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Event: 1554

Key Event Title

A descriptive phrase which defines a discrete biological change that can be measured. More help

Increase Chromosomal Aberrations

Short name

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Increase chromosomal aberrations

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Biological Context

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Level of Biological Organization
Molecular

Cell term

The location/biological environment in which the event takes place.The biological context describes the location/biological environment in which the event takes place. For molecular/cellular events this would include the cellular context (if known), organ context, and species/life stage/sex for which the event is relevant. For tissue/organ events cellular context is not applicable. For individual/population events, the organ context is not applicable. Further information on Event Components and Biological Context may be viewed on the attached pdf. More help

Organ term

Key Event Components

The KE, as defined by a set structured ontology terms consisting of a biological process, object, and action with each term originating from one of 14 biological ontologies (Ives, et al., 2017; https://aopwiki.org/info_pages/2/info_linked_pages/7#List). Biological process describes dynamics of the underlying biological system (e.g., receptor signalling).Biological process describes dynamics of the underlying biological system (e.g., receptor signaling). The biological object is the subject of the perturbation (e.g., a specific biological receptor that is activated or inhibited). Action represents the direction of perturbation of this system (generally increased or decreased; e.g., ‘decreased’ in the case of a receptor that is inhibited to indicate a decrease in the signaling by that receptor). Note that when editing Event Components, clicking an existing Event Component from the Suggestions menu will autopopulate these fields, along with their source ID and description. To clear any fields before submitting the event component, use the 'Clear process,' 'Clear object,' or 'Clear action' buttons. If a desired term does not exist, a new term request may be made via Term Requests. Event components may not be edited; to edit an event component, remove the existing event component and create a new one using the terms that you wish to add. Further information on Event Components and Biological Context may be viewed on the attached pdf. More help

Key Event Overview

AOPs Including This Key Event

All of the AOPs that are linked to this KE will automatically be listed in this subsection. This table can be particularly useful for derivation of AOP networks including the KE.Clicking on the name of the AOP will bring you to the individual page for that AOP. More help

AOP Name	Role of event in AOP	Point of Contact	Author Status	OECD Status
DNA damage and metastatic breast cancer	KeyEvent	Usha Adiga (send email)	Under development: Not open for comment. Do not cite	Under Development

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) that help to define the biological applicability domain of the KE.In many cases, individual species identified in these structured fields will be those for which the strongest evidence used in constructing the AOP was available in relation to this KE. More help

Term	Scientific Term	Evidence	Link
human	Homo sapiens	High	NCBI
rat	Rattus norvegicus	High	NCBI
mouse	Mus musculus	High	NCBI

Life Stages

An indication of the the relevant life stage(s) for this KE. More help

Life stage	Evidence
All life stages	High

Sex Applicability

An indication of the the relevant sex for this KE. More help

Term	Evidence
Unspecific	High

Key Event Description

A description of the biological state being observed or measured, the biological compartment in which it is measured, and its general role in the biology should be provided. More help

The term "structural chromosomal aberrations" refers to chromosome damage caused by breaks in the DNA that can result in the deletion, addition, or rearrangement of chromosomal segments. According to whether one or both chromatids are affected, chromosomal aberrations can be classified into two main groups: chromatid-type and chromosome-type. Additionally, they can be divided into rejoined and non-rejoined aberrations. Translocations, insertions, dicentrics, and rings are examples of rejoined aberrations, whereas acentric fragments and breaks are examples of unrejoined aberrations (Savage, 1976). Some of these abnormalities, like reciprocal translocations, are long-lasting and can last for many years (Tucker and Preston, 1996). Others, such as dicentrics and acentric fragments, are unstable and weaken with each cell division due to cell death (Boei et al., 1996). After cell division, these activities might still be visible, and the DNA is irreversibly damaged. The occurrence of chromosomal abnormalities is linked to cancer development and cell death (Mitelman, 1982).

A missing, excess, or asymmetrical part of chromosomal DNA is referred to as a chromosomal aberration (CA). There are various double-strand break (DSB) repair mechanisms that could be responsible for these DNA modifications in the chromosome structure (Obe et al., 2002).

