A Simple, Sensitive, and Generalizable Plate Assay for Screening PARP Inhibitors
Poly-ADP-ribose polymerases (also known as ADP-ribosyltransferases or ARTDs) are a family of 17 enzymes in humans that catalyze the reversible posttranslational modification known as ADP-ribosylation. PARPs are implicated in diverse cellular processes, from DNA repair to the unfolded protein response. Small-molecule inhibitors of PARPs have improved our understanding of PARP-mediated biology and, in some cases, have emerged as promising treatments for cancers and other human diseases. However these advancements are hindered, in part, by a poor understanding of inhibitor selectivity across the PARP family. Here, we describe a simple, sensitive, and generalizable plate assay to test the potency and selectivity of small molecules against several PARP enzymes in vitro. In principle, this assay can be extended to all active PARPs, providing a convenient and direct comparison of inhibitors across the entire PARP enzyme family.
Key wordsADP-ribosylation PARPs ARTDs Small-molecule inhibitor Screen Click chemistry 6-a-NAD+
We thank members of the Cohen lab for many helpful discussions. We thank H. Schuler for the construct for PARP14wwe-cat. We thank J. Pascal (Université de Montréal) for helpful discussions regarding PARP3 enzyme activity. This work was funded by the NIH (NIH 1R01NS088629) and a grant from the Pew Charitable Trust (M.S.C.).
- 4.Venkannagari H, Verheugd P, Koivunen J et al (2016) Small-molecule chemical probe rescues cells from mono-ADP-ribosyltransferase ARTD10/PARP10-induced apoptosis and sensitizes cancer cells to DNA damage. Cell Chem Biol 23:1251–1260. https://doi.org/10.1016/j.chembiol.2016.08.012 CrossRefPubMedGoogle Scholar