Training Seminar: Introduction to Analytical Method Development and Validation for Therapeutic Proteins Wednesday, January 21, 8:30 AM-4:25 PM - Thursday, January 22, 8:30 AM-12:30 PM Part of PepTalk: The Protein Science Week This course is a panoramic review of analytical method development and validation for therapeutic proteins, including antibodies and enzymes. It is intended for scientists working on therapeutic proteins in AD, QC, PD or related functional areas. It starts with basic knowledge of work on therapeutic proteins: manufacturing of proteins drugs, regulatory affair knowledge and protein chemistry. It then discusses fundamentals and practical aspects of commonly used analytical methods for proteins, including methods for structure elucidation, glycan characterization, biophysical characterization, potency measurement, purity and impurity analyses. The course concludes with the strategy and common practice in method validation and method transfer, including regulatory compliance at different stages of product development, application of DOE and QbD. The course emphasizes practical applications, real-world examples and useful tips. For full details visit www.chi-peptalk.com/peptalk_content.aspx?id=140090&libID=140061

1. AAnnaallyyttiiccaall MMeetthhoodd DDeevveellooppmmeenntt &&
VVaalliiddaattiioonn ffoorr TThheerraappeeuuttiicc PPrrootteeiinnss
1
Jichao (Jay) Kang, Ph.D., RAC
Director, Analytical and Formulation Development
Gallus Biopharmaceuticals NJ, LLC

2. CCoouurrssee OObbjjeeccttiivveess
Basic scientific and industrial knowledge of
working on proteins (including antibodies)
Get familiar with commonly used analytical
methods: purpose, advantages, limits, pitfalls
Get familiar with Protein Characterization
Get familiar with method validation and method
transfer: principle, common practice, and
potential problems
2

3. BBaassiicc SSttrruuccttuurree ooff MMAAbb
3

4. MMaajjoorr NN--lliinnkkeedd GGllyyccaannss
4
Classified depending on the extent of processing:
High mannose: lack Gal or GlcNAc at the end of antennae
Complex: GlcNAc after the Man at the antennae
Hybrid: have both Man and Gal/GlcNac at end of the antennae
Hybrid and Complex types can have two or more branches, termed Bi-antennary,
tri-antennary, or Tetra-antennary

5. RReedduucceedd CCEE--SSDDSS ooff MMAAbb
5

6. SSEECC--HHPPLLCC--MMAALLSS
Three detector monitoring
6

7. Pr Hydroxylated Proolliinnee aanndd LLyyssiinnee bbyy AAAAAA
7

8. Str Protein Struuccttuurree EElluucciiddaattiioonn bbyy MMSS//MMSS
8
Roepstorff Nomenclature Scheme

9. SSeeccoonnddaarryy SSttrruuccttuurree bbyy CCDD
9
T Estey, J kang et al., Pharm Sci , 2006, 95: 1626–1639

10. TTiimmiinngg ffoorr MMeetthhoodd
QQuuaalliiffiiccaattiioonn//VVaalliiddaattiioonn
10
Preclinical Clinical Commercial
Phase I Phase II Phase III
Qualification Validation Verification
Method Development and Qualification
Measurement
defined
Scientifically
Defendable
Ready for
validation
Fully Validated
Method Transfer
Methods related to product safety require qualification even at phase I
Other methods need to be validated prior to BLA, but by phase III, you should
have the methods that can be validated

11. Bi Case Study: Binnddiinngg AAssssaayy QQuuaalliiffiiccaattiioonn
11
1 10 100 1000 10000 100000
x axis
3
2
1
0
Parallel Line Analysis
4-P Fit: y = (A - D)/( 1 + (x/C)^B ) + D: A B C D Rel. Pot.
Standard (Standard: Concentration vs Mean... 0.0395 1.31 202 2.61 1
Sample3 (150%: Concentration vs MeanVal... 0.0395 1.31 127 2.61 1.59
Sample1 (50%: Concentration vs MeanValue) 0.0395 1.31 372 2.61 0.541
Sample2 (100%: Concentration vs MeanVal... 0.0395 1.31 200 2.61 1.01
__________
Curve Fit Option - Fixed Weight Value
PLA (Std. Curve: Standard) Degrees of Freedom: parallel = 37 free = 28 non-parallel = 9
R^2 = 1 F-stat = 2.14 F-prob = 0.712
Binding ELISA curves at different level

12. QQuuiizz ## 1155
For an identification method, what are the
validation parameters?
How to determine if a method is stability-indicating?
How to determine LOQ?
How to calculate method supported
specification (SS)?
List three considerations in setting SST
List three method transfer pitfalls
12