Refinement of a Methodology for Untargeted Detection of Serum Albumin Adducts in Human Populations.

George W Preston,Lilian Bastian,Marc Chadeau-Hyam,Karin Van Veldhoven,Osman Sozeri,Tim S Nawrot,David H Phillips,Michelle Plusquin

doi:10.1021/acs.chemrestox.7b00186

Abstract

Covalently modified blood proteins (e.g., serum albumin adducts) are increasingly being viewed as potential biomarkers via which the environmental causes of human diseases may be understood. The notion that some (perhaps many) modifications have yet to be discovered has led to the development of untargeted adductomics methods, which attempt to capture entire populations of adducts. One such method is fixed-step selected reaction monitoring (FS-SRM), which analyses distributions of serum albumin adducts via shifts in the mass of a tryptic peptide [Li et al. (2011) Mol. Cell. Proteomics 10, M110.004606]. Working on the basis that FS-SRM might be able to detect biological variation due to environmental factors, we aimed to scale the methodology for use in an epidemiological setting. Development of sample preparation methods led to a batch workflow with increased throughput and provision for quality control. Challenges posed by technical and biological variation were addressed in the processing and interpretation of the data. A pilot study of 20 smokers and 20 never-smokers provided evidence of an effect of smoking on levels of putative serum albumin adducts. Differences between smokers and never-smokers were most apparent in putative adducts with net gains in mass between 105 and 114 Da (relative to unmodified albumin). The findings suggest that our implementation of FS-SRM could be useful for studying other environmental factors with relevance to human health.

Highlights

Non-enzymatic covalent modifications to macromolecules represent a source of potential biomarkers with which to study human health and disease.[1−4] In some cases, modifications might be causally related to biological end points.[5]
When we attempted enrichment of a model adduct spiked into commercial serum, only a modest proportion of the albumin was removed
The fixed-step selected reaction monitoring (FS-SRM) methodology for analyzing distributions of HSA adducts was adapted for increased throughput

Summary

Introduction

Non-enzymatic covalent modifications to macromolecules represent a source of potential biomarkers with which to study human health and disease.[1−4] In some cases, modifications might be causally related to biological end points (e.g., a mutagenic lesion in DNA causing cancer).[5] Alternatively, modifications might be found in “off-pathway” products from which “on-pathway” events may be inferred This latter possibility has motivated researchers to investigate adducts of blood proteins as potential biomarkers, in studies seeking to understand the effects of environmental factors on human populations (biomonitoring, exposome studies). In which the origin of the reactive chemical is endogenous rather than exogenous, may occur.[9]

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Results

Conclusion