PubMed Health⌕ Search

Biomedical subjects

M R Boyd

Publications and source records attributed to M R Boyd.

At least 55 records · Page 3Linked to original sources
PubMed2000

Running away to nowhere: rural women's experiences of becoming alcohol dependent.

The purpose of this study was to describe women's perspectives in becoming and being alcohol dependent. Part 1, "Alienation From Self and Others," on page 134 in this issue of Archives, describes the study methods, the sample, and the basic psychosocial problem faced by rural, alcoholic women. Part 2, "Running Away to Nowhere," focuses on the basic psychosocial process that women used to resolve the pain caused by their "Alienation From Self and Others." The article concludes with suggestions for nursing intervention.

Adaptation, Psychological↗
PubMed2000

Analysis of the interaction between the HIV-inactivating protein cyanovirin-N and soluble forms of the envelope glycoproteins gp120 and gp41.

The novel virucidal protein cyanovirin-N (CV-N) binds with equally high affinity to soluble forms of either H9 cell-produced or recombinant glycosylated HIV-1 gp120 (sgp120) or gp160 (sgp160). Fluorescence polarization studies showed that CV-N is also capable of binding to the glycosylated ectodomain of the HIV-envelope protein gp41 (sgp41) (as well as SIV glycoprotein 32), albeit with considerably lower affinity than the sgp120/CV-N interaction. Pretreatment of CV-N with either sgp120 or sgp41 abrogated the neutralizing activity of CV-N against intact, infectious HIV-1 virions. Isothermal calorimetry and optical biosensor binding studies showed that CV-N bound to recombinant sgp120 with a K(d) value ranging from 2 to 45 nM and to sgp41 with a K(d) value of 606 nM; furthermore, they indicated an approximate 5:1 stoichiometry for CV-N binding to sgp120 and a 1:1 stoichiometry for CV-N binding to sgp41. Circular dichroism studies additionally illuminated the binding of CV-N with both sgp120 and sgp41, providing the first direct evidence that conformational changes are a consequence of CV-N interactions with both HIV-1 envelope glycoproteins.

Anti-HIV Agents↗
PubMed2000

Multiple antiviral activities of cyanovirin-N: blocking of human immunodeficiency virus type 1 gp120 interaction with CD4 and coreceptor and inhibition of diverse enveloped viruses.

Cyanovirin-N (CV-N) is a cyanobacterial protein with potent neutralizing activity against human immunodeficiency virus (HIV). CV-N has been shown to bind HIV type 1 (HIV-1) gp120 with high affinity; moreover, it blocks the envelope glycoprotein-mediated membrane fusion reaction associated with HIV-1 entry. However, the inhibitory mechanism(s) remains unclear. In this study, we show that CV-N blocked binding of gp120 to cell-associated CD4. Consistent with this, pretreatment of gp120 with CV-N inhibited soluble CD4 (sCD4)-dependent binding of gp120 to cell-associated CCR5. To investigate possible effects of CV-N at post-CD4 binding steps, we used an assay that measures sCD4 activation of the HIV-1 envelope glycoprotein for fusion with CCR5-expressing cells. CV-N displayed equivalently potent inhibitory effects when added before or after sCD4 activation, suggesting that CV-N also has blocking action at the level of gp120 interaction with coreceptor. This effect was shown not to be due to CV-N-induced coreceptor down-modulation after the CD4 binding step. The multiple activities against the HIV-1 envelope glycoprotein prompted us to examine other enveloped viruses. CV-N potently blocked infection by feline immunodeficiency virus, which utilizes the chemokine receptor CXCR4 as an entry receptor but is CD4 independent. CV-N also inhibited fusion and/or infection by human herpesvirus 6 and measles virus but not by vaccinia virus. Thus, CV-N has broad-spectrum antiviral activity, both for multiple steps in the HIV entry mechanism and for diverse enveloped viruses. This broad specificity has implications for potential clinical utility of CV-N.

Animals↗
PubMed2000

Development of a cyanovirin-N-HIV-1 gp120 binding assay for high throughput screening of natural product extracts by time-resolved fluorescence.

