no two are ever the same: approaches to the nonclinical safety assessment of biologics (slides)
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Copyright © 2014 Covance. All Rights Reserved
NO TWO ARE THE SAME: APPROACHES TO THE NONCLINICAL SAFETY ASSESSMENT OF BIOLOGICS
General Considerations for Study DesignJanice Lansita, PhD, DABT, Senior Managing Scientist, ToxStrategies, Inc.
The Value of Understanding the Biologic for Non-GLP Candidate Optimization and Group Size Considerations for FIH GLP StudiesShawn Heidel, DVM, PhD, Executive Director, Global Lead Optimization and Program ManagementCovance
General Considerations for Study Design
Overview
Introduction to Biological Products (biologics)• Regulatory guidance ICH S6(R1), Preclinical Safety Evaluation of
Biotechnology-Derived Pharmaceuticals
Toxicology Study Design• Dose• Dose Selection• Recovery Duration
Species Selection
Assessment of Immunogenicity
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Definition of Biologics
Biologics – protein pharmaceuticals derived from living organisms including:
Humans, animals, plants, microorganisms
Biotechnology methods (recombinant DNA/cell culture technology)
• Recombinant human proteins (enzymes, cytokines, growth factors)
• Monoclonal antibodies (mAb)• Fusion proteins• Antibody-drug conjugates (mAb + cytotoxic)• Bispecifics• Biosimilars
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Small Molecules vs. Biologics
Property Small Molecules Biologics
Molecular weight
Small• < 1000 daltons
Large• ≥ 1000 daltons, e.g. mAb ≈ 150 kDA
Target specificity
Less (compared to biologics)• Toxicities generally non-specific/not related to target (“off-target toxicity”)
High target specificity• Toxicity generally related to target/pharmacology or “on-target toxicity”; expected
Species Specificity
Generally not a concern• 2 species (rodent; non-rodent)
May be a challenge• Only 1 animal species may be relevant
Half-life Short (compared to antibodies)• Minutes - hours - days
Long – especially molecules with Fc of IgG FcRn receptor, protects IgG from catabolism• Avastin® ≈ 20 days
Distribution Potential for extensive distribution within the body
More limited distribution within body • Initially, largely confined to vascular space
Immunogenicity Generally not a concern Common challenge in animals and humans
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Biologics Regulatory Guidance: ICH S6 Addendum/ICH S6(R1) Key Points
ICH S6(R1) (2011), Preclinical Safety Evaluation of Biotechnology-Derived Pharmaceuticals
Guidance addresses biologics-specific issues.
For today’s discussion, we will focus on:
Study design • Dose selection• Duration • Recovery
Species selection
Immunogenicity
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Toxicology Studies: Study Design Objectives
Dose Selection• High-dose: generally a dose that produces some toxicity. However, with
biologics, often no overt toxicity is observed.– From ICH S6(R1): (1) a dose that provides the maximum intended
pharmacological effect in the preclinical species and (2) a dose that provides an approximately 10-fold exposure multiple over the maximum exposure to be achieved in the clinic
• Mid-dose: generally an even multiple between the low and high dose.• Low-dose: generally a dose that results in a similar exposure as the
anticipated clinically efficacious dose.
Clinical Start Dose• Define a NOAEL (STD 10 or HNSTD for oncology products)• NOAEL used to define the starting dose for clinical trials• Clinical trial starting dose depends on overall risk vs. benefit
NOAEL = No observable adverse effect levelSTD 10 (rodents) = severely toxic dose in 10% of animals HNSTD (non-rodents) = highest non-severely toxic dose
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Toxicology Studies: Study Design Objectives
Duration• Duration should support clinical trial. For chronic toxicity testing, 6-month
duration generally adequate
Dose Schedule/Frequency • Mimic clinical trial • Example: if biologic is to be administered twice weekly in patients, it
should be administered twice weekly in animals• Recovery and Duration of Recovery Period: – Address reversibility not delayed toxicity
– Complete recovery not essential
– Recovery duration should consider T1/2 of biologic
– Biologic product may interfere with detection of ADAs
Key objective of toxicology study is to define
a safe clinical dose
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Toxicology Studies: Key Objectives
Identify target organ(s) of toxicity: • Related to pharmacological action of drug? • Dose-response and time-course • Biomarkers or clinical parameters
• Monitorable, manageable and/or predictable?• Mechanism of toxicity• Reversibility
Clinical Risk vs. Benefit Assessment • Patient population and indication • Mitigate risks with clinical monitoring, inclusion/exclusion criteria, smaller
N, more conservative dose escalation, e.g., waiting period between patients and dose cohorts
• Risk and benefit become more defined; however, rare events difficult to predict until post-marketing
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Species Specificity
Species specificity - specific activity in a pharmacologically relevant species
• Species in which biologic is pharmacologically active due to expression of the intended target (e.g., receptor or epitope)
• Rats and dogs standard for small molecules
• Biologics – specific to human target• Nonhuman primate may be only relevant species• Cynomolgus monkey most common
ICH S6(R1) – “Toxicity studies in nonrelevant species may be misleading and are discouraged.”
