1. Principles and Applications of Lyophilization to Biotechnology
Michael J. Akers, Ph.D.
Baxter BioPharma Solutions
Bloomington, Indiana
BIOMAN 2010 Conference
Ivy Tech Community College
Bloomington, Indiana
July 13, 2010
Baxter Confidential

2. Organization of Presentation
Baxter Briefs
Biologics Market
Lyophilization
Baxter Confidential

3. Technologies Services
Bloomington, Indiana - Capabilities Overview
Technologies Services
Prefilled syringes
Lyophilization
Aseptic liquid vials
Cartridges
Diluent syringes
Formulation development
Process development
Lyo cycle development
Analytical development
Regulatory support
Labeling and Packaging
Clinical to Commercial
Baxter Confidential

4. Other Interesting Facts about Baxter BioPharma in Bloomington
Produce more prefilled syringes than any other manufacturer in North America
Small molecules, vaccines, proteins, diluents
Produce injectables marketed throughout the world
No history of FDA Warning Letters
308 employees in 2001; 993 end of 2009
Newest manufacturing building won ISPE facility of the year in 2006

5. Focus of Bloomington R&D
Formulation Development of Small Volume Injectables
Liquid
Lyophilized
Extensive experience with
Proteins
Monoclonal Antibodies
Small Molecules
Analytical Method Development
Method development and validation services available
Responsible for validation of transfer of all incoming QC methods for Bloomington facility
Expertise in cleaning validation limit calculations and method development/validation
Development stability studies
Lyophilization Process Optimization Research Programs
Approximately 80%
of projects in past 2 y
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6. Bloomington R&D Known As Baxter Lyophilization Center of Excellence
We characterize APIs by thermal analyses and freeze-dry microscopy to develop formulations and lyophilization cycles
We use freeze-dry microscopy and FTIR to evaluate formulation effects on protein stability
We coordinate transfer and scale up of lab-developed product to production freeze-dry operations.
Our research programs include mechanisms of heat and mass transfer during freeze-drying, nucleation optimization, process monitoring by Tunable Diode Laser Absorption Spectroscopy
Since 2001, 35 peer-review research articles, 3 books, 5 book chapters, dozens of short courses, and 80 presentations at conferences and universities
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7. Baxter BioPharma Solutions
Pharmaceutical Research and Development
Bloomington, IN
Top: Tim Paul, Lisa Hardwick, Nathan Pease, Wendy Saffell-Clemmer, Larry Callahan
Middle: Rhonda Haley,Wei Kuu, Melissa Beard, Mike Hildreth, Karen Abram, Kevin O’Bryan, Elizabeth Oslos, Michael Akers
Bottom: Lindsay Stanford, Angie Kruszynski, Nathaniel Fair, Jackie Karty, Greg Sacha, Hoang Mitchell, Dan Staples, Kelly Roby, Steven Nail
Baxter Confidential

8. Market segment by therapeutic area and molecule class
Source: Baxter Analysis
Baxter Confidential 8

9. Current/Future Facts About Biologics
Significant growth
2005 sales: $33B
2005 predicted 2010 sales: $82B
1 out of 4 new drugs introduced in US/EU is a biopharm product
Monoclonal antibodies (MoAbs)
$30B of 2010 sales expected to be MoAbs
> 200 MoAbs in clinical development right now
About half of current and new biologic products require lyophilization
Many of these products will be self-administered
Obviously, there is a huge market potential for simple, cost-effective devices to be used in home health care to prepare and deliver lyophilized biologics (and other lyo products)
Especially if product is for chronic health care
Baxter Confidential 9

10. Best Known (Best Selling) Lyophilized Biologics (Biopharmaceuticals)
Product Brand (Indication)
Company
2009 U.S. Sales
Enbrel (Arthritis)
Amgen
$6,580,000,000
Remicade (Arthritis, Crohn’s)
J&J
$5.934,000,000
Herceptin (Breast cancer)
Genentech
$4.890,000,000
7 Human Growth Hormone Brands (Growth hormone deficiency)
Pfizer, Novo, Lilly, Genentech, Serono, Sandoz, Ipsen
$2,886,000,000
Avonex (Multiple sclerosis)
Biogen Idec
$2,323,000,000
Advate (Hemophilia)
Baxter
$2,058,000,000
Betaferon (Muliple sclerosis)
Berlex/Bayer/Merck
$1,649,000,000
Total Biologic Sales
---
$91,780,000,000
Baxter Confidential

11. Lyophilization
The terms “lyophilization” and “freeze-drying” used interchangeably
Started in the 1930s, importance grew in WWII
Lyophilization is the conversion of a liquid to a solid through the process of sublimation
A sterile solution is prepared, filled into primary containers, placed into a freeze-dryer, and frozen
The solvent in the solution is removed by directly converting it from frozen ice to water vapor.
What remains in the container are the solute components in the solid state containing very low residual moisture (typically %).
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12. Schematic Overview of Processing Solution and Freeze-Dried Biopharmaceutical Dosage Forms
Dispense raw materials
(active and excipients)
Prepare solution in appropriate
mixing tank (add ingredients to Water For Injection)
Thaw and pool
active biopharmaceutical
Wash and sterilize
primary containers and closures
Add active to solution,
pH adjustment, final QS
This is formulation bulk solution
ISO 8, Grade D,
Class 100,000
ISO 5, Grade A
Class 100
Sterile filter formulated
bulk solution
Transfer to Freeze-dryers
and lyophilize
Aseptically fill formulated bulk solution
into primary package and stopper
(partial stopper if product is to be freeze-dried
Fully insert stopper, remove from freeze dryer
Apply aluminum overseal
Storage at appropriate
temperature (usually 2-8ºC)
Label, sec package, storage, distribution
100% inspection
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13. Preparing Product for Lyophilization
Prepare the sterile solution—compound, mix, filter
Fill into containers, typically vials
Partially insert a special designed rubber closure onto the vials
Aseptically load the vials into a freeze dry chamber
Freeze every single solution in every vial below a pre-determine critical temperature
Using appropriate application of temperature and pressure, sublime the ice from the product
Using further application of temperature and pressure, remove the necessary amount of bound water from the product
Automatically stopper the vials, neutralize the chamber
Aseptically remove the vials from the chamber and apply aluminum seals
Baxter Confidential

