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D A Adams

Publications and source records attributed to D A Adams.

At least 19 recordsLinked to original sources
PubMed2001

Two new classes of conopeptides inhibit the alpha1-adrenoceptor and noradrenaline transporter.

Cone snails use venom containing a cocktail of peptides ('conopeptides') to capture their prey. Many of these peptides also target mammalian receptors, often with exquisite selectivity. Here we report the discovery of two new classes of conopeptides. One class targets alpha1-adrenoceptors (rho-TIA from the fish-hunting Conus tulipa), and the second class targets the neuronal noradrenaline transporter (chi-MrIA and chi-MrIB from the mollusk-hunting C. marmoreus). rho-TIA and chi-MrIA selectively modulate these important membrane-bound proteins. Both peptides act as reversible non-competitive inhibitors and provide alternative avenues for the identification of inhibitor drugs.

Amino Acid Sequence↗
PubMed2000

Novel omega-conotoxins from Conus catus discriminate among neuronal calcium channel subtypes.

omega-Conotoxins selective for N-type calcium channels are useful in the management of severe pain. In an attempt to expand the therapeutic potential of this class, four new omega-conotoxins (CVIA-D) have been discovered in the venom of the piscivorous cone snail, Conus catus, using assay-guided fractionation and gene cloning. Compared with other omega-conotoxins, CVID has a novel loop 4 sequence and the highest selectivity for N-type over P/Q-type calcium channels in radioligand binding assays. CVIA-D also inhibited contractions of electrically stimulated rat vas deferens. In electrophysiological studies, omega-conotoxins CVID and MVIIA had similar potencies to inhibit current through central (alpha(1B-d)) and peripheral (alpha(1B-b)) splice variants of the rat N-type calcium channels when coexpressed with rat beta(3) in Xenopus oocytes. However, the potency of CVID and MVIIA increased when alpha(1B-d) and alpha(1B-b) were expressed in the absence of rat beta(3), an effect most pronounced for CVID at alpha(1B-d) (up to 540-fold) and least pronounced for MVIIA at alpha(1B-d) (3-fold). The novel selectivity of CVID may have therapeutic implications. (1)H NMR studies reveal that CVID possesses a combination of unique structural features, including two hydrogen bonds that stabilize loop 2 and place loop 2 proximal to loop 4, creating a globular surface that is rigid and well defined.

Alternative Splicing↗
PubMed2000

Conotoxin TVIIA, a novel peptide from the venom of Conus tulipa 1. Isolation, characterization and chemical synthesis.

A novel conotoxin belonging to the 'four-loop' structural class has been isolated from the venom of the piscivorous cone snail Conus tulipa. It was identified using a chemical-directed strategy based largely on mass spectrometric techniques. The new toxin, conotoxin TVIIA, consists of 30 amino-acid residues and contains three disulfide bonds. The amino-acid sequence was determined by Edman analysis as SCSGRDSRCOOVCCMGLMCSRGKCVSIYGE where O = 4-transL-hydroxyproline. Two under-hydroxylated analogues, [Pro10]TVIIA and [Pro10,11]TVIIA, were also identified in the venom of C. tulipa. The sequences of TVIIA and [Pro10]TVIIA were further verified by chemical synthesis and coelution studies with native material. Conotoxin TVIIA has a six cysteine/four-loop structural framework common to many peptides from Conus venoms including the omega-, delta- and kappa-conotoxins. However, TVIIA displays little sequence homology with these well-characterized pharmacological classes of peptides, but displays striking sequence homology with conotoxin GS, a peptide from Conus geographus that blocks skeletal muscle sodium channels. These new toxins and GS share several biochemical features and represent a distinct subgroup of the four-loop conotoxins.

Amino Acid Sequence↗
PubMed1999

Effects of chirality at Tyr13 on the structure-activity relationships of omega-conotoxins from Conus magus.

