Translesion DNA synthesis polymerases (TLS pols) are vital to bypass and tolerance of DNA damage. The mis-regulation of TLS pols occurs in many types of cancer, including malignant brain tumors of glial origin. Human DNA polymerase kappa (hpol ?) was recently shown to be an independent prognostic indicator of poor outcome in glioma patients. However, the mechanism(s) leading to up-regulation of hpol ? in gliomas and the precise effects upon DNA replication remain unknown. The experiments outlined herein will examine whether tumor-promoting activation of the aryl hydrocarbon receptor (AhR) leads to constitutive up-regulation of hpol ? in gliomas and then investigate mechanistic features associated with hpol ? activity in these deadly tumors. The central objectives include investigating mechanisms that promote hpol ? over-expression in glioma cells, determining the effect of this mis-regulation upon replication stress responses, studying protein-protein interactions that alter hpol ? activity and examining potential ways of improving glioma treatment through targeted inhibition of TLS pols in brain tumors. The current proposal will address important questions, such as: 1) Does the AhR pathway regulate hpol ? in gliomas? 2) Do exogenous or endogenous AhR ligands affect hpol expression and/or activity? 3) How does blocking endogenous AhR-signaling in gliomas affect replication stress? 4) Can modulation of TLS activity in gliomas potentiate anti-cancer treatments? 5) Do protein-protein interactions between hpol ? and the genomic caretaker Werner's syndrome protein (WRN) limit mutagenic bypass of DNA damage in glioma cells? The experimental strategies utilize in vitro cell culture, mutagenesis assays, structure-function analyses, small-molecule inhibitor studies and proteomics to ascertain the role of hpol ? in gliomas. The long-term goal of the application is to produce models for replication stress and TLS activity that inform our understanding of genomic maintenance and mutagenesis in gliomas, while also providing insights into how we might better treat patients suffering from this disease. The conclusions derived from these studies will dramatically improve our fundamental definition of processes that occur in malignant brain tumors, a major public health concern.

Public Health Relevance

Accurate copying of DNA is a barrier to cancer. Defects in accurate DNA replication can drive tumor development and limit the efficacy of anti-cancer treatments. We are interested in studying cellular mechanisms that lead to mis-regulation of the error-prone human DNA polymerase kappa (hpol ?) in gliomas and then examining the effect of hpol ? mis-regulation upon DNA replication events in these tumors. Experimental approaches to develop small-molecule inhibitors of hpol ? and potentiate anti-cancer treatments are also proposed. Additional studies will investigate interactions between hpol ? and the genomic caretaker Werner's syndrome protein (WRN), specifically focusing on mutagenic bypass of DNA damage. The research outlined in the current proposal will elucidate new mechanisms promoting genomic instability in brain tumors and may provide insight into how existing anti-cancer treatments may be improved through targeted inhibition of cellular pathways that activate error-prone polymerases. These results will elucidate fundamental properties related to DNA replication and may serve to improve our understanding of and ability to treat processes that are engaged in malignant brain tumors.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Cancer Etiology Study Section (CE)
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Okano, Paul
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University of Arkansas for Medical Sciences
Schools of Medicine
Little Rock
United States
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Ketkar, Amit; Voehler, Markus; Mukiza, Tresor et al. (2017) Residues in the RecQ C-terminal Domain of the Human Werner Syndrome Helicase Are Involved in Unwinding G-quadruplex DNA. J Biol Chem 292:3154-3163
Eoff, Robert L; Raney, Kevin D (2017) A catch and release program for single-stranded DNA. J Biol Chem 292:13085-13086
Zafar, Maroof K; Eoff, Robert L (2017) Translesion DNA Synthesis in Cancer: Molecular Mechanisms and Therapeutic Opportunities. Chem Res Toxicol 30:1942-1955
Eddy, Sarah; Tillman, Magdalena; Maddukuri, Leena et al. (2016) Human Translesion Polymerase ? Exhibits Enhanced Activity and Reduced Fidelity Two Nucleotides from G-Quadruplex DNA. Biochemistry 55:5218-29
Bostian, April C L; Eoff, Robert L (2016) Aberrant Kynurenine Signaling Modulates DNA Replication Stress Factors and Promotes Genomic Instability in Gliomas. Chem Res Toxicol 29:1369-80
Yeom, Mina; Kim, In-Hyeok; Kim, Jae-Kwon et al. (2016) Effects of Twelve Germline Missense Variations on DNA Lesion and G-Quadruplex Bypass Activities of Human DNA Polymerase REV1. Chem Res Toxicol 29:367-79
Madadi, Nikhil R; Ketkar, Amit; Penthala, Narsimha R et al. (2016) Dioxol and dihydrodioxin analogs of 2- and 3-phenylacetonitriles as potent anti-cancer agents with nanomolar activity against a variety of human cancer cells. Bioorg Med Chem Lett 26:2164-9
Bostian, April C L; Maddukuri, Leena; Reed, Megan R et al. (2016) Kynurenine Signaling Increases DNA Polymerase Kappa Expression and Promotes Genomic Instability in Glioblastoma Cells. Chem Res Toxicol 29:101-8
Madadi, Nikhil R; Zong, Hongliang; Ketkar, Amit et al. (2015) Synthesis and evaluation of a series of resveratrol analogues as potent anti-cancer agents that target tubulin. Medchemcomm 6:788-794
Eddy, Sarah; Maddukuri, Leena; Ketkar, Amit et al. (2015) Evidence for the kinetic partitioning of polymerase activity on G-quadruplex DNA. Biochemistry 54:3218-30

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