title: A strategy for selective O(6)-alkylguanine-DNA alkyltransferase depletion under hypoxic conditions.
abstract: Cellular resistance to chemotherapeutics that alkylate the O-6 position of guanine residues in DNA correlates with their O(6)-alkylguanine-DNA alkyltransferase activity. In normal cells high [O(6)-alkylguanine-DNA alkyltransferase] is beneficial, sparing the host from toxicity, whereas in tumor cells high [O(6)-alkylguanine-DNA alkyltransferase] prevents chemotherapeutic response. Therefore, it is necessary to selectively inactivate O(6)-alkylguanine-DNA alkyltransferase in tumors. The oxygen-deficient compartment unique to solid tumors is conducive to reduction, and could be utilized to provide this selectivity. Therefore, we synthesized 2-nitro-6-benzyloxypurine, an analog of O(6)-benzylguanine in which the essential 2-amino group is replaced by a nitro moiety, and 2-nitro-6-benzyloxypurine is >2000-fold weaker than O(6)-benzylguanine as an O(6)-alkylguanine-DNA alkyltransferase inhibitor. We demonstrate oxygen concentration sensitive net reduction of 2-nitro-6-benzyloxypurine by cytochrome P450 reductase, xanthine oxidase, and EMT6, DU145, and HL-60 cells to yield O(6)-benzylguanine. We show that 2-nitro-6-benzyloxypurine treatment depletes O(6)-alkylguanine-DNA alkyltransferase in intact cells under oxygen-deficient conditions and selectively sensitizes cells to laromustine (an agent that chloroethylates the O-6 position of guanine) under oxygen-deficient but not normoxic conditions. 2-Nitro-6-benzyloxypurine represents a proof of concept lead compound; however, its facile reduction (E(1/2) - 177 mV versus Ag/AgCl) may result in excessive oxidative stress and/or the generation of O(6)-alkylguanine-DNA alkyltransferase inhibitors in normoxic regions in vivo.
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA. philip.penketh@yale.edu
authors.forename: Philip G
authors.initials: PG
authors.lastname: Penketh
authors.forename: Krishnamurthy
authors.initials: K
authors.lastname: Shyam
authors.forename: Raymond P
authors.initials: RP
authors.lastname: Baumann
authors.forename: Kimiko
authors.initials: K
authors.lastname: Ishiguro
authors.forename: Eric V
authors.initials: EV
authors.lastname: Patridge
authors.forename: Rui
authors.initials: R
authors.lastname: Zhu
authors.forename: Alan C
authors.initials: AC
authors.lastname: Sartorelli
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D015687:Cell Hypoxia / Q000187:drug effects; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D006147:Guanine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D018922:HL-60 Cells; D006801:Humans; D006834:Hydrazines / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000378:metabolism; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism*; D011687:Purines / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D013449:Sulfonamides / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D006834:Hydrazines; D011687:Purines; D013449:Sulfonamides; C064976:O(6)-benzylguanine; D013481:Superoxides; C483604:laromustine; D006147:Guanine; D006861:Hydrogen Peroxide; D019853:O(6)-Methylguanine-DNA Methyltransferase; D010100:Oxygen