1. CSI Biopharmaceutical Manufacturing: How Manufacturing Failures are Investigated Katie Stewart – Technical Training Manager

2. 2 Biopharmaceuticals What are Biopharmaceuticals? – Protein or nucleic acid based pharmaceutical therapeutics – Produced by means other than: 1) Direct extraction from a biological source 2) Or chemical synthesis Biopharmaceutical examples – Recombinant protein – Insulin from E.coli – Antisense DNA – blocks protein production – Monoclonal Antibody – to work with immune system

3. 3 Example of an Antibody Pharmaceutical  BENLYSTA is a Monoclonal Antibody  Treatment for Lupus (autoimmune disease)  Controls immune system

4. 4 Various types of cells can be used to produce biopharmaceuticals Mammalian/Animal – typically used for antibody production Yeast Saccharomyces cerevisiae (primarily used by HGS) Pichia pastoris Plant Bacteria Insect How do we create new Biopharmaceuticals?

5. 5 A “gene,” which codes for the drug product (protein or antibody) is inserted into a cell’s DNA Gene Cell Nucleus of cell where the DNA exists

6. 6 The drug product (protein or antibody) is made by the cell using the inserted DNA and the cells machinery (inside the cell) through several steps The cell then secretes the product outside of itself Cell How do we create new Biopharmaceuticals?

7. Manufacturing Biopharmaceuticals 7 Upstream Production Cells are grown to large quantities to produce a lot of product Downstream (Purification) Product is collected, purified (cleaned) and concentrated (large to small volume) through multiple Cells Product Production

8. 8 Manufacturing Biopharmaceuticals - Upstream Centrifuge Filter Vial Thaw / Inoculum Expansion 500-Liter Bioreactor 50-Liter Bioreactor Filtration Media Prep 5,000-Liter Bioreactor 20,000-Liter Bioreactor Material is Transferred to Purification - “Downstream” START FINISH

9. What is a Bioreactor? An apparatus for growing organisms (yeast, bacteria, or animal cells) – Controls environment around cells - temperature, pH, and oxygen levels – Create homogeneous environment (same throughout) environment by constantly stirring the contents.

10. Deviation Investigating in Manufacturing

11. Deviations Deviation – Any variance that may have the potential to affect the identity, strength, quality or purity of a product – FDA Regulation - thorough deviation investigations must be conducted Why investigate? Consequences of incomplete investigations: – No FDA approval Loss of hundreds (+) jobs Unmet patient needs Impact to shareholders ($$$$$$) – Endangering of patients Impure or potentially dangerous product

12. Investigation Process Outline 1.Define the problem 2.Brainstorm potential causes 3.Investigate potential causes 4.Identify cause 5.Implement corrections

13. 13 1. Define the Problem 5 Steps to Developing a Problem Describe the problem in terms of 5 factors: 1.What is the problem? 2.Where is it observed? 3.When is (was) it occurring? 4.What is the magnitude of the problem (Trend or Extent)? 5.What might have been the cause?

14. 14 2. Brainstorm Potential Causes Fishbone - Investigation Tool List potential causes under the following 6 categories – Man (& Woman), – Machine, – Mother nature, – Methods, – Materials, – Measures

15. 15 3. Investigate Potential Causes 4. Determine Root Cause From the Fishbone list gather all of the facts to determine whether each item is or is not the root cause Photo(s) Material affected Operator Interviews Equipment Condition and Maintenance Training Records Recent Changes Documentation and Procedures

16. 5. Implementing Corrections Corrective and Preventative Actions – Corrective Action: Corrects the root cause – Preventative Action: Ensure the deviation does not reoccur – Example: A procedure was incorrectly written and caused the test to fail Corrective Action - Revise the procedure Preventative Action - Review all documents in the area to ensure they are accurate and complete

17. 17 Fishbone Example Mother Nature Cleanliness of room Power outage Machine Equipment not sterile Poor equipment condition Broken controller Materials Solution sterility Cell culture sterility Filter integrity Bioreactor Bag & line integrity Measures Temperature probe broken False positive bacteria result Man Procedures weren’t followed Operators were not trained Methods Procedures weren’t clear Process flaw Deviations occurred Problem: The presence of bacteria was found in a sterile cell culture solution in a bioreactor