The effects of chirality inversions of Tyr13 on the structure-activity relationships of omega-conotoxins MVIIA and MVIIC were examined using a combination of 2D 1H NMR spectroscopy and radioligand binding studies specific for N-type ([125I]GVIA) and P/Q-type ([125I]MVIIC) voltage-sensitive calcium channels (VSCCs). A comparison of the Halpha secondary shifts suggests that the structural scaffolds of MVIIA and MVIIC are little altered by the L- to D- inversion of Tyr13; however, the conformations of several residues in loop 2 (residues 9-14) are significantly altered. The experimentally determined 3D structure of [D-Y13]MVIIA indicates that the positions of key residues in this loop which are involved in the binding of MVIIA to the N-type VSCC (Tyr13, Arg10, and Leu11) are so changed as to render the peptide unrecognizable by its cognate ion channel. The large reduction in potency observed for MVIIA and MVIIC at both N-type and P/Q-type VSCCs is likely to stem from the change in conformation and orientation of loop 2.

Amino Acid Sequence↗
PubMed1999

Structure-activity studies of conantokins as human N-methyl-D-aspartate receptor modulators.

The activities of conantokin-G (con-G), conantokin-T (con-T), and several novel analogues have been studied using polyamine enhancement of [3H]MK-801 binding to human glutamate-N-methyl-D-aspartate (NMDA) receptors, and their structures have been examined using CD and 1H NMR spectroscopy. The potencies of con-G[A7], con-G, and con-T as noncompetitive inhibitors of spermine-enhanced [3H]MK-801 binding to NMDA receptor obtained from human brain tissue are similar to those obtained using rat brain tissue. The secondary structure and activity of con-G are found to be highly sensitive to amino acid substitution and modification. NMR chemical shift data indicate that con-G, con-G[D8, D17], and con-G[A7] have similar conformations in the presence of Ca2+. This consists of a helix for residues 2-16, which is kinked in the vicinity of Gla10. This is confirmed by 3D structure calculations on con-G[A7]. Restraining this helix in a linear form (i.e., con-G[A7,E10-K13]) results in a minor reduction in potency. Incorporation of a 7-10 salt-bridge replacement (con-G[K7-E10]) prevents helix formation in aqueous solution and produces a peptide with low potency. Peptides with the Leu5-Tyr5 substitution also have low potencies (con-G[Y5,A7] and con-G[Y5,K7]) indicating that Leu5 in con-G is important for full antagonist behavior. We have also shown that the Gla-Ala7 substitution increases potency, whereas the Gla-Lys7 substitution has no effect. Con-G and con-G[K7] both exhibit selectivity between NMDA subtypes from mid-frontal and superior temporal gyri, but not between sensorimotor and mid-frontal gyri. Asn8 and/or Asn17 appear to be important for the ability of con-G to function as an inhibitor of polyamine-stimulated [3H]MK-801 binding, but not in maintaining secondary structure. The presence of Ca2+ does not increase the potencies of con-G and con-T for NMDA receptors but does stabilize the helical structures of con-G, con-G[D8,D17], and, to a lesser extent, con-G[A7]. The NMR data support the existence of at least two independent Ca2+-chelating sites in con-G, one involving Gla7 and possibly Gla3 and the other likely to involve Gla10 and/or Gla14.

Amino Acid Sequence↗
PubMed1998

Development of a comprehensive panel of antibodies against the major xenobiotic metabolising forms of cytochrome P450 in humans.

