Download Degradation Studies

Practical Design and Application of Forced
Degradation Studies
Session 8
Steven S. Kuwahara, Ph.D.
GXP BioTechnology
6336 N. Oracle Rd. #326-313
Tucson, AZ 85704-5480
e-Mail: [email protected]
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What Do the Guidelines Say?
• Forced Degradation is also known as Stress Testing.
• Stress Testing is conducted under conditions that exceed
those employed in accelerated testing.
• Degradation Product (Defined in ICH Q5C) A molecule
resulting from a change in the drug substance (bulk
material) brought about over time. For the purpose of
stability testing of the products described in this guideline,
such changes could occur as a result of processing or
storage (e.g., by deamidation, oxidation, aggregation,
proteolysis). For biotechnological/biological products, some
degradation products may be active.
– Note that this refers to the API. For certain products, degradants
may arise from excipients or the container-closure system.
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Stress Testing (from Q1A(R2) 2.1 Drug Substance. I.
• 2.1.2. Stress Testing
• Stress testing of the drug substance can help identify the
likely degradation products, which can in turn help establish
the degradation pathways and the intrinsic stability of the
molecule and validate the stability indicating power of the
analytical procedures used. The nature of the stress testing
will depend on the individual drug substance and the type of
drug product involved.
• Stress testing is likely to be carried out on a single batch of
the drug substance. It should include the effect of
temperatures (in 10°C increments (e.g., 50°C, 60°C, etc.)
above that for accelerated testing), humidity (e.g., 75% RH
or greater) where appropriate, oxidation, and photolysis on
the drug substance.
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Stress Testing (from Q1A(R2)) 2.1 Drug Substance. II.
• The testing should also evaluate the susceptibility of the
drug substance to hydrolysis across a wide range of pH
values when in solution or suspension. Photostability testing
should be an integral part of stress testing. The standard
conditions for photostability testing are described in ICH
Q1B.
• Examining degradation products under stress conditions is
useful in establishing degradation pathways and developing
and validating suitable analytical procedures. However, it
may not be necessary to examine specifically for certain
degradation products if it has been demonstrated that they
are not formed under accelerated or long term storage
conditions. Results from these studies will form an integral
part of the information provided to regulatory authorities.
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Photostability Testing (from Q1B)
II. DRUG SUBSTANCE
• For drug substances, photostability testing should consist of
two parts: Forced degradation testing and confirmatory
testing.
• The purpose of forced degradation testing studies is to
evaluate the overall photosensitivity of the material for
method development purposes and/or degradation pathway
elucidation. This testing may involve the drug substance
alone and/or in simple solutions/suspensions to validate the
analytical procedures. In these studies, the samples should
be in chemically inert and transparent containers. In these
forced degradation studies, a variety of exposure conditions
may be used, depending on the photosensitivity of the drug
substance involved and the intensity of the light sources
used.
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Photostability Testing (from Q1B) II. DRUG SUBSTANCE. B.
• For development and validation purposes, it is appropriate
to limit exposure and end the studies if extensive
decomposition occurs. For photostable materials, studies
may be terminated after an appropriate exposure level has
been used. The design of these experiments is left to the
applicant's discretion although the exposure levels used
should be justified.
• Under forcing conditions, decomposition products may be
observed that are unlikely to be formed under the conditions
used for confirmatory studies. This information may be
useful in developing and validating suitable analytical
methods. If in practice it has been demonstrated they are not
formed in the confirmatory studies, these degradation
products need not be examined further.
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Photostability Testing (from Q1B) II. DRUG
SUBSTANCE. C.
• Confirmatory studies should then be undertaken to provide
the information necessary for handling, packaging, and
labeling (see section I.C., Procedure, and II.A., Presentation,
for information on the design of these studies).
• Normally, only one batch of drug substance is tested during
the development phase, and then the photostability
characteristics should be confirmed on a single batch
selected as described in the parent guideline if the drug is
clearly photostable or photolabile. If the results of the
confirmatory study are equivocal, testing of up to two
additional batches should be conducted. Samples should be
selected as described in the parent guideline.
