5-Lipoxygenase as a drug target: A review on trends in inhibitors structural design,SAR and mechanism based approach

The most common inflammatory disease of the airways is asthma among children affecting around 235 million people worldwide. 5-Lipoxygenase (5-LOX) is a crucial enzyme which helps in the conversion of arachidonic acid (AA) to leukotrienes (LTs), the lipid mediators. It is associated with several inflammation related disorders such as asthma, allergy, and atherosclerosis. Therefore, it is considered as a promising target against inflammation and asthma. Currently, the only drug against 5-LOX which is available is Zileuton, while a few inhibitors are in clinical trial stages such as Atreleuton and Setileuton. So, there is a dire requirement in the area of progress of novel 5-LOX inhibitors which necessitates an understanding of their structure activity relationship and mode of action. In this review, novel 5-LOX inhibitors reported so far, their structural design, SAR and developmental strategies along with clinical updates are discussed over the last two decades.     

Effect of Lactobacillus rhamnosus NCDC 298 with FOS in Combination on Viability and Toxin Production of Enterotoxigenic Escherichia coli

Probiotics and Antimicrobial Proteins - The present study was to investigate the utilization of prebiotics by Lactobacillus rhamnosus NCDC 298 and its synergistic adversary effect on both...     

Alpha-actinin-4 is required for normal podocyte adhesion

Mutations in the alpha-actinin-4 gene ACTN4 cause an autosomal dominant human kidney disease. Mice deficient in alpha-actinin-4 develop a recessive phenotype characterized by kidney failure, proteinuria, glomerulosclerosis, and retraction of glomerular podocyte foot processes. However, the mechanism by which alpha-actinin-4 deficiency leads to glomerular disease has not been defined. Here, we examined the effect of alpha-actinin-4 deficiency on the adhesive properties of podocytes in vivo and in a cell culture system. In alpha-actinin-4-deficient mice, we observed a decrease in the number of podocytes per glomerulus compared with wild-type mice as well as the presence of podocyte markers in the urine. Podocyte cell lines generated from alpha-actinin-4-deficient mice were less adherent than wild-type cells to glomerular basement membrane (GBM) components collagen IV and laminin 10 and 11. We also observed markedly reduced adhesion of alpha-actinin-4-deficient podocytes under increasing shear stresses. This adhesion deficit was restored by transfecting cells with alpha-actinin-4-GFP. We tested the strength of the integrin receptor-mediated linkages to the cytoskeleton by applying force to microbeads bound to integrin using magnetic pulling cytometry. Beads bound to alpha-actinin-4-deficient podocytes showed greater displacement in response to an applied force than those bound to wild-type cells. Consistent with integrin-dependent alpha-actinin-4-mediated adhesion, phosphorylation of beta1-integrins on alpha-actinin-4-deficient podocytes is reduced. We rescued the phosphorylation deficit by transfecting alpha-actinin-4 into alpha-actinin-4-deficient podocytes. These results suggest that alpha-actinin-4 interacts with integrins and strengthens the podocyte-GBM interaction thereby stabilizing glomerular architecture and preventing disease.     

Proteomic approach for identification and characterization of novel immunostimulatory proteins from soluble antigens of Leishmania donovani promastigotes

Visceral leishmaniasis (VL) caused by Leishmania donovani is a major parasitic disease prevalent in endemic regions of Bihar in India. In the absence of good chemotherapeutic options, there is a need to develop an effective vaccine against VL which should be dependent on the generation of a T helper type 1 (Th1) immune response. We have shown that soluble proteins from promastigote of a new clinical isolate of L. donovani (2001) ranging from 68 to 97.4 kDa (F2 fraction), induce Th1 responses in the peripheral blood mononuclear cells of cured Leishmania patients and hamsters and also showed significant prophylactic potential. To understand the nature of F2 proteins, it was further characterized using 2-DE, MALDI-TOF and MALDI-TOF/TOF-MS. In all, 63 spots were cut from a CBB stained gel for analysis and data was retrieved for 52 spots. A total of 33 proteins were identified including six hypothetical/unknown proteins. Major immunostimulatory proteins were identified as elongation factor-2, p45, heat shock protein (HSP)70, HSP83, aldolase, enolase, triosephosphate isomerase, protein disulfideisomerase and calreticulin. This study substantiates the usefulness of proteomics in characterizing a complex protein fraction (F2) map of soluble L. donovani promastigote antigen identified as Th1 stimulatory for its potential as vaccine targets against VL.     

Light-controlled gene silencing in zebrafish embryos

Functional genomic studies in zebrafish frequently use synthetic oligonucleotides called morpholinos that block RNA splicing or translation. However, the constitutive activity of these reagents limits their experimental utility. We report here the synthesis of a photoactivatable morpholino targeting the no tail (ntl) gene. This caged reagent permits spatiotemporal gene regulation in vivo and the photochemical generation of functionally mosaic organisms.     

