The recently discovered enzyme lysine-specific demethylase 1 (LSD1) plays an important role in the epigenetic control of gene expression, and aberrant gene silencing secondary to LSD1 over expression is thought to contribute to the development of cancer. We recently reported a series of (bis)guanidines and (bis)biguanides that are potent inhibitors of LSD1, and induce the reexpression of aberrantly silenced tumor suppressor genes in tumor cells in vitro. We now report a series of isosteric ureas and thioureas that are also potent inhibitors of LSD1. These compounds induce increases in methylation at the histone 3 lysine 4 (H3K4) chromatin mark, a specific target of LSD1, in Calu-6 lung carcinoma cells. In addition, these analogues increase cellular levels of secreted frizzle-related proteins (SFRP) 2 and 5, and transcription factor GATA4. These compounds represent an important new series of epigenetic modulators with the potential for use as antitumor agents.

The recently discovered enzyme lysine-specific demethylase 1 (LSD1) plays an important role in the epigenetic control of gene expression, and aberrant gene silencing secondary to LSD1 dysregulation is thought to contribute to the development of cancer. We reported that (bis)guanidines, (bis)biguanides and their urea- and thiourea isosteres are potent inhibitors of LSD1, and induce the re-expression of aberrantly silenced tumor suppressor genes in tumor cells in vitro. We now report a series of small molecule amidoximes that are moderate inhibitors of recombinant LSD1, but that produce dramatic changes in methylation at the histone 3 lysine 4 (H3K4) chromatin mark, a specific target of LSD1, in Calu-6 lung carcinoma cells. In addition, these analogues increase cellular levels of secreted frizzle-related protein (SFRP) 2, H-cadherin (HCAD) and transcription factor GATA4. These compounds represent leads for an important new series of drug-like epigenetic modulators with the potential for use as antitumor agents.

Activation of spermine/spermidine-N 1 -acetyltransferase (SSAT) leads to DNA damage and growth arrest in mammalian cells, and its ablation reduces the severity of ischemic and endotoxic injuries. Here we have examined the role of SSAT in the pathogenesis of toxic liver injury caused by carbon tetrachloride (CCl 4). The expression and activity of SSAT increase in the liver subsequent to CCl 4 administration. Furthermore, the early liver injury after CCl 4 treatment was significantly attenuated in hepatocyte-specific SSAT knockout mice (Hep-SSAT-Cko) compared with wild-type (WT) mice as determined by the reduced serum alanine aminotransferase levels, decreased hepatic lipid peroxidation, and less severe liver damage. Cytochrome P450 2e1 levels remained comparable in both genotypes, suggesting that SSAT deficiency does not affect the metabolism of CCl 4. Hepatocyte-specific deficiency of SSAT also modulated the induction of cytokines involved in inflammation and repair as well as leukocyte infiltration. In addition, Noxa and activated caspase 3 levels were elevated in the livers of WT compared with Hep-SSAT-Cko mice. Interestingly, the onset of cell proliferation was significantly more robust in the WT compared with Hep-SSAT Cko mice. The inhibition of polyamine oxidases protected the animals against CCl4-induced liver injury. Our studies suggest that while the abrogation of polyamine back conversion or inhibition of polyamine oxidation attenuate the early injury, they may delay the onset of hepatic regeneration. hepatotoxicity; carbon tetrachloride; polyamine POLYAMINES INTERACT WITH NUCLEIC acids and proteins and through these interactions play important roles in gene transcription, signal transduction, and cell proliferation (6,12,14,15,20). The cellular levels of polyamines are tightly regulated through import, export, synthesis and catabolism (Fig. 1). The initial step in polyamine synthesis is the decarboxylation of ornithine by ornithine decarboxylase (ODC) to form putrescine (Put). Sequential addition of aminopropyl residues to Put and spermidine (Spd) lead to the formation of Spd and spermine (Spm). Polyamines are degraded through their back conversion by Spm/Spd-N 1 -acetyltransferase (SSAT) and N 1 -acetylpolyamine oxidase (APAO) or by oxidation of Spm by Spm oxidase (SMO). Oxidation of acetylated Spm and Spd by APAO and Spm by SMO generates cytotoxic molecules such as H 2 O 2 and aldehydes (i.e., 3-aminopropanal and 3-acetoaminopropanal; Fig. 1).The expression and activity of SSAT increases in organs (e.g., liver, kidney, and brain) subjected to ischemia-reperfusion and septic and traumatic injuries (4,35,37,39). The ablation of the SSAT gene reduces the severity of renal and hepatic ischemia-reperfusion and endotoxin-induced renal injuries (35,37). Transgenic animals that express high levels of SSAT develop several pathologies, including skin lesions and pancreatitis (1,24,25). In vitro, expression of SSAT causes oxidative stress, DNA damage, cell cycle arrest, and apoptosis (7, 11). These results sugg...