A disulfide tether stabilizes the block of sodium channels by the conotoxin µO§-GVIIJ.

A cone snail venom peptide, µO§-conotoxin GVIIJ from Conus geographus, has a unique posttranslational modification, S-cysteinylated cysteine, which makes possible formation of a covalent tether of peptide to its target Na channels at a distinct ligand-binding site. µO§-conotoxin GVIIJ is a 35-aa peptide, with 7 cysteine residues; six of the cysteines form 3 disulfide cross-links, and one (Cys24) is S-cysteinylated. Due to limited availability of native GVIIJ, we primarily used a synthetic analog whose Cys24 was S-glutathionylated (abbreviated GVIIJSSG). The peptide-channel complex is stabilized by a disulfide tether between Cys24 of the peptide and Cys910 of rat (r) NaV1.2. A mutant channel of rNaV1.2 lacking a cysteine near the pore loop of domain II (C910L), was >10(3)-fold less sensitive to GVIIJSSG than was wild-type rNaV1.2. In contrast, although rNaV1.5 was >10(4)-fold less sensitive to GVIIJSSG than NaV1.2, an rNaV1.5 mutant with a cysteine in the homologous location, rNaV1.5[L869C], was >10(3)-fold more sensitive than wild-type rNaV1.5. The susceptibility of rNaV1.2 to GVIIJSSG was significantly altered by treating the channels with thiol-oxidizing or disulfide-reducing agents. Furthermore, coexpression of rNaVß2 or rNaVß4, but not that of rNaVß1 or rNaVß3, protected rNaV1.1 to -1.7 (excluding NaV1.5) against block by GVIIJSSG. Thus, GVIIJ-related peptides may serve as probes for both the redox state of extracellular cysteines and for assessing which NaVß- and NaVα-subunits are present in native neurons.

Synthesis, Characterization, and Interaction with Biomolecules of Platinum(II) Complexes with Shikimic Acid-Based Ligands.

Starting from the active ingredient shikimic acid (SA) of traditional Chinese medicine and NH2(CH2) n OH, (n = 2-6), we have synthesized a series of new water-soluble Pt(II) complexes PtL(a-e)Cl2, where L(a-e) are chelating diamine ligands with carbon chain covalently attached to SA (L(a-e) = SA-NH(CH2) n NHCH2CH2NH2; L(a), n = 2; L(b), n = 3; L(c), n = 4; L(d), n = 5; L(e), n = 6). The results of the elemental analysis, LC-MS, capillary electrophoresis, and (1)H, (13)C NMR indicated that there was only one product (isomer) formed under the present experimental conditions, in which the coordinate mode of PtL(a-e)Cl2 was two-amine bidentate. Their in vitro cytotoxic activities were evaluated by MTT method, where these compounds only exhibited low cytotoxicity towards BEL7404, which should correlate their low lipophilicity. The interactions of the five Pt(II) complexes with DNA were investigated by agarose gel electrophoresis, which suggests that the Pt(II) complexes could induce DNA alteration. We also studied the interactions of the Pt(II) complexes with 5'-GMP with ESI-MS and (1)H NMR and found that PtL(b)Cl2, PtL(c)Cl2, and PtL(d)Cl2 could react with 5'-GMP to form mono-GMP and bis-GMP adducts. Furthermore, the cell-cycle analysis revealed that PtL(b)Cl2, PtL(c)Cl2 cause cell G2-phase arrest after incubation for 72 h. Overall, these water-soluble Pt(II) complexes interact with DNA mainly through covalent binding, which blocks the DNA synthesis and replication and thus induces cytotoxicity that weakens as the length of carbon chain increases.

Synthesis, characterization, and in vitro antitumor properties of gold(III) compounds with the traditional Chinese medicine (TCM) active ingredient liriodenine.

Liriodenine, an oxoaporphine alkaloid with anticancer activity isolated from Zanthoxylum nitidum (rutaceous anticancer traditional Chinese medicine), was selected as a bioactive ligand to react with HAuCl(4) and NaAuCl(4) to afford [LH][AuCl(4)] (1) and [AuCl(3)L] (2), respectively (where L is liriodenine). The structures of 1 and 2 were characterized by IR spectroscopy, electrospray ionization mass spectrometry, (1)H-NMR spectroscopy, and elemental analysis. The single-crystal X-ray diffraction analysis of 1 revealed that it is an ionic compound consisting of protonated liriodenine cation [LH](+) and [AuCl(4)](-) anion. The spectroscopic analysis showed that 2 is a coordination compound, in which one liriodenine coordinates to gold via its 7-N donor. In aqueous solution, 1 is relatively stable, but 2 undergoes rapid hydrolysis. The in vitro cytotoxicity towards five human tumor cell lines shows that 1 and 2 manifest roughly similar biological behavior and appreciable antiproliferative properties, with IC(50) values falling in the 2-16 µM range. The flow-cytometric analysis of 1 and 2 suggests that both compounds induced an S-phase arrest. Compounds 1 and 2 significantly poison topoisomerase I in vitro at low concentration (25 µM or less). DNA binding studies indicate that both 1 and 2 interact with DNA mainly via intercalation between the neighboring base pairs of the DNA double helix. Electrostatic interactions of 1 and 2 with the polyanionic DNA phosphate backbone may reinforce the intercalation because both 1 and 2 are composed of planar cationic species.