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.
Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill 27599, USA.
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Sierakowska
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Sambade
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Agrawal
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Kole
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