During radiation therapy of human tumors the majority of DNA lesions originate from the radiolysis of water in the vicinity of the DNA molecule. These lesions can block DNA replication, which ultimately can lead to cell death and therapeutic efficacy. Alternatively, DNA damage can be bypassed resulting in mutations and the possibility of secondary tumors at the site of treatment by ionizing radiation. Thus it is the interaction between a DNA polymerase and a DNA lesion that ultimately determines the fate of irradiated cells. In order to maximize therapeutic gain and minimize the risk of secondary tumors at the treatment site it is imperative to have a solid mechanistic understanding of the interactions between a human DNA polymerase and a radiation-induced DNA lesion. We propose to address this central issue by studying two types of human DNA polymerases, replicative and specialized, in the context of ionization-induced lesions such as thymine glycol and abasic sites. DNA polymerase ? is a high-fidelity enzyme that plays a crucial role in DNA replication and repair. Pol ? is only somewhat proficient at inserting and extending past abasic sites and is unable to bypass Tg. In contrast, two specialized low-fidelity enzymes, DNA polymerases ? and ?, are much more efficient at extending past these DNA lesions generated by ionizing radiation. We will use a combination of biochemical and structural methods to uncover the mechanisms underpinning replication block or lesion bypass in the context of a replicative or bypass polymerase, with the ultimate goal to design compounds that inhibit pol ?, a polymerase that is overexpressed in breast cancer tumors.

Public Health Relevance

DNA polymerase ? is one of the three human replicative DNA polymerase. Beyond its role in DNA replication it also plays a crucial role in DNA repair. The high fidelity of this replicative DNA polymerase is paramount for the maintenance of genome stability and for the avoidance of carcinogenesis. Pol ? and ? are two specialized human DNA polymerases, which are known to bypass ionization-generated lesions such as abasic sites or thymine glycols. Importantly pol ? is overexpressed in breast cancer tumors and knockdown of its gene makes tumor cells more sensitive to radiation, with little effect to normal tissues. The long-term goal of this proposal is to develop inhibitors of pol ? to aid in the radiosensitization of tumor cells.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
2R01CA052040-19A1
Application #
8442632
Study Section
Radiation Therapeutics and Biology Study Section (RTB)
Program Officer
Pelroy, Richard
Project Start
1991-01-01
Project End
2017-12-31
Budget Start
2013-01-01
Budget End
2013-12-31
Support Year
19
Fiscal Year
2013
Total Cost
$331,422
Indirect Cost
$114,096
Name
University of Vermont & St Agric College
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
066811191
City
Burlington
State
VT
Country
United States
Zip Code
05405
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Aller, Pierre; Duclos, Stéphanie; Wallace, Susan S et al. (2011) A crystallographic study of the role of sequence context in thymine glycol bypass by a replicative DNA polymerase serendipitously sheds light on the exonuclease complex. J Mol Biol 412:22-34
Zahn, Karl E; Tchesnokov, Egor P; Gotte, Matthias et al. (2011) Phosphonoformic acid inhibits viral replication by trapping the closed form of the DNA polymerase. J Biol Chem 286:25246-55

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