Biomedical subjects

D F Lewis

Publications and source records attributed to D F Lewis.

At least 91 records · Page 5Linked to original sources

PubMed1996

Molecular modelling of CYP1A subfamily members based on an alignment with CYP102: rationalization of CYP1A substrate specificity in terms of active site amino acid residues.

1. Using a novel amino acid sequence alignment, proteins of the CYP1A subfamily have been produced from the CYP102 crystal structure template via residue replacement and energy minimization procedures. 2. Known substrates and inhibitors of CYP1A1 and CYP1A2 are shown to fit their respective active sites via key interactions with complementary amino acid residues. Substrates used in the modelling studies include: caffeine, PhIP, oestradiol, 2,4- and 2,5-diaminotoluenes, Glu-P-1, phenacetin, acetanilide, 7-methoxy and 7-ethoxyresorufins, 11-methyl cyclopenta[a]phenanthren-17-one, 7-ethoxycoumarin, aflatoxin B1, benzo[a]pyrene, benzo[a]pyrene-7,8-diol and 1'-hydroxy 3-methylcholanthrene. 3. A number of aspects relating to CYP1A substrate specificity and metabolism can be explained in terms of the enzyme models, as it is found that key interactions with active site amino acid residues direct CYP1A-mediated metabolism in the known positions.

Amino Acid Sequence↗

PubMed1996

Molecular modelling of CYP3A4 from an alignment with CYP102: identification of key interactions between putative active site residues and CYP3A-specific chemicals.

1. A structural model of CYP3A4 is reported on the basis of a novel amino acid sequence alignment between the CYP3 family and CYP102, a bacterial P450 of known crystal structure. 2. Construction of the CYP3A4 model from CYP102 is facilitated by the relatively high sequence homology between the two protein (52% homology; 27% identity) with many conservative amino acid changes, yielding a structure of low internal energy. 3. A considerable number of specific substrates, and some specific inhibitors, are shown to occupy the putative CYP3A4 active site via interactions with the same amino acid residues in almost all cases investigated. 4. The CYP3A4 model rationalizes the known positions of metabolism for many substrates of this major human P450 such that the route of metabolism in novel development compounds can be predicted.

Amino Acid Sequence↗

PubMed1995

Induction of hepatic CYP1A2 by the oral administration of caffeine to rats: lack of association with the Ah locus.

Caffeine was administered to male Wistar albino rats for two weeks at three concentrations, namely 0.1, 0.2 and 0.3%, and hepatic cytochrome P450-dependent mixed-function oxidase determined. Caffeine administration gave rise to a marked, dose-dependent increase in the O-deethylation of ethoxyresorufin and, to a lesser extent, in the O-depentylation of pentoxyresorufin. Erythromycin N-demethylase, p-nitrophenol hydroxylase and lauric acid hydroxylase activities, as well as total cytochrome P450 content were unaffected by this treatment. Immunoblot analysis revealed that caffeine gave rise to a dose-dependent increase in the hepatic CYP1A2, and at the highest dose only, CYP2B apoprotein levels. Apoprotein levels of CYP3A and CYP2E1 were not modulated by the treatment with caffeine at all dose levels studied. Caffeine could not displace [3H]TCDD from the rat hepatic cytosolic Ah receptor. Computer analysis showed that caffeine is essentially a planar molecule with an area/depth ratio 4.8, characteristic of CYP1A substrates/inducers. Molecular modelling revealed that the caffeine molecule could orientate itself within the putative CYP1A2 active site so as to facilitate demethylation of the N-1, N-3 and N-7 positions. However, at physiological pH, the N-9 nitrogen atom is likely to be partially protonated, allowing it to participate in an electrostatic interaction with the negatively-charged glutamate 318-residue, favouring N-3 demethylation, the major pathway of metabolism in both humans and animals. In conclusion caffeine, being essentially planar, is an inducer of CYP1A2 in rat liver.

Administration, Oral↗

PubMed1995

A quantitative structure-activity relationship study on a series of 10 para-substituted toluenes binding to cytochrome P4502B4 (CYP2B4), and their hydroxylation rates.

