Novel Techniques for Mapping DNA Damage and Repair in the Brain.

DNA damage in the brain is influenced by endogenous processes and metabolism along with exogenous exposures. Accumulation of DNA damage in the brain can contribute to various neurological disorders, including neurodegenerative diseases and neuropsychiatric disorders. Traditional methods for assessing DNA damage in the brain, such as immunohistochemistry and mass spectrometry, have provided valuable insights but are limited by their inability to map specific DNA adducts and regional distributions within the brain or genome. Recent advancements in DNA damage detection methods offer new opportunities to address these limitations and further our understanding of DNA damage and repair in the brain. Here, we review emerging techniques offering more precise and sensitive ways to detect and quantify DNA lesions in the brain or neural cells. We highlight the advancements and applications of these techniques and discuss their potential for determining the role of DNA damage in neurological disease.

DNA adductomics aided rapid screening of genotoxic impurities using nucleosides and 3D bioprinted human liver organoids.

Current genotoxicity assessment methods are mainly employed to verify the genotoxic safety of drugs, but do not allow for rapid screening of specific genotoxic impurities (GTIs). In this study, a new approach for the recognition of GTIs has been proposed. It is to expose the complex samples to an in vitro nucleoside incubation model, and then draw complete DNA adduct profiles to infer the structures of potential genotoxic impurities (PGIs). Subsequently, the genotoxicity is confirmed in human by 3D bioprinted human liver organoids. To verify the feasibility of the approach, lansoprazole chloride compound (Lanchlor), a PGI during the synthesis of lansoprazole, was selected as the model drug. After confirming genotoxicity by Comet assay, it was exposed to different models to map and compare the DNA adduct profiles by LC-MS/MS. The results showed Lanchlor could generate diverse DNA adducts, revealing firstly its genotoxicity at molecular mechanism of action. Furthermore, the largest variety and content of DNA adducts were observed in the nucleoside incubation model, while the human liver organoids exhibited similar results with rats. The results showed that the combination of DNA adductomics and 3D bioprinted organoids were useful for the rapid screening of GTIs.

Map of benzoapyrene metabolites-DNA adducts in human bronchial epithelial-like cells: Based on chromatin immunoprecipitation followed by sequencing technology / 环境与职业医学

Background The active metabolite of benzo[a]pyrene (BaP), 7,8-dihydroxy-9,10-epoxybenzo[a]pyrene (BPDE), can form adducts with DNA, but the spectrum of BPDE-DNA adducts is unclear. Objective To identify the distribution of BPDE adduct sites and associated genes at the whole-genome level by chromatin immunoprecipitation followed by sequencing (ChIP-Seq), and serve as a basis for further exploring the toxicological mechanisms of BaP. Methods Human bronchial epithelial-like cells (16HBE) were cultured to the fourth generation inthe logarithmic growth phase. Cells were harvested and added to chromatin immunoprecipitation lysis buffer. The lysate was divided into experimental and control groups. The experimental group received a final concentration of 20 μmol·L−1 BPDE solution, while the control group received an equivalent volume of dimethyl sulfoxide solution. The cells were then incubated at 37 °C for 24 h. Chromatin fragments of 100-500 bp were obtained through sonication. BPDE-specific antibody (anti-BPDE 8E11) was used to enrich DNA fragments with BPDE adducts. High-throughput sequencing was conducted to detect BPDE adduct sites. The top 1000 peak sequences were subjected to motif analysis using MEME and DREME software. BPDE adduct target genes at the whole-genome level were annotated, and Gene Ontology (GO) functional analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of BPDE adduct target genes were conducted using bioinformatics techniques. Results The high-throughput sequencing detected a total of 842 BPDE binding sites, distributed across various chromosomes. BPDE covalently bound to both coding and non-coding regions of genes, with 73.9% binding sites located in intergenic regions, 19.6% in intronic regions, and smaller proportions in upstream 2 kilobase, exonic, downstream 2 kilobase, and 5' untranslated regions. Regarding the top 1000 peak sequences, four reliable motifs were identified, revealing that sites rich in adenine (A) and guanine (G) were prone to binding. Through the enrichment analysis of binding sites, a total of 199 BPDE-adduct target genes were identified, with the majority located on chromosomes 1, 5, 7, 12, 17, and X. The GO analysis indicated that these target genes were mainly enriched in nucleic acid and protein binding, participating in the regulation of catalytic activity, transport activity, translation elongation factor activity, and playing important roles in cell division, differentiation, motility, substance transport, and information transfer. The KEGG analysis revealed that these target genes were primarily enriched in pathways related to cardiovascular diseases, cancer, and immune-inflammatory responses. Conclusion Using ChIP-Seq, 199 BPDE adduct target genes at genome-wide level are identified, impacting biological functions such as cell division, differentiation, motility, substance transport, and information transfer. These genes are closely associated with cardiovascular diseases, tumors, and immune-inflammatory responses.

