8 results on '"Earnshaw CJ"'
Search Results
2. Amoxicillin and Clavulanate Form Chemically and Immunologically Distinct Multiple Haptenic Structures in Patients.
- Author
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Meng X, Earnshaw CJ, Tailor A, Jenkins RE, Waddington JC, Whitaker P, French NS, Naisbitt DJ, and Park BK
- Subjects
- Amoxicillin-Potassium Clavulanate Combination pharmacology, Haptens pharmacology, Humans, Mass Spectrometry, Models, Molecular, Molecular Conformation, Serum Albumin chemistry, Serum Albumin isolation & purification, T-Lymphocytes drug effects, T-Lymphocytes immunology, Amoxicillin-Potassium Clavulanate Combination chemistry, Amoxicillin-Potassium Clavulanate Combination immunology, Haptens chemistry, Haptens immunology
- Abstract
Amoxicillin-clavulanate (AC) is one of the most common causes of drug induced liver injury (DILI). The association between AC-DILI and HLA alleles and the detection of drug-specific T cells in patients with AC-DILI indicate that the adaptive immune system is involved in the disease pathogenesis. In this study, mass spectrometric methods were employed to characterize the antigen formed by AC in exposed patients and the antigenic determinants that stimulate T cells. Amoxicillin formed penicilloyl adducts with lysine residues on human serum albumin (HSA) in vitro, with K190 and K199 being the most reactive sites. Amoxicillin-modified K190 and K199 have also been detected in all patients, and more extensive modification was observed in patients exposed to higher doses of amoxicillin. In contrast, the binding of clavulanic acid to HSA was more complicated. Multiple adducts were identified at high concentrations in vitro, including those formed by direct binding of clavulanic acid to lysine residues, novel pyrazine adducts derived from binding to the degradation products of clavulanic acid, and a cross-linking adduct. Stable adducts derived from formylacetic acid were detected in all patients exposed to the drug. Importantly, analysis of hapten-protein adducts formed in the cell culture medium revealed that the highly drug-specific T-cell responses were likely driven by the markedly different haptenic structures formed by these two drugs. In this study, the unique haptenic structures on albumin in patients formed by amoxicillin and clavulanic acid have been characterized and shown to function as chemically distinct antigens which can stimulate separate, specific T-cell clones.
- Published
- 2016
- Full Text
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3. Mass spectrometric characterization of circulating covalent protein adducts derived from a drug acyl glucuronide metabolite: multiple albumin adductions in diclofenac patients.
- Author
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Hammond TG, Meng X, Jenkins RE, Maggs JL, Castelazo AS, Regan SL, Bennett SN, Earnshaw CJ, Aithal GP, Pande I, Kenna JG, Stachulski AV, Park BK, and Williams DP
- Subjects
- Adult, Aged, Female, Humans, Male, Middle Aged, Protein Binding, Anti-Inflammatory Agents, Non-Steroidal metabolism, Diclofenac metabolism, Glucuronides metabolism, Mass Spectrometry methods, Serum Albumin metabolism
- Abstract
Covalent protein modifications by electrophilic acyl glucuronide (AG) metabolites are hypothetical causes of hypersensitivity reactions associated with certain carboxylate drugs. The complex rearrangements and reactivities of drug AG have been defined in great detail, and protein adducts of carboxylate drugs, such as diclofenac, have been found in liver and plasma of experimental animals and humans. However, in the absence of definitive molecular characterization, and specifically, identification of signature glycation conjugates retaining the glucuronyl and carboxyl residues, it cannot be assumed any of these adducts is derived uniquely or even fractionally from AG metabolites. We have therefore undertaken targeted mass spectrometric analyses of human serum albumin (HSA) isolated from diclofenac patients to characterize drug-: derived structures and, thereby, for the first time, have deconstructed conclusively the pathways of adduct formation from a drug AG and its isomeric rearrangement products in vivo. These analyses were informed by a thorough understanding of the reactions of HSA with diclofenac AG in vitro. HSA from six patients without drug-: related hypersensitivities had either a single drug-: derived adduct or one of five combinations of 2-8 adducts from among seven diclofenac N-acylations and three AG glycations on seven of the protein's 59 lysines. Only acylations were found in every patient. We present evidence that HSA modifications by diclofenac in vivo are complicated and variable, that at least a fraction of these modifications are derived from the drug's AG metabolite, and that albumin adduction is not inevitably a causation of hypersensitivity to carboxylate drugs or a coincidental association., (Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.)