Molecular structural and molecular orbital calculations (AM1 method) are reported on a series of 10 para-substituted toluene derivatives and this structural information has been used to rationalize the differences between both rates of hydroxylation catalysed by cytochrome P4502B4 and binding to the same cytochrome P450, via the generation of quantitative structure-activity relationships (QSARs). It was found that the rate constant for hydroxylation can be described by a two-variable expression involving the dipole moment and volume of the solvent-accessible molecular surface (r = 0.98), whereas binding free energies are well characterized by combinations of molecular volume and various electronic frontier orbital parameters (r = 0.98 and 0.99). This study represents an advance on a previous evaluation by White and McCarthy (Arch Biochem Biophys 246: 19-32, 1986) who used empirical physico-chemical parameters to obtain similar results which were generally of lower statistical significance to those of the present work. The QSAR expressions suggest that both binding to P450 and metabolism for this series of compounds are dependent on the relative ability of the molecules to desolvate and occupy the heme binding site, together with electronic properties of the whole molecule and of the methyl group which undergoes hydroxylation.

Aryl Hydrocarbon Hydroxylases↗

PubMed1995

Recurrence rate of shoulder dystocia.

OBJECTIVE: Shoulder dystocia continues to be a major complication of obstetrics, and several factors have been identified to help predict its occurrence. A previous shoulder dystocia is one of the risk factors. However, the recurrence rate is unknown. The purpose of this study is to report the recurrence rate of shoulder dystocia. STUDY DESIGN: Our obstetric database was used to identify all vaginal deliveries between January 1983 through December 1992. A subset of vaginal deliveries complicated by shoulder dystocia was selected from this database. These records were reviewed to identify subsequent pregnancies, outcomes, risk factors, and demographic data. RESULTS: During the study period there were 37,465 total vaginal deliveries, with shoulder dystocia complicating 747 (overall rate 2%). Of these 747 cases, 101 patients had 123 subsequent vaginal deliveries, with shoulder dystocia complicating 17 of these pregnancies (13.8% recurrence rate, p < 0.0001). Comparisons were made between those patients with recurrent shoulder dystocia. CONCLUSION: Shoulder dystocia recurred at a rate approximately seven times higher than our primary rate. Whether patients with a history of shoulder dystocia should be offered an elective abdominal delivery requires further investigation.

Adolescent↗

PubMed1995

Preterm premature rupture of membranes and abruptio placentae: is there an association between these pregnancy complications?

OBJECTIVE: Our purpose was to determine whether the incidence of abruptio placentae is increased in pregnancies with preterm premature rupture of membranes and to assess whether certain clinical risk factors in this group predispose them to have abruptio placentae. STUDY DESIGN: A retrospective cohort study over a 2.5-year period was performed. The study group consisted of 756 singleton pregnancies between 20 and 36 weeks' gestation complicated by preterm premature rupture of membranes and managed expectantly. The control group consisted of 11,240 pregnancies not complicated by preterm premature rupture of membranes and delivered during the same time period. The incidence of abruptio placentae was compared between the two groups. The study group of patients with preterm premature rupture of membranes was further subdivided into cases with (n = 38) and without abruptio placentae (n = 718) and compared. Clinical factors such as admission amniotic fluid index, history of bleeding before or after rupture of membranes, incidence of intrapartum fetal distress, and low 5-minute Apgar scores (< 6), latency-to-delivery interval, gestational age and weight at delivery, and incidence of amnionitis and endometritis were compared. RESULTS: The incidence of abruptio placentae in the study group (38/756, 5%) was significantly higher than that in the control group (97/11, 240, 0.9%) (p < 0.001, odds ratio = confidence interval). Comparison of cases with preterm premature rupture of membranes with and without abruptio placentae demonstrated both groups to have a similar gestational age at delivery, birth weight, latency-to-delivery interval, amniotic fluid index, and infectious morbidity. The group with abruptio placentae had a significantly higher incidence of bleeding before rupture of membranes (six of 38, 15% vs eight of 718, 1%; p < 0.005) and of intrapartum fetal distress (18/38, 46% vs 49/718, 7%; p < 0.0009). CONCLUSIONS: Pregnancies complicated by preterm premature rupture of membranes that are managed expectantly are at significant risk for abruptio placentae. Preterm premature rupture of membranes in such cases is more often preceded by bleeding. These abruptions may predispose the patient to intrapartum fetal distress.

Abruptio Placentae↗

PubMed1995

The genotoxicity of benzanthracenes: a quantitative structure-activity study.

