A re-examination of the difluoromethylenesulfonic acid group as a phosphotyrosine mimic for PTP1B inhibition

Protein tyrosine phosphatase 1B (PTP1B) is involved in the down-regulation of insulin signaling and is a well-validated therapeutic target for the treatment of diabetes and obesity. Key to the design of potent inhibitors of PTP1B is a moiety that effectively mimics the phosphate group of the natural phosphotyrosine substrate. Difluoromethylsulfonomethylphenylalanine (F(2)Smp) is one of the best monoanionic pTyr mimics reported to date. However, the difluoromethylenesulfonic acid (DFMS) group as a phosphate mimic has not been carefully evaluated in the context of a non-peptidyl platform. Here we present a careful examination of the DFMS group as a phosphate mimic. This was achieved by first constructing an analog of a previously reported high affinity, non-peptidyl PTP1B inhibitor (compound 2, IC(50)=8nM) in which a difluoromethylenephosphonic acid group is replaced with the DFMS moiety (compound 6). We also report the synthesis of its non-fluorinated methylenesulfonic analog (compound 7), as well as two other derivatives in which a distal sulfonamide moiety is replaced with a difluoromethylenesulfonamide group (compounds 8 and 9). Compounds 2 and 6-9 were examined as PTP1B inhibitors. Replacing the distal sulfonamide moiety with a difluoromethylenesulfonamide group had only a modest effect on inhibitor potency. However, compound 6 was approximately a 1000-fold poorer inhibitor than compound 2. Most significantly, inhibition studies with compound 7 and a peptide bearing sulfonomethylphenylalanine revealed that the fluorines have little effect on the potency of the DFMS-bearing inhibitors. This is in contrast to a previous assumption that the fluorines in DFMS-bearing inhibitors contributed significantly to their potency. This may in part explain the large difference in potency between the DFMS and DFMP-bearing compounds. These results also demonstrate that sulfonomethylphenylalanine, a pTyr mimic that is readily constructed, is a relatively good pTyr mimic in comparison to most others that have been reported when examined in the context of the DADE-X-LNH(2) peptide platform.     

Modulation of glucose transporter (GLUT4) by vanadate and Trigonella in alloxan-diabetic rats

Oral administration of vanadate is an effective treatment for diabetes in animal models. However, vanadate exerts these effects at high doses and several toxic effects are produced. Low doses of vanadate are relatively safe but are unable to elicit any antidiabetic effect. The present study explored the prospect of using low doses of vanadate in combination with Trigonella seed powder (TSP) to evaluate their antidiabetic effect in alloxan-diabetic rats. Alloxan-diabetic rats were treated with insulin, vanadate, TSP and vanadate and TSP in combination for 3 weeks. The effect of these antidiabetic compounds was examined on general physiological parameters and distribution of glucose transporter (GLUT4) in skeletal muscle by immunoblotting and immunohistochemistry. Treatment of alloxan-diabetic rats with insulin, vanadate, TSP and vanadate in combination with TSP revived normoglycemia and restored the disturbances in the distribution of GLUT4 in skeletal muscle. TSP treatment was only partially effective in the restoration of diabetic alterations. The treatment of diabetic rats with combined doses of vanadate and TSP was most effective in the normalization of plasma glucose levels and correction of altered GLUT4 distribution.     

Rapid recovery of a population of the cryptic and evolutionarily distinct Hochstetter's Frog,Leiopelma hochstetteri,in a pest‐free environment

New Zealand's native frogs (genus Leiopelma) display a very high degree of endemism and belong to a distinctive and ancient evolutionary lineage. All four extant species are considered to be threatened or endangered, but the mechanisms behind their decline are poorly understood. Many of the potential factors causing population declines are confounded for the two species living on the mainland; in particular, habitat degradation and the presence of introduced mammalian pests have been difficult to disentangle in the past. Here we present data on a population of Hochstetter's Frog (Leiopelma hochstetteri) living in a pest‐free mainland sanctuary, the 3363‐ha Maungatautari Ecological Island. Survey data from this population shows a fourfold increase in numbers between 2009 and 2012 from a very low initial density. Our results suggest that for Hochstetter's Frog at least, and possibly the other mainland species of Leiopelma, introduced mammals are a major driving factor in population declines. Given that the frog population at Maungatautari was discovered by serendipity when planning for the pest‐free sanctuary was already well underway, this also highlights the conservation value of large pest‐free areas and ecosystem‐level management.     

