Methylating agents are mainstay drugs for the treatment of cancer and are amongst the few agents available against brain tumors. Cells responsible for keeping the blood cell supply, """"""""blood progenitor cells"""""""", are highly sensitive to these agents, which results in severe side-effects, including bleeding and infection. Blood progenitor cells can be protected from the toxic effects of methylation by resistance genes. The most effective one is AGT, which directly removes toxic modifications caused by methylation of the cell's genetic material (DNA). However, AGT does not recognize certain toxic DNA modifications. Here I propose the use of a different gene (AlkB), which targets some of these alternative lesions, to improve protection against the toxic effects of methylation. This gene, which has only recently been characterized, should therefore enhance AGT protection. Moreover, unlike AGT, AlkB remains active after mediating a repair reaction, which implies that AlkB repair should be more amenable to modulation. Here I propose a powerful approach to identify variants of one human homologue of AlkB (ABH2) with improved repair performance. Specifically, I want to enhance the enzyme's ability to recognize lesions generated by agents used in the clinic. Selected variants will be tested in human progenitor cells in culture and in a mouse animal model for protection against treatment with high doses of these agents. Ultimately, the mutants identified in this work should improve the management of side effects of methylating agents and would also allow the use of higher doses for more effective treatment.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Clinical Investigator Award (CIA) (K08)
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Subcommittee G - Education (NCI)
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Myrick, Dorkina C
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University of California Santa Cruz
Public Health & Prev Medicine
Schools of Arts and Sciences
Santa Cruz
United States
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Standley, Melissa S; Million-Weaver, Samuel; Alexander, David L et al. (2018) Genetic control of ColE1 plasmid stability that is independent of plasmid copy number regulation. Curr Genet :
Lilly, Joshua; Camps, Manel (2015) Mechanisms of Theta Plasmid Replication. Microbiol Spectr 3:
Lilly, Joshua; Camps, Manel (2015) Mechanisms of Theta Plasmid Replication. Microbiol Spectr 3:PLAS-0029-2014
Troll, Christopher J; Adhikary, Suraj; Cueff, Marie et al. (2014) Interplay between base excision repair activity and toxicity of 3-methyladenine DNA glycosylases in an E. coli complementation system. Mutat Res 763-764:64-73
Million-Weaver, Samuel; Camps, Manel (2014) Mechanisms of plasmid segregation: have multicopy plasmids been overlooked? Plasmid 75:27-36
Troll, Christopher; Yoder, Jordan; Alexander, David et al. (2014) The mutagenic footprint of low-fidelity Pol I ColE1 plasmid replication in E. coli reveals an extensive interplay between Pol I and Pol III. Curr Genet 60:123-34
Alexander, David L; Lilly, Joshua; Hernandez, Jaime et al. (2014) Random mutagenesis by error-prone pol plasmid replication in Escherichia coli. Methods Mol Biol 1179:31-44
Million-Weaver, Samuel; Alexander, David L; Allen, Jennifer M et al. (2012) Quantifying plasmid copy number to investigate plasmid dosage effects associated with directed protein evolution. Methods Mol Biol 834:33-48
Allen, Jennifer M; Simcha, David M; Ericson, Nolan G et al. (2011) Roles of DNA polymerase I in leading and lagging-strand replication defined by a high-resolution mutation footprint of ColE1 plasmid replication. Nucleic Acids Res 39:7020-33
Troll, Chris; Alexander, David; Allen, Jennifer et al. (2011) Mutagenesis and functional selection protocols for directed evolution of proteins in E. coli. J Vis Exp :

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