Journal article
The canine factor VIII 3'-untranslated region and a concatemeric hepatocyte nuclear factor 1 regulatory element enhance factor VIII transgene expression in vivo
Human gene therapy, Vol.13(13), pp.1583-1593
2002
DOI: 10.1089/10430340260201671
PMID: 12228013
Abstract
If gene therapy is to be an effective treatment modality for hemophilia A, therapeutic levels and tissue-restricted expression of factor VIII (FVIII) must be achieved through optimization of transgene expression. To this end, we incorporated three types of sequence elements into a canine B domain-deleted FVIII transgene cassette and individually evaluated their effect on FVIII transgene expression. Functional FVIII activity was initially assessed in vitro and hydrodynamic injection of the different transgene constructs into mice was subsequently used as a model to compare in vivo expression of the various modified transgenes. Our results demonstrate that in vitro transgene expression is, in these studies, not a good predictor of in vivo transgene performance. In vivo analysis of a hybrid tissue-restricted promoter element, consisting of a concatemer of five hepatocyte nuclear factor 1 (HNF-1) consensus-binding motifs juxtaposed to the human FVIII proximal promoter, indicates that it is as efficient at mediating expression of the FVIII protein as the cytomegalovirus promoter. Addition of the full-length canine FVIII 3′-UTR also enhances transgene expression of FVIII in vivo. Sequence analysis of the canine FVIII 3′-UTR and human FVIII 3′-UTR indicates that the former lacks instability sequences and may therefore be more effective in stabilizing FVIII mRNA. Subsequent inclusion of FVIII introns 16 and 17 into the natural locations of the transgene disrupted mRNA processing and abolished expression of the FVIII protein. Introduction of intron 17 proximal to the FVIII cDNA did not enhance in vivo expression of canine FVIII from the transgene.
Details
- Title: Subtitle
- The canine factor VIII 3'-untranslated region and a concatemeric hepatocyte nuclear factor 1 regulatory element enhance factor VIII transgene expression in vivo
- Creators
- Colleen NOTLEY - Department of Pathology, Richardson Laboratories, Queen's University, Kingston, Ontario, K7L 3N6, CanadaAnnie KILLORAN - Department of Pathology, Richardson Laboratories, Queen's University, Kingston, Ontario, K7L 3N6, CanadaCherie CAMERON - Department of Pathology, Richardson Laboratories, Queen's University, Kingston, Ontario, K7L 3N6, CanadaKimberly WYND - Department of Pathology, Richardson Laboratories, Queen's University, Kingston, Ontario, K7L 3N6, CanadaChristine HOUGH - Department of Pathology, Richardson Laboratories, Queen's University, Kingston, Ontario, K7L 3N6, CanadaDavid LILLICRAP - Department of Pathology, Richardson Laboratories, Queen's University, Kingston, Ontario, K7L 3N6, Canada
- Resource Type
- Journal article
- Publication Details
- Human gene therapy, Vol.13(13), pp.1583-1593
- Publisher
- Liebert; Larchmont, NY
- DOI
- 10.1089/10430340260201671
- PMID
- 12228013
- ISSN
- 1043-0342
- eISSN
- 1557-7422
- Language
- English
- Date published
- 2002
- Academic Unit
- Neurology; Psychiatry
- Record Identifier
- 9984015007802771
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