Biomedical subjects

S D Taylor

Publications and source records attributed to S D Taylor.

At least 19 recordsLinked to original sources

PubMed2005

Topically applied antibody fragments penetrate into the back of the rabbit eye.

AIMS: Antibody fragments have been shown to penetrate into the anterior chamber when applied to the cornea. The aim of this study was to investigate whether such fragments could penetrate into the vitreous cavity after topical administration to the ocular surface of rabbits. METHODS: An engineered single-chain variable-domain antibody fragment with specificity for an irrelevant rat determinant was applied as a 50 microl eye drop to the eyes of live rabbits at 20-min intervals over 12 h. Eye drops contained 0.8-1.1 mg/ml protein in a buffered salt solution supplemented with penetration and viscosity enhancers. Samples were collected by paracentesis from the vitreous cavity immediately postmortem. Antibody fragments in these samples were quantified by measuring the binding activity to specific antigen, using flow cytometry. RESULTS: Topically applied antibody fragments were detectable in the vitreous of rabbit eyes after 4-12 h but had cleared at 12 h following the final eye drop. Concentrations of the antibody fragment in the vitreous samples were estimated to be 50-150 ng/ml at 12 h. Penetration of the parental whole antibody into the vitreous was not observed. CONCLUSION: Antibody fragments penetrate into the vitreous chamber of the rabbit eye after topical administration to the ocular surface. Such fragments may have therapeutic potential for diseases affecting the posterior segment.

Administration, Topical↗

PubMed2005

Influence of format on in vitro penetration of antibody fragments through porcine cornea.

AIM: Antibody fragments, appropriately formulated, can penetrate through the ocular surface and thus have potential as therapeutic agents. The aim was to investigate the influence of protein fragment format on the kinetics and extent of ocular penetration in vitro. METHODS: Immunoglobulin single chain variable domain fragments of a murine monoclonal antibody with specificity for rat CD4 were engineered with a 20 or 11 amino acid linker by assembly polymerase chain reaction, expressed in Escherichia coli and purified by chromatography. Fab fragments of the parental antibody were prepared by papain digestion. Antibody fragments were formulated with a penetration and a viscosity enhancer and were applied to the surface of perfused pig corneas for up to 10 hours in vitro. Penetration was quantified by flow cytometry on rat thymocytes. RESULTS: 20-mer antibody fragments formed natural monomers and dimers following purification that could be separately isolated, while 11-mer fragments were dimeric. All formats of fragment (20-mer monomers and dimers, 11-mer dimers, Fab) showed penetration through the pig cornea after 6 hours of intermittent topical administration. CONCLUSION: Antibody fragments of different shapes and sizes can penetrate the cornea after topical administration, thereby increasing the potential of this class of proteins for topical ophthalmic use.

Administration, Topical↗

PubMed2001

Structure of protein tyrosine phosphatase 1B in complex with inhibitors bearing two phosphotyrosine mimetics.

Protein tyrosine phosphatases (PTPases) are signal-transducing enzymes that dephosphorylate intracellular proteins that have phosphorylated tyrosine residues. It has been demonstrated that protein tyrosine phosphatase 1B (PTP1B) is an attractive therapeutic target because of its involvement in regulating insulin sensitivity (Elcheby et al. Science 1999, 283, 1544-1548). The identification of a second binding site in PTP1B (Puius et al., Proc. Natl. Acad. Sci. U.S.A.1997, 94, 13420-13425) suggests a new strategy for inhibitor design, where appropriate compounds may be made to simultaneously occupy both binding sites to gain much higher affinity and selectivity. To test this hypothesis and gain further insights into the structural basis of inhibitor binding, we have determined the crystal structure of PTP1B complexed with two non-peptidyl inhibitors, 4 and 5, both of which contain two aryl difluoromethylenephosphonic acid groups, a nonhydrolyzable phosphate mimetic. The structures were determined and refined to 2.35 and 2.50 A resolution, respectively. Although one of the inhibitors seems to have satisfied the perceived requirement for dual binding, it did not bind both the active site and the adjacent noncatalytic binding site as expected. The second or distal phosphonate group instead extends into the solvent and makes water-mediated interactions with Arg-47. The selectivity of the more potent of these two inhibitors, as well as four other inhibitors bearing two such phosphate mimetics for PTP1B versus seven other PTPases, was examined. In general, selectivity was modest to good when compared to PTPases Cdc25a, PTPmeg-1, PTPbeta, and CD45. However, selectivity was generally poor when compared to other PTPases such as SHP-1, SHP-2, and especially TCPTP, for which almost no selectivity was found. The implications these results have concerning the utility of dual-binding inhibitors are discussed.