Aflatoxin B1-DNA adducts modify the effects of post-operative adjuvant transarterial chemoembolization improving hepatocellular carcinoma prognosis.

Aim: DNA damage involves in the carcinogenesis of some cancer and may act as a target for therapeutic intervention of cancers. However, it is unclear whether aflatoxin B1 (AFB1)-DNA adducts (ADAs), an important kind of DNA damage caused by AFB1, affect the efficiency of post-operative adjuvant transarterial chemoembolization (po-TACE) treatment improving hepatocellular carcinoma (HCC) survival. Methods: A hospital-based retrospective study, including 318 patients with Barcelona Clinic Liver Cancer (BCLC)-C stage HCC from high AFB1 exposure areas, to investigate the potential effects of ADAs in the tissues with HCC on po-TACE treatment. The amount of ADAs in the cancerous tissues was tested by competitive enzyme-linked immunosorbent assay (c-ELISA). Results: Among these patients with HCC, the average amount of ADAs was 3.00 µmol/mol ± 1.51 µmol/mol DNA in their tissues with cancer. For these patients, increasing amount of ADAs was significantly associated with poorer overall survival (OS) and tumor reoccurrence-free survival (RFS), with corresponding death risk (DR) of 3.69 (2.78-4.91) and tumor recurrence risk (TRR) of 2.95 (2.24-3.88). The po-TACE therapy can efficiently improve their prognosis [DR = 0.59 (0.46-0.76), TRR = 0.63 (0.49-0.82)]. Interestingly, this improving role was more noticeable among these patients with high ADAs [DR = 0.36 (0.24-0.53), TRR = 0.40 (0.28-0.59)], but not among those with low ADAs (P > 0.05). Conclusions: These results suggest that increasing ADAs in the cancerous tissues may be beneficial for po-TACE in ameliorating the survival of patients with HCC.

BPDE-DNA adduct formation and alterations of mRNA, protein, and DNA methylation of CYP1A1, GSTP1, and GSTM1 induced by benzo[a]pyrene and the intervention of aspirin in mice.

Benzo[a]pyrene (B[a]P), one typical environmental pollutant, the toxicity mechanisms, and potential prevention remain perplexing. Available evidence suggests cytochrome P450 1A1 (CYP1A1) and glutathione S-transferases (GSTs) metabolize B[a]P, resulting in metabolic activation and detoxification of B[a]P. This study aimed to reveal the impact of B[a]P exposure on trans-7,8-diol-anti-9,10-epoxide DNA (BPDE-DNA) adduct formation, level of CYP1A1, glutathione S-transferase pi (GSTP1) and glutathione S-transferase mu1 (GSTM1) mRNA, protein and DNA methylation in mice, and the potential prevention of aspirin (ASP). This study firstly determined the BPDE-DNA adduct formation in an acute toxicity test of a large dose in mice induced by B[a]P, which subsequently detected CYP1A1, GSTP1, and GSTM1 at levels of mRNA, protein, and DNA methylation in the organs of mice in a subacute toxicity test at appropriate doses and the potential prevention of ASP, using the methods of real-time quantitative PCR (QPCR), western blotting, and real-time methylation-specific PCR (MSP), respectively. The results verified that B[a]P induced the formation of BPDE-DNA adduct in all the organs of mice in an acute toxicity test, and the order of concentration of which was lung > kidney > liver > brain. In a subacute toxicity test, following B[a]P treatment, mice showed a dose-dependent slowdown in body weight gain and abnormalities in behavioral and cognitive function and which were alleviated by ASP co-treatment. Compared to the controls, following B[a]P treatment, CYP1A1 was significantly induced in all organs in mice at mRNA level (P < 0.05), was suppressed in the lung and cerebrum of mice at protein level, and inhibited at DNA methylation level in the liver, lung, and cerebrum, whereas GSTP1 and GSTM1 at mRNA, protein, and DNA methylation levels showed organ-specific changes in mice following B[a]P treatment, which was generally alleviated by ASP intervention. In conclusion, B[a]P induced BPDE-DNA adduct formation in all organs in mice and altered the mRNA, protein, and DNA methylation levels in CYP1A1, GSTP1, and GSTM1 in an organ-dependent pattern, which could be related to the organ toxicity and mechanism of B[a]P. ASP intervention may be an effective measure to prevent B[a]P toxicity. The findings provide scientific evidence for further study on the organ toxicity and mechanisms of B[a]P.

