title: Hyperactive piggyBac transposons for sustained and robust liver-targeted gene therapy.
abstract: The development of robust nonviral vectors could facilitate clinical gene therapy applications and may overcome some of the immune complications of viral vectors. Nevertheless, most nonviral gene deliver approaches typically yield only transient and/or low gene expression. To address these caveats, we have explored piggyBac transposons to correct hemophilia B by liver-directed factor IX (FIX) gene therapy in hemophilic mice. To achieve this, we combined the use of: (i) a hyperactive codon-optimized piggyBac transposase, (ii) a computationally enhanced liver-specific promoter, (iii) a hyperfunctional codon-optimized FIX transgene (FIX R338L Padua), and (iv) a modification of the transposon terminal repeats. This combination strategy resulted in a robust 400-fold improvement in vector performance in hepatocytes, yielding stable supraphysiologic human FIX activity (>1 year). Liver-specific expression resulted in the induction of FIX-specific immune tolerance. Remarkably, only very low transposon/transposase doses were required to cure the bleeding diathesis. Similarly, PB transposons could be used to express supraphysiologic factor VIII levels using low transposon/transposase doses. PB transposition did not induce tumors in a sensitive hepatocellular carcinoma-prone mouse model. These results underscore the potency and relative safety of the latest generation PB transposons, which constitutes a versatile platform for stable and robust secretion of therapeutic proteins.
authors.affiliation: 1] Department of Gene Therapy and Regenerative Medicine, Free University of Brussels, Brussels, Belgium [2] Center for Molecular and Vascular Biology, Department of Cardiovascular Medicine, University of Leuven, Leuven, Belgium.
authors.forename: Mario
authors.initials: M
authors.lastname: Di Matteo
authors.affiliation: Department of Gene Therapy and Regenerative Medicine, Free University of Brussels, Brussels, Belgium.
authors.forename: Emira
authors.initials: E
authors.lastname: Samara-Kuko
authors.affiliation: Molecular Immunology Unit, Institute of Child Health, University College London, London, UK.
authors.forename: Natalie J
authors.initials: NJ
authors.lastname: Ward
authors.affiliation: 1] Institute for Women's Health Gene Transfer Technology Group, University College London, London, UK [2] School of Pathology, Faculty of Health Sciences, University of the Witwatersrand, South Africa.
authors.forename: Simon N
authors.initials: SN
authors.lastname: Waddington
authors.forename: Simon N
authors.initials: SN
authors.lastname: Waddingon
authors.affiliation: University of Surrey, Molecular Medicine Group, Surrey, UK.
authors.forename: John H
authors.initials: JH
authors.lastname: McVey
authors.affiliation: 1] Department of Gene Therapy and Regenerative Medicine, Free University of Brussels, Brussels, Belgium [2] Center for Molecular and Vascular Biology, Department of Cardiovascular Medicine, University of Leuven, Leuven, Belgium.
authors.forename: Marinee K L
authors.initials: MK
authors.lastname: Chuah
authors.affiliation: 1] Department of Gene Therapy and Regenerative Medicine, Free University of Brussels, Brussels, Belgium [2] Center for Molecular and Vascular Biology, Department of Cardiovascular Medicine, University of Leuven, Leuven, Belgium.
authors.forename: Thierry
authors.initials: T
authors.lastname: VandenDriessche
mesh_terms: D000818:Animals; D004251:DNA Transposable Elements; D004195:Disease Models, Animal; D005164:Factor IX / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage* / Q000627:therapeutic use; D002836:Hemophilia B / Q000276:immunology / Q000628:therapy*; D022781:Hepatocytes / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D009928:Organ Specificity; D019895:Transposases / Q000235:genetics / Q000378:metabolism
chemical_list: D004251:DNA Transposable Elements; D005164:Factor IX; D019895:Transposases