Alkylation of Human Serum Albumin by Sulfur Mustard in Vitro and in Vivo:  Mass Spectrometric Analysis of a Cysteine Adduct as a Sensitive Biomarker of Exposure

Noort, Daan; Hulst, A.G.; Jong, L.P.A. de; Benschop, H. P.

doi:10.1021/tx9900369

Cited by 130 publications

(130 citation statements)

References 26 publications

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“…After 2 h of incubation at 37°C, the mixture was passed through molecular mass cutoff filters (10 kDa) under centrifugation at 2772g to remove the enzyme. Under these conditions, pronase E digestion of albumin adducts is completed (Noort et al, 1999). The filtrate was subsequently analyzed by LC/MS/MS.…”

Section: Methodsmentioning

confidence: 99%

“…This methodology, which has been applied successfully for the biomonitoring of exposure of humans to chemical warfare agents (Noort et al, 1999(Noort et al, , 2002, is based on the digestion of albumin by pronase E and subsequent selective detection of covalent adducts to the tripeptide cysteine 34 -proline-phenylalanine (CPF) by LC/MS/MS. Cysteine 34 is the only free thiol group in HSA and is capable of reacting with electrophiles.…”

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confidence: 99%

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Liquid Chromatography/Tandem Mass Spectrometry Detection of Covalent Binding of Acetaminophen to Human Serum Albumin

Damsten¹,

Commandeur²,

Fidder³

et al. 2007

Drug Metab Dispos

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ABSTRACT:Covalent binding of reactive electrophilic intermediates to proteins is considered to play an important role in the processes leading to adverse drug reactions and idiosyncratic drug reactions. Consequently, both for the discovery and the development of new drugs, there is a great interest in sensitive methodologies that enable the detection of covalent binding of drugs and drug candidates in vivo. In this work, we present a strategy for the generation and analysis of drug adducts to human serum albumin. Our methodology is based on the isolation of albumin from blood, its digestion to peptides by pronase E, and the sensitive detection of adducts to the characteristic cysteine-proline-phenylalanine (CPF) tripeptide by liquid chromatography/tandem mass spectrometry. We chose acetaminophen (APAP) as a model compound because this drug is known to induce covalent binding to proteins when bioactivated by cytochromes P450 to its reactive N-acetyl-p-benzoquinoneimine metabolite. First, by microsomal incubations of APAP in presence of CPF and/or intact albumin, in vitro reference adducts were generated to determine the mass spectrometric characteristics of the expected CPF adducts and to confirm their formation on pronase E digestion of the alkylated protein. When applying this methodology to albumin isolated from blood of patients exposed to APAP, we were indeed able to detect the corresponding CPF adducts. Therefore, this strategy could be seen as a potential biomonitoring tool to detect in vivo reactive intermediates of drugs and drug candidates, e.g., in the preclinical and clinical development phase.

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Liquid Chromatography/Tandem Mass Spectrometry Detection of Covalent Binding of Acetaminophen to Human Serum Albumin

Damsten¹,

Commandeur²,

Fidder³

et al. 2007

Drug Metab Dispos

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“…30,31) Mustard gas has been shown to alkylate a cysteine residue in human serum albumin, and the alkylation site was identified in a tryptic digest of mustard-gas-exposed albumin. 32) From an in vitro experiment using [ 14 C]-labeled mustard gas and cultured human cells, 15-20% of radioactivity was combined with keratin. 33) A time course study of the mustard gas adduct showed that the albumin adduct (S-2-hydroxyethylthioethyl)-Cys-ProTyr was detected until 7 days after exposure, and the N-terminal valine adduct in hemoglobin was detected after 28 days.…”

Section: Analysis Of Mustard Gas and Nitrogen Mustard Adductsmentioning

confidence: 99%

Mass Spectrometric Identification of Chemical Warfare Agent Adducts with Biological Macromolecule for Verification of Their Exposure