Regulation of Xenobiotic-Metabolizing Enzymes by 5-Demethylnobiletin and Nobiletin to Mitigate Benzo[a]pyrene-Induced DNA Damage In Vitro and In Vivo.

Benzo[a]pyrene (B[a]P) is a genotoxic polycyclic aromatic hydrocarbon that is metabolized by cytochrome P450 family 1 enzymes (CYP 1s) and can bind to DNA to form DNA adducts, leading to DNA damage and increased colorectal cancer risk. Previous studies have shown polymethoxyflavones to have a high potential for anticancer effects by regulating CYP 1s, especially nobiletin (NBT) and 5-demethylnobiletin (5-DMNB). However, the effects of NBT and 5-DMNB on B[a]P metabolism remain unclear. Therefore, this study aimed to clarify the effects of NBT and 5-DMNB on B[a]P-induced DNA damage in vitro and in vivo. In NCM460 cells, 5-DMNB and NBT appeared to reduce the metabolic conversion of B[a]P by regulating the aryl hydrocarbon receptor (AhR)/CYP 1s signaling pathway. This process protected NCM460 cells from B[a]P's cytotoxic effects by decreasing DNA damage and suppressing B[a]P diol-epoxide-DNA adduct formation. In BALB/c mice, 5-DMNB and NBT also protected against B[a]P-induced DNA damage. Altogether, these findings indicate that 5-DMNB and NBT attenuate B[a]P-induced DNA damage by modulating biotransformation, highlighting their chemopreventive potential against B[a]P-induced carcinogenesis. Therefore, 5-DMNB and NBT are promising agents for colorectal cancer chemoprevention in the future.

Exploration and optimization of extraction, analysis and data normalization strategies for mass spectrometry-based DNA adductome mapping and modeling.

Although interest in characterizing DNA damage by means of DNA adductomics has substantially grown, the field of DNA adductomics is still in its infancy, with room for optimization of methods for sample analysis, data processing and DNA adduct identification. In this context, the first objective of this study was to evaluate the use of hydrophilic interaction (HILIC) vs. reversed phase liquid chromatography (RPLC) coupled to high resolution mass spectrometry (HRMS) and thermal acidic vs. enzymatic hydrolysis of DNA followed by DNA adduct purification and enrichment using solid-phase extraction (SPE) or fraction collection for DNA adductome mapping. The second objective was to assess the use of total ion count (TIC) and median intensity (MedI) normalization compared to QC (quality control), iQC (internal QC) and quality control-based robust locally estimated scatterplot smoothing (LOESS) signal correction (QC-RLSC) normalization for processing of the acquired data. The results demonstrate that HILIC compared to RPLC allowed better modeling of the tentative DNA adductome, particularly in combination with thermal acidic hydrolysis and SPE (more valid models, with an average Q2(Y) and R2(Y) of 0.930 and 0.998, respectively). Regarding the need for data normalization and the management of (limited) system instability and signal drift, QC normalization outperformed TIC, MedI, iQC and LOESS normalization. As such, QC normalization can be put forward as the default data normalization strategy. In case of momentous signal drift and/or batch effects however, comparison to other normalization strategies (like e.g. LOESS) is recommended. In future work, further optimization of DNA adductomics may be achieved by merging of HILIC and RPLC datasets and/or application of 2D-LC, as well as the inclusion of Schiff base stabilization and/or fraction collection in the thermal acidic hydrolysis-SPE sample preparation workflow.

