Branched or lobed DNAs with single-stranded ends are formed during DNA replication and DNA repair in healthy individuals. These single-stranded flap DNAs can be excised by an enzyme such as FEN1 to repair the integrity of the double-stranded state of the DNA.
Higher levels of FEN1 have been observed in cancer cells with worse clinical outcomes, and these cells are more dependent on this repair enzyme in the absence of other supporting DNA damage response proteins. These cells are sensitive and will die when FEN1 is blocked - synthetically lethal. Preventing the action of FEN1 such enzymes may be a way to selectively kill these cancer cells.
An interdisciplinary team of scientists from the University of Sheffield and AstraZeneca Pharmaceuticals confirmed in detail the combination of inhibitors that block and block FEN1 to provide clues as to how "the function of the protein is blocked."
They present the first crystal structure of inhibitor-bound hFEN1 and show a cyclic N-hydroxyurea bound in the active site coordinated to two magnesium ions. Three such compounds had similar IC50 values but differed subtly in mode of action. One had comparable affinity for protein and protein–substrate complex and prevented reaction by binding to active site catalytic metal ions, blocking the unpairing of substrate DNA necessary for reaction. Other compounds were more competitive with substrate. Cellular thermal shift data showed engagement of both inhibitor types with hFEN1 in cells with activation of the DNA damage response evident upon treatment. However, cellular EC50s were significantly higher than in vitro inhibition constants.
The researchers said: "The results of this study have indeed deepened our understanding of how 'a group of specific compounds previously identified as FEN1 inhibitors work.' This insight can help us develop methods to block nucleases, nucleases are A family of important enzymes in cellular DNA damage response, AstraZeneca's strategic focus is on oncology drug development."
All in all, this international team of scientists has discovered how compounds block FEN1, which is critical for DNA damage response and cancer therapeutic targets.
EIAAB SCIENCE INC, WUHAN has developed FEN1 protein, antibody and ELISA kit.