Diffuse large cell lymphoma presenting as a sacral mass and lupus anticoagulant

A 67-year-old gentleman presented to Yale-New Haven Hospital (YNHH) for assessment of a supratherapeutic INR and sacral lesion. Hematologic workup revealed elevated ESR, PT, INR, PTT, and CRP, mixing studies that failed to correct, and a positive Russell Viper Venom Test (RVVT), which confirmed the presence of lupus anticoagulant (LA), a subtype of antiphospholipid syndrome (APA). Pathology of the patient's sacral lesion revealed diffuse large B-cell lymphoma. This case provides insight into the association between APA and lymphoid neoplasm. The patient's unique presentation is in marked contrast to other reports of APA and lymphoid malignancy, which are typically associated with elevated PTT, normal PT, minimal extranodal disease, and potential thrombotic complications. Further, treatment with Rituximab-CHOP chemotherapy led to excellent clinical response with tumor remission and normalization of PT and PTT.     

Molecular dynamics simulation of mammalian 15S-lipoxygenase with AMBER force field

A molecular dynamics simulation study of mononuclear iron 15S-lipoxygenase (15S-LOX) from rabbit reticulocytes was performed to investigate its structure and dynamics; newly developed AMBER force field parameters were employed for the first coordination sphere of the catalytic iron (II). The results obtained from this study demonstrate that the structural features of the catalytic iron coordination site are in good agreement with available data obtained from experiments. The motional flexibility of the N-terminal β-barrel domain is greater than the C-terminal catalytic domain; flexibility was assessed in terms of B-factors and secondary structure calculations. The significant features obtained for the relative motional flexibility of these two domains of 15S-LOX in solution as well as the isolated C-terminal domain were analyzed in terms of radius of gyration and maximum diameter, which correlated well with the structural flexibility of 15-lipoxygenase-1 in solution as probed by small-angle X-ray scattering. The motional flexibility indicates interdomain motion between the N-terminal β-barrel and the C-terminal catalytic domain; this was further verified by the evaluation of central bending in the solvated LOX molecule, which identified an unstructured stretch of amino acids as the interdomain linker. The average bending angle confirmed significant central bending between these two domains, which was linked to the high degree of motional freedom of the N-terminal β-barrel domain in aqueous solutions. This can be considered to have biological relevance for membrane binding as well as for regulating the catalytic domain.     

Protective Effect of Selenomethionine on Aflatoxin B1-Induced Oxidative Stress in MDCK Cells

AFB1 is a mycotoxin which exerts their cytotoxicity through increasing oxidative damage in target organ. Kidney is one of target organs vulnerable to damage caused by AFB1. In this study, Madin-Darby canine kidney (MDCK) cells were used to evaluate the AFB1-induced cell damage by the MTT assay. The results revealed that the toxic effect of AFB1 on MDCK cells is both dose and time dependent. Half maximal toxic concentration (IC₅₀) was noted at 0.25 μg/ml of AFB1. Further, protective effect of six different concentrations (0.2, 0.8, 1, 2, 4, and 8 μM) of selenomethionine (SeMet) was observed against 0.25 μg/ml of AFB1-induced damage. The results showed that 0.25 μg/ml of AFB1 caused significant increase in oxidative stress, which was demonstrated by significant increase of malondialdehyde (MDA) level, reduction of intracellular GSH level, as well as GPX1 activity and mRNA level in MDCK cells when compared with control. SeMet protected the cells from AFB1-induced oxidative damage in a dose-dependant manner. Good protection could be achieved between 1 and 4 μM of concentration. Amid this range, MDA level significantly decreased while intracellular GSH level and GPX1 activity in addition to mRNA level significantly increased. Moreover, cell viability was significantly improved. It could be concluded that SeMet is a potential antioxidative agent to alleviate AFB1-induced oxidative stress.     

