Structure based design and synthesis of peptide inhibitor of human LOX-12: in vitro and in vivo analysis of a novel therapeutic agent for breast cancer

Human breast cancer cell proliferation involves a complex interaction between growth factors, steroid hormones and peptide hormones. The interaction of growth factors, such as epidermal growth factor (EGF), with their receptors on breast cancer cells can lead to the hydrolysis of phospholipids and release of fatty acid such as arachidonic acid, which can be further metabolized by cyclooxygenase (COX) and lipoxygenase (LOX) pathways to produce prostaglandins. The high concentration of prostaglandins has been associated with chronic inflammatory diseases and several types of human cancers. This is due to the over expression COX, LOX and other inflammatory enzymes. Ten peptides were designed and synthesized by solid phase peptide synthesis and analyzed in vitro for enzyme inhibition. Out of these peptides, YWCS had shown significant inhibitory effects. The dissociation constant (K(D)) was determined by surface plasmon resonance (SPR) analysis and was found to be 3.39 × 10(-8) M and 8.6 × 10(-8) M for YWCS and baicalein (positive control), respectively. The kinetic constant Ki was 72.45 × 10(-7) M as determined by kinetic assay. The peptide significantly reduced the cell viability of estrogen positive MCF-7 and estrogen negative MDA-MB-231 cell line with the half maximal concentration (IC(50)) of 75 μM and 400 μM, respectively. The peptide also induced 49.8% and 20.8% apoptosis in breast cancer cells MCF-7 and MDA-MB-231, respectively. The YWCS was also found to be least hemolytic at a concentration of 358 μM. In vivo studies had shown that the peptide significantly inhibits tumor growth in mice (p<0.017). This peptide can be used as a lead compound and complement for ongoing efforts to develop differentiation therapies for breast cancer.     

The diversity of life-form type, habitat preference and phenology of the endemics in the Goa region of the Western Ghats, India

Aim To carry out (1) a floristic survey of endemic flowering plants of the Western Ghats occurring in Goa, (2) identify their habitat preference and diversity of life‐form type, (3) observe flowering phenology of the endemics and (4) to correlate factors that affect their phenological pattern. Location Goa state is located between 15°48′ N and 14°53′54′′ N and 74°20′13′′ E and 73°40′33′′ E, in the northern part of the Western Ghats, India. Method A list of endemic plants from the study area was prepared using available floristic works and checklists of endemic plants of India. Based on preliminary field observations carried out in the study area, major habitats such as plateaus, moist deciduous forests, semi‐evergreen forests, evergreen forests and mangroves were identified for the subsequent intensive survey of endemics. Voucher specimens for all the endemic species were collected, processed using conventional herbarium techniques and deposited in the Herbarium, Department of Botany, Goa University. Species were identified using local and regional floras and their identity was confirmed at various herbaria along the Western Ghats. Data on their life‐form types, habitat and phenology was recorded in the field. Phenological observations were made every fortnight. A computerized data base was generated incorporating details on their life‐form type, phenology and habitat. Results A floristic survey of endemic plants of the Western Ghats in Goa resulted in the collection of 113 endemic species. Life‐form analysis reveals that herbaceous endemics are the most dominant followed by trees, shrubs and climbers. Plateaus in the study area harbour the largest number of endemic species, especially herbs. Endemic trees are distributed in the semi‐evergreen and evergreen forests. Endemic species in the study area show different peak and lean seasons of flowering depending on their life‐form type, habitat and ecological factors like temperature and rainfall/moisture content in the soil. Main conclusions The plateaus in the northern part of the Western Ghats are unique, being species rich with herbaceous endemics. These ephemerals are closely associated with the rainfall patterns thus; any change of moisture regime over the long‐term will have an impact on the distribution of these endemics.     

Expedient synthesis of the pentasaccharide repeating unit of the <Emphasis Type="Italic">O</Emphasis>-antigen of <Emphasis Type="Italic">Escherichia coli</Emphasis> O86 and its conformational analysis

Synthesis of the pentasaccharide with a 2-aminoethyl linker attached to the reducing end corresponding to the cell wall O-antigen of Escherichia coli O86 strain is reported. The synthetic strategy involves sequential glycosylation of suitably protected monosaccharide intermediates under similar glycosylation reaction conditions. Thioglycosides have been used as glycosyl donor throughout the synthetic strategy. Conformational analysis of the synthesized pentasaccharide has been carried out using 2D ROESY NMR spectral analysis and all atom explicit molecular dynamics (MD) simulation technique.

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Graphical abstract Facile synthesis of the pentasaccharide with a 2-aminoethyl linker attached to the reducing end corresponding to the cell wall O-antigen of Escherichia coli O86 strain is reported. Conformational analysis of the synthesized pentasaccharide has been carried out using 2D ROESY NMR spectral analysis and all atom explicit molecular dynamics (MD) simulation technique.