The unique, high-affinity binding of cyanovirin-N (CV-N), a potent anti-human immunodeficiency virus (HIV) protein, to the HIV envelope glycoprotein gp120, was exploited to develop an HTS assay in an attempt to discover small-molecule mimetics of CV-N. A competition binding assay was developed using CV-N labeled with europium (Eu(3+)). The labeling protocol did not significantly alter the gp120 binding properties or the antiviral activity of CV-N. This report describes the assay development, validation, and results of screening a large library of aqueous and organic natural product extracts. The extracts were incubated with immobilized recombinant gp120 in 96-well plates prior to the addition of Eu(3+)-labeled CV-N. Following a wash step, bound CV-N was measured by dissociation-enhanced time-resolved fluorometry of Eu(3+). The assay proved to be robust, rapid, and reproducible, and was used to screen over 50,000 natural product extracts, and has resulted in the identification of several aqueous natural product extracts that inhibited CV-N-gp120 binding and also had anti-HIV activity.

Anti-HIV Agents↗
PubMed2000

Properties of cyanovirin-N (CV-N): inactivation of HIV-1 by sessile cyanovirin-N (sCV-N).

Cyanovirin-N (CV-N) is a novel anti-HIV protein isolated and characterized from a cyanobacterium Nostoc ellipsosporum. CV-N protein is a single 101 amino acid chain containing two intrachain disulphide bonds and considerable internal sequence duplication, but no significant homology to previously described proteins or to the transcription products of known nucleotide sequences. In solution, CV-N exists largely as a beta-sheet protein with internal two-fold pseudosymmetry. CV-N irreversibly inactivates diverse laboratory strains, primary isolates and clades of HIV-1, as well as strains of HIV-2 and simian immunodeficiency virus (SIV). CV-N binds with extremely high affinity to highly conserved binding site(s) on the viral envelope glycoprotein gp120, preventing virus-to-cell fusion, viral entry and infection of cells. The CV-N binding site appears to overlap, but is not identical with, the unique carbohydrate-dependent epitope 2G12, and may lie predominantly within an immunologically "silent" region of gp120. CV-N is undergoing preclinical development for topical anti-HIV prophylactic (e.g., microbicidal) applications to prevent sexual transmission of HIV. Since CV-N may be immunogenic in humans, methods for using CV-N for ex vivo inactivation of HIV in blood, plasma, or putative vaccines preferably would allow for its exclusion from biologicals for parenteral use. To explore this concept we biotinylated CV-N (bCV-N) and coupled it to streptavidin coated magnetic beads to provide a product which we termed sessile CV-N (sCV-N). When reacted with a laboratory strain and a primary isolate of HIV- 1, the sCV-N completely inactivated 100 TCID50 of the virus. However RT-PCR of the viral extracts indicated that only a fraction of the virus was removed by the sCV-N, leaving behind a relatively larger fraction of non-infectious virus in the supernatant which we designated as replication incompetent virions (RIV). It would be worthwhile investigating the role of RIV as a putative HIV vaccine.

Administration, Intravaginal↗
PubMed2000

Antineoplastic agents 442. Synthesis and biological activities of dioxostatin.

A high-yield regioselective synthesis of (E)-combretastatin A-1 2b was completed using methoxymethyl (MOM) protection and a Wadsworth-Emmons reaction as key steps. In turn, (E)-stilbene 11 was converted by convenient syntheses to both (S,S)- and (R,R)-1,3-dioxolanes 5a and 6a. A Sharpless asymmetric dihydroxylation reaction was employed for preparation of intermediates (S,S)-12 and (R,R)-13. The (4S,5S)-4-(2',3'-dihydroxy4'-methoxyphenyl)-5-(3",4",5"-trimethoxyphenyl)-1, 3-dioxolane 5a was found to be a highly potent inhibitor of microtubule assembly (IC50 = 0.59 microM) and was designated dioxostatin. Conversion to sodium phosphate 17 (P388 lymphocytic leukemia cell line: ED50 = 0.2 microg/ml) provided a very useful water-soluble prodrug.

Angiogenesis Inhibitors↗
PubMed2000

Antineoplastic agents 463. Synthesis of combretastatin A-3 diphosphate prodrugs.

A new and more efficient synthesis of combretastatin A-3 (2a) was completed (8.4% overall yield) starting from methyl gallate and isovanillin with aldehyde 5 and phosphonium salt 8 as key intermediates. Conversion of combretastatin A-3 (2a) to a series of diphosphate prodrugs (10a-l) was readily achieved. Both the diphosphate sodium (10a) and potassium salts (10c) displayed aqueous solubility in excess of 220 mg/ml at room temperature and good cancer cell line inhibitory activity.

Anisoles↗
PubMed1999

Effect of thiol modification on brain mitochondrial complex I activity.

