Chronic Hepatitis B 
Virus Infection:  
Developing Drugs for 
Treatment 
Guidance for Industry 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Drug Evaluation and Research (CDER) 
 
April 2022 
Clinical/Antimicrobial 
 
  Chronic Hepatitis B 
Virus Infection:  
Developing Drugs for 
Treatment 
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/guidance-compliance-regulatory-information/guidances-drugs 
 
 
 
 
 
 
 
 
 
 
 
 
U.S. Department of Health and Human Services 
Food and Drug Administration 
Center for Drug Evaluation and Research (CDER) 
 
April 2022 
Clinical/Antimicrobial 
  TABLE OF CONTENTS 
 
I.  INTRODUCTION............................................................................................................ 1 
II.  BACKGROUND .............................................................................................................. 2 
III.  DEVELOPMENT PROGRAM ...................................................................................... 2 
A.  General Drug Development Considerations .............................................................................. 2 
1. Early Phase Development Considerations .................................................................................... 3 
a.  Pharmacology/toxicology development considerations .......................................................... 3 
b.  Nonclinical virology development considerations .................................................................. 4 
c.  Clinical pharmacology considerations .................................................................................... 6 
2. Drug Development Population ...................................................................................................... 7 
3. Safety Considerations .................................................................................................................... 8 
B.  Phase 3 Efficacy Trial Considerations ....................................................................................... 9 
1. Trial Design ................................................................................................................................... 9 
a.  Chronic suppressive therapy ................................................................................................... 9 
b.  Finite duration therapy .......................................................................................................... 10 
2. Trial Population ........................................................................................................................... 11 
3. Entry Criteria ............................................................................................................................... 11 
4. Randomization, Stratification, and Blinding................................................................................ 12 
5. Specific Populations..................................................................................................................... 12 
a.  Patients with HBV/HIV-1 coinfection .................................................................................. 12 
b.  Patients with HBV/HDV coinfection .................................................................................... 12 
c.  Pediatric patients ................................................................................................................... 13 
6. Dose Selection .............................................................................................................................. 14 
7. Efficacy Endpoints ....................................................................................................................... 15 
8. Trial Procedures and Timing of Assessments .............................................................................. 16 
9. Statistical Considerations ............................................................................................................ 16 
a.  Analysis populations ............................................................................................................. 16 
b.  Efficacy analyses .................................................................................................................. 17 
c.  Handling missing data ........................................................................................................... 17 
10.  Accelerated Approval (Subpart H/I) Considerations .............................................................. 17 
11.  Benefit-Risk Considerations .................................................................................................... 17 
C.  Other Considerations................................................................................................................. 18 
1. Clinical Virology Considerations ................................................................................................ 18 
2. Pharmacokinetic/Pharmacodynamic Considerations .................................................................. 20 
3. Labeling Considerations .............................................................................................................. 20 
GLOSSARY OF ACRONYMS ................................................................................................ 22 
REFERENCES ........................................................................................................................... 23 
  Contains Nonbinding Recommendations 
Chronic Hepatitis B Virus Infection: 
Developing Drugs for Treatment  
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 clinical development of drugs and 
biologics for the treatment of chronic hepatitis B virus (HBV) infection from the initial 
investigational new drug application (IND) through the new drug application (NDA)/biologics 
license application (BLA) and postmarketing phases.2  Sponsors are also encouraged to 
communicate with the Division of Antivirals through the pre-IND consultation program to obtain 
advice in the development of drugs with unique considerations based on mechanism of action, 
novel treatment approaches, or the use of novel biomarkers.3   
 
This guidance does not address development of vaccines or blood-derived products, which are 
regulated by the Center for Biologics Evaluation and Research.  This guidance also does not 
discuss general issues of statistical analysis or clinical trial design.  Those topics are addressed in 
the ICH guidances for industry E9 Statistical Principles for Clinical Trials (September 1998) 
and E10 Choice of Control Group and Related Issues in Clinical Trials (May 2001), 
respectively.4   
 
The contents of this document do not have the force and effect of law and are not meant to bind 
the public in any way, unless specifically incorporated into a contract.  This document is 
intended only to provide clarity to the public regarding existing requirements under the law.  
FDA guidance documents, including this guidance, should be viewed only as recommendations, 
 
1 This guidance has been prepared by the Division of Antivirals 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 unless otherwise specified.  
 
3 See the Division of Antivirals Pre-IND Letter of Instruction web page at https://www.fda.gov/drugs/pre-ind-
consultation-program/division-anti-viral-dav-pre-ind-letter-instruction. 
 
 
4 We update guidances periodically.  For the most recent version of a guidance, check the FDA guidance web page 
at https://www.fda.gov/regulatory-information/search-fda-guidance-documents.   
1  Contains Nonbinding 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  
 
HBV is an enveloped DNA virus belonging to the Hepadnavirus family.  The highly stable 
covalently closed circular viral DNA (cccDNA) functions as a nonreplicative minichromosome 
and persists throughout the lifespan of infected hepatocytes.  The cccDNA is not eliminated by 
currently approved therapies that include drugs from the nucleoside/nucleotide reverse 
transcriptase inhibitor (NrtI) class, and pegylated interferon.    
 
Chronic HBV (CHB) infection results in progressive liver disease ranging from asymptomatic to 
severe disease with complications including cirrhosis, liver failure, and the development of 
hepatocellular carcinoma (HCC).  In untreated adults with CHB, the cumulative 5-year incidence 
of cirrhosis is 8 to 20 percent; and among those with cirrhosis, the 5-year cumulative risk of 
hepatic decompensation is 20 percent, and risk of HCC is 2 to 5 percent (Terrault et al. 2016).  
An effective vaccine and antiviral therapies are approved for the prevention of HBV infection 
and treatment of CHB, respectively.   
 
Currently available therapies achieve sustained suppression of HBV DNA while on treatment, 
but rates of HBV surface antigen (HBsAg) loss with or without seroconversion to anti-HBsAg 
(HBsAb) remain low.  Sustained HBV DNA suppression is associated with serum alanine 
aminotransferase (ALT) normalization and improvement in liver histology including regression 
of hepatic fibrosis and cirrhosis (Chang et al. 2010; Marcellin et al. 2013; Buti et al. 2015).  
Effective antiviral therapy for CHB reduces disease-related complications, such as hepatic 
decompensation and liver failure, and decreases risk of HCC (Lok et al. 2016; Papatheodoridis et 
al. 2017).  Clearance of HBsAg is associated with reduced risk of hepatic decompensation and 
improved survival (Terrault et al. 2016).  HBsAg loss is considered the best predictor of 
sustained remission off-treatment (Terrault et al. 2016).  New finite duration therapies target 
treatment regimens that can achieve sustained suppression of HBV DNA off-treatment with 
HBsAg loss (with or without HBsAb seroconversion) with low risk of virologic relapse (as 
defined by HBV DNA) and minimal risk of liver disease progression after the treatment is 
stopped (Lok et al. 2017).    
 
