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Connecting People, Science and Regulation ®
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Risk Assessment and DoE must be usedin Synergy for the success of QbD
Alain Poncin
Process Development Unit Manager
LFB Biotechnologies
Quality by Design, Frankfurt
Connecting People, Science and Regulation ®
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Quality by Design
� Science
� Statistics
� Risk Management
Knowledge
- of the productionprocess
- of the product
summarized in the RiskAssessment Report
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Risk Management� Technical report 42, PDA (2005)
� ICH Q8 : Pharmaceutical Development (2005) and annex(2007)
� ICH Q9 : Quality Risk Management (2005)
� ICH Q10 : Pharmaceutical Quality System (2008)
� ICH Q11 : Development and Manufacture of Drug Substance (concept paper, 2008, draft expected in 2009)
Focused on Product Quality
� ISO13485 : Medical devices – Quality management systems– Requirementsfor regulatory purposes
� ISO14971 : Medical devices – Application of Risk Management to medicaldevices
Focused on Product Quality + Product Availability
Pharma :
Compliance
Medical Device
Compliance
Economy
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Risk Management at LFB Biotechnology
WhenWhenStandard initial Risk analysis (FMEA) as soon as a first lab process gives
satisfactory resultsUpdated during product/process development, clinical development and post
approval.HowBased on standardised ‘’blocks’’ (Upstream, Harvest, Chromatography,
Ultra/diafiltration,…) ‘’Personnalised’’ using what is known : protein stability, ease/difficulty of steps,
occurrence of hasard,…
Ranks the work to be performed during development and process characterisationReflects and summarisesall what is known about the protein and the process
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Initial Risk analysis
1- Process Flow ChartPreculture Fermentation Harvest Filtration
Chromatography 1Viral inactivationChromatography 2
Ultrafiltration FiltrationFiltration
Vialing
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2- Table of content
Cleaning after purification5.6
Purification5.5
Sample preparation5.4
Storage of intermediates5.3
System assembly5.2
Cleaning beforechromatography
5.1
Capture of targetproduct after harvest
Capture by Chromatography
5
JustificationSub systemSystemN°
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3- Risk Analysis
� Based on previous experience, building of new blocks for thisfirst Risk Analysis at LFB Biotechnologies
� Copy and paste to a ‘’white’’ Risk Assessment
� Missing blocks (viral inactivation, nanofiltration,…) speciallyfor plasma product
� Not adapted to LFB Biotechnologies (history,…)
� Final analysis not homogeneous
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Lack of homogeneity:
1055Written SOP
Trained staff
3Human error7Contamination/loss of sample
Wrong buffer for equilibration
Ultra/
Diafiltration
6.8
243Trained staff
WrittenSOP/method of production
2Human error
(inversion of buffers)
4Contamination of targetproduct
Wrong buffer for equilibration/wash and or elution
Purification5.5
543Qualifiedequipment, preventivemaintenance
Trained staff
Calibration beforeuse
3Defectiveequipment (pH, scale,…)
6Ultra/diafiltration failure, contamination or degradationof targetproduct
Wrong buffer (pH, conductivity)
Buffer preparation
2.2
RPNDRisk controlPPossible causeSPossible Effect(harm)
Possible Hazard/
Failure
Product, Part, System, Fonction
N#
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To increase homogeneity and quality of RA:� Define risks
� For the Product safety/efficacy/availability-contamination (physical, chemical,…)-degradation (lower yield, production failure, immunogenicity)
� Identify risks for a general ‘’process’’
� Identify Possibles causes (human, material,…)
� Identify the measures to reduce the risk
� Adapt for each kind of process : fermentation, chromatography, ultrafiltration,…
Connecting People, Science and Regulation ®
10Storage of Matrix/column5.8
Cleaning after purification5.7
Product recovery5.6
Purification5.5
Sample preparation5.4
Storage and expiration time of intermediates
5.3
Cleaning beforechromatography
5.2
System assembly and calibration5.1
Capture of targetproduct after harvest
Capture by Chromatography
5
JustificationSub systemSystemN°
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2- Table of content
Membrane/carter Storage6.8
Cleaning afterultra/diafiltration
6.7
Product recovery6.6
Ultra/diafiltration6.5
Sample preparation6.4
Storage and expiration time of intermediates
6.3
Cleaning beforeultra/diafiltration
6.2
System assembly and calibration
6.1
Buffer ExchangeUltra/diafiltration6
JustificationSub systemSystemN°
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5- Quantification of Residual Risk
247
30
30
RPN
7
3
3
D
7
5
5
S
5
2
2
P
Identification of critical factors
Trained staff
WrittenSOP/method of production
Automation
Description/QC of raw material
Approvedsuppliers
Risk Control, Measures of Risk reduction, Tests
To be determinedContamination of Drug Product
Ineffective purification
Purification5.5.6
Human errorContamination of Drug product
Wrongbuffer (pH, conductivity)
Purification5.5.1
Reagentsidentity/Quality
Purification failure,
Production stopped