Solid state and solution conformations of a helical peptide with a central gly-gly segment

The influence of amino acids with contrasting conformational tendencies on the stereochemistry of oligopeptides has been investigated using an octapeptide Boc-Leu-Aib-Val-Gly-Gly-Leu-Aib-Val-OMe, which contains two helix-promoting Aib residues and a central helix-destabilizing Gly-Gly segment. Single crystal x-ray diffraction studies reveal that a 3 ₁₀-helix is formed up to the penultimate Aib residue, at which point there is a helix reversal in the backbone, reminiscent of a C-terminal 6 → I hydrogen bond. The curious feature in the crystal is the solvation of the possible 6 → 1 bond by a CH₃OH molecule, where the OH is inserted between O(3) and N(8) and participates in hydrogen bonds with both. The cell parameters are as follows: space group P2₁2₁2₁, a = 10.649(4) Å, b = 15.694(5) Å, c = 30.181(8) Å, R = 6.7% for 3427 data (| F₀| > 3σF) observed to 0.9 Å. Nuclear magnetic resonance studies in CDCl₃ using NH group solvent accessibility and nuclear Overhauser effects as probes are consistent with a 3 ₁₀-helical conformation. In contrast, in (CD₃)₂SO, unfolding of the central segment results in a multiple β-turn structure, with β-turn conformations populated at residues 1–2, 3–4, and 6–7. CD studies in methanol-2,2,2-trifluoroethanol (TFE) mixtures also provide evidence for a solvent-dependent structural transition. Helical conformations are populated in TFE, while type II β-turn structures are favored in methanol. © 1996 John Wiley & Sons, Inc.     

Omega amino acids in peptide design: incorporation into helices

Incorporation of easily available achiral ω-amino acid residues into an oligopeptide results in substitution of amide bonds by polymethylene units of an aliphatic chain, thereby providing a convenient strategy for constructing a peptidomimetic. The central Gly-Gly segment of the helical octapeptide Boc-Leu-Aib-Val-Gly-Gly-Leu-Aib-Val-Ome(1) has been replaced by δ-amino-valeric acid (δ-Ava) residue in the newly designed peptide Boc-Leu-Aib-Val-δ-Ava-Leu-Aib-Val-OMe(2). ¹H-nmr results clearly suggest that in the apolar solvent CDCl₃, the δ-Ava residue is accommodated into a folded helical conformation, stabilized by successive hydrogen bonds involving the NH groups of Val(3), δ-Ava(4), and Leu(5). The δ-Ava residue must adopt a gauche-gauche-trans-gauche-gauche conformation along the central polymethylene unit of the aliphatic segment, a feature seen in an energy-minimized model conformation based on nmr parameters. The absence of hydrogen bonding functionalities, however, limits the elongation of the helix. In fact, in CDCl₃, the folded conformation consists of an N-terminal helix spanning residues 1–4, followed by a Type II β-turn at residues 5 and 6, whereas in strongly solvating media like (CD₃)₂SO, the unfolding of the N-terminal helix results in β-turn conformations at Leu(1)-Aib(2). The Type II β-turn at the Leu(5)-Aib(6) segment remains intact even in (CD₃)₂SO. CD comparisons of peptides 1 and 2 reveal a “nonhelical” spectrum for 2 in 2,2,2-trifluoroethanol. © 1996 John Wiley & Sons, Inc.     

Incidence of lab-confirmed dengue fever in a pediatric cohort in Delhi,India

BackgroundOur aim was to estimate the overall and age-specific incidence of lab-confirmed dengue fever using ELISA based assays among children 6 months to 15 years in Delhi.MethodsWe enrolled a cohort of 984 children aged 6 months to <14 years in South Delhi and followed-up weekly for fever for 24 months or till 15 completed years of child-age. Households of the enrolled children were geo-tagged. NS1, IgM and IgG assays were conducted using ELISA method to confirm dengue fever in children with ≥3 consecutive days of fever. Molecular typing was done in a subset of NS1 positive cases to identify the circulating serotypes.Principal findingsWe had a total of 1953 person-years (PY) of follow up. Overall, there were 4208 episodes of fever with peaks during June to November. The overall incidence (95%CI) of fever was 215/100 PY (209 to 222). A total of 74/1250 3-day fever episodes were positive for acute dengue fever (NS1 and/or IgM positive). The overall incidence (95%CI) of acute dengue fever was 37.9 (29.8 to 47.6) per 1000 PY; highest among children aged 5 to 10 years (50.4 per 1000 PY, 95% CI 36.5 to 67.8). Spatial autocorrelation analysis suggested a clustering pattern for the dengue fever cases (Moran’s Index 0.35, z-score 1.8, p = 0.06). Dengue PCR was positive in 16 of the 24 specimens tested; DEN 3 was the predominant serotype identified in 15/24 specimens.ConclusionsWe found a high incidence of dengue fever among under 15-year children with clustering of cases in the community. DEN 3 was the most commonly circulating strain encountered. The findings underscore the need for development of affordable pre-vaccination screening strategy as well as newer dengue vaccines for young children while continuing efforts in vector control.