# Identification and quantification of drug-protein adducts by mass spectrometry

> **NIH NIH R01** · UNIVERSITY OF WASHINGTON · 2024 · $80,362

## Abstract

Unanticipated adverse drug reactions remain a major cause of post marketing withdrawals of drugs and of
restricted access to new medications. Adverse drug reactions are often caused by reactive metabolites that form
covalent adducts with proteins, but the identity and extent of protein adducts formed is often difficult to predict
and characterize. Despite decades of research, current methods are unable to produce a comprehensive and
quantitative catalog of xenobiotic protein adducts. This hinders progress in optimizing safety of novel medications
and limits our ability to define mechanisms of clinically observed adverse drug reactions. To bridge this gap, we
have developed innovative proteomic methods for discovery, characterization and quantification of protein
adducts in simple and complex biological matrices. The goal of this proposal is to establish these methods for
rapid, reliable and quantitative identification and characterization of drug-protein adducts, and uniquely
customize these methods for human adductomics research impacting drug safety assessment. We will focus on
covalent protein modifications resulting from metabolic oxidative activation of drugs. We will use a set of model
compounds that are known to cause adverse events in patients and form reactive metabolites that likely result
in protein adducts. In our aim 1 we will test the hypothesis that the modification masses and chemical
characteristics of protein adducts formed by reactive metabolites in recombinant enzyme systems predict adduct
formation in more complex systems such as liver microsomes and S9 fractions. Through this work we will
optimize our proteomics methods for complex human liver preparations from individual donors. In aim 2 we will
test the hypothesis that adduct formation varies quantitatively between individuals and due to differences in
metabolic activity and individual genotype. In this aim we will establish quantitative adductomics for individual
donors and define the basis for inter-individual variability in drug-protein adduct formation. In aim 3 we will test
the hypothesis that human hepatocytes exposed to reactive metabolites generated in situ secrete proteins that
have been adducted by the reactive intermediates. In this aim we will establish the in vitro relationship between
hepatocyte adductomic burden and secretion of adducted proteins as biomarkers. When completed, the
proposed studies will be transformative in integrating novel suite of cutting-edge tools and high-dimensional
proteomics data to characterize the deep adductomic profiles of key drugs resulting in adverse drug
reactions. The methods developed will enable the assessment and quantification of a broad range of adducts
across the human proteome. The results will generate unprecedented insight into mechanisms of enzyme
inactivation, liver adductomes formed after exposure to reactive metabolites and quantitative relationships
between adduct formation and metabolic activity in the liver. ...

## Key facts

- **NIH application ID:** 11099164
- **Project number:** 3R01GM147947-03S1
- **Recipient organization:** UNIVERSITY OF WASHINGTON
- **Principal Investigator:** Nina Isoherranen
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $80,362
- **Award type:** 3
- **Project period:** 2022-09-01 → 2026-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/11099164

## Citation

> US National Institutes of Health, RePORTER application 11099164, Identification and quantification of drug-protein adducts by mass spectrometry (3R01GM147947-03S1). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/11099164. Licensed CC0.

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