Medicinal Chemistry - Volume 1, Issue 2, 2005

The synthesis of C-12 deuterated and tritiated deoxoartemisinins is described. Tritiated deoxoartemisinin which could be incubated with the protein of Plasmodium falciparum for binding mechanism study was obtained by direct reduction of the carbonyl group of artemisinin using NaBT4 and BF3×Et2O in dried THF.

The membrane-embedded, ligand-gated P2X glycoprotein receptor is a monovalent-bivalent cation channel that is activated by physiological concentrations of extracellular ATP. A quantitative structure-activity relationship (QSAR) analysis was developed to model the cation permeability of the P2X2 channel and its mutants. As chemical properties, the helix-coil equilibrium constants and the distribution coefficients of the system octanol/water at pH 7.4 were applied and modified (sliding windows) according to Eroshkin et al. (Comput. Appl. Biosci., 1995, 11, 49-44). The results were visualized by a dimeric P2X2 channel construct. The results support the hypothesis that residues which put into the cavity and contribute to hydrogen bonding forces are involved to a control of the transport of hydrated cations through the P2X2 channel. The model may be useful to develop P2X2 receptor antagonists.

A novel series of 5-substituted-3-(1H-pyrrol-2-yl)-2-oxazolidinones 2a-s has been described as pyrrole analogues of toloxatone and befloxatone, two phenyl-oxazolidinones active as anti-MAO agents and used in antidepressant therapy. Tested against MAO-A and MAO-B enzymes, the majority of 2a-s show highly potent inhibitory effect against the A isoform of the enzyme, with Ki values in the range 0.52-0.004 μM, whilst their anti-MAO-B activity is considerably lower (Ki = >100-0.5 μM). Structurally, 2a-s differs for the substituent inserted at the C5 position of the 2- oxazolidinone ring (hydroxymethyl (2a-d), methoxymethyl (2e-h), azidomethyl (2i-l), methylaminomethyl (2m-p), and aminomethyl (2q-s)), and the size of the alkyl chain at the pyrrole N1 position (methyl, ethyl, allyl, or benzyl). As a rule, apart from the C5 substitution, the bulkier is the alkyl group at the pyrrole-N1, the lower is the anti-MAO-A activity of the compounds, being the N1-methyl derivatives 2a, 2e, 2i, and 2q among the most potent (KiMAO-A = 0.087-0.004 μM) and A-selective (A-selectivity ratio: >11,111-41) compounds in this series. Exceptions are represented by the N1-benzyl derivative 2d (KiMAO-A = 0.009 μM) and the N1-allylpyrrole 2o (KiMAO-A = 0.04 μM). In comparison with the reference drugs, these highly active derivatives are more potent than toloxatone, slightly less potent than befloxatone, and several times more A-selective than both the references. Such results indicate that 2a-s may represent a new promising series of antidepressant agents.

Recent advances in the design and preclinical evaluations of promising new generation taxane anticancer agents are reviewed in this article. Paclitaxel and docetaxel are two of the most important anticancer drugs today. However, recent reports have shown that treatment with these drugs often encounters undesirable side effects as well as drug resistance. Therefore, it is important to develop new taxane anticancer agents with fewer side effects, superior pharmacological properties, and improved activity against drug-resistant human cancers. Structure-activity relationship (SAR) studies led to the discovery of a series of highly active second-generation taxanes. One of them, “Ortataxel” (SB-T-101131, IDN5109, BAY59-8862), exhibits excellent activity against a variety of drugsensitive and drug-resistant cancer cell lines, as well as human tumor xenografts in mice. It is orally active and is currently in phase II clinical trials. Photoaffinity labeling of microtubules and P-glycoprotein using photoreactive radiolabeled taxoids has disclosed the drugbinding domain of tubulin as well as Pgp. Together with information on microtubule-bound fluorine-labeled taxoids obtained by solid-state NMR studies, the bioactive conformation of paclitaxel and taxoids appears to emerge. Novel taxane-monoclonal antibody (mAb) immunoconjugates, have shown highly promising results for the tumor-specific delivery and release of an extremely cytotoxic, second-generation taxane. Also, another novel series of second generation taxanes conjugated with n-3 polyunsaturated fatty acids, e.g. decosahexaenoic acid (DHA), has exhibited impressive antitumor activity with minimum general toxicity against the highly drug-resistant DLD-1 human colon cancer xenografts in SCID mice.