 
III.  DEVELOPMENT PROGRAM  
 
A.  General Drug Development Considerations 
 
This section discusses nonclinical and early phase clinical development considerations, followed 
by issues related to the target population for drug development, assessment of activity in early 
phase trials, and safety considerations.     
 
2  Contains Nonbinding Recommendations 
1.  Early Phase Development Considerations 
 
Early clinical evaluation should provide sufficient data to establish safety and evidence of 
antiviral activity to support the phase 3 trials.    
 
a.  Pharmacology/toxicology development considerations  
 
Pharmacology/toxicology development considerations for single investigational drugs intended 
to treat CHB should follow the approaches outlined in existing guidances for drug development.5   
 
When a combination of two or more early stage investigational drugs to treat CHB is being 
developed, sponsors should discuss with the FDA whether combination toxicology studies are 
needed (including the design of such studies) to support clinical trials for the intended 
combination.  When combination toxicology studies are conducted, usually no more than two 
drugs should be tested simultaneously in a particular arm of a toxicology study.  Nonclinical 
combination studies of an investigational drug plus an approved drug or licensed biological 
product generally are not needed unless data from nonclinical studies of an investigational drug 
suggest a potential for serious synergistic toxicity with an approved drug or licensed biological 
product.  
 
In general, sponsors developing drugs intended to treat CHB with proposed treatment durations 
of 6 months or more should conduct carcinogenicity studies.6  Sponsors should submit 
carcinogenicity studies with an initial NDA.  However, under limited circumstances and with 
prior written agreement, the FDA may consider allowing sponsors to submit the completed 
carcinogenicity studies during the postmarketing period under section 505(o)(3) of the Federal 
Food, Drug, and Cosmetic Act (FD&C Act)7, as long as the studies have been initiated before 
NDA submission. 
 
Sponsors developing biological products should follow approaches outlined in ICH S6(R1) and 
discuss their proposals for a carcinogenicity risk assessment with the FDA during clinical 
development to facilitate a final assessment needed to support a BLA.  
 
 
5 See the ICH guidances for industry M3(R2) Nonclinical Safety Studies for the Conduct of Human Clinical Trials 
and Marketing Authorization for Pharmaceuticals (January 2010) and S6(R1) Preclinical Safety Evaluation of 
Biotechnology-Derived Pharmaceuticals (May 2012).  
  
6 See the ICH guidance for industry S1A The Need for Long-Term Rodent Carcinogenicity Studies of 
Pharmaceuticals (March 1996).  
 
 
7 See also the draft guidance for industry Postmarketing Studies and Clinical TrialsImplementation of Section 
505(o)(3) of the Federal Food, Drug, and Cosmetic Act (October 2019).  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/regulatory-information/search-fda-guidance-documents.  
  
3  Contains Nonbinding Recommendations 
b.  Nonclinical virology development considerations 
 
Sponsors should consider recommendations for general antiviral drug development in the draft 
guidance for industry Antiviral Product DevelopmentConducting and Submitting Virology 
Studies to the Agency (February 2014).8  The FDA encourages detailed study reports describing 
the mechanism of action, antiviral activity in cell culture, cytotoxicity, and mitochondrial 
toxicity (Marroquin et al. 2007; Arnold et al. 2012), animal models,9 and resistance studies.  
Additionally, sponsors are advised to provide the following nonclinical virology data for 
investigational drugs developed specifically for the treatment of CHB.    
 
Resistance and cross-resistance 
 
HBV is differentiated into 10 genotypes (A through J) with many different subtypes.  Drugs 
targeting viral proteins or sequence targets will likely be impacted by genotype- and subtype-
specific variations that occur in the drug target as well as variations that evolve in the presence of 
the drug that allow the virus to develop resistance.  Therefore, characterizing resistance pathways 
and their relationship to HBV genomic variations helps identify subjects who will benefit from 
the drug and also helps identify potential cross-resistance with other drugs having a similar 
mechanism of action. 
 
HBV does not generally grow well enough in cell culture to select for resistant virus in the 
presence of drug.  When possible, we recommend that resistance assessments be performed for 
all animal studies that assess the antiviral activity of an investigational drug in infected animals 
receiving the drug that experience viral breakthrough.  In addition, a resistance monitoring plan 
for identifying changes in viral targets associated with treatment failure should be included in the 
protocols or can be submitted separately for each clinical trial that will treat subjects with CHB.    
 
  Amino acid substitutions or nucleotide mutations associated with the development of 
resistance (i.e., changes from baseline) to an investigational drug should be determined 
by sequencing the drug target and validated by introducing resistance-associated 
substitutions or mutations into the HBV reference genome using site-directed 
mutagenesis and determining the fold-shift in susceptibility.  Results from these studies 
help identify resistance pathways and support the drugs proposed mechanism of action.  
Lack of a shift in susceptibility does not exclude a resistance association for a specific 
substitution or mutation that occurs in two or more independent events.   
 
Cross-resistance should be assessed to determine if resistance against approved HBV drugs with 
the same target confers resistance to the drug being developed and vice versa.  The development 
of cross-resistance to epitopes of approved HBV vaccines should be assessed.  
 
 
8 When final, this guidance will represent the FDAs current thinking on this topic. 
 
9 We support the principles of the 3Rs (reduce/refine/replace) for animal use in testing when feasible.  FDA 
encourages sponsors to consult with review divisions when considering a nonanimal testing method believed to be 
suitable, adequate, validated, and feasible.  FDA will consider if the alternative method could be assessed for 
equivalency to an animal test method. 
4  Contains Nonbinding Recommendations 
Considerations for oligonucleotide-based investigational drugs 
 
Knockdown of viral protein expression via oligonucleotide-based therapeutics is an active area 
for the development of antiviral drugs.  These drugs, which have a nucleic acid target, present 
potential off-target binding at mismatched sequences that could lead to species-specific toxicities 
not necessarily detected in toxicology studies.  Therefore, we recommend that sequence-
dependent off-target assessments be conducted for such drugs using appropriate in silico and in 
vitro methodologies to identify potential off-target mismatches that can be investigated and 
potentially monitored during clinical development.  The Division recognizes that many 
additional factors, such as temporal and cell type specific expression, pharmacokinetic (PK) 
properties, and hybridization-dependent efficiencies may be considered in the overall risk 
assessment of potential off-target effects (refer to Lindow et al. 2012 for a general approach). 
The following points should be considered when designing sequence-dependent off-target 
assessments: 
 
  Several different classes of oligonucleotide-based therapeutics (small interfering RNAs 
and antisense oligonucleotides, etc.) exist.  The criteria for defining high-risk off-target 
mismatches are expected to differ for each class; appropriate criteria should be provided 
and justified for each drug. 
 
  All elements of the oligonucleotide drug product that are potentially available for 
mismatch binding should be assessed, including both the sense and antisense strands, 
overlapping ends, etc. 
 
  There are multiple in silico approaches that may be considered, and we recommend 
sponsors justify the methods and approaches used.  In silico studies can be used to do the 
following: 
 
  Identify all potential off-target mismatches, regardless of tissue expression, in the 

human transcriptome, including the mitochondrial transcriptome; for each of these, 
describe available information on mouse knockouts and human genetic diseases.  
Sponsors should specify and justify the criteria for defining a potential off-target 
mismatch.  
 
