Implementation of validated pharmacodynamic assays in multiple laboratories: challenges, successes, and limitations

Abstract

There is a "life cycle" of pharmacodynamic (PD) biomarker assays that guides the development and clinical implementation in our laboratories. The well-recognized elements of analytical assay validation and demonstration of fitness-for-purpose of the biomarker, specimen collection, handling, and assay methods are only a part of the required activities. Assay transfer across laboratories and testing on actual human clinical specimens are vital for understanding assay performance and robustness. In our experience, this patient specimen-centered approach has required assay method modifications, some unexpected, but which were critical to successful implementation in clinical trials. In addition, dispersing assays throughout the National Cancer Institute's clinical trials network has required the development of calibrator and control materials as well as formal training courses for smooth implementation. One measure of success of this approach has been that a number of the assays developed at NCI's Frederick National Laboratory have ultimately reached the stage of commercialization, enabling wide accessibility of the PD biomarker assays by the research community. See all articles in this ccr focus section, "Progress in pharmacodynamic endpoints."

Fig. 1

Functional Flow Chart of the PD assay development, implementation and support strategy used in the Division of Cancer Treatment and Diagnosis for support of early stage clinical trials.

Figure 2

γH2AX detection in CTC A, Photomicrograph of γH2AX-postive MCF-7 breast cancer cells used as the positive control. Cells were treated with 1 εmol/L topotecan for 2 hours and stained with γH2AX-AF488 (green) and DAPI (blue). Image capture was conducted using Leica 5000 DM. Final Magnification 200x. B & C, Images of γH2AX-postive CTCs from a breast cancer patient treated with ABT-888 combined with Cytoxan. The blood samples were obtained from the patient in cycle one on (B) day 2, 24 hours after drug administration and (C) day 5, 24 hours after drug administration. The EpCAM-enriched CTCs were stained with conjugated antibodies for CK-PE, CD45-APC, DAPI, and γH2AX-AF488 and analyzed on the CellSearch system.