A metabolically stable and centrally acting analog of pGlu-Glu-Pro-NH2 ([Glu2]TRH, a tripeptide structurally related to TRH (thyrotropin-releasing hormone)) was designed by replacing the amino-terminal pyroglutamyl residue with a pyridinium moiety. The analeptic action of the analog was used to optimize the efficacy of this novel CNS agent when administered intravenously in its CNS-permeable prodrug forms obtained via the reduction of the pyridinium moiety to the nonionic dihydropyridine and esterifying the central Glu with various alcohols. The maximum effect in antagonizing pentobarbital-induced narcosis in mice was achieved with the hexyl ester that was used subsequently for a comparative evaluation with a prodrug of the parent neuropeptide in the Porsolt swim test as a paradigm for antidepressant effect. The novel analog maintained its antidepressant potency but showed reduced analeptic action compared to [Glu2]TRH; thus, an increase in the selectivity of CNS-action was obtained by the incorporation of the pyridinium moiety.

Negatively charged gold nanoparticles featuring 2-(10-mercapto-decyl)-malonic acid were synthesized using the Murray place-displacement reaction. These water-soluble malonate gold mixed monolayer protected clusters (MMPCs) effectively bind and inhibit chymotrypsin based on complementary electrostatic surface recognition. The effect of increasing ionic strength on inhibition was also studied. It was observed that addition of high ionic strength solutions to protein-nanoparticle complexes show almost complete restoration of protein activity. The conformational change of chymotrypsin upon binding to the MMPC was investigated using fluorescence spectrometry and circular dichroism, thus correlating structural changes with enzyme activity.

Six analogs of bidentate 1-[pyridoxylidene]-2-phenyl]hydrazine, twelve analogs of N2O-tridentate 1- [pyridoxylidene]-2-[heteroaryl]hydrazine, and four O2N-tridentate analogs of 1-[pyridoxylidene]-2-[heteroaroyl] hydrazines were synthesized and characterized. Their solutions in water and DMSO were assayed in vitro for activity against a chloroquine-resistant species of P. falciparum obtained from Hadassah Hospital Blood Bank in Jerusalem. The O2N-tridentate group was essentially inactive, whereas the bidentate group, with N and O liganding atoms, exhibited slight activity against late-stage trophozoites and schizonts of P. falciparum. The N2O-tridentate group, by contrast, was remarkably active against resistant P. falciparum, highlighting the importance of the Denticity Effect in this system. It is assumed that the pyridoxal-based chelator acts as an iron redox mediator, controlling the first coordination sphere and, therefore, the immediate chemical environment of the iron.Chelation of iron-(II) presumably facilitates its oxidation..The Fe(II) (r) Fe(III) intra-electron transfer, may be viewed as a switch (“redox switch”), controlling the thermodynamic stability and kinetic lability of the coordination shell. The redox-switch is accompanied by the appearance of a carbonbased Fe-(III)-chelate radical, capable of donating its free electron to the parasite-DNA, thus causing death. The antimalarial N2O-tridentate Fe(III)-chelates appear to be prone to redox-switch, and tend to be converted into their Fe(II) species, whereas the inactive O2N-tridentate analogs apparently cannot do so.

Quantitative Structure Activity Relationship (QSAR) techniques are used routinely by computational chemists in drug discovery and development to analyze datasets of compounds. Quantitative numerical methods like Partial Least Squares (PLS) and Artificial Neural Networks (ANN) have been used on QSAR to establish correlations between molecular properties and bioactivity. However, ANN may be advantageous over PLS because it considers the interrelations of the modeled variables. This study focused on the HIV-1 Protease (HIV-1 Pr) inhibitors belonging to the peptidomimetic class of compounds. The main objective was to select molecular descriptors with the best predictive value for antiviral potency (Ki). PLS and ANN were used to predict Ki activity of HIV-1 Pr inhibitors and the results were compared. To address the issue of dimensionality reduction, Genetic Algorithms (GA) were used for variable selection and their performance was compared against that of ANN. Finally, the structure of the optimum ANN achieving the highest Pearson;s-R coefficient was determined. On the basis of Pearson;s-R, PLS and ANN were compared to determine which exhibits maximum performance. Training and validation of models was performed on 15 random split sets of the master dataset consisted of 231 compounds. For each compound 192 molecular descriptors were considered. The molecular structure and constant of inhibition (Ki) were selected from the NIAID database. Study findings suggested that non-covalent interactions such as hydrophobicity, shape and hydrogen bonding describe well the antiviral activity of the HIV-1 Pr compounds. The significance of lipophilicity and relationship to HIV-1 associated hyperlipidemia and lipodystrophy syndrome warrant further investigation.