  Determine whether each off-target site in the human transcriptome is conserved in the 

animal species in which antiviral assessments and pivotal toxicology studies are 
performed.  
 
  Determine the variation within the off-target mismatches in the transcriptomes of 

different ethnic populations in the United States, when feasible, to assess whether 
certain populations may be more susceptible to off-target effects than others. 
 
  In vitro assessments of possible off-target binding (cell-based analysis, RNA-seq, etc.) 
should be conducted to confirm or eliminate the importance of each off-target mismatch 
determined via in silico assessments. 
 
5  Contains Nonbinding Recommendations 
  An overall risk assessment of potential off-target effects should be provided along with 
the off-target assessment data. 
 
Considerations for drugs developed to modulate innate and adaptive immune responses  
 
Drugs developed to modulate innate and adaptive immune responses to chronic HBV infection 
are likely to target host factors and induce or repress immune biomarkers before having an 
impact on HBV replication or clearance of HBV infected hepatocytes.  Nonclinical studies 
should describe the specific mechanism of action of the drug and demonstrate that immune 
modulation in cell culture and animal models of HBV infection results in suppression of HBV 
replication, as measured by HBV DNA or HBsAg loss, or that the HBV cccDNA reservoir is 
reduced by assessing cccDNA levels.  In addition, given that these drugs may target host factors, 
it is important to determine that the target of the drug is conserved, having similar affinity 
between the animal species being assessed and the human target. 
 
Targeting host factors 
 
For drugs targeting host factors, polymorphisms in the gene encoding the target should be 
assessed to determine whether the drug will be more effective or less effective in different 
populations.  Sponsors should identify key racial and ethnic groups in the United States who will 
be part of the proposed indication at the initiation of drug development.  If a nonclinical assay to 
assess the drug effect is available, multiple samples from each of these groups in the United 
States should be evaluated to determine whether race or ethnicity may be a factor contributing to 
efficacy (Forde et al. 2013).  Samples should be collected during clinical trials to determine the 
virus genotype of subjects who respond less favorably to treatment.   
 
c.  Clinical pharmacology considerations 
 
In general, dose selection for early efficacy trials should be predicted to provide plasma drug 
exposures that exceed by severalfold the protein binding-adjusted, cell culture EC50 value of the 
drug for the relevant HBV genotype/subtype.  In cases where either total (protein bound plus 
unbound) or free (unbound) drug concentrations in plasma may not be related to antiviral 
activity, sponsors may provide justification for dose selection based on drug concentration in 
other relevant tissues.  The dose selection should also consider the safety data from the previous 
phase 1 trials and animal studies.    
 
Sponsors should refer to the appropriate clinical pharmacology guidances for industry to inform 
the need and design of drug-drug interaction studies and PK studies in patients with renal or 
hepatic impairment.10  We encourage sponsors to conduct these studies, if needed, early in 
 
10 See the guidance for industry Pharmacokinetics in Patients With Impaired Hepatic Function:  Study Design, Data  
Analysis, and Impact on Dosing and Labeling (May 2003).  See also the guidances for industry Clinical Drug 
Interaction StudiesCytochrome P450 Enzyme- and Transporter-Mediated Drug Interactions  (January 2020), In 
Vitro Drug Interaction StudiesCytochrome P450 Enzyme- and Transporter-Mediated Drug Interactions (January 
2020), and the draft guidance for industry Pharmacokinetics in Patients With Impaired Renal FunctionStudy 
Design, Data Analysis, and Impact on Dosing (September 2020).  When final, this guidance will represent the 
FDAs current thinking on these topics.  
6  Contains Nonbinding Recommendations 
development to inform the management of drug interactions and the inclusion of patients with 
renal and hepatic impairment in phase 3 trials as appropriate.  See section III. B. 6., Dose 
Selection, for dose selection for phase 2 and 3 trials and section III. C. 2.,  
Pharmacokinetic/Pharmacodynamic Considerations, for other PK and pharmacodynamic 
considerations.  
  
2.  Drug Development Population 
 
Therapies should be developed for use in a wide range of patients with CHB, including pediatric 
populations.    
  
Early phase clinical trials should focus on the adult population without cirrhosis.  Initial trials can 
be conducted in HBV e antigen positive (HBeAg-positive) or HBV e antigen negative (HBeAg-
negative) patients with active disease who are treatment-naive, or those who were previously 
treated but currently viremic, off-treatment.  Trials can also be conducted in HBeAg-positive or 
HBeAg-negative patients who are virally suppressed on NrtIs.  In addition to endpoints discussed 
in section III. B., sponsors can evaluate exploratory endpoints in early phase trials to gather data 
to inform and support the choice of appropriate endpoints in late phase trials, particularly those 
evaluating treatments of finite durations.  Some of these exploratory endpoints may include the 
following:  
  
  Change in quantitative HBsAg (qHBsAg) concentration at various time points on 
treatment  
  Quantitative HBeAg levels  
  Quantitative HBV RNA levels 
  Quantitative HBV core-related antigen (HBcrAg) levels 
  cccDNA quantification  
  Quantitative HBsAg levels from integrated HBV genome fragments  
  Quantitative HBsAg-anti-HBs immune complex levels    
  
Also depending on the drugs mechanism of action, liver biopsy findings can be used in certain 
proof-of-concept studies to further understand the effect of the drug on the cccDNA reservoir 
and/or to help better understand potential surrogate markers of antiviral activity.   
  
CHB is a global disease, and clinical trials are often conducted in multiple countries.  FDA will 
accept a well-designed and well-conducted foreign clinical study not conducted under an IND as 
support for an IND or application for marketing approval if the trial was conducted in 
accordance with good clinical practice and if the FDA is able to validate the data from the trial 
through an onsite inspection if the Agency deems it necessary.11  When sponsors rely on foreign 
data, these should be supported with information about predominant HBV genotypes and 
subtypes in the region or regions (Schweitzer et al. 2015).  Development programs should 
include a sufficient number of U.S. patients to ensure prevalent genotypes in the U.S. population 
are represented.  FDA strongly encourages sponsors to provide a plan to address inclusion of 
 
11 See 21 CFR 312.120; 21 CFR 314.106. 
 
7  Contains Nonbinding Recommendations 
clinically relevant subpopulations with regard to age, gender, race, and ethnicity in the clinical 
trials to support an NDA or BLA, no later than the end-of-phase 2 meeting.12    
  
3.  Safety Considerations 
  
In general, we recommend that initial marketing applications for drugs intended to treat CHB 
contain a total safety database of about 1,000 to 1,500 subjects exposed to the proposed dose and 
duration of treatment.  Depending on the drug safety profile and concerns identified during the 
development process, a larger database or long durations of posttreatment follow-up may be 
needed.    
  
