HTS-Compatible CometChip Enables Genetic Screening for Modulators of Apoptosis and DNA Double-Strand Break Repair.

Dysfunction of apoptosis and DNA damage response pathways often drive cancer, and so a better understanding of these pathways can contribute to new cancer therapeutic strategies. Diverse discovery approaches have identified many apoptosis regulators, DNA damage response, and DNA damage repair proteins; however, many of these approaches rely on indirect detection of DNA damage. Here, we describe a novel discovery platform based on the comet assay that leverages previous technical advances in assay precision by incorporating high-throughput robotics. The high-throughput screening (HTS) CometChip is the first high-throughput-compatible assay that can directly detect physical damage in DNA. We focused on DNA double-strand breaks (DSBs) and utilized our HTS CometChip technology to perform a first-of-its-kind screen using an shRNA library targeting 2564 cancer-relevant genes. Conditions of the assay enable detection of DNA fragmentation from both exogenous (ionizing radiation) and endogenous (apoptosis) sources. Using this approach, we identified LATS2 as a novel DNA repair factor as well as a modulator of apoptosis. We conclude that the HTS CometChip is an effective assay for HTS to identify modulators of physical DNA damage and repair.

Duke Scholars

Author Scott Richard Floyd Radiation Oncology