Molecular orbital (MO) evaluations of a series of 14 methyl-substituted benz[alpha]anthracenes, calculated by the complete neglect of differential overlap (CNDO/2) method, are reported. By quantitative structure-activity relationship (QSAR) analysis, the carcinogenic and mutagenic potencies of these compounds have been shown to be correlated with their electronic structures, namely, with the magnitude of the energy of the lowest unoccupied molecular orbital (LUMO). The log mutagenicity potencies for the series of 14 benz[alpha]anthracenes are negatively dependent on E(LUMO), with a correlation coefficient of 0.82, which is increased to 0.90 by inclusion in the QSAR of a second variable, namely Q3H, the electronic density in the highest occupied molecular orbital, E(HOMO), of carbon-3. E(LUMO) is also negatively correlated with mouse carcinogenicity of the benzanthracenes, with a correlation coefficient of 0.88 for tumour incidence, and of 0.83 for log carcinogenicity index. The carcinogenicity and mutagenicity of the individual members of this series of polycyclic aromatic hydrocarbons are discussed in terms of the relationships between molecular structure, electron density, metabolic activation and covalent binding of reactive intermediates.

Animals↗

PubMed1995

Latency period after preterm premature rupture of membranes: a comparison of ampicillin with and without sulbactam.

OBJECTIVE: To compare ampicillin with and without sulbactam with respect to the effect on the latency period after preterm premature rupture of membranes (PROM). METHODS: Patients with PROM at 25-35 weeks' gestation were offered participation in a randomized blinded trial comparing ampicillin-sulbactam with ampicillin. Evaluations for cervical pathogens were performed on admission and patients were followed-up with daily maternal and fetal evaluation. Maternal and neonatal outcomes were analyzed using indicated techniques. RESULTS: Fifty-three women were studied, with 25 receiving ampicillin-sulbactam and 28 receiving ampicillin. The ampicillin-sulbactam group had a significantly longer latency period (433 +/- 625 versus 143 +/- 165 hours, P = .03) and significantly fewer neonatal complications (five versus 20, P < .001). Although no neonatal infectious complications were observed in sulbactam-treated cases, there were four cases of neonatal sepsis and two of neonatal pneumonia in the ampicillin group. Also, significantly more neonates in the ampicillin group required prolonged oxygen and ventilatory support. There was no significant difference in maternal morbidity. CONCLUSIONS: In our population with preterm PROM, a broad-spectrum antibiotic that provides anaerobic coverage appears to extend latency and decrease neonatal morbidity without increasing adverse maternal outcome.

Adult↗

PubMed1995

Computer graphics analysis of the interaction of alkoxy methylenedioxybenzenes with cytochromes P4501.

A quantitative structure-activity relationship (QSAR) in a homologous series of alkoxy methylenedioxybenzenes (MDBs) is reported. Measurements of molecular dimensions from computer-generated space-filling structures have provided values for the shape parameter area/depth2. These have been shown to correlate with the extent of inhibition of ethoxyresorufin O-deethylase activity by a series of MDBs. The implication of this is that the MDB nucleus fits the cytochrome P4501 substrate binding site and that this ability decreases with increase in the alkyl chain length of the alkoxy substituent. These findings are in agreement with previous results relating to the spatial dimensions of the cytochrome P4501 binding site, showing that substrate specificity can be rationalized in terms of overall molecular shape.

Animals↗

PubMed1995

Molecular modelling of the human estrogen receptor and ligand interactions based on site-directed mutagenesis and amino acid sequence homology.

A molecular model of the human estrogen receptor is reported based on a new alignment with the alpha 1-antitrypsin sequence, a homologous protein of known crystal structure. The putative ligand binding site is situated roughly equidistant between the DNA binding and dimerization regions. This binding site contains a number of amino acid residues shown by site-directed mutagenesis to be associated with the binding of agonists and antagonists. This putative ligand binding pocket is well-defined within a loop of peptide, containing complementary amino acids for binding interactions with agonists and antagonists. A leucine-rich region, common to most steroid-binding proteins, is in an optimum position for dimerization leading to DNA interaction. It is likely that ligand binding influences dimerization and DNA interaction by a conformational change in the receptor via the transcriptional activation residues. This model suggests that ligand binding may affect the hydrogen bonding pattern such that transpeptide signalling is initiated. The model accommodates steroidal estrogens and antiestrogens as well as the non-steroidal partial antagonist, hydroxytamoxifen.

Computer Simulation↗

PubMed1995

Species variation in the metabolism of 15,16-dihydro-11-methylcyclopenta[a]phenanthren-17-one to its 3,4-dihydrodiol, the proximate carcinogen.

