Azacytidine Inhibits RNA Methylation at DNMT2 Target Sites in Human Cancer Cell Lines

Abstract

The cytosine analogues azacytidine and decitabine are currently being developed as drugs for epigenetic cancer therapy. Although various studies have shown that both drugs are effective in inhibiting DNA methylation, it has also become clear that their mode of action is not limited to DNA demethylation. Because azacytidine is a ribonucleoside, the primary target of this drug may be cellular RNA rather than DNA. We have now analyzed the possibility that azacytidine inhibits the RNA methyltransferase DNMT2. We found that DNMT2 is variably expressed in human cancer cell lines. RNA bisulfite sequencing showed that azacytidine, but not decitabine, inhibits cytosine 38 methylation of tRNA Asp , a major substrate of DNMT2. Azacytidine caused a substantially stronger effect than decitabine on the metabolic rate of all the cancer cell lines tested, consistent with an effect of this drug on RNA metabolism. Of note, drug-induced loss of RNA methylation seemed specific for DNMT2 target sites because we did not observe any significant demethylation at sites known to be methylated by other RNA methyltransferases. Our results uncover a novel and quantifiable drug activity of azacytidine and raise the possibility that tRNA hypomethylation might contribute to patient responses.

Inhibiting DNA Methylation Causes an Interferon Response in Cancer via dsRNA Including Endogenous Retroviruses

“…The induced responses may not be a general stress phenomenon as they do not occur with carboplatin, a cytotoxic agent commonly used in EOC treatment (Figure 1b). Aza and Dac incorporate into DNA, inhibiting 3 DNA methyltransferases (Schaefer et al, 2009), but Aza also incorporates into RNA, inhibiting the RNA methyltransferase DNMT2. Aza thus can demethylate RNA, and unmethylated RNA may activate TLR3 and the interferon response (Kariko et al, 2005).…”

Perspectives and future directions for epigenetics in hematology

“…60 Indeed, Lyko and colleagues have shown that 5-azacitidine is incorporated into RNA species and results in hypomethylation of transfer RNA. 61 Although dozens of covalently modified RNA species are recognized currently, studying how 5-azacitidine changes RNA species is technically challenging, and we suggest this area is ripe for reexamination. 5-azacitidine-modified RNA could have an even greater impact than in DNA.…”

Enhanced in Vitro Anti-Tumor Activity of 5-Azacytidine by Entrapment into Solid Lipid Nanoparticles

“…5 Furthermore, 5-azacytidine inhibits protein production by being incorporated into RNA and interrupting its performance. 6,7 Although a few investigations have illustrated the effectiveness of experiments, 8,9 most clinical trials of the effects of 5-azacytidine on solid tumors are disappointing. 10 Due to its short half-life, 5-azacytidine is unable to permeate solid tumors and remain long enough to affect cancer cells.…”