• Animal species that does not express the target• Unreliable safety data • Not predictive of potential human toxicity• Example: early biologics drug development of interferons in rodents
(nonrelevant species) vs. non-human primates (pharmacologically relevant species).
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Determination of a Pharmacologically Relevant Species
How to determine a pharmacologically relevant species?
Comparison of animal species vs. human:• Sequence homology
• Expression and distribution of target • Binding affinity • In vitro functional bioassay – e.g., neutralization/activation of target
– Cell proliferation, tyrosine kinase phosphorylation
• In vivo assay (if available)– E.g., serum marker of pharmacological activity
– Up-regulation/down-regulation of cytokine or
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Example of Data Used to Determine Pharmacologically Relevant Species
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Species Sequence Homology
Binding Affinity (pM)
In vitro Functional Activity IC50 (μM)
Human --- 160 10
Cynomolgus
Monkey98% 80 5
Rat 95% 140 15
Hamster 89% 1700 550
Mouse 87% 2000 980
Monkey & rat are pharmacologically relevant
Data set consists of: target sequence homology, binding affinity of mAb to target, and in vitro functional activity.
Immunogenicity
Immunogenicity is a common challenge for biologics and is a normal and expected response
• Animal’s immune system recognizes the biologic as foreign • Develops anti-drug antibodies (ADA) to eliminate from body
Measuring animal ADA responses in toxicology studies is necessary to aid in study interpretation
Primary concern for development of clearing and/or neutralizing ADA in toxicology studies:
• Lower exposure of target organs to the drug
• Fewer treatment-related toxicities or false-negative results
ADA responses in animals are not relevant to predict human ADA responses
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Immunogenicity: Various Types of ADA Responses Can Occur
Binding: minimal to no impact
Clearing: increase drug clearance (decrease exposure)
Sustaining: decrease drug clearance (increase exposure)
Neutralizing: interfere with biologic binding to the target
Cross-reactive: binds to biologic and endogenous protein• Neutralizes pharmacological activity of biologic and endogenous protein• Endogenous protein mediates a unique biological action – Toxicity may be a concern if function is neutralized
Erythropoietin (pure red cell aplasia)
Thrombopoietin (thrombocytopenia)
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Day 1 Day 28
Time (days)
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Time (days)
0 2 4 6 8 10 12 14
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Antibody positive
Clearing Antibody: Impact on Exposure
Antibody negative
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Immunogenicity: Take-home Messages
ADA responses can make long-term studies difficult
ADA responses do not necessarily invalidate a study• Sufficient number of animals exposed to active drug
Potential solutions: • Higher doses or more frequent dosing– “Saturate” or “dose through” the ADA response
– Sufficient drug “on board” to maintain systemic exposure
– Do not include low dose group if unlikely to maintain exposure throughout toxicology study
• Other approaches
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THE VALUE OF UNDERSTANDING THE BIOLOGIC FOR NON-GLP CANDIDATE OPTIMIZATION AND GROUP SIZE CONSIDERATIONS FOR FIH GLP STUDIES
Shawn Heidel, DVM, PhD Executive Director, Global Lead Optimization and Program ManagementCovance
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Goal for Candidate Optimization Studies (Non-GLP)
► Eliminate molecules with an unacceptable margin of safety
► Provide data to set doses for GLP toxicology studies
► Establish a dose response relationshipTarget saturation plays a key role
in dose selection for biologics
► Maximum saturation = maximum effect at the intended target
Toxicity associated with biopharmaceuticals is usually due to sustained, saturating pharmacology
Exceeding maximum saturation of the intended target can result in activating related targets
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Low Dose
Mid Dose
High Dose
Simple Pharmacodynmic (PD) Curve with Recommended Doses For GLP Studies
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Dose Selection
“…PK-PD…can assist in high dose selection by identifying 1) a dose which provides the maximum intended pharmacological effect in the preclinical species; and 2) a dose which provides an approximately 10-fold exposure multiple over the maximum exposure to be achieved in the clinic. The higher of these two doses should be chosen for the high dose group ....”