The four basic categories of CAs are inversions, translocations, duplications, and deletions. When a section of a chromosome's genetic material is destroyed, deletions take place. When a chromosome's end portion is cut, terminal deletions result.

When a chromosome splits into two different places and wrongly rejoins, leaving the middle portion out, interstitial deletions result. Duplications occur when excess genetic material is added to or rearranged; they can take the forms of transpositions, tandem duplications, reverse duplications, and misplaced duplications (Griffiths et al., 2000). A segment of one chromosome is transferred to a non-homologous chromosome in translocations (Bunting and Nussenzweig, 2013). A reciprocal translocation occurs when regions of two non-homologous chromosomes are switched. When an inversion occurs, the DNA sequence is effectively reversed because both ends of the chromosome split and are ligated at the opposite ends.

The copy number variant is a fifth type of CA that can exist in the genome (CNV). CNVs are deletions or duplications that can range in size from 50 base pairs (Arlt et al., 2012; Arlt et al., 2014; Liu et al., 2013) up into the megabase pair range and may make up more than 10% of the human genome (Shlien et al., 2009; Zhang et al., 2016; Hastings et al., 2009). (Arlt et al., 2012; Wilson et al., 2015; Arlt et al., 2014; Zhang et al., 2016). According to Wilson et al. (2015), CNV regions are particularly abundant in large active transcription units and genes, and they are especially problematic when they result in the duplication of oncogenes or the loss of tumor suppressor genes (Liu et al., 2013; Curtis et al., 2012).

Recurrent and non-recurrent CNVs are two different types. Non-allelic homologous recombination (NAHR), a recombination process that occurs during meiosis, is hypothesised to be the cause of recurrent CNVs (Arlt et al., 2012; Hastings et al., 2009). These germline CNVs, also known as recurrent CNVs, may be inherited and are hence prevalent in various people (Shlien et al., 2009; Liu et al., 2013). It is thought that non-recurrent CNVs are created in mitotic cells during the replication process. It has been proposed that replication-related stress, particularly stalled replication forks, triggers microhomology-mediated processes to break the replication stall, which frequently leads to duplications or deletions, despite the fact that the mechanism is not well understood.

Two models that have been proposed to explain this mechanism include the Fork Stalling and Template Switching (FoSTeS) model, and the Microhomology-Mediated Break-Induced Replication (MMBIR) model (Arlt et al., 2012; Wilson et al., 2015; Lee et al., 2007; Hastings et al., 2009).

Depending on whether the aberration affects the chromatid or the chromosome, CAs can be categorized. Chromosome-type aberrations (CSAs) are chromosome breakage and chromatid swaps; ring chromosomes, marker chromosomes, and dicentric chromosomes are examples of chromatid-type aberrations (CTAs) (Bonassi et al., 2008; Hagmar et al., 2004). Micronuclei (MN; small nucleus-like structures that contain a chromosome or a fragment of a chromosome that was lost during mitosis) and nucleoplasmic bridges (NPBs; physical linkages between the two nuclei) are visible in binucleated cells when cells are halted at the cytokinesis step (El-Zein et al., 2014). The DNA sequence can be examined to evaluate other CAs, as it is for identifying copy number variants (CNVs) (Liu et al., 2013).

Essentiality of the key event

Chromosomal aberrations, such as mutations, deletions, and translocations, are indicative of genetic damage, which can result from exposure to genotoxic agents. This key event represents a mechanistic step that contributes to the overall progression of the pathway, helping to bridge the gap between the initial exposure and the manifestation of adverse effects.

By showcasing experimental evidence that supports the occurrence of increased chromosomal aberrations in response to the MIE, the AOP gains scientific credibility and biological plausibility. Studies demonstrating the genotoxic effects of certain substances provide empirical support for the connectivity of events within the pathway. For example, genotoxicity assays that detect structural changes in chromosomes can serve as evidence of chromosomal aberrations (e.g., Ames test, in vitro micronucleus assay).

