Population pharmacokinetic and pharmacogenetic analysis of 6-mercaptopurine in paediatric patients with acute lymphoblastic leukaemia

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT center dot The cytotoxic effects of 6-mercaptopurine (6-MP) were found to be due to drug-derived intracellular metabolites (mainly 6-thioguanine nucleotides and to some extent 6-methylmercaptopurine nucleotides) rather than the drug itself. center dot Current empirical dosing methods for oral 6-MP result in highly variable drug and metabolite concentrations and hence variability in treatment outcome. WHAT THIS STUDY ADDS center dot The first population pharmacokinetic model has been developed for 6-MP active metabolites in paediatric patients with acute lymphoblastic leukaemia and the potential demographic and genetically controlled factors that could lead to interpatient pharmacokinetic variability among this population have been assessed. center dot The model shows a large reduction in interindividual variability of pharmacokinetic parameters when body surface area and thiopurine methyltransferase polymorphism are incorporated into the model as covariates. center dot The developed model offers a more rational dosing approach for 6-MP than the traditional empirical method (based on body surface area) through combining it with pharmacogenetically guided dosing based on thiopurine methyltransferase genotype. To investigate the population pharmacokinetics of 6-mercaptopurine (6-MP) active metabolites in paediatric patients with acute lymphoblastic leukaemia (ALL) and examine the effects of various genetic polymorphisms on the disposition of these metabolites. Data were collected prospectively from 19 paediatric patients with ALL (n = 75 samples, 150 concentrations) who received 6-MP maintenance chemotherapy (titrated to a target dose of 75 mg m(-2) day(-1)). All patients were genotyped for polymorphisms in three enzymes involved in 6-MP metabolism. Population pharmacokinetic analysis was performed with the nonlinear mixed effects modelling program (nonmem) to determine the population mean parameter estimate of clearance for the active metabolites. The developed model revealed considerable interindividual variability (IIV) in the clearance of 6-MP active metabolites [6-thioguanine nucleotides (6-TGNs) and 6-methylmercaptopurine nucleotides (6-mMPNs)]. Body surface area explained a significant part of 6-TGNs clearance IIV when incorporated in the model (IIV reduced from 69.9 to 29.3%). The most influential covariate examined, however, was thiopurine methyltransferase (TPMT) genotype, which resulted in the greatest reduction in the model's objective function (P

Response of vulval lichen sclerosus and squamous hyperplasia to photodynamic treatment using sustained topical delivery of aminolevulinic acid from a novel bioadhesive patch system

This study evaluated the clinical and histopathological responses of vulval lichen sclerosus (LS) and squamous hyperplasia (SH) to photodynamic therapy (PDT). A novel bioadhesive patch containing aminolevulinic acid (ALA) at a dose of (38 mg/cm(2)) was used to treat 10 patients before irradiation with light of 630 nm. Clinical, histopathological and pathological responses to treatment were assessed at 6 weeks post-treatment. After 17 cycles of PDT, six patients reported significant symptomatic relief and no cutaneous photosensitivity. Histopathological differences were not demonstrated, but statistically significant induction of apoptosis was seen. It can be concluded that ALA-PDT patch-based formulation is pragmatic and primarily offers symptomatic management of vulval LS and SH.

The role of previous experience and attitudes toward statistics in statistics assessment outcomes among undergraduate psychology students

Previous research has demonstrated that students’ cognitions about statistics are related to their performance in statistics assessments. The purpose of this research is to examine the nature of the relationships between undergraduate psychology students’ previous experiences of maths, statistics and computing; their attitudes toward statistics; and assessment on a statistics course. Of the variables examined, the strongest predictor of assessment outcome was students’ attitude about their intellectual knowledge and skills in relation to statistics at the end of the statistics curriculum. This attitude was related to students’ perceptions of their maths ability at the beginning of the statistics curriculum. Interventions could be designed to change such attitudes with the aim of improving students’ learning of statistics.

