Main Article Content

Seyed Mehdi Hassanzadeh
Azadeh Mirfeizollahi
Jafar Nikzad
Elahe Omidi
Hooman Kaghazian


Transposon, cell line development, Elosulfase alfa, Imiglucerase, PiggyBac


lysosomal storage diseases are treated using recombinant enzyme replacement in Morquio A syndrome by Elosulfase alfa and Gaucher disease via Imiglucerase. Production of these recombinant enzymes with high quality need to appropriate genetic manipulation of the host genome. In this work, PiggyBac transposon system was designed for expression of elosulfase alfa and imiglucerase in the Chinese hamster ovary (CHO) cells. Three transposon vectors including PB::GALNS, PB::GBA, and PB::GFP were constructed separately and together with SuperPBase vector were transfected into the CHO-K1 cells. Then recombinant cell lines were selected by exposure to zeocin. The appearance of green fluorescent cells by PB: GFP/Super Base vectors showed the efficiency of the designed transposon system. The expression of elosulfase alfa in the GALNS recombinant cell lines and imiglucerase in GBA recombinant cells were confirmed by ELISA. In conclusion, The PiggyBac transposon system can be considered as an efficient tool to modification the CHO host genome for the development of recombinant cell lines to produce therapeutic enzymes.

Abstract 100 | pdf Downloads 90


1. Winchester B, Vellodi A, Young E. The molecular basis of lysosomal storage diseases and their treatment. Biochemical Society Transactions. 2000;28(2):150-4.
2. Tomatsu S YE, Patel P, et al. Morquio A syndrome: diagnosis and current and future therapies. Pediatric endocrinology reviews 2014;12:141-51.
3. Starzyk K, Richards S, Yee J, Smith SE, Kingma W. The long-term international safety experience of imiglucerase therapy for Gaucher disease. Molecular Genetics and Metabolism. 2007;90(2):157-63.
4. Dandana A, Ben Khelifa S, Chahed H, Miled A, Ferchichi S. Gaucher Disease: Clinical, Biological and Therapeutic Aspects. Pathobiology. 2015;83(1):13-23.
5. Kazazian H. Mobile elements: drivers of genome evolution. Science. 2004;303(5664):1626-32.
6. Lopez M, Perez G. DNA transposons: nature and applications in genomics. Current genomics. 2010;11(2):115-28.
7. Ivics Z, Li MA, Mátés L, Boeke JD, Nagy A, Bradley A, et al. Transposon-mediated genome manipulation in vertebrates. Nature methods. 2009;6(6):415-22.
8. Sandoval-Villegas N, Nurieva W, Amberger M, Ivics Z. Contemporary Transposon Tools: A Review and Guide through Mechanisms and Applications of Sleeping Beauty, piggyBac and Tol2 for Genome Engineering. International journal of molecular sciences. 2021;22(10).
9. Cary LC, Goebel M, Corsaro BG, Wang HG, Rosen E, Fraser MJ. Transposon mutagenesis of baculoviruses: analysis of Trichoplusia ni transposon IFP2 insertions within the FP-locus of nuclear polyhedrosis viruses. Virology. 1989;172(1):156-69.
10. Ding S, Wu X, Li G, Han M, Zhuang Y, Xu T. Efficient transposition of the piggyBac (PB) transposon in mammalian cells and mice. Cell. 2005;122(3):473-83.
11. Nakanishi H HY, Kawakami S, et al. piggyBac transposon-mediated long-term gene expression in mice. Mol Ther 2010;18(4):707-14.
12. Elick TA, Bauser CA, Fraser MJ. Excision of the piggyBac transposable element in vitro is a precise event that is enhanced by the expression of its encoded transposase. Genetica. 1996;98(1):33-41.
13. Li MA, Turner DJ, Ning Z, Yusa K, Liang Q, Eckert S, et al. Mobilization of giant piggyBac transposons in the mouse genome. Nucleic acids research. 2011;39(22):e148.
14. Lee CY LJ, Liou JS, et al. . A gene delivery system for human cells mediated by both a cell-penetrating peptide and a piggyBac transposase. Biomaterials. 2011;32:6264-76.
15. Balasubramanian S, Matasci M, Kadlecova Z, Baldi L, Hacker DL, Wurm FM. Rapid recombinant protein production from piggyBac transposon-mediated stable CHO cell pools. Journal of biotechnology. 2015;200:61-9.
16. Walsh G. Biopharmaceutical benchmarks 2014. Nature Biotechnology. 2014;32(10):992-1000.
17. Wurm F. Production of recombinant protein therapeutics in cultivated mammalian cells. Nat Biotechnol. 2004;22:1393-8.
18. Di Matteo M, Belay E, Chuah MK, Vandendriessche T. Recent developments in transposon-mediated gene therapy. Expert opinion on biological therapy. 2012;12(7):841-58.
19. Park TS HJ. piggyBac transposition into primordial germ cells is an efficient tool for transgenesis in chickens. . Proc Natl Acad Sci U S A. 2012;12(24):9337-41.
20. Li R ZY, Han M, et al. . piggyBac as a high-capacity transgenesis and gene-therapy vector in human cells and mice. Dis Model Mech. 2013;6:828-33.
21. Wilber A LJ, Tian X, et al. Efficient and stable transgene expression in human embryonic stem cells using transposon-mediated gene transfer. Stem Cells. 2007;25:2919-27.
22. Balasubramanian S. Recombinant CHO Cell Pool Generation Using piggyBac Transposon System. Methods in molecular biology (Clifton, NJ). 2018;1850:69-78.
23. Matasci M, Baldi L, Hacker DL, Wurm FM. The PiggyBac transposon enhances the frequency of CHO stable cell line generation and yields recombinant lines with superior productivity and stability. Biotechnology and bioengineering. 2011;108(9):2141-50.
24. Cosma MP, Pepe S, Annunziata I, Newbold RF, Grompe M, Parenti G, et al. The Multiple Sulfatase Deficiency Gene Encodes an Essential and Limiting Factor for the Activity of Sulfatases. Cell. 2003;113(4):445-56.
25. Dierks T, Schmidt B, Borissenko LV, Peng J, Preusser A, Mariappan M, et al. Multiple sulfatase deficiency is caused by mutations in the gene encoding the human C(alpha)-formylglycine generating enzyme. Cell. 2003;113(4):435-44.
26. Puentes-Tellez MA, Sánchez OF, Rojas-Rodriguez F and et al. Evaluation of HIV–1 derived lentiviral vectors as transductors of Mucopolysaccharidosis type IV a fibroblasts. Gene. 2021;780:145527.