- Published
- 2014
- Full Text
- View/download PDF
4. T cell responses to drugs and drug metabolites.
- Author
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Earnshaw CJ, Pecaric-Petkovic T, Park BK, and Naisbitt DJ
- Subjects
- Animals, Drug Hypersensitivity etiology, Drug-Related Side Effects and Adverse Reactions immunology, Humans, Immunologic Techniques, Drug Hypersensitivity immunology, T-Lymphocytes immunology
- Abstract
Understanding the chemical mechanisms by which drugs and drug metabolites interact with cells of the immune system is pivotal to our knowledge of drug hypersensitivity as a whole.In this chapter, we will discuss the currently accepted mechanisms where there is scientific and clinical evidence to support the ways in which drugs and their metabolites interact with T cells. We will also discuss bioanalytical platforms, such as mass spectrometry, and in vitro test assays such as the lymphocyte transformation test that can be used to study drug hypersensitivity; the combination of such techniques can be used to relate the chemistry of drug antigen formation to immune function. Ab initio T cell priming assays are also discussed with respect to predicting the potential of a drug to cause hypersensitivity reactions in humans in relation to the chemistry of the drug and its ability to form haptens, antigens and immunogens in patients.
- Published
- 2014
- Full Text
- View/download PDF
5. Detection of drug bioactivation in vivo: mechanism of nevirapine-albumin conjugate formation in patients.
- Author
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Meng X, Howarth A, Earnshaw CJ, Jenkins RE, French NS, Back DJ, Naisbitt DJ, and Park BK
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- Adult, Aged, Anti-HIV Agents blood, Biotransformation, Cysteine metabolism, HIV-1, Histidine metabolism, Humans, Middle Aged, Nevirapine blood, Protein Binding, Anti-HIV Agents pharmacokinetics, Glutathione S-Transferase pi metabolism, HIV Infections metabolism, Nevirapine pharmacokinetics, Serum Albumin metabolism
- Abstract
The non-nucleoside reverse transcriptase inhibitor nevirapine (NVP) is widely used for the treatment of human immunodeficiency virus type 1 (HIV-1), particularly in developing countries. Despite its therapeutic benefits, NVP has been associated with skin and liver injury in exposed patients. Although the mechanism of the tissue injury is not yet clear, it has been suggested that reactive metabolites of NVP may be involved. The detection of NVP mercapturate in the urine of patients undergoing standard antiretroviral chemotherapy indicates that NVP undergoes bioactivation in vivo. However, covalent binding of drug to protein in patients remains to be determined. In this study, we investigate the chemical basis of NVP protein adduct formation by using human serum albumin (HSA) and glutathione S-transferase pi (GSTP) as model proteins in vitro. In addition, HSA was isolated from serum samples of HIV-1 patients undergoing NVP therapy to measure NVP haptenation. Mass spectrometric analysis of 12-sulfoxyl-NVP-treated HSA revealed that the drug bound selectively to histidine (His146, His242, and His338) and a cysteine residue (Cys34). The reaction proceeds most likely by a concerted elimination-addition mechanism. This pathway was further confirmed by the observation of NVP-modified Cys47 in GSTP. Importantly, the same adduct (His146) was detected in HSA isolated from the blood of patients receiving NVP, providing direct evidence that NVP modifies protein in vivo, via the formation of a reactive metabolite.
- Published
- 2013
- Full Text
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6. Human leukocyte antigen (HLA)-B*57:01-restricted activation of drug-specific T cells provides the immunological basis for flucloxacillin-induced liver injury.
