The number of people seeking help for mental illness is increasing across all ages, creating a major burden for individuals, families, and the society. While personalized medicine is advancing in other fields, diagnosis and treatment of mental disorders remain largely symptom-based and fail to capture individual, sex, and gender differences in risk, manifestation, and treatment response. Early signs of illness often go unnoticed due to the lack of monitoring tools, and stigma continues to hinder prevention and care. In some phases of life, an individual’s susceptibility to mental illness is heightened and may be influenced by changes in endocrine signalling. To address these challenges, the research project Building REsilience against MEntal illness during ENDocrine-sensitive life stages (RE-MEND) has implemented an interdisciplinary approach focusing on four critical endocrine-sensitive life stages: prenatal, puberty, peripartum, and older age. The project integrates longitudinal population-based cohorts with experimental and clinical studies to identify genetic, environmental, and endocrine factors shaping susceptibility and resilience to mental illness. Multi-omics data (genomics, epigenomics, transcriptomics, proteomics, metabolomics, lipidomics, and adductomics) will be combined with neurobiological, clinical, and behavioural measures, analysed using advanced biostatistics and machine learning. RE-MEND seeks to i) identify risk and resilience factors affecting mental health; ii) deliver biomarker panels for susceptibility, disease progression, and treatment response across sensitive life stages; iii) discover novel drug targets through repurposing strategies, and iv) promote mental health literacy and reduce stigma. The integration of biological research with communication science is anticipated to result in translatable findings, supporting earlier intervention and more effective care.

The growing use of environmental DNA adductomics for exposure assessment emphasizes the need for improved methods that are adapted to chemically and biologically diverse samples. While liquid chromatography coupled to high-resolution mass spectrometry (LC–HRMS) has advanced DNA adductome analysis, sample cleanup methods remain challenging, especially in nonmammalian models. Here, we used amphipod Monoporiea affinis, a sentinel species in biological effect studies, as a surrogate crustacean species for the method development. It is particularly suitable because its chitinous tissues and high lipid content present a challenging matrix for DNA extraction. We addressed the following objectives: (i) to propose adduct structures by combining multiple tools for DNA adduct screening and (ii) to evaluate the applicability of dispersive solid-phase extraction (d-SPE) as a cleanup step for DNA adductome analysis. Toward the first objective, we integrated open-source software nLossFinder with a recently introduced DNA adductomics database (GitLab) to enhance the structural identification of unknown adducts. A combination of accurate mass data and MS2-fragmentation allowed differentiation of the complex mixture of nucleoside adducts, facilitating the structural identification of 16 DNA adducts, including 10 modifications on amphipod DNA reported for the first time. Toward the second objective, we introduced d-SPE as a novel cleanup approach for DNA adduct analysis in whole-body crustacean samples. Using Z-sep+ as the d-SPE sorbent, we demonstrated a major reduction of matrix interferences, including phospholipids, and enhanced sensitivity toward DNA adducts detection, leading to improved LC–HRMS signal response by up to 170%. d-SPE offers a simple alternative to conventional methods like standard SPE and liquid–liquid extraction, making it a valuable tool for DNA adductomics in environmental monitoring and aiding high-throughput capacity, especially while handling large numbers of samples. Further studies should include validation of this method for other species and DNA modifications. These advancements underscore the potential of the proposed data analysis workflow and d-SPE for improving mass spectrometry-based DNA adductomics in environmental monitoring, paving the way for more accurate and comprehensive exposure assessments across diverse species and environmental conditions.

Exposure to chemical pollution can induce genetic andepigeneticalterations, developmental changes, and reproductive disorders, leadingto population declines in polluted environments. These effects aretriggered by chemical modifications of DNA nucleobases (DNA adducts)and epigenetic dysregulation. However, linking DNA adducts to thepollution load in situ remains challenging, and thelack of evidence-based DNA adductome response to pollution hampersthe development and application of DNA adducts as biomarkers for environmentalhealth assessment. Here, we provide the first evidence for pollutioneffects on the DNA modifications in wild populations of Baltic sentinelspecies, the amphipod Monoporeia affinis. A workflow based on high-resolution mass spectrometry to screenand characterize genomic DNA modifications was developed, and itsapplicability was demonstrated by profiling DNA modifications in theamphipods collected in areas with varying pollution loads. Then, thecorrelations between adducts and the contaminants level (polycyclicaromatic hydrocarbons (PAHs), trace metals, and pollution indices)in the sediments at the collection sites were evaluated. A total of119 putative adducts were detected, and some (5-me-dC, N-6-me-dA, 8-oxo-dG, and dI) were structurally characterized. The DNAadductome profiles, including epigenetic modifications, differed betweenthe animals collected in areas with high and low contaminant levels.Furthermore, the correlations between the adducts and PAHs were similaracross the congeners, indicating possible additive effects. Also,high-mass adducts had significantly more positive correlations withPAHs than low-mass adducts. By contrast, correlations between theDNA adducts and trace metals were stronger and more variable thanfor PAHs, indicating metal-specific effects. These associations betweenDNA adducts and environmental contaminants provide a new venue forcharacterizing genome-wide exposure effects in wild populations andapply DNA modifications in the effect-based assessment of chemicalpollution. DNA adductome analysis identifiesexposure to environmentalcontaminants in a sentinel species in the Baltic Sea.

