Bulky DNA adducts, increase leads to Increase, Chromosomal aberrations

This KER describes the relationship between increased Bulky DNA adduct formation (upstream KD/MIE) and increased chromosomal aberrations (downstream KE/AO2). The likelihood of chromosomal aberrations (downstream KE/AO2) increases with the accumulation and persistence of DNA adducts, particularly bulky adducts that substantially distort DNA structure and interfere with normal DNA replication and repair. Under these conditions, DNA damage is more likely to be converted into structural chromosomal alterations, resulting in an increased frequency of chromosomal aberrations. 

Evidence was collected for this KER in two methods. The initial method being through expert knowledge from Dr. Carole Yauk, Emmanuelle Demuynck, and the research team at PARC. The second strategy followed that described in Huliganga (2025).  

The initial articles collected were from expert knowledge through presentation as well as previous searches conducted by the PARC team with Emmanuelle as a lead. Notably, the paper provided by Dr. Yauk was used to look for direct connections in connectedpapers.com. Any paper with direct connections, that being a forward or backward citation, were collected. The papers provided by Emmanuelle and the PARC team were already sorted in an Excel spreadsheet. These were then sorted for which were labeled as having “relevance” to the overall AOP and collected. The final two collections were conducted on 6/26/2026 & 7/15/2026, using OMNI (provided by the University of Ottawa). The searches consisted of Boolean strings using either “any field contains” or “any phrase contains exact phrase” requirements with the titles of the KEs as the terms (e.g. “any field contains Bulky adducts & chromosomal aberrations”). A full list of terms used has been included below. 

All articles from expert sources were added to covidence.com for tracking systematic review of the articles. For the searches using OMNI, only the top 50 results were downloaded to RIS file and submitted to covidence.com for review. During this time, covidence was able to remove duplicates as well as help organize inclusion criteria following a modified PECO like that in Huliganga (2025). The inclusion criteria, for this specific KER, included: Bulky DNA adducts, Chromosomal aberrations (or any combination of these two), All species, all life stages. During review of the titles and abstracts it was noted on each paper the reason for inclusion to the next step or the reason for exclusion from the review. Once they reached full text review, the paper was reviewed for relevant data, graphs, methods, as well as background of the KER. Once all papers were reviewed, the data was extracted and placed into the corresponding KER. The final search day, on the 15th of July, was not included in Covidence.com, as there were only 24 articles and it was found easier to read through each paper separately and determine inclusion at the point of reading using the same inclusion criteria.  

The scientific evidence supporting this KER is generally strong particularly with respect to dose-response and incidence concordance. Multiple studies have demonstrated consistent increases in upstream and downstream events across a range of exposure levels and biological tissues, while epidemiological observations show an association between adduct formation and chromosomal aberrations in exposed populations with polymorphisms. Temporal concordance remains less well established, as relatively few studies have directly measured both key events within the same experimental design. This limitation introduces some uncertainty regarding the precise progression of events. Nevertheless, the mechanistic relationship is supported by substantial biological plausibility and convergent empirical evidence from both experimental and observational studies. Overall, despite gaps in temporal evidence and direct event-to-event measurements, the weight of evidence supports a high level of confidence in this KER.  

Available evidence suggests that a substantial increase in DNA adduct formation is required before detection of chromosomal aberrations. In the study conducted by Long et al., increases in DNA adducts were observed at doses as low as 0.20 mg/kg bw/day. While statistically significant increases in micronucleated reticulocytes and normochromatic erythrocytes were not observed until 3.13 mg/kg bw/day. Micronucleus frequencies increased approximately 4.5-fold and 3.7-fold above control levels at this dose, indicating that substantial accumulation of DNA adducts is required before measurable chromosomal damage is induced. This threshold-like relationship is further supported by benchmark dose modelling, where the BMD100 for DNA adduct formation (0.0286 mg/kg bw/day), demonstrating that DNA adduct formation is a considerably more sensitive endpoint.  

