title: Specific targeting of a DNA-alkylating reagent to mitochondria. Synthesis and characterization of [4-((11aS)-7-methoxy-1,2,3,11a-tetrahydro-5H-pyrrolo[2,1-c][1,4]benzodiazepin-5-on-8-oxy)butyl]-triphenylphosphonium iodide.
abstract: The selective manipulation of the expression and replication of mitochondrial DNA (mtDNA) within mammalian cells has proven difficult. In progressing towards this goal we synthesized a novel mitochondria-targeted DNA-alkylating reagent. The active alkylating moiety [(11aS)-8-hydroxy-7-methoxy-1,2,3,11a-tetrahydro-5H-pyrrolo[2,1-c][1,4]benzodiazepin-5-one (DC-81)], irreversibly alkylates guanine bases in DNA (with a preference for AGA triplets), preventing its expression and replication. To target this compound to mitochondria it was covalently coupled to the lipophilic triphenylphosphonium (TPP) cation to form a derivative referred to as mitoDC-81. Incorporation of this lipophilic cation led to the rapid uptake of mitoDC-81 by mitochondria, driven by the large membrane potential across the inner membrane. This compound efficiently alkylated isolated supercoiled, relaxed-circular or linear plasmid DNA and isolated mtDNA. However mitoDC-81 did not alkylate mtDNA within isolated mitochondria or cells, even though it accessed the mitochondrial matrix at concentrations up to 100-fold higher than those required to alkylate isolated DNA. This surprising finding suggests that mtDNA within intact mitochondria may not be accessible to this class of alkylating reagent. This inability to alkylate mtDNA in situ has significant implications for the design of therapies for mtDNA diseases and for studies on the packaging, expression and turnover of mtDNA in general.
authors.affiliation: MRC-Dunn Human Nutrition Unit, Wellcome Trust-MRC Building, Cambridge, UK.
authors.forename: Andrew M
authors.initials: AM
authors.lastname: James
authors.forename: Frances H
authors.initials: FH
authors.lastname: Blaikie
authors.forename: Robin A J
authors.initials: RA
authors.lastname: Smith
authors.forename: Robert N
authors.initials: RN
authors.lastname: Lightowlers
authors.forename: Paul M
authors.initials: PM
authors.lastname: Smith
authors.forename: Michael P
authors.initials: MP
authors.lastname: Murphy
mesh_terms: D000477:Alkylating Agents / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D001569:Benzodiazepines / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism*; D002412:Cations / Q000737:chemistry / Q000378:metabolism; D019069:Cell Respiration / Q000502:physiology; D004272:DNA, Mitochondrial / Q000378:metabolism*; D006801:Humans; D007454:Iodides / Q000737:chemistry / Q000378:metabolism*; D008930:Mitochondria, Liver / Q000378:metabolism*; D015394:Molecular Structure; D009943:Organophosphorus Compounds / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism*; D051381:Rats; D014407:Tumor Cells, Cultured
chemical_list: C474072:(11aS)-8-hydroxy-7-methoxy-1,2,3,11a-tetrahydro-5H-pyrrolo(2,1-c)(1,4)benzodiazepin-5-one; D000477:Alkylating Agents; D002412:Cations; D004272:DNA, Mitochondrial; D007454:Iodides; D009943:Organophosphorus Compounds; D001569:Benzodiazepines