The peroxidation-derived DNA adduct, 6-oxo-M1dG, is a strong block to replication by human DNA polymerase η.

The DNA adduct 6-oxo-M1dG, (3-(2'-deoxy-ß-D-erythro-pentofuranosyl)-6-oxo-pyrimido(1,2alpha)purin-10(3H)-one) is formed in the genome via oxidation of the peroxidation-derived adduct M1dG. However, the effect of 6-oxo-M1dG adducts on subsequent DNA replication is unclear. Here we investigated the ability of the human Y-family polymerase hPol η to bypass 6-oxo-M1dG. Using steady-state kinetics and analysis of DNA extension products by liquid chromatography-tandem mass spectrometry, we found hPol η preferentially inserts a dAMP or dGMP nucleotide into primer-templates across from the 6-oxo-M1dG adduct, with dGMP being slightly preferred. We also show primer-templates with a 3'-terminal dGMP or dAMP across from 6-oxo-M1dG were extended to a greater degree than primers with a dCMP or dTMP across from the adduct. In addition, we explored the structural basis for bypass of 6-oxo-M1dG by hPol η using X-ray crystallography of both an insertion-stage and an extension-stage complex. In the insertion-stage complex, we observed that the incoming dCTP opposite 6-oxo-M1dG, although present during crystallization, was not present in the active site. We found the adduct does not interact with residues in the hPol η active site but rather forms stacking interactions with the base pair immediately 3' to the adduct. In the extension-stage complex, we observed the 3' hydroxyl group of the primer strand dGMP across from 6-oxo-M1dG is not positioned correctly to form a phosphodiester bond with the incoming dCTP. Taken together, these results indicate 6-oxo-M1dG forms a strong block to DNA replication by hPol η and provide a structural basis for its blocking ability.

Characterization and quantitation of busulfan DNA adducts in the blood of patients receiving busulfan therapy.

DNA alkylating drugs have been used as cancer chemotherapy with variable outcomes. The establishment of predictive biomarkers to identify patients who will effectively respond to treatment would allow for the development of personalized therapies. As the degree of interaction of alkylating drug with DNA plays a key role in their mechanism of action, our hypothesis is that the measurement of the DNA adducts formed by alkylating drugs could be used to inform patient stratification. Beginning with busulfan, we took advantage of our DNA adductomic approach to characterize DNA adducts formed by reacting busulfan with calf-thymus DNA. Samples collected from six patients undergoing busulfan-based chemotherapy prior to allogeneic hematopoietic cell transplantation were analyzed for the presence of busulfan-derived DNA adducts. Among the 15 adducts detected in vitro, 12 were observed in the patient blood confirming the presence of a large profile of DNA adducts in vivo. Two of the detected adducts were structurally confirmed by comparison with synthetic standards and quantified in patients. These data confirm our ability to comprehensively characterize busulfan-derived DNA damage and set the stage for the development of methods to support personalized chemotherapy.

A new toxicity mechanism of N-(1,3-Dimethylbutyl)-N'-phenyl-p-phenylenediamine quinone: Formation of DNA adducts in mammalian cells and aqueous organisms.

N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine-quinone (6PPDQ), one of the oxidation products of rubber antioxidant 6PPD, has been identified as a novel toxicant to many organisms. However, an understanding of its underlying toxicity mechanisms remained elusive. In this study, we reported that 6PPDQ could react with deoxyguanosine to form one isomer of 3-hydroxy-1, N2-6PPD-etheno-2'-deoxyguanosine (6PPDQ-dG). Next, by employing an ultra-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) method, we found that 6PPDQ-dG could be detected in genomic DNA from 6PPDQ-treated mammalian cells and Chlamydomonas reinhardtii. We observed positive correlations between concentrations of exogenous 6PPDQ and the amounts of 6PPDQ-dG, and a recovery period after removal of 6PPDQ also led to decreased levels of the adduct in both organisms, which suggested potential repair pathways for this adduct in mammalian cells and unicellular algae. Additionally, we extracted the genomic DNA from tissues of frozen capelin and observed substantial amounts of the adduct in roe and gills, as well as livers at a relatively lower level. These results provided insights into the target organs and tissues that 6PPDQ might accumulate or harm fish. Overall, our study provides a new understanding of the mechanisms of toxicity of 6PPDQ in mammalian cells and aqueous organisms.

