TY - JOUR AU - Portier, I AU - Vanhoorelbeke, K AU - Verhenne, S AU - Pareyn, I AU - Vandeputte, N AU - Deckmyn, H AU - Goldenberg, DS AU - Samal, HB AU - Singh, M AU - Ivics, Zoltán AU - Izsvak, Z AU - De Meyer, SF TI - High and long-term von Willebrand factor expression after Sleeping Beauty transposon-mediated gene therapy in a mouse model of severe von Willebrand disease. JF - JOURNAL OF THROMBOSIS AND HAEMOSTASIS J2 - J THROMB HAEMOST VL - 16 PY - 2018 IS - 3 SP - 592 EP - 604 PG - 13 SN - 1538-7933 DO - 10.1111/jth.13938 UR - https://m2.mtmt.hu/api/publication/3415331 ID - 3415331 AB - Essentials von Willebrand disease (VWD) is the most common inherited bleeding disorder. Gene therapy for VWD offers long-term therapy for VWD patients. Transposons efficiently integrate the large von Willebrand factor (VWF) cDNA in mice. Liver-directed transposons support sustained VWF expression with suboptimal multimerization. SUMMARY: Background Type 3 von Willebrand disease (VWD) is characterized by complete absence of von Willebrand factor (VWF). Current therapy is limited to treatment with exogenous VWF/FVIII products, which only provide a short-term solution. Gene therapy offers the potential for a long-term treatment for VWD. Objectives To develop an integrative Sleeping Beauty (SB) transposon-mediated VWF gene transfer approach in a preclinical mouse model of severe VWD. Methods We established a robust platform for sustained transgene murine VWF (mVWF) expression in the liver of Vwf(-/-) mice by combining a liver-specific promoter with a sandwich transposon design and the SB100X transposase via hydrodynamic gene delivery. Results The sandwich SB transposon was suitable to deliver the full-length mVWF cDNA (8.4 kb) and supported supra-physiological expression that remained stable for up to 1.5 years after gene transfer. The sandwich vector stayed episomal (~60 weeks) or integrated in the host genome, respectively, in the absence or presence of the transposase. Transgene integration was confirmed using carbon tetrachloride-induced liver regeneration. Analysis of integration sites by high-throughput analysis revealed random integration of the sandwich vector. Although the SB vector supported long-term expression of supra-physiological VWF levels, the bleeding phenotype was not corrected in all mice. Long-term expression of VWF by hepatocytes resulted in relatively reduced amounts of high-molecular-weight multimers, potentially limiting its hemostatic efficacy. Conclusions Although this integrative platform for VWF gene transfer is an important milestone of VWD gene therapy, cell type-specific targeting is yet to be achieved. LA - English DB - MTMT ER - TY - JOUR AU - Smith, K AU - Li, YP AU - Piccinini, F AU - Csucs, G AU - Balazs, C AU - Bevilacqua, A AU - Horváth, Péter TI - CIDRE: an illumination-correction method for optical microscopy JF - NATURE METHODS J2 - NAT METHODS VL - 12 PY - 2015 IS - 5 SP - 404 EP - 406 PG - 3 SN - 1548-7091 DO - 10.1038/NMETH.3323 UR - https://m2.mtmt.hu/api/publication/2896546 ID - 2896546 AB - Uneven illumination affects every image acquired by a microscope. It is often overlooked, but it can introduce considerable bias to image measurements. The most reliable correction methods require special reference images, and retrospective alternatives do not fully model the correction process. Our approach overcomes these issues for most optical microscopy applications without the need for reference images. LA - English DB - MTMT ER - TY - JOUR AU - Mátés, Lajos AU - Chuah, MKL AU - Belay, E AU - Jerchow, B AU - Manoj, N AU - Acosta-Sanchez, A AU - Grzela, DP AU - Schmitt, A AU - Becker, K AU - Matrai, J AU - Ma, L AU - Samara-Kuko, E AU - Gysemans, C AU - Pryputniewicz, D AU - Miskey, C AU - Fletcher, B AU - VandenDriessche, T AU - Ivics, Zoltán AU - Izsvák, Zsuzsa TI - Molecular evolution of a novel hyperactive Sleeping Beauty transposase enables robust stable gene transfer in vertebrates JF - NATURE GENETICS J2 - NAT GENET VL - 41 PY - 2009 IS - 6 SP - 753 EP - 761 PG - 9 SN - 1061-4036 DO - 10.1038/ng.343 UR - https://m2.mtmt.hu/api/publication/1274890 ID - 1274890 N1 - Megjegyzés-20816409 FU: Volkswagen Stiftung [EU FP5, EU FP6, EU FP7]; Bundesministerium fur : Bildung und Forschung [NGFN-2]; FWO [G.0632.07]; [VIB GOA/2004/09] FX: We thank C. Judis, V. Gillijns and M. Shrahna for technical assistance. : Z. Izsvak