We have studied the effect of altered thiol status on brain mitochondrial complex I activity. Exposure of mouse brain slices to diethyl maleate resulted in significant loss in complex I activity with concomitant loss of thiol antioxidant, glutathione. However, incubation of mitochondria isolated from diethyl maleate treated slices with dithiothreitol completely regenerated activity of complex I. Incubation of mouse brain slices with iodoacetic acid, a thiol modifier, also resulted in inhibition of complex I activity, which was reversed by glutathione. The present study demonstrates sensitivity of complex I to thiol modifying agents. Reversal of this inhibition by thiol reductants indicates that secondary oxidation of thiol groups in complex I as a consequence of thiol modification is responsible for the loss in activity.

Animals↗
PubMed1999

On topography and functionality in the B-D rings of cephalostatin cytotoxins.

Analogues 12'beta-hydroxycephalostatin 1 (9), 7'-deoxyritterazine G (10), and 14-epi-7'-deoxyritterazine B (11) were prepared via our protocol for unsymmetrical pyrazine synthesis. Cytotoxicity against human tumors was also determined for the first time for ritterazines, with femtomolar potency and a high correlation to cephalostatins observed. The SAR of these and related compounds provide insight into the importance of topography and certain chemical functionality in the B-D and B'-D' rings of cephalostatin type antineoplastics.

Antineoplastic Agents↗
PubMed1999

Crystal structure of cyanovirin-N, a potent HIV-inactivating protein, shows unexpected domain swapping.

The crystal structure of cyanovirin-N (CV-N), a protein with potent antiviral activity, was solved at 1.5 A resolution by molecular replacement using as the search model the solution structure previously determined by NMR. The crystals belong to the space group P3221 with one monomer of CV-N in each asymmetric unit. The primary structure of CV-N contains 101 residues organized in two domains, A (residues 1 to 50) and B (residues 51 to 101), with a high degree of internal sequence and structural similarity. We found that under the conditions of the crystallographic experiments (low pH and 26 % isopropanol), two symmetrically related monomers form a dimer by domain swapping, such that domain A of one monomer interacts with domain B' of its crystallographic symmetry mate and vice versa. Because the two swapped domains are distant from each other, domain swapping does not result in additional intramolecular interactions. Even though one of the protein sample solutions that was used for crystallization clearly contained 100 % monomeric CV-N molecules, as judged by various methods, we were only able to obtain crystals containing domain-swapped dimers. With the exception of the unexpected phenomenon of domain swapping, the crystal structure of CV-N is very similar to the NMR structure, with a root-mean-square deviation of 0.55 A for the main-chain atoms, the best agreement reported to date for structures solved using both techniques.

Antiviral Agents↗
PubMed1999

Antineoplastic agents. 410. Asymmetric hydroxylation of trans-combretastatin A-4.

The South African willow tree Combretum caffrum has yielded a number of potent cancer cell growth inhibitors. The present SAR studies of the antineoplastic agent combretastatin A-4 (1c) were focused mainly on the olefinic bridge to determine the effects on cancer cell growth and, potentially, to better define the combretastatin A-4 binding site on tubulin. The geometric trans-isomer 3a of combretastatin A-4 was converted to the (1S,2S)- and (1R,2R)-vicinal diols 4c and 4d, respectively, under Sharpless' asymmetric dihydroxylation conditions. Cancer cell line testing showed the (1S, 2S)-diol 4c to be more potent than its enantiomer 4d. Diol 4c weakly inhibited tubulin polymerization (IC50 = 22 microM, versus 1.2 microM for combretastatin A-4), while 4d was inactive (IC50 > 40 microM). Esterification of either stereoisomer at the diol and/or phenolic positions resulted in elimination of inhibitory activity.

Animals↗
PubMed1999

Solution structure by NMR of circulin A: a macrocyclic knotted peptide having anti-HIV activity.

The three-dimensional solution structure of circulin A, a 30 residue polypeptide from the African plant Chassalia parvifolia, has been determined using two-dimensional 1H-NMR spectroscopy. Circulin A was originally identified based upon its inhibition of the cytopathic effects and replication of the human immunodeficiency virus. Structural restraints consisting of 369 interproton distances inferred from nuclear Overhauser effects, and 21 backbone dihedral and nine chi1 angle restraints from spin-spin coupling constants were used as input for simulated annealing calculations and energy minimisation in the program X-PLOR. The final set of 12 structures had mean pairwise rms differences over the whole molecule of 0.91 A for the backbone atom, and 1.68 A for all heavy atoms. For the well-defined region encompassing residues 2-12 and 18-27, the corresponding values were 0.71 and 1.66 A, respectively. Circulin A adopts a compact structure consisting of beta-turns and a distorted segment of triple-stranded beta-sheet. Fluorescence spectroscopy provided additional evidence for a solvent-exposed Trp residue. The molecule is stabilised by three disulfide bonds, two of which form an embedded loop completed by the backbone fragments connecting the cysteine residues. A third disulfide bond threads through the centre of this loop to form a "cystine-knot" motif. This motif is present in a range of other biologically active proteins, including omega-contoxin GVIA and Cucurbita maxima trypsin inhibitor. Circulin A belongs to a novel class of macrocyclic peptides which have been isolated from plants in the Rubiaceae family. The global fold of circulin A is similar to kalata B1, the only member of this class for which a structure has previously been determined.