DNA adducts as link between in vitro and in vivo carcinogenicity - A case study with benzo[a]pyrene.

To reduce the need for animal tests, in vitro assays are often used as alternative methods. To derive toxic doses for higher tier organisms from in vitro assay results, quantitative in vitro-in vivo extrapolation (qIVIVE) based on physiological-based toxicokinetic (PBTK) models is typically the preferred approach. Such PBTK models require many input parameters to address the route from dose to target site concentration. However, respective data is very often not available. Hence, our aim is to call attention to an alternative way to build a link between animal (in vivo) and cell-derived (in vitro) toxicity data. To this end, we selected the carcinogenic chemical benzo[a]pyrene (BaP) for our study. Our approach relates both in vitro assay and in vivo data to a main intermediate marker structure for carcinogenicity on the subcellular level - the BaP-DNA adduct BaP-7,8-dihydrodiol-9,10-epoxide-deoxyguanosine. Thus, BaP dose is directly linked to a measure of the toxicity-initiating event. We used Syrian hamster embryo (SHE) and Balb/c 3T3 cell transformation assay as in vitro data and compared these data to outcomes of in vivo carcinogenicity tests in rodents. In vitro and in vivo DNA adduct levels range within three orders of magnitude. Especially metabolic saturation at higher doses and interspecies variabilities are identified and critically discussed as possible sources of errors in our simplified approach. Finally, our study points out possible routes to overcome limitations of the envisaged approach in order to allow for a reliable qIVIVE in the future.

Evaluation of DNA adduct damage using G-quadruplex-based DNAzyme.

Toxicity assessment is a major problem in pharmaceutical candidates and industry chemicals development. However, due to the lack of practical analytical methods for DNA adduct analysis, the safety evaluation of drug and industry chemicals was severely limited. Here, we develop a DNAzyme-based method to detect DNA adduct damage for toxicity assessment of drugs and chemicals. Among 18 structural variants of G4 DNAzyme, EA2 DNAzyme exhibits an obvious DNA damaging effect of styrene oxide (SO) due to its unstable structure. The covalent binding of SO to DNAzyme disrupts the Hoogsteen hydrogen bonding sites of G-plane guanines and affects the formation of the G4 quadruplex. DNA damage chemicals reduce the peroxidase activity of the G4 DNAzyme to monitor the DNA adduct damage by disrupting the structural integrity of the G4 DNAzyme. Our method for genotoxic assessment of pharmaceutical candidates and industrial chemicals can elucidate the complex chemical pathways leading to toxicity, predict toxic effects of chemicals, and evaluate possible risks to human health.

Aristolochic acid-containing Chinese herbal medicine and upper urinary tract urothelial carcinoma in Taiwan: a narrative review.

PURPOSE: The high incidence of upper urinary tract urothelial carcinoma (UTUC) in Taiwan is largely due to exposure to aristolochic acid (AA), a principal component of Aristolochia-based herbal medicines. Here we systematically review the molecular epidemiology, clinical presentation and biomarkers associated with AA-induced UTUC. METHODS: This is a narrative review. Medline, Embase, and Web of Science were searched from inception to December 31, 2021. Studies evaluating the association, detection, and clinical characteristics of AA and UTUC were included. RESULTS: A nationwide database revealed 39% of the Taiwanese population had been exposed to AA-containing herbs between 1997 and 2003. Epidemiological reports revealed AA posed a significantly higher hazard for renal failure and UTUC in herbalists and the general population who ingested AA-containing herbs. The presence of aristolactam-DNA adducts and a distinctive signature mutation, A:T to T:A transversions, located predominantly on the non-transcribed DNA strand, with a strong preference for deoxyadenosine in a consensus sequence (CAG), was observed in many UTUC patients. Clinically, AA-related UTUC patients were characterized by a younger age, female gender, impaired renal function and recurrence of contralateral UTUC. To date, there are no preventive measures, except prophylactic nephrectomy, for subjects at risk of AA nephropathy or AA-related UTUC. CONCLUSION: AA exposure via Aristolochia-based herbal medicines is a problem throughout Taiwan, resulting in a high incidence of UTUC. Aristolactam-DNA adducts and a distinctive signature mutation, A:T to T:A transversions, can be used as biomarkers to identify AA-related UTUC. AA-related UTUC is associated with a high recurrence rate of contralateral UTUC.