In addition to routine safety monitoring, the safety monitoring plans should consider the 
mechanism of action of the drug, as there may be different safety concerns associated with 
specific drugs both during treatment and after treatment cessation.  Some of the notable safety 
concerns associated with immune modulatory therapies are immune-mediated hepatitis flares and 
autoimmunity (e.g., immune-related adverse events observed with checkpoint inhibitors).  The 
specific criteria for defining and monitoring for hepatitis flares during treatment and after 
stopping therapy should be prespecified in the clinical trial protocols.  The safety profile 
observed in early phase trials should guide the monitoring plan for late phase trials. 
   
Severe acute exacerbations of HBV infection may occur after discontinuation of anti-HBV 
therapy, particularly in the absence of HBsAg loss.  Subjects should be monitored closely with 
both laboratory and clinical follow-up after discontinuation of anti-HBV therapy.  The duration 
of follow-up should consider the mechanism of action and the half-life of the specific 
investigational drug.  In certain circumstances, resumption of anti-HBV therapy may be 
warranted.  The detailed plan and criteria for treatment reinitiation should be prespecified in the 
clinical trial protocols. 
 
Clinical trial protocols should include predefined algorithms for data collection in the setting of 
significant hepatic events to ensure that the relevant data are available for further assessment and 
adjudication of these cases to differentiate between potential etiologies.  The outcomes for all 
serious hepatic events should be systematically evaluated during clinical development.  
Evaluation by an independent adjudication committee is encouraged.  
 
For a drug approved for use in patients without cirrhosis or with compensated cirrhosis, the 
database needed to extend use to the decompensated cirrhotic population would depend on the 
safety profile of the investigational drug and the overall benefit-risk profile for the indicated 
population.  Similarly, obtaining safety data in other subpopulations, such as in patients 
coinfected with hepatitis D virus (HDV), may be important for certain clinical development 
programs.  We encourage sponsors to discuss with the FDA safety-related considerations, 
including but not limited to the size of the safety database, before the initiation of phase 3 trials.  
 
 
12 See the guidance for industry Collection of Race and Ethnicity Data in Clinical Trials (October 2016). 
8  Contains Nonbinding Recommendations 
B.  Phase 3 Efficacy Trial Considerations 
 
Sponsors can submit an NDA/BLA to support marketing approval of a drug in a single patient 
population (e.g., treatment-naive patients or patients who are virally suppressed on NrtIs).  Such 
an application should include at least two adequate and well-controlled trials conducted in the 
proposed population.  Alternatively, sponsors can choose to pursue an indication for different 
populations (e.g., a trial in treatment-naive subjects and a second trial in subjects who are virally 
suppressed on NrtIs).  In these situations, the NDA/BLA should contain at least one adequate 
and well-controlled trial in each patient population, with adequate supporting data.  
  
1.  Trial Design 
  
Randomized controlled trials are recommended to establish efficacy because of the heterogeneity 
of the natural course of CHB.  Appropriate trial designs depend on whether the therapeutic is 
intended for chronic suppressive therapy or therapy of finite duration as discussed below.   
  
a.  Chronic suppressive therapy 
 
Sponsors developing drugs for chronic suppressive therapy of CHB should consider the 
following trial design options: 
  
  A randomized controlled noninferiority (NI) (or superiority) trial comparing the 
investigational drug with an approved active control armThe primary efficacy endpoint 
should be undetectable HBV DNA13 after 48 weeks on-treatment in HBeAg-positive 
subjects, HBeAg-negative subjects, or both.  The active comparator should be an antiviral 
drug that is recommended for treatment of CHB and reflects current practice at the time 
of trial initiation.  The patient population may be treatment-naive or previously treated 
subjects with detectable HBV DNA.  Potential concerns related to the development of 
resistance when evaluating an investigational drug with a low barrier to resistance as a 
monotherapy should be addressed.  For an NI design, determination of the NI margin 
should be discussed with the FDA. 
 
  A randomized controlled add-on superiority trial comparing an investigational drug plus 
an approved NrtI with an NrtI aloneHowever, for this trial design, it is not clear what 
the most appropriate primary endpoint should be to demonstrate the investigational 
drugs clinical contribution.  Presently, HBV DNA is not recommended as a primary 
endpoint for an add-on trial because it is unclear what incremental numerical benefits in 
HBV DNA, over the substantial HBV DNA suppression achieved by an NrtI alone, are 
predictive of clinical benefit.  Other endpoints such as HBsAg clearance or another 
surrogate found to be predictive of clinical benefit may be considered; however, the trial 
design and endpoints should be discussed with the FDA in advance of trial initiation. 
 
 
13 Defined as less than the lower limit of quantification (LLOQ), specifying if the results are target detected (TD) or 
target not detected (TND). 
 
9  Contains Nonbinding Recommendations 
b.  Finite duration therapy 
 
The appropriate trial design depends on the patient population being studied and the treatment 
regimen being evaluated.  See section III. B. 7., Efficacy Endpoints for a description of efficacy 
endpoints that could be used for evaluation of finite duration therapies in clinical trials. 
 
Sponsors developing drugs for finite duration therapy of CHB should consider the following trial 
design options: 
 
Virally suppressed on NrtIs  
  
To evaluate the primary efficacy outcome of sustained HBV DNA suppression off-treatment 
with HBsAg loss in subjects with active disease (HBeAg-positive or HBeAg-negative CHB) who 
are virally suppressed on NrtIs, sponsors can consider an add-on superiority trial against placebo 
with a current NrtI treatment regimen as the background therapy.  The co-primary efficacy 
endpoints14 of HBsAg loss and sustained HBV DNA suppression should be assessed at the 6-
month posttreatment time point with additional follow-up to monitor for durability of response 
(i.e., sustained HBV DNA suppression and HBsAg loss) off-treatment.  In general, off-treatment 
refers to discontinuation of all therapies (i.e., investigational agent and background NrtI 
regimen).  To demonstrate sustained response off-treatment, sponsors should systematically 
assess the duration of a treatment consolidation period, defined as the duration of continued 
treatment needed after achieving HBsAg loss, during late phase trials.  This may vary based on 
the mechanism of action and half-life of the specific investigational drug.  
  
Sponsors should use the following criteria for stopping NrtI therapy at the end of the 
investigational treatment period when evaluating a finite duration therapy:  (1) applied equally 
across treatment arms, as applicable; (2) well-defined in the protocol; and (3) stringent, such as 
HBsAg loss or substantial HBsAg decline or marked reduction in other important biomarkers 
identified in phase 2 trials.  It is expected that few subjects would meet such criteria on the 
placebo arm.  The criteria for stopping NrtI therapy should be based on clinical evidence that 
also reflects current practice guidelines recommended by the authoritative scientific bodies to 
ensure that the discontinuation of NrtI therapy does not pose undue safety risk to the trial 
participants.  Sponsors should discuss with the FDA the use of biomarkers as a trigger for 
treatment interruption in advance of trial initiation.  
 
Alternatively, an outcome of sustained HBV DNA suppression off-treatment without HBsAg 
clearance can be evaluated after a finite treatment duration using a superiority trial design 
comparing the investigational drug plus an NrtI with an NrtI alone. 
  
