Nicole A. Kratochwil

A NOVEL DINUCLEAR DIAMINOPLATINUM(II) GLUTATHIONE MACROCHELATE

Both oxidized and reduced glutathione (gamma-L-Glu-L-Cys-Gly) react with the anticancer complex [Pt(en)Cl(2)] to form the bicyclic complex illustrated (en=ethylenediamine). This unprecedented structure, which was determined from extensive NMR experiments, contains a ten-membered macrochelate ring.

cis-[PtCl2(NH3){2-(2-hydroxyethyl)pyridine}] − an Analogue of the Anticancer Drug AMD473: Unusual Hydrolysis Rates and pKa Values for the Diaqua Adduct

The X-ray crystal structure of cis-[PtCl2(NH3){2-(2-hydroxyethyl)pyridine}] shows that it has similar structural features to the 2-picoline analogue, the anticancer drug AMD473 (ZD0473), but undergoes aquation much more rapidly. The diaqua adduct has unusually low pKa values which have been rationalised by molecular modelling. (© Wiley-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002)

Relationships between Reduction Properties and Cancer Cell Growth Inhibitory Activities of cis‐Dichloro‐ and cis‐Diiodo‐Pt(IV)‐ethylenediamines

The chemical reactivities and cancer cell growth inhibitory activities of a new series of cis‐diiodo‐Pt(IV)‐ethylenediamines were compared and contrasted with their cis‐dichloro‐Pt(IV)‐counterparts. cis‐Diiodo‐Pt(IV)‐ethylenediamines bearing various axial ligands (i.e., OH, OAc, OCOCF3, OSO2CH3) were prepared by oxidizing [PtI2(en)] with 30% H2O2 to yield trans,cis‐[PtOH2I2(en)], which was then reacted with either Ac2O, (CF3CO)2O, or (SO2CH3)2O in CH2Cl2. The cis‐diiodo‐Pt(IV) complexes were readily reduced by biological thiols such as L‐cysteine, glutathione (GSH), and bovine serum albumin (BSA) at pH 6.9 and 37 °C; the kinetics of reduction were second‐order with respect to thiol concentration. In contrast, the cis‐dichloro analogues were stable in the presence of GSH. The reduction potentials estimated by means of cyclovoltammetry for the Pt(IV) complexes are useful for obtaining a ranking order of reactivity towards biological thiols; however, the reduction potentials alone cannot be used to predict whether a Pt(IV) complex will be reduced by GSH at biologically relevant concentrations. GSH greatly facilitated the platination of calf thymus DNA by the diiodo‐Pt(IV) complexes, which was >90% complete after 24 h at 37 °C when the ratio of GSH to Pt(IV) was 2:1. DNA‐platination by trans,cis‐[Pt(OH)2I2(en)] and trans,cis‐[Pt(OAc)2I2(en)] were dependent on the presence of GSH while trans,cis‐[Pt(OSO2CH3)2I2(en)] showed 23% DNA platination after 24 h in the absence of GSH. In contrast, the dichloro analogues trans,cis‐[Pt(OH)2Cl2(en)] and trans,cis‐[Pt(OAc)2Cl2(en)] failed to react with DNA in the presence of either low (0.015 mM) to high (3.0 mM) concentrations of GSH. Cell culture experiments with four human cancer cell lines showed that the maximal growth inhibitory activity of the cis‐diiodo‐Pt(IV)‐ethylenediamines was reached within a 24 h exposure to platinum complex, while the dichloro‐Pt(IV) analogues required a much longer drug‐exposure time (i.e., 96 h) to reach maximal activity.

G protein-coupled receptor transmembrane binding pockets and their applications in GPCR research and drug discovery: a survey.

G protein-coupled receptors (GPCRs) share a common architecture consisting of seven transmembrane (TM) domains. Various lines of evidence suggest that this fold provides a generic binding pocket within the TM region for hosting agonists, antagonists, and allosteric modulators. Hence, an automated method was developed that allows a fast analysis and comparison of these generic ligand binding pockets across the entire GPCR family by providing the relevant information for all GPCRs in the same format. This methodology compiles amino acids lining the TM binding pocket including parts of the ECL2 loop in a so-called 1D ligand binding pocket vector and translates these 1D vectors in a second step into 3D receptor pharmacophore models. It aims to support various aspects of GPCR drug discovery in the pharmaceutical industry. Applications of pharmacophore similarity analysis of these 1D LPVs include definition of receptor subfamilies, prediction of species differences within subfamilies in regard to in vitro pharmacology and identification of nearest neighbors for GPCRs of interest to generate starting points for GPCR lead identification programs. These aspects of GPCR research are exemplified in the field of melanopsins, trace amine-associated receptors and somatostatin receptor subtype 5. In addition, it is demonstrated how 3D pharmacophore models of the LPVs can support the prediction of amino acids involved in ligand recognition, the understanding of mutational data in a 3D context and the elucidation of binding modes for GPCR ligands and their evaluation. Furthermore, guidance through 3D receptor pharmacophore modeling for the synthesis of subtype-specific GPCR ligands will be reported. Illustrative examples are taken from the GPCR family class C, metabotropic glutamate receptors 1 and 5 and sweet taste receptors, and from the GPCR class A, e.g. nicotinic acid and 5-hydroxytryptamine 5A receptor.

Effect of thiols exported by cancer cells on the stability and growth-inhibitory activity of Pt(IV) complexes

Purpose: The role of thiols in the reduction of Pt(IV) antitumor agents to Pt(II) is well recognized and it is widely thought that this reaction is required for activity. The sources of extracellular thiols in cell culture have been less studied. The purpose of the present work was to determine whether the stability of Pt(IV) complexes in culture medium can be affected by thiols that are released by cancer cells. Methods: A two-column HPLC assay with UV/visible detection was used to determine the stability of two Pt(IV) complexes in culture medium with and without cells. The kinetics of the thiol release from a human ovarian cancer cell line SK-OV-3 and a human glioblastoma cell line U-87 MG were determined by a modification of the Ellmans method. Results: The stability of a Pt(IV) complex with equatorial iodo ligands, trans,cis-[Pt(en)(OAc)2I2], was dramatically lower in culture medium in the presence of cells than in fresh culture medium, whereas the half-life of the dichloro analog, trans,cis-[Pt(en)(OAc)2Cl2], was somewhat increased. Although both complexes showed similar in vitro cell-growth-inhibitory activity, trans, cis-[Pt(en)(OAc)2Cl2] required a longer incubation time than the iodo analog to reach its maximal effect. The thiol content of the culture medium in the presence of cells was measured after 2 days: the concentrations from cultures of U-87 MG and SK-OV-3 cells were 3.6 ± 0.1 μM and 9.3 ± 0.1 μM respectively, compared to 0.07 ± 0.04 μM in fresh medium. During the rapid growth phase, the extracellular thiol content reached a maximum of 20.0 ± 0.5 μM and 47.8 ± 0.2 μM for U-87 MG and SK-OV-3 cells respectively. Conclusions: These findings show that the culture medium conditioned by cancer cells can influence the stabilities of certain Pt(IV) complexes in cytotoxicity studies.