Use of DNA adduct and histopathological defects as indications for bio-persistence potency of zinc oxide nanoparticles in gastropod, Monacha cartusiana (Mǜller) after short-term exposure.

The cytotoxic effects of metallic nanoparticles (MNPs) might be revealed in genomic and histopathological defects. Therefore current study aimed to assess the bio-persistence and adverse effects potency of zinc oxide nanoparticles (ZnONPs) in the gastropod, Monacha cartusiana. Gastropods were exposed to 74 µg/mL for 14 d then the DNA adduct and histopathological defect profiles were assessed. The results demonstrated significant decline in the estimated genomic template stability (GTS%) in haemolymph and digestive gland ranging from 10.0 to 42.9% in treated animals compared to controls. In the treated and recovered snails, randomly amplified polymorphic (RAPD)-DNA showed the appearance and/or disappearance of DNA bands, indicating DNA damage due to the cytotoxicity of ZnONPs on gastropods. Significant defects in microvilli (MV), nucleus (N), mitochondria (M), and execratory glands (EXG) were noticed in the treated individuals with respect to controls. The remaining live animals were subjected to a recovery period (14 d, without treatment) and slight recovery profiles were reported for both measures compared to the control group. The recovery pattern was recognized in the nucleus/cytoplasm ratio with 0.186 and 0.428 in the treated and recovered groups concerning their control (0.176). The studied parameters reported herein might be reliable tools to assess accumulation and bio-persistence patterns of NPs in the organisms for short-term exposure indicating the cytotoxic and genotoxic effects. Also, gastropods may provide simple models for evaluating the ecotoxicological effects of nanomaterials.

Label-free and de-conjugation-free workflow to simultaneously quantify trace amount of free/conjugated and protein-bound estrogen metabolites in human serum.

Different chemical forms of sex hormones including free/conjugated metabolites as well as their protein/DNA adducts in human serum are a panel of important indicators of health conditions. It is, however, hard to quantify all species simultaneously due to the lack of general extraction, derivatization, and de-conjugation methods. Here we developed a label-free and de-conjugation-free workflow to quantify 11 free/conjugated estrogen metabolites including depurinating DNA and protein adduct forms of 4-hydroxyestradiol (4OHE2) in human serum. Acetonitrile acts as an excellent solvent to purify adducted and non-adducted human serum albumin (HSA) by precipitation as well as to extract free/conjugated metabolites and depurinating DNA adducts from the supernatant by salting-out effect. The adduction level of 4OHE2 on HSA was determined by proteomics; free/conjugated metabolites were quantified by a newly developed microflow liquid chromatography (microflow LC)-nanoelectrospray ionization (nanoESI)-multiple reaction monitoring (MRM) method with high reproducibility (7-22% RSD, n > 3) and sub-picogram levels (0.6-20 pg/mL) of quantification limits (S/N = 8) by using non-pulled capillary as nano-ESI emitter. This workflow was demonstrated to reveal endogenous adduction level of 4OHE2 on HSA as well as circulation levels of free/conjugated metabolites in clinical samples. 4OHE2 in human serum were solely detected as protein-bound form, indicating the merit of such integrated platform covering unstable or active metabolites. Compared to traditional methods using labeling or de-conjugation reaction, this workflow is much simplier, more sensitive, and more specific. Moreover, it can be widely applied in omics to concurrently access various bio-transformed known and un-known markers or drugs.

Boosting cisplatin chemotherapy by nanomotor-enhanced tumor penetration and DNA adducts formation.

