Abstract 1763: Quantitative gamma-H2AX immunoassay for pharmacodynamic monitoring anticancer cytotoxic agents

Abstract Phosphorylated H2AX (gamma-H2AX) has been reported as a biomarker for DNA double-strand breaks and programmed cell death. We have developed a 96-well plate based ELISA test for quantifying gamma-H2AX with the lowest limit quantiation at 16 pg/mL. The gamma-H2AX immunoassay is a cell- free assay with chemiluminescence readout using one unique pair of anti- gamma-H2AX antibodies as a sandwich bracketing the gamma-H2AX antigen from cell lysis extracts. The high affinity antibodies used in the assay provide high sensitivity and specificity allowing simple cell lysis procedures for antigen capture from crude extracts. The gamma-H2AX quantitation is normalized based on either cell numbers or total protein through BCA assay. We have performed gamma-H2AX quantitation in selected NCI60 cell lines from nine groups of human cancers. The gamma-H2AX levels among the NCI60 cell lines ranged from undetectable (< 16 pg/ml) to 6325 pg/10 million cells with an average of 711 pg. The gamma-H2AX quantitation results from ionic radiation and topotecan treated cell lines, topotecan treated human PBMC and selected patient specimens will be reported. The applications of the assay include target discovery and drug screening with selected human cancer cell lines, preclinical development and pharmacodynamic evaluation with xenograft tumors and animal models under treatment of investigational drugs, as well as clinic testing for patient enrollment, regiment selection and therapy monitoring of anti-cancer agents involving in DNA replication, repair, cell cycle and apoptosis. Funded by NCI Contract No. HHSN261200800001E Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1763.