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Published on: March 31, 2022
Discovery of novel DNA-damaging agents through phenotypic screening for DNA double-strand break
Doudou Zhang1, Takashi Shimokawa2, Qianqian Guo1,3
1Department of Molecular Genetics, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan.
Abstract:
DNA double-strand breaks (DSBs) seriously damage DNA and promote genomic instability that can lead to cell death. They are the source of conditions such as carcinogenesis and aging, but also have important applications in cancer therapy. Therefore, rapid detection and quantification of DSBs in cells are necessary for identifying carcinogenic and anticancer factors. In this study, we detected DSBs using a flow cytometry-based high-throughput method to analyze γH2AX intensity. We screened a chemical library containing 9600 compounds and detected multiple DNA-damaging compounds, although we could not identify mechanisms of action through this procedure. Thus, we also profiled a representative compound with the highest DSB potential, DNA-damaging agent-1 (DDA-1), using a bioinformatics-based method we termed "molecular profiling." Prediction and verification analysis revealed DDA-1 as a potential inhibitor of topoisomerase IIα, different from known inhibitors such as etoposide and doxorubicin. Additional investigation of DDA-1 analogs and xenograft models suggested that DDA-1 is a potential anticancer drug. In conclusion, our findings established that combining high-throughput DSB detection and molecular profiling to undertake phenotypic analysis is a viable method for efficient identification of novel DNA-damaging compounds for clinical applications.
Insights
This study developed a high-throughput method to detect DNA double-strand breaks (DSBs) and identified a novel anticancer drug candidate. The approach combines flow cytometry and bioinformatics for efficient discovery of DNA-damaging compounds.
Area of Science:
- Molecular Biology
- Genetics
- Pharmacology
Background:
- DNA double-strand breaks (DSBs) are critical DNA lesions linked to genomic instability, aging, and cancer.
- Accurate detection of DSBs is vital for identifying carcinogenic and anticancer agents.
- DSBs are also therapeutically relevant in cancer treatment strategies.
Purpose of the Study:
- To establish a high-throughput method for detecting and quantifying DSBs in cells.
- To screen a chemical library for novel DNA-damaging compounds.
- To identify and characterize a potential anticancer drug candidate.
Main Methods:
- Utilized flow cytometry to analyze γH2AX intensity for DSB detection.
- Screened a 9600-compound chemical library for DNA-damaging activity.
- Employed a bioinformatics approach termed "molecular profiling" for compound mechanism analysis.
Main Results:
- Identified multiple DNA-damaging compounds from the chemical library.
- Characterized DNA-damaging agent-1 (DDA-1) as a potential topoisomerase IIα inhibitor.
- DDA-1 and its analogs showed promise as anticancer agents in preclinical models.
Conclusions:
- Combined high-throughput DSB detection and molecular profiling offer an effective strategy for identifying novel DNA-damaging compounds.
- This integrated approach facilitates phenotypic analysis for drug discovery.
- The findings support the potential clinical application of DDA-1 as an anticancer therapeutic.
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