Managing Raw Material Variability Over The Life-Cycle Of A Molecule, Sigma S. Mostafa, Ph.D.Director, Process Development, Upstream KBI Biopharma, Inc. Session : Selection of Source Materials (Biological products)

1. Managing Raw Material Variability Over
The Life-Cycle Of A Molecule
Sigma S. Mostafa, Ph.D.
Director, Process Development, Upstream
KBI Biopharma, Inc.
Session : Selection of Source Materials (Biological products)

2. Overview
• Raw material types
• Common sources of variability
• Case studies
• Managing raw material variability

3. Types of Raw Materials
• Chemical Raw Material
 Culture media, feed, supplements
 Base and acid used for pH control
 Purification buffers
• Physical Raw Material
 Storage bags for media, feed, supplements
 Tubings, connectors,
 Disposable cell culture bags
• Biological Raw Material
 Master/Working cell bank, primary virus stock
 Serum
 Chicken eggs, tobacco plants

4. Common Sources Of Variabilities Over
Life-Cycle Of A Molecule
• Change in secondary vendors for RMs with multiple
components
• Multiple manufacturing sites receiving RM from vendor’s
multiple sites (e.g. US vs. EU sites of media vendors)
• Multiple lots of hydrolysates in media
• In case of physical RMs, product line may be discontinued
• Multiple WCBs

5. Case Studies

6. Case Study #1
Media performance Variability
Description
PD and GMP campaign for Phase I showed no problem with cell growth.
When Phase II GMP started after 1 yr of no cell culture activities, little to no
growth of cells.
Investigation
All Phase I cell culture done with a single medium lot (no lot to lot
variability data available).
Poor cell growth apparent after few passages, not immediately after vial
thaw.
A long list of items investigated – hydrolysates (2 types), cholesterol, iron supplement,
insulin, order of addition during liquid medium preparation, pH set points at different
steps of medium preparation, age of WCB, etc.

7. Case Study #1
Media performance Variability
Root Cause
The iron supplement used in the medium had a history of variable
performance. Vendor had changed the iron source in their newer products
but did not notify clients who used the older products of this potential
problem.
Investigation and resolution of the problem took 6 months and 8 FTEs from
client’s 3 sites

8. Case Study #2
Media performance Variability
Description
PD work showed no problem with cell growth. GMP campaign at a CMO
site showed half peak cell density and half titer compared to PD.
Investigation
For CMO work, liquid medium was transferred in bags across country. PD
work was done by hydrating powder few days before experiment.
Cell culture was done by a long perfusion process. Medium degradation in
Stainless steel reactor was never a problem.
Thorough investigation of raw materials, WCBs, seed train steps, and
production process.

9. Case Study #2
Media performance Variability
Root Cause
Loss of vitamin K during shipment of liquid medium across country.
Addition of extra vitamin K at CMO site prior to start of culture fully restored
culture performance.
Investigation took 1 month and 6 FTE.

10. Other Causes for Media/Feed Variability
• Aged liquid media/feed
• Precipitation in feed concentrate
- Often caused by pH/temperature interaction
• Water used for hydration of media/feed
- pH of water source affects solubility of powder
• Selective loss of components during sterile filtration
- larger components are some times lost especially if 1 micron
filter is used to prevent mycoplasma

11. Case Study #3
Disposable Bioreactor Bag Performance Variability
Description
Poor cell growth in disposable bioreactor.
Investigation
Cell growth tested in multiple types of disposable bioreactors.
Leachable content measured.
Root Cause
High leachable content in certain types of bags caused poor cell growth.

12. Case Study #4
Disposable Bioreactor Bag Performance Variability
Description
Breakage of impellers during mixing study.
Investigation
Impellers broke while mixing study was being performed at vendor
suggested RPM.
Investigation on going.

13. Case Study #5
Particulates in Buffer Bag
Description
Particulates found in buffer bag
Investigation
Testing of particulates indicated that origin of particulates was c-flex
tubing.
Root Cause
Buffers of certain characteristics (e.g. high pH/low conductivity)
when interfacing with air can degrade c-flex tubing over time.

14. Case Study #6
Cell bank affected during transfer
Description
WCB vials had poor viability upon thaw
Investigation
WCB vials thawed in PD had good viability; whereas vials thawed at
manufacturing had poor viability
Root Cause
Vials were shipped to PD and manufacturing separately. Vials at
manufacturing were compromised during shipment/handling/storage

15. Managing Raw Material Variabilities
• Comprehensive RM management plan
• In-depth understanding of the supply-chain of each RM
• At least one back-up supplier for each RM
• Use of platform process to minimize change of RM
• In house data on expiration dates of key RMs
• Information on RM production process differences from site to site
• During PD use RMs planned to be used during GMP
• If possible, in house media/feed formulation development
• If project is restarted after a long gap, do a demo run with raw materials before
actual run.

16. Questions?