is an EURYI Awardee. We acknowledge the financial support of : EU FP5 ( JUMPY), EU FP6 (INTHER) and EU FP7 ( PERSIST), grants from the : Volkswagen Stiftung and from the Bundesministerium fur Bildung und : Forschung (NGFN-2), VIB GOA/2004/09 and FWO ( G.0632.07). We thank E. : Zeira, E. Galun ( Goldyne Savad Institute of Gene Therapy) and M. Rhee : ( Chunguam University of Korea) for providing the pT2/CAGGS-GFP and the : pcGlobin2 constructs, respectively. We further thank M. J. Fraser ( : University of Notre Dame) and A. Bradley ( Sanger Centre) for providing : the pXL-BacII and codon-optimized piggyBac transposase constructs, : respectively, and A. Leutz and N. Rajewsky for critical reading of the : manuscript. A. Schmitt is affiliated with Medical Faculty of Charite, : Berlin. Megjegyzés-20817348 FU: Volkswagen Stiftung [EU FP5, EU FP6, EU FP7]; Bundesministerium fur : Bildung und Forschung [NGFN-2]; FWO [G.0632.07]; [VIB GOA/2004/09] FX: We thank C. Judis, V. Gillijns and M. Shrahna for technical assistance. : Z. Izsvak is an EURYI Awardee. We acknowledge the financial support of : EU FP5 ( JUMPY), EU FP6 (INTHER) and EU FP7 ( PERSIST), grants from the : Volkswagen Stiftung and from the Bundesministerium fur Bildung und : Forschung (NGFN-2), VIB GOA/2004/09 and FWO ( G.0632.07). We thank E. : Zeira, E. Galun ( Goldyne Savad Institute of Gene Therapy) and M. Rhee : ( Chunguam University of Korea) for providing the pT2/CAGGS-GFP and the : pcGlobin2 constructs, respectively. We further thank M. J. Fraser ( : University of Notre Dame) and A. Bradley ( Sanger Centre) for providing : the pXL-BacII and codon-optimized piggyBac transposase constructs, : respectively, and A. Leutz and N. Rajewsky for critical reading of the : manuscript. A. Schmitt is affiliated with Medical Faculty of Charite, : Berlin. Megjegyzés-20818067 FU: Volkswagen Stiftung [EU FP5, EU FP6, EU FP7]; Bundesministerium fur : Bildung und Forschung [NGFN-2]; FWO [G.0632.07]; [VIB GOA/2004/09] FX: We thank C. Judis, V. Gillijns and M. Shrahna for technical assistance. : Z. Izsvak is an EURYI Awardee. We acknowledge the financial support of : EU FP5 ( JUMPY), EU FP6 (INTHER) and EU FP7 ( PERSIST), grants from the : Volkswagen Stiftung and from the Bundesministerium fur Bildung und : Forschung (NGFN-2), VIB GOA/2004/09 and FWO ( G.0632.07). We thank E. : Zeira, E. Galun ( Goldyne Savad Institute of Gene Therapy) and M. Rhee : ( Chunguam University of Korea) for providing the pT2/CAGGS-GFP and the : pcGlobin2 constructs, respectively. We further thank M. J. Fraser ( : University of Notre Dame) and A. Bradley ( Sanger Centre) for providing : the pXL-BacII and codon-optimized piggyBac transposase constructs, : respectively, and A. Leutz and N. Rajewsky for critical reading of the : manuscript. A. Schmitt is affiliated with Medical Faculty of Charite, : Berlin. Megjegyzés-20818075 FU: Volkswagen Stiftung [EU FP5, EU FP6, EU FP7]; Bundesministerium fur : Bildung und Forschung [NGFN-2]; FWO [G.0632.07]; [VIB GOA/2004/09] FX: We thank C. Judis, V. Gillijns and M. Shrahna for technical assistance. : Z. Izsvak is an EURYI Awardee. We acknowledge the financial support of : EU FP5 ( JUMPY), EU FP6 (INTHER) and EU FP7 ( PERSIST), grants from the : Volkswagen Stiftung and from the Bundesministerium fur Bildung und : Forschung (NGFN-2), VIB GOA/2004/09 and FWO ( G.0632.07). We thank E. : Zeira, E. Galun ( Goldyne Savad Institute of Gene Therapy) and M. Rhee : ( Chunguam University of Korea) for providing the pT2/CAGGS-GFP and the : pcGlobin2 constructs, respectively. We further thank M. J. Fraser ( : University of Notre Dame) and A. Bradley ( Sanger Centre) for providing : the pXL-BacII and codon-optimized piggyBac transposase constructs, : respectively, and A. Leutz and N. Rajewsky for critical reading of the : manuscript. A. Schmitt is affiliated with Medical Faculty of Charite, : Berlin. Megjegyzés-20818183 FU: Volkswagen Stiftung [EU FP5, EU FP6, EU FP7]; Bundesministerium fur : Bildung und Forschung [NGFN-2]; FWO [G.0632.07]; [VIB GOA/2004/09] FX: We thank C. Judis, V. Gillijns and M. Shrahna for technical assistance. : Z. Izsvak is an EURYI Awardee. We acknowledge the financial support of : EU FP5 ( JUMPY), EU FP6 (INTHER) and EU FP7 ( PERSIST), grants from the : Volkswagen Stiftung and from the Bundesministerium fur Bildung und : Forschung (NGFN-2), VIB GOA/2004/09 and