Notes from Round Table 5. January 26 and 27, 2016

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1 Notes from Round Table 5 January 26 and 27, 2016 Topic: Facilitator: Scribe: Potency Assays: Cell Based vs. Non Cell Based Formats Sally Seaver, Seaver Associates LLC Cheryl Blaise, Bristol-Myers Squibb NOTES: A test for potency is one of the required, listed tests (21CFR610.10) and which shall consist of either in vitro or in vivo tests, or both, which have been specifically designed for each product so as to indicate its potency in a manner adequate to satisfy the interpretation of potency given by the definition in 600.3(s). 21CFR600.3(s) defines potency as the specific ability or capacity of the product, as indicated by appropriate laboratory tests or by adequately controlled clinical data obtained through the administration of the product in the manner intended, to effect a given result. In other words a potency test shows that the sample/lot/batch is active. Originally most potency tests were in vivo (animal based) but as biologica/biotech products became well characterized in vivo tests were replaced with in vitro (cell based) tests. The later are more accurate, precise, faster and more reliable. Now many groups start with non-cell based assays such as binding to an antigen, receptor or other target molecule, which they find to be even more reliable (faster, more accurate and precise). Many regulators consider the non cell based test not a true test of potency. BULLET POINTS FOR DISCUSSION Does your company have platform formats for potency tests whether cell and non-cell based? How many formats does your company consider for binding tests? For cell based tests? o Typically a challenge to have platform assays since potency assays are specific o Some have platformed the Promega (ADCC and/or CDC) assays and it works very well o Promega assay being used as release/stability GMP assays FDA OK with approach; being used at CRO o Most people happier with readout cell lines as opposed to primary cell lines o Use same technology for all assays competitive binding assay format (automated binding assay) all through the development process from pre-clinical to commercial (allergan products, substitute skin test for the binding assay); in this case representative of MOA o Depending on product, selectively use binding assay format as seeing some issues with the CBA o For CBA development, have tried many things; cannot get the CBA to work o Ideal scenario, use binding and CBA, demonstrate comparability between two assays and drop CBA (faster, easier, more consistent) is possible. More likely than not, need to keep CBA

2 1. Linked to MOA hard to have platform methods 2. Maybe more applicable to FC effector functions 3. Assessment to platform may be whole new development 4. Most companies typically start with a binding assay then add in a CBA later in development 5. need to have a CBA that is truly reflective of the MOA 6. really need to understand the action of the drug to develop a CBA that is reflective of what you want to measure; understand the biology behind what is going on Formats: Binding assays: ELISA and Biacore and FACS CBA: Depends on the cell type involved for the assay When do you start work on cell based potency tests? Non-cell based tests. How do you show (or do you show) that the binding test leads to a desired therapeutic action? o Small company perspective: binding assay is part of the initial screens; stay with binding as long as possible; bring in CBA when possible; Use CBA as a characterization assay with report results until more knowledge is gained. Binding assays typically not stability indicating o Binding assays in Ph 1 and 2. CBA in Ph 3 o File IND with a binding assay; feedback from FDA need a CBA. research has an assay but not a QC method so will develop a CBA as development progresses o usually only know that the antibody is binding to the antigen target, but not if there is a biological activity o do not know the MOA this early on to show that the desired therapeutic action is there If you have several cell or non-cell based tests, how do you choose which one(s) o To qualify toxicology material? o For early clinical material? o For pivotal clinical trial material? o To submit for approved product material? when to switch between binding and CBA? use a gradual approach? what about multiple CBA assays? how to pick between? o Start with qualified assays and collect the data to help to decide which assay to keep; need to consider what is going to wind up in the QC lab o Good idea to have more than one potency assay especially at the beginning where MOA may not be always known with certainty Commercial reagents and kits for potency: are they OK to use? justified? how to mitigate risk o try to avoid one vendor o working ok for now for Promega (ADCC/CDC) o cell lines are slightly different; may be a better situation than for a kit; can purchase rights to the cell line for use

3 o (complement reagent) may be a problem as the reproducibility from vendor to vendor may be problematic o Need to be aware of product quality o Need vendors to notify of changes when primary reagents change o start to have a CBA in the queue as soon as possible o need risk assessment on the CQAs o cell culture conditions, then start to evaluate the cell assay o CBA needed ASAP even if only a characterization assay o usually a characterization assay early on o binding assay is for QC early on o sometimes don t have any clue as to the MOA As you transition between cell based and non-cell based formats, what does your company do to show the comparability of the old and new formats? o Before tox lots o Before clinical lots o Before pivotal clinical lots o After approval changing from one assay to another use stats based on assay precision to determined how many samples to test uses stats to determine if assay results are comparable; set up criteria to see if precision is better with new assay use different lots if available get a pile of data and do some stats great if you have a lot of lots to test is phase appropriate; onus is greater later on if switch is made late in development may need forced deg material to help with testing to indicate stability indicating method When to drop one assay: typically after one is demonstrated to not be needed Direct binding assay does not indicate MOA; CBA is typically more variable but will tell you something is working or not working CBA is where more work is placed as MOA is key to assay Just b/c something binds does not mean it is going to be active in the cell It is possible to have just a binding assay w/o CBA with proper justification How does the qualification of operators for the specific test formats change?

4 o Before tox lots o Before clinical lots o Before pivotal clinical lots o After approval QC difficult to teach some technical challenging assays Set acceptance criteria for the training based on the performance of the assay Depends on the type of assay reporter gene assay for one product and another product o read and understand o demonstrate two or three assay runs with acceptable outcomes Most of all need an assay that is easy to train on operators come to innovator site to train first operators then return home to demonstrate their ability training protocol used to bring on new operators developers will work with the QC analysts to train Other topics for discussion: Is everyone performing relative potency? o Yes; Monitor EC50 for assay performance Is EC50 used as a sys suit criteria o No, use it as a trending tool to determine if something is going on with the assay How to qualify new lot of material eg - antigen or antibody o test old lot and new lot side by side o qualify anything that is coming in that is changed o can use the concept of a comparability protocol to help pre-justify changes document outlines how you will qualify new lot of material Can remove data points but need to be prescriptive on how to do so (one point or two, but not next to each other, not on the ends...) especially in phase 1 and 2 o need to make sure that accuracy and precision are not affected What is most appropriate assay to measure potency? consider MOA first and foremost o whatever it most appropriate for measuring the potency as defined by CQAs Can testing for CBA stop at some point? o on a case by case basis maybe o need to build a correlation between the content and the potency o although potency and content is always directly related o how well do you know your product and what other assays do you have how are you able to develop an precise bioassay o best precision achieved for CBA is 3%, 10%, <10%; typically 10% is best precision achieved o all about indicator cell line o cryopreserved cells helps too o banking is critical

5 o need set of parameters to establish banks to avoid drift; trending helps o General precision of binding assay is around 5% Any success in not implementing a CBA: o No experience around the table o Suggestion to talk to the agency and relay all of the information of the work that was attempted; show good scientific effort o Especially common with primary cells o Potentially mimic the biology in another cell line o Show patient benefit for drug When to drop one assay: typically after one is demonstrated to not be needed o Direct binding assay does not indicate MOA; CBA is typically more variable but will tell you something is working or not working o CBA is where more work is placed as MOA is key to assay o Just b/c something binds does not mean it is going to be active in the cell o It is possible to have just a binding assay w/o CBA with proper justification