Treatment-naive  
  
An outcome of sustained HBV DNA suppression off-treatment with HBsAg loss can be 
evaluated in treatment-naive subjects by demonstrating superiority to an active control, or, in 
 
14 See draft guidance for industry Multiple Endpoints in Clinical Trials (January 2017).  When final, this guidance 
will represent the FDAs current thinking on this topic. 
 
10  Contains Nonbinding Recommendations 
patients in whom treatment is currently not indicated per treatment guidelines, superiority to 
placebo.  In certain patient populations (e.g., for patients in the immune-tolerant phase with mild 
necroinflammation or fibrosis) comparison with placebo may be feasible as current treatment 
guidelines do not recommend treatment for these patients.  In some of the trial design scenarios, 
subjects in the placebo group may be rolled over to an active investigational drug before the 
completion of the trial based on a prespecified interim analysis.  Sponsors should prospectively 
plan these analyses15 and should discuss with the FDA before trial enrollment.  
  
Sponsors considering an NI trial design should discuss in advance with the FDA their trial 
designs and justifications of the proposed NI margin based on historical evidence of treatment 
effect of the active control.  In general, the active comparator in an NI trial should be an FDA-
approved drug that is considered the standard of care for the specific indication and population 
being studied.  A detailed protocol and statistical analysis plan (SAP) should be submitted for 
review.   
  
2.  Trial Population 
  
Patients fulfilling one of the following two criteria for CHB should be enrolled (Centers for 
Disease Control and Prevention 2012):  
  
(1)  Negative immunoglobulin M (IgM) antibodies to HBV core antigen and a positive result 
on one of the following tests:  HBsAg, HBeAg, or nucleic acid test for hepatitis B virus 
DNA (including qualitative, quantitative, and genotype testing); or  
  
(2)  Positive HBsAg result or positive nucleic acid test for HBV DNA (including qualitative, 
quantitative, and genotype testing) or positive HBeAg on two occasions at least 6 months 
apart (any combination of these tests performed 6 months apart is acceptable).  
  
Sponsors should consider evaluating drug efficacy in key CHB subpopulations, including 
but not limited to the following:   
  
  HBeAg-positive and HBeAg-negative subjects  
  Subjects with cirrhosis  
  Subjects with decompensated liver disease  
  
3.  Entry Criteria 
  
The presence or absence of cirrhosis at study entry should be documented based on clinical 
assessment or previous liver biopsy, when available.  As the criteria for diagnosing cirrhosis 
using noninvasive methods are not yet well defined, particularly in patients who are virally 
suppressed, the use of a noninvasive modality to define presence or absence of cirrhosis in a trial 
protocol should be supported by references that summarize performance characteristics and 
sensitivity and specificity of the modality for identifying patients with cirrhosis.   
  
 
15 See the guidance for industry Adaptive Designs for Clinical Trials of Drugs and Biologics (November 2019). 
 
11  Contains Nonbinding Recommendations 
4.  Randomization, Stratification, and Blinding 
 
Sponsors should conduct randomized, double-blind trials whenever feasible to reduce the 
likelihood of potential biases.  In general, trials should be designed to evaluate the effect of 
investigational therapies in subjects with key disease characteristics.  Patient subpopulations 
could be evaluated in separate trials or as separate strata in a single trial.  If multiple patient 
populations are included in a single trial, sponsors should consider stratifying groups at 
randomization based on baseline variables (e.g., HBeAg status, HBsAg level, presence or 
absence of cirrhosis, and HBV DNA level) and ensure that an adequate number of subjects are in 
each stratum to provide informative data.    
  
5.  Specific Populations 
 
a.  Patients with HBV/HIV-1 coinfection 
 
The overall treatment goals for patients with HBV/HIV coinfection remain identical to those 
described for the HBV-monoinfected population.  Sponsors should consider including subjects 
coinfected with HIV in the clinical trials evaluating finite duration HBV therapies.  In cases 
where there is a strong rationale for exclusion of patients with HBV/HIV coinfection, the 
rationale should be addressed in the trial protocol.  The concurrent use of HIV antiretroviral 
drugs that are also effective against HBV (e.g., HIV nucleoside/nucleotide reverse transcriptase 
inhibitors) may have implications for treatment cessation when evaluating finite duration HBV 
therapies and possibly confound interpretation of efficacy outcome as subjects continue their 
HIV nucleoside/nucleotide reverse transcriptase inhibitor -based antiretroviral regimen.  Because 
of the various interactions between HIV and HBV therapies, we recommend sponsors discuss 
their plans with and obtain feedback from the FDA.    
 
b.  Patients with HBV/HDV coinfection 
 
Infection with HDV only occurs in the setting of concurrent HBV infection (Wranke and 
Wedemeyer 2016).  According to the World Health Organization, an estimated 15 million to 20 
million people worldwide are living with HBV/HDV coinfection (World Health Organization 
2017).  A meta-analysis reported a much higher worldwide HBV/HDV coinfection prevalence of 
62 million to 72 million (Chen et al. 2019).  Relative to HBV monoinfection, HBV/HDV 
coinfection leads to more severe liver disease resulting in a greater risk of cirrhosis, HCC, and 
hepatic decompensation/failure. 
 
The ultimate goal of treatment in patients with HBV/HDV coinfection is clearance or long-term 
suppression of both viruses.  CHB treatment leading to loss of HBsAg may ultimately lead to the 
clearance of HDV infection (Wranke and Wedemeyer 2016).  HDV superinfection frequently 
leads to spontaneous suppression of HBV (Huang and Lo 2014), and the effect of specific HBV 
therapies on the interplay between the two viruses cannot be predicted.  The possibility of HBV 
reactivation needs to be considered when HDV is suppressed after HDV treatment initiation.  To 
mitigate the risk of HBV reactivation and associated hepatitis flares, it is preferred to have 
patients on NrtI suppressive therapy before they receive HDV treatment.  Recommendations for 
trials in patients with HBV/HDV coinfection are beyond the scope of this guidance and are 
12  Contains Nonbinding Recommendations 
included in a separate draft guidance.16  Sponsors should discuss development plans directly with 
the FDA. 
 
c.  Pediatric patients  
 
Pediatric assessments are required under section 505B of the FD&C Act as part of the overall 
drug development program for a new active ingredient, new indication, new dosage form, new 
dosing regimen, or new route of administration,17 unless those assessments are waived.18  
Sponsors should discuss their plans for pediatric assessments with the review division and be 
aware of timing and content requirements for pediatric study plans under section 505B(e) of the 
FD&C Act.  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 the FDA and the sponsor.19  
Because progressive liver disease is uncommon in young children with HBV infection, it is 
generally not recommended to include patients younger than 2 years of age in most development 
programs.  Further, treatment generally is not recommended in children younger than 2 years of 
age, as per current treatment guidelines (Terrault et al. 2016).   
 
In general, pediatric clinical trials can be initiated after phase 2 adult data characterizing the 
safety profile and preliminary evidence of efficacy are available.  Given the natural history of 
CHB infection, the dynamic relationship between viral replication and the host immune response 
in pediatric patients may differ from that observed in adults, hence extrapolation of efficacy may 
not be possible in all scenarios and could be considered on a case-by-case basis depending on the 
mechanism of action of the investigational drug or drugs and the pediatric age group being 
evaluated. 
 