Adductomics studies are used for the detection and characterizationof various chemical modifications (adducts) of nucleic acids and proteins.The advancements in liquid chromatography coupled with high-resolutiontandem mass spectrometry (HRMS/MS) have resulted in efficient methodsfor qualitative and quantitative adductomics. We developed an HRMS-basedmethod for the simultaneous analysis of RNA and DNA adducts in a singlerun and demonstrated its application using Baltic amphipods, usefulsentinels of environmental disturbances, as test organisms. The noveltyof this method is screening for RNA and DNA adducts by a single injectionon an Orbitrap HRMS instrument using full scan and data-independentacquisition. The MS raw files were processed with an open-source program, nLossFinder, to identify and distinguish RNA and DNA adductsbased on the characteristic neutral loss of ribonucleosides and 2 & PRIME;-deoxyribonucleosides,respectively. In the amphipods, in addition to the nearly 150 putativeDNA adducts characterized earlier, we detected 60 putative RNA adducts.For the structural identification of the detected RNA adducts, theMODOMICS database was used. The identified RNA adducts included simplemono- and dimethylation and other larger functional groups on differentribonucleosides and deaminated product inosine. However, 54 of theseRNA adducts are not yet structurally identified, and further workon their characterization may uncover new layers of information relatedto the transcriptome and help understand their biological significance.Considering the susceptibility of nucleic acids to environmental factors,including pollutants, the developed multi-adductomics methodologywith further advancement has the potential to provide biomarkers fordiagnostics of pollution effects in biota.

Electrophilic diol epoxide metabolites are involved in the carcinogenicity of benzo[a]pyrene, one of the widely studied polycyclic aromatic hydrocarbons (PAHs). The exposure of humans to this PAH can be assessed by measuring stable blood protein adducts, such as to histidine and lysine in serum albumin, from their reactive metabolites. In this respect, measurement of the adducts originating from the genotoxic (+)-anti-benzo[a]pyrene diol epoxide is of interest. However, these are difficult to measure at such low levels as are expected in humans generally exposed to benzo[a]pyrene from air pollution and the diet. The analytical methods detecting PAH-biomarkers still suffer from low selectivity and/or detectability to enable generation of data for calculation of in vivo doses of specific stereoisomers, for evaluation of risk factors and assessing risk from exposures to PAH. Here, we suggest an analytical methodology based on high-pressure liquid chromatography (HPLC) coupled to high-resolution tandem mass spectrometry (MS) to lower the detection limits as well as to increase the selectivity with improvements in both chromatographic separation and mass determination. Method development was performed using serum albumin alkylated in vitro by benzo[a]pyrene diol epoxide isomers. The (+)-anti-benzo[a]pyrene diol epoxide adducts could be chromatographically resolved by using an HPLC column with a pentafluorophenyl stationary phase. Interferences were further diminished by the high mass accuracy and resolving power of Orbitrap MS. The achieved method detection limit for the (+)-anti-benzo[a]pyrene diol epoxide adduct to histidine was approximately 4 amol/mg serum albumin. This adduct as well as the adducts to histidine from (−)-anti- and (+/−)-syn-benzo[a]pyrene diol epoxide were quantified in the samples from benzo[a]pyrene-exposed mice. Corresponding adducts to lysine were also quantified. In human serum albumin, the anti-benzo[a]pyrene diol epoxide adducts to histidine were detected in only two out of twelve samples and at a level of approximately 0.1 fmol/mg.

