Simulations reveal that the HIV-1 gp120-CD4 complex dissociates via complex pathways and is a potential target of the polyamidoamine (PAMAM) dendrimer

The polyamidoamine (PAMAM) dendrimer prevents HIV-1 entry into target cells in vitro. Its mechanism of action, however, remains unclear and precludes the design of potent dendrimers targeting HIV-1 entry. We employed steered molecular dynamics simulations to examine whether the HIV-1 gp120-CD4 complex is a target of PAMAM. Our simulations mimicked single molecule force spectroscopy studies of the unbinding of the gp120-CD4 complex under the influence of a controlled external force. We found that the complex dissociates via complex pathways and defies the standard classification of adhesion molecules as catch and slip bonds. When the force loading rate was large, the complex behaved as a slip bond, weakening gradually. When the loading rate was small, the complex initially strengthened, akin to a catch bond, but eventually dissociated over shorter separations than with large loading rates. PAMAM docked to gp120 and destabilized the gp120-CD4 complex. The rupture force of the complex was lowered by PAMAM. PAMAM disrupted salt bridges and hydrogen bonds across the gp120-CD4 interface and altered the hydration pattern of the hydrophobic cavity in the interface. In addition, intriguingly, PAMAM suppressed the distinction in the dissociation pathways of the complex between the small and large loading rate regimes. Taken together, our simulations reveal that PAMAM targets the gp120-CD4 complex at two levels: it weakens the complex and also alters its dissociation pathway, potentially inhibiting HIV-1 entry.

Propyl-2-(8-(3,4-Difluorobenzyl)-2 `,5 `-Dioxo-8-AzaspiroBicyclo3.2.1] Octane-3,4 `-Imidazolidine]-1 `-yl) Acetate Induces Apoptosis in Human Leukemia Cells through Mitochondrial Pathway following Cell Cycle Arrest

Background: Due to the functional defects in apoptosis signaling molecules or deficient activation of apoptosis pathways, leukemia has become an aggressive disease with poor prognosis. Although the majority of leukemia patients initially respond to chemotherapy, relapse is still the leading cause of death. Hence targeting apoptosis pathway would be a promising strategy for the improved treatment of leukemia. Hydantoin derivatives possess a wide range of important biological and pharmacological properties including anticancer properties. Here we investigated the antileukemic activity and mechanism of action of one of the potent azaspiro hydantoin derivative, (ASHD). Materials and Methods: To investigate the antileukemic efficacy of ASHD, we have used MTT assay, cell cycle analysis by FACS, tritiated thymidine incorporation assay, Annexin V staining, JC1 staining and western blot analysis. Results: Results showed that ASHD was approximately 3-fold more potent than the parent compounds in inducing cytotoxicity. Tritiated thymidine assay in conjunction with cell cycle analysis suggests that ASHD inhibited the growth of leukemic cells. The limited effect of ASHD on cell viability of normal cells indicated that it may be specifically directed to cancer cells. Translocation of phosphatidyl serine, activation of caspase 3, caspase 9, PARP, alteration in the ratio of BCL2/BAD protein expression as well as the loss of mitochondrial membrane potential suggests activation of the intrinsic pathway of apoptosis. Conclusion: These results could facilitate the future development of novel hydantoin derivatives as chemotherapeutic agents for leukemia.

Aza-induced cardiomyocyte differentiation of P19 EC-cells by epigenetic co-regulation and ERK signaling

Stem cells in cell based therapy for cardiac injury is being potentially considered. However, genetic regulatory networks involved in cardiac differentiation are not clearly understood. Among stem cell differentiation models, mouse P19 embryonic carcinoma (EC) cells, are employed for studying (epi)genetic regulation of cardiomyocyte differentiation. Here, we comprehensively assessed cardiogenic differentiation potential of 5-azacytidine (Aza) on P19 EC-cells, associated gene expression profiles and the changes in DNA methylation, histone acetylation and activated-ERK signaling status during differentiation. Initial exposure of Aza to cultured EC-cells leads to an efficient (55%) differentiation to cardiomyocyte-rich embryoid bodies with a threefold (16.8%) increase in the cTnI(+) cardiomyocytes. Expression levels of cardiac-specific gene markers i.e., Isl-1, BMP-2, GATA-4, and alpha-MHC were up-regulated following Aza induction, accompanied by differential changes in their methylation status particularly that of BMP-2 and alpha-MHC. Additionally, increases in the levels of acetylated-H3 and pERK were observed during Aza-induced cardiac differentiation. These studies demonstrate that Aza is a potent cardiac inducer when treated during the initial phase of differentiation of mouse P19 EC-cells and its effect is brought about epigenetically and co-ordinatedly by hypo-methylation and histone acetylation-mediated hyper-expression of cardiogenesis-associated genes and involving activation of ERK signaling.

