PubMed Health⌕ Search

Biomedical subjects

D R Phillips

Publications and source records attributed to D R Phillips.

At least 163 records · Page 9Linked to original sources
PubMed1989

A novel photoproduct of 2'-deoxyguanosine induced by acetone photosensitization: 8-(2,3,4-trihydroxybutyl)guanine.

Acetone photosensitisation of 2'-deoxyguanosine in deaerated aqueous solution gives 8-(2,3,4-trihydroxybutyl)guanine as a major photoproduct. Its structure and that of its tetraacetate have been determined primarily by high resolution 1H NMR and mass spectrometry; a di-isopropylidene derivative has also been prepared. Mechanistic aspects of this novel photochemical transformation are discussed, particularly in relation to the alkaline cleavage of acetone photosensitised DNA at the sites of guanine bases.

Acetone↗
PubMed1989

Drug-DNA dissociation kinetics. In vitro transcription and sodium dodecyl sulphate sequestration.

The rate of dissociation of actinomycin D from DNA was measured by sodium dodecyl sulphate (SDS) sequestration (37 degrees) from calf thymus DNA and a 24 base pair (bp) synthetic DNA containing one high affinity AGCT site for the drug. The time constants were 276 and 142 sec, respectively, and suggest a stabilising effect though positive cooperativity in heterogenous DNA, or from specific neighbouring sequences. The time constant for dissociation of actinomycin D from the AGCT site was 2900 sec as measured by an in vitro transcription assay at 37 degrees, and suggests that, under conditions of active transcription of the DNA, the drug-DNA complex has additional stabilising contributions, possibly by a cage effect from RNA polymerase, or by additional drug-RNA polymerase contacts.

DNA↗
PubMed1989

Synthesis and expression of the fibroblast fibronectin receptor in human monocytes.

Human monocytes adhere to fibronectin, but the receptor (or receptors) mediating this interaction has not been clearly identified. To examine the nature of this receptor, human monocytes were obtained by counter-current elutriation and were found to adhere to immobilized fibronectin but not to vitronectin or von Willebrand factor. Antibodies and peptides were used to determine whether monocyte adherence to immobilized fibronectin was mediated by a receptor similar to that which mediates the adherence of fibroblasts to fibronectin. Antibodies against both the alpha and beta subunits of the fibroblast fibronectin receptor (VLA-5) inhibited monocyte adherence to fibronectin. These antibodies immunoprecipitated two surface proteins from monocytes that migrated at 140 and 120 kD under nonreducing conditions. Surface proteins with identical apparent molecular weights were immunoprecipitated from surface-labeled MG-63 cells, a fibroblast-like cell line. A synthetic peptide containing the Arg-Gly-Asp-Ser (RGDS) sequence also inhibited monocyte adherence to fibronectin. cDNA hybridization experiments revealed the presence of mRNA for both the alpha and beta subunits of the fibroblast fibronectin receptor in human monocytes. We conclude that circulating monocytes express a fibronectin receptor that is structurally and functionally very similar, if not identical, to the fibronectin receptor in adherent fibroblasts, and that this receptor mediates monocyte adherence to immobilized fibronectin.

Antibodies, Monoclonal↗
PubMed1989

Synthesis of polyamine-linked bis-daunomycin hydrazones and their interaction with DNA.