In cases in which extrapolation of efficacy from adults to pediatrics is possible, after critical PK 
parameters of a drug are identified in adults, pediatric development programs can focus on 
identifying dosing regimen(s) that achieve pediatric exposures similar to adults to demonstrate 
effectiveness in pediatric populations in which treatment is indicated as per current treatment 
guidelines.  Additional clinical data should be obtained to assess whether antiviral activity is 
comparable to that observed in adult trials.  Early discussion with the FDA on the appropriate 
endpoints and pediatric trial design is encouraged. 
 
 
16 See the draft guidance for industry Chronic Hepatitis D Virus Infection:  Developing Drugs for Treatment 
(November 2019).  When final, this guidance will represent the FDAs current thinking on this topic. 
 
17 See section 505B(a)(1)(A) of the FD&C Act; 21 U.S.C. 355c(a)(1)(A).  
 
18 See section 505B(a)(5) of the FD&C Act.  
 
19 See section 505B(e)(2)(A)(ii) of the FD&C Act; see also the guidance for industry Pediatric Study Plans:  
Content of and Process for Submitting Initial Pediatric Study Plans and Amended Initial Pediatric Study Plans (July 
2020).  
 
13  Contains Nonbinding Recommendations 
In the absence of a serious safety signal in adults, sponsors should enroll adolescents20 in phase 3 
adult trials or conduct a dedicated adolescent trial concurrently with the phase 3 adult trials.  
Confirmatory PK, safety, and efficacy data from this age group should be included as part of the 
data included at the time of filing of the original NDA/BLA.21   
  
Typically, the nonadolescent pediatric population (for the purpose of this guidance, 2 to younger 
than 12 years of age) is divided into several groups or cohorts according to age or weight for 
enrollment into trials.  Weight, rather than age, is the preferred criterion for enrollment because 
dosing recommendations for most antiviral drugs are weight-based.  In addition, within clinical 
studies, sponsors should enroll the cohorts in parallel rather than in series, unless a drug has a 
specific safety or drug disposition factor that warrants a different approach.  Dose selection for 
pediatric treatment trials (e.g., phase 2 or 3) should be supported by PK data generated from the 
initial PK/safety phase of the pediatric study and results of available modeling and simulation.  
Sponsors should discuss the dose selection with the FDA before initiating the phase 2 or 3 
pediatric trials.    
  
The pediatric trials should also obtain data to support safety in pediatric populations.  In general, 
a safety database of about 100 subjects receiving the proposed dose and treatment duration is 
recommended.  The participants should be adequately distributed across the pediatric age group 
for which studies are required.  Long-term follow-up data should be collected from pediatric 
trials to assess long-term safety as well as durability of treatment response.  If clinical trials in 
adults have demonstrated differences in safety profile or dosing based on fibrosis stage, pediatric 
subjects should be assessed for presence or absence of cirrhosis using the most appropriate 
modality for each study location.    
  
Section 505B of the FD&C Act also mandates that the requisite pediatric assessments be 
conducted using a formulation of the drug that is appropriate for each pediatric group being 
studied.22  Adult formulations generally are considered appropriate for adolescent patients 
(Momper et al. 2013), but younger patients, who may not be able to swallow pills, may require 
different formulations.  Therefore, pediatric formulation development should begin as early as 
possible to enable the development of appropriate pediatric formulations of investigational drugs.  
 
6.  Dose Selection 
 
The results from proof-of-concept antiviral activity trials can be used to guide the selection of 
doses to be evaluated in subsequent trials.  Sponsors are encouraged to use various quantitative 
clinical pharmacology approaches that leverage the totality of available information on exposure 
and pharmacodynamic assessments such as biomarkers that measure target engagement, HBsAg 
 
20 For the purpose of this guidance, adolescents are defined as age 12 to younger than 18 years of age. 
 
21 We note that, for applications to which section 505B applies, all pediatric assessments must be submitted with the 
application unless those assessments have been deferred (section 505B(a)(1)(A)).  
 
22 See section 505B(a)(2)A) of the FD&C Act.  
 
14  Contains Nonbinding Recommendations 
decline, HBV DNA decline, HBV RNA decline, and HBcrAg decline to select doses for phase 3 
trials.   
 
7.  Efficacy Endpoints 
 
New finite duration therapies could be evaluated in clinical trials using any of the following 
efficacy endpoints:   
  
  Sustained suppression (6 months or longer) of HBV DNA (less than LLOQ, TD, or TND) 
off-treatment after a finite duration of therapy   
  
  Sustained suppression (6 months or longer) of HBV DNA (less than LLOQ, TD, or TND) 
off-treatment with HBsAg loss (less than 0.05 international unit/milliliter (IU/mL)) with 
or without HBsAb seroconversion after a finite duration of therapy    
 
At present, utility of reduction in HBsAg from baseline (without complete clearance) for 
assessing response to CHB therapies is unclear because of inconsistent correlations between 
qHBsAg and clinical response (Hu et al. 2018; Thompson et al. 2010; Chan et al. 2011).    
 
A limited number of secondary endpoints (e.g., HBeAg loss, anti-HBe seroconversion in HBeAg 
positive patients, ALT normalization) should be considered for testing using appropriate 
statistical methods for multiplicity.  Biochemical serum markers such as ALT values vary 
between laboratories, and lack of normalization of ALT may often be confounded by the 
presence of other chronic liver diseases such as nonalcoholic fatty liver disease.  
 
Other important endpoints:  Assessing progression of liver disease  
 
Except for patients with advanced or decompensated cirrhosis, a statistically rigorous evaluation 
of endpoints of liver progression can be challenging because these events occur infrequently 
until late in the course of CHB.  However, treatment effects on these endpoints provide useful 
clinical information, and trials evaluating them could be used to support an expanded indication 
or patient population and could be summarized in appropriate sections of the label.   
 
Some of the parameters or clinical outcomes that sponsors can consider include the following:  
 
  Change in Model for End Stage Liver Disease scores  
  Change in Child-Turcotte-Pugh scores  
  Progression to liver failure requiring transplantation or resulting in death  
  Occurrence of HCC  
  
Treatment-related regression of fibrosis or cirrhosis, as assessed by liver biopsy or noninvasive 
methods, may be appropriate for display in the label.  However, sponsors should discuss with the 
Division plans for labeling and performance characteristics of the noninvasive modality when 
protocols evaluating these endpoints are being designed.  
 
15  Contains Nonbinding Recommendations 
Patient-Reported Outcome Measures 
 
Sponsors who are considering incorporating patient-reported outcome measures23 as endpoints in 
clinical trials to measure treatment benefit should discuss patient-reported outcome instruments 
with the Agency during the clinical development process. 
 
