Andrew Lees PhD Scientific Director Fina BioSolutions LLC wwwFinaBiocom Fina Chemical drugs vs Biologicals Chemical drugs can be precisely defined Physical chemical characterization ID: 461281
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Slide1
Quality Control in Biotechnology
Andrew Lees, Ph.D
.
Scientific Director
Fina BioSolutions LLC
www.FinaBio.comSlide2
FinaSlide3
Chemical drugs vs
Biologicals
Chemical drugs can be precisely definedPhysical chemical characterization
NMR- structureMass Spectrometry- molecular weightChromatography- purity, quantityPotencyFormulation
Relatively easy to create “generics”Slide4
Chemical drugs vs
Biologicals
Biologicals are produced by living cellsImpossible to
control every variablecompletely characterizePrecisely replicateTraditionally,
biologicals are defined by “product by process”Product is defined by the manufacturing processQuality is compliance-driven
Goal is a “well-characterized biological”Slide5
c
urrent Good Manufacturing Process
cGMP
Doing what you said you were going to do
Proving that you did what you said
D
ocumenting that you did it.
Following SOPs
Validated methodsSlide6
Examples of Biologicals
Insulin
EPO
InterferonsToxinsAntibodyConjugates
Antibody-drug conjugatesFusion proteinsSlide7
Protein Structure
Secondary Structure:
3-dimensional
structure of
segments
Tertiary structure
:
Final
specific geometric shape that a protein assumes
Quaternary structure:
Arrangement
of multiple folded protein or coiling protein molecules in a multi-subunit complex.
Primary Structure:
Amino
acid sequenceSlide8
Insulin Slide9
AntibodySlide10
M
odifications
Post-translation modifications
Chemical changes not directly coded in DNAGlycosylation
PhosphorylationLipidationChemical degradations
OxidationsDe-amidationRearrangementsSlide11
Genetic Engineering (bacterial)Slide12
Monoclonal
AntibodiesSlide13
Bioprocessing
Unit Operations
Fermentation
Centrifugation
ChromatographySlide14
Variable inputs
Cells
“black box”
Process variables
Biologically derived
Chemically derived
Variable
c
rude product
Inherent heterogeneity
Stochastic processes
Heterogeneous
p
roduct
FormulationSlide15
Herceptin
(anti-cancer antibody)
Seven different
glycoforms
, each with different levels ofbiological
acitivity.
Variability of materialsSlide16
Quality by Design (
QbD
)
Operate within a specified design space Target Product Profile Critical Quality Attributes
Critical Product Attributes Better understanding of process and productDefining design spaceAllows for more variability
Process Analytical Technologies Real time monitoring & feedback.
Product by process
Lot release
criteria
- pass/
fail
Very difficult to make process improvementsSlide17
Design of
Experiment
Understanding the operating spaceSlide18
Design of Experiment (DOE)
A statistical method to model a process
Many fewer experiments than “one factor at a time”
Allows for modeling interactions
Useful to determine critical parameters
Critical for “Quality by Design”Slide19
Application of Design of Experiment to Conjugate
VaccinesSlide20
Design of Experiment
Concentration
CDAP Ratio
% conjugateSlide21
Process Analytical Technologies (PAT)
Real time feedback to control processSlide22
Types of Vaccines
Subunit (protein) vaccines
Tetanus toxoid
Diptheria toxoidPneumococcal (PneumoVax)
Killed vaccinesRubella, Measles, Polio (Salk)Flu
Hepatitis A
Live attenuated
vaccines
Polio (Sabin
)
Flu (
Flumist
)
Rotavirus (
Rotarix
)
Virus Like Particles (VLP
)
HPV (
Gardisil
)
New Generation Vaccines
DNA vaccines
Conjugate vaccines
Pneumococcal (
Prevnar
)
Meningicoccal
(
Menactra
)
Haemophilus
b (
Hib)Slide23
Polysaccharide Vaccine
Polysaccharide
Protein
+
Poorly immunogenic in infants
No boosting or memory
No class switching
No affinity maturation
Immunogenic in infants
Boosting and memory
Class
switching
Affinity maturation
Conjugate
VaccineSlide24
Conjugate Vaccines are EffectiveSlide25
Why Conjugate Vaccines?
