Restoration of correct splicing in thalassemic pre-mRNA by antisense oligonucleotides.

Z Dominski, R Kole - … of the National Academy of Sciences, 1993 - National Acad Sciences
Z Dominski, R Kole
Proceedings of the National Academy of Sciences, 1993National Acad Sciences
Antisense 2'-O-methylribooligonucleotides were targeted against specific sequence
elements in mutated human beta-globin pre-mRNAs to restore correct splicing of these
RNAs in vitro. The following mutations of the beta-globin gene, A--> G at nt 110 of the first
intron (beta 110), T--> G at nt 705 and C--> T at nt 654 of the second intron (IVS2 (705) and
IVS2 (654), respectively), which led to aberrant splicing of the corresponding pre-mRNAs,
were previously identified as the underlying causes of beta-thalassemia. Aberrant splicing of …
Antisense 2'-O-methylribooligonucleotides were targeted against specific sequence elements in mutated human beta-globin pre-mRNAs to restore correct splicing of these RNAs in vitro. The following mutations of the beta-globin gene, A-->G at nt 110 of the first intron (beta 110), T-->G at nt 705 and C-->T at nt 654 of the second intron (IVS2(705) and IVS2(654), respectively), which led to aberrant splicing of the corresponding pre-mRNAs, were previously identified as the underlying causes of beta-thalassemia. Aberrant splicing of beta 110 pre-mRNA was efficiently reversed by an oligonucleotide targeted against the branch point sequence in the first intron of the pre-mRNA but not by an oligonucleotide targeted against the aberrant 3' splice site. In both IVS2(705) and IVS2(654) pre-mRNAs, correct splicing was restored by oligonucleotides targeted against the aberrant 5' splice sites created by the mutations in the second intron or against a cryptic 3' splice site located upstream and activated in the mutated background. These experiments represent an approach in which antisense oligonucleotides are used to restore the function of a defective gene and not, as usual, to down-regulate the expression of an undesirable gene.
National Acad Sciences