8.  Trial Procedures and Timing of Assessments 
 
Biochemical, serological, virological, and histological endpoints can be used to assess the 
effectiveness of therapy.  For drugs with finite treatment durations, the optimal time point to 
assess the primary efficacy endpoint of sustained virologic response is 6 months or longer after 
cessation of therapy.  Additionally, the most appropriate time point to assess efficacy endpoints 
depends on the mechanism of action and half-life of the drug.  Longer term follow-up may be 
useful to confirm durability of treatment response and to measure clinical outcomes.  
 
9.  Statistical Considerations 
 
In general, a detailed protocol and SAP stating the trial hypotheses, analysis methods, and all 
other relevant details should be provided to the Division of Antivirals before trial initiation.  To 
improve the precision of treatment effect estimation and inference, sponsors should consider 
adjusting for prespecified baseline factors that are anticipated to be prognostic of the outcome 
(e.g., HBeAg status, HBV DNA level, and HBsAg level).  If randomization is stratified by 
baseline covariates, the analysis should account for the stratification factors.  A discussion of 
how the analyses will account for the stratified randomization should be included in the protocol.  
For details see the draft guidance for industry Adjusting for Covariates in Randomized Clinical 
Trials for Drugs and Biological Products (May 2021).24  For statistical analysis methods and 
issues, see the guidances for industry Providing Clinical Evidence of Effectiveness for Human 
Drug and Biological Products (May 1998), Non-Inferiority Clinical Trials to Establish 
Effectiveness (November 2016), and Adaptive Designs for Clinical Trials of Drugs and Biologics 
and the FDA white paper Statistical Considerations on Subgroup Analysis in Clinical Trials 
(Alosh et al. 2015).    
 
a.  Analysis populations   
 
All subjects who are randomized and received at least one dose of assigned therapy during the 
trial should be included in the primary efficacy analysis for all fully blinded trials.  For 
unblinded/partially blinded trials, all randomized subjects should be considered.  Any possibility 
of randomized subjects who do not receive treatment in either or both arms should be minimized.   
 
 
23 See the guidance for industry Patient-Reported Outcome Measures:  Use in Medical Product Development to 
Support Labeling Claims (December 2009). 
 
24 When final, this guidance will represent the FDAs current thinking on this topic. 
16  Contains Nonbinding Recommendations 
b.  Efficacy analyses 
 
The primary analysis should compare the proportion of responders across trial treatment arms.  
This analysis determines whether effectiveness has been demonstrated.   
  
For subgroup analyses, the analysis of the primary efficacy endpoint should be performed within 
important demographic and baseline characteristics (e.g., geographic region, sex, race, age 
group, HBV genotype, HBeAg status, screening HBV DNA, baseline weight, body mass index, 
baseline ALT, baseline fibrosis/cirrhosis, and (if applicable) prior response to previous treatment 
regimens).  The purpose of these analyses is to explore the consistency of the primary efficacy 
endpoint result across these subgroups.    
 
Treatment-by-region interaction should be investigated and reported to assess consistency of the 
efficacy results.  Treatment-by-HBeAg status interaction should also be investigated if HBeAg-
positive and HBeAg-negative subjects are enrolled in the trial.  
 
c.  Handling missing data 
 
Sponsors should make every attempt to limit discontinuation of subjects from the trial.  When the 
loss is unavoidable, sponsors should explain the causes of missing data and attempt to determine 
the final status of a subject who does not complete the protocol.  Analyses excluding subjects 
with missing data or other posttreatment outcomes can be biased because subjects who do not 
complete the trial may differ substantially in both measured and unmeasured ways compared 
with subjects who remain in the trial.  The primary method of handling missing data in the 
analysis should be prespecified in the protocol and SAP.  Sensitivity analyses should 
demonstrate that the primary analysis results are robust to the assumptions regarding missing 
data.    
 
10.  Accelerated Approval (Subpart H/I) Considerations 
 
For CHB, HBV DNA suppression with or without HBsAg loss is considered a validated 
surrogate endpoint that has been demonstrated to predict clinical outcomes, and this endpoint 
could be used to support a traditional approval.  Sponsors should discuss with the FDA plans to 
use any other surrogate endpoints that are reasonably likely to predict clinical benefit to support 
accelerated approval.25  Accelerated approval is subject to the requirement that postmarketing 
confirmatory trials are conducted to verify and describe the anticipated effect on irreversible 
morbidity or mortality or other clinical benefit.26  
 
11.  Benefit-Risk Considerations 
  
A thorough and comprehensive benefit-risk assessment ensures that the benefits outweigh 
potential risks to the intended population.  A benefit-risk assessment takes into consideration the 
 
25 See section 506(c) of the FD&C Act; 21 CFR part 314, subpart H; 21 CFR part 601, subpart E.    
  
26 See 21 CFR 314.510; 21 CFR 601.41.  
17  Contains Nonbinding Recommendations 
demonstrated therapeutic effect of the new drug and observed safety profile in the context of 
underlying disease and current treatment options available for the indication.    
 
C.  Other Considerations  
 
1.  Clinical Virology Considerations 
 
Samples for HBV quantification, genotypic, and phenotypic analysis should be obtained at 
different time points during treatment and follow-up.  Timing of sample collection should be 
based on initial observations of potency and on-treatment and off-treatment durability.  The 
genotypes and phenotypes of baseline and virologic failure isolates should be determined 
(virologic failure defined as a confirmed increase of greater than or equal to 1 log  HBV DNA 
10
copies/mL above nadir, quantifiable HBV DNA of greater than or equal to 1 log  copies/mL 
10
above LLOQ after being less than LLOQ, or never achieved HBV DNA levels less than LLOQ 
for two consecutive visits).  Transient increases in HBV DNA after the completion of treatment 
with a finite regimen may occur before sustained suppression.  A strategy for distinguishing 
treatment failures from subjects experiencing a temporary increase should be agreed upon with 
the Division during protocol development. 
 
Clinical assessment during the development of activators of the immune system is likely to be 
challenging, given that the greatest impact of these types of drugs will likely be a reduction in 
infected cells resulting in depletion of the cccDNA reservoir.  Complete depletion of the 
cccDNA reservoir to below the limit of detection may take a long time and will vary depending 
on the mechanism of action of the drug.  Currently, the only biomarker sufficient to predict a 
sustained response off-treatment is HBsAg loss, the assessment of which may be complicated by 
HBsAg expressed from integrated HBV DNA genome fragments.  In addition to host immune 
markers and serum HBV DNA, clinical trial protocols should assess several exploratory HBV 
endpoints (HBcrAg, HBV RNA, etc.) early in the development program in an attempt to identify 
potential markers that correlate with the drugs activity and may predict response to therapy.  
 
Genotypes of baseline and on-therapy virologic failure isolates should be compared and newly 
emerged drug resistance-associated substitutions/mutations should be identified.  HBV DNA 
from subjects with genotypic resistance to the investigational drug should be cloned in an HBV 
genome background and susceptibility to the investigational drug should be determined.  
 