The title compound is a strong carcinogen, similar in potency to benzo[a]pyrene in mouse skin assay. This paper describes a comparison of its in vitro metabolism by hepatic microsomal preparations from mouse, rat, rabbit, hamster, dog, monkey and man. Metabolites were isolated by preparative high pressure liquid chromatography from the ethyl acetate extractable material and their structures tentatively assigned on the basis of their retention times and ultraviolet spectra, when possible by direct comparison with authentic synthetic specimens. Mass spectrometry was then used to confirm these assignments. All these animals produce the same range of metabolites derived exclusively from oxidation at the benzo-ring A, the five-membered ring D, and at the 11-methyl group. However, the amounts of individual metabolites varied substantially. In particular all the animals yielded the proximate carcinogen 3,4-dihydroxy-11-methyl-3,4,15, 16-tetrahydrocyclopenta[a]phenanthren-17-one, from which it is reasoned that all might be susceptible to its carcinogenic action. A rationalization for the observed distribution of the metabolites is proposed on the basis of a molecular model of the active site of cytochrome P450 1A1, the oxidative enzyme involved.

Animals↗

PubMed1995

A comparison of magnesium sulfate and indomethacin to magnesium sulfate only for tocolysis in preterm labor with advanced cervical dilation.

Preterm labor becomes more difficult to inhibit as the degree of cervical dilation increases. Indeed, some physicians do not even attempt tocolysis with advanced cervical dilation. We compared single- versus double-agent tocolytic therapy when the cervix was dilated 3 cm or greater. We conducted a retrospective study of 44 patients with preterm labor of unknown etiology and with cervical dilation of greater than 3 cm. At the admitting physician's discretion, patients were treated with either magnesium sulfate or with magnesium sulfate and indomethacin in combination. Longer duration of successful tocolysis was noted in the group that received both magnesium sulfate and indomethacin (368.3 hours versus 70.9 hours). No maternal complications occurred in either group. These pilot data suggest that tocolysis with magnesium sulfate and indomethacin is a safe, effective method of tocolysis in patients with advanced cervical dilation.

Adult↗

PubMed1995

Structural requirements at the melatonin receptor.

1. High affinity, specific binding sites for the pineal hormone, melatonin (5-methoxy N-acetyltryptamine) can be detected in chick brain membranes by use of the radiolabelled agonist, 2-[125I]-iodomelatonin (2-[125I]-aMT). 2. The affinity of a number of analogues of melatonin at the 2-[125I]-aMT binding site was determined and compared with an analysis of their electronic structure and significant quantitative relationships obtained. 3. The best correlations indicated that binding affinity was correlated with delta E, the difference between the frontier orbital energies, and QNH, the electron density in the highest occupied molecular orbital of the side-chain nitrogen atom. 4. These findings suggest that ligand binding may involve hydrogen bonding between the 5-methoxy and amide moieties of melatonin and complementary amino acid residues, and charge transfer interactions between the indole ring of melatonin and an aromatic amino acid in the receptor binding site. 5. A molecular model of a putative binding site is proposed based on the predicted amino acid sequence of the cloned Xenopus laevis melanophore melatonin receptor and the quantitative structure-affinity relationships observed in the present study.

Animals↗

PubMed1995

A retrospective evaluation of COMPACT predictions of the outcome of NTP rodent carcinogenicity testing.

The carcinogenic potentials of 40 National Toxicology Program chemicals previously predicted by Computer Optimised Molecular Parametric Analysis for Chemical Toxicity (COMPACT), based on the identification of potential substrates of cytochromes P4501A and 2E (CYP1A and CYP2E), have been compared with new rodent carcinogenicity results. The COMPACT predictions have also been compared with published Ames mutagenicity data and with our own Hazardexpert predictions for carcinogenicity. Concordance evaluations between rodent carcinogenicity (1/4 segments positive) and predictions by COMPACT or Hazardexpert were 64% for COMPACT (CYP1A only), 72% for COMPACT (CYP1A plus CYP2E), 70% for Hazardexpert alone, and 86% for COMPACT (CYP1A plus CYP2E) plus Hazardexpert. Sensitivities of the predictions were for COMPACT, 75%; Hazardexpert, 60%; and Ames, 54%. Positive predictivities were for COMPACT, 75%; Hazardexpert, 78%; and Ames 81%. Negative predictivites were for COMPACT, 62%; Hazardexpert, 52%; and Ames, 42%.

Animals↗

PubMed1995

Three-dimensional models of human and other mammalian microsomal P450s constructed from an alignment with P450102 (P450bm3).