Analytical methods and tools for the characterization of the human exposome by untargeted mass spectrometry approaches are advancing rapidly. Adductomics methods have been developed for untargeted screening of short-lived electrophiles, in the form of adducts to proteins or DNA, in vivo. The identification of an adduct and its precursor electrophile in the blood is more complex than that of stable chemicals. The present work aims to illustrate procedures for the identification of an adduct to N-terminal valine in hemoglobin detected with adductomics, and pathways for the tracing of its precursor and possible exposure sources. Identification of the adduct proceeded via preparation and characterization of standards of adduct analytes. Possible precursor(s) and exposure sources were investigated by measurements in blood of adduct formation by precursors in vitro and adduct levels in vivo. The adduct was identified as hydroxypropanoic acid valine (HPA-Val) by verification with a synthesized reference. The HPA-Val was measured together with other adducts (from acrylamide, glycidamide, glycidol, and acrylic acid) in human blood (n = 51, schoolchildren). The HPA-Val levels ranged between 6 and 76 pmol/g hemoglobin. The analysis of reference samples from humans and rodents showed that the HPA-Val adduct was observed in all studied samples. No correlation of the HPA-Val level with the other studied adducts was observed in humans, nor was an increase in tobacco smokers observed. A small increase was observed in rodents exposed to glycidol. The formation of the HPA-Val adduct upon incubation of blood with glycidic acid (an epoxide) was shown. The relatively high adduct levels observed in vivo in relation to the measured reactivity of the epoxide, and the fact that the epoxide is not described as naturally occurring, suggest that glycidic acid is not the only precursor of the HPA-Val adduct identified in vivo. Another endogenous electrophile is suspected to contribute to the in vivo HPA-Val adduct level. 

DNA adductomics is a relatively new omics approach aiming to measure known and unknown DNA modifications, called DNA adducts. Liquid chromatography–tandem mass spectrometry (LC-MS/MS) has become the most common method for analyzing DNA adducts. Recent advances in the field of mass spectrometry have allowed the possibility to perform a comprehensive analysis of adducts, for instance, by using a nontargeted data-independent acquisition method, with multiple precursor m/z windows as an inclusion list. However, the generated data are large and complex, and there is a need to develop algorithms to simplify and automate the time-consuming manual analysis that has hitherto been used. Here, a graphical user interface (GUI) program was developed, with the purpose of tracking a characteristic neutral loss reaction from tandem mass spectrometry of the nucleoside adducts. This program, called nLossFinder, was developed in the MATLAB platform, available as open-source code. Calf thymus DNA was used as a model for method optimization, and the overall adductomics approach was applied to DNA from amphipods (Monoporeia affinis) collected within the Swedish National Marine Monitoring Program. In the amphipod DNA, over 150 putative adducts were found in comparison to 18 using a manual approach in a previous study. The developed program can improve the processing time for large MS data, as it processes each sample in a few seconds, and hence can be applicable for high-throughput screening of adducts.

The instability of electrophilic reactive metabolites in in vitro metabolism studies makes their accurate analysis challenging. To stabilise the reactive compounds prior to their analysis, different trapping agents, such as thiols, amines and cob(I)alamin, have earlier been tested depending on the metabolites to be analysed and the type of study. In the present work, DNA is introduced as a trapping agent for measuring the formation of bulky electrophilic metabolites. Benzo[a]pyrene (B[a]P), a polycyclic aromatic hydrocarbon (PAH), was used as a model compound in a rat liver S9 metabolic system. Under physiological incubation conditions, B[a]P metabolises to diol epoxide (BPDE) metabolites which were trapped by DNA resulting in the formation of covalently bound DNA adducts. The methodology for analysis of these adducts included extraction of the DNA from the metabolic system, digestion of the DNA to yield nucleosides and analysis of the BPDE-adduct to deoxyguanosine (BPDE-dG) by liquid chromatography coupled to high resolution mass spectrometry (HRMS). The chromatographic conditions in combination with the high mass accuracy data (±3 ppm) was useful in resolving BPDE-dG in its protonated form from the complex set of ions present in the metabolic matrix. The method was validated in terms of sensitivity, specificity, accuracy, precision and recovery, and applied to provide a preliminary estimate of BPDE-dG levels from the metabolism of B[a]P in rat S9. The use of DNA as a trapping agent for in vitro metabolites has a potential to aid in cancer risk assessment procedure of PAHs, for instance, in inter-species comparison of metabolism to reactive metabolites and can be adapted for screening of genotoxic metabolites, e.g., from emerging environmental contaminants.