Kinetic parameters for the reaction of 1,2,4‐trioxolanes and artemisinin with iron(II): New evidence for the source of antimalarial activity

The Fenton‐like reductive cleavage of antimalarial peroxides like artemisinin by iron(II) species is a chemical reaction whose mechanistic pathway has not been yet fully understood; it is, however, known that there is considerable production of radical species centered at both the oxygen and carbon, which are important to the therapeutical effects of those compounds. This article reports kinetic data for the reaction of artemisinin and two model 1,2,4‐trioxolanes with iron(II) species and also a mechanistic interpretation of this reductive cleavage from transition state thermodynamics. The suggestion of the presence of an enhancing specific factor inside the plasmodium is made.     

Bax-associated mechanisms underlying the response of embryonic cells to methotrexate

Bax was shown previously to regulate apoptotic cell death in various experimental systems, however, its involvement in teratogen-induced apoptosis is not clear yet. Therefore, we explored the involvement of Bax in the response of mouse embryonic fibroblasts (MEFs) to the anti cancer drug methotrexate (MTX), using Bax wild type (WT) and knockout (Bax^−/−) MEFs. Our results demonstrated a significant teratogen-induced dose- and time-dependant decrease in the survival and culture density of both cell lines, which were found to be somewhat more prominent in WT cells. Exposure to MTX resulted also in decreased cell proliferation of WT but not Bax^−/− cells and accordingly, we observed an accumulation of cells in the S phase and an increased percentage of cells in the Sub-G₁ phase of the cell cycle and the appearance of condensed nuclei, which were found to be somewhat more prominent in WT MEFs. In parallel, WT MEFs demonstrated a MTX-induced increase in the percentage of Bax-positive cells and a significant decrease in the percentage of bcl-2-, p65- or IκBα-positive cells, which were not detected in Bax^−/− MEFs. Altogether, the differential sensitivity of WT or Bax^−/− MEFs to MTX suggests a possible involvement of this molecule in the response of embryonic cells to teratogens.     

From Zn to Mn: the study of novel manganese-binding groups in the search for new drugs against tuberculosis

In most eubacteria, apicomplexans, and most plants, including the causal agents for diseases such as malaria, leprosy, and tuberculosis, the methylerythritol phosphate pathway is the route for the biosynthesis of the C(5) precursors to the essential isoprenoid class of compounds. Owing to their absence in humans, the enzymes of the methylerythritol phosphate pathway have become attractive targets for drug discovery. This work investigates a new class of inhibitors against the second enzyme of the pathway, 1-deoxy-D-xylulose 5-phosphate reductoisomerase. Inhibition of this enzyme may involve the chelation of a crucial active site Mn ion, and the metal-chelating moieties studied here have previously been shown to be successful in application to the zinc-dependent metalloproteinases. Quantum mechanics and docking calculations presented in this work suggest the transferability of these metal-chelating compounds to Mn-containing 1-deoxy-D-xylulose 5-phosphate reductoisomerase enzyme, as a promising starting point to the development of potent inhibitors.     

Effects of a glyphosate‐based herbicide on the uterus of adult ovariectomized rats

Glyphosate is the active ingredient of several herbicide formulations. Different reports suggest that glyphosate‐based herbicides (GBHs) may act as endocrine disruptors. We evaluated the potential estrogenic effects of a GBH formulation using the uterotrophic assay. Adult ovariectomized rats were sc injected for 3 consecutive days with: saline solution (vehicle control), 2.10^?5 g E₂/kg/day (uterotrophic dose; UE₂), 2.10^?7 g E₂/kg/day (nonuterotrophic dose; NUE₂), or 0.5, 5, or 50 mg GBH/kg/day of the. Twenty‐four hours after the last injection, the uterus was removed and weighed and processed for histopathology and mRNA extraction. Epithelial cell proliferation and height and expression of estrogen‐responsive genes were evaluated (estrogen receptors, ERα and ERβ; progesterone receptor, PR; complement 3, C3). Uterine weight and epithelial proliferation were not affected by GBH. However, the luminal epithelial cell height increased at GBH0.5. ERα mRNA was downregulated by all GBH doses and E₂ groups, whereas PR and C3 mRNA were diminished by GBH0.5. GBH5‐, GBH50‐, and UE₂‐treated rats showed downregulated ERα protein expression in luminal epithelial cells, while the receptor was upregulated in the stroma. GBH upregulated ERβ (GBH0.5–50) and PR (GBH5) expressions in glandular epithelial cells, similar effect to that of NUE₂ group. These results indicate that, although the uterine weight was not affected, GBH modulates the expression of estrogen‐sensitive genes. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1191–1201, 2017.     