From stress to distress: Conceptualizing the British family farming patriarchal way of life

‘Rural stress’ and ‘farming stress’ are terms that have become commonly appropriated by British health-based academic disciplines, the medical profession and social support networks, especially since the agricultural ‘crises’ of B.S.E. and Foot and Mouth Disease. Looking beyond the media headlines, it is apparent that the terms in fact are colloquial catch-alls for visible psychological and physiological outcomes shown by individuals. Seldom have the underlying causes and origins of presentable medical outcomes been probed, particularly within the context of the patriarchal and traditionally patrilineal way of life which family forms of farming business activity in Britain encapsulate. Thus, this paper argues that insufficient attention has been paid to the conceptualization of the terms. They have become both over-used and ill-defined in their application to British family farm individuals and their life situations. A conceptual framework is outlined that attempts to shift the stress research agenda into the unilluminated spaces of the family farming ‘way of life’ and focus instead on ‘distress’. Drawing upon theorization from agricultural and feminist geography together with cultural approaches from rural geography, four distinct clusters of distress originate from the thoughts of individuals and the social practices now required to enact patriarchal family farming gender identities. These are explored using case study evidence from ethnographic repeated life history interviews with members of seven farming families in Powys, Mid Wales, an area dominated by family forms of farming business. Future research agendas need to be based firmly on the distressing reality of patriarchal family farming and also be inclusive of those who, having rejected the associated way of life, now lie beyond the farm gate.<br/><br/>

Construction of cDNA libraries from trifluoroacetic acid-solvated amphibian skin secretions: molecular cloning of multiple bombinin-like peptide precursor transcripts from a library of yellow-bellied toad (Bombina variegata) secretion

Amphibian skin secretions are renowned as complex mixtures of bioactive peptides many of which are analogues of endogenous regulatory peptides. While skin secretions can be obtained non-invasively for peptidome analysis, parallel studies on the granular gland transcriptome required specimen sacrifice. The aim of the present study was to analyse archived skin secretions to determine the robustness of bioactive peptide precursor-encoding polyadenylated mRNAs in an attempt to extract maximum molecular information from rare samples. A range of solvated skin secretion samples were examined after lyophilisation for their potential to generate viable and comprehensive cDNA libraries based upon polyadenylated mRNA capture and amplification/cloning using appropriate commercial kits. Here we present unequivocal data that the granular gland transcriptome persists in a PCR amenable format even after storage for as long as 12 years in 0.1%(v/v) aqueous trifluoroacetic acid (TFA). We used a pooled skin secretion sample (2 ml) from the yellow-bellied toad, Bombina variegata (n = 14), containing the equivalent of 5 mg/ml of lyophilised skin secretion, that had been used in part for peptide isolation purposes in 1998 and had been stored at - 20 °C since that time. In the first cloning experiment, 12 different bombinin-like peptide precursor cDNAs were cloned encoding 17 different bombinins, the majority of which were novel. Subsequently, bombesin and bradykinin-related peptide precursor transcripts have been cloned successfully. These data illustrate the unexpected stability/longevity of the transcriptome in these secretions — a finding with implications for both this field of research and for the wider field of molecular biology.

Effect of growth-promoting 17ß estradiol, 19 nortestosterone and dexamethasone on circulating levels of nine potential biomarker candidates in veal calves

The use of screening methods based on the detection of biological effects of growth promoters is a promising approach to assist residue monitoring. To reveal useful effects on protein metabolism, male and female veal calves at 10 weeks of age were treated thrice with a combination of 25 mg 17ß-estradiol 3-benzoate and 150 mg 19-nortestosterone decanoate with 2 weeks intervals and finally once with 4 mg dexamethasone. Hormone-treated calves showed a significant accelerated growth rate over 6 weeks. Plasma samples of treated and control calves were analysed for immunoreactive inhibin (ir-inhibin), osteocalcin, insulin-like growth factor 1 (IGF-1), insulin-like growth factor-binding protein 2 (IGFBP-2), IGFBP-3, luteinzing hormone (LH), follicle-stimulating hormone (FSH) and prolactin using immunoaffinity assays. Hormone treatment did not affect levels of IGF-1, IGFBP-2, IGFBP-3, LH, FSH and prolactin. The concentration of circulating ir-inhibin decreased, however, significantly (P &lt; 0.05) in bull calves upon administration of the sex steroids, whereas it remained unchanged in the female animals. Dexamethasone treatment decreased significantly (P &lt; 0.05) circulating levels of osteocalcin in both female and male animals. Ir-inhibin and osteocalcin were, therefore, considered as candidates for a protein biomarker-based screening assay for detection of abuse of estrogens, androgens and/or glucocorticoids in cattle fattening, which is being developed in the framework of EU research project BioCop