Binding Sites↗

PubMed2001

Synthesis of protected L-4-[sulfono(difluoromethyl)phenylalanine and its incorporation into a peptide.

[reaction: see text] A protected form of L-4-[sulfono(difluoromethyl)]phenylalanine (F(2)Smp), a novel non-hydrolyzable phospho- and sulfotyrosine mimetic, was synthesized via electrophilic fluorination of a benzylic sulfonate followed by a Pd-catalyzed cross-coupling reaction between the fluorinated sulfonate and the zincate of protected iodoalanine. F(2)Smp was incorporated into a peptide using solid-phase peptide synthesis techniques.

Combinatorial Chemistry Techniques↗

PubMed2001

Gender differences in the self-regulation of hypertension.

The theory of planned behavior was adapted and revised to explain efforts made by patients to regulate their hypertension. A test of this theory of self-regulation revealed significant differences between men (N = 105) and women (N = 102) in their decision making and goal pursuit activities. For men, attitude toward success had both direct and indirect (through desire and intentions) effects on efforts to regulate hypertension. Attitudes had no influence on effort for women. Perceived behavioral control had both direct and indirect (through desire and intentions) effects on effort for men but no effects for women. Intentions influenced effort for men but not women. In contrast, perceived subjective norms had direct effects on effort for women but not for men. Effort was found also to be a function of past efforts at trying to regulate hypertension for both men and women. The study results confirm the existence of differences in the manner in which men and women perceive health maintenance. Additionally, the theory of self-regulation was found to serve as a useful framework in understanding the role of goal-setting and goal-striving in the management of chronic conditions.

Attitude↗

PubMed2000

Selective deuterium exchange during superheated heavy water chromatography-nuclear magnetic resonance spectroscopy-mass spectrometry of sulfonamides.

Superheated deuterium oxide has been investigated as an eluent for reversed-phase HPLC on a polystyrene-divinylbenzene column with UV, 1H NMR and MS detection using a series of sulfonamides as model compounds. In the course of these studies, a selective, specific and efficient deuteration of the methyl groups on a pyrimidine ring was observed during chromatography of certain of the sulfonamides. The potential of this methodology for producing deuterium-labelled compounds from substances bearing suitable substituents is considered. The utility of HPLC-NMR-MS as a means for studying on-column reactions is discussed.

Chromatography, Liquid↗

PubMed2000

Application of directly coupled HPLC-NMR-MS/MS to the identification of metabolites of 5-trifluoromethylpyridone (2-hydroxy-5-trifluoromethylpyridine) in hydroponically grown plants.

Directly coupled HPLC-NMR-MS was used to characterize two major metabolites of 5-trifluoromethylpyridone (2-hydroxy-5-trifluoromethylpyridine), a model compound for herbicides, after it had been dosed into hydroponically grown maize plants. The combination of NMR and MS data allowed the identification of both of these metabolites, namely, the N-glucoside and O-malonylglucoside conjugates of the parent pyridone. This work demonstrates the efficiency and the potential application of HPLC-NMR-MS to the investigation of the metabolism of agrochemicals. The work also indicates that combination of the use of hydroponically grown plants and directly coupled HPLC-NMR-MS allows rapid identification of metabolites with little sample preparation.

Chromatography, High Pressure Liquid↗

PubMed2000

An assessment of home remedy use by African Americans.

This analysis represents the first national look at family and individual use of home remedies by African Americans. The purpose is to examine home remedy usage by African-American individuals and their families and assess the relationship between sociodemographic characteristics and home remedy usage for African-American families and African-American individuals. Using logistic regression, a secondary analysis of the National Survey on Black Americans (NSBA) data (N = 2107) was conducted to examine factors associated with home remedy use. Multivariate analysis indicated that parent's education, importance of religion, living with a grandparent, and living in a rural area were associated with families' use of home remedies. Age, gender, living with a grandparent, education, and geographic region were associated with individual home remedy use. The results of this research may provide insight to health care practitioners in their challenge of appropriately integrating self-care practices (i.e., home remedy use) and the use of the formal health care system among the patients that utilize both "scientific" and "folk" medical systems. When possible, treatment plans should be adapted to consider patients' demographics, health beliefs, and self-care practices. Health care providers should encourage patient and family involvement and dialogue regarding therapeutic approaches. As more information becomes available, health care practitioners will be better able to ascertain the possible health consequences of concurrent usage of home remedies and prescription drug therapies.

