Clinical Pharmacology 
Data to Support a 
Demonstration of 
Biosimilarity to a 
Reference Product 
 
Guidance for Industry 
 
 
 
 
 
 
 
 
 
 
 
 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Drug Evaluation and Research (CDER) 
Center for Biologics Evaluation and Research (CBER) 
 
December 2016 
Biosimilars 
 
C:\Users\CAROLANM\AppData\Local\Microsoft\Windows\Temporary Internet Files\Content.Outlook\NRXMQ6DI\FRDTS 2015-739_FINAL 
clin pharm biosimilar guidance _111616_for RPMS.docx   
Clinical Pharmacology Data 
to Support a Demonstration 
of Biosimilarity to a 
Reference Product 
 
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  
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http://www.fda.gov/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/default.htm 
 
and/or 
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Food and Drug Administration 
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Phone: 800-835-4709 or 240-402-8010; Email: ocod@fda.hhs.gov 
http://www.fda.gov/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInformation/Guidances/default.htm 
 
 
 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Drug Evaluation and Research (CDER) 
Center for Biologics Evaluation and Research (CBER) 
 
December 2016 
Biosimilars 
   
TABLE OF CONTENTS 
I.  INTRODUCTION............................................................................................................. 1 
II.  THE ROLE OF CLINICAL PHARMACOLOGY STUDIES IN THE 
DEMONSTRATION OF BIOSIMILARITY ............................................................................. 2 
III.  CRITICAL CONSIDERATIONS IN THE USE OF CLINICAL 
PHARMACOLOGY STUDIES TO SUPPORT BIOSIMILARITY ....................................... 3 
A.  Exposure and Response Assessment to Support a Demonstration of Biosimilarity ................ 3 
B.  Evaluation of Residual Uncertainty ............................................................................................. 4 
C.  Analytical Quality and Similarity ................................................................................................. 4 
D.  Integrity of the Bioanalytical Methods Used in PK and PD Studies ......................................... 6 
E.  Safety and Immunogenicity .......................................................................................................... 8 
IV.  DEVELOPING CLINICAL PHARMACOLOGY DATA FOR SUPPORTING A 
DEMONSTRATION OF BIOSIMILARITY ............................................................................. 9 
A.  Study Design ................................................................................................................................... 9 
B.  Reference Product ........................................................................................................................ 10 
C.  Study Population .......................................................................................................................... 10 
D.  Dose Selection ............................................................................................................................... 11 
E.  Route of Administration .............................................................................................................. 11 
F.  Pharmacokinetic Measures ......................................................................................................... 12 
G.  Pharmacodynamic Measures ...................................................................................................... 12 
H.  Defining the Appropriate Pharmacodynamic Time Profile ..................................................... 13 
I.  Statistical Comparison of PK and PD Results ........................................................................... 13 
V.  UTILITY OF SIMULATION TOOLS IN STUDY DESIGN AND DATA 
ANALYSIS .................................................................................................................................. 14 
VI.  CONCLUSION ............................................................................................................... 14 
DEFINITIONS ............................................................................................................................ 15 
  Contains Nonbinding Recommendations 
Clinical Pharmacology Data to Support a Demonstration of 
Biosimilarity to a Reference Product 
 
1
Guidance for Industry  
 
 
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 
 
This guidance is intended to assist sponsors with the design and use of clinical pharmacology 
studies to support a decision that a proposed therapeutic biological product is biosimilar2 to its 
reference product.  This guidance pertains to those productssuch as therapeutic biological 
productsfor which pharmacokinetic (PK) and pharmacodynamic (PD) data are needed to 
support a demonstration of biosimilarity.  Specifically, the guidance discusses some of the 
overarching concepts related to clinical pharmacology testing for biosimilar products, 
approaches for developing the appropriate clinical pharmacology database, and the utility of 
modeling and simulation for designing clinical trials.   
 
This guidance is one in a series that FDA is developing to implement the Biologics Price 
Competition and Innovation Act of 2009 (BPCI Act).3  This guidance is intended to assist 
sponsors in designing clinical pharmacology studies that can support an application submitted 
under section 351(k) of the Public Health Service Act (PHS Act) (42 U.S.C. 262(k)) (a 351(k) 
application) as part of a stepwise approach to support a demonstration of biosimilarity.4  
 
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 
                                                 
 
1 This guidance has been prepared by the Center for Drug Evaluation and Research (CDER) and the Center for 
Biologics Evaluation and Research (CBER) at the Food and Drug Administration.  
2 Terms that appear in bold type are defined in the Definitions section at the end of this guidance. 
3 Sections 7001 through 7003 of the Patient Protection and Affordable Care Act (Affordable Care Act), Public Law 
111-148. 
4 See FDAs guidance for industry Scientific Considerations in Demonstrating Biosimilarity to a Reference Product. 
We update guidances periodically.  To make sure you have the most recent version of a guidance, check the FDA 
Drugs guidance Web page at 
http://www.fda.gov/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/default.htm.    
1  Contains Nonbinding Recommendations 
the word should in Agency guidances means that something is suggested or recommended, but 
not required.  
 
