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Dysfunction or progressive degeneration of retinal pigment epithelium (RPE) contributes in the initial pathogenesis of age-related macular degeneration (AMD) causing irreversible vision loss, which makes RPE the prime target of the disease. The present study aimed to identify compounds to protect 4-hydroxynonenal (4-HNE)-induced RPE cell death by inhibiting NADPH oxidase 4 (NOX4) activity, not just as free radical scavengers, using ARPE-19, a human adult retinal pigment epithelial cell line, as a RPE representative. Novel thirty-two 6-ureido/thioureido-2,4,5-trimethylpyridin-3-ol derivatives 17 were synthesized and tested. We found that there was a strong correlation between level of protective effect of compounds 17 against 4-HNE-induced APRE-19 cell death and that of inhibitory activity against 4-HNE-induced superoxide production, and that most of the compounds 17 showed minimal DPPH radical scavenging activity. Compound 17–28 showed the best protective activity against 4-HNE-induced superoxide production (79.5% inhibition) and cell death (85.1% recovery) at 10 μM concentration, which was better than that of VAS2870, a NOX2/4 inhibitor. In addition, compound 17–28 blocked 4-HNE-induced apoptosis of ARPE-19 cells in a concentration-dependent manner. The results indicate that compound 17–28 may be a lead compound to develop AMD therapeutics.

Tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) are the critical pro-inflammatory cytokines involved in the pathogenesis of inflammatory bowel disease (IBD). Inhibition of these cytokines and related signaling pathways has been a target for the development of IBD therapeutics. In the current study, 6-acetamido-2,4,5-trimethylpyridin-3-ol (1) and various analogues with the amido scaffold were synthesized and examined for their inhibitory activities in in vitro and in vivo IBD models. The parent compound 1 (1 μM) showed an inhibitory activity against TNF-α- and IL-6-induced adhesion of monocytes to colon epithelial cells, which was similar to tofacitinib (1 μM), a JAK inhibitor, but much better than mesalazine (1,000 μM). All the analogues showed a positive relationship (R2 = 0.8943 in a linear regression model) between the inhibitory activities against TNF-α-induced and those against IL-6-induced adhesion. Compound 2–19 turned out to be the best analogue and showed much better inhibitory activity against TNF-α- and IL-6-induced adhesion of the cells than tofacitinib. In addition, oral administration of compound 1 and 2–19 resulted in a significant suppression of clinical signs of TNBS-induced rat colitis, including weight loss, colon tissue edema, and myeloperoxidase activity, a marker for inflammatory cell infiltration in colon tissues. More importantly, compound 2–19 (1 mg/kg) was more efficacious in ameliorating colitis than compound 1 and sulfasalazine (300 mg/kg), the commonly prescribed oral IBD drug. Taken together, the results suggest that compound 2–19 can be a novel platform for dual-acting IBD drug discovery targeting both TNF-α and IL-6 signaling.

We recently reported that 6-aminoalkyl-2,4,5-trimethylpyridin-3-ols, novel series of 6-aminopyridin-3-ol-based antioxidants, have high antiangiogenic activities. In pursuit of wider variety in the analogues, we here report the synthesis and antiangiogenic activities of 6-amidoalkyl-2,4,5-trimethylpyridin-3-ols, which would not be considered excellent antioxidants because of the poorer electron-donating effect of the C(6)-amido group than the corresponding C(6)-amino group. The selected 6-amido compounds showed up to several fold-higher antiangiogenic activities and up to an order of magnitude better antitumor activities in the chick embryo chorioallantoic membrane (CAM) assay than SU4312, a positive control. We also found that paracetamol, as a direct phenolic analogue of our simplest 6-amidopyridin-3-ol, showed a moderate level of antiangiogenic activity. We propose this study will offer a basis for a scaffold of novel angiogenesis inhibitors that can perturb angiogenesis-related pathologies.