Many human diseases are caused by dysregulated gene expression. The oversupply or over activity of one or more transcription factors may be required for the survival, growth and metastatic behavior of all human cancers. Recent technological advances have spurred a revolution of in-depth analytical methods available for determining transcription factor-DNA interactions on a genome-wide scale;most notably through the coupling of chromatin immunoprecipitation to massively parallel sequencing (ChIP-Seq). Such genome-wide analyses are now capable of routinely identifying thousands of transcription factor binding sites in a single experiment. However, there remains a large gap between the prolific generation of genome-wide protein-DNA binding data, and the paucity of methods to functionally probe these binding events in living cells. No technology exists that would allow the selective inhibition of subsets of DNA-binding and transcription activation events. The central hypothesis of this research program is that cell permeable small molecules which can be programmed to bind a broad repertoire of DNA sequences can disrupt transcription factor-DNA interfaces and modulate aberrant gene expression pathways. The Dervan laboratory has pioneered the development of Py-Im polyamides as programmable oligomers for targeting double-strand DNA. Minor groove DNA binding polyamides containing aromatic amino acids form the basis of a modular code to control sequence specificity. These small molecules achieve affinities and specificities of DNA binding proteins, inhibit DNA binding of a broad range of transcription factors, localize to the cell nucleus, bind to chromatin, and down regulate endogenous gene expression in cell culture.
Two specific aims are focused on transcription factors important in human disease, hypoxia inducible factor (HIF-11) and glucocorticoid receptor (GR). HIF is a principle transcriptional activator stabilized in response to low oxygen (hypoxia) whose aberrant regulation is associated with cancer invasion and metathesis. The DNA sequence of the hypoxic response element is highly conserved with variability in the 5'nucleotide. The hypothesis is that targeting single-nucleotide variations in the canonical HRE will allow genomic perturbation of subsets of HIF occupied sites and result in distinct gene expression profiles, thereby providing insight into both Py-Im polyamide mechanism as well as the role of single nucleotide variations in the hypoxic response. Glucocorticoid receptor (GR) is a nuclear hormone receptor that modulates the inflammatory response through a variety of mechanisms, including DNA binding of glucocorticoid response elements (GREs). These GREs are composed of distinct conserved and variable half sites, often located distal relative to target genes. The hypothesis is that the Py-Im polyamides designed to target a subset of variable GR half-sites may be used to link candidate GREs to the expression of GR-regulated genes, and provide a new method for connecting distal protein-DNA binding events to their downstream transcriptional responses. The approach is highly innovative because it represents a unique departure from protein-directed and siRNA based methods of transcription factor inhibition by allowing direct antagonization of the protein-DNA binding event in living cells. The proposed research is significant because it represents the only known chemical method for specifically disrupting the protein-DNA interface in a genetically unmodified cell.
Cancer cells have a different and pathological transcriptional pattern compared with normal cells from which they originated. A number of transcription factors are overactive in most human cancers. Cell permeable DNA binding small molecules can be programmed to bind a broad repertoire of DNA sequences. Selective inhibition of transcription activity could be achieved by disrupting transcription factor-DNA binding interfaces. The proposed research is relevant to public health because it provides a deeper mechanistic understanding of dysregulated gene networks underpinning human diseases. Thus the proposed research is relevant to NIH's mission that pertains to developing fundamental knowledge that will create new therapeutic targets for alleviating disease.
|Mysore, Veena S; Szablowski, Jerzy; Dervan, Peter B et al. (2016) A DNA-binding Molecule Targeting the Adaptive Hypoxic Response in Multiple Myeloma Has Potent Antitumor Activity. Mol Cancer Res 14:253-66|
|Kang, JeenJoo S; Dervan, Peter B (2015) A sequence-specific DNA binding small molecule triggers the release of immunogenic signals and phagocytosis in a model of B-cell lymphoma. Q Rev Biophys 48:453-64|
|Kang, JeenJoo S; Meier, Jordan L; Dervan, Peter B (2014) Design of sequence-specific DNA binding molecules for DNA methyltransferase inhibition. J Am Chem Soc 136:3687-94|
|Raskatov, Jevgenij A; Szablowski, Jerzy O; Dervan, Peter B (2014) Tumor xenograft uptake of a pyrrole-imidazole (Py-Im) polyamide varies as a function of cell line grafted. J Med Chem 57:8471-6|
|Martínez, Thomas F; Phillips, John W; Karanja, Kenneth K et al. (2014) Replication stress by Py-Im polyamides induces a non-canonical ATR-dependent checkpoint response. Nucleic Acids Res 42:11546-59|
|Raskatov, Jevgenij A; Puckett, James W; Dervan, Peter B (2014) A C-14 labeled Py-Im polyamide localizes to a subcutaneous prostate cancer tumor. Bioorg Med Chem 22:4371-5|
|Nickols, Nicholas G; Szablowski, Jerzy O; Hargrove, Amanda E et al. (2013) Activity of a Py-Im polyamide targeted to the estrogen response element. Mol Cancer Ther 12:675-84|
|Edwards, Jonathan S; Betts, Laurie; Frazier, Monica L et al. (2013) Molecular basis of antibiotic multiresistance transfer in Staphylococcus aureus. Proc Natl Acad Sci U S A 110:2804-9|
|Hargrove, Amanda E; Raskatov, Jevgenij A; Meier, Jordan L et al. (2012) Characterization and solubilization of pyrrole-imidazole polyamide aggregates. J Med Chem 55:5425-32|
|Meier, Jordan L; Yu, Abigail S; Korf, Ian et al. (2012) Guiding the design of synthetic DNA-binding molecules with massively parallel sequencing. J Am Chem Soc 134:17814-22|
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