 
II.  THE ROLE OF CLINICAL PHARMACOLOGY STUDIES IN THE 
DEMONSTRATION OF BIOSIMILARITY 
 
The BPCI Act, which was enacted as part of the Affordable Care Act, established an abbreviated 
pathway for FDA licensure of biological products that are demonstrated to be biosimilar to or 
interchangeable with an FDA-licensed reference product. This pathway is described in section 
351(k) of the PHS Act.  
 
Under section 351(k)(2) of the PHS Act, a 351(k) application must contain, among other things, 
information demonstrating that the biological product is biosimilar to a reference product based 
on data derived from analytical studies, animal studies, and a clinical study or clinical studies, 
including an assessment of immunogenicity, PK, and PD,5 unless FDA determines, in its 
discretion, that certain studies are unnecessary in a 351(k) application.6   
 
Biosimilarity is defined at section 351(i)(2) of the PHS Act to mean that the biological product is 
highly similar to the reference product notwithstanding minor differences in clinically inactive 
components and that there are no clinically meaningful differences between the biological product 
and the reference product in terms of the safety, purity, and potency of the product.  
 
Comparative analytical data provide the foundation for a development program for a proposed 
biosimilar product intended for submission under section 351(k) of the PHS Act.  Clinical 
pharmacology studies build on the comparative analytical studies in the stepwise approach to 
support a demonstration of biosimilarity, and are normally a critical part of demonstrating 
biosimilarity by supporting a demonstration that there are no clinically meaningful differences 
between the proposed biosimilar product and the reference product.  These studies provide the 
data that describe the degree of PK similarity  between the proposed biosimilar product and the 
reference product.  In addition, clinical pharmacology studies often include PD endpoints (both 
therapeutic and toxic) and pharmacometric analysis to assess whether there are clinically 
meaningful differences between the proposed biosimilar product and the reference product.  
These clinical pharmacology studies may address residual uncertainties that remain after the 
analytical evaluation, can add to the totality of the evidence supporting a demonstration of 
biosimilarity, and can guide both the need for and design of subsequent clinical testing to support 
a demonstration of no clinically meaningful differences in the overall demonstration of 
biosimilarity.  Clinical pharmacology data can be an important component of the scientific 
justification supporting extrapolation of data to one or more additional conditions of use.7 
 
                                                 
5 Section 351(k)(2)(A)(i)(I) of the PHS Act. 
6 Section 351(k)(2)(A)(iii) of the PHS Act. 
7 See FDAs guidance for industry Biosimilars: Questions and Answers Regarding Implementation of the Biologics 
Price Competition and Innovation Act of 2009 for more information on this topic.   
2  Contains Nonbinding Recommendations 
The types of clinical pharmacology studies to be conducted will depend on the residual 
uncertainties about biosimilarity that these studies can address to add to the totality of evidence 
for biosimilar product development. 
 
 
III.  CRITICAL CONSIDERATIONS IN THE USE OF CLINICAL 
PHARMACOLOGY STUDIES TO SUPPORT BIOSIMILARITY 
 
Three key conceptsnamely a PK and PD response assessment, an evaluation of residual 
uncertainty, and assumptions about analytical quality and similarityare especially relevant to 
the stepwise development of proposed biosimilar products and are discussed in more detail in 
this section.  Bioanalytical methodology and the use of clinical pharmacology studies to gain 
safety and immunogenicity information are also examined. 
 
A.  Exposure and Response Assessment to Support a Demonstration of 
Biosimilarity 
 
The objective of a well-designed clinical PK and PD study in a biosimilar development program 
is to evaluate the similarities and differences in the PK and PD profiles between the proposed 
biosimilar product and the reference product.  A well-designed clinical PK and PD study should 
include information about the exposure and, when possible, the exposure-response to the 
biological products, which are important for assessing whether there are any potential clinically 
meaningful differences between two products.  Determining the exposure-response to a 
biological product can be particularly challenging because of the complex nature and 
heterogeneity of biological products. An evaluation of clinical pharmacology similarity should 
include assessments of PK similarity, and if applicable, PD similarity. 
 
The PD biomarker(s) used to measure PD response should be a single biomarker or a composite 
of biomarkers that effectively demonstrate the characteristics of the products target effects.  Use 
of a single scientifically appropriate PD biomarker or a composite of more than one relevant PD 
biomarker can reduce residual uncertainty regarding the existence of any clinically meaningful 
differences between products and can significantly add to the overall demonstration of 
biosimilarity.  Using broader panels of PD biomarkers (e.g., by conducting a protein or mRNA 
microarray analysis) that capture multiple pharmacological effects of the product can be of 
additional value.  When determining which biomarkers should be used to measure response, it is 
important to consider the following five characteristics: 
 
  The time of onset of change in the PD biomarker relative to dosing and its return to 
baseline with discontinuation of dosing 
 
  The dynamic range of the PD biomarker over the exposure range to the biological 
product 
 
  The sensitivity of the PD biomarker to differences between the proposed biosimilar 
product and the reference product 
 
3  Contains Nonbinding Recommendations 
  The relevance of the PD biomarker to the mechanism of action of the drug (to the 
extent that the mechanism of action is known for the reference product) 
 
