High-Throughput PARP Inhibitor Screening with a Mix-and-Read Biochemical Assay

Описание: Poly(ADP-ribose) polymerases (PARPs) play a central role in DNA damage repair by binding to DNA double-strand breaks and generating poly(ADP-ribose) (PAR) chains that act as a scaffold for recruitment of DNA repair proteins. The clinical success of PARP inhibitors—first demonstrated in BRCA1-mutant ovarian cancer through the concept of synthetic lethality—has expanded their use to breast, prostate, pancreatic, and ovarian cancers, making PARP enzymes high-value drug discovery targets.

PARP enzymes use NAD⁺ as a donor to sequentially add ADP-ribose units to DNA-associated proteins such as histones, as well as to PARP itself. Discovering new PARP inhibitors requires robust, automation-friendly biochemical assays. However, many existing PARP assays rely on complex workflows, modified NAD substrates, or multiple binding proteins, increasing cost and limiting scalability for high-throughput screening (HTS).

In this video, we introduce a high-throughput, mix-and-read Transcreener® poly-ADPR PARP assay that enables direct, quantitative detection of poly(ADP-ribose) formation. The assay works by enzymatically converting poly(ADP-ribose) into AMP, which is then detected using the Transcreener AMP/GMP assay with either fluorescence polarization (FP) or time-resolved FRET (TR-FRET) readouts. This homogeneous format integrates seamlessly into automated HTS workflows.

The assay delivers exceptional sensitivity, detecting PARP activity at sub-nanomolar enzyme concentrations. Because each poly(ADP-ribose) chain is converted into multiple AMP molecules, the signal is amplified, providing greater sensitivity than traditional poly-ADPR detection methods. Raw data can be directly converted into AMP product formation, enabling accurate, quantitative measurements of enzyme activity and reliable IC₅₀ determination—even for highly potent inhibitors.

Fully validated for HTS, the Transcreener poly-ADPR PARP assay consistently delivers Z′ values greater than 0.8, with low compound interference and strong confirmation of hits in dose-response studies. The assay supports inhibitor screening, mechanistic studies, and lead optimization with confidence.

For a complete PARP biochemical screening solution, the assay is designed to be used with BellBrook Labs’ Enzolution™ PARP1 and PARP2 Assay Systems, which include purified enzymes, NAD⁺, DNA substrate, and optimized buffers. Together, these tools provide an extensively validated, HTS-ready platform for discovering and characterizing next-generation PARP modulators.