To clarify the biological role of the 90K/Mac-2BP glycoprotein, we evaluated the ability of two MAbs SP-2 and 1A4.22, to reveal this glycoprotein in both serum and tissue from hepatocellular carcinoma (HCC) patients. Tissue expression of 90K was detected by the immunohistochemical method in 20 HCC patients, while the 90K serum level was assessed by the ELISA assay in 13 HCC patients. MAb SP-2 was reactive only in serum, with a mean value of 12.8± 6.7 μg/ml . On the contrary, MAb 1A4.22 revealed immunoreactivity both in 92% of sera and in 60% of neoplastic samples. Positive staining was seen only in the epithelial cells and was cytoplasmic and granular in all instances. The mean 90K serum level assayed with MAb 1A4.22 was 29.4 ± 13.7 μg/ml. .Patients with a 90K serum level ≤ 30 mg/ml had positive tissue samples in 71% of cases versus 20% of patients with a serum level > 30 μg/ml. Moreover, a possible poor prognostic role was observed for negative 90K in tissue. Our results suggest that only MAb 1A4.22 could demonstrate 90K glycoprotein expression in paraffin-embedded tissue and that this MAb could have a diagnostic and prognostic role in both sera and tissues from HCC patients.

Cancer chemotherapy often fails due to acquired drug resistance. One of the most critical biochemical changes observed in drug-resistant tumor cells is over-expression of glutathione S-transferase Pi isozyme (GSTP1). Glutathione Stransferase inhibitors have been used as potentiating agents of chemotherapeutic drugs. Earlier we reported haloenol lactone 1 as a site-directed GSTP1 inactivator. We proposed that enzymatic hydrolysis of the haloenol lactone may be the initial step of GSTP1 chemical modification, resulting in the inactivation of the enzyme. Enzyme inactivation is initiated through addition of Cys-47 to the lactone ring, which is opened in the process to form an α-bromoketone adduct. The acidity of Cys-47 confers good leaving group properties, and rapid hydrolysis occurs to generate an α-bromoketoacid intermediate. The reaction may proceed via alkylation of the transient thioester to form a six-membered ring episulfonium ion intermediate which would be yet more reactive toward hydrolysis, with either process leading to the observed mass increase of 230 Da. To probe the importance of the bromine of the lactone in GST inactivation, we designed and synthesized compound 2. Unlike lactone 1, lactone 2 did not show time-dependent inhibitory effect on GSTP1. Incubation of compounds 1 and 2 with excess of N-acetyl cysteine produced the corresponding di-N-acetyl cysteine conjugate and mono-N-acetyl cysteine conjugate, respectively. To probe the role of Cys-47 in the inactivation of GSTP1 by compound 1, we prepared mutant C47A GSTP1. The mutant GSTP1 still showed good activity toward CDNB, but it lost susceptibility to the inactivation by compound 1. In addition, LC-MS/MS technique allowed us to identify the modified Cys-47 after the enzyme was exposed to compound 1.

The SELEX technique (systematic evolution of ligands by exponential enrichment) is a combinatorial library approach in which DNA or RNA molecules are selected by their ability to bind their protein targets with high affinity and specificity. The isolated molecules are referred to as aptamers (from aptus = Latin “to fit”). First, RNA and DNA aptamers were identified that bind to proteins naturally interacting with nucleic acids, or to small organic molecules such as ATP. In the following years, the use of the SELEX technique was extended to isolate oligonucleotide ligands for a wide range of proteins of importance for therapy, and diagnostics. Since these RNA and DNA molecules bind their targets with similar affinities as antibodies, and are able to distinguish between isotypes of an enzyme, aptamers have been also called synthetic antibodies. Recently, the use of in vitro selection methods to isolate protein inhibitors has been extended to complex targets, such as receptors that are only functional in their membrane-bound form, cells, and trypanosomes. RNA aptamers have been expressed in living cells where they inhibit a protein implicated in intracellular signal transduction. The utility of aptamers for in vivo experiments, and diagnostic and therapeutic purposes, is considerably enhanced by introducing chemical modifications into the oligonucleotides to provide resistance against enzymatic degradation in body fluids. Recently, such inhibitors have been evolved for a great variety of targets, including receptors, growth factors, and adhesion molecules implicated in disease. Furthermore, some results were already obtained in animal models and clinical trials.