  The analytical validity of the PD biomarker assay  
 
If these characteristics are addressed, through the submission of PK and PD results, the extent of 
the clinical development program can be refined in both the design and extent of additional 
clinical trials necessary to assess whether there are clinically meaningful differences between the 
proposed biosimilar product and the reference product.  In some instances, PD biomarkers with 
the relevant characteristics listed above are not identified, but the sponsor is still encouraged to 
incorporate PD biomarkers that achieve a large dynamic range over the concentration range in 
the PK evaluation because these PD biomarkers represent potential orthogonal tests that can 
support similarity.  When PD biomarkers are not sensitive or specific enough to detect clinically 
meaningful differences, the derived PK parameters should be used as the primary basis for 
evaluating similarity from a clinical pharmacology perspective, and the PD biomarkers can be 
used to augment the PK data.  A combination of PK and PD similarity can be an important 
assessment in demonstrating that there are no clinically meaningful differences between the 
proposed biosimilar product and the reference product. 
 
B.  Evaluation of Residual Uncertainty 
 
In evaluating a sponsors data to support a demonstration of biosimilarity, FDA will consider the 
totality of the data and information submitted using a risk-based approach, including data from 
the structural and functional characterizations, nonclinical evaluations, clinical PK and PD 
studies, clinical immunogenicity testing and an investigation of clinical safety, and, when 
appropriate, clinical effectiveness.  These data should be collected in a stepwise manner.  
Especially pertinent to FDAs clinical pharmacology evaluation are the clinical PK and PD data 
and immunogenicity and other safety data obtained in conjunction with the clinical 
pharmacology studies.  The need for additional studies at each step in this progressive approach 
will be determined by the degree of residual uncertainty that remains at each step regarding the 
similarity of the products and whether the study can address these uncertainties. 
 
C.  Analytical Quality and Similarity 
 
In a stepwise assessment of biosimilarity, extensive and robust comparative structural and 
functional studies (e.g., bioassays, binding assays, and studies of enzyme kinetics) should be 
performed to evaluate whether the proposed biosimilar product and the reference product are 
highly similar.  A meaningful assessment of biosimilarity depends on, among other things, the 
capabilities of available state-of-the-art analytical assays to evaluate, for example, the molecular 
weight, the higher-order structure and post-translational modifications, heterogeneity, functional 
properties, impurity profiles, and degradation profiles denoting stability of the protein.  The 
sponsor should describe the capabilities and limitations of the methods used in the analytical 
assessment.  
 
An extensive analytical characterization can reveal differences between the proposed biosimilar 
product and the reference product.  The type, nature, and extent of any differences between the 
4  Contains Nonbinding Recommendations 
two products should be clearly identified, and the potential effect of these differences should be 
addressed and supported by appropriate data.  In some cases, additional studies can demonstrate 
that the identified difference is within an appropriate range to consider the proposed biosimilar 
product to be highly similar to the reference product.8  However, certain differences in the results 
of the analytical characterization can preclude a determination by FDA that the proposed 
biosimilar product is highly similar to the reference product and, therefore, the further 
development of the proposed biosimilar product through the 351(k) regulatory pathway is not 
recommended. 
 
It may be useful to compare the quality attributes of the proposed biosimilar product with those 
of the reference product using a meaningful fingerprint-like analysis algorithm that covers a 
large number of product attributes and their combinations with high sensitivity using orthogonal 
methods.  Comparison of quality attributes in this manner can further quantify the overall 
similarity between two products and might provide a basis for a more selective and targeted 
approach to subsequent animal and/or clinical studies. 
 
The result of the comparative analytical characterization during product development can lead to 
one of the following four assessments within a development-phase continuum, with the 
understanding that FDA does not make the ultimate determination that the proposed biosimilar 
product is highly similar to the U.S.-licensed reference product until the time of licensure. 
 
  Insufficient analytical similarity:  Certain differences in the results of the analytical 
characterization are sufficiently significant such that further development through the 
351(k) regulatory pathway is not recommended unless, for example, modifications 
are made to the manufacturing process for the proposed biosimilar product that are 
likely to lead to the minimization or elimination of such differences. 
 
  Analytical similarity with residual uncertainty:   Further information, additional 
analytical data, or other studies are needed to determine if observed analytical 
differences are likely to fall within an appropriate range when the 351(k) application 
for the proposed biosimilar product is submitted.  As an example, glycosylation plays 
an important role in the PK of certain protein products.  Manufacturing process 
conditions can affect glycosylation and in some cases PK.  Comparative PK and PD 
studies of the proposed biosimilar product and the reference product, in addition to 
supporting a demonstration of no clinically meaningful differences, may address 
residual uncertainties regarding certain glycosylation differences and the impact on 
PK.  Thus PK and PD studies could support that some differences in glycosylation 
identified in the analytical studies might fall within an appropriate range.  
 
  Tentative analytical similarity:  At this stage of development, the results of the 
comparative analytical characterization permit high confidence in the analytical 
similarity of the proposed biosimilar product and the reference product, and it can be 
                                                 
8 See FDAs guidance for industry Quality Considerations in Demonstrating Biosimilarity of a Therapeutic Protein 
Product to a Reference Product.   
5  Contains Nonbinding Recommendations 
appropriate for the sponsor to conduct targeted and selective animal and/or clinical 
studies to resolve residual uncertainty and support a demonstration of biosimilarity. 
 