Despite many nano-based strategies devoted to delivering cisplatin for tumor therapy, its clinical benefits are compromised by poor tissue penetration and limited DNA adducts formation of the drug. Herein, a cisplatin loading nanomotor based janus structured Ag-polymer is developed for cisplatin delivery of deeper tissue and increased DNA adducts formation. The nanomotor displayed a self-propelled tumor penetration fueled by hydrogen peroxide (H2O2) in tumor tissues, which is catalytically decomposed into a large amount of oxygen bubbles by Ag nanoparticles (NPs). Notably, cisplatin could elevate the intracellular H2O2 level through cascade reactions, further promote the degradation of Ag NPs accompanied with the Ag+ release, which could downregulate intracellular Cl- through the formation of AgCl precipitate, thereby enhancing cisplatin dechlorination and Pt-DNA formation. Moreover, polymer can also inhibit the activity of ALKBH2 (a Fe2+-dependent DNA repair enzyme) by chelating intracellular Fe2+ to increase the proportion of irreparable Pt-DNA cross-links. It is found that deep tissue penetration, as well as the increased formation and maintenance of Pt-DNA adducts induced by the nanomotor afford 80% of tumor growth inhibition with negligible toxicity. This work provides an important perspective of resolving chemotherapeutic barriers for boosting cisplatin therapy.

Fluorescent adenine analogues with ESPT characteristic utilized for real-time detecting DNA adduct.

The 8-oxo-7,8-dihydro-2'-deoxyguanine (8-oxoG) is the representative damaged nucleoside that may increase the risk of developing diseases. Accordingly, the selective detection of 8-oxoG in DNA with minimal disturbance to the native structure is important to have an in-depth understanding of the formation mechanism and becomes an attractive tool for genomic research. To identify the DNA adduct in real-time efficiently, a series of quasi-intrinsic optical probes are performed based on the natural adenine, which has preference to form a stable base pair with 8-oxoG in the syn conformation. The calculations revealed that the A-analogues in solution could bring red-shifted absorption spectra and bright photoluminescence arisen from the additional π-conjugation by means of fluorophore modification and the ring expansion. Especially, A1 possesses large Stokes shifts and the highest fluorescence intensity in emission, which is proposed as the biosensor to monitor the optical changes in the presence and absence of the considered 8-oxoG. It is found that the fluorescence is insensitive to base pairing with thymine, while the excited state intermolecular proton transfer (ESPT) induced efficient fluorescence quenching is observed upon pairing with the 8-oxoG. To evaluate the direct usefulness of the bright adenine analogues in biological environment, we further examined the influences of linking deoxyribose on the absorption and emission, which are consistent with the experimental data.

Aristolochic acids exposure was not the main cause of liver tumorigenesis in adulthood.

Aristolochic acids (AAs) have long been considered as a potent carcinogen due to its nephrotoxicity. Aristolochic acid I (AAI) reacts with DNA to form covalent aristolactam (AL)-DNA adducts, leading to subsequent A to T transversion mutation, commonly referred as AA mutational signature. Previous research inferred that AAs were widely implicated in liver cancer throughout Asia. In this study, we explored whether AAs exposure was the main cause of liver cancer in the context of HBV infection in mainland China. Totally 1256 liver cancer samples were randomly retrieved from 3 medical centers and a refined bioanalytical method was used to detect AAI-DNA adducts. 5.10% of these samples could be identified as AAI positive exposure. Whole genome sequencing suggested 8.41% of 107 liver cancer patients exhibited the dominant AA mutational signature, indicating a relatively low overall AAI exposure rate. In animal models, long-term administration of AAI barely increased liver tumorigenesis in adult mice, opposite from its tumor-inducing role when subjected to infant mice. Furthermore, AAI induced dose-dependent accumulation of AA-DNA adduct in target organs in adult mice, with the most detected in kidney instead of liver. Taken together, our data indicate that AA exposure was not the major threat of liver cancer in adulthood.

A Comprehensive Database for DNA Adductomics.

