Because the focus of the meeting was biomolecules, the speakers were chosen based on experience in the relevant fields. The questions raised were relevant to regulatory compliance and exhibited a general concern about agency wishes versus corporate actions.
Dr. Rob Garnick (Genentech) related his experiences in the development of therapeutic proteins at Genentech over the past twelve years. He emphasized the need to know the product molecule in every aspect and then to evaluate each method used to gain this knowledge. One aspect he discussed was their attempt to replace a very expensive bioassay requiring an animal colony with an in vitro receptor binding assay to characterize the potency of human growth hormone. Even with significant effort by Genentech and cross-validation of the two assays by the FDA, Genentech is presently required to perform the more expensive bioassay. Validation of the in vitro approach and full characterization and quality control of the receptor integral to the in vitro assay were critical to gaining this level of cooperation from the FDA. A clear message that came from Dr. Garnick's talk was to scientifically know your system, set up a rigid set of quality control methods based on the key parameters of the molecule, and document everything.
Augustine Smith (Abbott) spoke in detail about how to validate a complicated derivitization and separation method such as amino acid analysis. She discussed each step in a logical fashion, emphasizing the strategy behind methods validation rather than specifics applicable only to amino acid analysis. Although not heavily employed, she has found statistically designed experimentation to improve efficiency in validating the key parameters of a method. Rather than spending significant resources on software validation at the "code-level", Augustine suggested that a more efficient and acceptable approach was functional testing of the system as a whole using a NIST standard and comparing with expected values. As to choice of amino acid analysis method, Augustine had no preference other than use of the system that exists in house.
Muriel Doleman (Sanofi) gave a concise presentation on the validation of a multiple-step enzyme map of an enzyme-polymer conjugate, emphasizing strategy and logic rather than details. As part of this validation, every proteolytic fragment was characterized by mass spectrometry, amino acid analysis, and Edman degradation. Additionally, a restricted time-window and peak height was specified for each peptide in order for the system to meet "system suitability" requirements. These requirements test the system for its ability to digest and separate the peptide fragments derived from a reference standard of the enzyme-polymer conjugate prior to analyzing the sample. Lot-to-lot variability in the function of the enzyme (Asp-N) generated the need for this suitability test to ensure that results obtained are meaningful. Due to its complexity, it is not used as a release method but rather a characterization tool capable of determining the percent modification by PEG at each lysine residue within the protein.
As part of the committee's movement towards determining the needs of the ABRF and Protein Society members, a survey was distributed to attendees. This survey probed methods for determining system suitability as well as methods validation. The results are presently being compiled.
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