Abstract
Qualitative trapping profile of reactive metabolites arising from six structurally different compounds was tested with three different d-peptide isomers (Peptide 1, gly–tyr–pro–cys–pro–his-pro; Peptide 2, gly–tyr–pro–ala–pro–his–pro; Peptide 3, gly–tyr–arg–pro–cys–pro–his–lys–pro) and glutathione (GSH) using mouse and human liver microsomes as the biocatalyst. The test compounds were classified either as clinically “safe” (amlodipine, caffeine, ibuprofen), or clinically as “risky” (clozapine, nimesulide, ticlopidine; i.e., associated with severe clinical toxicity outcomes). Our working hypothesis was as follows: could the use of short different amino acid sequence containing d-peptides in adduct detection confer any add-on value to that obtained with GSH? All “risky” agents’ resulted in the formation of several GSH adducts in the incubation mixture and with at least one peptide adduct with both microsomal preparations. Amlodipine did not form any adducts with any of the trapping agents. No GSH and peptide 2 and 3 adducts were found with caffeine, but with peptide 1 one adduct with human liver microsomes was detected. Ibuprofen produced one Peptide 1-adduct with human and mouse liver microsomes but not with GSH. In conclusion, GSH still remains the gold trapping standard for reactive metabolites. However, targeted d-peptides could provide additional information about protein binding potential of electrophilic agents, but their clinical significance needs to be clarified using a wider spectrum of chemicals together with other safety estimates.
Highlights
The metabolic activation of a drug to an electrophilic reactive metabolite and its covalent binding to cellular macromolecules, such as proteins or nucleic acids, is considered to be one of many ways that drugs exert their toxicity
We tested the qualitative trapping properties of three synthetic d-peptides with different sequences and compared their binding properties to that of GSH using mouse and human liver microsomes as catalyzing enzyme source
The adducts formed were analyzed by liquid chromatography–mass spectrometry (LC/MS) ion-trap mass-spectrometry with an ESI source
Summary
The metabolic activation of a drug to an electrophilic reactive metabolite and its covalent binding to cellular macromolecules, such as proteins or nucleic acids, is considered to be one of many ways that drugs exert their toxicity. The adducts formed can cause either acute or long-term toxicity, e.g., degenerative diseases [5,25,54]. It is difficult to monitor the formation of electrophilic reactive metabolites because they are unstable. Usually the amounts of these compounds produced are very small compared to major metabolites or the parent molecule. Different types of trapping agents are needed to measure the formation of electrophilic compounds. Glutathione (GSH) is the most commonly used trapping agent [3,37,38] because it forms conjugates with many different kinds of electrophiles. A bromine-containing glutathione analog [23] has been claimed to increase sensitivity compared to unmodified GSH
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