  Fingerprint-like analytical similarity:  The results of integrated, multi-parameter 
approaches that are extremely sensitive in identifying analytical differences (i.e., 
fingerprint-like analyses) permit a very high level of confidence in the analytical 
similarity of the proposed biosimilar product and the reference product, and it would 
be appropriate for the sponsor to use a more targeted and selective approach to 
conducting animal and/or clinical studies to resolve residual uncertainty and to 
support a demonstration of biosimilarity.  
 
The outcome of the comparative analytical characterization should inform the next steps in the 
demonstration of biosimilarity.   
 
D.  Integrity of the Bioanalytical Methods Used in PK and PD Studies  
 
When performing an evaluation of clinical pharmacology similarity, it is critical to use the 
appropriate bioanalytical methods to evaluate the PK and PD properties of a proposed biosimilar 
product and the reference product.  Because of the generally complex molecular structure of 
biological products, conventional analytical methods might not be suitable for biological 
products.  The bioanalytical methods used for PK and PD evaluations should be accurate, 
precise, specific, sensitive, and reproducible.  The scientific requirements of bioanalytical 
methods are described in a separate FDA guidance document.9   
 
1.  General PK Assay Considerations 
 
A sponsor should design or choose an assay based on a thorough understanding of the 
mechanism of action (to the extent that the mechanism of action is known for the reference 
product) and/or structural elements of the proposed biosimilar product and reference product that 
are critical for activity.  An assay producing concentration data that correlate to the 
pharmacological/PD activity is preferred.  The same assay should be used for measuring 
concentrations of the proposed biosimilar product and the reference product and validated for use 
with both products.  Analytical assays should have design and performance parameters that are 
consistent with current industry best practices. 
 
2.  General PD Assay Considerations 
 
Sponsors should make every effort to employ the most suitable assays and methodologies to 
obtain data that are meaningful and reflective of PK, the biological activity, and/or the PD effect 
of the proposed biosimilar product and the reference product.  Furthermore, in submissions to the 
FDA, the sponsor should provide a rationale for the choice of assay and the relevance of the 
assay to drug activity.   
                                                 
9 See FDAs revised draft guidance for industry, Bioanalytical Method Validation, for more information on this 
topic.  When final, this guidance will represent FDAs current thinking on this issue.  
 
6  Contains Nonbinding Recommendations 
 
3.  Specific Assays 
 
Three types of assays are of particular importance for biosimilar product development: ligand 
binding assays, concentration and activity assays, and PD assays.   
 
  Ligand binding assays 
 
Currently, the concentration of most biological products in circulation is measured 
using ligand binding assays.  These assays are analytical methods for quantifying 
high affinity and selective macromolecular interactions between assay reagents (e.g. 
antibodies, receptors or ligands) and the biological product. The ligand binding assay 
reagents chosen for detecting the biological product should be carefully evaluated 
with the goal of producing product concentration data that are meaningful to, and 
reflective of, the pharmacological activity and/or PD effect of the biological product 
of interest. Assays that rely upon antibody reagents and epitopes involved in 
pharmacological/biochemical interactions with targets are most likely to produce 
concentration data that are meaningful for target binding activity. 
 
Some biological products exert pharmacological effects only after multiple molecular 
interactions.  For example, in some cases, the in vivo mechanism of action of 
monoclonal antibodies, bispecific antibodies, or fusion proteins involves binding 
mediated by different regions of the protein product (e.g. binding to both a ligand or 
receptor through a target antigen binding epitope of the protein and to Fc gamma 
receptors with the fragment crystallizable (Fc) region of the protein.  A sponsor 
should choose the most appropriate interactions to measure.  Generally, assays for 
monoclonal antibody product concentrations rely on molecular interactions involving 
the antigen binding (Fab) region, in particular epitopes in the complementarity 
determining regions (CDRs).   
 
  Concentration and activity assays 
 
Bioanalytical methods that are not based on ligand binding can be used for 
quantification of the proposed biosimilar product and the reference product 
concentrations.  For some biological products, such as those that are used to achieve 
enzyme replacement, the drug availability measurements may rely on activity and 
should be captured through an appropriate activity assay.  Depending on the 
complexity of the structural features, some biological products should have more than 
one assay to characterize the systemic exposure of the proposed biosimilar product 
and the reference product.  If more than one assay is used, mass spectrometry and 
other assays can be useful for distinguishing the structures of product variants, if 
relevant.  
 
  PD assays 
 
7  Contains Nonbinding Recommendations 
Relevant PD biomarkers might not always be available to support a proposed 
biosimilar products development through clinical pharmacology studies.  However, 
when PD assessment is a component of the biosimilarity evaluation, sponsors should 
submit (1) a rationale for the selection of the PD endpoints and/or biomarkers and (2) 
data to demonstrate the quality of the assay. PD assays should be sensitive for a 
product or product class and designed to quantitatively evaluate the pharmacological 
effects of the biologic product.  The use of multiple complementary PD assays that 
reflect different aspects of pharmacologic activity of the product might be particularly 
useful to reduce residual uncertainty regarding clinically meaningful differences 
between the products.  Because the PD assay is highly dependent on the 
pharmacological activity of the product, the approach for assay validation and the 
characteristics of the assay performance might differ depending on the specific PD 
assay.  However, the general guiding principles for choosing PK assays (i.e., 
demonstration of specificity, reliability, and robustness) also apply to PD assays. 
 