Amino Acid Sequence↗
PubMed1999

Inhibition of mitochondrial complex I by haloperidol: the role of thiol oxidation.

We have examined the effects of a variety of classical and atypical neuroleptic drugs on mitochondrial NADH ubiquinone oxido-reductase (complex I) activity. Sagittal slices of mouse brain incubated in vitro with haloperidol (10 nM) showed time- and concentration-dependent inhibition of complex I. Similar concentrations of the pyridinium metabolite of haloperidol (HPP+) failed to inhibit complex I activity in this model; indeed, comparable inhibition was obtained only at a 10000-fold higher concentration of HPP+ (100 microM). Treatment of brain slices with haloperidol resulted in a loss of glutathione (GSH), while pretreatment of slices with GSH and alpha-lipoic acid abolished haloperidol-induced loss of complex I activity. Incubation of mitochondria from haloperidol treated brain slices with the thiol reductant, dithiothreitol, completely regenerated complex I activity demonstrating thiol oxidation as a feasible mechanism of inhibition. In a comparison of different neuroleptic drugs, haloperidol was the most potent inhibitor of complex I, followed by chlorpromazine, fluphenazine and risperidone while the atypical neuroleptic, clozapine (100 microM) did not inhibit complex I activity in mouse brain slices. The present studies support the view that classical neuroleptics such as haloperidol inhibit mitochondrial complex I through oxidative modification of the enzyme complex.

Animals↗
PubMed1999

Cytotoxic and tubulin-interactive hemiasterlins from Auletta sp. and Siphonochalina spp. sponges.

Chemical and biological investigations of extracts from the sponge genus Auletta and two collections of Siphonochalina sp. have shown these organisms to be producers of the potent hemiasterlin class of antitumor agents. In addition to the previously known hemiasterlin (1) and hemiasterlin A (2), a new analogue, hemiasterlin C (3), was isolated and identified. The structures of 1 and 2 were assigned based on comparison to literature values, and 3 was identified on the basis of 1H NMR, 13C NMR, COSY, HSQC, and HMBC experiments. The cytotoxic and antitubulin activities of 1-3 were evaluated. In a comparative assay for inhibition of tubulin polymerization, the hemiasterlins were more potent than dolastatin 15 and equipotent with cryptophycin 1, but were somewhat less potent than dolastatin 10.

Animals↗
PubMed1999

Guttiferone F, the first prenylated benzophenone from Allanblackia stuhlmannii.

The HIV-inhibitory activity in extracts of Allanblackia stuhlmannii was tracked, via bioassay-guided fractionation, to a new member of the camboginol/guttiferone class of prenylated benzophenones, guttiferone F (1). The structure was solved by extensive NMR analyses and by acid-catalyzed conversion to 30-epi-cambogin (4). This is the first report of this compound type in the genus Allanblackia.

Anti-HIV Agents↗
PubMed1999

Vismiaphenones D-G, new prenylated benzophenones from Vismia cayennensis.

Following anti-HIV bioassay-guided fractionation, four new prenylated benzophenones, vismiaphenones D-G (7-10), were isolated from extracts of leaves of Vismia cayennensis. The structures were elucidated by spectral analyses. Only vismiaphenone D (7) exhibited HIV-inhibitory activity in the NCI primary screen.

Anti-HIV Agents↗
PubMed1999

Alertenone, a dimer of suberosenone from Alertigorgia sp.

Bioassay-guided fractionation of organic extracts of the gorgonian Alertigorgia sp. has yielded the previously known suberosenone (1), a cytotoxic tricyclic sesquiterpene of the quadrone class, and alertenone (2), a dimer of suberosenone. The structure of 2 was determined by spectral analysis; the 1D TOCSY experiment was particularly useful in the structure elucidation. Comparison of the in vitro cytotoxicity of alertenone and suberosenone revealed that the dimeric alertenone was devoid of cytotoxicity below 35 microg/mL. In a hollow-fiber assay model of in vivo activity, suberosenone exhibited some growth inhibition of two of six tumor cell lines tested.

Animals↗