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[First AACR International Conference on Molecular Diagnostics in Cancer Therapeutic Development, Sep 12-15, 2006]


Diagnostic Technologies and Molecular and Cellular Profiling: Biomarker Assay Development, Validation, and Qualification

Development of a quantitative enzyme immunoassay for measurement of PAR as a pharmacodynamic biomarker of PARP activity

Robert J. Kinders, Joann Palma, Xuesong Liu, Milagros Colon-Lopez, Yan Luo, Luiz E. Rodriguez, Yan Shi, Ran Guan, Shimin Wei, Jennifer Low, Anthony Murgo, Oxana Pickerel, Shivaani Kumar, Martin Gutierrez, Larry Rubinstein, Sherry Yang, Jay Ji, Gurmeet Kaur, Ralph Parchment, Tiziano DiPaolo, Jerry Collins, Melinda Hollingshead, Joseph Tomaszewski, James Doroshow and Vincent Giranda

Applied/Developmental Research Directorate, SAIC-Frederick, Inc., NCI Frederick, Frederick, MD, Pharmaceutical Products Division, Abbott Laboratories, Abbott Park, IL, Divison of Cancer Treatment and Diagnostics, NCI, Bethesda, MD, Divison of Cancer Treatment and Diagnostics, NCI; Center for Cancer Research, NCI, Bethesda, MD, Developmental Therapeutics Program, National Cancer Institute, Frederick, MD, Biological Testing Branch, NCI-Frederick, Frederick, MD, Developmental Therapeutics Program, National Cancer Institute, Bethesda, MD

Abstract

A5

A biochemical marker for activity of a target enzyme could provide an important readout of target inhibition and be a useful parameter in determining dosing regimens of a candidate clinical agent. Our team from Abbott Pharmaceuticals and the NCI-Frederick laboratories has developed and cross-validated a quantitative immunoassay for PAR (poly ADP-ribose polymerase) to monitor activity of PARP (PAR polymerase) as a biomarker of pharmacodynamic activity of the experimental agent ABT-888, a selective inhibitor of PARP1 and PARP2. The validated assay is a two-site enzyme chemiluminescence immunoassay employing commercially-obtained antibodies to PAR, and pure PAR as a standard. Assay dynamic range is 31 to 2000 pg/ml PAR, with an LLQ of approximately 15 pg/ml PAR. The standard curve is linear throughout the range (adjusted R2 typically better than 0.98). The assay uses High, Midrange and Low controls produced from the human melanoma line Colo 829. Specimen handling was optimized for both PBMCs and tumor needle biopsies (18 ga), and harmonized for use with the same standards and controls. Specimens could be subjected to at least 3 freeze-thaw cycles without a detectable loss of antigen binding. Assay precision was determined at both Abbott and the NCI-Frederick, to be better than 20% (estimated total imprecision at Abbott, 7% or less). Accuracy, as assessed by spike recovery of pure PAR into PBMC lysates, was 100% +/- 20%. Assay dilution linearity was established for the Colo829 controls, although deviations from linearity are observed in some tumor homogenates, and assay conditions are controlled to compensate for that lack of linearity. The validated assay was used to measure PAR levels in PBMCs of healthy donors, and in animal models after administration of a single dose of ABT-888. This assay will be used in real time to measure PAR in PBMCs and tumor biopsies in a Phase 0 clinical trial at the NCI.Funded by NCI Contract N01-CO-12400.







HOME HELP FEEDBACK HOW TO CITE ABSTRACTS ARCHIVE CME INFORMATION SEARCH
Cancer ResearchClinical Cancer Research
Cancer Epidemiology Biomarkers & PreventionMolecular Cancer Therapeutics
Molecular Cancer ResearchCancer Prevention Research
Cancer Prevention Journals PortalCancer Reviews Online
Annual Meeting Education BookMeeting Abstracts Online
Copyright © 2006 by the American Association for Cancer Research.