E.  Safety and Immunogenicity 
 
When immunogenicity results in, for example, either loss of PD effect or efficacy (e.g., 
neutralizing antibodies) or immune-mediated toxicity, the incidence and severity of the response 
should be assessed.10  Safety and immunogenicity data from the clinical pharmacology studies 
should be collected and evaluated.  FDA recognizes that safety and immunogenicity data derived 
from these studies may need to be supplemented by additional evaluations.  The overall 
immunogenicity assessment should include relevant patient populations that are not 
immunocompromised and thus are able to mount an immune response. However, as part of their 
role in the overall assessment of biosimilarity, clinical pharmacology studies can sometimes 
suggest that there are clinically meaningful differences between the products that can inform the 
design and the details of additional investigations and/or clinical studies conducted to investigate 
these potential differences.  The extent of such potential differences will determine whether or 
not further development of the proposed biosimilar product should continue, and if so, what 
studies should be conducted.  
 
Publicly available information on the safety and immunogenicity profile of the reference product 
should be considered when incorporating safety and immunogenicity measurements in the 
clinical pharmacology studies.11  For example, when the reference product is known to have the 
potential for immune-mediated toxicity, assays capable of detecting binding antibodies (and their 
neutralizing potential) should be developed in advance to analyze samples obtained from PK and 
PD studies, so that immunogenicity can be evaluated in real time.  Generally, samples can be 
stored for future analysis if such assays are not yet developed.12  In all cases, sponsors should 
carefully consider assay confounders such as the systemic presence of the proposed biosimilar 
                                                 
10 See FDAs guidance for industry Immunogenicity Assessment for Therapeutic Protein Products. 
11 See footnote 4 for more information on this topic.    
12 FDA has issued the draft guidance for industry Assay Development and Validation for Immunogenicity Testing of 
Therapeutic Protein Products.   
8  Contains Nonbinding Recommendations 
product or the reference product.  Recommendations for immunogenicity assay development are 
described in a separate FDA guidance document.13 
 
When evaluating data (e.g., safety or immunogenicity) collected during the PK and PD studies, 
sponsors should have an understanding of the time course of the appearance and resolution of 
safety signals or immune responses.  The PK profile of the proposed biosimilar product and/or 
the publicly available PK data for the reference product can be used to inform the duration of 
follow-up for safety signals or immunogenicity.   
 
 
IV.  DEVELOPING CLINICAL PHARMACOLOGY DATA FOR SUPPORTING A 
DEMONSTRATION OF BIOSIMILARITY  
 
Sponsors are encouraged to discuss their clinical pharmacology development plan with FDA in 
the early stages of the biosimilar product development program. Critical topics that should be 
discussed with FDA are set forth below. 
  
A.  Study Design 
 
To evaluate clinical PK and PD similarity for the development of proposed biosimilar 
products, two study designs are of particular relevance: crossover designs and parallel study 
designs.  All clinical pharmacology studies of the proposed biosimilar product should be 
performed using materials from the final manufacturing process expected to be used for the 
marketed product if approval is granted.  The relevance of data submitted from studies using 
materials from different manufacturing processes may need to be adequately justified, for 
example, by establishing an analytical and PK bridge to the to-be-marketed product. 
 
  Crossover design 
 
  For PK similarity assessments, a single-dose, randomized, crossover study is 
generally preferred.  A crossover study is recommended for a product with a short 
half-life (e.g., shorter than 5 days), a rapid PD response (e.g., the time of onset, 
maximal effect, and disappearance in conjunction with drug exposure), and a low 
anticipated incidence of immunogenicity.  This design is considered the most 
sensitive to assess PK similarity, and can provide reliable estimates of differences in 
exposure with a minimum number of subjects.  For PD similarity assessments, a 
multiple-dose design may be appropriate when the PD effect is delayed or otherwise 
not parallel to the single-dose drug PK profile.  The time course of appearance and 
disappearance of immunogenicity and its relation to the washout period should be 
considered for studies using a crossover design.   
 
  Parallel design 
 
                                                 
13 See footnote 10. 
9  Contains Nonbinding Recommendations 
  Many biological products have a long half-life and elicit immunogenic responses.  A 
parallel group design is appropriate for products that have a long half-life or for 
products where repeated exposures can lead to an increased immune response that can 
affect the PK and/or PD similarity assessments.  This design is also appropriate for 
diseases that exhibit time-related changes associated with exposure to the drug.  
 