Adolescent↗

PubMed1998

Novel phosphate mimetics for the design of non-peptidyl inhibitors of protein tyrosine phosphatases.

Benzylic alpha,alpha-difluorosulfonates, alpha,alpha-difluorotetrazoles, and alpha,alpha-difluorocarboxylates of type 5 and 6 were synthesized and examined as potential phosphate biosteres for PTP1B inhibition. The alpha,alpha-difluorosulfonates and alpha,alpha-difluorotetrazoles were found to be more effective inhibitors than the analogous compounds bearing the fluoromalonyl group, a phosphate biostere currently being used for PTP inhibition.

Drug Design↗

PubMed1998

Naphthalenebis[alpha,alpha-difluoromethylenephosphonates] as potent inhibitors of protein tyrosine phosphatases.

A series of naphthalenebis(difluoromethylenephosphonates) were prepared and compared to their monosubstituted counterparts as inhibitors of the protein phosphatases, PTP1B, CD45 and PP2A. In general, the bissubstituted compounds were better inhibitors than the mono derivatives and some of these are among the most potent, nonpeptidyl inhibitors of protein tyrosine phosphatases, PTP1B and CD45, reported to date.

Catalytic Domain↗

PubMed1998

Potent non-peptidyl inhibitors of protein tyrosine phosphatase 1B.

The development of inhibitors of protein tyrosine phosphatases (PTPs) has recently been the subject of intensive investigation due to their potential as chemotherapeutics and as tools for studying signal transduction pathways. Here we report the evaluation of a variety of small molecule, non-peptidyl inhibitors of protein tyrosine phosphatase 1B (PTP1B), bearing the alpha, alpha-difluoromethylenephosphonic acid (DFMP) group, a non-hydrolyzable phosphate mimetic. A series of phenyl derivatives bearing a single DFMP group were initially surveyed. In general, these were not significantly more potent inhibitors than the parent compound, alpha, alpha-difluorobenzylphosphonic acid, with the exception being the meta-phenyl substituted species which decreased the IC50 by approximately 17-fold relative to alpha, alpha-difluorobenzylphosphonic acid. However, certain compounds bearing two DFMP moieties were very potent inhibitors. Some of these are among the most potent small molecule inhibitors of any PTP reported to date with the best one exhibiting a Ki of 1.5 microM. The structural basis for these results are discussed. One of the bis-DFMP inhibitors was examined in detail and it was found that the fluorines were essential for potent inhibition. Inhibition was independent of pH between pH 5.5-7.2 suggesting that both the mono and dianionic forms of the individual DFMP groups bind equally well. The trends observed in the inhibitory potency of these compounds with PTP1B were very similar to the trends observed by other workers on the K(m)'s of the analogous phenylphosphate substrates with rat PTP1. This indicates that studies of non-peptidyl substrates with rat PTP1 can be used as a guide for the development of human PTP1B inhibitors.

Enzyme Inhibitors↗

PubMed1996

Evidence for a causal role of parathyroid hormone-related protein in the pathogenesis of human breast cancer-mediated osteolysis.

Breast cancer almost invariably metastasizes to bone in patients with advanced disease and causes local osteolysis. Much of the morbidity of advanced breast cancer is a consequence of this process. Despite the importance of the problem, little is known of the pathophysiology of local osteolysis in the skeleton or its prevention and treatment. Observations in patients with bone metastases suggest that breast cancer cells in bone express parathyroid hormone-related protein (PTHrP) more frequently than in soft tissue sites of metastasis or in the primary tumor. Thus, the role of PTHrP in the causation of breast cancer metastases in bone was examined using human breast cancer cell lines. Four of eight established human breast cancer cell lines expressed PTHrP and one of these cell lines, MDA-MB-231, was studied in detail using an in vivo model of osteolytic metastases. Mice inoculated with MDA-MB-231 cells developed osteolytic bone metastasis without hypercalcemia or increased plasma PTHrP concentrations. PTHrP concentrations in bone marrow plasma from femurs affected with osteolytic lesions were increased 2.5-fold over corresponding plasma PTHrP concentrations. In a separate experiment, mice were treated with either a monoclonal antibody directed against PTHrP(1-34), control IgG, or nothing before tumor inoculation with MDA-MB-231 and twice per week for 26 d. Total area of osteolytic lesions was significantly lower in mice treated with PTHrP antibodies compared with mice receiving control IgG or no treatment. Histomorphometric analysis of bone revealed decreased osteoclast number per millimeter of tumor/bone interface and increased bone area, as well as decreased tumor area, in tumor-bearing animals treated with PTHrP antibodies compared with respective controls. These results indicate that tumor-produced PTHrP can cause local bone destruction in breast cancer metastatic to bone, even in the absence of hypercalcemia or increased circulating plasma concentrations of PTHrP. Thus, PTHrP may have an important pathogenetic role in the establishment of osteolytic bone lesions in breast cancer. Neutralizing antibodies to PTHrP may reduce the development of destructive bone lesions as well as the growth of tumor cells in bone.