A series of bis-anthracycline hydrazones were prepared, compounds 9, containing either two or three secondary amines in the linker chain as well as the corresponding acetyl (10), or benzoyl (11) protected derivatives. Compounds 9 could not be isolated in pure form as they decomposed during semi-preparative h.p.l.c. Compounds 10a-g and 11b were characterized by f.a.b. mass spectrometry. All compounds bis-intercalated into DNA as judged by viscometric lengthening of DNA and by decreased dissociation kinetics from DNA under conditions of SDS sequestration. The secondary amine containing derivatives (9) exhibited dissociation rates less than 10(-5) that of daunomycin. The apparent affinity of these compounds was so great that the dissociation constants could not be quantitated, and they were only liberated from DNA to monomeric species by a slow hydrolysis process of the hydrazone links. Protection of those derivatives containing two amines in the linker by acetyl (10) and benzoyl (11) groups, decreased the dissociation time constant to 490-2900 s (1400-8000 fold slower than daunomycin) with maximal DNA residence time corresponding to a linker length of 14 atoms (approximately 14 A). Addition of a third protected amine in the linker, compound 10g, enhanced the DNA residence time a further three-fold. In vitro inhibition of transcription analysis showed that all of the bis-anthracyclines exhibited a DNA sequence specificity for 5'-CpA-3' sites, with adjacent intercalation sites decreasing in the order CA greater than AC, TC greater than CT greater than GC, CG, CC, TA, and it is apparent that the presence of the basic amino functions in the bridging chain provides for greatly enhanced formation of the drug-DNA complex.

Antibiotics, Antineoplastic↗
PubMed1988

Transcriptional analysis of multisite drug-DNA dissociation kinetics: delayed termination of transcription by actinomycin D.

An in vitro transcription assay was used to measure the relative occupancy, sequence specificity, and dissociation kinetics of six actinomycin D binding sites on DNA during conditions of active transcription of the DNA from the lac UV5 promoter. Five of the sites contained a GpC sequence, with three of these having a common AGCT sequence that differed by up to an order of magnitude in affinity for the drug, as indicated by their relative occupany and dissociation kinetics. Positive cooperativity was observed by higher occupancy and slower dissociation kinetics for neighboring GpC sites on a different DNA fragment (UV5-lambda PL). Termination of transcription was observed at some drug binding sites, while complete drug-induced termination of transcription was seen 7-10 nucleotides downstream of two drug sites. This delayed termination was minimized when ITP was incorporated into the transcripts and suggests that a time delay is required to enable stable RNA hairpin helices to form. A model is presented of the role of RNA hairpin helices in delayed, drug-induced termination of transcription. The classical picture of DNA-binding drugs as inhibitors of transcription now appears too simplistic as it does not accommodate this phenomenon. It will be important to gain a greater understanding of the mechanism of this phenomenon of drug-induced termination of transcription, as there are many implications for the design of DNA-acting drugs.

Bacterial Proteins↗
PubMed1988

The 5'-CA DNA-sequence preference of daunomycin.

The DNA-sequence specificity of daunomycin was investigated by DNase I footprinting and an E. coli RNA polymerase transcription-inhibition assay. The 5'-CA sequence was identified as being the highest affinity binding site, although other modest affinity (5'-GC, CG, CT, TC, AC) and poor affinity sites (5'-AA, AT, TA) were also observed. The preference of daunomycin for 5'-CA nucleotide sequence suggests that its biological activity may arise from association with the 5'-CA-containing sequences thought to be associated with genetic regulatory elements in eukaryotes.

Adenine↗
PubMed1988

Fast atom bombardment mass spectrometry of some anthracycline and bisanthracycline derivatives.

Fast atom bombardment (FAB) mass spectra of daunomycin, four of its derivatives, seven bisanthracyclines and three mixed-functional daunomycin-acridine derivatives are reported. These anthracyclines all exhibited their expected [MH]+ ions and peaks corresponding to the fragmentations which are characteristic of the anthracycline moiety, and in addition the spectra showed enhanced [MH + n]+ (n = 1-4) ions which were attributed to reductive processes occurring in the liquid matrix under FAB conditions. Daunomycin was also observed to form a dimeric cluster ion [M2H]+ together with associated reduced ions under FAB conditions. We have found that FAB mass spectrometry is an ideal method for the qualitative analysis of large, non-volatile derivatives of anthracyclines.

Antibiotics, Antineoplastic↗
PubMed1988

Privatising residential care for elderly people: the geography of developments in Devon, England.