Design of New and Potent Diethyl Thiobarbiturates as Urease Inhibitors: A Computational Approach

Urease is an important enzyme both in agriculture and medicine research. Strategies based on urease inhibition is critically considered as the first line treatment of infections caused by urease producing bacteria. Since, urease possess agro-chemical and medicinal importance, thus, it is necessary to search for the novel compounds capable of inhibiting this enzyme. Several computational methods were employed to design novel and potent urease inhibitors in this work. First docking simulations of known compounds consists of a set of arylidine barbiturates (termed as reference) were performed on the Bacillus pasteurii (BP) urease. Subsequently, two fold strategies were used to design new compounds against urease. Stage 1 comprised of the energy minimization of enzyme-ligand complexes of reference compounds and the accurate prediction of the molecular mechanics generalized born (MMGB) interaction energies. In the second stage, new urease inhibitors were then designed by the substitution of different groups consecutively in the aryl ring of the thiobarbiturates and N, N-diethyl thiobarbiturates of the reference ligands.. The enzyme-ligand complexes with lowest interaction energies or energies close to the calculated interaction energies of the reference molecules, were selected for the consequent chemical manipulation. This was followed by the substitution of different groups on the 2 and 5 positions of the aryl ring. As a result, several new and potent diethyl thiobarbiturates were predicted as urease inhibitors. This approach reflects a logical progression for early stage drug discovery that can be exploited to successfully identify potential drug candidates.     

Extracellular biosynthesis of silver nanoparticles: effects of shape-directing cetyltrimethylammonium bromide,pH, sunlight and additives

The work reported in this paper describes the preparation, morphology, stability and sensitivity of Ag-nanoparticles towards sunlight using Allium sativum, garlic extract for the first time. The synthesized silver particles show an intense surface plasmon resonance band in the visible region at 410 nm. The position of the wavelength maxima, blue and red shift, strongly depends on the sunlight and pH. TEM analysis revealed the presence of spherical, different size (from 5.0 to 30 nm) and garlic constituents bio-conjugated, stabilized and/or layered silver nanoparticles. The concentrations of garlic extract, cetyltrimethylammonium bromide, Ag⁺ ions and reaction time play vital roles for nucleus formation and the growth processes. Sulfur-containing biomolecules of extract, especially cysteine, are responsible for the reduction of Ag⁺ ions into metallic Ag⁰. The agglomeration number of the silver nanoparticles (N _Ag) and the average number of free electrons per particle (n _fe) are calculated and discussed.     

GMF-Knockout Mice are Unable to Induce Brain-Derived Neurotrophic Factor after Exercise

We earlier reported that overexpression of glia maturation factor (GMF) in cultured astrocytes enhances the production of brain-derived neurotrophic factor (BDNF). The current study was conducted to find out whether BDNF production is impaired in animals devoid of GMF. To this end GMF-knockout (KO) mice were subjected to exercise and the neurotrophin mRNAs were determined by real-time RT-PCR. Compared to wild-type (WT) mice, there is a decrease in exercise-induced BDNF in the KO mice. The observation was correlated with the finding that, in WT mice, exercise increases GMF expression. The results are consistent with the hypothesis that GMF is necessary for exercise-induction of BDNF, and that GMF may promote neuroprotection through BDNF production.     

Antiproliferative function of glia maturation factor beta.

Recombinant human glia maturation factor beta (GMF-beta) reversibly inhibits the proliferation of neoplastic cells in culture by arresting the cells in the G0/G1 phase. This phenomenon is not target-cell specific, as neural and nonneural cells are equally inhibited. When tested simultaneously, GMF-beta suppresses the mitogenic effect of acidic fibroblasts growth factor (aFGF), but the two are synergistic in promoting the morphologic differentiation of cultured astrocytes. GMF-beta also counteracts the growth-stimulating effect of pituitary extract and cholera toxin on Schwann cells. The results underscore the regulatory role of GMF-beta and its intricate interaction with the mitogenic growth factors.