Novel high-throughput deoxyribonuclease 1 assay.
Dae Song Jang1, Narsimha R Penthala2, Eugene O Apostolov1
1Department of Pharmacology & Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR, USA.
Journal of Biomolecular Screening
|October 19, 2014
Summary
Researchers developed a novel high-throughput assay to find deoxyribonuclease I (DNase I) inhibitors. This assay identified compounds that protect kidney cells from toxic injury, paving the way for new therapeutic strategies.
Area of Science:
- Biochemistry
- Molecular Biology
- Pharmacology
Background:
- Deoxyribonuclease I (DNase I) is a key endonuclease involved in cellular damage from toxic, hypoxic, and radiation injuries.
- The absence of high-throughput screening methods and DNase I inhibitors hinders research into its role in cellular injury.
Purpose of the Study:
- To develop a high-throughput assay for identifying DNase I inhibitors.
- To screen chemical libraries for potential DNase I inhibitors.
- To evaluate the therapeutic potential of identified inhibitors against cellular damage.
Main Methods:
- Developed a 96-well plate-based assay utilizing fluorescence intensity changes upon oligonucleotide degradation by DNase I.
- Screened a chemical library using the developed high-throughput assay.
- Validated hit compounds in vitro for their protective effects against cisplatin-induced kidney cell death.
Main Results:
- The DNase I assay demonstrated high sensitivity, reliability (Z' ≥ 0.5), and low variability.
- Screening identified several potential DNase I inhibitors.
- Two selected hit compounds effectively protected kidney cells from cisplatin-induced death in vitro.
Conclusions:
- The developed high-throughput DNase I assay is suitable for screening chemical libraries and identifying potent inhibitors.
- Identified DNase I inhibitors show promise in protecting cells from toxic injury, suggesting therapeutic applications.
- The assay may be adaptable for identifying inhibitors of other endonucleases.