Tsuge

Seto

2009

JOURNAL OF HEALTH SCIENCE

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Exposure to chemical warfare agents (CWAs) can be verified by detecting their degradation products in biological samples. However, the half-lives of these CWA degradation products in the body are short. CWAs are known to combine with biological macromolecules and the covalently bound compounds are called adducts. The residence time of adducts in the human body is longer than that of the corresponding CWA degradation products. Therefore, development of analytical methods to detect these adducts is a more realistic way of verifying CWA exposure. In this mini review, we describe the present state of research on analytical methods for identifying CWA adducts, and mainly introduce our research on nerve gas adducts using a direct method. Novel analytical methods for verifying low level exposure to nerve gases use liquid chromatography-mass spectrometry (LC-MS). Butyrylcholinesterase (BuChE) has been purified by affinity chromatography and sodium dodecylsulfate poly-acrylamide gel electrophoresis. The purified enzyme is inhibited by nerve gases (sarin, VX and soman) and digested by chymotrypsin, and then the peptides are analyzed by LC-MS and LC tandem MS. The peptide fragment that is combined with nerve gas is detected by LC-MS. Furthermore, the chemical structure of the adduct peptide characterized by MS provides structural information about the analyte. The detection limit of BuChE or BuChE adduct is estimated at 4 ng/injection (53 fmol/injection). If at least 1% of BuChE activity is inhibited by nerve gas, the adduct can be detected at a level of 1% BuChE inhibition in the victim's serum.

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“…Methodologies for retrospective detection of exposure to CWs are available 29 . However, these advanced techniques were not available to the team to use for verification.…”

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confidence: 99%

Long-term health effects of chemical warfare agents on children following a single heavy exposure

Talabani

Ali²,

Kadir

et al. 2017

Hum Exp Toxicol

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In the 1980s, villages in the Kurdistan region of Iraq were exposed to chemical weapons (CWs), which killed and injured thousands of civilians. There has been no clinical assessment of the long-term effects of CWs exposure on those injured. We report the first such evaluation of CW effects on long-term health of children. Patients from the CW-exposed areas were interviewed to assess previous and current clinical history and underwent clinical examination. The status of organs known to be targets of CWs, including skin, eyes, respiratory and neuromuscular systems, was assessed. Children of similar age and social background, but with no history of CW exposure, were selected as a control population. Results showed that 70% of children in the CWs group had chronic health problems in contrast to 3.3% in the unexposed group (p < 0.0001). Fifty-five per cent of the CWexposed group had long-term visual impairment but none in the unexposed population. Thirty-six per cent of the CW-exposed group had chronic dermatological conditions compared with 0.8% of the unexposed group (p < 0.0001), 31% of the CWs group had neurological sequelae compared with 0.4% of the unexposed group (p < 0.0001) and 51% of the CWs group had long-term respiratory problems compared with 1.5% of the unexposed group (p < 0.0001). Respiratory complaints including asthma, chronic bronchitis and bronchiectasis were particularly common. Our study suggests that CWs used were probably a combination of sulphur mustard and organophosphate nerve agents. Results also indicate that the prevalence of acute and chronic health problems following exposure to CW agents appear to be higher in children compared with reported data in adults.

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Alkylation of Human Serum Albumin by Sulfur Mustard in Vitro and in Vivo: Mass Spectrometric Analysis of a Cysteine Adduct as a Sensitive Biomarker of Exposure

Cited by 130 publications

References 26 publications

Liquid Chromatography/Tandem Mass Spectrometry Detection of Covalent Binding of Acetaminophen to Human Serum Albumin

Liquid Chromatography/Tandem Mass Spectrometry Detection of Covalent Binding of Acetaminophen to Human Serum Albumin

Mass Spectrometric Identification of Chemical Warfare Agent Adducts with Biological Macromolecule for Verification of Their Exposure

Long-term health effects of chemical warfare agents on children following a single heavy exposure

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