Quantitative DNA methylation analysis identifies a single CpG dinucleotide important for ZAP-70 expression and predictive of prognosis in chronic lymphocytic leukemia

Rainer Claus, David M. Lucas, Stephan Stilgenbauer, Amy S. Ruppert, Lianbo Yu, Manuela Zucknick, Daniel Mertens, Andreas Buḧler, Christopher C. Oakes, Richard A. Larson, Neil E. Kay, Diane F. Jelinek, Thomas J. Kipps, Laura Z. Rassenti, John G. Gribben, Hartmut Doḧner, Nyla A. Heerema, Guido Marcucci, Christoph Plass, John C. Byrd

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90 Scopus citations

Abstract

Purpose: Increased ZAP-70 expression predicts poor prognosis in chronic lymphocytic leukemia (CLL). Current methods for accurately measuring ZAP-70 expression are problematic, preventing widespread application of these tests in clinical decision making. We therefore used comprehensive DNA methylation profiling of the ZAP-70 regulatory region to identify sites important for transcriptional control. Patients and Methods: High-resolution quantitative DNA methylation analysis of the entire ZAP-70 gene regulatory regions was conducted on 247 samples from patients with CLL from four independent clinical studies. Results: Through this comprehensive analysis, we identified a small area in the 5′ regulatory region of ZAP-70 that showed large variability in methylation in CLL samples but was universally methylated in normal B cells. High correlation with mRNA and protein expression, as well as activity in promoter reporter assays, revealed that within this differentially methylated region, a single CpG dinucleotide and neighboring nucleotides are particularly important in ZAP-70 transcriptional regulation. Furthermore, by using clustering approaches, we identified a prognostic role for this site in four independent data sets of patients with CLL using time to treatment, progression-free survival, and overall survival as clinical end points. Conclusion: Comprehensive quantitative DNA methylation analysis of the ZAP-70 gene in CLL identified important regions responsible for transcriptional regulation. In addition, loss of methylation at a specific single CpG dinucleotide in the ZAP-70 5′ regulatory sequence is a highly predictive and reproducible biomarker of poor prognosis in this disease. This work demonstrates the feasibility of using quantitative specific ZAP-70 methylation analysis as a relevant clinically applicable prognostic test in CLL.

Original languageEnglish (US)
Pages (from-to)2483-2491
Number of pages9
JournalJournal of Clinical Oncology
Volume30
Issue number20
DOIs
StatePublished - Jul 10 2012

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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    Claus, R., Lucas, D. M., Stilgenbauer, S., Ruppert, A. S., Yu, L., Zucknick, M., Mertens, D., Buḧler, A., Oakes, C. C., Larson, R. A., Kay, N. E., Jelinek, D. F., Kipps, T. J., Rassenti, L. Z., Gribben, J. G., Doḧner, H., Heerema, N. A., Marcucci, G., Plass, C., & Byrd, J. C. (2012). Quantitative DNA methylation analysis identifies a single CpG dinucleotide important for ZAP-70 expression and predictive of prognosis in chronic lymphocytic leukemia. Journal of Clinical Oncology, 30(20), 2483-2491. https://doi.org/10.1200/JCO.2011.39.3090