The exposure of human DNA to genotoxic compounds induces the formation of covalent DNA adducts, which may contribute to the initiation of carcinogenesis. Liquid chromatography (LC) coupled with high-resolution mass spectrometry (HRMS) is a powerful tool for DNA adductomics, a new research field aiming at screening known and unknown DNA adducts in biological samples. The lack of databases and bioinformatics tool in this field limits the applicability of DNA adductomics. Establishing a comprehensive database will make the identification process faster and more efficient and will provide new insight into the occurrence of DNA modification from a wide range of genotoxicants. In this paper, we present a four-step approach used to compile and curate a database for the annotation of DNA adducts in biological samples. The first step included a literature search, selecting only DNA adducts that were unequivocally identified by either comparison with reference standards or with nuclear magnetic resonance (NMR), and tentatively identified by tandem HRMS/MS. The second step consisted in harmonizing structures, molecular formulas, and names, for building a systematic database of 279 DNA adducts. The source, the study design and the technique used for DNA adduct identification were reported. The third step consisted in implementing the database with 303 new potential DNA adducts coming from different combinations of genotoxicants with nucleobases, and reporting monoisotopic masses, chemical formulas, .cdxml files, .mol files, SMILES, InChI, InChIKey and IUPAC nomenclature. In the fourth step, a preliminary spectral library was built by acquiring experimental MS/MS spectra of 15 reference standards, generating in silico MS/MS fragments for all the adducts, and reporting both experimental and predicted fragments into interactive web datatables. The database, including 582 entries, is publicly available (https://gitlab.com/nexs-metabolomics/projects/dna_adductomics_database). This database is a powerful tool for the annotation of DNA adducts measured in (HR)MS. The inclusion of metadata indicating the source of DNA adducts, the study design and technique used, allows for prioritization of the DNA adducts of interests and/or to enhance the annotation confidence. DNA adducts identification can be further improved by integrating the present database with the generation of authentic MS/MS spectra, and with user-friendly bioinformatics tools.

Influences of polycyclic aromatic hydrocarbon on the epigenome toxicity and its applicability in human health risk assessment.

The existence of polycyclic aromatic hydrocarbons (PAHs) in ambient air is an escalating concern worldwide because of their ability to cause cancer and induce permanent changes in the genetic material. Growing evidence implies that during early life-sensitive stages, the risk of progression of acute and chronic diseases depends on epigenetic changes initiated by the influence of environmental cues. Several reports deciphered the relationship between exposure to environmental chemicals and epigenetics, and have known toxicants that alter the epigenetic states. Amongst PAHs, benzo[a]pyrene (B[a]P) is accepted as a group 1 cancer-causing agent by the International Agency for the Research on Cancer (IARC). B[a]P is a well-studied pro-carcinogen that is metabolically activated by the aryl hydrocarbon receptor (AhR)/cytochrome P450 pathway. Cytochrome P450 plays a pivotal role in the stimulation step, which is essential for DNA adduct formation. Accruing evidence suggests that epigenetic alterations assume a fundamental part in PAH-promoted carcinogenesis. This interaction between PAHs and epigenetic factors results in an altered profile of these marks, globally and locus-specific. Some of the epigenetic changes due to exposure to PAHs lead to increased disease susceptibility and progression. It is well understood that exposure to environmental carcinogens, such as PAH triggers disease pathways through changes in the genome. Several evidence reported due to the epigenome-wide association studies, that early life adverse environmental events may trigger widespread and persistent variations in transcriptional profiling. Moreover, these variations respond to DNA damage and/or a consequence of epigenetic modifications that need further investigation. Growing evidence has associated PAHs with epigenetic variations involving alterations in DNA methylation, histone modification, and micro RNA (miRNA) regulation. Epigenetic alterations to PAH exposure were related to chronic diseases, such as pulmonary disease, cardiovascular disease, endocrine disruptor, nervous system disorder, and cancer. This hormetic response gives a novel perception concerning the toxicity of PAHs and the biological reaction that may be a distinct reliance on exposure. This review sheds light on understanding the latest evidence about how PAHs can alter epigenetic patterns and human health. In conclusion, as several epigenetic change mechanisms remain unclear yet, further analyses derived from PAHs exposure must be performed to find new targets and disease biomarkers. In spite of the current limitations, numerous evidence supports the perception that epigenetics grips substantial potential for advancing our knowledge about the molecular mechanisms of environmental toxicants, also for predicting health-associated risks due to environmental circumstances exposure and individual susceptibility.