B.  Reference Product 
 
Analytical studies, clinical PK and, if appropriate, PD studies that are intended to support a 
demonstration of biosimilarity should include an adequate comparison of the proposed biosimilar 
product directly with the U.S.-licensed reference product.  However, a sponsor could use a non-
U.S.-licensed comparator product in certain studies to support a demonstration that the proposed 
biological product is biosimilar to the U.S.-licensed reference product.  If a sponsor seeks to use 
data from a clinical study comparing its proposed biosimilar product to a non-U.S.-licensed 
comparator product to address, in part, the requirements under section 351(k)(2)(A) of the PHS 
Act, the sponsor should provide adequate data or information to scientifically justify the 
scientific relevance of these comparative data to an assessment of biosimilarity and establish an 
acceptable bridge to the U.S.-licensed reference product. 14  
 
C.  Study Population 
 
Healthy Subject vs. Patient:  The study population selected should be the most informative for 
detecting and evaluating differences in PK and PD profiles between the proposed biosimilar 
product and the reference product.  Clinical PK and PD studies should be conducted in healthy 
subjects if the product can be safely administered to them.  A study in healthy subjects is 
considered to be more sensitive in evaluating the product similarity because it is likely to 
produce less PK and/or PD variability compared with a study in patients with potential 
confounding factors such as underlying and/or concomitant disease and concomitant 
medications.  If safety or ethical considerations preclude the participation of healthy subjects in 
human PK and PD studies for certain products (e.g., immunogenicity or known toxicity from 
the reference product), or if PD biomarkers can only be relevant in patients with the relevant 
condition or disease, the clinical pharmacology studies should be conducted in such patients.  A 
population that is representative of the patient population to which the drug is targeted will be 
appropriate unless a study in a different population would be more sensitive to detect potential 
differences between the proposed biosimilar product and the reference product. 
 
Demographic Group:  Clinical pharmacology studies should be conducted in the subject or 
patient demographic group most likely to provide a sensitive measure of differences between the 
proposed biosimilar product and the reference product.  The sponsor should justify why the 
subject or patient group chosen for clinical pharmacology studies will provide an adequately 
sensitive measure of difference between the proposed biosimilar product and the reference 
product.  The total number of subjects studied should provide adequate statistical power for PK, 
and, when relevant, PD similarity assessment.  Analysis of the data should be conducted 
                                                 
14 See footnote 4 and footnote 7 for more information on the bridging data needed and examples of issues that a 
sponsor may need to address.    
10  Contains Nonbinding Recommendations 
according to the pre-specified analysis plan, and any post hoc statistical analysis is exploratory 
only.   
 
D.  Dose Selection 
 
As in the selection of study population, the most sensitive dose should be selected to detect and 
to evaluate differences in the PK and PD profiles between the proposed biosimilar product and 
the reference product.  The dose selected should be one most likely to provide clinically 
meaningful and interpretable data.  If a study is conducted in a patient population, the approved 
dose for the reference product can be the appropriate choice, because this approved dose can best 
demonstrate the pharmacological effects in a clinical setting.  However, a lower dose on the 
steep part of the exposure-response curve is generally appropriate when PD is being measured or 
when healthy subjects are selected for evaluation (See section V, Utility of Simulation Tools in 
Study Design and Data Analysis). 
 
In certain cases, a dose selected from a range of doses can be useful for a clinical PK and PD 
similarity assessment.  For example, if the concentration-effect relationship of the reference 
product is known to be highly variable or nonlinear, a range of doses can be used to assess dose-
response (See Section V, Utility of Simulation Tools in Study Design and Data Analysis).   
 
If the product can only be administered to patients, an alternative dosing regimen such as a single 
dose for a chronic indication or a lower dose than the approved dose may be preferable to 
increase the sensitivity for detecting differences if the approved dose either results in nonlinear 
PK or exceeds the dose required for maximal PD effect.  The appropriateness of an alternative 
dosing regimen will depend on certain factors, e.g., whether the lower dose is known to have the 
same effect as the approved dose and whether it is ethically appropriate to give lower doses 
notwithstanding differences in effect.  An adequate justification for the selection of an alternative 
dosing regimen should be provided.  
 
When appropriate, PD biomarkers should be used to assess PK/PD similarity between the 
proposed biosimilar product and the reference product.  Development of an exposure-response 
profile that includes the steep part of the exposure-response curve is a sensitive test for PK/PD 
similarity between products; if clinical pharmacology similarity between products is 
demonstrated, in some instances, the exposure-response profile might support an adequate 
assessment of whether there are clinically meaningful differences between the products, and in 
others, the exposure-response profile might support a more targeted clinical development 
program to address residual uncertainty regarding whether there are any such clinically 
meaningful differences.   
 
E.  Route of Administration 
 
Clinical PK and PD studies should be conducted using the same route of administration for the 
proposed biological product and the reference product.  If more than one route of administration 
(e.g., both intravenous and subcutaneous) is approved for the reference product, the route 
selected for the assessment of PK and PD similarity should be the one most sensitive for  
detecting clinically meaningful differences.  In most cases, the most sensitive route is likely to be 
11  Contains Nonbinding Recommendations 
the subcutaneous or other extravascular routes of administration, because extravascular routes 
can provide insight into potential PK differences during the absorption phase in addition to the 
distribution and elimination phases.  In addition, extravascular routes of administration may 
provide a more sensitive assessment for differences in immunogenicity. 
 