FWO ( G.0632.07). We thank E. : Zeira, E. Galun ( Goldyne Savad Institute of Gene Therapy) and M. Rhee : ( Chunguam University of Korea) for providing the pT2/CAGGS-GFP and the : pcGlobin2 constructs, respectively. We further thank M. J. Fraser ( : University of Notre Dame) and A. Bradley ( Sanger Centre) for providing : the pXL-BacII and codon-optimized piggyBac transposase constructs, : respectively, and A. Leutz and N. Rajewsky for critical reading of the : manuscript. A. Schmitt is affiliated with Medical Faculty of Charite, : Berlin. Megjegyzés-20818257 FU: Volkswagen Stiftung [EU FP5, EU FP6, EU FP7]; Bundesministerium fur : Bildung und Forschung [NGFN-2]; FWO [G.0632.07]; [VIB GOA/2004/09] FX: We thank C. Judis, V. Gillijns and M. Shrahna for technical assistance. : Z. Izsvak is an EURYI Awardee. We acknowledge the financial support of : EU FP5 ( JUMPY), EU FP6 (INTHER) and EU FP7 ( PERSIST), grants from the : Volkswagen Stiftung and from the Bundesministerium fur Bildung und : Forschung (NGFN-2), VIB GOA/2004/09 and FWO ( G.0632.07). We thank E. : Zeira, E. Galun ( Goldyne Savad Institute of Gene Therapy) and M. Rhee : ( Chunguam University of Korea) for providing the pT2/CAGGS-GFP and the : pcGlobin2 constructs, respectively. We further thank M. J. Fraser ( : University of Notre Dame) and A. Bradley ( Sanger Centre) for providing : the pXL-BacII and codon-optimized piggyBac transposase constructs, : respectively, and A. Leutz and N. Rajewsky for critical reading of the : manuscript. A. Schmitt is affiliated with Medical Faculty of Charite, : Berlin. Megjegyzés-20818339 FU: Volkswagen Stiftung [EU FP5, EU FP6, EU FP7]; Bundesministerium fur : Bildung und Forschung [NGFN-2]; FWO [G.0632.07]; [VIB GOA/2004/09] FX: We thank C. Judis, V. Gillijns and M. Shrahna for technical assistance. : Z. Izsvak is an EURYI Awardee. We acknowledge the financial support of : EU FP5 ( JUMPY), EU FP6 (INTHER) and EU FP7 ( PERSIST), grants from the : Volkswagen Stiftung and from the Bundesministerium fur Bildung und : Forschung (NGFN-2), VIB GOA/2004/09 and FWO ( G.0632.07). We thank E. : Zeira, E. Galun ( Goldyne Savad Institute of Gene Therapy) and M. Rhee : ( Chunguam University of Korea) for providing the pT2/CAGGS-GFP and the : pcGlobin2 constructs, respectively. We further thank M. J. Fraser ( : University of Notre Dame) and A. Bradley ( Sanger Centre) for providing : the pXL-BacII and codon-optimized piggyBac transposase constructs, : respectively, and A. Leutz and N. Rajewsky for critical reading of the : manuscript. A. Schmitt is affiliated with Medical Faculty of Charite, : Berlin. Megjegyzés-22179212 DI: 10.1038/ng.343 Megjegyzés-22220293 DI: 10.1038/ng.343 AB - The Sleeping Beauty (SB) transposon is a promising technology platform for gene transfer in vertebrates; however, its efficiency of gene insertion can be a bottleneck in primary cell types. A large-scale genetic screen in mammalian cells yielded a hyperactive transposase (SB100X) with B100-fold enhancement in efficiency when compared to the first-generation transposase. SB100X supported 35-50% stable gene transfer in human CD34(+) cells enriched in hematopoietic stem or progenitor cells. Transplantation of gene-marked CD34(+) cells in immunodeficient mice resulted in long-term engraftment and hematopoietic reconstitution. In addition, SB100X supported sustained (> 1 year) expression of physiological levels of factor IX upon transposition in the mouse liver in vivo. Finally, SB100X reproducibly resulted in 45% stable transgenesis frequencies by pronuclear microinjection into mouse zygotes. The newly developed transposase yields unprecedented stable gene transfer efficiencies following nonviral gene delivery that compare favorably to stable transduction efficiencies with integrating viral vectors and is expected to facilitate widespread applications in functional genomics and gene therapy. LA - English DB - MTMT ER - TY - JOUR AU - Görög, Dénes AU - Regöly-Mérei, J AU - Paku, Sándor AU - Kopper, László AU - Nagy, Péter TI - Alpha-fetoprotein expression is a potential prognostic marker in hepatocellular carcinoma JF - WORLD JOURNAL OF GASTROENTEROLOGY J2 - WORLD J LGASTROENTEROL VL - 11 PY - 2005 SP - 5015 EP - 5018 PG - 4 SN - 1007-9327 UR - https://m2.mtmt.hu/api/publication/1046984 ID - 1046984 N1 - Journal Article; Research Support, Non-U.S. Gov't LA - English DB - MTMT ER -