Skip to main content
SpringerLink
Log in

Polyallylamine Derivatives: Novel NonToxic Transfection Agents

  • Protocol
  • First Online:
Non-Viral Gene Delivery Vectors

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1445))

Abstract

Cationic polymers have shown great potential for the delivery of proteins, nucleic acids forming complexes, called polyplexes. The most important issue in the context of using cationic polymers as carriers is the balance between the high transfection efficiency and low cytotoxicity. In this chapter, we report the preparation of polyallylamine derivatives mainly based on substitution of amino groups by glycidyltrimethylammonium chloride. The resulting polyplexes enhance the transfection of HeLa cell line without cytotoxic effects. Here, we describe methods for preparation and characterization of polyplexes using dynamic light scattering, ζ-potential measurements, gel retardation assay, and atomic force microscopy. Moreover, we provide protocols for the transfection of HeLa cell line by polyplexes, determination of their cytotoxicity, cell uptake, and intracellular trafficking.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 109.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Miguel MG, Pais AACC, Dias RS, Leal C, Rosa M, Lindman B (2003) DNA-cationic amphiphile interactions. Colloid Surface A 228(1-3):43–55

    Article  CAS  Google Scholar 

  2. Yang J, Liu HM, Zhang X (2014) Design, preparation and application of nucleic acid delivery carriers. Biotechnol Adv 32(4):804–817

    Article  CAS  PubMed  Google Scholar 

  3. Kwoh DY, Coffin CC, Lollo CP, Jovenal J, Banaszczyk MG, Mullen P, Phillips A, Amini A, Fabrycki J, Bartholomew RM, Brostoff SW, Carlo DJ (1999) Stabilization of poly-L-lysine/DNA polyplexes for in vivo gene delivery to the liver. Biochim Biophys Acta 1444(2):171–190

    Article  CAS  PubMed  Google Scholar 

  4. Agirre M, Zarate J, Ojeda E, Puras G, Desbrieres J, Pedraz JL (2014) Low molecular weight chitosan (LMWC)-based polyplexes for pDNA delivery: from bench to bedside. Polymers 6(6):1727–1755

    Article  Google Scholar 

  5. Singh D, Han SS, Shin EJ (2014) Polysaccharides as nanocarriers for therapeutic applications. J Biomed Nanotechnol 10(9):2149–2172

    Article  CAS  PubMed  Google Scholar 

  6. Pourianazar NT, Mutlu P, Gunduz U (2014) Bioapplications of poly(amidoamine) (PAMAM) dendrimers in nanomedicine. J Nanopart Res 16 (4):2342 1-38.

    Google Scholar 

  7. Bishop CJ, Ketola TM, Tzeng SY, Sunshine JC, Urtti A, Lemmetyinen H, Vuorimaa-Laukkanen E, Yliperttula M, Green JJ (2013) The effect and role of carbon atoms in poly(beta-amino ester)s for DNA binding and gene delivery. J Am Chem Soc 135(18):6951–6957

    Article  CAS  PubMed  Google Scholar 

  8. Godbey WT, Wu KK, Mikos AG (1999) Poly(ethylenimine) and its role in gene delivery. J Control Release 60(2-3):149–160

    Article  CAS  PubMed  Google Scholar 

  9. Lungwitz U, Breunig M, Blunk T, Gopferich A (2005) Polyethylenimine-based non-viral gene delivery systems. Eur J Pharm Biopharm 60(2):247–266

    Article  CAS  PubMed  Google Scholar 

  10. Wytrwal M, Leduc C, Sarna M, Goncalves C, Kepczynski M, Midoux P, Nowakowska M, Pichon C (2015) Gene delivery efficiency and intracellular trafficking of novel poly(allylamine) derivatives. Int J Pharm 478(1):372–382

    Article  CAS  PubMed  Google Scholar 

  11. Tang MX, Szoka FC (1997) The influence of polymer structure on the interactions of cationic polymers with DNA and morphology of the resulting complexes. Gene Ther 4(8):823–832

