Diverse Small Molecule Inhibitors of Human Apurinic/Apyrimidinic Endonuclease APE1 Identified from a Screen of a Large Public Collection

Abstract

The major human apurinic/apyrimidinic endonuclease APE1 plays a pivotal role in the repair of base damage via participation in the DNA base excision repair (BER) pathway. Increased activity of APE1, often observed in tumor cells, is thought to contribute to resistance to various anticancer drugs, whereas down-regulation of APE1 sensitizes cells to DNA damaging agents. Thus, inhibiting APE1 repair endonuclease function in cancer cells is considered a promising strategy to overcome therapeutic agent resistance. Despite ongoing efforts, inhibitors of APE1 with adequate drug-like properties have yet to be discovered. Using a kinetic fluorescence assay, we conducted a fully-automated high-throughput screen (HTS) of the NIH Molecular Libraries Small Molecule Repository (MLSMR), as well as additional public collections, with each compound tested as a 7-concentration series in a 4 µL reaction volume. Actives identified from the screen were subjected to a panel of confirmatory and counterscreen tests. Several active molecules were identified that inhibited APE1 in two independent assay formats and exhibited potentiation of the genotoxic effect of methyl methanesulfonate with a concomitant increase in AP sites, a hallmark of intracellular APE1 inhibition; a number of these chemotypes could be good starting points for further medicinal chemistry optimization. To our knowledge, this represents the largest-scale HTS to identify inhibitors of APE1, and provides a key first step in the development of novel agents targeting BER for cancer treatment.

Structure‐based virtual screening toward the discovery of novel inhibitors of the DNA repair activity of the human apurinic/apyrimidinic endonuclease 1

“…In fact, fluorescence‐based assays for the identification of small‐molecule inhibitors may be affected by the inherent fluorescence of a given compound. In this case, by adopting an assay with an APE1 substrate detected in the red‐shifted spectral region, this interference can be minimized …”

“…Lucanthone has also been regarded as a non‐specific direct inhibitor of APE1 as it also inhibits topoisomerase II and intercalates into the DNA . Therefore, several authors proposed other classes of APE1 inhibitors using different approaches, which include the HTS of commercially available chemical libraries, the rational design with structure–activity relationship studies and also ligand‐based pharmacophore models, structure‐based virtual screening (SBVS) and molecular docking studies . Nevertheless, there is still a need for additional studies to identify novel pharmacologically active APE1 inhibitors with clinical application as combination therapy in cancer treatment.…”

Role of apurinic/apyrimidinic nucleases in the regulation of homologous recombination in myeloma: mechanisms and translational significance

“…AP nuclease activity was assessed using a fluorescence based assay reported previously 51 . Briefly, the oligonucleotides carrying AP site mimic (Tetrahydrofuran; THF) and a fluorescein label on one strand and a quenching moiety (Dabcyl-Q) on other strand are synthesized commercially (Eurogentec Ltd., San Diego, CA).…”

DNA repair targeted therapy: The past or future of cancer treatment?

“…An in silico screening was used to focus on the endonuclease region of APE1 and resulted in the discovery of a series of APE1 inhibitors with varying potency (Mohammed et al, 2011). Similarly, a HTS approach was also used and resulted in the identification of a large number of hits that was subsequently culled to identify cellular activity (Abbotts et al, 2014; Dorjsuren et al, 2012). Srinivasan et.…”