title: Impairment of APE1 function enhances cellular sensitivity to clinically relevant alkylators and antimetabolites.
abstract: Base excision repair (BER) is the major pathway for removing mutagenic and cytotoxic oxidative and alkylation DNA modifications. Using a catalytically inactive, dominant negative protein form of human APE1, termed ED, which binds with high affinity to substrate DNA and blocks subsequent repair steps, we assessed the role of BER in mediating cellular resistance to clinically relevant alkylating drugs and antimetabolites. Colony formation assays revealed that ED expression enhanced cellular sensitivity to melphalan not at all; to decarbazine, thiotepa, busulfan and carmustine moderately (1.2- to 2.4-fold); and to streptozotocin and temozolomide significantly (2.0- to 5.3-fold). The effectiveness of ED to promote enhanced cytotoxicity generally correlated with the agent's (a) monofunctional nature, (b) capacity to induce N(7)-guanine and N(3)-adenine modifications, and (c) inability to generate O(6)-guanine adducts or DNA cross-links. ED also enhanced the cell killing potency of the antimetabolite troxacitabine, apparently by blocking the processing of DNA strand breaks, yet had no effect on the cytotoxicity of gemcitabine, results that agree well with the known efficiency of APE1 to excise these nucleoside analogues from DNA. Most impressively, ED expression produced an approximately 5- and 25-fold augmentation of the cell killing effect of 5-fluorouracil and 5-fluorodeoxyuridine, respectively, implicating BER in the cellular response to such antimetabolites; the increased 5-fluorouracil sensitivity was associated with an accumulation of abasic sites and active caspase-positive staining. Our data suggest that APE1, and BER more broadly, is a potential target for inactivation in anticancer treatment paradigms that involve select alkylating agents or antimetabolites.
authors.affiliation: Biomedical Research Center, National Institute on Aging, NIH, Baltimore, MD 21224, USA.
authors.forename: Daniel R
authors.initials: DR
authors.lastname: McNeill
authors.forename: Wing
authors.initials: W
authors.lastname: Lam
authors.forename: Theodore L
authors.initials: TL
authors.lastname: DeWeese
authors.forename: Yung-Chi
authors.initials: YC
authors.lastname: Cheng
authors.forename: David M
authors.initials: DM
authors.lastname: Wilson
mesh_terms: D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D016466:CHO Cells; D002453:Cell Cycle / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D003412:Cricetulus; D004260:DNA Repair; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D005472:Fluorouracil / Q000494:pharmacology; D016193:G1 Phase / Q000187:drug effects; D006801:Humans; D009154:Mutation; D009369:Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism / Q000473:pathology
chemical_list: D000964:Antimetabolites, Antineoplastic; D018906:Antineoplastic Agents, Alkylating; C085108:APEX1 protein, human; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D005472:Fluorouracil