14. Today’s State of the Art Technology Uses Automated Systems To Transfer Product From Production Line Into the Freeze-Dryer

15. Baxter Pharmaceutical Solutions
Freeze-Dryer Subsystems (What Must Be Operating Smoothly and Maintained for Lyophilization Process To Succeed?)
Heat Transfer System
Refrigeration System
Vacuum System
Stopper-in-Place System
Clean-in-Place System
Control System
Loading/Unloading System
Typical Batch of a Lyophilized Biopharmaceutical Product Worth Several Million $
Baxter Pharmaceutical Solutions
Baxter Confidential

16. Courtesy of Samir U. Sane, Ph.D., Genentech, Inc.
Baxter Confidential

17. Product temperature lower than shelf temperature during primary drying because energy is consumed as ice sublimes to vapor.
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18. Mushroom Valve (open position)
Flow of Vapor from Chamber to Condenser T
Vacuum Pump T P
Water Vapor Flow
Door
Product Chamber
Condenser Chamber
Connecting Duct
Condenser Coils
Mushroom Valve (open position)
Shelves with Vials
Baxter Confidential

19. Schematic of Heat and Mass Transfer in the Freeze Dryer
Temperature difference between chamber and condenser and pressure differential between solution in vials and vacuum pump drives ice out of vial and onto the condenser
Conversion of solid (ice) to vapor in chamber called sublimation
Vacuum Pump
Mass Transfer
Condenser ΔP
Heat Transfer
Dry Cake
Pressure gradient between
sublimation front and chamber
Sublimation Front
Frozen Solution
Thermal Fluid
Shelf
Thermal fluid circulates within the shelves to control temperature in chamber
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20. Desired Freeze Dried Product Characteristics
Intact cake
Sufficient strength
Uniform color
Sufficiently dry
Sufficiently porous
Sterile
Free of pyrogens
Free of particulates
Chemically stable—both in dry
state and after reconstitution
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21. Advantages of Lyophilization
Removal of water at low temperature
Ease of reconstitution
Compatible with aseptic operations
More precise fill weight control
Done properly, the freeze-dried solid has relatively high specific surface area, which promotes rapid, complete reconstitution
Baxter Confidential

22. Disadvantages of Lyophilization
The drug may not be stable as a freeze-dried solid
Many biological molecules are damaged by the stresses associated with freezing, freeze-drying, or both
Not all solutes can be freeze-dried to form a pharmaceutically acceptable cake
Cost may be an issue, depending on the product
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23. Common Lyophilized Products
Pharmaceuticals – large and small molecules
Bacteria
Viruses
Vaccines
Plasma
Small Zoological Specimens (Taxidermy)
Fruit
Coffee
Flowers
Water-Damaged Documents
Baxter Confidential

24. Main Challenges of Lyophilization
Development of formulation that meets all the necessary critical product attribute requirements (quality appearance, potency, stability, recon time, etc)
Accurate determination of the “critical temperature” of final formulation necessary to determine conditions of lyophilization cycle (Tg’, Te, Tc)
Establishment of temperature, pressure, and time cycle settings that can achieve best product quality in shortest possible time
Transfer and scale-up of lab-developed process to production scale process
Keep all the equipment running smoothly
Special challenges in product development of lyophilized biologic formulations and cycles

25. Challenges in Lyophilization of Protein Pharmaceuticals
Baxter Pharmaceutical Solutions
Baxter Confidential

26. Challenges in Lyophilization of Protein Pharmaceuticals
Measurable differences in recovery of activity can be associated with differences in the thermal history of freezing
Some proteins can be subject to overdrying; that is dryer is not necessarily better
Stability of the freeze dried solid is often a concern. Most freeze dried proteins require refrigerated storage.
Solid state stability can be affected by small differences in residual moisture content.
Baxter Confidential

27. Why Does Lyophilization Take So Long?
Sometimes, the properties of the formulation require that the temperature be very low, often below –30oC. This decreases the driving force
The heat required is very high, and heat transfer in freeze drying is very inefficient
Resistance to mass transfer – transport of water vapor from the sublimation front through the porous bed of partially dried solids – can be significant.
Huge batch sizes, takes time to complete the 3 stages of lyophilization

28. Final Comments
Roughly 50% of all commercial biologic (therapeutic protein products) are lyophilized.
Lyophilization technology and the expertise to use it are vital to the ability of the biopharmaceutical industry to prepare and market life-saving injectable medicines
Lyophilization is the most challenging (and most expensive) of all sterile product manufacturing unit operations
Baxter Confidential

29. Ms. Wendy Saffell-Clemmer
Acknowledgements
Thanks to all my Baxter R&D colleagues for the great collegiality we have and what makes my job so enjoyable.
Thanks to the following individuals for providing some of the slides I showed
Dr. Steven Nail
Ms. Lisa Hardwick
Ms. Wendy Saffell-Clemmer
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