Discovery of anticancer inhibitors of DNA polymerase

CD437 is a retinoid-like small molecule that selectively induces apoptosis in cancer but not normal cells through an unknown mechanism. We used a forward genetic strategy to discover mutations in POLA1 that coincide with CD437 resistance (POLA1R). Introduction of one of these mutations into cancer cells by CRISPR/Cas9 genome editing conferred CD437 resistance demonstrating causality. POLA1 encodes DNA polymerase α, the enzyme responsible for initiating DNA synthesis during the S phase of the cell cycle. CD437 inhibits DNA replication in cells and recombinant POLA1 activity in vitro. Both effects are abrogated by mutations associated with POLA1R. In addition, we detected an increase in the total fluorescence intensity and anisotropy of CD437 in the presence of increasing concentrations of POLA1 consistent with a direct binding interaction. The discovery of POLA1 as the direct anti-cancer target for CD437 has the potential to catalyze its development into an anti-cancer therapeutic.

There are therapeutic implications for CD437 as a newly discovered inhibitor of POLA1 and DNA replication. Many cancer drugs act by targeting any of a variety of aspects of DNA replication. Three classes of agents block DNA replication by influencing the template: anthracyclines through DNA intercalation, topoisomerase inhibitors through alteration of the supercoiled state of DNA, and alkylators through covalent modification of DNA. Other approved drugs inhibit replication by affecting nucleotides, which are essential for DNA synthesis. For instance, hydroxyurea, methotrexate, and pemetrexed reduce the overall level of nucleotides, and gemcitabine and cytarabine block replication by substituting for deoxycytidine. Finally, drugs that target enzymes that regulate DNA replication, such as inhibitors of cyclin-dependent kinases (CDK4 and CDK6), also have therapeutic efficacy. Surprisingly, despite the demonstrable clinical utility of anti-cancer therapies that target essential components of DNA replication, there are no currently available drugs that directly inhibit DNA polymerase.

A major goal in our laboratory is to build on our discovery and develop and test DNA polymerase alpha inhibitors in pre-clinical models of cancer. The goal of the project is to develop first-in-class DNA polymerase alpha for the treatment of cancer.