Mono-specific antibodies against the human cytochrome P450 (P450) enzymes CYP1A1, CYP1A2, CYP1B1, CYP2A6, CYP2B6, CYP2D6, CYP2E1, CYP3A4, CYP3A5 and CYP4A11 and an antibody that binds to CYP2C8, CYP2C9 and CYP2C19 have been produced by immunising rabbits with synthetic peptides representing small regions of each of these P450 enzymes. The specificity of the antibodies was confirmed by immunoblotting using recombinant P450 enzymes and samples of human hepatic microsomal fraction. Each of the antibodies bound only to their respective target P450 enzyme(s). The relative intensity of immunoreactive bands was compared with a variety of P450 activities and correlations were found between CYP1A2 and phenacetin O-deethylase activity, CYP2A6 and coumarin 7-hydroxylase activity, CYP2C9 and tolbutamide 4-hydroxylase activity, CYP2C19 and S-mephenytoin 4-hydroxylase activity, CYP2D6 and debrisoquine 4-hydroxylase activity, CYP2E1 and chlorzoxazone 6-hydroxylase activity, CYP3A4 and midazolam 1'-hydroxylase activity, and CYP4A11 and lauric acid 12-hydroxylase activity. A proportion of the 30 liver samples examined lacked CYP2A6 (7%), CYP2C19 (10%) or CYP2D6 (13%), consistent with the polymorphic expression of these P450 enzymes in human liver. Although CYP3A5 was detected in most individuals (97%), expression was polymorphic with 20% containing substantially higher levels. CYP2B6 was expressed in 20% of the human liver samples, with one sample containing a particularly high level. No immunodetectable CYP1A1 or CYP1B1 was found, consistent with the low level of expression of these P450 enzymes in human liver. The results demonstrate the utility of the antipeptide approach for producing specific antibodies against human P450 enzymes, enabling a comprehensive panel of antibodies against human P450 enzymes to be produced.

Amino Acid Sequence↗
PubMed1998

Interlaboratory comparison of the assessment of P450 activities in human hepatic microsomal samples.

1. Although the importance of in vitro technology in supporting drug development is widely accepted, there is no real consensus about which approaches should be taken, which substrates should be used, or on the reliability and application of in vitro data. Consequently, as part of a collaborative project to characterize human liver with respect to the major forms of cytochrome P450, an interlaboratory comparison of the analysis of samples for form-specific activities was undertaken. 2. Microsomal fractions were isolated from five different human liver samples taken from the liver bank maintained at the Royal Postgraduate Medical School (RPMS). Aliquots from the five samples were sent to the 11 collaborating laboratories for characterization using their in-house, form-specific assays for cytochrome P450 activities. Although each laboratory assayed protein concentration, total cytochrome P450 content and enzyme activities were calculated using the protein estimation generated by RPMS to eliminate this possible source of variability. 3. With the exception of one laboratory, all estimates of protein concentration were similar (coefficient of variation, CoV, 9-13%) and the rank-order of the five samples was consistent across the laboratories. There was greater variability in the estimates of total cytochrome P450 content (CoV 28-43%), although again rank order of the samples across laboratories was fairly consistent. 4. The various laboratories used a number of different probe substrates, together with a range of conditions (substrate concentration, time of incubation, amount of protein), to assay for activity of CYP1A2, CYP2C9, CYP2D6, CYP2E1 and CYP3A4. However, apart from the occasional outlier, the five samples were ranked for activity of all these forms of cytochrome P450 with a high degree of consistency by the various laboratories and the choice of substrate had no appreciable effect on the ranking of the samples. 5. While this interlaboratory comparison has shown that greater consistency in the approach to in vivo determination of drug-metabolizing activity is desirable, there was little indication that any particular approach or substrate was superior to the others.

Aryl Hydrocarbon Hydroxylases↗
PubMed1997

Specific inhibition of human CYP1A2 using a targeted antibody.

