The specific inhibitory effect of demethoxydehydroaltenusin, a derivative of dehydroaltenusin, on mammalian DNA polymerase α

Abstract

Abstract. In the screening of selective inhibitors of eukaryotic DNA polymerases (pols), dehydroaltenusin from the fungus Acremonium sp. was found to be an inhibitor of pol α. The present study succeeded in chemically synthesizing dehydroaltenusin, and the compound strongly inhibited calf pol α activity and weakly suppressed rat pol ß activity, with IC 50 values of 0.68 and 64 μM, respectively. We purified or synthesized various slightly modified derivatives of dehydroaltenusin, and using these, investigated the relationship between chemical structure and the inhibitory effects. These results suggest that the ketone group at the 5'-position in dehydroaltenusin is essential for pol inhibitory activity, and the group at the 5-position is important for the specificity of pol α inhibition. Demethoxydehydroaltenusin was found to be the most specific pol α inhibitor among the prepared derivatives, and the IC 50 values for pols α and ß were 0.24 and 89 μM, respectively. This compound did not influence the activities of other replicative pols such as pols δ and ε, and also demonstrated no effect on pol α activity from another vertebrate, fish and a plant species. Demethoxydehydroaltenusin also had no influence on the other pols and DNA metabolic enzymes tested. Therefore, demethoxydehydroaltenusin is of interest as a mammalian pol α-selective inhibitor as a 'chemical knockout agent' in vitro and in vivo. IntroductionThe human genome encodes 16 DNA polymerases (pols) that control cellular DNA synthesis (1). Eukaryotic cells reportedly contain three replicative types: pols α, δ and ε, mitochondrial pol γ, and at least twelve repair types: pols ß, δ, ε, ζ, η, θ, ι, κ, λ, μ, and σ and REV1 (2). Selective inhibitors of each pol are useful tools and molecular probes for distinguishing pols and for clarifying their biological and in vivo functions (3). For example, aphidicolin is a selective inhibitor of both pol α and eukaryotic DNA replicative pols δ and ε, indicating that these pols are essential for DNA replication (4), and this inhibitor has been very useful for studying the DNA replication system (5); however, aphidicolin is not capable of distinguishing pols α, δ and ε.Therefore, we established an assay method to detect pol inhibitors, and have screened for natural sources of inhibitors for more than 10 years. An inhibitor was isolated that selectively influences the activity of mammalian pol α, dehydroaltenusin, from a fungus (Acremonium sp.) collected from fields in the vicinity of Noda city in Chiba prefecture, Japan (6). A total chemical synthesis method was established for dehydroaltenusin and succeeded in completely synthesizing the compound (7-9).Subsequently, the slightly modified derivatives were purified or chemically synthesized to examine the structural relationship between dehydroaltenusin and pols. Dehydroaltenusin and its analogues represent a group of potentially useful agents to examine the precise role of each pol in vivo, and to develop a drug design strategy for cancer chemotherapy agents. The compou...