Synthetic glutathione peroxidase mimics: effect of nucleophilicity of the aryl thiol cofactor on the antioxidant activity

Catalytic activity of a series of potent amide- and amine-based organoselenium compounds are studied in the presence of various aromatic thiols having electron donating and electron withdrawing substituents on the phenyl ring. This study suggests that the antioxidant activities of the synthetic GPx mimics can be significantly increased by the incorporation of a suitable electron donating group on the phenyl ring of an aromatic thiol.

MPK-09, a small molecule inspired from bioactive styryllactone restores the wild-type function of mutant p53

In the search for more efficacious and less toxic cancer drugs, the tumor suppressor p53 protein has long been a desirable therapeutic target. In the recent past, few independent studies have demonstrated that the antitumor activity of wild-type p53 can be restored in cancer cells harboring mutant form of p53 using small molecule activators. In this study, we describe a novel small molecule MPK-09, which is selective and highly potent against allele specific p53 mutations mainly, R175H, R249S, R273H, R273C, and E285K. Except E285K, all other mutations tested are among the six ``hot spot'' p53 mutations reported in majority of human cancer. Furthermore, our study conclusively demonstrates that the apoptotic activity of the small molecule MPK-09 against cancer cells harboring R273C and E285K mutations is due to restoration of the wild-type conformation to the corresponding mutant form of p53.

Gene Transfection in High Serum Levels: Case Studies with New Cholesterol Based Cationic Gemini Lipids

Background: Six new cationic gemini lipids based on cholesterol possessing different positional combinations of hydroxyethyl (-CH2CH2OH) and oligo-oxyethylene -(CH2CH2O)(n)- moieties were synthesized. For comparison the corresponding monomeric lipid was also prepared. Each new cationic lipid was found to form stable, clear suspensions in aqueous media. Methodology/Principal Findings: To understand the nature of the individual lipid aggregates, we have studied the aggregation properties using transmission electron microscopy (TEM), dynamic light scattering (DLS), zeta potential measurements and X-ray diffraction (XRD). We studied the lipid/DNA complex (lipoplex) formation and the release of the DNA from such lipoplexes using ethidium bromide. These gemini lipids in presence of a helper lipid, 1, 2-dioleoyl phophatidyl ethanol amine (DOPE) showed significant enhancements in the gene transfection compared to several commercially available transfection agents. Cholesterol based gemini having -CH2-CH2-OH groups at the head and one oxyethylene spacer was found to be the most effective lipid, which showed transfection activity even in presence of high serum levels (50%) greater than Effectene, one of the potent commercially available transfecting agents. Most of these geminis protected plasmid DNA remarkably against DNase I in serum, although the degree of stability was found to vary with their structural features. Conclusions/Significance: -OH groups present on the cationic headgroups in combination with oxyethylene linkers on cholesterol based geminis, gave an optimized combination of new genera of gemini lipids possessing high transfection efficiency even in presence of very high percentage of serum. This property makes them preferential transfection reagents for possible in vivo studies.

De novo synthesis of 1-deoxythiosugars

A short and efficient chemical synthesis of biologically potent and novel 1-deoxythiosugars is accomplished. Introduction of sulfur mediated by benzyltriethylammonium tetrathiomolybdate, as a sulfur transfer reagent through nucleophilic double displacement of tosylate in alpha,omega-di-O-tosyl aldonolactones in an intramolecular fashion is the key feature. The subsequent reduction of thiosugar lactones with borohydride exchange resin (BER) offers a number of deoxythiosugars in good overall yield. (C) 2013 Elsevier Ltd. All rights reserved.

Highly Stable Hexacoordinated Nonoxidovanadium(IV) Complexes of Sterically Constrained Ligands: Syntheses, Structure, and Study of Antiproliferative and Insulin Mimetic Activity

Three highly stable, hexacoordinated nonoxidovanadium(IV), V-IV(L)(2), complexes (1-3) have been isolated and structurally characterized with tridentate aroylhydrazonates containing ONO donor atoms. All the complexes are stable in the open air in the solid state as well as in solution, a phenomenon rarely observed in nonoxidovanadium(IV) complexes. The complexes have good solubility in organic solvents, permitting electrochemical and various spectroscopic investigations. The existence of nonoxidovanadium(IV) complexes was confirmed by elemental analysis, ESI mass spectroscopy, cyclic voltammetry, EPR, and magnetic susceptibility measurements. X-ray crystallography showed the N3O3 donor set to define a trigonal prismatic geometry in each case. All the complexes show in vitro insulin mimetic activity against insulin responsive L6 myoblast cells, with complex 3 being the most potent, which is comparable to insulin at the complex concentration of 4 mu M, while the others have moderate insulin mimetic activity. In addition, the in vitro antiproliferative activity of complexes 1-3 against the He La cell line was assayed. The cytotoxicity of the complexes is affected by the various functional groups attached to the bezoylhydrazone derivative and 2 showed considerable antiproliferative activity compared to the most commonly used chemotherapeutic drugs.

Identification and characterization of novel indole based small molecules as anticancer agents through SIRT1 inhibition

In our pursuit to develop new potential anticancer leads, we designed a combination of structural units of indole and substituted triazole; and a library of 1-{1-methyl-2-4-phenyl-5-(propan-2-ylsulfanyl)-4H-1,2,4-triazol-3-yl ]-1H-indol-3-yl}methanamine derivatives was synthesized and characterized. Cytotoxic evaluations of these molecules over a panel of three human cancer cell lines were carried out. Few molecules exhibited potent growth inhibitory action against the treated cancer cell lines at lower micro molar concentration. An in vitro assay investigation of these active compounds using recombinant human SIRT1 enzyme showed that one of the compounds (IT-14) inhibited the deacetylation activity of the enzyme. The in vivo study of IT-14 exemplified its promising action by reducing the prostate weight to the body weight ratio in prostate hyperplasia animal models. A remarkable decrease in the disruption of histoarchitecture of the prostate tissues isolated from IT-14 treated animal compared to that of the positive control was observed. The molecular interactions with SIRT1 enzyme were also supported by molecular docking simulations. Hence this compound can act as a lead molecule to treat prostatic hyperplasia. (C) 2013 Elsevier Masson SAS. All rights reserved.

Isolation and characterization of biofumigant from leaves of Lantana camara for control of stored grain insect pests

Due to environmental concerns, health hazards to man and the evolution of resistance in insect pests, there have been constant efforts to discover newer insecticides both from natural sources and by chemical synthesis. Natural sources for novel molecules hold promise in view of their eco-friendly nature, selectivity and mammalian safety. We have isolated one natural bioactive molecule from the leaves of Lantana camara named Coumaran, based on various physical-chemical and spectroscopic techniques (IR, H-1 NMR, C-13 NMR and MS). Coumaran is highly toxic and very low concentration is needed for control of stored product insects. This molecule has potent grain protectant potential and caused significant reduction in F1 progeny of all the three species in the treated grain and the progeny was completely suppressed at 30 mu g/l. The differences in germination between the control and treated grains were not significant. The lack of any adverse effect of Coumaran on the seed germination is highly desirable for a grain protectant, becoming a potential source of biofumigant for economical and environmentally friendly pest control strategies against stored grain pests during storage of grains or pulses. (C) 2013 Elsevier B.V. All rights reserved.

Modulation of glyceraldehyde-3-phosphate dehydrogenase activity by surface functionalized quantum dots

Enzymatic regulation is a fast and reliable diagnosis tool via identification and design of inhibitors for modulation of enzyme function. Previous reports on quantum dots (QDs)-enzyme interactions reveal a protein-surface recognition ability leading to promising applications in protein stabilization, protein delivery, bio-sensing and detection. However, the direct use of QDs to control enzyme inhibition has never been revealed to date. Here we show that a series of biocompatible surface-functionalized metal-chalcogenide QDs can be used as potent inhibitors for malignant cells through the modulation of enzyme activity, while normal cells remain unaffected. The in vitro activity of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), an enzyme involved critically in the glycolysis of cancer cells, is inactivated selectively in a controlled way by the QDs at a significantly low concentration (nM). Cumulative kinetic studies delineate that the QDs undergo both reversible and irreversible inhibition mechanisms owing to the site-specific interactions, enabling control over the inhibition kinetics. These complementary loss-of-function probes may offer a novel route for rapid clinical diagnosis of malignant cells and biomedical applications.

Molecular Basis for the Differential Quinolone Susceptibility of Mycobacterial DNA Gyrase

DNA gyrase is a type II topoisomerase that catalyzes the introduction of negative supercoils in the genomes of eubacteria. Fluoroquinolones (FQs), successful as drugs clinically, target the enzyme to trap the gyrase-DNA complex, leading to the accumulation of double-strand breaks in the genome. Mycobacteria are less susceptible to commonly used FQs. However, an 8-methoxy-substituted FQ, moxifloxacin (MFX), is a potent antimycobacterial, and a higher susceptibility of mycobacterial gyrase to MFX has been demonstrated. Although several models explain the mechanism of FQ action and gyrase-DNA-FQ interaction, the basis for the differential susceptibility of mycobacterial gyrase to various FQs is not understood. We have addressed the basis of the differential susceptibility of the gyrase and revisited the mode of action of FQs. We demonstrate that FQs bind both Escherichia coli and Mycobacterium tuberculosis gyrases in the absence of DNA and that the addition of DNA enhances the drug binding. The FQs bind primarily to the GyrA subunit of mycobacterial gyrase, while in E. coli holoenzyme is the target. The binding of MFX to GyrA of M. tuberculosis correlates with its effectiveness as a better inhibitor of the enzyme and its efficacy in cell killing.

Endoplasmic Reticulum Stress-Mediated Activation of p38 MAPK, Caspase-2 and Caspase-8 Leads to Abrin-Induced Apoptosis

Abrin from Abrus precatorius plant is a potent protein synthesis inhibitor and induces apoptosis in cells. However, the relationship between inhibition of protein synthesis and apoptosis is not well understood. Inhibition of protein synthesis by abrin can lead to accumulation of unfolded protein in the endoplasmic reticulum causing ER stress. The observation of phosphorylation of eukaryotic initiation factor 2 alpha and upregulation of CHOP (CAAT/enhancer binding protein (C/EBP) homologous protein), important players involved in ER stress signaling by abrin, suggested activation of ER stress in the cells. ER stress is also known to induce apoptosis via stress kinases such as p38 MAPK and JNK. Activation of both the pathways was observed upon abrin treatment and found to be upstream of the activation of caspases. Moreover, abrin-induced apoptosis was found to be dependent on p38 MAPK but not JNK. We also observed that abrin induced the activation of caspase-2 and caspase-8 and triggered Bid cleavage leading to mitochondrial membrane potential loss and thus connecting the signaling events from ER stress to mitochondrial death machinery.

Evaluation of benzothiophene carboxamides as analgesics and anti-inflammatory agents

Nonsteroid anti-inflammatory drugs (NSAIDs) represent standard therapy for the alleviation of pain and inflammation. At present various classes of compounds have been reported as selective inhibitors of cyclooxygenase-2 (COX-2). However, they are associated with adverse side effects. To address these issues, we report here a new class of compounds that exhibit potent analgesic and anti-inflammatory response. Substituted bromo-benzothiophene carboxamides (4-11) were examined for their analgesic and anti-inflammatory properties. Our findings demonstrate that newly synthesized bromo-benzothiophene carboxamide derivatives 4, 6, and 8 attenuate nociception and inflammation at lower concentration than classical NSAIDs, such as ibuprofen. These compounds act by selectively inhibiting COX-2 and by disrupting the prostaglandin-E2-dependent positive feedback of COX-2 regulation, which was further substantiated by reduction in the levels of cytokines, chemokines, neutrophil accumulation, synthesis of prostaglandin-E2, expression of COX-2, and neutrophil activation at lower concentration than the classic NSAID ibuprofen. Toxicological study reveals that these compounds are well tolerated and metabolized to avoid any toxicity. Thus, these molecules represent a new class of analgesic and anti-inflammatory agents. (c) 2014 IUBMB Life, 66(3):201-211, 2014

Synthesis, antimicrobial and cytotoxic activities of pyrimidinyl benzoxazole, benzothiazole and benzimidazole

A variety of pyrimidinyl benzoxazoles, benzothiazoles and benzimidazoles linked by thio, methylthio and amino moieties were prepared and studied their antimicrobial and cytotoxic activities. The compound pyrimidinyl bis methylthio benzimidazole 22 was a potent antimicrobial agent particularly against Staphylococcus aureus (29 mm, MIC 12.5 mu g/mL) and Penicillium chrysogenum (38 mm, MIC 12.5 mu g/mL). The amino linked pyrimidinyl bis benzothiazole 24 exhibited cytotoxic activity on A549 cells with IC50 value of 10.5 mu M. (C) 2014 Elsevier Masson SAS. All rights reserved.