title: Mass spectrometric characterization of circulating covalent protein adducts derived from a drug acyl glucuronide metabolite: multiple albumin adductions in diclofenac patients.
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.
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Thomas G
authors.initials: TG
authors.lastname: Hammond
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Meng
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Rosalind E
authors.initials: RE
authors.lastname: Jenkins
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: James L
authors.initials: JL
authors.lastname: Maggs
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Anahi Santoyo
authors.initials: AS
authors.lastname: Castelazo
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Sophie L
authors.initials: SL
authors.lastname: Regan
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Stuart N L
authors.initials: SN
authors.lastname: Bennett
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Caroline J
authors.initials: CJ
authors.lastname: Earnshaw
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Guruprasad P
authors.initials: GP
authors.lastname: Aithal
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Ira
authors.initials: I
authors.lastname: Pande
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: J Gerry
authors.initials: JG
authors.lastname: Kenna
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: Andrew V
authors.initials: AV
authors.lastname: Stachulski
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom.
authors.forename: B Kevin
authors.initials: BK
authors.lastname: Park
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Institute of Translational Medicine, (T.G.H., X.M., R.E.J., J.L.M., A.S.C., S.L.R., C.J.E., B.K.P., D.P.W.) and Department of Chemistry (A.V.S.), University of Liverpool, Liverpool, United Kingdom; Nottingham Digestive Diseases Centre, NIHR Nottingham Digestive Diseases-Biomedical Research Unit, Nottingham University Hospitals NHS Trust and University of Nottingham (G.P.A.) and Department of Rheumatology, Nottingham University Hospitals NHS Trust (I.P.), Nottingham, United Kingdom; and AstraZeneca U.K. Ltd (S.N.L.B.) and Safety Assessment, AstraZeneca U.K. Ltd (J.G.K.), Alderley Park, Macclesfield, Cheshire, United Kingdom dominic.williams@astrazeneca.com.
authors.forename: Dominic P
authors.initials: DP
authors.lastname: Williams
mesh_terms: D000328:Adult; D000368:Aged; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000378:metabolism*; D004008:Diclofenac / Q000378:metabolism*; D005260:Female; D020719:Glucuronides / Q000378:metabolism*; D006801:Humans; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D008875:Middle Aged; D011485:Protein Binding; D012709:Serum Albumin / Q000378:metabolism*
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D020719:Glucuronides; D012709:Serum Albumin; D004008:Diclofenac