
each. 
2.  Pharmacodynamics 
We recommend that you obtain information about the onset and duration of the 
antibody response in pilot studies because these data may help in selecting the 
proper species, study design, and dosing schedules. Initial information can be 
derived from non-pregnant animals.  However, it may also be necessary to 
perform these pilot studies in a small group of pregnant animals to evaluate 
antibody formation in relation to test article exposure and placental antibody 
transfer to the fetus if there is evidence that antibody formation may differ in 
pregnant versus non-pregnant animals.   
We recognize that antibody induction presents only one aspect of the overall 
immune response induced by the vaccine and that other immunological 
parameters, such as cytokines and induction of cytotoxic T cells, may be as 
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important.  However, given the relative lack of validated assays to assess the 
induction of these other parameters, antibody assessments are currently used as a 
marker for vaccine induced effects in these studies.  This does not exclude the 
evaluation of additional immunologic parameters on a case-by-case basis.  For 
example, if data are available which indicate that a vaccine antigen induces a 
particular cytokine response, respective cytokine measurements may be included, 
especially if the cytokine is one that may affect pregnancy.   
3.  Experimental procedure 
In order to detect adverse effects on the pregnant/lactating female animal and 
development and growth of the embryo/fetus and the offspring, we recommend 
that the female be exposed to the vaccine during the interval from implantation 
through closure of the hard palate and also at later stages of pregnancy. The 
offspring should be followed to weaning and observed for normal growth and 
development.  We recommend that you submit one subgroup of pregnant females 
to Caesarean examination at the end of pregnancy for routine uterine and fetal 
examinations, and allow another subgroup to litter and rear their offspring to 
weaning in order to monitor the post-natal development of the offspring up to 
weaning. 
4.  Dosage 
We recommend that you assess a single dose level that is capable of inducing an 
immune response in the animal model.  Where possible, we recommend that you 
administer animals the maximum human dose (e.g., 1 human dose = 1 rabbit 
dose) regardless of body weight. If it is not feasible to administer the maximum 
human dose (e.g., limitation in total volume that can be administered; dosing 
induced local toxicity affecting pregnancy), we recommend that you administer a 
dose that exceeds the human dose on a mg/kg bases while still capable of 
inducing an immune response in the animal.  
5.  Frequency and route of administration 
We recommend that the vaccination regimen optimize maternal antibody titers 
throughout the embryonic, fetal, and early post-natal periods.  Timing and number 
of doses will depend on the onset and duration of the immune response of the 
particular product. Because of concerns that a daily dosing regimen may lead to 
overexposure to the vaccine antigen that could potentially result in immune 
tolerance, we recommend episodic dosing of pregnant animals rather than daily 
dosing. In addition, episodic dosing appears to be more relevant, as it better 
mimics the clinically proposed immunization schedule for most preventive and 
therapeutic vaccines for infectious disease indications. Considering the short 
gestational period of animal species most frequently used, it may be necessary to 
administer priming doses to the animals several days or weeks prior to mating in 
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order to elicit a peak antibody production during the critical phases of pregnancy, 
i.e., the period of organogenesis. 
When dosing prior to implantation, stress reactions may be observed in the animal 
that may affect the pregnancy status.  Therefore, with treatment of animals prior 
to mating/insemination, it may be necessary to add more animals to the study to 
ensure that sufficient animals become pregnant for evaluation.   
We recommend that you administer one or several additional doses during 
organogenesis (i.e., implantation to closure of the hard palate) to evaluate 
potential direct embryotoxic effects of the components of the vaccine formulation 
and to maintain a high level of antibody throughout the remainder of gestation.  In 
certain cases, subgroups of animals that are dosed at certain time points may be 
included to evaluate if the vaccine acts as a selective toxicant, bearing in mind 
that it may be difficult to adjust vaccine administration with gestational timelines.  
We recommend that the route of administration mimic the clinical intended route 
of administration. 
6.  Control groups 
We recommend that you dose control animals with placebo at the same time and 
frequency as test group animals.  Since the potential toxicity of each of the 
components presented in the vaccine formulation will need to be evaluated, we 
recommend that you consider additional groups if components other than the 
vaccine antigen contained in the vaccine formulation (e.g. excipients, 
preservatives) cause effects or affect the activity of the test substance. In 
addition, if the vaccine is formulated with adjuvant, we recommend that you 
consider the inclusion of an adjuvant-only control arm, particularly if the adjuvant 
is novel. 
7.  Endpoints 
In general, the study endpoints should include those recommended for studies for 
effects on prenatal and postnatal development including maternal functions as 
stated in the ICH S5A document (see Ref. 1).  When deciding on the endpoints to 
be evaluated, we recommend that you take into consideration the nature of the 
vaccine and particular concerns associated with that product. The following 
parameters listed are intended to provide a basic panel of endpoints to be 
evaluated that are not meant to be all-inclusive. 
a.  Premating/preinsemination period   
Clinical observations including data on general appearance and body weights 
should be obtained weekly and on days of test article administration. 
