Drug Discovery 2022: driving the next life science revolution

A sensitive, homogeneous, and high-throughput assay for homologous recombination DNA repair (HRR) modulators using the engineered luminescent enzyme


R Lubis1
1 Lead Discovery Center, Germany


Targeting and deterring DNA damage repair using small molecules is considered a valuable strategy to increase the effect of chemotherapy or ionizing radiation when treating cancer. Homologous recombination repair (HRR) is used in human cells to repair DNA double-strand breaks. A traditional method to study HRR events consists of inducing DNA damage with an endonuclease, and using green fluorescent protein (GFP) as a reporter. However, this method is not easily adaptable to a high-throughput system due to its numerous, time-consuming sample separation and washing steps.

We established a novel, sensitive, homogeneous and high-throughput cell-based assay to quantify the levels of HRR using an engineered nano luciferase enzyme. This method showed a higher sensitivity to DNA damage repair, with an approximate 31-fold signal upon DNA damage induction, compared to the GFP-based method, which produced a 4-fold signal. This gain in sensitivity enables the detection of HRR event modulators, promoters or inhibitors. Furthermore, the assay takes less than 5 minutes to complete in a homogeneous “add and read” format, and it has been successfully used to monitor RecQ-like helicases involved in the activation of cellular DNA repairpathways (e.g. suppressing and promoting HRR events). This novel method has been validated for the study of DNA helicases and proved suitable for inhibitor profiling and high-throughput screening.

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