A competitive enzyme-linked immunosorbent assay for determination of chloramphenicol

Chloramphenicol is a broad-spectrum antibiotic shown to have specific activity against a wide variety of organisms that are causative agents of several disease conditions in domestic animals. Chloramphenicol has been banned for use in food-producing animals for its serious adverse toxic effects in humans. Due to the harmful effects of chloramphenicol residues livestock products should be free of any traces of these residues. Several analytical methods are available for chloramphenicol analysis but sensitive methods are required in order to ensure that no traces of chloramphenicol residues are present in edible animal products. In order to prevent the illegal use of chloramphenicol, regulatory control of its residues in food of animal origin is essential. A competitive enzyme-linked immunosorbent assay for chloramphenicol has been locally developed and optimized for the detection of chloramphenicol in sheep serum. In the assay, chloramphenicol in the test samples and that in chloramphenicol-horseradish peroxidase conjugate compete for antibodies raised against the drug in camels and immobilized on a microtitre plate. Tetramethylbenzidine-hydrogen peroxide (TMB/H2O2) is used as chromogen-substrate system. The assay has a detection limit of 0.1 ng/mL of serum with a high specificity for chloramphenicol. Cross-reactivity with florfenicol, thiamphenicol, penicillin, tetracyclines and sulfamethazine was not observed. The assay was able to detect chloramphenicol concentrations in normal sheep serum for at least 1 week after intramuscular injection with the drug at a dose of 25 mg/kg body weight (b.w.). The assay can be used as a screening tool for chloramphenicol use in animals.

Development and validation of a fast monoclonal based disequilibrium enzyme-linked immunosorbent assay for the detection of triphenylmethane dyes and their metabolites in fish

Malachite Green (MG), Crystal Violet (CV) and Brilliant Green (BC) are antibacterial, antifungal and antiparasitic agents that have been used for treatment and prevention of diseases in fish. These dyes are metabolized into reduced leuco forms (LMG, LCV, LBG) that can be present in fish muscles for a long period. Due to the carcinogenic properties they are banned for use in fish for human consumption in many countries including the European Union and the United States. HPLC and LC-MS techniques are generally used for the detection of these compounds and their metabolites in fish. This study presents the development of a fast enzyme-linked immunosorbent assay (ELISA) method as an alternative for screening purposes. A first monoclonal cell line producing antibodies to MG was generated using a hybridoma technique. The antibody had good cross-reactivates with related chromatic forms of triphenylmethane dyes such as CV, BC, Methyl Green, Methyl Violet and Victoria Blue R. The monoclonal antibody (mAb) was used to develop a fast (20 min) disequilibrium ELISA screening method for the detection of triphenylmethanes in fish. By introducing an oxidation step with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) during sample extraction the assay was also used to detect the presence of the reduced metabolites of triphenylmethanes. The detection capability of the assay was 1 ng g(-1) for MG, LMG, CV, LCV and BC which was below the minimum required performance limit (MRPL) for the detection method of total MG (sum of MG and LMG) set by the Commission Decision 2004/25/EC (2 ng g(-1)). The mean recoveries for fish samples spiked at 0.5 MRPL and MRPL levels with MG and LMG were between 74.9 and 117.0% and inter- and intra-assay coefficients of variation between 4.7 and 25.7%. The validated method allows the analysis of a batch of 20 samples in two to three hours. Additionally, this procedure is substantially faster than other ELISA methods developed for MG/LMG thus far. The stable and efficient monoclonal cell line obtained is an unlimited source of sensitive and specific antibody to MG and other triphenylmethanes. (C) 2011 Elsevier B.V. All rights reserved.

