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

ABSTRACT: Human DNA polymerase kappa (pol ?) 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 ? TLS activity can reduce the cellular toxicity of chemotherapeutic agents and since gliomas overexpress pol ?, small molecule library screens targeting pol ? 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 ? by primer extension assays using DNAs containing a site-specific acrolein-derived, ring-opened reduced form of ?-HOPdG. Furthermore, candesartan cilexetil could enhance ultraviolet light-induced cytotoxicity in xeroderma pigmentosum variant cells, suggesting its inhibitory effect against intracellular pol ?. In summary, this investigation represents the first high-throughput screening designed to identify inhibitors of pol ?, with the characterization of biochemical and biologically relevant endpoints as a consequence of pol ? inhibition. These approaches lay the foundation for the future discovery of compounds that can be applied to combination chemotherapy.

SUBMITTER: Yamanaka K 

PROVIDER: S-EPMC3466269 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature