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Sulforaphane [1-isothiocyanato-4-(methylsülfinyl)butane], a naturally occurring isothiocyanate derived from cruciferous vegetables, is a highly potent inducer of phase 2 cytoprotective enzymes and can protect against electrophiles including carcinogens. oxidative stress, and inflammation. The mechanism of action of sulforaphane is believed to involve modifications of critical cysteine residues of Keap1, which lead to stabilization of Nr12 to activate the antioxidant response element of phase 2 enzymes. However, the dithiocarbamate functional group formed by a reversible reaction between isothiocyanate of sulforaphane and sulfhydryl nucleophiles of Keap1 is kinetically labile, and such modification in intact cells has not yet been demonstrated. Here we designed sulforaphane analogs with replacement of the reactive isothiocyanate by the more gentle electrophilic sulfoxythiocarbamate group that also selectively targets cysteine residues in proteins but forms stable thiocarbamate adducts. Twenty-four sulfoxythiocarbamate analogs were synthesized that retain the structural features important for high potency in sulforaphane analogs: the sulfoxide or keto group and its appropriate distance to electrophilic functional group. Evaluation in various cell lines including hepatoma cells, retinal pigment epithelial cells, and keratinocytes as well as in mouse skin shows that these analogs maintain high potency and efficacy for phase 2 enzyme induction as well as the inhibitory effect on lipopolysaccharide-induced nitric oxide formation like sulforaphane. We further show in living cells that a sulfoxythiocarbamate analog can label Keap1 on several key cysteine residues as well as other cellular proteins offering new insights into the mechanism of chemoprotection. [ABSTRACT FROM AUTHOR]

Isothiocyanates and phenolic antioxidants can prevent cancer through activation of Nrf2 (NF-E2 p45-related factor 2), a transcription factor that controls expression of cytoprotective genes through the antioxidant response element (ARE) enhancer. Using a human mammary MCF7-derived AREc32 reporter cell line, we now report that all-trans retinoic acid (ATRA), and other retinoic acid receptor alpha (RARα) agonists, markedly reduces the ability of Nrf2 to mediate induction of ARE-driven genes by cancer chemopreventive agents including the metabolite of butylated hydroxyanisole, tert-butylhydroquinone (tBHQ). The basal and tBHQ-inducible expression of aldo-keto reductase (AKR) AKR1C1 and AKR1C2 genes, which are regulated by Nrf2, was also repressed by ATRA in AREc32 cells. Antagonists of RARα augmented induction of ARE-driven gene expression by tBHQ, as did knockdown of RARα by using RNAi. The expression of the ARE-gene battery was increased in the small intestine of mice fed on a vitamin A-deficient diet, and this increase was repressed by administration of ATRA. By contrast, in the small intestine of Nrf2 null mice, the expression of ARE-driven genes was not affected by vitamin A status. In MCF7 cells, ATRA did not block the nuclear accumulation of Nr12 but reduced the binding of Nrf2 to the ARE enhancer as a consequence of forming a complex with RARα. These data suggest that cross-talk between Nrf2 and RARα could markedly influence the sensitivity of cells to electrophiles and oxidative stressors and, as a consequence, to carcinogenesis. [ABSTRACT FROM AUTHOR]