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Development of a Validated Immunofluorescence Assay for gamma H2AX as a Pharmacodynamic Marker of Topoisomerase I Inhibitor Activity

  1. Author:
    Kinders, R. J.
    Hollingshead, M.
    Lawrence, S.
    Ji, J. P.
    Tabb, B.
    Bonner, W. M.
    Pommier, Y.
    Rubinstein, L.
    Evrard, Y. A.
    Parchment, R. E.
    Tomaszewski, J.
    Doroshow, J. H.
    Natl Canc Inst Phase 0 Clinical, T.

  2. Author Address

    [Kinders, Robert J.] NCI, Lab Human Toxicol & Pharmacol, Appl Dev Res Support Directorate, Sci Applicat Int Corp Frederick Inc, Frederick, MD 21702 USA. [Lawrence, Scott; Tabb, Brian] NCI, Pathol Histotechnol Lab, Lab Anim Sci Program, Frederick, MD 21702 USA. [Hollingshead, Melinda] NCI, Biol Testing Branch, Dev Therapeut Program, Frederick, MD 21702 USA. [Bonner, William M.; Pommier, Yves; Doroshow, James H.] NCI, Mol Pharmacol Lab, Ctr Canc Res, Bethesda, MD 20892 USA. [Rubinstein, Larry; Tomaszewski, Joseph; Doroshow, James H.] NCI, Div Canc Treatment & Diag, Bethesda, MD 20892 USA.;Kinders, RJ, NCI, Lab Human Toxicol & Pharmacol, Appl Dev Res Support Directorate, Sci Applicat Int Corp Frederick Inc, POB B, Frederick, MD 21702 USA.;kindersr@mail.nih.gov
    1. Year: 2010
    2. Date: Nov
  2. Journal: Clinical Cancer Research
    1. 16
    2. 22
    3. Pages: 5447-5457
  4. Type of Article: Article
  5. ISSN: 1078-0432
  1. Abstract:

    Purpose: Phosphorylated histone H2AX (gamma H2AX) serves as a biomarker for formation of DNA double-strand break repair complexes. A quantitative pharmacodynamic immunofluorescence assay for gamma H2AX was developed, validated, and tested in human tumor xenograft models with the use of clinically relevant procedures. Experimental Design: The gamma H2AX immunofluorescence assay uses a novel data quantitation and image processing algorithm to determine the extent of nuclear-specific gamma H2AX staining in tumor needle biopsies and hair follicles collected from mice bearing topotecan-responsive A375 xenografts. After method validation with the topoisomerase I (Top1) inhibitor topotecan, the assay was used to compare pharmacodynamic properties of three structurally related indenoisoquinoline Top1 inhibitors. Results: gamma H2AX response to topotecan was quantified over a 60-fold dose range (0.016-1.0 times the murine single-dose maximum tolerated dose), and significant pharmacodynamic response was measured at the mouse equivalent of the 1.5 mg/m(2) clinical dose as well as the lowest dose tested. Responses were within a time window amenable for biopsy collection in clinical trials. These studies enabled characterization of dose and time responses for three indenoisoquinolines, resulting in selection of two for clinical evaluation. gamma H2AX response to Top1 inhibitors in hair follicles was also observable above a minimal dose threshold. Conclusions: Our gamma H2AX assay is sufficiently accurate and sensitive to quantify gamma H2AX in tumor samples and will be used in correlative studies of two indenoisoquinolines in a phase I clinical trial at the National Cancer Institute. Data suggest that hair follicles may potentially serve as a surrogate tissue to evaluate tumor gamma H2AX response to Top1 inhibitors. Clin Cancer Res; 16(22); 5447-57. (C) 2010 AACR.

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External Sources

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  2. DOI: 10.1158/1078-0432.ccr-09-3076
  3. View record in Web of Science®
  4. WOS: 000284173600009

Library Notes

  1. Fiscal Year: FY2010-2011
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