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Photostability Testing (from Q1B) II. DRUG
SUBSTANCE. D.
• C. Judgment of Results
• The forced degradation studies should be designed to
provide suitable information to develop and validate test
methods for the confirmatory studies. These test methods
should be capable of resolving and detecting photolytic
degradants that appear during the confirmatory studies.
When evaluating the results of these studies, it is important
to recognize that they form part of the stress testing and are
not therefore designed to establish qualitative or quantitative
limits for change.
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Photostability Testing (from Q1B) II. DRUG
SUBSTANCE. E.
• V. GLOSSARY
• Forced degradation testing studies are those undertaken to
degrade the sample deliberately. These studies, which may
be undertaken in the development phase normally on the
drug substances, are used to evaluate the overall
photosensitivity of the material for method development
purposes and/or degradation pathway elucidation.
•
Confirmatory studies are those undertaken to establish photostability
characteristics under standardized conditions. These studies are used to
identify precautionary measures needed in manufacturing or formulation
and whether light-resistant packaging and/or special labeling is needed
to mitigate exposure to light. For the confirmatory studies, the batch(es)
should be selected according to batch selection for long-term and
accelerated testing which is described in the parent guideline.
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Stress Testing (from Q1A(R2) Other Statements
• Specification, which is a list of tests, reference to analytical
procedures, and proposed acceptance criteria, is addressed
in ICH Q6A and Q6B. In addition, specification for
degradation products in a drug substance is discussed in
Q3A.
• 2.2.2. Photostability Testing
• Photostability testing should be conducted on at least one
primary batch of the drug product if appropriate. The
standard conditions for photostability testing are described
in ICH Q1B.
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What About Biologics?
• Guideline for Industry
• Quality of Biotechnological Products: Stability Testing
of Biotechnological/Biological Products
• July 1996, ICH Q5C
•
I. INTRODUCTION (1)
• The guidance stated in the ICH harmonized tripartite
guideline entitled “Stability Testing of New Drug
Substances and Products” (ICH Q1A) applies in general to
biotechnological/biological products. However,
biotechnological/biological products have distinguishing
characteristics to which consideration should be given in
any well-defined testing program designed to confirm their
stability during the intended storage period.
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ICH Q5C: A.
• For such products in which the active components
are typically proteins and/or polypeptides,
maintenance of molecular conformation and, hence,
of biological activity, is dependent on noncovalent
as well as covalent forces. The products are
particularly sensitive to environmental factors such
as temperature changes, oxidation, light, ionic
content, and shear. To ensure maintenance of
biological activity and to avoid degradation,
stringent conditions for their storage are usually
necessary.
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ICH Q5C: B.: B. Potency (5.2)
•
•
•
•
When the intended use of a product is linked to a definable and
measurable biological activity, testing for potency should be part of the
stability studies.
For the purpose of stability testing of the products described in this
guideline, potency is the specific ability or capacity of a product to
achieve its intended effect. It is based on the measurement of some
attribute of the product and is determined by a suitable in vivo or in vitro
quantitative method.
In general, potencies of biotechnological/biological products tested by
different laboratories can be compared in a meaningful way only if
expressed in relation to that of an appropriate reference material.
For that purpose, a reference material calibrated directly or indirectly
against the corresponding national or international reference material
should be included in the assay.
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ICH Q5C: C.: C
• Wherever significant qualitative or quantitative
changes indicative of degradation product
formation are detected during long-term,
accelerated, and/or stress stability studies,
consideration should be given to potential hazards
and to the need for characterization and
quantification of degradation products within the
long-term stability program. Acceptable limits
should be proposed and justified, taking into
account the levels observed in material used in
preclinical and clinical studies.
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ICH Q5C: D: C. Accelerated and Stress Conditions
(6.3)
•
However, it is strongly suggested that studies be conducted on the drug
substance and drug product under accelerated and stress conditions.