The structural similarity of related forms of P450 makes selective immunoinhibition of individual forms notoriously difficult to achieve. We report the use of a targeted antibody to overcome this problem. An antibody was raised against the synthetic peptide, Ser-Lys-Lys-Gly-Pro-Arg-Ala-Ser-Gly-Asn-Leu-Ile, corresponding to residues 291-302 of human CYP1A2. This sequence of human CYP1A2 is located in a similar position to a proinhibitory region previously identified in rat CYP1A1 and CYP1A2. The antibody bound strongly and specifically to CYP1A2 in human hepatic microsomal fraction. Binding was unaffected by denaturation of the protein. The specificity of the antibody was demonstrated by immunoblotting of human hepatic microsomal fraction where a single immunoreactive band was identified at Mr 54,000. The intensity of this band correlated strongly with high-affinity phenacetin O-deethylase activity of the microsomal fractions. In addition, the antibody bound to a single protein at Mr 54,000 in the microsomal fraction of lymphoblastoid cells expressing human CYP1A2, but not to any other recombinant P450 enzyme. CYP1A2-dependent activity (high-affinity phenacetin O-deethylase) of human hepatic microsomal fraction was inhibited >90% by whole antiserum or purified immunoglobulin. This decrease in activity represents complete inhibition of CYP1A2 activity, residual phenacetin O-deethylase activity being due to low-affinity enzymes. In contrast, the antibody, which does not bind to rat CYP1A2, had no effect on CYP1A2-dependent activity (high-affinity phenacetin O-deethylase) of rat hepatic microsomal fraction. The antiserum also had no effect on human hepatic microsomal debrisoquine 4-hydroxylase (CYP2D6) or coumarin 7-hydroxylase (CYP2A6) activities, indicating that inhibition was specific to human CYP1A2. These results demonstrate the importance of the region comprising residues 291-302 of human CYP1A2 in the catalytic activity of this enzyme.

Amino Acid Sequence↗
PubMed1997

Analysis of terfenadine in human plasma using microbore high-performace liquid chromatography-electrospray ionisation mass spectrometry.

An assay based on combined microbore high-performance liquid chromatography-positive ion electrospray ionisation mass spectrometry with selected ion recording has been developed for the measurement of the antihistamine drug terfenadine in human plasma. A deuterated analogue of terfenadine was synthesised for use as an internal standard and extraction of terfenadine was carried out on C18 solid phase extraction columns. The limit of detection of terfenadine in plasma is 0.1 ng/ml and the intra-assay coefficient of variation at 1 ng/ml is 10.1%. Plasma concentrations of terfenadine measured in six normal subjects following a 120 mg oral dose are reported.

Anti-Allergic Agents↗
PubMed1997

The cardiac effects of terfenadine after inhibition of its metabolism by grapefruit juice.

OBJECTIVE: To determine whether the pharmacokinetics and electrocardiographic pharmacodynamics of terfenadine are affected by the concomitant administration of grapefruit juice. METHODS: Six healthy volunteers were recruited for a balanced cross-over study. Each volunteer received 120 mg terfenadine 30 min after drinking 300 ml of either water or freshly squeezed grapefruit juice. The alternative treatment was administered on the second study day 2 weeks later. Measurements of the area under the terfenadine plasma concentration-time curve (AUC), maximum terfenadine concentration (Cmax) and the time to maximum concentration (tmax) were made, and the corrected QT (QTc) interval was measured from the surface electrocardiogram. RESULTS: Terfenadine was quantifiable in plasma in all 6 subjects on both study days for up to 24 h post-dosing. The AUC of terfenadine was significantly increased by concomitant grapefruit administration (median values 40.6 vs 16.3 ng.ml-1.h), as was the Cmax (median values 7.2 vs 2.1 ng.ml-1). The tmax was not significantly increased and there was no significant change in the median QTc interval despite the increased terfenadine levels. The 95% confidence interval for the difference in the change in QTc interval at Cmax was -13 to +38 ms. CONCLUSION: Administration of grapefruit juice concomitantly with terfenadine may lead to an increase in terfenadine bioavailability, but the increase observed in this study did not lead to significant cardiotoxicity in normal subjects. However, this does not exclude the risk of cardiotoxicity in high-risk subjects given greater doses of grapefruit juice over longer periods of time.

Adult↗
PubMed1996

Auditory brainstem response screening for hearing loss in high risk neonates.