Fasciola hepatica: time-dependent disruption of spermatogenesis following in vivo treatment with triclabendazole

Sheep infected with the Cullompton isolate of Fasciola hepatica were treated with triclabendazole at a concentration of 10 mg/kg at 12 weeks post-infection. Adult flukes were recovered from the liver and, where present, from the gall bladder at 48, 72 and 96 h post-treatment (pt). Gross changes to the spermatogenic cells of the testis were examined by histology and ultrastructural alterations were visualised via transmission electron microscopy. Disruption was progressive in nature, with the testis tubules becoming shrunken, vacuolated and gradually more denuded of cellular content over the 96-h time period. From 48 h pt, the number of primary and secondary spermatogonia decreased and multinucleate spermatogonial cells were frequent. Later, developmental stages were uncommon, giving rise to much empty space within the tubules. By 72 h pt, the tubules contained many apoptotic and degraded cells and had an extremely disorganised appearance. At 96 h pt, the tubules were almost completely empty, with the exception of the remains of degraded spermatogenic cells. These results indicate that triclabendazole severely disrupts spermatogenesis in the liver fluke from 48 h pt in vivo.

Enhancement of triclabendazole action in vivo against a triclabendazole-resistant isolate of Fasciola hepatica by co-treatment with ketoconazole

An in vivo study in the laboratory rat model was carried out to monitor morphological changes in adult Fasciola hepatica over a 4-day period resulting from combination treatment of triclabendazole (TCBZ) and the metabolic inhibitor, ketoconazole (KTZ). Rats were infected with the TCBZ-resistant Oberon isolate of F. hepatica and divided into 3 groups at 12 weeks post-infection. The first group was dosed orally with TCBZ at a dosage of 10 mg/kg and KTZ at a dosage of 10 mg/kg. Flukes were recovered at 24, 48, 72 and 96h post-treatment (p.t.). A second group of rats was treated with TCBZ alone (10 mg/kg) and sacrificed at 96 h p.t. The third group acted as untreated controls. Surface changes were monitored by scanning electron microscopy (SEM). In flukes from the TCBZ+ KTZ-treated group, the results showed a progressive and time-dependent increase in the level of disruption to the tegumental syncytium. Swelling, furrowing, blebbing and sloughing of the syncytium increased with time p.t. Another feature seen was a thick layer of tegumental shedding in some fluke samples at different times p.t. By comparison, flukes treated with TCBZ alone remained unaffected. The results demonstrated that the Oberon isolate is only sensitive to drug action in the presence of ketoconazole, indicating that combining triclabendazole with a metabolic inhibitor could be used to preserve the effectiveness of the drug against TCBZ-resistant populations of F. hepatica. (C) 2010 Elsevier B.V. All rights reserved.

Reproductive disruption in Fasciola hepatica associated with incomplete efficacy of a new experimental formulation of triclabendazole

The objective of the present study was to analyse the reproductive viability (using histopathologic studies) of Fasciola hepatica from cattle artificially infected and treated subcutaneously with a new experimental formulation of triclabendazole (8 mg/kg b.w.). The results of the efficacy controlled test, which only takes into account the presence of live adult flukes, indicated that, whilst in the control group (n=7)533 live specimens were recovered, in the test groups (doses of 8 and 12 mg/kg b.w.) only 195 and 47 adults were recovered, respectively. These numbers indicate efficacies of 69% and 95.6%, respectively. It was observed in that dose of 8 mg/kg b.w. some specimens remained viable, but they were infertile, which severely compromises the biological cycle of the trematode. in the testis tubules of flukes treated with the low dose of TCBZ (8 mg/kg), very few cells were present and the vitelline follicles were markedly reduced in size and each follicle contained very few cells. This would have direct implications for the pathogenesis of the parasitosis since the remaining parasites would produce little clinical-productive manifestations, would stimulate the immune response and would find it difficult to establish future re-infestations/re-infections. Consequently, these observations will also prompt a review of certain methodological and interpretative aspects related to efficacy tests, where the only discriminative factor is the reduction of the adult parasite load. On one hand, histopathological studies could be complementary to the efficacy controlled test for TCBZ or other BZD formulations. (C) 2010 Elsevier B.V. All rights reserved.