F.  Pharmacokinetic Measures 
 
All PK measures should be obtained for both the proposed biosimilar product and the reference 
product.  The sponsor should obtain measures of  peak concentration (C ) and total area under 
max
the curve (AUC) in a relevant biological fluid.  For single-dose studies, AUC should be 
calculated as the area under the biological product concentration-time curve from time zero to 
time infinity (AUC ), where AUC  = AUC  + C/k  (or C (concentration at the last 
0- 0- 0-t t el t
measurable timepoint) divided by k  (elimination rate constant)) is calculated based on an 
el
appropriate method.  C  should be determined from the data without interpolation.  For 
max
intravenous studies, AUC  will be considered the primary endpoint.  For subcutaneous studies, 
0-
C  and AUC will be considered coprimary study endpoints.  For multiple dose studies, the 
max
measurement of total exposure should be the area under the concentration-time profile from time 
zero to the end of the dosing interval at steady-state (AUC ), and is considered the primary 
0-tau
endpoint.  Both the concentration prior to the next dose during multiple dosing (C ) and C
trough ss max 
are considered secondary endpoints.  Population PK data will not provide an adequate 
assessment for PK similarity. 
 
G.  Pharmacodynamic Measures 
 
In certain circumstances, clinical PK and PD data that demonstrate similar exposure and 
response between a proposed biosimilar product and the reference product can be sufficient to 
completely assess whether there are clinically meaningful differences between products, 
notwithstanding the need for an adequate assessment of immunogenicity.  The biosimilarity 
assessment should be based on similarity in PD using a biomarker that reflects the mechanism of 
drug action when the PD measure has a wide dynamic range over the range of drug 
concentrations achieved during the PK study.  In such instances, a full evaluation of safety and 
immunogenicity should still be conducted.  When human PD data in a PK/PD study are 
insufficient to fully assess whether there are clinically meaningful differences between the 
proposed biosimilar and the reference product, human PD data can nonetheless be helpful to 
support a more targeted approach for the collection of subsequent clinical safety and 
effectiveness data.  Selection of appropriate time points and durations for the measure of PD 
biomarkers will depend on the characteristics of the PD biomarkers (e.g., the timing of the PD 
response after administration of the product based on the half-life of the product and the 
anticipated duration of the products effect).  When a PD response lags after initiation of product 
administration, a study of multiple-dose and steady state conditions can be important, especially 
if the proposed therapy is intended for long-term use.  The PD biomarker(s) evaluated for the 
proposed biosimilar product and the reference product should be compared by determining the 
area under the effect curve (AUEC).  If only one PD measurement is available because of the 
characteristics of the PD biomarker, the measurement should be linked to a simultaneous drug 
concentration measurement. The relationship of drug concentration and the PD biomarker should 
then be used as a basis for comparison between products.   
12  Contains Nonbinding Recommendations 
 
Use of a single, scientifically appropriate PD biomarker as described above, or a composite of 
more than one relevant PD biomarkers, can reduce any residual uncertainty about whether there 
are clinically meaningful differences between products and add significantly to the overall 
demonstration of biosimilarity.  Using broader panels of biomarkers (e.g., by conducting a 
protein or mRNA microarray analysis) that capture multiple pharmacological effects of the 
product can also add value.   
 
When available and appropriate, clinical endpoints in clinical pharmacology studies can also 
provide useful information about the presence of clinically meaningful differences between two 
products.   
 
H.  Defining the Appropriate Pharmacodynamic Time Profile 
 
The optimal sampling strategy for determining PD measures can differ from the strategy used for 
PK measures.  For PK sampling, frequent sampling at early time points following product 
administration with decreased frequency later is generally most effective to characterize the 
concentration-time profile.  However, the PD-time profile might not mirror the PK-time profile.  
In such cases, the PD sampling should be well justified.  When both PK and PD data are to be 
obtained during a clinical pharmacology study, the sampling strategy should be optimized for 
both PK and PD measures.   
 
I.  Statistical Comparison of PK and PD Results  
 
The assessment of the clinical pharmacology similarity of a proposed biosimilar product and the 
reference product in PK and PD studies is based on statistical evaluation.  The recommended 
clinical pharmacology similarity assessment relies on: (1) a criterion to allow the comparison, (2) 
a confidence interval for the criterion, and (3) an acceptable limit for the biosimilarity 
assessment.  FDA recommends that log-transformation of the exposure measures be performed 
before the statistical analysis.  Sponsors should use an average equivalence statistical 
approach15 to compare PK and PD parameters for both replicate and nonreplicate design studies.  
This average equivalence approach involves a calculation of a 90% confidence interval for the 
ratio between the geometric means of the parameters of the proposed biosimilar product and the 
reference product.  To establish PK and/or PD similarity, the calculated confidence interval 
should fall within an acceptable limit.  Selection of the confidence interval and the acceptable 
limits can vary among products.  An appropriate starting point for an acceptable limit for the 
confidence interval of the ratio is 80125%; if other limits are proposed, the sponsor should 
justify the limits selected for the proposed biosimilar product.  There can be situations in which 
the results of the PK and/or PD study fall outside the pre-defined limits.  Because such results 
can suggest existence of underlying differences between the proposed biosimilar product and the 
reference product that can preclude development under the 351(k) pathway, FDA encourages 
sponsors to analyze and explain such findings and discuss them with the FDA before proceeding 
to the next step in the development program.   
                                                 