Furthermore, the presence of increased chromosomal aberrations is indicative of potential genetic harm, which aligns with the adverse outcome. This insight aids in risk assessment and regulatory decision-making, as the occurrence of genotoxicity informs the evaluation of the potential health risks associated with exposure to certain agents.

Fischer et al., in their mRNA expression profiles showed that the tumor subtypes of neuroblastoma had significantly more segmental genomic imbalances, indicating that a combination of expression profiling (miRNAs and mRNAs) with analysis of DNA copy number alterations, will lead to improved prognostication of this often fatal tumor subtype (Fischer et al., 2010)

How It Is Measured or Detected

A description of the type(s) of measurements that can be employed to evaluate the KE and the relative level of scientific confidence in those measurements.These can range from citation of specific validated test guidelines, citation of specific methods published in the peer reviewed literature, or outlines of a general protocol or approach (e.g., a protein may be measured by ELISA). Do not provide detailed protocols. More help

Assay	References	Description
Fluorescent In Situ Hybridization (FISH)	Beaton et al., 2013; Pathak et al., 2017	Fluorescent assay of metaphase chromosomes that can detect CAs through chromosome painting and microscopic analysis
Cytokinesis Block Micronucleus (CBMN) Assay with Microscopy in vitro	Fenech, 2000; OECD, 2016a	Cells are cultured with cytokinesis blocking agent, fixed to slides, and undergo MN quantification using microscopy.
Micronucleus (MN) Assay by Microscopy in vivo	OECD, 2016b	Cells are fixed on slides and MN are scored using microscopy. Red blood cells can also be scored for MN using flow cytometry (see below)
CBMN with Imaging Flow Cytometry	Rodrigues et al., 2015	Cells are cultured with cytokinesis blocking agent, fixed in solution, and imaged with flow cytometry to quantify MN
Flow cytometry detection of MN	Dertinger et al., 2004; Bryce et al., 2007; OECD 2016a, 2016b	In vivo and in vitro flow cytometry-based, automated micronuclei measurements are also done without cytokinesis block. MN analysis in vivo is performed in peripheral blood cells to detect MN in erythrocytes and reticulocytes.
High-throughput biomarker assays (indirect measures to confirm clastogenicity)	Bryce et al. 2014, 2016, 2018 Khoury et al., 2013, Khoury et al., 2016) Hendriks et al., 2012, 2016; Wink et al., 2014	Multiplexed biomarkers can be measured by flow cytometry are used to discern clastogenic and aneugenic mechanisms for MN induction. Flow cytometry-based quantification of γH2AX foci and p53 protein expression (Bryce et al., 2016). Prediscreen Assay– In-Cell Western-based quantification of γH2AX Green fluorescent protein reporter assay to detect the activation of stress signaling pathways, including DNA damage signaling including a reporter porter that is associated with DNA double strand breaks.
Dicentric Chromosome Assay (DCA)	Abe et al., 2018	Cells are fixed on microscope slides, chromosomes are stained, and the number of dicentric chromosomes are quantified
High content imaging	Shahane et al., 2016	DNA can be stained using fluorescent dyes and micronuclei can be scored high-throughput microscopy image analysis.
Chromosomal aberration test	OECD, 2016c; 2016d; 20l16e	In vitro, the cell cycle is arrested at metaphase after 1.5 cell cycle following 3-6 hour exposure In vivo, the test chemical is administered as a single treatment, bone marrow is collected 18-24 hrs later (TG 475), while testis is collected 24-48 hrs later (TG 483). The cell cycle is arrested with a metaphase-arresting chemical (e.g., colchicine) 2-5 hours before cell collection. Once cells are fixed and stained on microscope slides, chromosomal aberrations are scored
Array Comparative Genomic Hybridization (aCGH) or SNP Microarray	Adewoye et al., 2015; Wilson et al., 2015; Arlt et al., 2014; Redon et al., 2006; Keren, 2014; Mukherjee, 2017	CNVs are most commonly detected using global DNA microarray technologies; This method, however, is unable to detect balanced CAs, such as inversions
Next Generation Sequencing (NGS): Whole Genome Sequencing (WGS) or Whole Exome Sequencing (WES)	Liu, 2013; Shen, 2016; Mukherjee, 2017	CNVs are detected by fragmenting the genome and using NGS to sequence either the entire genome (WGS), or only the exome (WES); Challenges with this methodology include only being able to detect CNVs in exon-rich areas if using WES, the computational investment required for the storage and analysis of these large datasets, and the lack of computational algorithms available for effectively detecting somatic CNVs

Domain of Applicability

A description of the scientific basis for the indicated domains of applicability and the WoE calls (if provided). More help

Chromosomal aberrations indicating clastogenicity can occur in any eukaryotic or prokaryotic cell. However, dose-response curves can differ depending on the cell cycle stage when the DSB agent was introduced (Obe et al., 2002).

References

List of the literature that was cited for this KE description. More help

Abe, Y et al. (2018), “Dose-response curves for analyzing of dicentric chromosomes and chromosome translocations following doses of 1000 mGy or less, based on irradiated peripheral blood samples from five healthy individuals”, J Radiat Res. 59(1), 35-42. doi:10.1093/jrr/rrx052

Adewoye, A.B.et al. (2015), “The genome-wide effects of ionizing radiation on mutation induction in the mammalian germline”, Nat. Commun. 6:66-84. doi: 10.1038/ncomms7684.

Arlt MF, Wilson TE, Glover TW. (2012), “Replication stress and mechanisms of CNV formation”, Curr Opin Genet Dev. 22(3):204-10. doi: 10.1016/j.gde.2012.01.009.

Arlt, MF. Et al. (2014), “Copy number variants are produced in response to low-dose ionizing radiation in cultured cells”, Environ Mol Mutagen. 55(2):103-13. doi: 10.1002/em.21840.

Beaton, L. A. et al. (2013), “Investigating chromosome damage using fluorescent in situ hybridization to identify biomarkers of radiosensitivity in prostate cancer patients”, Int J Radiat Biol. 89(12): 1087-1093. doi:10.3109/09553002.2013.825060

Boei, J.J., Vermeulen, S., Natarajan, A.T. (1996), “Detection of chromosomal aberrations by fluorescence in situ hybridization in the first three postirradiation divisions of human lymphocytes”, Mutat Res, 349:127-135. Doi: 10.1016/0027-5107(95)00171-9.

Bonassi, S. (2008),”Chromosomal aberration frequency in lymphocytes predicts the risk of cancer: results from a pooled cohort study of 22 358 subjects in 11 countries”, Carcinogenesis. 29(6):1178-83. doi: 10.1093/carcin/bgn075.

Bryce SM, Bemis JC, Avlasevich SL, Dertinger SD. In vitro micronucleus assay scored by flow cytometry provides a comprehensive evaluation of cytogenetic damage and cytotoxicity. Mutat Res. 2007 Jun 15;630(1-2):78-91. doi: 10.1016/j.mrgentox.2007.03.002. Epub 2007 Mar 19. PMID: 17434794; PMCID: PMC1950716.

Bryce, S. et al. (2014), “Interpreting In VitroMicronucleus Positive Results: Simple Biomarker Matrix Discriminates Clastogens, Aneugens, and Misleading Positive Agents”, Environ Mol Mutagen, 55:542-555. Doi:10.1002/em.21868.

Bryce, S. et al.(2016), “Genotoxic mode of action predictions from a multiplexed flow cytometric assay and a machine learning approach”, Environ Mol Mutagen, 57:171-189. Doi: 10.1002/em.21996.

Bryce SM, Bernacki DT, Smith-Roe SL, Witt KL, Bemis JC, Dertinger SD. Investigating the Generalizability of the MultiFlow ® DNA Damage Assay and Several Companion Machine Learning Models With a Set of 103 Diverse Test Chemicals. Toxicol Sci. 2018 Mar 1;162(1):146-166. doi: 10.1093/toxsci/kfx235. PMID: 29106658; PMCID: PMC6059150.