Association of HMGB1 and HMGB2 genetic polymorphisms with lung cancer chemotherapy response

The aim of the present study was to investigate the association of genetic polymorphisms in high mobility group box 1 and 2 (HMGB1 and HMGB2, respectively) with platinum‐based chemotherapy responses in Chinese lung cancer patients. In total, 338 Chinese lung cancer patients (154 responders and 184 non‐responders) were recruited to the study. All patients received at least two cycles of first‐line platinum‐based chemotherapy. Three tagging single nucleotide polymorphisms (SNPs) of HMGB1 and two tagging SNPs of HMGB2 were detected in patients. We found that rs1412125 and rs2249825 of HMGB1 were significantly associated with the platinum‐based chemotherapy response in both recessive and genotypic models. In addition, rs1412125 showed significant association with platinum‐based chemotherapy response for the subgroup of patients aged >55 years in additive, recessive and genotypic models. No significant associations were detected between other SNPs and the platinum‐based chemotherapy response. The HMGB1 SNPs (rs1412125 and rs2249825) were associated with platinum‐based chemotherapy responses in Chinese lung cancer patients. In conclusion, HMGB1 SNPs may serve as potential biomarkers for predicting the efficacy of platinum‐based chemotherapy.     

Pharmacokinetics of dietary cancer chemopreventive compound dibenzoylmethane in rats and the impact of nanoemulsion and genetic knockout of Nrf2 on its disposition

The pharmacokinetic disposition of a dietary cancer chemopreventive compound dibenzoylmethane (DBM) was studied in male Sprague‐Dawley rats after intravenous (i.v.) and oral (p.o.) administrations. Following a single i.v. bolus dose, the mean plasma clearance (CL) of DBM was low compared with the hepatic blood flow. DBM displayed a high volume of distribution (Vss). The elimination terminal t_1/2 was long. The mean CL, Vss and AUC_0?∞/dose were similar between the i.v. 10 and 10 mg/kg doses. After single oral doses (10, 50 and 250 mg/kg), the absolute oral bioavailability (F^*) of DBM was 7.4%–13.6%. The increase in AUC was not proportional to the oral doses, suggesting non‐linearity. In silico prediction of oral absorption also demonstrated low DBM absorption in vivo. An oil‐in‐water nanoemulsion containing DBM was formulated to potentially overcome the low F^* due to poor water solubility of DBM, with enhanced oral absorption. Finally, to examine the role of Nrf2 on the pharmacokinetics of DBM, since DBM activates the Nrf2‐dependent detoxification pathways, Nrf2 wild‐type (+/+) mice and Nrf2 knockout (?/?) mice were utilized. There was an increased systemic plasma exposure of DBM in Nrf2 (?/?) mice, suggesting that the Nrf2 genotype could also play a role in the pharmacokinetic disposition of DBM. Taken together, the results show that DBM has low oral bioavailability which could be due in part to poor water solubility and this could be overcome by a nanotechnology‐based drug delivery system and furthermore the Nrf2 genotype could also play a role in the pharmacokinetics of DBM. Copyright © 2010 John Wiley & Sons, Ltd.     

Syntheses of four peptides from the immunodominant region of hepatitis C viral pathogens using PS‐TTEGDA support for the investigation of HCV infection in human blood

Abstract: Four peptides were designed and synthesized on a highly solvating copolymer of tetraethyleneglycol diacrylate cross‐linked polystyrene (PS‐TTEGDA) support with very high purity and yield. The polymer was synthesized in various cross‐linking densities (1, 2, 3, 4, 5 and 10%) using radical aqueous suspension polymerization. Four per cent PS‐TTEGDA resin showed rigidity and mechanical characteristics comparable with those of divinylbenzene cross‐linked polystyrene (PS‐DVB) support. Swelling and solvation characteristics of PS‐TTEGDA were much higher than PS‐DVB support in all solvents used in solid‐phase peptide synthesis. Forty‐eight hour treatment of the support with neat trifluoroacetic acid did not show any change in its infrared spectra. PS‐TTEGDA could be functionalized with chloromethyl, aminomethyl and hydroxymethyl functional groups under various controlled conditions. Synthetic utility of the support was demonstrated by the synthesis of four peptides selected from the envelope and nonstructural protein region of the prototype hepatitis C virus (HCV). These peptides were later used successfully to develop a peptide‐based immunoassay (PBEIA) for the detection of HCV immunity. Peptides designed from the NS1 and NS4 protein regions were found to be very promising for the development of a new diagnostic kit to detect HCV infection in human blood. Peptide purity was tested by RP‐FPLC and the peptide identity was confirmed by amino acid analysis.     