The growth in numbers of very elderly people is becoming a trend in many Western societies. Often, these people may come to require some kind of assisted living environment. In Britain during the 1980s the overwhelming growth of residential accommodation has been in the private rather than the public sector. This has links with a number of other trends in health care and other sectors of the economy which are moving towards privatisation. The reasons for this are discussed and a case study of the county of Devon introduced. A survey of about one-quarter of all 450 homes in the county in mid-1984 revealed that they had important characteristics as small businesses. Countywide, a marked concentration of private residential homes has developed in some coastal 'holiday' locations. However, there have recently been changes in this pattern and growth of numbers of homes in some main towns also. There have been certain adverse reactions to the growth of homes and in a few areas, planning authorities have attempted to prevent the development of local concentrations of homes. This has been related to other policies elsewhere to prevent the concentration and ghettoisation of service-dependent groups. The nature and results of such planning policies are briefly considered. The paper addresses the overall questions of the type of care we wish to provide for our elderly people and whether privatisation of this aspect of health and welfare services is justified. This poses an important area of research for medical geographers interested in service delivery and aspects of equity in health care provision.

Aged↗
PubMed1988

Accommodation for elderly persons in newly industrializing countries: the Hong Kong experience.

The newly industrializing countries seem set to follow many developed countries with a rapid growth in numbers of elderly people. This will throw considerable strain on their resources, particularly in the provision of services and accommodation for this group in society. Hong Kong is a leader amongst the newly industrializing countries, both in terms of its aging population (more than 11 percent of its people are now over 60 years of age) and in terms of economic growth. It has, however, only relatively recently embarked upon a comprehensive and integrated program to provide a wide range of accommodation for the elderly. This includes sheltered housing within Hong Kong's well-known public housing schemes, old people's homes, care-and-attention homes, and infirmaries. The private sector has also been growing rapidly in the 1980s with a considerable recent increase of old people's homes. A voluntary code of conduct for this sector was introduced in late 1986, but concerns remain about the quality of care and the rapid growth of private homes in the territory. The approaches to the provision of accommodation for the elderly are set in the context of Hong Kong's overall social policy development. Whilst the experience of relatively wealthy Hong Kong may not prove to be of relevance for the majority of Third World countries, it is argued that it may provide a model for other newly industrializing countries.

Adult↗
PubMed1988

Elucidation of the DNA sequence preferences of daunomycin.

The sequence specificity of daunomycin was assessed using competition equilibrium dialysis, DNAse I footprinting and an E. coli RNA polymerase transcription inhibition assay; similar studies were performed on adriamycin and a new bis-intercalating daunomycin dimer. The results clearly demonstrate that the highest affinity sites are CA for daunomycin and adriamycin, and CACA for the bis-daunomycin. Other modest affinity (GC, CG, CT, TC, CC, AC) and poor affinity binding sites (AA, AT, TA) were also observed. Our results are in agreement with (a) the observed 5'-pyrimidine-purine-3' sequence preference of intercalating drugs, (b) the reported role played by OH(9) of daunomycin in the stabilization of the drug/DNA intercalation complex, and (c) the thermodynamics of nearest neighbour base-pair unstacking at the intercalation site. The CA specificity of daunomycin and adriamycin suggests that their biological activity may arise from association with the CA containing sequences which are thought to be associated with genetic regulatory elements in eukaryotes. The implications for future anthracycline drug design are presented in this context.

Base Sequence↗
PubMed1988

Daunomycin-DNA dissociation kinetics.