Animals↗

PubMed1995

Acid secretion during indomethacin therapy. Effect of misoprostol.

Secretin is an important physiologic regulator of gastric acid and pancreatic bicarbonate secretion. Endogenous prostaglandins play an important intermediary role, because indomethacin pretreatment prevents the physiological actions of secretin. Misoprostol is a prostaglandin E1 (PGE1) analog used for the prevention of nonsteroidal antiinflammatory drug (NSAID)-induced ulceration. This study sought to determine whether cotherapy with misoprostol reverses the NSAID-induced blockade of secretin's inhibition of human gastric acid secretion. Seven healthy volunteers were studied. Gastric acid secretion was measured during the basal, pentagastin-stimulated and intravenous secretin--inhibited states. The studies were then repeated after 3 days of oral indomethacin (50 mg p.o. t.i.d. for 10 doses) with and without concomitant misoprostol (200 micrograms q.i.d. for 13 doses). Exogenous secretin reduced pentagastrin-stimulated acid secretion by 35%, and this inhibition was reduced to 9% during indomethacin treatment (p < 0.05). Cotherapy with misoprostol during indomethacin treatment did not restore secretin's inhibition of gastric acid secretion; paradoxically, during secretin administration, acid secretion increased. When subjects received misoprostol treatment only, secretin failed to inhibit gastric acid secretion. We conclude that during indomethacin treatment, cotherapy with misoprostol cannot restore the inhibitory action of secretin to inhibit gastric acid secretion. These results suggest that PGE1 is an unlikely intermediate in secretin's physiological action. Although misoprostol can prevent NSAID-induced ulcers, it does not ameliorate all physiological perturbations induced by cyclooxygenase inhibition.

Adult↗

PubMed1995

Effects of parathyroid hormone (PTH)-related protein and PTH on osteoclasts and osteoclast precursors in vivo.

Increased production of PTH-related protein (PTHrP) and PTH is frequently responsible for hypercalcemia and its associated morbidity. However, it is unclear whether these peptides produce identical effects on cells in the osteoclast lineage in vivo. To examine the effects of continuous in vivo exposure to these factors on both the osteoclast precursors and mature osteoclasts, we inoculated Chinese hamster ovarian cells expressing PTH-(1-84), PTHrP-(1-141), or nontransfected Chinese hamster ovarian cells into nude mice. The effects of these tumors on blood ionized calcium, plasma PTH and PTHrP concentrations, and osteoclast formation were then determined. PTH and PTHrP tumor-bearing mice became hypercalcemic (1.90 +/- 0.04 and 1.97 +/- 0.16 mmol/liter, respectively) compared with control mice (1.29 +/- 0.015 mmol/liter). After 4 days of hypercalcemia, mice were killed, and bone marrow cells were harvested to examine cells at three discrete stages of osteoclast development: multipotent osteoclast precursors, the granulocyte/macrophage colony-forming unit; more committed marrow mononuclear osteoclast precursors; and mature osteoclasts. Neither PTH nor PTHrP had an effect on granulocyte/macrophage colony-forming unit, but similarly increased the number of more committed mononuclear osteoclast progenitors as well as mature osteoclasts in the calvaria. No differences were detected between the effects of PTH and PTHrP on cells in the osteoclast lineage in vivo. Thus, PTH and PTHrP appear to affect only more differentiated cells in the osteoclast lineage, and the differences in osteoclastic bone resorption between primary hyperparathyroidism and humoral hypercalcemia of malignancy are probably due to mechanisms other than effects on osteoclast precursor cells in vivo.

Animals↗

PubMed1994

Demand for predictive genetic testing for Huntington's disease in Australia, 1987 to 1993.