Bunting, S. F., & Nussenzweig, A. (2013), “End-joining, translocations and cancer”, Nature Reviews Cancer.13 (7): 443-454. doi:10.1038/nrc3537

Curtis, C. et al. (2012), “The genomic and transcriptomic architecture of 2,000 breast tumours reveals novel subgroups”, Nature. 486(7403):346-52. doi: 10.1038/nature10983.

Dertinger, S.D. et al.(2004), “Three-color labeling method for flow cytometric measurement of cytogenetic damage in rodent and human blood”, Environ Mol Mutagen, 44:427-435. Doi: 10.1002/em.20075.

El-Zein, RA. Et al. (2014), “The cytokinesis-blocked micronucleus assay as a strong predictor of lung cancer: extension of a lung cancer risk prediction model”, Cancer Epidemiol Biomarkers Prev. 23(11):2462-70. doi: 10.1158/1055-9965.EPI-14-0462.

Fenech, M. (2000), “The in vitro micronucleus technique”, Mutation Research. 455(1-2), 81-95. Doi: 10.1016/s0027-5107(00)00065-8

Fischer, M., Bauer, T., Oberthür, A., Hero, B., Theissen, J., Ehrich, M., ... & Berthold, F. (2010). Integrated genomic profiling identifies two distinct molecular subtypes with divergent outcome in neuroblastoma with loss of chromosome 11q. Oncogene, 29(6), 865-875.

Griffiths, A. J. F., Miller, J. H., & Suzuki, D. T. (2000), “An Introduction to Genetic Analysis”, 7th edition. New York: W. H. Freeman. Available from: https://www.ncbi.nlm.nih.gov/books/NBK21766/

Hagmar, L. et al. (2004), “Impact of types of lymphocyte chromosomal aberrations on human cancer risk: results from Nordic and Italian cohorts”, Cancer Res. 64(6):2258-63.

Hastings PJ, Ira G & Lupski JR. (2009), “A microhomology-mediated break-induced replication model for the origin of human copy number variation”. PLoS Genet. 2009 Jan;5(1): e1000327. doi: 10.1371/journal.pgen.1000327.

Hendriks, G. et al. (2012), “The ToxTracker assay: novel GFP reporter systems that provide mechanistic insight into the genotoxic properties of chemicals”, Toxicol Sci, 125:285-298. Doi: 10.1093/toxsci/kfr281.

Hendriks, G. et al. (2016), “The Extended ToxTracker Assay Discriminates Between Induction of DNA Damage, Oxidative Stress, and Protein Misfolding”, Toxicol Sci, 150:190-203. Doi: 10.1093/toxsci/kfv323.

Keren, B. (2014),”The advantages of SNP arrays over CGH arrays”, Molecular Cytogenetics.7( 1):I31. Doi: 10.1186/1755-8166-7-S1-I31.

Khoury, L., Zalko, D., Audebert, M. (2016), “Evaluation of four human cell lines with distinct biotransformation properties for genotoxic screening”, Mutagenesis. 31:83-96. Doi: 10.1093/mutage/gev058.

Khoury, L., Zalko, D., Audebert, M. (2013), “Validation of high-throughput genotoxicity assay screening using cH2AX in-cell Western assay on HepG2 cells”, Environ Mol Mutagen, 54:737-746. Doi: 10.1002/em.21817.

Lee JA, Carvalho CM, Lupski JR. (2007). “Replication mechanism for generating nonrecurrent rearrangements associated with genomic disorders”, Cell. 131(7):1235-47. Doi: 10.1016/j.cell.2007.11.037.

Liu B. et al. (2013). “Computational methods for detecting copy number variations in cancer genome using next generation sequencing: principles and challenges”, Oncotarget. 4(11):1868-81. Doi: 10.18632/oncotarget.1537.

Mitelman, F. (1982), “Application of cytogenetic methods to analysis of etiologic factors in carcinogenesis”, IARC Sci Publ, 39:481-496.

