title: Targeted gene correction with 5' acridine-oligonucleotide conjugates.
abstract: Single-stranded oligonucleotides (SSOs) mediate gene repair of punctual chromosomal mutations at a low frequency. We hypothesized that enhancement of DNA binding affinity of SSOs by intercalating agents may increase the number of corrected cells. Several biochemical modifications of SSOs were tested for their capability to correct a chromosomally integrated and mutated GFP reporter gene in human 293 cells. SSOs of 25 nucleotide length conjugated with acridine at their 5' end increased the efficiency of gene correction up to 10-fold compared to nonmodified SSOs. Acridine and psoralen conjugates were both evaluated, and acridine-modified SSOs were the most effective. Conjugation with acridine at the 3' end of the SSO inhibited gene correction, whereas flanking the SSO by acridine on both sides provided an intermediate level of correction. These results suggest that increasing the stability of hybridization between SSO and its target without hampering a 3' extension improves gene targeting, in agreement with the "annealing-integration" model of DNA repair.
authors.affiliation: INSERM U733, Laboratoire de Thérapie Génique Hématopoïétique, Institut Universitaire d'Hématologie, Hôpital Saint-Louis, 75010 Paris, France.
authors.forename: G
authors.initials: G
authors.lastname: de Piédoue
authors.forename: C
authors.initials: C
authors.lastname: Andrieu-Soler
authors.forename: J P
authors.initials: JP
authors.lastname: Concordet
authors.forename: R
authors.initials: R
authors.lastname: Maurisse
authors.forename: J-S
authors.initials: JS
authors.lastname: Sun
authors.forename: B
authors.initials: B
authors.lastname: Lopez
authors.forename: I
authors.initials: I
authors.lastname: Kuzniak
authors.forename: P
authors.initials: P
authors.lastname: Leboulch
authors.forename: J-P
authors.initials: JP
authors.lastname: Feugeas
mesh_terms: D000166:Acridines / Q000378:metabolism*; D005363:Ficusin / Q000378:metabolism; D015316:Genetic Therapy; D006801:Humans; D007364:Intercalating Agents / Q000378:metabolism*; D009841:Oligonucleotides / Q000235:genetics* / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism; D052416:Targeted Gene Repair / Q000379:methods*; D014162:Transfection
chemical_list: D000166:Acridines; D007364:Intercalating Agents; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; D005363:Ficusin