The dissociation of daunomycin from DNA was examined using a sodium dodecyl sulfate-sequestered stopped flow procedure. Two dissociation processes were observed with calf thymus and bacterial DNA, with approximately 45% of the amplitude associated with the faster process. Both processes were largely independent of DNA sequence for bacterial DNA, comprising 30-70% (G + C) content. The rate of both processes increased by a similar amount with increasing ionic strength. The faster process decreased with decreasing drug loading, whereas the slower process was independent of drug loading. Only one dissociation event was observed for the dissociation of daunomycin from four different synthetic polydeoxynucleotides. All observations are consistent with a parallel model of sodium dodecyl sulfate-induced dissociation of daunomycin from DNA, where the two processes observed reflect two resolvable processes that may be comprised of a series of rate constants for the dissociation of drug from differing environments. The slower process observed with bacterial DNA (0.5-0.7 sec-1) is related to dissociation from preferential 5'-CA DNA-binding sites, whereas the faster process reflects dissociation of drug from lower affinity sites on heterogenous DNA (3.2-4.1 sec-1). Dissociation of daunomycin from four different synthetic polydeoxynucleotides (which did not contain the 5'-CA preferential daunomycin-binding site) exhibited dissociation rates characteristic of low affinity sites (3.3-4.8 sec-1).

Base Composition↗
PubMed1988

The DNA-association and biological activity of a new bis(14-thiadaunomycin).

A new bis-daunomycin has been synthesized and characterized by 13C-n.m.r. and reversed-phase h.p.l.c. The compound was found to be highly self-associated in aqueous solution and to bis-intercalate into DNA with a residence time about 200-fold greater than the parent compound daunomycin. Although the bis-daunomycin and its parent congener were taken up by V79 Chinese hamster ovarian leukaemia cells to similar extents, the cytotoxicity of the former was considerably lower. The possible in vivo relationship between DNA binding affinity and cytotoxicity is discussed.

Animals↗
PubMed1987

Comparison of cDNA-derived protein sequences of the human fibronectin and vitronectin receptor alpha-subunits and platelet glycoprotein IIb.

The fibronectin receptor (FnR), the vitronectin receptor (VnR), and the platelet membrane glycoprotein (GP) IIb-IIIa complex are members of a family of cell adhesion receptors, which consist of noncovalently associated alpha- and beta-subunits. The present study was designed to compare the cDNA-derived protein sequences of the alpha-subunits of human FnR, VnR, and platelet GP IIb. cDNA clones for the alpha-subunit of the FnR (FnR alpha) were obtained from a human umbilical vein endothelial (HUVE) cell library by using an oligonucleotide probe designed from a peptide sequence of platelet GP IIb. cDNA clones for platelet GP IIb were isolated from a cDNA expression library of human erythroleukemia cells by using antibodies. cDNA clones of the VnR alpha-subunit (VnR alpha) were obtained from the HUVE cell library by using an oligonucleotide probe from the partial cDNA sequence for the VnR alpha. Translation of these sequences showed that the FNR alpha, the VnR alpha, and GP IIb are composed of disulfide-linked large (858-871 amino acids) and small (137-158 amino acids) chains that are posttranslationally processed from a single mRNA. A single hydrophobic segment located near the carboxyl terminus of each small chain appears to be a transmembrane domain. The large chains appear to be entirely extracellular, and each contains four repeated putative Ca2+-binding domains of about 30 amino acids that have sequence similarities to other Ca2+-binding proteins. The identity among the protein sequences of the three receptor alpha-subunits ranges from 36.1% to 44.5%, with the Ca2+-binding domains having the greatest homology. These proteins apparently evolved by a process of gene duplication.

Amino Acid Sequence↗
PubMed1987

Synthetic peptides derived from fibrinogen and fibronectin change the conformation of purified platelet glycoprotein IIb-IIIa.