1. A novel modelling alignment for P450s, utilizing NADPH-P450 reductase for electron transfer, is proposed on the basis of analysis of their amino acid sequences. 2. Information used to facilitate the alignment process includes: the recent X-ray crystal structure of P450102 (P450bm3), site-directed mutagenesis experiments, chemical modification of specific residues, and antibody recognition studies. 3. The alignment has been used to construct a number of microsomal P450s of relevance to xenobiotic and endogenous metabolism.

Amino Acid Sequence↗

PubMed1995

Molecular modelling of members of the P4502A subfamily: application to studies of enzyme specificity.

1. Using the recently published crystal structure of a bacterial P450, namely 102 (also termed P450bm3), as a template molecular models of mammalian 2A1, 2A4, 2A5 and 2A6 were constructed. 2. Substrate interaction studies demonstrated that in keeping with known catalytic activities the putative binding sites of mouse hepatic P4502A4 and 2A5 oriented testosterone for 15 alpha-hydroxylation and coumarin for 7-hydroxylation respectively. 3. Substrate interaction studies with the putative binding site of human liver P4502A6 demonstrated that coumarin was oriented for 7-hydroxylation. However, in keeping with previous site-directed mutagenesis studies with P4502A4 and 2A5, changing a single phenylalanine residue to leucine in 2A6 gave rise to a mutant enzyme, which could bind testosterone as a substrate for 15 alpha-hydroxylation rather than coumarin. 4. Substrate interaction studies with the putative binding site of rat hepatic P4502A1 suggested that this isoenzyme would hydroxylate coumarin at the 3- rather than at the 7-position. 5. The results of these molecular modelling studies demonstrate that apparently minor modifications to P4502A subfamily amino acid sequences can result in major alterations in enzyme specificity. 6. Molecular modelling is thus a useful technique that can aid in elucidating substrate specificities of P450 isoenzymes and species differences in xenobiotic metabolism. The technique can also be utilized to complement site-directed mutagenesis studies in order to identify critical structural features of P450s and other enzymes.

Amino Acid Sequence↗

PubMed1995

Molecular orbital-generated QSARs in a homologous series of alkoxyresorufins and studies of their interactive docking with P450s.

1. Molecular and electronic structural parameters have been determined, by molecular orbital (MO) calculations, for a homologous series of 8 alkoxyresorufins (methoxy- to octoxy-). 2. Quantitative structure-activity relationships (QSARs) between these structural parameters and the rates of metabolism of the alkoxyresorufins in hepatic microsomes from the 3-methylcholanthrene (MC)-, and phenobarbital (PB)-pretreated mouse, and the beta-naphthoflavone (beta NF)-pretreated rat have been established. 3. The most significant single relationship is between beta NF-induction of cytochrome P4501 (CYP1A) and the total nucleophilic superdelocalizability (sigma SN) for the eight compounds in the series. 4. For double regressions, the electronic charge on the alkoxy oxygen, Q(O), or alpha-carbon Q(C), is important when combined with the hydrophobic substituent constant (pi). 5. These findings indicate that the rates of metabolism of these alkoxyresorufins are dependent upon their ability to cross cellular membranes, to fit the relevant CYP1A binding site, and on their ability to accept electrons from a donor nucleophilic species. 6. A different set of parameters correlated with CYP2B activity, namely, parameters of overall shape, which indicates that the way in which the alkoxyresorufins fit the CYP2B site, determines their differences in specificity. 7. Computer graphic interactive docking studies of the alkoxyresorufins with their affinity-specific cytochromes P450, namely, methoxy- with CYP1A2; ethoxy- with CYP1A1; pentoxy- with CYP2B1; and benzyloxy- with CYP3A, have also been undertaken to show the specific interactions of the alkoxyresorufins with the binding sites of the individual P450s.

Amino Acid Sequence↗

PubMed1994

Comparison between rodent carcinogenicity test results of 44 chemicals and a number of predictive systems.

A comparison is made between a number of predictive systems including bacterial mutagenicity, structure alert and chronic toxicity (Tennant et al., 1990), COMPACT, Hazardexpert, and DEREK, with the outcome of the two species rodent carcinogenicity bioassay for 44 chemicals conducted under the NTP protocol. It is shown that, following minor updating of the rodent data in the light of pathology reports, there is a generally good agreement between various predictions and the actual carcinogenicity results. In particular, a combination of two methods of prediction produces an over 80% concordance with the rodent bioassay. Possible reasons for various discrepancies are discussed in light of xenobiotic metabolism and activation mechanisms of chemical carcinogenesis.

Animals↗