Detection of monensin residues in poultry liver using an enzyme immunoassay

Monensin, a carboxylic acid ionophore, is commonly fed to poultry to control coccidiosis. A method for the detection and quantification of monensin residues in liver has been developed, Samples (3 g) were extracted with acetonitrile-water and applied to a competitive enzyme immunoassay using a polyclonal antiserum raised against a monensin-transferrin conjugate, The limit of detection (mean + 3s) calculated from the analysis of 12 known negative samples was 2.91 ng g(-1). Intra- and inter-assay RSD were determined as 8.5 and 10.6%, respectively, using a liver sample fortified with 20 ng g(-1) monensin, A pharmacokinetic study in which 70 six week old broilers were fed monensin at a rate of 120 mg kg(-1) in their feed for 14 d resulted in mean monensin liver residues of 102 ng g(-1). However these had fallen below the limit of detection of the assay within the 3 d withdrawal period recommended by the manufacturer.

Early onset of changes to the reproductive system of Fasciola hepatica following in vivo treatment with triclabendazole

Lambs infected with the Cullompton triclabendazole (ICBZ)-susceptible isolate of Fasciola hepatica were treated with TCBZ at a dosage of 10 mg/kg at 16 weeks post-infection. Adult flukes were recovered from the liver at 3 h, 24 h, 48 h and 60 h post-treatment (pt). They were processed for histological analysis of the uterus, Mehlis' gland, vitellaria, ovary and testis. At 3 h pt, the flukes were essentially similar to the controls and were producing normal eggs. Egg production had ceased by 24 h pt. At this time period, the cells of the Mehlis' gland showed some evidence of vacuolation, but otherwise were relatively normal. A shift in the population of vitelline cells towards mature cells was observed at 24 h pt, and this trend continued at later time-periods. It was accompanied by a breakdown of the cells and the presence of apoptotic bodies. Marked changes to the ovary were first noted at 48 h Pt, as evidenced by vacuolation and the presence of apoptotic bodies. Some disruption to the testis was seen at 24 h pt, with a reduction in the population of spermatogenic cells, the appearance of apoptotic bodies and some peripheral vacuolation of the tubules. These abnormalities increased in severity with longer time periods pt. The results bring forward the time-line of cessation of egg production by 24 h, demonstrating that this process is affected very rapidly pt. (C) 2011 Elsevier B.V. All rights reserved.

Measurements of energetic proton transport through magnetized plasma from intense laser interactions with solids

Protons with energies up to 18 MeV have been measured from high density laser-plasma interactions at incident laser intensities of 5 X 10(19) W/cm(2). Up to 10(12) protons with energies greater than 2 MeV were observed to propagate through a 125 mu m thick aluminum target and measurements of their angular deflection were made. It is likely that the protons originate from the front surface of the target and are bent by large magnetic fields which exist in the target interior. To agree with our measurements these fields would be in excess of 30 MG and would be generated by the beam of fast electrons which is also observed.

The Emergence of Rural Support Organisations in the UK and Canada: Providing Support for Patrilineal Family Farming

A certain type of rural support has emerged since agricultural restructuring of the 1980s. The article draws on research from the UK and Canada to suggest that support in both countries is derived from a patrilineal culture that still dominates family farming in both countries. Such a way of life, it is argued, involves the majority of farming men and women across generations working increasingly hard to ensure farm survival in order to facilitate an overall pattern of farm succession via the male or ‘patrilineal’ line. The article begins by providing a conceptualisation of patrilineal family farming drawing on insights from gender-informed work on farming identities, political-economy approaches from agricultural geography and the cultural turn in rural studies. This section will provide theoretical direction for discussion of the research findings. Here the article presents a discussion of the context to and typologies of organisations that emerged and five key findings derived from research conducted with members of the organisations in the UK and Canada. This assists in developing the argument that the emergent organisations are responding to and supporting this way of life and highlights some of the potential implications of doing so. The article has two aims. Firstly, it suggests that family farming in the UK and Canada continues to be predominantly structured by a way of life transmitted across generations which has the overall prerequisite of maintaining farm survival to enable patrilineal succession. Secondly, it suggests that a particular type of support for farming families emerged as a response to perceived threats to this way of life and provides evidence of its enduring nature.