The present paper reports the findings of a 7 year study evaluating the use of the auditory brainstem response (ABR) as the basis of a hearing screening procedure in a group of newborns at increased risk of hearing impairment. A Special Care Baby Unit (SCBU) population of 417 infants with diverse clinical backgrounds and treatment histories was tested for hearing impairment at birth using ABR audiometry. Some 332 passed the original screen at 30 dBnHL test level in both ears. Of the failure group, 18 did not survive and 32 had some degree of hearing impairment confirmed, nine of which were sensorineural in origin. An increased incidence of persistent middle ear disease was also noted in the failure group. A detailed operational analysis demonstrates that provided appropriate pass/fail criteria are adopted, the ABR technique offers excellent sensitivity and specificity for the detection of significant hearing loss in the test population. Furthermore, the study establishes that implementation of an ABR-based screening programme could reduce the average age at detection of permanent hearing loss by 7 months. A cost assessment shows that the introduction of such a targetted screening procedure could be done at a reasonable outlay.

Cost-Benefit Analysis↗
PubMed1996

Dissecting the function of cytochrome P450.

1. The role that P450 plays in the metabolism of drugs and other chemicals is often pivotal in determining the duration and magnitude of their biological effects, so that it is important to identify the specific forms of the enzyme involved. 2. The similarity between different forms of P450 is such that polyclonal antibodies raised by conventional means, even against a homogeneous preparation of the enzyme will often react with several different P450s. 3. Whilst monoclonal antibodies are often more specific, their selectivity cannot be determined prior to their use. 4. The use of short sequences of amino acids (4-7 residues) predicted to occur in surface loop regions of the protein as immunogens, affords a means of targeting antibodies to specific forms of P450. 5. The terminal amino acid of a peptide, here the C-terminal residue, is very immunodominant in determining the specificity of the resultant antibody. Hence, antibodies against the C-terminal residues of P450 have a high affinity and, in most cases, will be specific. 6. Application of anti-peptide antibodies to the major forms of P450 in human liver revealed the anticipated interindividual variation. However, evidence was found that the polymorphism in CYP3A5 expression is quantitative rather than qualitative. 7. A putative pro-inhibitory region on the surface of mammalian P450, possibly involved in intramolecular electron transport, has been identified, using a combination of directed anti-peptide antibodies and sequence alignment based on secondary structure. 8. Antibodies directed against defined regions of P450 enzymes are proving invaluable in exploring the regulation, specificity and function of these key enzymes.

Amino Acid Sequence↗
PubMed1996

Perforation rates after ventilation tube insertion: does the positioning of the tube matter?

A prospective study was performed of children undergoing bilateral ventilation tube insertion. One hundred and twenty-one children aged between 9 months and 10 years 3 months were admitted for surgery for secretory otitis media (glue ear). Each child had a ventilation tube inserted anteriorly in the tympanic membrane of one ear and posteriorly in the tympanic membrane of the other. They underwent regular clinical and audiological assessment until extrusion of the ventilation tubes occurred. Perforations were noted in 2.75% of tympanic membranes (4.6% of the children). The rate with posteriorly placed ventilation tubes was higher than with the anteriorly placed ventilation tubes (3.7% compared with 1.8%) though this is not statistically significant.

Child↗
PubMed1995

Syphilitic labyrinthitis--an update.

Forty-eight patients with syphilitic labyrinthitis have been treated and followed up for periods of six months to 25 years; interpretation of the hearing results must be in the light of the natural history of this condition which is perceived to be progression of hearing loss to profound deafness. At the beginning of the series treatment was with ampicillin alone and on this treatment 11 ears with useful hearing were maintained at this level. The group with combined ampicillin and prednisone treatment, which included some failures from ampicillin alone, maintained useful hearing in 28 out of 29 ears. Those treated initially with ampicillin and prednisone and later also given regular injections of ACTH, maintained hearing in 16 out of 17 ears. At the outset, there were 29 profoundly deaf ears. One of these recovered some useful hearing but two ears with useful hearing at the outset became profoundly deaf so that at the end there were 30 profoundly deaf ears. However, all patients as opposed to ears, with useful hearing at the outset were left with some hearing; no patient became profoundly deaf.

Adrenocorticotropic Hormone↗
PubMed1995

Faculty workload and collegial support related to proportion of part-time faculty composition.

