Redox-active cysteine, a highly reactive sulfhydryl, is one of the major targets of ROS. Formation of disulfide bonds and other oxidative derivatives of cysteine including sulfenic, sulfinic, and sulfonic acids, regulates the biological function of various proteins. We identified novel lowabundant cysteine modifications in cellular GAPDH purified on 2-dimensional gel electrophoresis (2D-PAGE) by employing selectively excluded mass screening analysis for nano ultraperformance liquid chromatography-electrospray-quadrupole-time of flight tandem mass spectrometry, in conjunction with MOD i and MODmap algorithm. We observed unexpected mass shifts (⌬m ‫؍‬ ؊16, ؊34, ؉64, ؉87, and ؉103 Da) at redox-active cysteine residue in cellular GAPDH purified on 2D-PAGE, in oxidized NDP kinase A, peroxiredoxin 6, and in various mitochondrial proteins. Mass differences of ؊16, ؊34, and ؉64 Da are presumed to reflect the conversion of cysteine to serine, dehydroalanine (DHA), and Cys-SO 2 -SH respectively. To determine the plausible pathways to the formation of these products, we prepared model compounds and examined the hydrolysis and hydration of thiosulfonate (Cys-S-SO 2 -Cys) either to DHA (⌬m ‫؍‬ ؊34 Da) or serine along with Cys-SO 2 -SH (⌬m ‫؍‬ ؉64 Da). We also detected acrylamide adducts of sulfenic and sulfinic acids (؉87 and ؉103 Da). These findings suggest that oxidations take place at redox-active cysteine residues in cellular proteins, with the formation of thiosulfonate, Cys-SO 2 -SH, and DHA, and conversion of cysteine to serine, in addition to sulfenic, sulfinic and sulfonic acids of reactive cysteine.

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Cinnamaldehyde, a major active compound of cinnamon, is known to induce apoptotic cell death in numerous human cancer cells. Here, dual acid-responsive polymeric micelle-forming cinnamaldehyde prodrugs, poly[(3-phenylprop-2-ene-1,1-diyl)bis(oxy)bis(ethane-2,1-diyl)diacrylate]-co -4,4'(trimethylene dipiperidine)-co -poly(ethylene glycol), termed PCAE copolymers, are reported. PCAE is designed to incorporate cinnamaldehyde via acid-cleavable acetal linkages in its pH-sensitive hydrophobic backbone and self assemble to form stable micelles which can encapsulate camptothecin (CPT). PCAE self assembles to form micelles which release CPT and cinnamaldehyde in pHdependent manners. PCAE micelles induce apoptotic cell death through the generation of intracellular reactive oxygen species (ROS) and exert synergistic anticancer effects with a payload of CPT in vitro and in vivo model of SW620 human colon tumor-bearing mice. It is anticipated that dual acid-sensitive micelle-forming PCAE with intrinsic anticancer activities has enormous potential as novel anticancer therapeutics.

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Nucleoside diphosphate kinase (NDPK, Nm23), a housekeeping enzyme, is known to be a multifunctional protein, acting as a metastasis suppressor, transactivation activity on c-myc, and regulating endocytosis. The cellular mechanisms regulating Nm23 functions are poorly understood. In this study, we identified the modifications and interacting proteins of Nm23-H1 in response to oxidative stress. We found that Cys109 in Nm23-H1 is oxidized to various oxidation states including intra- and inter-disulfide crosslinks, glutathionylation, and sulfonic acid formation in response to H2O2 treatment both in vivo and in vitro. The cross-linking sites and modifications of oxidized Nm23-H1 were identified by peptide sequencing using UPLC-ESI-q-TOF tandem MS. Glutathionylation and oxidation of Cys109 inhibited the NDPK enzymatic activity of Nm23-H1. We also found that thioredoxin reductase 1 (TrxR1) is an interacting protein of Nm23-H1, and it binds specifically to oxidized Nm23-H1. Oxidized Nm23 is a substrate of NADPH-TrxR1-thioredoxin shuttle system, and the disulfide crosslinking is reversibly reduced and the enzymatic activity is recovered by this system. Oxidation of Cys109 in Nm23-H1 inhibited its metastatic suppressor activity as well as the enzymatic activities. The mutant, Nm23-H1 C109A, retained both the enzymatic and metastasis suppressor activities under oxidative stress. This suggests that key enzymatic and metastasis suppressor functions of Nm23-H1 are regulated by oxido-reduction of its Cys109.

Skin as a delivery route for drugs has attracted a great attention in recent decades as it avoids many of the limitations of oral and parenteral administration. However, the excellent barrier property of skin is a major obstacle in the effective transport of drugs through this route. The topmost layer of skin, the "stratum corneum" is the tightest one and is responsible for most of the resistance offered. This necessitates breaching the resistance of the stratum corneum reversibly and transiently in order to achieve a therapeutically meaningful level in systemic circulation or local skin. In last few decades, a number of approaches have been developed to improve the limited drug permeability through stratum corneum. One promising approach is the use of nanoparticulate carriers as they not only facilitate drug delivery across skin but also avoid the drawbacks of conventional skin formulations. This review focuses on nanoparticulate carriers including conventional liposomes, deformable liposomes, ethosomes, niosomes and lipid nanoparticles developed for topical and transdermal drug delivery. A special emphasis is placed on their composition, structure, mechanism of penetration and recent application. The presented data demonstrate the potential of these nanoparticulate carriers for dermal and transdermal delivery.

An approach to the computer-assisted, pharmacophore design of nonsteroidal templates for the glucocorticoid receptor (GR) that contained an element of pseudo-C2 symmetry was developed. The enatiomer of the initial design, 1Ra, and not the designed molecule, 1S, showed the desired ligand binding to the GR. The pseudo-C2 symmetry of the template allowed for rapid improvements in GR activity resulting in potent, selective, nonsteroidal GR antagonists, CP-394531 and CP-409069.