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b.  Gestational period 
We recommend that you observe maternal animals during the study for signs of 
morbidity and mortality, and record clinical observations regarding general 
appearance and behavior. We recommend that the evaluations include body 
weight and body weight change, potential signs of local toxicity, food 
consumption, duration of pregnancy, abortions, premature deliveries, and 
parturition (for maternal animals not subjected to Caesarian sectioning). 
c.  Caesarean sectioning group 
i.  Maternal Observations 
At terminal examination of groups subjected to Caesarean sectioning, we 
recommend that you conduct a necropsy (macroscopic examination) and 
preserve maternal tissue with macroscopic findings for histological 
evaluations as deemed necessary by the gross findings.  For example, 
histological evaluations may be indicated if you observe effects on organ 
to body weight ratios. We recommend that you record the number and 
distribution of corpora lutea, implantation sites, viable and nonviable 
fetuses, and early and late resorptions and that you perform a gross 
evaluation of the placenta. 
ii.  Fetal examinations 
We recommend that you obtain individual body weights of live fetuses 
and examine each viable fetus for gross external, visceral, and skeletal 
alterations. Late resorptions and dead fetuses should also be examined for 
gross external alterations to the extent possible. All fetuses should be 
examined internally to determine sex. 
d.  Natural delivery group 
i.  Maternal observations 
In addition to the parameters outlined in section IV.B.7.b (Gestational 
period), we recommend that you determine the duration of gestation and 
parameters such as the fertility index, gestation index, and live birth index.  
Animals that deliver a litter should be sacrificed at the end of the pre 
weaning period. We recommend that you perform a gross necropsy of the 
thoracic, abdominal and pelvic viscera, and record the number and 
distribution of implantation sites as well as any observed abnormalities.  
Animals that die or are sacrificed because of moribund condition, abortion 
or premature delivery should be examined for the cause of death and 
pregnancy status recorded. We recommend that you also examine aborted 
fetuses and/or delivered pups to the extent possible. 
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ii.  F1 generation 
We recommend postnatal follow-up from birth to weaning to assess 
normal growth, body weight gain, and nursing activity as indicators for 
normal development.  We also recommend that you include into the study 
design tests to screen for normal neuro-development, for example, 
auditory and visual function tests. Viability and lactation indices should 
be determined and individual sexes should be recorded.  At terminal 
sacrifice, we recommend that you perform a necropsy, record any 
abnormalities and retain gross lesions for possible histological 
examinations.  We recommend that you evaluate pups that die before 
examination for vital status at birth and examine pups found dead for 
gross lesions and cause of death. 
8.  Immunological endpoints 
In addition to evaluating potential developmental adverse effects and adverse 
effects on the pregnant animal, we recommend that you include an assessment of 
the vaccine induced antibody response to verify exposure of the embryo/fetus to 
maternal antibody.  Serum specimen from maternal animals prior to and at 
additional time points following dosing should be collected to assess the 
development of antibodies.  Sampling is usually conducted prior to test article 
administration and at day of Caesarean sectioning (where applicable) and at the 
end of the weaning period. 
In addition, we recommend that you obtain cord blood samples to assess placental 
transfer of maternal antibodies from animals in the Caesarean subgroup.  We 
recommend that you also assess antibody levels from a representative number of 
pups/litters at the end of the weaning period to verify exposure of the neonates to 
maternal antibody induced.  Antibody evaluations in developmental toxicity 
studies serve the purpose of verifying an effect of the vaccine in the test species 
as opposed to evaluating potential immunotoxic effects.  You may evaluate 
additional immune parameters on a case-by-case basis.  For example, if evidence 
exists that the vaccine antigen or other vaccine components can trigger the release 
of a particular cytokine potentially affecting pregnancy, you may include 
respective assessments.   
9.  Additional assessments 
In cases where non-clinical developmental toxicity studies reveal vaccine-induced 
adverse effects on either the pregnant/lactating animal, embryo/fetal development, 
or development of the offspring, we recommend you conduct further animal 
studies to evaluate the cause of the effect. Such studies may include broader 
immunological evaluations, e.g., histochemical analysis for antibody depositions, 
as well as neurological assessments.   
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V.  R	 EFERENCES 
1. 	 International Conference on Harmonization (ICH) Harmonized Tripartite Guideline for 
Industry (ICH-S5A) Detection of Toxicity to Reproduction for Medicinal Products, (59 
FR 48746, September 22, 1994), http://www.fda.gov/cder/guidance/s5a.pdf. 
2. 	 Recommended adult immunization schedule-United States, 2002-2003, MMWR October 
11, 2002, Vol. 51 (40); 904-908. 
3. 	 Colley, Gilbert B., Brantley, M.D., Larson, M.K., Family Planning Practices and 
Pregnancy Intention, 1997. Atlanta, GA: Division of Reproductive Health, National 
Center for Chronic Disease Prevention and Health Promotion, Centers for Disease 
Control and Prevention, 2000, 
http://www.cdc.gov/reproductivehealth/prams/pdf/97/PRAMS_sr_97.pdf. 
4.  B	 arrow, P.C., Reproductive toxicology studies and immunotherapeutics. Toxicology 185 
(2003), 205-212. 
5. 	 Thellin, O., Heinen, E., Pregnancy and the immune system: between tolerance and 
rejection. Toxicology 185 (2003), 179-184. 
6. 	 Guidance for Industry: Content and Format of Chemistry, Manufacturing and Controls 
Information and Establishment Description Information for a Vaccine or Related Product 
(January 1999), http://www.fda.gov/cber/gdlns/cmcvacc.pdf. 
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