Note: It’s also important to ensure the low-dose isn’t overtly toxic and that the mid-dose identifies a dose response between the low and high doses.
FROM ICHS6(R1) SECTION 3.1 DOSE SELECTION AND APPLICATION OF PK/PD PRINCIPLES
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What Candidate Optimization Studies Should be Done?
Case #1 – only active in NHPs
No in vivo pharmacology data, because NHP are the only appropriate species and no pharmacology models exist in NHPs
► Sequence homology and in vitro binding completed
► Affinity for NHP target < 10-fold and rodent target > 30-fold compared to the human target
► No in vivo pharmacodynamic (PD) marker available
► No related targets that the biologic could bind to and cause toxicity
IT DEPENDS ON THE BIOLOGIC AND WHAT IS KNOWN
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What Candidate Optimization Studies Should be Done?
Case #1 continued
Reactive only in NHPs and target is not anticipated to have any toxicology findings, then conduct only a single dose Pharmacokinetic (PK) study► A good example is a anti-cytokine antibody to a cytokine that’s
expressed mostly during disease; anti-IL6 mAb
► Typically don’t need more than 3 NHPs/dose, since this is range finding
► Dose selection needs to start low to observe any target mediated clearance
► Usually a single dose, because of material requirements
► Collect clinical pathology and hematology to ensure no unanticipated toxicity
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Determining Target Saturation with PK Study
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Cynomolgus Monkey PK curve
Target-mediated CL
Normal antibody CL pathways after target is saturated
Target Saturation ~ 0.5 mg/kg
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NHPs the Only Appropriate Species
Reactive Only in NHPs and Target Activation Could Result in Toxicity, then NHP PK + Toxicology Study
► EGFR is also known as Erb-1 and belongs to a family of receptors that includes Erb-2, -3 and -4
► The family members can form homodimers or heterodimers with other members of the family
► Multiple ligands bind different receptors
► Binding to other receptors in the family may be intended for the desired pharmacological effect or unintended due to structural similarity across receptors in the family
► Very complex biology
ANTI-EGFR AS AN EXAMPLE
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Case #2
NHP = Non-Human Primate
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Complexity of EGF Receptor Biology
Ligand Receptor
ErbB-1 ErbB-2 ErbB-3 ErbB-4
EGF + - - -TGF-α + - - -HB-EGF + - - +
amphiregulin + - - -
betacellulin + - - +
epigen + - - -epiregulin + - - +
neuregulin 1 - - + +
neuregulin 2 - - + +
neuregulin 3 - - - +
neuregulin 4 - - - +
24 Table from Wikipedia
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NHPs the Only Appropriate Species
► EGFR is expressed on multiple normal tissues
Skin, adrenal, cecum, colon, heart, etc.
► EGFR is over-expressed on multiple tumor types
► Mouse xenograft models with different human tumors is a common pharmacology model
Each tumor responds to a different dose of anti-EGFR
Tumor responsiveness does not always correlate with EGFR expression level
ANTI-EGFR AS AN EXAMPLE
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It is difficult/impossible to use mouse xenograft data to model doses that may cause toxicity in monkeys
Case #2 – continued
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NHPs the Only Appropriate Species
► What doses should be selected for monkey toxicology studies?
► Does the monkey PK screen identify a dose that approximates target saturation?
► How much target occupancy is needed for toxicology?
► Given the complexity of the target biology, is it possible to estimate doses that have partial pharmacology, but not toxicity?
ANTI-EGFR AS AN EXAMPLE
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Case #2 – continued
Given the complexity of targets like EGFR, having a pharmacodynamic marker of target inhibition is very useful and perhaps critical
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What Can Happen if You Don’t Have Enough Data to Establish a Dose-Response?
► No dose-response!
► If target saturation is associated with toxicity, then the result is toxicity at all doses!
► What is a safe starting clinical dose?
► Clinical hold?