  There are 10 recognized HBV genotypes (genotypes A through J) as well as subtypes 
identified for genotypes A through F.  The different HBV genotypes/subtypes encode 
distinct viral proteins and may exhibit differential responses to an investigational drug, 
which could confound efficacy results in clinical trials if the drug is only effective against 
some genotypes/subtypes (Congly et al. 2013).  Therefore, we recommend determining 
the genotypes/subtypes of HBV infection present at baseline to determine if the 
investigational drug exhibits antiviral activity against all HBV genotypes/subtypes.  For 
subjects with suppressed HBV DNA at baseline, historical genotype assessments may be 
used if available.  The assay, with performance characteristics, used to genotype the HBV 
samples in enrolled subjects should be included with the clinical trial protocol.  It may be 
important to confirm the genotype/subtype by phylogenetic analysis.  
18  Contains Nonbinding Recommendations 
  
  For resistance analyses, any changes, including mixtures, in the amino acid sequence of 
the target protein or DNA sequence for genome targeting drugs, present in on-treatment 
or follow-up samples but not in the baseline sample, can be reported as having developed 
during therapy.  In addition, baseline samples should be analyzed to identify HBV 
genetic polymorphisms that are associated with differential antiviral activity against the 
investigational drug.  Sponsors should consult the FDA early for the most current format 
for submission of resistance data and if Next Generation Sequencing (NGS) will be used.  
  
  There is a risk of developing resistance against an antiviral drug that targets similar viral 
proteins in different virus species in patients coinfected with HIV and HBV.  Because of 
this risk, we recommend assessing for the development of resistance and cross-resistance 
in the viral proteins of both HIV-1 and HBV when appropriate.  
  
  For all virologic assessments in clinical trials, we recommend the use of FDA-approved 
or FDA-cleared assays, when available, and a central laboratory.  Sponsors can collect 
results from local lab tests, identifying the assays used.  If investigational assays are used, 
performance characteristics of the assays determined from analytical validation studies 
using geographically and temporally distinct isolates should be provided in addition to 
detailed descriptions of the methodology.27  Drugs that require assays to identify the 
infected population benefiting from treatment (e.g., specific genotypes or resistant 
populations) may require a companion diagnostic.  Additional recommendations can be 
found in the draft guidance for industry and Food and Drug Administration staff 
Principles for Codevelopment of an In Vitro Companion Diagnostic Device with a 
Therapeutic Product (July 2016).28  
  
  Sponsors are encouraged to submit a resistance monitoring plan early in development.  If 
resistance evaluation in clinical trials involves NGS, we recommend that sponsors discuss 
details of the NGS approach with the FDA.  Submission of NGS data in FASTQ format 
is strongly encouraged.  Additional recommendations can be found in the guidance for 
industry technical specifications document Submitting Next Generation Sequencing Data 
to the Division of Antiviral Products (July 2019). 
  
  HBV should be genotyped for any instances where HBV DNA is detected in long-term 
follow-up to distinguish relapse from reinfection.   
 
 
27 See the IDE (Investigational Device Exemption) web page, available at 
https://www.fda.gov/MedicalDevices/DeviceRegulationandGuidance/HowtoMarketYourDevice/InvestigationalDevi
ceExemptionIDE/ucm046164.htm.   
  
28 When final, this guidance will represent the FDAs current thinking on this topic.   
 
19  Contains Nonbinding Recommendations 
2.  Pharmacokinetic/Pharmacodynamic Considerations  
 
Trials conducted in HBV-infected subjects should include assessment of PK and the relationship 
between drug exposure (e.g., minimum or maximum plasma concentration (C  or C ), area 
min max
under the curve) and virologic success and toxicity in all subjects.  
 
Sponsors can use a combination of intensive and sparse sampling throughout development to 
characterize the PK of the investigational drug.  For example, sponsors should implement an 
intensive sampling schedule in early phase monotherapy trials.  In longer term trials, an intensive 
sampling schedule might not be feasible.  Alternatively, sponsors can combine sparse sampling 
from these trials with intensive PK data from earlier trials for population PK analysis.  Sponsors 
should obtain multiple sparse PK samples from as many subjects as possible, including at the 
time of key virologic assessments.  It is important to document dosing times and plasma 
sampling times.  
 
Sponsors can use the following two broad approaches to characterize the relationship between 
drug exposure and viral kinetics or virologic suppression of the investigational drug, depending 
on the development stage and purpose of the analysis.  Both approaches allow for exploration of 
relevant covariates.  
 
(1) To aid the design of phase 2b or phase 3 trials, with respect to selection of the dosage 
regimen, a mechanistic approach relating drug concentrations and viral kinetics should be 
considered.  A mechanistic modeling approach should also account for the development 
of resistance to the investigational drug and the intended patient population.  For 
combination therapy, the potential of additive or synergistic antiviral effects can be 
incorporated in the model to assist optimization of the dose combination.  
 
(2) A simplified analysis relating the proportion of subjects with virologic suppression or 
virologic failure and appropriate exposure variable (e.g., minimum concentration or area 
under the plasma drug concentration versus time curve) can be used to support evidence 
of activity and to support dose selection.   
 
Exposure-response safety analyses should consider the mechanistic on-target and off-target effects 
of the investigational drug and adverse events that are more frequent in the investigational drug 
arm.  The appropriate exposure parameter and modeling approach depends on the investigational 
drug and toxicity.  
 
3.  Labeling Considerations  
 
Severe acute exacerbations of CHB may occur after discontinuation of anti-HBV therapy.  
Hepatic function should be monitored closely with both clinical and laboratory follow-up for at 
least several months in subjects who discontinue anti-HBV therapy.  In certain circumstances, 
resumption of anti-HBV therapy may be warranted.  These concerns should be adequately 
conveyed in drug labeling.  
 
20  Contains Nonbinding Recommendations 
Development of HIV-1 resistance against anti-HBV drugs with activity against HIV-1 is a 
potential risk that should be conveyed in labeling.    
    
21  Contains Nonbinding Recommendations 
GLOSSARY OF ACRONYMS  
 
ALT      alanine aminotransferase  
CC       concentration inhibiting 50 percent cell growth  
50
cccDNA   covalently closed circular DNA  
CHB    chronic hepatitis B  
EC50/90    effective drug concentration inhibiting 50 or 90 percent virus replication  
FD&C Act   Federal Food, Drug, and Cosmetic Act  
HBeAg   HBV e-antigen  
HBsAb   antibody specific to HBsAg  
HBsAg   HBV surface antigen  
HBV    hepatitis B virus  
HBV DNA   hepatitis B virus DNA  
HCC    hepatocellular carcinoma  
HDV    hepatitis delta virus  
HIV      human immunodeficiency virus  
IFN      interferon  
IgM      immunoglobulin M  
IU      international unit  
LLOQ    lower limit of quantification  
mL  milliliter  
NRTI     HIV nucleoside/nucleotide reverse transcriptase inhibitor 
NrtI      HBV nucleoside/nucleotide reverse transcriptase inhibitor  
NGS      Next Generation Sequencing  
NI      noninferiority  
PK      pharmacokinetic  
qHBsAg   quantitative HBsAg  
RNA    ribonucleic acid  
rt      reverse transcriptase  
SAP  statistical analysis plan  
TD      target detected 
TND    target not detected  
    
   
22  Contains Nonbinding Recommendations 
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