Studies under accelerated conditions may provide useful support data
for establishing the expiration date, provide product stability information
or future product development (e.g., preliminary assessment of proposed
manufacturing changes such as change in formulation, scale-up), assist
in validation of analytical methods for the stability program, or generate
information that may help elucidate the degradation profile of the drug
substance or drug product. Studies under stress conditions may be useful
in determining whether accidental exposures to conditions other than
those proposed (e.g., during transportation) are deleterious to the
product and also for evaluating which specific test parameters may be
the best indicators of product stability. Studies of the exposure of the
drug substance or drug product to extreme conditions may help to reveal
patterns of degradation; if so, such changes should be monitored under
proposed storage conditions.
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ICH Q5C: E: Glossary
• Degradation Product
• A molecule resulting from a change in the drug
substance (bulk material) brought about over time.
For the purpose of stability testing of the products
described in this guideline (Biologics), such
changes could occur as a result of processing or
storage (e.g., by deamidation, oxidation,
aggregation, proteolysis).
• For biotechnological/biological products, some
degradation products may be active.
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What Are You Trying to Degrade? I.
• There is a need to understand the nature of the
product in the degradation study.
– The guidelines as originally written were for small
molecule drugs that degraded via simple one step
mechanisms with a defined activation energy.
– Newer drugs and biologics can degrade via multiple
pathways that may depend upon environmental
factors such as pH and temperature.
• Thus there may be several degradation pathways depending
one the environment.
• Some molecules, especially biologics, may have degradation
products that are, pharmacologically active.
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What Are You Trying to Degrade? II.
• In addition, the physical state of the molecule
can change.
– A molecule in a crystal may be susceptible to
different factor from a molecule in an oil or in water.
– Lyophilized products often degrade differently from
products after reconstitution.
• The stability of the pure drug in bulk may be
quite different from its stability in the final
formultation.
• YOU MUST UNDERSTAND THE
CHEMISTRY OF THE MOLECULE.
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Be Practical, but Also Sensitive to Nuances.
• Any organic molecule will degrade at 400o C in
air. An ester will hydrolyze in 5N acid or base.
– So if you show degradation under these conditions,
what’s the point?
– Autoclaving a 5% glucose solution can produce about
80 different products depending on pH.
• Some are secondary or even tertiary degradants.
• The problem here is that some degradants react with the
parent molecule or each other. The problem varies with the
concentration of the molecules and the time of exposure.
• You should design your study to see primary degradants,
but remember that some reactions may be fast.
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Designing the Study
• The idea of the degradation study is to provide
information on what degradants you may expect
to see in the course of the stability study.
– You especially want to observe the degradants that
may require a long time to become observable.
– Another type of important degradant would the type
that might develop during processing and
contaminate the product.
– You must account for the stoichiometry of the API
and the degradants.
• Otherwise you may miss the presence of a significant
degradant.
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General Studies. I.
• Generally, thermal degradation is studied at 60o C.;
acid/base sensitivity in 0.1 N HCl and NaOH; Redox
sensitivity in 0.1% H2O2, and 100 mM mercaptoethanol;
and photostability as given in ICH Q1B.
• However there are no hard and fast rules, and different
molecules and final products may be treated differently.
Companies are free to choose suitable methods.
• The idea is to see what types of products may form
under certain conditions, not to just degrade the
product.
• This will tell you what to look for or expect in the
product under various conditions.
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General Studies. II.
• One of the problems is that the degradation
studies are often conducted as “one factor at a
time” (OFAT) studies.
– The problem is that degradation may depend on a
combination of conditions.
• Sensitivity to oxidation may be pH dependent.
• Ionic strength may be a factor in stabilizing or denaturing a
protein.
• Some biologics may be contaminated with degrading
enzymes that are inactivated by pH extremes, but are active
under conditions near neutrality.
• The use of a designed, multifactor, experiment
should b considered.
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General Studies. III.
• Time is often a major factor, especially when
working out degradation pathways.
• Simply incubating a product for 10 min under a
given environmental condition is not enough.
– Slow-forming degradants may be missed in a short
incubation while long incubations may not find the
primary degradant and only find secondary or
tertiary degradants.
– Sampling intervals may be critical, depending on the
kinetics of the degradation reactions.