Wrongpreparation(saltaddition, …
Samplepreparation
5.4.5
Possible causePossible effect(harm) of the hazard/failure
Possible hazard/
failure
Product, Part, System, Function, Process
N#
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Risk Priority Number
� RPN = Severity x Probability x Detectability
� Require Internal Policy Definition� At LFB Biotechnology : 4 levels :
o 1 to 100 : broadly acceptable region o 101 to 150 : as low as reasonable practicable region (ALARP), part I o 151 to 250 : as low as reasonable practicable region (ALARP), par II
o 251 to 1000 : intolerable region
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6- Initial Risk Analysis conclusions
Final choice of filter type0.22 µm filtration8
Well known and controlled step, yet close to the optimum (load, wash and elution)
Chromatography7
Need first evaluation of viral clearance before First in Man
Viral Inactivation6
Not used in standard conditions, need identification of critical parameters to obtain a reproducibleprocess
Capture by Chromatography
5
Final choice of filter type0.22 µm filtration4
Need further comparability studies to assessstarting material equivalency
Clarification3
Current status/notesSystemStep
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� From initial Risk analysis : identification of capture purification step as a high risk Step
� To reduce this risk : identification of critical factor s and critical quality attributes.
Experience (int./ex.) Process development Initial Risk AssesmentPublication review First lab scale productionPatent review
Identification of critical factors by
DoE
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Particularity of DoE in Process Development
Protein folding : protein concentration, pH, Salt, Organic solvent,… 14 factors
Chromatography : pH and conductivity for equilibration and elution, sample load, matrix and
column resolution,… 10 factors
Ultrafiltration : pump speed, Pin, Pout, membrane, temperature, volume of buffer,…8 factors
Design space highly multidimensional
2 n experiments (full factorial) cannot be used,
2 n-k experiments (semi factorial) used for identification of critical factors
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Even 2 n-k experiments are a huge amount of work (process+ analytics)
Before starting experiments :
� Are Analytic tools sufficient ?
� Which Design : (semi) factorial ?
� Which factors to test ?
� Which Range (Design Space) ?
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Which Design ?
Full
Semi
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� Which factors and which Design space ?
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Experimental Work
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Output (Quality attributes)
� capture step : high yield, reproducible
Quantification of target protein biological activity
Quantification of total proteins
Purity by SDS-PAGE
On Load, Flow Through and Peak (23 samples)
Calculation of step yield and specific activity
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Results : usually nice graphics
Design-Expert® Software
Yield92
10
X1 = B: Conductivity sampleX2 = D: pH elution
Actual FactorsA: pH sample = 7.07C: Load = 30.00E: Gradient = 15.41
2.50
4.38
6.25
8.13
10.00
6.50
7.00
7.50
8.00
8.50
0
25
50
75
100
Y
ield
B: Conductivity sample D: pH elution
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But in another region of the space…
Design-Expert® Software
Yield92
10
X1 = B: Conductivity sampleX2 = D: pH elution
Actual FactorsA: pH sample = 7.07C: Load = 30.00E: Gradient = 15.41
2.50
4.38
6.25
8.13
10.00
6.50
7.00
7.50
8.00
8.50
0
25
50
75
100
Y
ield
B: Conductivity sample D: pH elution
Design-Expert® Software
Yield92
10
X1 = B: Conductivity sampleX2 = D: pH elution
Actual FactorsA: pH sample = 7.07C: Load = 10.00E: Gradient = 15.41
2.50
4.38
6.25
8.13
10.00
6.50
7.00
7.50
8.00
8.50
0
25
50
75
100
Y
ield
B: Conductivity sample D: pH elution
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Results : critical factors : half normal plot of effects
Design-Expert® SoftwareYield
Error from replicates
Shapiro-Wilk testW-value = 0.962p-value = 0.794A: pH sampleB: Conductivity sampleC: LoadD: pH elutionE: Gradient
Positive Effects Negative Effects
Half-Normal Plot
Ha
lf-N
orm
al
% P
rob
ab
ility
|Standardized Effect|
0.00 7.63 15.25 22.88 30.50
01020
30
50
70
80
90
95
99
A
C
E
- + Factor Effect = slope/2
E
A
C
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Or pareto chart
Design-Expert® SoftwareYield
A: pH sampleB: Conductivity sampleC: LoadD: pH elutionE: Gradient
Positive Effects Negative Effects
Pareto Chart
t-V
alu
e o
f |E
ffe
ct|
Rank
0.00
1.56
3.12
4.68
6.24
Bonf erroni Limit 4.38176
t-Value Limit 2.57058
1 2 3 4 5 6 7
E
A
C
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Are they significants ? ANOVA
ANOVA for selected factorial modelAnalysis of variance table [Partial sum of squares - Type III]
Sum of Mean F p-valueSource Squares df Square Value Prob > FModel 4129 3 1376 28,8 0.0014 A-pH sample 1301 1 1301 27,2 0.0034 C-Load 968 1 968 20,3 0.0064 E-Gradient 1861 1 1861 38,9 0.0015Curvature 960 1 960 20,1 0.0065Residual 239 5 48Lack of Fit 207 4 52 1,6 0.5244Pure Error 32 1 32Cor Total 5328 9
- +Factor
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ANOVA : Statistical analysis : can you do it ?