POSSIBLE TO SELECT ALL DOSES THAT SATURATE THE TARGET
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Low Dose
Mid DoseHigh Dose
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How to Utilize a Pharmacodynamic Marker to Set Doses ?► Concentrations of an anti-EGFR antibody in cynomolgus monkeys following a
single dose of 1, 10, or 100 mg/kg (not actual data)
► The effective concentrations that provide 100% or 50% target inhibition (TI) are shown (not actual data)
Time after dosing (hr)
0 10 20 30 40 50 60 70
Co
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Estimated TI 100
Estimated TI 50
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NHPs and Rodents, Toxicity Anticipated
Reactive in NHPs and Rodents, and is anticipated to have toxicity, then 14-day toxicity study in the rodent, 7 days in the non-rodent
►Establish target saturation in both species as in Case Study #1
►Pharmacology model(s), if available
Dose selection based on PK modeling and target saturation
Spread out the doses to cover the anticipated PKPD curve
►Use data from PK studies and pharmacology study(ies) to model doses for the non-GLP 14-day rodent and 7-day non-rodent toxicology studies
►Common to run 2 NHP/sex/gp for a “range finding” toxicology study
Typically no reversibility in Candidate Optimization studies unless a decision to discontinue development could be made
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Case #3
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Group Size Considerations for GLP Studies
► “It depends”
► Golimumab (humanized anti-TNF)Approved more than a decade after the first anti-TNF
mAb and therefore, not anticipating any serious findings
Only 2 dose groups in mouse and NHP studies
3 NHP/sex/group + 2/sex/group recovery
► When a robust data set exists, the study design can be minimal
► No absolute regulatory requirements for group size >3/sex/group
Most companies use 3-4/sex/group, a control and 3 dose groups
HOW MANY ANIMALS IS ENOUGH?
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Recovery Group Considerations
“Recovery from pharmacological and toxicological effects with potential adverse clinical impact should be understood when they occur at clinically relevant exposures. This information can be obtained by an understanding that the particular effect observed is generally reversible/non-reversible or by including a non-dosing period in at least one study, at least one dose level, to be justified by the sponsor. The purpose of the non-dosing period is to examine reversibility of these effects, not to assess delayed toxicity. The demonstration of complete recovery is not considered essential. The addition of a recovery period just to assess potential for immunogenicity is not required.”
RECOVERY: ICHS6(R1)
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Do not include recovery animals at this stage::
•Consider excluding recovery animals from initial studies•Only include recovery animals in later studies if there is an indication of need•Re-asseaa the need for recovery based on the outcome of the other studies i.e. once the toxicity profile has been determined How to include recovery animals:
Recovery Group ConsiderationsWHEN TO INCLUDE RECOVERY GROUPS AND HOW MANY ANIMALS?
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Sewell et al_Reg PharmTox_2014
When to include recovery animals:Are recovery animals necessary at this stage? i.e. does recovery need to be
assessed now, rather than in later studies, once more information is available?
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How Does this Compare to Actual Practice with FIH Studies for Biologics?
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Sewell et al_Reg PharmTox_2014
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Why Are Recovery Groups Usually Included for Biologics?
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Biologicals
Ranking Rationale for inclusion No. studies No. compounds No. companies
1234567891011
Company PracticePerceived Regulatory ExpectationBiotherapeuticLiterature/Known Class EffectsPrevious Internal Data on the CompoundPerceived First in ClassSignal from Prior In Vivo StudyOncology DrugClinical Request (internal or external)OtherFormal Regulatory Feedback
41363418131197221
(59%)(52%)(49%)(26%)(19%)(16%)(13%)(10%)(3%)(3%)(1%)
3125241511765211
(66%)(53%)(51%)(32%)(23%)(15%)(13%)(11%)(4%)(2%)(2%)
1161067554211
Sewell et al_Reg PharmTox_2014
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What are the Reasons for Not Including Recovery in Any Dose Group and the Impact?
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Biologicals
Ranking Rationale for exclusion No. studies No. compounds No. companies
123456
Company practice Signal (or lack thereof) from prior in vivo study Previous internal data on the compound No indication of need 5 Literature/known class effects (MOA) Other
11109981
(85%)(77%)(70%)(70%)(62%)(8%)
443331
(67%)(67%)(50%)(50%)(50%)(17%)
332211
Ranking Impact of exclusion No. studies No. compounds No. companies
1 None 13 (100%) 6 (100%) 4
Sewell et al_Reg PharmTox_2014
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Reversibility in All Dose Groups was Most Common. Consistent with “at clinically relevant exposures” from ICHS6(R1)?
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No
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Summary
► Consider each biologic and design the Candidate Optimization package of studies appropriately
► Establishing a pharmacodynamic curve is critical for selecting the appropriate doses for GLP studies
Dose selection must consider pharmacology and how that might relate to toxicity/adverse events
► Reversibility should be carefully considered at clinically relevant exposures
NO TWO ARE EVER THE SAME
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Let’s Start the Conversation
Shawn Heidel, DVM, PhD
Executive Director, Global Lead Optimization and Program Management
Covance
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