Expensive
Challenging to manufacture
Many serotypes
Haemophilus
influenzae b (Hib)
Neisseria
meningiditis
Streptococcus
pneumoniae
Salmonella
typhiSlide26
HARVARD STRAIN
Cl.
tetani
INTERMEDIATE SEED
HIGB (18 -32
hrs
)
FERMENTOR
35 ± 5°C
ANAEROBIC CULTURE
Revival On ATG medium
48 hours at 37 °C
Inculcation into production
Medium (Muller &Miller)
(PH – 7.5 ± 0.1)
CRUDE TOXOID SUBLOT
STERTILE TOXIN
CULTURE HARVEST 6\7 days
SMEAR
PH
LF/ML
Sterility test
LF/ML
MLD
MTV
Antigenic purity
LF/ML
Detoxification test in mice
Detoxification with 0.5 % Formalin
4 days at Room temperature & 4 weeks at 35 °C
Millipore Filter
Clarification
Sterilization
CONCENTRATED TOXOID
BULK PURIFIED TETANUS TOXOID
POOLED CONCETRATE
(1-2 SUBLOTS)
Millipore Filter [30
KDa
] 0.2 to 0.45 micron
SUPERNATANT
STERILE FILTRATION
PURIFIED TOXOID
PRECIPITATION
Sterility test
Antigenic purity
LF/ML
Specific Toxicity
Irreversibility test
Released
for
Blending
Final Filtration (Cartridge Filter)
Dialysi
s
0.2 micron bag pore size
LF Test
LF Test
Centrifugation
(4000 rpm @45 minutes)
Precipitation with ammonium
Sulphate – I
st
salting (10% - 12%)
5
th
Day
sample collection for SMEAR, pH, growth
lysis
Precipitate with high ammonium sulphate (22% - 24%)
Centrifuge
4000 rpm
@ 1 hour
pooling
TETANUS
TOXOID PRODUCTIONSlide27
Identity
Polysaccharide composition
Moisture content
Protein impurity
Nucleic acid impurity
Pyrogen
content
Molecular size
distribution
Extent of activation
Molecular size distribution
Identity
Purity
Toxicity
Extent of
derivatisation
(if
appropriate) NR
Identity
Residual reagents
Saccharide:protein
ratio & conjugation
markers
Capping markers
Saccharide content NR
Conjugated v. free saccharide
Protein content
Molecular size distribution
Sterility
Specific toxicity of carrier (if appropriate)
Endotoxin content
Carrier Protein
Polysaccharide
Activated saccharide
Bulk Conjugate
Conjugation
Synthesis of Conjugate Vaccines
WHO Recommendations for the production and control of pneumococcal
conjugate
vaccines, ECBS, October 2003. Updated 2009.Slide28
Bulk
Conjugate1
Bulk
Conjugate2
Bulk
Conjugate3
Bulk
Conjugate4
Bulk
Conjugate5
Bulk
Conjugate6
Bulk
Conjugate7
Bulk
Conjugate8
Bulk
Conjugate9
Bulk
Conjugate10
Bulk
Conjugate11
Bulk
Conjugate12
Bulk
Conjugate13
Formulated
Vaccine
Multivalent pneumococcal conjugate vaccineSlide29
Carrier Protein
Bulk Conjugate
Final Vaccine
Polysaccharide
Activated saccharide
Identity
Polysaccharide composition
Moisture content
Protein impurity
Nucleic acid impurity
Pyrogen
content
Molecular size
distribution
Extent of activation
Molecular size distribution
Identity
Purity
Toxicity
Extent of
derivatisation
(if
appropriate) NR
Identity
Residual reagents
Saccharide:protein
ratio & conjugation
markers
Capping markers
Saccharide content NR
Conjugated v. free saccharide
Protein content
Molecular size distribution
Sterility
Specific toxicity of carrier (if appropriate)
Endotoxin content
Identity
Sterility
Saccharide content (of each)
Residual moisture
Endotoxin content
Adjuvant content (if used)
Preservataive
content (if used)
General safety test
pH
Inspection
Conjugation
Formulation
Control testing of Pn conjugates
WHO Recommendations for the production and control of pneumococcal
conjugate vaccines, ECBS, October 2003. Updated 2009.Slide30
>300 GMP steps for Prevnar13
Managing
supply
chain & supply chain quality
Each ingredient must be ready at the right time
QA/QC for bulk and formulated vaccine
Complexity of Supply Chain & Quality ControlSlide31
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