title: DNA Adduct Profiles Predict in Vitro Cell Viability after Treatment with the Experimental Anticancer Prodrug PR104A.
abstract: PR104A is an experimental DNA-alkylating hypoxia-activated prodrug that can also be activated in an oxygen-independent manner by the two-electron aldo-keto reductase 1C3. Nitroreduction leads to the formation of cytotoxic hydroxylamine (PR104H) and amine (PR104M) metabolites, which induce DNA mono and cross-linked adducts in cells. PR104A-derived DNA adducts can be utilized as drug-specific biomarkers of efficacy and as a mechanistic tool to elucidate the cellular and molecular effects of PR104A. Toward this goal, a mass spectrometric bioanalysis approach based on a stable isotope-labeled adduct mixture (SILAM) and selected reaction monitoring (SRM) data acquisition for relative quantitation of PR104A-derived DNA adducts in cells was developed. Use of this SILAM-based approach supported simultaneous relative quantitation of 33 PR104A-derived DNA adducts in the same sample, which allowed testing of the hypothesis that the enhanced cytotoxicity, observed by preconditioning cells with the transcription-activating isothiocyanate sulforaphane, is induced by an increased level of DNA adducts induced by PR104H and PR104M, but not PR104A. By applying the new SILAM-SRM approach, we found a 2.4-fold increase in the level of DNA adducts induced by PR104H and PR104M in HT-29 cells preconditioned with sulforaphane and a corresponding 2.6-fold increase in cytotoxicity. These results suggest that DNA adduct levels correlate with drug potency and underly the possibility of monitoring PR104A-derived DNA adducts as biomarkers of efficacy.
authors.affiliation: Department of Health Sciences and Technology, ETH Zurich , Schmelzbergstrasse 9, 8092 Zurich, Switzerland.
authors.forename: Alessia
authors.identifier: 0000-0002-8287-9551
authors.initials: A
authors.lastname: Stornetta
authors.affiliation: Masonic Cancer Center, University of Minnesota , 2231 Sixth Street Southeast, Minneapolis, Minnesota 55455, United States.
authors.forename: Peter W
authors.initials: PW
authors.lastname: Villalta
authors.affiliation: Department of Health Sciences and Technology, ETH Zurich , Schmelzbergstrasse 9, 8092 Zurich, Switzerland.
authors.forename: Frederike
authors.initials: F
authors.lastname: Gossner
authors.affiliation: Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland , Auckland 92019, New Zealand.
authors.forename: William R
authors.initials: WR
authors.lastname: Wilson
authors.affiliation: Masonic Cancer Center, University of Minnesota , 2231 Sixth Street Southeast, Minneapolis, Minnesota 55455, United States.
authors.forename: Silvia
authors.initials: S
authors.lastname: Balbo
authors.affiliation: Department of Health Sciences and Technology, ETH Zurich , Schmelzbergstrasse 9, 8092 Zurich, Switzerland.
authors.forename: Shana J
authors.identifier: 0000-0001-6808-5950
authors.initials: SJ
authors.lastname: Sturla
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D018736:DNA Adducts; D019073:HT29 Cells; D006801:Humans; D066298:In Vitro Techniques; D009588:Nitrogen Mustard Compounds / Q000494:pharmacology*; D011355:Prodrugs / Q000494:pharmacology*
chemical_list: D000970:Antineoplastic Agents; D018736:DNA Adducts; D009588:Nitrogen Mustard Compounds; C524170:PR-104A; D011355:Prodrugs