Design-Expert® SoftwareYield
Color points by value ofYield:
82
5
Internally Studentized Residuals
No
rma
l %
Pro
ba
bil
ity
Normal Plot of Residuals
-1.94 -0.97 0.00 0.97 1.94
1
5
10
20
30
50
70
80
90
95
99
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What is the value of the model obtained ?
Design-Expert® SoftwareYield
Color points by value ofYield:
82
5
2
Actual
Pre
dic
ted
Predicted vs. Actual
5.00
24.25
43.50
62.75
82.00
5.00 24.25 43.50 62.75 82.00
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Finally, capture step can be optimised
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Capture step by DoE and rangesDesign-Expert® Software
AS
Design Points
X1 = D: Conductivity
Actual FactorsA: Contact Time = 90B: pH load = 7.5C: Column Volume = 8E: Elution temperature = 20
20 25 30 35 40
8
25.75
43.5
61.25
79
D: Conductivity
AS
One Factor
25 + 5 mS/cm
NOR
PAR
Edge of failure
?
Definition of Criticals
Parameters Nice results isn’t ?
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But following purification results did not correlated wellwith prediction from this model.
A new analysis was performed with more powerfull tools.
Two main raisons were implied in the failure of the first model
� One critical factor not identified� High collinearity of some factors
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How many factors to select ?
Adjusted R-Squared or Mallows’CP statistics
Selection of number of factor
0
20
40
60
80
100
0 1 2 3 4 5
Number of factor
R S
quar
ed
0
500
1000
1500
2000
Cp
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High collinearity : regression by least square not efficient
-0.2684
-2.9287
Factor 3
-0.1870
-1.5614
Factor 2
0.67410.01 (ridge regression)
4.26370.0 (classical regression)
Factor 1Ridge parameter
Use of Ridge statistics
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Lack of detection of one critical factor
- +
FactorEffect = 0 D : pH elution
The new Model is predictible for the purifications performed (up to now)
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Finally, update of the initial Risk Analysis
2477Identification of critical factors
5To be determined7Contamination of Drug Product
Ineffective purification
Purification5.5.6
303Description/QC of raw material
Approvedsuppliers
2Reagentsidentity/Quality
5Purification failure,
Production stopped
Wrongpreparation(saltaddition, …
Samplepreparation
5.4
RPNDRisk Control, PPossible causeSPossible effectHazardSystem, N#
1205Qualification of equipment, preventivemaintenance
Trained staff
4Contact Time tooshort (< 2 min)
Other factorsunder control
6Contamination of Drug Product
Ineffective purification
Purification5.5.6
1255Quantification of proteolyticactivities
Written SOP
Automation
5Reagentsidentity/Quality
5Increase of proteolyticactivities
Production failure
Wrongpreparation(buffer addition)
Samplepreparation
5.4
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Synergy of Risk Management and DoEExperience (int./ex.) First lab scale Initial Risk AssesmentPublication review ProductionPatent review Process
Identification of criticalfactors by DoE
Risk Assesment Update
Process optimisation
Process capability indices Risk Assesment Update(Six Sigma)
Process Characterisation
Risk Assesment update
Process Validation
Statistics Risk Assesment update Phamacovigilance(trends,…)
Process Development
Phase I
Phase II/IIII
First in Man
GMP for Phase I
CTDPhase IV
Product discontinuation
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And in the future
� Increased number of experiments (// chromatography, 96 wells technology)
� Application to ultra/diafiltration
� Introduction of additional statistic tools (bayesianstatistics, Monte Carlo simulation, …)
� More applications of Six Sigma
� PAT
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Economical impact of QbD : example
Eurogentec s.a.
010
203040
5060
87 88 89 90 91 92 93 94 95 96 97 98 99-00
00-01
01-02
02-03
03-04
04-05
05-06
06-07
07-08
Turnover in millions € Nber of employees x 10