- Author
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Monshi MM, Faulkner L, Gibson A, Jenkins RE, Farrell J, Earnshaw CJ, Alfirevic A, Cederbrant K, Daly AK, French N, Pirmohamed M, Park BK, and Naisbitt DJ
- Subjects
- Aged, Aged, 80 and over, CD4-Positive T-Lymphocytes immunology, CD8-Positive T-Lymphocytes immunology, Cell Movement drug effects, Chemical and Drug Induced Liver Injury immunology, Clone Cells immunology, Female, Floxacillin metabolism, HLA-B Antigens immunology, Humans, Interferon-gamma metabolism, Leukocytes, Mononuclear drug effects, Leukocytes, Mononuclear immunology, Lymphocyte Activation drug effects, Lysine metabolism, Male, Middle Aged, Receptors, CCR biosynthesis, Receptors, CCR4 biosynthesis, Serum Albumin metabolism, Chemical and Drug Induced Liver Injury etiology, Floxacillin adverse effects, HLA-B Antigens physiology, Lymphocyte Activation immunology
- Abstract
Unlabelled: The role of the adaptive immune system in adverse drug reactions that target the liver has not been defined. For flucloxacillin, a delay in the reaction onset and identification of human leukocyte antigen (HLA)-B*57:01 as a susceptibility factor are indicative of an immune pathogenesis. Thus, we characterize flucloxacillin-responsive CD4+ and CD8+ T cells from patients with liver injury and show that naive CD45RA+CD8+ T cells from volunteers expressing HLA-B*57:01 are activated with flucloxacillin when dendritic cells present the drug antigen. T-cell clones expressing CCR4 and CCR9 migrated toward CCL17 and CCL 25, and secreted interferon-gamma (IFN-γ), T helper (Th)2 cytokines, perforin, granzyme B, and FasL following drug stimulation. Flucloxacillin bound covalently to selective lysine residues on albumin in a time-dependent manner and the level of binding correlated directly with the stimulation of clones. Activation of CD8+ clones with flucloxacillin was processing-dependent and restricted by HLA-B*57:01 and the closely related HLA-B*58:01. Clones displayed additional reactivity against β-lactam antibiotics including oxacillin, cloxacillin, and dicloxacillin, but not abacavir or nitroso sulfamethoxazole., Conclusion: This work defines the immune basis for flucloxacillin-induced liver injury and links the genetic association to the iatrogenic disease., (Copyright © 2012 American Association for the Study of Liver Diseases.)
- Published
- 2013
- Full Text
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7. Direct analysis of pharmaceutical tablet formulations using Matrix-Assisted Laser Desorption/Ionisation Mass Spectrometry Imaging.
- Author
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Earnshaw CJ, Carolan VA, Richards DS, and Clench MR
- Subjects
- Chemistry, Pharmaceutical, Tablets analysis, Pharmaceutical Preparations analysis, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization methods
- Abstract
Matrix-Assisted Laser Desorption/Ionisation Mass Spectrometry Imaging (MALDI MSI) has been used to directly analyse a range of tablets in order to assess the homogeneity of the active drug compound throughout the excipients contained within the tablets studied. The information gained from the imaging experiments can be used to improve and gain a greater understanding of the manufacturing process; such knowledge will enable improvements in finished product quality to make safer and more efficacious tablet formulations. Commercially available and prescription tablet formulations have been analysed, including aspirin, paracetamol, sildenafil citrate (Viagra(R)) and a batch of tablets in development (tablet X: placebo; 1 mg; 3 mg and 6 mg). MALDI MSI provides semi-quantitative information that is related to ion abundance, therefore Principal Component Analysis (PCA), a multivariate analysis technique, has been used to differentiate between tablets containing different amounts of active drug ingredient. Aspects of sample preparation have also been investigated with regard to tablet shape and texture. The results obtained indicate that MALDI MSI can be used effectively to analyse the spatial distribution of the active pharmaceutical component (API) in pharmaceutical tablet formulations., (Copyright (c) 2010 John Wiley & Sons, Ltd.)
- Published
- 2010
- Full Text
- View/download PDF
8. Imaging Matrix Assisted Laser Desorption Ionization Mass Spectrometry: a technique to map plant metabolites within tissues at high spatial resolution.
- Author
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Burrell M, Earnshaw C, and Clench M
- Subjects
- Arginine analysis, Biological Factors analysis, Calibration, Hexosephosphates analysis, Seeds chemistry, Seeds growth & development, Sucrose analysis, Temperature, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization, Triticum chemistry, Triticum metabolism
- Abstract
Imaging Matrix Assisted Laser Desorption Ionization Mass Spectrometry provides a new and powerful tool to analyse the distribution of metabolites within plant tissues. The two matrices alpha-cyano-4-hydroxycinnamic acid (alpha-CHCA) and 9-aminoacridine provide a useful combination that allows the measurement of amino acids, sugars, and phosphorylated metabolites. Results are presented showing that representatives of these metabolites are unevenly distributed in wheat seeds at different stages of development and under temperature stress.
- Published
- 2007
- Full Text
- View/download PDF
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