A comprehensive strategy to discover inhibitors of the translesion synthesis DNA polymerase κ.

Yamanaka K, Dorjsuren D, Eoff RL, Egli M, Maloney DJ, Jadhav A, Simeonov A, Lloyd RS
PLoS One. 2012 7 (10): e45032

PMID: 23056190 · PMCID: PMC3466269 · DOI:10.1371/journal.pone.0045032

Human DNA polymerase kappa (pol κ) is a translesion synthesis (TLS) polymerase that catalyzes TLS past various minor groove lesions including N(2)-dG linked acrolein- and polycyclic aromatic hydrocarbon-derived adducts, as well as N(2)-dG DNA-DNA interstrand cross-links introduced by the chemotherapeutic agent mitomycin C. It also processes ultraviolet light-induced DNA lesions. Since pol κ TLS activity can reduce the cellular toxicity of chemotherapeutic agents and since gliomas overexpress pol κ, small molecule library screens targeting pol κ were conducted to initiate the first step in the development of new adjunct cancer therapeutics. A high-throughput, fluorescence-based DNA strand displacement assay was utilized to screen ∼16,000 bioactive compounds, and the 60 top hits were validated by primer extension assays using non-damaged DNAs. Candesartan cilexetil, manoalide, and MK-886 were selected as proof-of-principle compounds and further characterized for their specificity toward pol κ by primer extension assays using DNAs containing a site-specific acrolein-derived, ring-opened reduced form of γ-HOPdG. Furthermore, candesartan cilexetil could enhance ultraviolet light-induced cytotoxicity in xeroderma pigmentosum variant cells, suggesting its inhibitory effect against intracellular pol κ. In summary, this investigation represents the first high-throughput screening designed to identify inhibitors of pol κ, with the characterization of biochemical and biologically relevant endpoints as a consequence of pol κ inhibition. These approaches lay the foundation for the future discovery of compounds that can be applied to combination chemotherapy.

MeSH Terms (21)

Acrolein Benzimidazoles Biphenyl Compounds Cell Line, Transformed Cell Survival Deoxyguanosine DNA DNA-Directed DNA Polymerase DNA Adducts DNA Damage DNA Repair Dose-Response Relationship, Drug Drug Evaluation, Preclinical Enzyme Inhibitors Humans Indoles Nucleic Acid Synthesis Inhibitors Small Molecule Libraries Terpenes Tetrazoles Ultraviolet Rays

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