    Article  CAS  PubMed  Google Scholar 

  12. Khalil IA, Kogure K, Akita H, Harashima H (2006) Uptake pathways and subsequent intracellular trafficking in nonviral gene delivery. Pharmacol Rev 58(1):32–45

    Article  CAS  PubMed  Google Scholar 

  13. Pathak A, Patnaik S, Gupta KC (2009) Recent trends in non-viral vector-mediated gene delivery. Biotechnol J 4(11):1559–1572

    Article  CAS  PubMed  Google Scholar 

  14. Pichon C, Billiet L, Midoux P (2010) Chemical vectors for gene delivery: uptake and intracellular trafficking. Curr Opin Biotechnol 21(5):640–645

    Article  CAS  PubMed  Google Scholar 

  15. Parhamifar L, Larsen AK, Hunter AC, Andresen TL, Moghimi SM (2010) Polycation cytotoxicity: a delicate matter for nucleic acid therapy-focus on polyethylenimine. Soft Matter 6(17):4001–4009

    Article  CAS  Google Scholar 

  16. De Smedt SC, Demeester J, Hennink WE (2000) Cationic polymer based gene delivery systems. Pharm Res 17(2):113–126

    Article  PubMed  Google Scholar 

  17. Varkouhi AK, Scholte M, Storm G, Haisma HJ (2011) Endosomal escape pathways for delivery of biologicals. J Control Release 151(3):220–228

    Article  CAS  PubMed  Google Scholar 

  18. Bertin A (2014) Polyelectrolyte complexes of DNA and polycations as gene delivery vectors. Adv Polym Sci 256:103–195

    Article  CAS  Google Scholar 

  19. Wytrwal M, Bednar J, Nowakowska M, Wydro P, Kepczynski M (2014) Interactions of serum with polyelectrolyte-stabilized liposomes: Cryo-TEM studies. Colloid Surface B 120:152–159

    Article  CAS  Google Scholar 

  20. de Ilarduya CT, Sun Y, Duezguenes N (2010) Gene delivery by lipoplexes and polyplexes. Eur J Pharm Sci 40(3):159–170

    Article  Google Scholar 

  21. Cho YW, Kim JD, Park K (2003) Polycation gene delivery systems: escape from endosomes to cytosol. J Pharm Pharmacol 55(6):721–734

    Article  CAS  PubMed  Google Scholar 

  22. Pegg AE (2013) Toxicity of polyamines and their metabolic products. Chem Res Toxicol 26(12):1782–1800

    Article  CAS  PubMed  Google Scholar 

  23. Novo L, Rizzo LY, Golombek SK, Dakwar GR, Lou B, Remaut K, Mastrobattista E, van Nostrum CF, Jahnen-Dechent W, Kiessling F, Braeckmans K, Lammers T, Hennink WE (2014) Decationized polyplexes as stable and safe carrier systems for improved biodistribution in systemic gene therapy. J Control Release 195:162–175

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Min SH, Park KC, Yeom YI (2014) Chitosan-mediated non-viral gene delivery with improved serum stability and reduced cytotoxicity. Biotechnol Bioproc E 19(6):1077–1082

    Article  CAS  Google Scholar 

  25. Wytrwal M, Koczurkiewicz P, Wojcik K, Michalik M, Kozik B, Zylewski M, Nowakowska M, Kepczynski M (2014) Synthesis of strong polycations with improved biological properties. J Biomed Mater Res A 102(3):721–731

    Article  PubMed  Google Scholar 

  26. Bertrand E, Goncalves C, Billiet L, Gomez JP, Pichon C, Cheradame H, Midoux P, Guegan P (2011) Histidinylated linear PEI: a new efficient non-toxic polymer for gene transfer. Chem Commun 47(46):12547–12549

    Article  CAS  Google Scholar 

  27. Billiet L, Gomez JP, Berchel M, Jaffres PA, Le Gall T, Montier T, Bertrand E, Cheradame H, Guegan P, Mevel M, Pitard B, Benvegnu T, Lehn P, Pichon C, Midoux P (2012) Gene transfer by chemical vectors, and endocytosis routes of polyplexes, lipoplexes and lipopolyplexes in a myoblast cell line. Biomaterials 33(10):2980–2990