Part-time faculty use has become more prevalent in higher education in response to enrollment shifts and budgetary constraints. This descriptive, exploratory study used a mailed survey to investigate whether full-time nursing faculty perceptions of workload and collegial support differ with changes in the proportion of part-time faculty in Comprehensive I baccalaureate nursing programs. Workload was measured by Dick's Workload Instrument. Collegial support was measured by the Survey of Collegial Communication, adapted by Beyer, which was based on Likert's organizational model. Schools were partitioned into three strata based on the proportion of part-time faculty employed (low, medium, and high). A 30% sample of schools were randomly selected from each stratum (10 schools from each). Within each selected school, six full-time undergraduate faculty were chosen by their respective deans to participate. The total response rate was 89.4%. The results of this study did not support assertions about part-time faculty use in the literature and existing accreditation standards. Findings indicated that there were significant differences in reported total faculty workload when varying proportions of part-time faculty are employed. Faculty in nursing programs with medium proportions of part-time faculty reported higher average total workloads per week than faculty in programs with low and high proportions of part-timers. Another finding demonstrated that full-time faculty in nursing programs with high proportions of part-time faculty spend fewer hours in direct clinical supervision of their students when compared with faculty in the other two strata. There were, however, no differences in perceived collegial support among full-time faculty participants. It was recommended that further research be conducted to investigate specific workload differences found in this study using more precise quantitative measures. Communication and collegiality between part-time and full-time faculty should be further developed and researched under more controlled conditions. Case studies of arrangements that make part-time faculty use beneficial are needed. Other variables such as leadership style, scholarly productivity, and morale and their relationship to the proportion of part-time faculty employed in the nursing program should be investigated.

Adult↗
PubMed1995

Determination of human hepatic cytochrome P4501A2 activity in vitro use of tacrine as an isoenzyme-specific probe.

Oxidative metabolism of the cognition activator tacrine (1,2,3,4-tetrahydro-9-aminoacridine) is thought to be catalyzed by cytochrome P4501A2 (CYP1A2). In this study, the use of tacrine as a specific substrate to measure CYP1A2 activity in vitro was investigated. Tacrine metabolism was assessed in 16 human liver microsomal samples. Initially, the percentage conversion of tacrine to stable metabolites (i.e. 1-, 2-, 4-, and 7-hydroxytacrine) at a single time point was correlated with levels of CYP1A2 apoprotein. Apoprotein was detected by immunoquantification using a monospecific CYP1A2 antipeptide antibody. Significant correlations were seen between CYP1A2 content and the degree of 1-hydroxylation (r = 0.81, p < 0.001), 7-hydroxylation (r = 0.70, p < 0.001), and metabolism to all stable products (r = 0.82, p < 0.001). The major metabolite formed in all livers was 1-hydroxytacrine. The conversion of tacrine to this metabolite was examined in more detail. The rate of formation varied from 19.2 pmol min-1 mg-1 to 101.0 pmol min-1 mg-1. There was a significant correlation (r = 0.84, p < 0.001) between the rate of formation and CYP1A2 levels. Tacrine metabolism was also compared with the rate of formation of 3-methylxanthine, from theophylline, a reaction previously shown to be catalyzed by CYP1A2. Significant correlations were found between 3-methylxanthine formation and all quantified tacrine metabolites. The rate of 3-methylxanthine generation also correlated with CYP1A2 apoprotein levels. It is concluded, therefore, that tacrine is a valuable probe for the determination of human hepatic CYP1A2 activity in vitro.

Amino Acid Sequence↗
PubMed1994

In vivo detection of myelin phospholipids in multiple sclerosis with phosphorus magnetic resonance spectroscopic imaging.

The goal of this study was to investigate myelin phospholipids in vivo in multiple sclerosis lesions and normal-appearing white matter by evaluating the spectral broad component from phosphorus 31 magnetic resonance spectroscopic imaging data. The phospholipid broad component was reduced nearly 35% (p < 0.001) in both lesions and in normal-appearing white matter in multiple sclerosis subjects compared to control subjects, suggesting reduced myelin phospholipid concentration or altered relaxation times.

Adult↗