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Analytical Methods
• Test methods should be capable of detecting a
wide array of substances.
– Chromatographic, mass spectral, and U.V.-Vis and
I.R. spectral methods are common.
– More than one method should be used on any given
sample type, since each method has limitations.
– Test method sensitivity is important especially if
potential toxins are found.
– The test methods should be validated.
• If they are later, incorporated into stability studies, they will
need to be shown to be stability-indicating.
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The Five Types of Stability
Chemical: The API maintains chemical integrity and
labeled potency within specified limits.
Physical: The original properties including appearance,
palatability, uniformity, dissolution, and
suspendability are maintained.
Microbiological: Sterility or resistance to microbial growth
are retained according to specified requirements.
Antimicrobial agents that are present retain
effectiveness within specified limits.
Therapeutic: The therapeutic effect is unchanged.
Toxicological: No significant increase in toxicity occurs.
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Problems: Potency
• Since we are in the pharmaceutical industry,
the pharmacologic activity of a molecule is
important.
– Just because a molecule appears to be intact, it does
not mean that it is active. Even with small molecule
drugs, there can be small changes that seriously
affect biological activity.
– With amino acids and carbohydrates, simple changes from one
optical isomer to another can change biological activity.
– These changes may have very small activation energy barriers,
compared with the breakage of covalent bonds.
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Problems: Toxicity
• During forced degradation studies, toxic
products may be formed.
– In many cases, they may be the result of interactions
among or with excipients or the container-closure
system.
– These toxic products must be considered. They
should not be dismissed as unlikely to form under
normal conditions.
• There must be data showing that these products will not be
present under normal conditions.
– If a toxic degradant is detected through tissue culture
or mutagenicity tests, it should be identified.
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Biologic Stability
Complexities I.
• Activation energy changes: Hydrogen bond for
N – H····O=C is 4.76 kcal/mole in the gas phase,
but drops to 0.5 – 1.5 kcal/mole in water.
– Three dimensional configuration changes can occur
more rapidly in aqueous solutions than in the dried
state.
• The change is supposedly due to changes in the polarity in
the medium and entropy increases in water.
• Three dimensional configuration changes: A
macromolecule can unfold into many different
states, some of which are inactive.
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Biologic Stability
Complexities II.
• 3-D configuration problems: Macromolecules in
a random coil may be inactive or easily
degraded.
– Some proteases and nucleases prefer denatured
molecules.
– Unfolded molecules can still bind to target sites and
act as inhibitors.
– Natural molecules in unnatural configurations can be
highly antigenic and lead to antibody formation.
– If a patient is genetically deficient in a protein, a
therapeutic dose of the “natural” protein can cause
an immunologic response.
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Biologic Stability
Complexities III.
• PK/PD problems: Certain glyco-proteins,
especially those containing neuraminic acid
links, can have the side chain cleaved by mild
acid catalyzed hydrolysis or enzyme action in a
preparation. The resulting cleaved protein may
still have the correct potency but will be more
rapidly cleared from the circulation.
• Freeze-thaw problems: When a buffer-containing
solution is slowly frozen, at the last stages of freezing one
of the acid or base forms may freeze out faster than the
other. This can cause violent changes in the pH of the
solution.
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Biologic Stability
Complexities IV.
• Freeze-thaw problems: As a solution freezes, the
formation of the ice crystals can mechanically cause the
macromolecules to “stretch” or deform on the surface of
the crystals. This is similar to the foaming or bubble
problem. Where surface tension effects cause
deformation of macromolecules on the surface of the
bubbles and lead to denaturation.
• Solutions subjected to freeze-thaw or foaming may
initially appear to be clear, but will form particles
during long-tern storage.
• Freeze-thaw problems are usually minimized by rapid
freezing and rapid thawing.
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Biologic Stability
Complexities V.
• Turbulence problem: Vigorous mixing or
stirring of a solution containing macromolecules
can subject them to shear forces that result in
denatured material.
– Sometimes, vigorously sucking up a solution into a
pipette and strongly forcing the solution out can
cause a loss in activity or band (peak) spreading,
especially if this is done repeatedly.
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