Mukherjee. S. et al. (2017), “Addition of chromosomal microarray and next generation sequencing to FISH and classical cytogenetics enhances genomic profiling of myeloid malignancies, Cancer Genet. 216-217:128-141. doi: 10.1016/j.cancergen.2017.07.010.

Obe, G. et al. (2002), “Chromosomal Aberrations: formation, Identification, and Distribution”, Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis. 504(1-2), 17-36. Doi: 10.1016/s0027-5107(02)00076-3.

Savage, J.R. (1976), “Classification and relationships of induced chromosomal structual changes”, J Med Genet, 13:103-122. Doi: 10.1136/jmg.13.2.103.

Shahane SA, Nishihara K, Xia M. High-Throughput and High-Content Micronucleus Assay in CHO-K1 Cells. Methods Mol Biol. 2016;1473:77-85. doi: 10.1007/978-1-4939-6346-1_9. PMID: 27518626.

OECD (2016a), Test No. 487: In Vitro Mammalian Cell Micronucleus Test, OECD Guidelines for the Testing of Chemicals, Section 4, OECD Publishing, Paris, https://doi.org/10.1787/9789264264861-en.

OECD. (2016b), “Test No. 474: Mammalian Erythrocyte Micronucleus Test. OECD Guideline for the Testing of Chemicals, Section 4.”Paris: OECD Publishing.

OECD. (2016c), “In Vitro Mammalian Chromosomal Aberration Test 473.”

OECD. (2016d). Test No. 475: Mammalian Bone Marrow Chromosomal Aberration Test. OECD Guideline for the Testing of Chemicals, Section 4. Paris: OECD Publishing.

OECD. (2016e). Test No. 483: Mammalian Spermatogonial Chromosomal Aberration Test. Paris: OECD Publishing.

Pathak, R., Koturbash, I., & Hauer-Jensen, M. (2017), “Detection of Inter-chromosomal Stable Aberrations by Multiple Fluorescence In Situ Hybridization (mFISH) and Spectral Karyotyping (SKY) in Irradiated Mice”, J Vis Exp(119). doi:10.3791/55162.

Redon, R. et al. (2006), “Global variation in copy number in the human genome”, Nature. 444(7118):444-54. 10.1038/nature05329.

Rodrigues, M. A., Beaton-Green, L. A., & Wilkins, R. C. (2016), “Validation of the Cytokinesis-block Micronucleus Assay Using Imaging Flow Cytometry for High Throughput Radiation Biodosimetry”, Health Phys. 110(1): 29-36. doi:10.1097/HP.0000000000000371

Shahane S, Nishihara K, Xia M. (2016), “High-Throughput and High-Content Micronucleus Assay in CHO-K1 Cells”, In: Zhu H, Xia M, editors. High-Throughput Screening Assays in Toxicology. New York, NY: Humana Press. p 77-85.

Shen.TW, (2016),”Concurrent detection of targeted copy number variants and mutations using a myeloid malignancy next generation sequencing panel allows comprehensive genetic analysis using a single testing strategy”, Br J Haematol. 173(1):49-58. doi: 10.1111/bjh.13921.

Shlien A, Malkin D. (2009), “Copy number variations and cancer”, Genome Med. 1(6):62. doi: 10.1186/gm62.

Tucker, J.D., Preston, R.J. (1996), “Chromosome aberrations, micronuclei, aneuploidy, sister chromatid exchanges, and cancer risk assessment”, Mutat Res, 365:147-159.

Wilson, TE. et al. (2015), “Large transcription units unify copy number variants and common fragile sites arising under replication stress”, Genome Res. 25(2):189-200. doi: 10.1101/gr.177121.114.

Wink, S. et al. (2014), “Quantitative high content imaging of cellular adaptive stress response pathways in toxicity for chemical safety assessment”, Chem Res Toxicol, 27:338-355.

Zhang N, Wang M, Zhang P, Huang T. 2016. Classification of cancers based on copy number variation landscapes. Biochim Biophys Acta. 1860(11 Pt B):2750-5. doi: 10.1016/j.bbagen.2016.06.003.