Repair of thalassemic human beta-globin mRNA in mammalian cells by antisense oligonucleotides.
In one form of beta-thalassemia, a genetic blood disorder, a mutation in intron 2 of the beta-globin gene (IVS2-654) causes aberrant splicing of beta-globin pre-mRNA and, consequently, beta-globin deficiency. Treatment of mammalian cells stably expressing the IVS2-654 human beta-globin gene with antisense oligonucleotides targeted at the aberrant splice sites restored correct splicing in a dose-dependent fashion, generating correct human beta-globin mRNA and polypeptide. Both products persisted for up to 72 hr posttreatment. The oligonucleotides modified splicing by a true antisense mechanism without overt unspecific effects on cell growth and splicing of other pre-mRNAs. This novel approach in which antisense oligonucleotides are used to restore rather than to down-regulate the activity of the target gene is applicable to other splicing mutants and is of potential clinical interest.
Sierakowska
H
H
Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill 27599, USA.
Sambade
M J
MJ
Agrawal
S
S
Kole
R
R
D016475:3T3 Cells; D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D015870:Gene Expression / Q000187:drug effects; D015316:Genetic Therapy / Q000379:methods; D005914:Globins / Q000096:biosynthesis* / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D051379:Mice; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D012322:RNA Precursors / Q000378:metabolism*; D012326:RNA Splicing / Q000187:drug effects*; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis; D013873:Thionucleotides; D014162:Transfection; D017086:beta-Thalassemia / Q000235:genetics* / Q000628:therapy
D016376:Oligonucleotides, Antisense; D012322:RNA Precursors; D012333:RNA, Messenger; D011994:Recombinant Proteins; D013873:Thionucleotides; D005914:Globins