15 See FDAs guidance for industry Statistical Approaches to Establishing Bioequivalence.   
 
13  Contains Nonbinding Recommendations 
 
 
V.  UTILITY OF SIMULATION TOOLS IN STUDY DESIGN AND DATA 
ANALYSIS 
 
Modeling and simulation tools can be useful when designing a PK and/or PD study.  For 
instance, these tools can contribute to the selection of an optimally informative dose or doses for 
evaluating PD similarity.  When a biomarker-based comparison is used, it is preferable that the 
selected dose be on the steep portion of the dose-response curve of the reference product.  
Sponsors should provide data to support the claim that the selected dose is on the steep part of 
the dose-response curve and not on the plateau of the dose-response curve where it is not likely 
to detect differences between the two products.  Publicly available data for the dose (or 
exposure)-response relationship of the reference product can be analyzed using model-based 
simulations to justify the dose selected for the PK and/or PD study or studies.   
 
If the exposure-response data for the reference product are not available, the sponsor can decide 
to generate this information using a small study to determine an optimally informative dose (e.g., 
a dose representing the effective dose to achieve the 50% maximal response [ED ] of the 
50
reference product).  This small study can involve evaluating the PK/PD relationship at multiple 
dose levels (e.g., the low, intermediate, and highest approved dose) to obtain dose-response 
and/or exposure-response data.16  Alternatively, when possible, sponsors can conduct a PK/PD 
similarity study between the reference product and the proposed biosimilar product with low, 
intermediate, and the highest approved doses where a clear dose-response is observed.  If 
multiple doses are studied, PK/PD parameters such as EC , the maximum PD response (E ), 
50 max
and the slope of the concentration-effect relationship should be evaluated for similarity.  Such 
studies should be useful for the demonstration of PK, PK/PD, and PD similarity when the 
clinical pharmacology evaluation is likely to be the major source of information to assess 
clinically meaningful differences.  Publicly available information on biomarker-clinical endpoint 
relationships accompanied with modeling and simulation can also be used to define the 
appropriate limits for PD similarity.  
 
 
VI.  CONCLUSION 
 
Clinical pharmacology studies play a critical role in the development of biosimilar products.  
These studies are part of a stepwise process for demonstrating biosimilarity between a proposed 
biosimilar product and the reference product. These studies may support a demonstration that 
there are no clinically meaningful differences between the products.  These studies may address 
residual uncertainties that remain after the analytical evaluation, may add to the totality of the 
evidence supporting a demonstration of biosimilarity, and may also support a selective and 
targeted approach to the design of any recommended subsequent clinical studies to support a 
demonstration of biosimilarity.   
                                                 
16 For more information, see FDAs guidance for industry Topical Dermatologic Corticosteroids: In Vivo 
Bioequivalence.   
 
14  Contains Nonbinding Recommendations 
DEFINITIONS 
 
Biological product:  [A] virus, therapeutic serum, toxin, antitoxin, vaccine, blood, blood 
component or derivative, allergenic product, protein (except any chemically synthesized 
polypeptide), or analogous product, or arsphenamine or derivative of arsphenamine (or any other 
trivalent organic arsenic compound), applicable to the prevention, treatment, or cure of a disease 
or condition of human beings.17 
 
Biosimilar or biosimilarity:  In reference to a biological product that is the subject of an 
application under subsection 351(k) of the PHS Act, the term biosimilar or biosimilarity 
means that the biological product is highly similar to the reference product notwithstanding 
minor differences in clinically inactive components; and that there are no clinically meaningful 
differences between the biological product and the reference product in terms of the safety, 
purity, and potency of the product.18 
 
Exposure:  In this guidance, we use the broad term exposure to refer to PK variables, including 
input of all active components of the biological product as measured by dose (drug input to the 
body) and various measures of single or integrated drug concentrations in plasma and other 
biological fluid, e.g., peak concentration (C ), concentration prior to the next dose during 
max
multiple dosing (C ), and area under the plasma/blood concentration-time curve (AUC). 
trough ss
 
Fingerprint-like:  Integrated, multi-parameter approaches that are extremely sensitive in 
identifying analytical differences.  
 
Immunogenicity:  In this guidance immunogenicity refers to an immune response that a 
biological product elicits through formation of antibodies to the administered biological product. 
 
Reference product:  The single biological product licensed under section 351(a) of the PHS Act 
against which a biological product is evaluated in a 351(k) application.19   
 
Exposure-Response:  Pharmacodynamic response, referred to here as PD (a direct measure of 
the pharmacological or toxicological effect of a drug) in relationship to drug exposure (PK) 
variables. 
 
Average equivalence:  An approach to statistical analysis for pharmacokinetic measures, such 
as area under the curve (AUC) and peak concentration (C ).  It is based on the two one-sided 
max
tests procedure to determine whether the average values for the pharmacokinetic measures 
determined after administration of the Test (T) and Reference (R) products are comparable.  This 
approach involves the calculation of a 90% confidence interval for the ratio of the log-
transformed averages of the measures for the T and R products. 
 
 
                                                 
17 Section 351(i)(1) of the PHS Act. 
18 Section 351(i)(2) of the PHS Act. 
19 Section 351(i)(4) of the PHS Act. 
15