Validation strategies and which parameters to focus on€¦ · −ICH guideline Q2(R1) does not...
Transcript of Validation strategies and which parameters to focus on€¦ · −ICH guideline Q2(R1) does not...
CE Pharm 2018
Dr. Angelina Rafai, PD Dr. Maria Schwarz
12 September 2018
Validation strategies and which parameters to focus onSpecial characteristics of area percentage test methods
Introduction
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Method feasibility/
setup
Method development
Method
Qualification / Validation
Method performance verification
Method design
− Method purpose
− Separation mode
− Impurity identification
− Availability of impurities
− Evaluation of method characteristics (e.g.
precision, robustness)
− Future specifications?
− Can method meet these specifications?
� Preparation of method validation!
Routine analysis
− Stability
− Release
Change
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Validation of Analytical Procedures: Text and Methodology
ICH Q2(R1) - Analytical Validation
Type of testmethod
Identification Testing for impurities Assay
Characteristic Quantitative Limit
Specificity + + + +
Precision
Repeatability
Intermediate
precision
LOD
LOQ
Linearity
Accuracy
Range +
Robustness + +
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Quantitative methods
(response ~ conc.)
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Test methods in biopharmaceutical analysis
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Size variants
Variants differing
in hydrophobicity
Charge variants
fragments
aggregates
chemical modifications
deamidation
AA modificationssialylation
glycosylation
glycation
oxidation
peptide mapping
SEC
CGE
RP-HPLC/
HIC
HILIC
CZE/cIEF
IEX
Relative (e.g. area%)
Method characteristics
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Relative methods (e.g. area%)
− Response (e.g. area%) not related to
the test sample concentration
− Response related to relative content of
each component
− Extinction coefficient known for entire
protein solution, not for each single
component
− Protein variants might differ in
extinction coefficient (depending on
modification)
Quantitative methods
− Response (e.g. area) related to
concentration
− Extinction coefficient known
− All variants are identified and
characterized
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Test methods in biopharmaceutical analysis
Relative (e.g. area%)
Quantitative
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Common procedure for relative methods (e.g. area%)
Analytical Validation
Angelina Rafai | CE Pharm 2018
Type of testmethod
Identification Testing for impurities AssayPurity/Impurity
based on e.g. area%
Characteristic Quantitative Limit
Specificity
Precision
Repeatability
Intermediate
precision
LOD
LOQ
Linearity
Accuracy
Range
Robustness
ICH guideline does not cover
relative (area%) methods
(biopharmaceuticals)
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Common procedure for relative methods (e.g. area%)
Analytical Validation
Angelina Rafai | CE Pharm 2018
Type of testmethod
Identification Testing for impurities AssayPurity/Impurity
based on e.g. area%
Characteristic Quantitative Limit
Specificity
Precision
Repeatability
Intermediate
precision
LOD
LOQ
Linearity
Accuracy
Range
Robustness
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Common procedure for relative methods (e.g. area%)
Analytical Validation
Angelina Rafai | CE Pharm 2018
Type of testmethod
Identification Testing for impurities AssayPurity/Impurity
based on e.g. area%
Characteristic Quantitative Limit
Specificity Interfering signals
Precision
Repeatability RSD (area%), n = 6
Intermediate
precisionRSD (area%), n = 9
LOD S/N ≥ 3
LOQ S/N ≥ 10, RSD (area)
Linearity LOQ – 120 % (area)
Accuracy Recovery/Diff (area%)
RangeDeduced from Spec,
Lin, Prec
Robustness Recovery/Diff (area%)
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Common procedure for relative methods (e.g. area%)
Analytical Validation
Angelina Rafai | CE Pharm 2018
Type of testmethod
Identification Testing for impurities AssayPurity/Impurity
based on e.g. area%
Characteristic Quantitative Limit
Specificity Interfering signals
Precision
Repeatability RSD (area%), n = 6
Intermediate
precisionRSD (area%), n = 9
LOD S/N ≥ 3
LOQ S/N ≥ 10, RSD (area)
Linearity LOQ – 120 % (area)
Accuracy Recovery/Diff (area%)
RangeDeduced from Spec,
Lin, Prec
Robustness Recovery/Diff (area%)
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Suggested procedure for relative methods (e.g. area%)
Analytical Validation
Angelina Rafai | CE Pharm 2018
Type of testmethod
Identification Testing for impurities AssayPurity/Impurity
based on e.g. area%
Characteristic Quantitative Limit
Specificity
Precision
Repeatability
Intermediate
precision
LOD
LOQ
Linearity
Accuracy
Range
Robustness
− Sample dilution to LOQ level (e.g. 0.1 %)
− Evaluation of RSD for area (Main variant)
− Evaluation of S/N
− LOQ level is determined by serial dilutions (calculating the S/N)
− Determined LOQ is applied to test method (one LOQ for all variants)
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LOQ – common procedure and suggestion
suitable
signals for
LOQ
evaluation
− Undiluted test solution (100 % target load)
− Possibly using stressed material
− Differentiation between fragments/aggregates and acid/basic variants
− Evaluation of RSD for area% (suitable LOQ signals)
− Separate LOQs for different variants
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Linearity – common procedure and suggestion
− Sample dilution (LOQ – 150 % of target load)
− Evaluation of response (e.g. area)
− Linear plot %target load / response (e.g. area)
− Linear range of detector (also for relative methods)
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Linearity – suggestion
− Linear range of detector (also for relative methods)
− 50 – 150 % of target load
− If suitable material is available (purified variants)
− Differentiation between fragments/aggregates and acid/basic variants
− e.g. 60 – 120 % of specification
− Evaluation of residuals
− Deduce accuracy
Unspiked sample
Spiked sample
Accuracy - suggestion
Size-based separation (CGE, SEC)
− Origin and identity of impurities should
be clear
− Non-red: fragments
� spiking of reduced material
− Red: deglycosylated variants
� spiking of deglycosylated material
Charge-based separation (cIEF, CZE, IEX)
− Identity of impurities should be clear
− Charge variants may include fragments
(ε ≠ const.) and amino acid modifications(ε ≈ const.)
� spiking material with equal features
(produced material has to be
characterized first)
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Examples for mAb
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Summary
− Relative methods (e.g. area%) are not quantitative
− ICH guideline Q2(R1) does not cover relative test methods (e.g. area%)
− Validation parameters need to be aligned with analytical result (no evaluation of
absolute values, e.g. area)
− LOQ, linearity, and accuracy have to be addressed appropriately (extinction
coefficient)
Acknowledgements
− Solvias AG, Biopharma Department
− PD Dr. Maria Schwarz
− Dr. Alexander Beck
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Römerpark 2, 4303 Kaiseraugst, Switzerland
www.solvias.com
Angelina Rafai, [email protected], T +41 61 845 6501
Solvias AG