The glycoprotein IIb-IIIa complex (GP IIb-IIIa) is a platelet cell-surface receptor for fibrinogen and fibronectin. A carboxyl-terminal decapeptide of the fibrinogen gamma-chain (Leu-Gly-Gly-Ala-Lys-Gln-Ala-Gly-Asp-Val LGGAKQAGDV] and a tetrapeptide (Arg-Gly-Asp-Ser (RGDS] from the fibrinogen alpha-chain and the fibronectin cell-binding domain appear to mediate the binding of these ligands to GP IIb-IIIa. The present study was designed to examine the effects of these and related peptides on the structure of purified platelet GP IIb-IIIa. Treatment of GP IIb-IIIa with various synthetic peptides affected the glycoprotein so that GP IIb alpha became a substrate for hydrolysis by thrombin. The order of potency of these peptides was as follows: RGDS greater than LGGAKQAGDV greater than KGDS greater than RGES. This is the same order of potency in which these peptides inhibit fibrinogen binding to platelets. This effect was time-, temperature-, and concentration-dependent; RGDS induced a half-maximal effect at approximately 60 microM. In addition, RGDS, but not RGES, decreased the intensity of the intrinsic protein fluorescence of GP IIb-IIIa. Finally, the decapeptide or RGDS decreased the sedimentation coefficient of GP IIb-IIIa from 8.5 to 7.7 or 7.4 S, respectively, whereas RGES had a minimal effect. This decrease was accompanied by an increase in the Stoke's radius from 74 to 82 A with RGDS or 85 A with the decapeptide, indicating a peptide-induced unfolding of the GP IIb-IIIa complex. This change in conformation may be related to changes in the distribution and function of GP IIb-IIIa on the platelet surface that occur when adhesive proteins or peptides from the GP IIb-IIIa binding domains of these proteins bind to GP IIb-IIIa.

Binding Sites↗
PubMed1987

Platelet glycoprotein IIb-IIIa-like proteins mediate endothelial cell attachment to adhesive proteins and the extracellular matrix.

The adherence of human umbilical vein endothelial (HUVE) cells to adhesive matrix proteins was examined to determine if cell attachment and spreading were mediated by the glycoprotein (GP) IIb-IIIa complex on endothelial cells. The HUVE cells adhered well to glass slides that had been coated with fibronectin, vitronectin, fibrinogen, or von Willebrand factor but failed to adhere to albumin-coated or to uncoated slides. The HUVE cell attachment and spreading on vitronectin, fibrinogen, and von Willebrand factor were greatly inhibited by a GP IIb-IIIa monoclonal antibody (7E3). In contrast, HUVE cell attachment to fibronectin was not inhibited by 7E3 but was inhibited by a fibronectin-receptor antibody (alpha GP140), which had no effect on cell attachment to the other adhesive proteins. The 7E3 antibody, but not alpha GP140, disrupted HUVE cell monolayers by detaching cells from their naturally occurring extracellular matrix. These data indicate that platelet GP IIb-IIIa-like proteins mediate the adherence of HUVE cells to specific adhesive proteins and to the extracellular matrix.

Cell Adhesion↗
PubMed1987

Protein sequence of endothelial glycoprotein IIIa derived from a cDNA clone. Identity with platelet glycoprotein IIIa and similarity to "integrin".

Platelet membrane glycoprotein (GP) IIIa forms a Ca2+-dependent heterodimer complex with GP IIb. The GP IIb-IIIa complex constitutes the fibrinogen and fibronectin receptor on stimulated platelets. A biochemically and immunologically similar membrane glycoprotein complex is present on endothelial cells. A human umbilical vein endothelial cell cDNA library was screened using oligonucleotide probes designed from peptide sequences obtained from platelet GP IIIa. A cDNA clone was sequenced and found to encode a protein of 84.5 kDa. The translated endothelial cDNA contained five sequences that corresponded to peptide sequences in platelet GP IIIa, including the amino-terminal 19 residues. Thus, the endothelial and platelet forms of GP IIIa are apparently identical. Glycoprotein IIIa consists of a long amino-terminal extracellular domain with several potential N-linked glycosylation sites and four cysteine-rich tandem repeats, a 29-residue hydrophobic transmembrane segment, and a short carboxyl-terminal cytoplasmic domain. Glycoprotein IIIa has a 47% amino acid sequence homology to "integrin," a fibronectin receptor from chicken embryo fibroblasts. This homology suggests that GP IIIa is a member of a family of cell-surface adhesion receptors.

Amino Acid Sequence↗