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Inhibition of DNA Methyltransferase Activates Tumor Necrosis Factor –Induced Monocytic Differentiation in Acute Myeloid Leukemia Cells

¹ Segal Cancer Center and Lady Davis Institute of the Jewish General Hospital for Medical Research and ² Department of Oncology, McGill University, Montreal, Quebec, Canada; ³ Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, New York; ⁴ Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill, North Carolina; and ⁵ Department of Experimental Pathology and Oncology, University of Florence, Florence, Italy

Requests for reprints: Wilson H. Miller, Jr., Lady Davis Institute for Medical Research, Sir Mortimer B. Davis Jewish General Hospital Segal Cancer Center, 3755 Côte Ste-Catherine Road, Montreal, Quebec, Canada H3T 1E2. Phone: 514-340-8260; Fax: 514-340-7576; E-mail: wmiller{at}ldi.jgh.mcgill.ca.

Key Words: 5-aza-2'-deoxycytidine • monocytic differentiation • TNF

Transcriptional silencing via promoter methylation of genes important for cell growth and differentiation plays a key role in myeloid leukemogenesis. We find that clinically achievable levels of 5-aza-2'-deoxycytidine (5-AZA-dC), a potent inhibitor of DNA methylation, can modify chromatin and restore the ability of tumor necrosis factor (TNF) to induce monocytic differentiation of the acute myeloid leukemia cells NB4 and U937. Although 5-AZA-dC cannot fully induce differentiation, we show that 5-AZA-dC acts directly on TNF-responsive promoters to facilitate TNF-induced transcriptional pathways leading to differentiation. 5-AZA-dC regulates the expression of Dif-2, a TNF target gene, by deacetylating chromatin domains in a methylation-dependent manner. Chromatin immunoprecipitation analyses of the Dif-2 promoter show histone hyperacetylation and a recruitment of the nuclear factor-B transcription factor in response to 5-AZA-dC. Furthermore, 5-AZA-dC plus TNF enhances the level of phosphorylated RNA Pol II at the Dif-2 promoter via synergistic recruitment of TFIIH. We conclude that nonspecific changes in chromatin can allow a specific transcriptional inducer to overcome blocks in leukemic cell differentiation. Our results support the concept of low doses of 5-AZA-dC acting in combination with other agents to target epigenetic changes that drive malignant growth in leukemic cells. [Cancer Res 2009;69(1):55–64]