     

Inheritance of gynandromorphism in the parasitic wasp Nasonia vitripennis

The parasitic wasp Nasonia vitripennis has haplo-diploid sex determination. Males develop from unfertilized eggs and are haploid, whereas females develop from fertilized eggs and are diploid. Females and males can be easily distinguished by their morphology. A strain that produces individuals with both male and female features (gynandromorphs) is studied. We provide data on female/male patterning within and between individuals, on environmental effects influencing the occurrence of gynandromorphism, and on its pattern of inheritance. A clear anterior/posterior pattern of feminization is evident in gynandromorphic individuals that developed from unfertilized haploid eggs. The proportion of gynandromorphic individuals can be increased by exposing the mothers to high temperature and also by exposing embryos at early stages of development. Selection for increased gynandromorph frequency was successful. Backcross and introgression experiments showed that a combination of a nuclear and a heritable cytoplasmic component causes gynandromorphism. Analyses of reciprocal F(2) and F(3) progeny indicate a maternal effect locus (gyn1) that maps to chromosome IV. Coupled with previous studies, our results are consistent with a N. vitripennis sex determination involving a maternal/zygotic balance system and/or maternal imprinting. Genetics and temperature effects suggest a temperature-sensitive mutation of a maternally produced masculinizing product that acts during a critical period in early embryogenesis.     

Hypothalamic Control of Conspecific Self-Defense

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The effect of dietary carbohydrate on the rise in plasma glutamate concentrations following oral glutamate ingestion in rats

Plasma glutamate concentrations were examined in male rats following oral intubation of monosodium L-glutamic acid (MSG, 250 mg/kg) soon after ingesting one of several meals differing in carbohydrate content. Intubation of MSG alone produced a 4-fold rise in plasma glutamate that peaked at 15 min, and returned to baseline by 60 min. Red blood cell glutamate concentrations were unchanged. The ingestion of a meal lacking carbohydrate produced a modest attenuation of the post-MSG intubation rise in plasma glutamate concentrations. This attenuating effect increased progressively with the carbohydrate content of the meal (and as the protein content declined, to maintain isocaloric meals), though as little as 5% carbohydrate marked attenuated the plasma glutamate rise. This effect diminished as the time interval between the meal and MSG intubation increased from 1 to 4 hrs. Similar, but not identical effects were noted when meals substituted fat (instead of protein) for carbohydrate. The intubation of MSG alone produced a slight increase in plasma alanine concentrations over the 60-min post-intubation period examined. The ingestion of any of the meals just prior to intubation did not influence this effect. Overall, the results indicate that although the ingestion of carbohydrate can markedly attenuate the rise in plasma glutamate that follows MSG consumption in rats, this effect is also influenced by the other macronutrients present. The absence of notable, meal related changes in plasma alanine suggests that this parameter does not provide a useful indication of gut glutamate transamination.     

On a model and the kinetics of photo-enhanced enzyme reactions

The recently observed enhancement, by laser irradiation, of the specific activity of the enzyme chymotrypsin (which hydrolyses Benzoyl-L-tyrosine-ethyl ester) at low enzyme concentration is considered. The enhancement of the reaction rate is attributed to a coherently excited state of the enzyme molecule (activated through Raman scattering of the laser light) following a prediction due to Fröhlich. The model is described, the kinetics of the process is framed and the observed enzyme-concentration dependence of the specific activity is reproduced. Predictions of the model are delineated to urge verification of the main contentions through further experimentation.     

A novel palindromic triple-stranded structure formed by homopyrimidine dodecamer d-CTTCTCCTCTTC and homopurine hexamer d-GAAGAG.

We have carried out NMR and molecular mechanics studies on a complex formed when a palindromic homopyrimidine dodecamer (d-CTTCTCCTCTTC) and a homopurine hexamer (d-GAAGAG) are mixed in 1:1 molar ratio in aqueous solutions. Such studies unequivocally establish that two strands of each oligomer combine to form a triple-stranded DNA structure with a palindromic symmetry and with six T.A:T and six C+. G:C hydrogen-bonded base triads. The two purine strands are placed head to head, with their 3' ends facing each other in the center of the structure. One-half of each pyrimidine strand contains protonated and the other half contains non-protonated cytosines. The two half segments containing protonated cytosines are hydrogen bonded to each of the two purine hexamers through Hoogsteen T.A and C+.G base pairing. The segments containing non-protonated cytosines are involved in Watson-Crick (A:T and G:C) base pairing. This leads to a palindromic triplex with a C2-dyad symmetry with respect to the center of the structure. The complex is less stable at neutral pH, but the cytosines involved in Hoogsteen base pairing remain protonated even under these conditions. Molecular mechanics calculations using NMR constraints have provided a detailed three-dimensional structure of the complex. The entire stretches of purine, and the pyrimidine nucleotides have a conformation close to B-DNA.