Repair of thalassemic human β-globin mRNA in mammalian cells by antisense oligonucleotides

Repair of thalassemic human β-globin mRNA in mammalian cells by antisense oligonucleotides

^*Lineberger Comprehensive Cancer Center and Department of Pharmacology, University of North Carolina, Chapel Hill, NC 27599; and ^‡Hybridon, Inc., Worcester, MA 01605

Abstract

In one form of β-thalassemia, a genetic blood disorder, a mutation in intron 2 of the β-globin gene (IVS2-654) causes aberrant splicing of β-globin pre-mRNA and, consequently, β-globin deficiency. Treatment of mammalian cells stably expressing the IVS2-654 human β-globin gene with antisense oligonucleotides targeted at the aberrant splice sites restored correct splicing in a dose-dependent fashion, generating correct human β-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.

Footnotes

↵ † On leave from: The Institute of Biochemistry and Biophysics, Warsaw, Poland.
↵ § To whom reprint requests should be addressed at: Lineberger Comprehensive Cancer Center, University of North Carolina, CB# 7295, Chapel Hill, NC 27599. e-mail: kole{at}med.unc.edu.
Y. W. Kan, University of California, San Francisco, CA
Abbreviation: RT-PCR, reverse transcription–PCR.