A ligand-based model is reported that predicts the Ki values for cytochrome P450 2C9 (CYP2C9) inhibitors. This CoMFA model was used to predict the affinity of 14 structurally diverse compounds not in the training set and appears to be robust. The mean error of the predictions is 6 microM. The experimentally measured Ki values of the 14 compounds range from 0.1 to 48 microM. Leave-one-out cross-validated partial least-squares gives a q2 value of between 0.6 and 0.8 for the various models which indicates internal consistency. Random assignment of biological data to structure leads to negative q2 values. These models are useful in that they establish a pharmacophore for binding to CYP2C9 that can be tested with site-directed mutagenesis. These models can also be used to screen for potential drug interactions and to design compounds that will not bind to this enzyme with high affinity.
4-Ipomeanol (IPO) is a pneumotoxin that is bioactivated to a reactive intermediate that binds to DNA and other cellular macromolecules. Despite over 30 years of research in this area, detailed structural information on the nature of the IPO reactive intermediate is still lacking. In the present study, we reacted IPO with rabbit CYP4B1 in the presence of exogenous nucleophiles and analyzed the products by liquid chromatography/electrospray ionization-mass spectrometry. Coincubation of IPO and rabbit CYP4B1 with glutathione gave rise to multiple products due likely to the presence of both sulfur and nitrogen nucleophiles in the same trapping molecule. Reaction mixtures containing equimolar N-acetyl cysteine (NAC) and N-acetyl lysine (NAL) provided a major NADPH- and CYP4B1-dependent product. A combination of high-resolution mass spectrometry and two-dimensional NMR analysis following large-scale isolation of the biologically derived material provided evidence for an N-substituted cysteinyl pyrrole derivative of IPO, analogous to that characterized previously in model chemical studies conducted with cis-2-butene-1,4-dial. Purified native rabbit lung CYP4B1 and purified recombinant rabbit CYP4B1 produced the trapped NAC/NAL-IPO pyrrole adduct at rates of 600-700 nmol/nmol P450/30 min. A panel of 14 commercially available recombinant human CYPs was also studied, and substantial rates of IPO bioactivation (>100 nmol/nmol/30 min) were observed with CYP1A2, CYP2C19, CYP2D6, and CYP3A4. These studies provide evidence for the formation of an enedial reactive intermediate during CYP-mediated IPO bioactivation, identify multiple human liver P450s capable of IPO bioactivation, and demonstrate that the same reactive intermediate is formed by both rabbit CYP4B1 and human P450s.
Previously, it was shown that beta-elimination of selenocysteine Se-conjugates by rat renal cytosol leading to pyruvate formation was not solely catalyzed by pyridoxal phosphate-dependent enzymes. It was hypothesized that selenoxidation of the selenocysteine Se-conjugates, followed by syn-elimination, may be an alternative mechanism for pyruvate formation. In this study, selenoxidation of selenocysteine Se-conjugates was studied using rat liver microsomes and recombinant human oxidative enzymes. For all six selenocysteine Se-conjugates that were tested, it was found that rat liver microsomal incubations led to the formation of pyruvate, whereas the corresponding selenoxides were not observed. Microsomal pyruvate formation from Se-benzyl-L-selenocysteine (SeBC) was NADPH-dependent, but only marginally inhibited by several P450 inhibitors. Inhibition by methimazole and by heat pretreatment and stimulation by n-octylamine indicated that flavin-containing monooxygenases are mainly responsible for pyruvate formation from the selenocysteine Se-conjugates in rat liver microsomes. In the case of S-benzyl-L-cysteine, the sulfur analogue of SeBC, pyruvate formation was not observed. For this substrate, a chemically stable sulfoxide could be observed, as previously described. By using recombinantly expressed human flavin-containing monooxygenases and P450 enzymes, it was delineated that SeBC is selenoxidized by human FMOs, but not by human P450s. The k(cat)/K(m) of selenoxidation was 3.8-fold higher for FMO-1 than for FMO-3. In conclusion, selenoxidation of selenocysteine Se-conjugates catalyzed by FMOs and subsequently syn-elimination has taken place as an alternative route for the formation of pyruvate from selenocysteine Se-conjugates. Although selenoxides are known to be easily reduced by thiol compounds, microsomal pyruvate formation from SeBC was only 75% inhibited in the presence of an excess of glutathione. This indicates that even in the presence of physiological concentrations of reducing thiol compounds, selenoxides of selenocysteine Se-conjugates may undergo syn-elimination to some extent. Whether selenoxides and/or selenenic acids that are formed are involved in the activity of chemopreventive selenocysteine Se-conjugates remains to be established.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.