OBJECTIVE: To assess the demand for genetic testing for Huntington's disease among at-risk individuals and for prenatal testing among parents at risk of transmitting the disease, from January 1987 to March 1993. METHODS: Questionnaires were sent to all State coordinators of genetic testing services. RESULTS: One hundred and ninety adults (5.5% of those at risk) and 56 fetuses were tested. Nine per cent more women than men used the service. Most people tested were married and had offspring. Men generally presented for testing at an older age than women. CONCLUSION: The low uptake for testing is not surprising given the severity and untreatability of the illness. Careful monitoring is required, particularly of the effects on individuals told of their genetic status and of possible abuse of the information by third parties.

Adolescent↗

PubMed1994

Peptide synthesis catalyzed by an antibody containing a binding site for variable amino acids.

Monoclonal antibodies, induced with a phosphonate diester hapten, catalyzed the coupling of p-nitrophenyl esters of N-acetyl valine, leucine, and phenylalanine with tryptophan amide to form the corresponding dipeptides. All possible stereoisomeric combinations of the ester and amide substrates were coupled at comparable rates. The antibodies did not catalyze the hydrolysis of the dipeptide product nor hydrolysis or racemization of the activated esters. The yields of the dipeptides ranged from 44 to 94 percent. The antibodies were capable of multiple turnovers at rates that exceeded the rate of spontaneous ester hydrolysis. This achievement suggests routes toward creating a small number of antibody catalysts for polypeptide syntheses.

Antibodies, Catalytic↗

PubMed1994

Duodenal acidification and secretin, but not intraduodenal fat, inhibit human gastric acid secretion via prostaglandins.

BACKGROUND/AIMS: Acid and fat in the duodenum inhibit gastric acid secretion and increase plasma secretin. The role of prostaglandins and secretin in the inhibition of gastric acid secretion by duodenal infusion of hydrochloric acid and fat in healthy human volunteers was studied. METHODS: Gastric acid secretion was submaximally stimulated with intravenous pentagastrin followed by duodenal infusion of 0.1N hydrochloric acid, oleic acid, or intravenous secretin. To inhibit endogenous prostaglandins, the protocol was then repeated after indomethacin treatment. RESULTS: Duodenal fat infusion inhibited acid secretion 80% +/- 5% and was unaffected by indomethacin treatment. Intraduodenal acidification inhibited acid secretion by 43% +/- 8% and was reduced by indomethacin treatment to 15% +/- 4% (P < 0.01). Similarly, intravenous secretin inhibited acid secretion by 34% +/- 3%, which was decreased to 13% +/- 6% by indomethacin treatment (P < 0.01). The increase in plasma secretin levels after intraduodenal hydrochloric acid treatment was significantly greater than that observed with intravenous secretin or introduodenal oleic acid treatment; all were within the physiological range. Acid in the duodenum releases secretin, which inhibits gastric acid secretion at least in part via prostaglandins. In contrast, fat in the duodenum strongly inhibits gastric acid secretion via a nonprostaglandin pathway. CONCLUSIONS: Secretin is the predominant mediator for the inhibition of human gastric acid secretion induced by the presence of acid, but not fat, in the duodenum.

Adolescent↗

PubMed1993

Glutathione peroxidase protects cultured mammalian cells from the toxicity of adriamycin and paraquat.

Dihydrofolate reductase-minus mutants of Chinese hamster ovary cells were depleted of glutathione peroxidase by transcription of the transfected bovine cDNA in inverted orientation upstream from the cDNA for dihydrofolate reductase to engender a bicistronic mRNA. In a clone of cells selected for expression of dihydrofolate reductase by the ability to grow in nucleoside-free medium the activity of glutathione peroxidase was reduced to 20% of the activity in the untransfected parental line of cells (DG44). The cells depleted of glutathione peroxidase were more sensitive to the toxicities of paraquat and adriamycin than the untransfected parental cells from which they derived but not more sensitive to bleomycin, menadione, or phenazine methosulfate. That the mildly increased sensitivity to paraquat and adriamycin was the consequence of the diminished cellular content of glutathione peroxidase was confirmed by the increase in sensitivity of untransfected cells after treatment with buthionine sulfoximine, an agent which depletes cells of glutathione. These and other data strongly suggest that the enzymic action of glutathione peroxidase protects cells from the toxicity of paraquat and adriamycin. The toxin which these agents engender is likely to be hydrogen peroxide or another hydroperoxide upon which glutathione peroxidase acts.

Animals↗