Cell-based assays for identification of novel double-strand break-inducing agents
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Updated: Sep 26, 2026
Analysis of DNA Double-strand Break (DSB) Repair in Mammalian Cells
Published on: September 8, 2010
Heather M Dunstan1, Catherine Ludlow, Sondra Goehle
1Program in Molecular Pharmacology, Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA.
Background:
We are developing cell-based assays to identify anticancer agents that are selectively toxic to cells with defined mutations. As a test, we used a three-stage strategy to screen compounds from the National Cancer Institute's repository for agents that are selectively toxic to double-strand break repair-deficient yeast cells.
Methods:
Compounds identified in the screen were further analyzed by use of yeast and vertebrate cell-based and in vitroassays to distinguish between topoisomerase I and II poisons.
Results:
Of the more than 85 000 compounds screened, 126 were selectively toxic to yeast deficient in DNA double-strand break repair. Eighty-seven of these 126 compounds were structurally related to known topoisomerase poisons, and 39 were not. Twenty-eight of the 39 were characterized, and we present data for eight of the compounds. Among these eight compounds, we identified two novel topoisomerase II poisons (NSC 327929 and NSC 638432) that were equipotent to etoposide in biochemical tests and in cells, five (NSC 63599, NSC 65601, NSC 380271, NSC 651646, and NSC 668370) with topoisomerase I-dependent toxicity in yeast that induced DNA damage and toxicity in mammalian cells, and one (NSC 610898) that directly bound to DNA and induced strand breaks.
Conclusions:
Cell-based assays can be used to identify molecules that are selectively toxic to cells with a predetermined genetic background, including mutations in genes involved in the cell cycle and its checkpoints, for which there are currently no selectively toxic compounds.
Insights
This study screened over 85,000 compounds to find anticancer agents selectively toxic to DNA repair-deficient cells. Researchers identified novel topoisomerase poisons with potential for targeted cancer therapy.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Developing cell-based assays for targeted anticancer agent discovery.
- Utilizing a screening strategy for compounds selectively toxic to DNA double-strand break repair-deficient cells.
Purpose of the Study:
- To identify novel anticancer agents with selective toxicity against specific cellular mutations.
- To screen the National Cancer Institute's compound repository using a three-stage strategy.
Main Methods:
- Screening over 85,000 compounds for selective toxicity in DNA repair-deficient yeast.
- Analyzing identified compounds using yeast and vertebrate cell-based assays.
- Distinguishing between topoisomerase I and II poisons through in vitro assays.
Main Results:
- 126 compounds showed selective toxicity to DNA double-strand break repair-deficient yeast.
- Identified two novel topoisomerase II poisons (NSC 327929, NSC 638432) equipotent to etoposide.
- Discovered five compounds with topoisomerase I-dependent toxicity and one DNA-binding agent.
Conclusions:
- Cell-based assays effectively identify molecules selectively toxic to cells with specific genetic mutations.
- This approach enables the discovery of targeted therapies for cancers with defects in cell cycle and checkpoint genes.
