CREDENTIALING THE DNA METHYLATION MAINTENANCE PROTEIN UHRF1 AS A TARGET FOR ANTICANCER THERAPY
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Date
2014-10-01
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Johns Hopkins University
Abstract
Cancer is a disease of genome alterations that drive uncontrolled growth. The most challenging aspect of cancer treatment is that there are many avenues to enable proliferation within the cell, including alterations to epigenetic elements. DNA methylation—a methyl modification on the 5-carbon of cytosine bases—has been pinpointed to be one of these avenues, and as such is an option for targeted therapy. DNA methylation at the promoter regions of genes can silence their transcription, and normally-silenced genes that are hypomethylated can be re-expressed. Current approved cancer therapies targeting DNA methylation focus on inhibiting DNA methyltransferases, enzymes responsible for maintaining DNA methylation during division. These therapies, which are effective at reducing methylation levels on DNA globally and at tumor suppressor genes, come at a cost: they are nucleoside analogs that exhibit serious toxicities. The central hypothesis for this thesis is that another DNA methylation maintenance protein, ubiquitin-like containing PHD and RING finger domains 1 (UHRF1) is an attractive candidate for anticancer therapy. UHRF1 directly interacts with DNMT1 and is thought of as a “helper” molecule that recruits DNMTs to newly synthesized DNA. DNA is methylated symmetrically on both strands, but during replication new copies of DNA contain only one strand with the proper methylation information; this so-called hemimethylated state is the target for the Set- and RING-Associated (SRA) domain of UHRF1, which binds this hemimethylated DNA and allows UHRF1 to recruit the DNA methyltransferase machinery to maintain methylation on the newly synthesized strand. To address our hypothesis, we have developed a suite of molecular tools to study the impact of removing UHRF1 on cellular growth, DNA methylation patterns, gene expression and survivability in vitro. We have developed a mouse model for heterozygous deletion of UHRF1 in vivo. We have successfully implemented a UHRF1 inhibitor assay to use on small molecule libraries to identify potential inhibitors. We show that UHRF1 is overexpressed in cancer cell lines and tissue, and when knocked down, reduces growth and survival. We also demonstrate that UHRF1 is necessary for global and local methylation, and that methylation loss due to lower UHRF1 levels results in re-expression of silenced genes. Our mouse model of UHRF1 deletion shows that it is necessary for tumorigenesis in the APCmin model. Finally, we determine that anthracycline derivatives like mitoxantrone and doxorubicin interfere with UHRF1 binding DNA, decrease global methylation in cells and synergize with decitabine in cell killing. Synthesizing these results, we believe that UHRF1 is a viable target for cancer treatment, and effectively inhibiting it can enhance approved DNA methylation chemotherapies for better cancer control.
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Keywords
UHRF1, cancer, epigenetics, epigenetic therapy