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This paper tests the hypothesis that cytosine DNA methyltransferase (DNA MeTase) is a candidate target for anticancer therapy. Several observations have suggested recently that hyperactivation of DNA MeTase plays a critical role in initiation and progression of cancer and that its up-regulation is a component of the Ras oncogenic signaling pathway. We show that a phosphorothioate-modified, antisense oligodeoxynucleotide directed against the DNA MeTase mRNA reduces the level of DNA MeTase mRNA, inhibits DNA MeTase activity, and inhibits anchorage independent growth of Y1 adrenocortical carcinoma cells ex vivo in a dosedependent manner. Injection of DNA MeTase antisense oligodeoxynucleotides i.p. inhibits the growth of Y1 tumors in syngeneic LAF1 mice, reduces the level of DNA MeTase, and induces demethylation of the adrenocortical-specific gene C21 and its expression in tumors in vivo. These results support the hypothesis that an increase in DNA MeTase activity is critical for tumorigenesis and is reversible by pharmacological inhibition of DNA MeTase.Modification of DNA by methylation is now recognized as an important mechanism of epigenetic regulation of genomic functions (1-3). Methylation of DNA is a postreplication event catalyzed by the DNA methyltransferase (DNA MeTase) enzyme using S-adenosyl methionine as a methyl donor (4). Approximately 80% of cytosines located in the CpG dinucleotide sequence are methylated in the genome of most vertebrate cells, but the distribution of methylated sites is cell-and tissue-specific (5). Patterns of methylation are generated during development by enzymatic de novo methylation and demethylation processes (1-7) and are maintained in somatic cells.A number of observations have suggested that the pattern of DNA methylation is disrupted in cancer cells (8,9). Both hypomethylation (9) and hypermethylation (10-12) of different CpG sites in cancer cells and tissues relative to the cognate normal tissue have been documented. Some of the sites that are hypermethylated in tumors are located in tumor-suppressor loci such as p16 (13), retinoblastoma (14), von Hippel-Lindau (15), and Wilms tumor (16), and, recently, a new candidate tumorsuppressor gene was cloned by molecular analysis of the hypermethylated region in chromosome 17p13.3 (17). One possible explanation that has been proposed to explain the changes in DNA methylation observed in cancer cells is that they are the end result of a change in the enzymatic machinery controlling DNA methylation in the cell (7,12,(18)(19)(20). In accordance with this hypothesis, cancer cell lines (21) and human tumors (22) have been shown to express elevated levels of DNA MeTase. Recently, Belinsky et al. (23) showed that increased DNA MeTase activity is an early event in carcinogen-initiated lung cancer in the mouse. Forced expression of DNA MeTase cDNA in murine NIH 3T3 cells leads to genomic hypermethylation and neoplastic transformation (24), and expression of an antisense mRNA to the DNA MeTase leads to loss of tumorigenicity of t...

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This paper tests the hypothesis that expression of the DNA methyltransferase, dnmt1, gene is regulated by a methylation-sensitive DNA element. Methylation of DNA is an attractive system for feedback regulation of DNA methyltransferase as the final product of the reaction, methylated DNA, can regulate gene expression in cis. We show that an AP-1-dependent regulatory element of dnmt1 is heavily methylated in most somatic tissues and in the mouse embryonal cell line, P19, and completely unmethylated in a mouse adrenal carcinoma cell line, Y1. dnmt1 is highly over expressed in Y1 relative to P19 cell lines. Global inhibition of DNA methylation in P19 cells by 5-azadeoxycytidine results in demethylation of the AP-1 regulatory region and induction of dnmt1 expression in P19cells, but not Y1 cells. We propose that this regulatory region of dnmt1 acts as a sensor of the DNA methylation capacity of the cell. These results provide an explanation for the documented coexistence of global hypomethylation and high levels of DNA methyltransferase activity in many cancer cells and for the carcinogenic effect of hypomethylating diets.

Two receptors [estrogen receptor (ER)alpha and ERbeta] mediate the manifold effects of estrogens throughout the body. Although a clear role has been established for ERalpha in the classical effects of estrogen activity, the physiological role of ERbeta is less well understood. A small-molecule ERbeta selective agonist, ERB-041, has potent antiinflammatory activity in the Lewis rat model of adjuvant-induced arthritis. To characterize the response of target organs and pathways responsible for this antiinflammatory effect, mRNA expression profiling of the spleen, lymph node, and liver was performed, in conjunction with a global analysis of the plasma proteome. We find that the expression of a large number of genes and proteins are altered in the disease model and the majority of these are partially or fully reversed by ERB-041 treatment. Regulated pathways include the acute-phase response, eicosanoid synthesis, fatty acid metabolism, and iron metabolism. In addition, many of the regulated genes and proteins are known to be dysregulated in human rheumatoid arthritis, providing further evidence that the manifestations of the Lewis rat adjuvant-induced arthritis model bear similarity to the human disease.

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