• Drug Lot Release Potency Assay / Testing

    Posted on May 9, 2022

    Why have Lot Release Potency Testing?

    The US FDA requires that licensed products pass critical analysis before release by the manufacturer for clinical use. These include potency, sterility, purity and identity. Potency is used to ensure consistency and efficacy in the manufacturer’s product used for stability and lot release. A Potency Assay is typically reported as an EC50, which is the 50% of the maximal biological effect. Another typical reported value for the Potency Assay is the activity of a new lot of drug compared to a Reference Standard titrated in the same assay. This is reported as Relative Potency (RP).

    The goal for the potency assay is to represent the mechanism of action for the intended therapeutic activity. However, many new therapies use different modalities of activity (e.g. gene therapy requires transfer of genetic sequence into a cell and biological effect of this sequence). The traditional approach is to develop a quantitative biological assay (e.g. cell based/bioassay) that measures the specific intended activity with regards to the amount of drug. A surrogate assay (e.g. ELISA binding assay) can be used only if the measurement and biological activity is well established.

    In summary, potency assays will allow for product characterization and understanding of what affects the quality, potency, lot to lot consistency and stability over time. These assays should be initiated early during the development and pre-clinical stages in order to establish a baseline knowledge of the drug.

    What Methods Can be Used to Measure Potency?

    The bioassay or cell-based assay is the traditional approach whereupon the targeted mechanism of action is employed in the assay readout (e.g. synthesis of a cytokine or cell death or proliferation). The assay should be specific so that it is not affected by degraded drug, or inactive ingredients (e.g. matrix). At clinical Phase III and BLA / NDA stage, linearity, robustness (including at least two operators), and documentation of acceptable range of critical parameters and parallelism showing qualitative equivalence of the test sample and Standard, are required.

    Early phase studies, Phase 1 or 2, the assay criteria will have wider acceptance criteria than later phase investigations. In Phase 3, pivotal studies, the acceptance criteria will be narrower to support efficacy and assure lot-to-lot drug preparations are biologically active and consistently manufactured.

    Development and Validation of Potency Assays

    The strategy for development of Potency Assays (Fig. 1) is to ensure that the assay will be accurate and robust through validation and reproducible in subsequent use

    Fig. 1: Strategies for development of potency assays for drug lot release testing.

    Issues to be investigated

    • Precision (Repeatability, Intermediate Precision), each assay must be shown to give a precise RP with a small 95% confidence interval
    • Accuracy, the method must be shown to measure a lot accurately, target within 10% at the BLA stage
    • Specificity, not affected by interference, (e.g. drug incipient, drug degradants, or other drugs in the manufacturing pipeline) should have negligible response
    • Linearity Range, and Accurate Concentrations, the method should be shown to have a linear dose response (e.g. 70% to 130% of the nominal 100% value)
    • Stability Indicating, the method should be able to distinguish purposefully degraded material
    • Parallelism, a new lot should have a dose response curve that parallels the Reference Standard, to demonstrate that it is quantitatively the same as the Reference Standard
    • System Suitability, from the above criteria, a set of system suitability criteria are developed to demonstrate that the assay passes
    • Robustness, from the above criteria, the assay should be reproducible from operator to operator, laboratory to laboratory, time to time

    Marin Bio has validated many different kinds of Potency Assays which are used for drug lot release and stability studies over many years. Let our scientists help you with your Potency Assay design, development, qualification/validation and implementation throughout your drug’s life cycle.

    Potency Assays for Lot-Release Testing

    • Cell-based assays: (dependent on drug types, functions, read out)
    • Ligand binding assays – In vitro assays
    • ELISA
    • Plasmid, viral transfection assays
    • Customized assays

    Case Study: Cell-based potency assay using a bispecific antibody drug which binds a killer T-cell line and binds to the cancer cell line and measures cell killing. This experiment compares Reference Standard to Test Sample (bifunctional or bispecific antibody)

    The effector and target cell lines are incubated at a specific effector cell-to-target cell concentration. The target cancer cells are pre-loaded with a fluorescent dye. Increasing concentrations of bispecific antibody are added. Incubation of cells with drugs is 2.5 hours and the fluorescence released into the supernatant from dead target cells is measured.

    Fig. 2: Graph of bispecific antibody directing effector T cells killing a cancer target.  Fluorochromes released from killed targets cells vs ng/mL bispecific antibody, Reference Standard compared to a Test Sample

    Our Unique Services

    • Offers GMP, GLP and non-GMP potency assays.
    • MarinBio scientists develop customized and license-free innovative lot release potency assays for clients.

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