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Updated: Jun 3, 2026
A High-Throughput Comet Assay Approach for Assessing Cellular DNA Damage
Published on: May 10, 2022
Measurement of drug-induced DNA interstrand crosslinking using the single-cell gel electrophoresis (comet) assay
V J Spanswick1, J M Hartley, T H Ward
1CRC Drug-DNA Interactions Research Group, Department of Oncology, University College London Medical School, London, UK.
Abstract:
DNA damaging agents have been widely used in cancer chemotherapy for many years and have proved successful in the treatment of both solid tissue and haematological malignancies. Many commonly used clinical agents, such as members of the nitrogen mustard, chloroethylnitrosourea, dimethane-sulphonate and platinum classes, are bifunctional. DNA interstrand crosslinks (ISC) formed in cells are clearly critical cytotoxic lesions and the formation of DNA ISC has been shown to correlate with cytotoxicity in vitro (1-5). Acquired resistance in vitro to such agents can occur by a number of mechanisms, for example altered drug transport (6), intracellular detoxification via enhanced glutathione and glutathione-S-transferase activity (7), but enhanced DNA repair capacity can also play an important role (3). Clinically the mechanisms of acquired resistance to DNA damaging agents are less clear but enhanced repair of ISC has been suggested to play a role in the acquired resistance of some cancers, e.g., chronic lymphocytic leukaemia to nitrogen mustards (8). In addition, the inherent sensitivity (and curability) of some tumors, e.g., testicular cancer, to DNA damaging agents may result in part from their inability to repair critical DNA lesions (9).
Insights
DNA damaging agents are crucial in cancer chemotherapy, forming DNA interstrand crosslinks (ISC) that are cytotoxic. Enhanced DNA repair capacity can lead to acquired resistance, impacting treatment efficacy in various malignancies.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- DNA damaging agents are mainstays in cancer chemotherapy for solid and hematological malignancies.
- Bifunctional agents, like nitrogen mustards and platinum compounds, induce DNA interstrand crosslinks (ISC).
- DNA ISC are critical cytotoxic lesions, correlating with cell death in vitro.
Purpose of the Study:
- To explore the role of DNA interstrand crosslinks (ISC) in chemotherapy efficacy.
- To investigate mechanisms of acquired resistance to DNA damaging agents.
- To understand the contribution of DNA repair capacity to drug resistance and sensitivity.
Main Methods:
- Review of existing literature on DNA damaging agents and cancer chemotherapy.
- Analysis of in vitro studies correlating DNA ISC formation with cytotoxicity.
- Examination of proposed mechanisms of acquired resistance, including drug transport, detoxification, and DNA repair.
Main Results:
- DNA interstrand crosslinks (ISC) are critical cytotoxic lesions induced by common chemotherapy agents.
- Acquired resistance can involve altered drug transport, enhanced detoxification (glutathione/glutathione-S-transferase), and increased DNA repair.
- Enhanced repair of ISC is implicated in clinical resistance (e.g., chronic lymphocytic leukemia) and inability to repair ISC may contribute to tumor sensitivity (e.g., testicular cancer).
Conclusions:
- DNA interstrand crosslinks (ISC) are key cytotoxic lesions in cancer chemotherapy.
- Enhanced DNA repair capacity is a significant mechanism of acquired resistance to DNA damaging agents.
- Understanding DNA repair is crucial for optimizing cancer treatment strategies and overcoming drug resistance.