Evidence for the temporal relationship between these key events is more limited. Mimmler et al., demonstrated that DNA adducts were detectable within 2-8 hours of exposure and remained elevated for up to 48 hours depending on the compound and concentration used. In contrast, chromosomal aberrations were only evaluated after approximately 38 hours (24-hours exposure plus 14-hour mitotic arrest), at which point significant increases were observed. These findings indicate that DNA adduct formation precedes chromosomal aberration induction. However, the minimum duration or magnitude of adduct accumulation required to trigger chromosomal damage cannot be quantitatively defined because both key events were not assessed concurrently across multiple time points. Consequently, while there is strong evidence that DNA adduct formation occurs earlier and at lower doses than chromosomal aberrations, the precise temporal and quantitative relationship between the two key events remains uncertain.   

Due to the conserved structure of DNA across cellular organisms, the potential for bulky adduct formation exists across taxa, sexes, and life stages. Bulky DNA adducts can form in a wide range of cell types following exposure to endogenous and exogenous genotoxic agents (Long et al., 2018). The cellular processes involved in DNA damage recognition and repair are also broadly conserved across species. While quantitative differences in adduct formation and repair may occur among taxa, sexes, and developmental stages, elevated levels of persistent DNA adduct increase the likelihood of chromosomal aberrations in dividing cells. Therefore, this KER is considered broadly applicable across taxa, sexes, and life stages. 

Bi, Xiaohui, et al. “DNA Polymerase κ Is Specifically Required for Recovery from the Benzo[a]Pyrene-Dihydrodiol Epoxide (BPDE)-Induced S-Phase Checkpoint.” The Journal of Biological Chemistry [United States], vol. 280, no. 23, June 2005, pp. 22343–55, https://doi.org/10.1074/jbc.M501562200. 

Georgiadis, P., et al. “Impact of Phase I or Phase II Enzyme Polymorphisms on Lymphocyte DNA Adducts in Subjects Exposed to Urban Air Pollution and Environmental Tobacco Smoke.” Toxicology Letters [Shannon], vol. 149, no. 1, April 2004, pp. 269–80, https://doi.org/10.1016/j.toxlet.2003.12.038. 

Georgiadis, Panagiotis, et al. “Interactions between CYP1A1 Polymorphisms and Exposure to Environmental Tobacco Smoke in the Modulation of Lymphocyte Bulky DNA Adducts and Chromosomal Aberrations.” Carcinogenesis (New York) [Oxford], vol. 26, no. 1, January 2005, pp. 93–101, https://doi.org/10.1093/carcin/bgh294. 

Long, Alexandra S., et al. “Benchmark Dose Analyses of Multiple Genetic Toxicity Endpoints Permit Robust, Cross-Tissue Comparisons of MutaMouse Responses to Orally Delivered Benzo[a]Pyrene.” Archives of Toxicology [Berlin/Heidelberg], vol. 92, no. 2, February 2018, pp. 967–82, https://doi.org/10.1007/s00204-017-2099-2. 

Mimmler, Maximilian, et al. “DNA Damage Response Curtails Detrimental Replication Stress and Chromosomal Instability Induced by the Dietary Carcinogen PhIP.” Nucleic Acids Research [England], vol. 44, no. 21, December 2016, pp. 10259–76, https://doi.org/10.1093/nar/gkw791. 

Nikolova, Teodora, et al. “The γH2AX Assay for Genotoxic and Nongenotoxic Agents: Comparison of H2AX Phosphorylation with Cell Death Response.” Toxicological Sciences [United States], vol. 140, no. 1, July 2014, pp. 103–17, https://doi.org/10.1093/toxsci/kfu066. 

Pavel Rossner, et al. “Genetic Toxicology and Carcinogenesis.” Toxicology for the Health and Pharmaceutical Sciences, edited by Antonio Peña-Fernández et al., 1st ed., CRC Press, 2022, pp. 81–97, https://doi.org/10.1201/9780203730584-5. 

Rossner, Pavel, et al. “Molecular Epidemiology Focused on Airborne Carcinogens.” Air Pollution and Health Effects, edited by Srikanth S. Nadadur and John W. Hollingsworth, Springer London, Limited, 2015, pp. 185–212, https://doi.org/10.1007/978-1-4471-6669-6_7. 

Tu, Lan Chun, et al. “Hedamycin, a DNA Alkylator, Induces γH2AX and Chromosome Aberrations: Involvement of Phosphatidylinositol 3-Kinase–Related Kinases and DNA Replication Fork Movement.” Molecular Cancer Therapeutics, vol. 4, no. 8, August 2005, pp. 1175–85, https://doi.org/10.1158/1535-7163.MCT-05-0054.