Inhibitors incorporating zinc-binding groups target the GlcNAc-PI de-N-acetylase in Trypanosoma brucei, the causative agent of African sleeping sickness

Disruption of glycosylphosphatidylinositol biosynthesis is genetically and chemically validated as a drug target against the protozoan parasite Trypanosoma brucei, the causative agent of African sleeping sickness. The N-acetylglucosamine-phosphatidylinositol de-N-acetylase (deNAc) is a zinc metalloenzyme responsible for the second step of glycosylphosphatidylinositol biosynthesis. We recently reported the synthesis of eight deoxy-2-C-branched monosaccharides containing carboxylic acid, hydroxamic acid, or N-hydroxyurea substituents at the C2 position that may act as zinc-binding groups. Here, we describe the synthesis of a glucocyclitol-phospholipid incorporating a hydroxamic acid moiety and report the biochemical evaluation of the monosaccharides and the glucocyclitol-phospholipid as inhibitors of the trypanosome deNAc in the cell-free system and against recombinant enzyme. Monosaccharides with carboxylic acid or hydroxamic acid substituents were found to be the inhibitors of the trypanosome deNAc with IC(50) values 0.1-1.5mM and the glucocyclitol-phospholipid was found to be a dual inhibitor of the deNAc and the α1-4-mannose transferase with an apparent IC(50)= 19±0.5μm.     

Crystal structures of Candida albicans dihydrofolate reductase bound to propargyl-linked antifolates reveal the flexibility of active site loop residues critical for ligand potency and selectivity

Candida albicans and Candida glabrata cause fungal bloodstream infections that are associated with significant mortality. As part of an effort to develop potent and selective antifolates that target dihydrofolate reductase (DHFR) from Candida species, we report three ternary crystal structures of C. albicans DHFR (CaDHFR) bound to novel propargyl‐linked analogs. Consistent with earlier modeling results, these structures show that hydrophobic pockets in the binding site may be exploited to increase ligand potency. The crystal structures also confirm that loop residues Thr 58‐ Phe 66, which flank the active site and influence ligand potency and selectivity, adopt multiple conformations. To aid the development of a dual Candida spp. inhibitor, three new crystal structures of C. glabrata DHFR (CgDHFR) bound to similar ligands as those bound in the ternary structures of CaDHFR are also reported here. Loop residues 58–66 in CgDHFR and human DHFR are 1 and 3 Å closer to the folate binding site, respectively, than loop residues in CaDHFR, suggesting that a properly size ligand could be a potent and selective dual inhibitor of CaDHFR and CgDHFR.     

Proteomic response of the rat liver in differential swimming modes

Moderate exercise helps improve competition results, providing a balanced muscle tone and biochemical activity, whereas excessive training disrupts the balance between training and recovery, causes harm to the organism, and leads to overtraining syndrome (OTS). To explore the mechanisms of different protein expressions during training and acquisition of immunity, we used proteomic analyses to investigate the differences of liver‐protein expressions between 2 swimming modes. Sprague‐Dawley rats were randomly divided into control (CT), fatigue training (FT), and exhaustive training (ET) groups, and liver tissues from each group were subjected to 2‐dimensional electrophoresis (2DE). A total of 4518 protein spots were detected in 9 replicates, and 45 protein spots exhibited a >2‐fold difference in expression (P < .05), 31 of which was successfully identified by mass spectrometry. SERPINA3K expression decreased markedly during 2 stages from CT → FT and FT → ET, while DDT, RHOT1, and RBP4 decreased significantly only from CT → ET but not from the former 2 stages. By contrast, KRT8, PCBD1, KRT18, PRDX1, and ACY1A showed significant >2‐fold increase in expression in either the CT → FT or FT → ET stages. Bioinformatic analyses showed that among the identified proteins, 30.2%, 54.18%, and 15.62% were involved in biological processes, molecular functions, and cell composition, respectively. Notably, PCBD1, PRDX1, and PPP1CB were involved in redox processes, while PPP1CB was only expressed in the FT group. RGN, PSMB9, and AGT, commonly recognized as oxidative stress biomarkers, may involve in regulating homeostasis in the locomotor mode and may provide diagnostic criteria for the occurrence and prevention of exercise‐induced fatigue and OTS.     

Structure‐Based Design of a Novel Class of Potent Inhibitors of InhA,the Enoyl Acyl Carrier Protein Reductase from Mycobacterium Tuberculosis: A Computer Modelling Approach

The NADH‐dependent Enoyl‐ACP reductase (InhA) of Mycobacterium tuberculosis has been shown to be the primary target of the frontline drug isoniazid (INH). However, INH must be first activated by katG gene, mutations in which have mediated resistance to INH. Recently, direct inhibitors of InhA have been reported. Using a structure‐based approach, we have identified a tripeptide inhibitor with the sequence WYW, which is 100 times more potent than the existing inhibitors. It is therefore, a potential lead compound for the development of new anti‐TB drugs.