Anti-cancer drugs of the DNA intercalator class, which includes doxorubicin (adriamycin), are among the most effective and widely used agents in systemic cancer chemotherapy. Numerous mechanisms have been proposed to be responsible for intercalator-mediated cytotoxicity, including inhibition of DNA topoisomerase II and DNA replication, yet none of these can completely account for the myriad effects these agents have seen shown to exert on cells. Our incomplete understanding of the molecular mechanism of action of DNA intercalators prevents them from being used as effectively as possible. Epigenetic genome modifications, in particular DNA methylation, have now been recognized as major regulators of gene activity and genomic stability. Indeed, the DNA methylation machinery and cellular DNA methylation patterns are disrupted in nearly all tumor cells. Emerging evidence indicates that chemotherapeutic drugs can alter DNA methylation patterns. How anti-cancer drugs such as DNA intercalators alter the efficacy of the DNA methylation machinery - and vice versa - has been largely unexplored. The central hypothesis we plan to test in this application is that inhibition of DNA methyltransferases (DNMTs) by DNA intercalating agents leads to alterations in genomic DNA methylation patterns and contributes directly to intercalator-mediated cell growth inhibition and apoptosis of human cancer cells. The three aims of this proposal are designed to test this hypothesis.
In aim 1, the inhibition properties of a group of DNA intercalating agents on the major DNMTs will be characterized in vitro using both DNA and chromatin substrates.
Our second aim entails characterizing the effects of DNA intercalating agents on both DNA methylation patterns (global and gene-specific) and the DNMTs in vivo in tumor cell lines. Lastly, we aim to determine whether cellular DNA methyltransferase levels influence the capacity of DNA intercalating drugs to induce cell cycle arrest and apoptosis in human tumor cell lines. Addressing these questions will advance our understanding of how DNA intercalating agents exert their powerful anticancer effects and how their inhibition of DNA methylation affects this process. This is expected to positively affect human health by allowing for the development of new therapies that maximize the anti-tumor properties of DNA intercalating agents and minimize their side-effects. Cancer is one of the leading causes of death in the United States and chemotherapy is frequently employed to treat cancer patients, however the success rate is variable and our current repertoire of drugs is limited. It is therefore critical to understand how these drugs function to kill tumor cells and determine how they can be used more effectively. Elucidating the molecular mechanism of a widely utilized class of anticancer drugs known as DNA intercalators is the focus of this proposal.