    Article  CAS  PubMed  Google Scholar 

  28. Nuutila J, Lilius EM (2005) Flow cytometric quantitative determination of ingestion by phagocytes needs the distinguishing of overlapping populations of binding and ingesting cells. Cytometry A 65A(2):93–102

    Article  Google Scholar 

  29. Monsigny M, Roche AC, Midoux P (1984) Uptake of Neoglycoproteins Via Membrane Lectin(S) of L1210 Cells Evidenced by Quantitative Flow Cytofluorometry and Drug Targeting. Biol Cell 51(2):187–196

    Article  CAS  PubMed  Google Scholar 

  30. Midoux P, Roche AC, Monsigny M (1987) Quantitation of the binding, uptake, and degradation of fluoresceinylated neoglycoproteins by flow-cytometry. Cytometry 8(3):327–334

    Article  CAS  PubMed  Google Scholar 

  31. Goncalves C, Mennesson E, Fuchs R, Gorvel JP, Midoux P, Pichon C (2004) Macropinocytosis of polyplexes and recycling of plasmid from clathrin-dependent pathway impair the transfection efficiency into human hepatocarcinoma cells. Mol Ther 10:373–385

    Article  CAS  PubMed  Google Scholar 

  32. Steinman RM, Mellman IS, Muller WA, Cohn ZA (1983) Endocytosis and the recycling of plasma-membrane. J Cell Biol 96(1):1–27

    Article  CAS  PubMed  Google Scholar 

  33. Erbacher P, Roche AC, Monsigny M, Midoux P (1996) Putative role of chloroquine in gene transfer into a human hepatoma cell line by DNA lactosylated polylysine complexes. Exp Cell Res 225(1):186–194

    Article  CAS  PubMed  Google Scholar 

  34. de Bruin K, Ruthardt N, von Gersdorff K, Bausinger R, Wagner E, Ogris M, Brauchle C (2007) Cellular dynamics of EGF receptor-targeted synthetic viruses. Mol Ther 15(7):1297–1305

    Article  PubMed  Google Scholar 

  35. Breuzard G, Tertil M, Goncalves C, Cheradame H, Geguan P, Pichon C, Midoux P (2008) Nuclear delivery of N kappa B-assisted DNA/polymer complexes: plasmid DNA quantitation by confocal laser scanning microscopy and evidence of nuclear polyplexes by FRET imaging. Nucleic Acids Res 36(12), e17

    Article  Google Scholar 

Download references

Acknowledgment

This work was supported by the National Science Centre (NCN) Poland (Grant No. 2012/05/N/ST5/00809).

Author information

Authors and Affiliations

  1. Academic Centre for Materials and Nanotechnology, AGH University of Science and Technology, al. Mickiewicza 30, 30-059, Krakow, Poland

    Magdalena Wytrwal

  2. Department of Physical Chemistry and Electrochemistry, Faculty of Chemistry, Jagiellonian University, Ingardena 3, 30-060, Kraków, Poland

    Magdalena Wytrwal

  3. Centre de Biophysique Moléculaire, UPR4301 CNRS affiliated to the University of Orléans, Rue Charles Sadron, 45071, Orléans Cedex 2, France

    Chantal Pichon

Authors
  1. Magdalena Wytrwal
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Chantal Pichon
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Magdalena Wytrwal or Chantal Pichon .

Editor information

Editors and Affiliations

  1. Polytechnic University of Milan, Milan, Italy

    Gabriele Candiani

Rights and permissions

Reprints and permissions

Copyright information

© 2016 Springer Science+Business Media New York

About this protocol

Cite this protocol

Wytrwal, M., Pichon, C. (2016). Polyallylamine Derivatives: Novel NonToxic Transfection Agents. In: Candiani, G. (eds) Non-Viral Gene Delivery Vectors. Methods in Molecular Biology, vol 1445. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3718-9_10

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-3718-9_10

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3716-5

  • Online ISBN: 978-1-4939-3718-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation