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Physical Chemical and Biomolecular Methods for the Optimization of Cationic Lipid-Based Lipoplexes In Vitro for the Gene Therapy Applications

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Non-Viral Gene Delivery Vectors

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

Abstract

Preparation and application protocols play a very important role while optimizing the cationic lipid-based lipoplexes in vitro. These protocols serve as the basis for the betterment of the lipoplexes with regard to their successful application in animals and eventually human subjects. Starting from the chemical structures of used cationic lipids (CLs), optimization of the additive inclusions, methods of nanoparticle (lipoplex) formation, presence of blood serum, time intervals of lipoplex incubation, and type of efficiency read-outs in various conditions play important roles in reaching insightful conclusions. Such steps of summarizing protocols and requirements of the pertinent events focus on getting improved lipoplexes for achieving optimal effects in terms of post transfection gene and protein expression. The progression of optimization and efficiency evaluation lead to predictable structure-method-activity relationship with involvement of various feedback principles including physical chemical and biomolecular evaluations before and after the use of lipoplexes in biological systems. This chapter discusses some of the focused strategies for the establishment of lipoplexes for a better post transfection activity with reduced risk of failure.

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References

  1. “Disease” at Dorland’s Medical Dictionary

    Google Scholar 

  2. “Mental Illness—Glossary”. US National Institute of Mental Health. Retrieved 18 Apr 2010

    Google Scholar 

  3. Anderson WF (1998) Human gene therapy. Nature 392:25–30

    Article  CAS  PubMed  Google Scholar 

  4. Hattori Y, Maitani Y (2005) Folate-linked nanoparticle-mediated suicide gene therapy in human prostate cancer and nasopharyngeal cancer with herpes simplex virus thymidine kinase. Cancer Gene Ther 12:796–809

    Article  CAS  PubMed  Google Scholar 

  5. Chae HY, Lee BW, Oh SH, Ahn YR, Chung JH, Min YK, Lee MS, Lee MK, Kim KW (2005) Effective glycemic control achieved by transplanting non-viral cationic liposome-mediated VEGF-transfected islets in streptozotocin-induced diabetic mice. Exp Mol Med 37:513–523

    Article  CAS  PubMed  Google Scholar 

  6. Yang Y, Nunes FA, Berencsi K, Goenczoel E, Engelhardt JF, Wilson JM (1994) Inactivation of E2a in recombinant adenoviruses improves the prospect for gene therapy in cystic fibrosis. Nat Genet 7:362–369

    Article  CAS  PubMed  Google Scholar 

  7. Curiel TJ, Piche A, Kasono K, Curiel DT (1997) Gene therapy strategies for AIDS-related malignancies. Gene Ther 4:1284–1288

    Article  CAS  PubMed  Google Scholar 

  8. Yla-Herttuala S, Martin JF (2000) Cardiovascular gene therapy. Lancet 355:|213–222

    Article  CAS  PubMed  Google Scholar 

  9. Vasir JK, Labhestwar V (2007) Biodegradable nanoparticles for cytosolic delivery of therapeutics. Adv Drug Deliv Rev 59:718–728

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Ow H, Larson DR, Srivastava M, Baird BA, Webb WW, Wiesner U (2005) Bright and stable core − shell fluorescent silica nanoparticles. Nano Lett 5:113–117

    Article  CAS  PubMed  Google Scholar 

  11. Jin S, Ye K (2007) Nanoparticle-mediated drug delivery and gene therapy. Biotechnol Prog 23:32–41

    Article  CAS  PubMed  Google Scholar 

  12. Breyer B, Jiang W, Cheng H, Zhou L, Paul R, Feng T, He TC (2001) Adenoviral vector-mediated gene transfer for human gene therapy. Curr Gene Ther 1:149–162

    Google Scholar 

  13. Anderson WF (1984) Prospects for human gene therapy. Science 226:401–409

    Article  CAS  PubMed  Google Scholar 

  14. Robbins PD, Tahara H, Ghivizzani SC (1998) Viral vectors for gene therapy. Trends Biotechnol 16:35–40

    Article  CAS  PubMed  Google Scholar 

  15. Felgner PL, Gadek TR, Holm M, Roman R, Chan HW, Wenz M, Northrop JP, Ringold GM, Danielsen M (1987) Lipofection: a highly efficient, lipid-mediated DNA-transfection procedure. Proc Natl Acad Sci U S A 84:7413–7417

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Bhattacharya S, Bajaj A (2009) Advances in gene delivery through molecular design of cationic lipids. Chem Commun 31:4632–4656

    Article  Google Scholar 

  17. Bhattacharya S, De S (1999) Synthesis and vesicle formation from dimeric pseudoglyceryl lipids with (CH2)m spacers: pronounced m‐value dependence of thermal properties, vesicle fusion, and cholesterol complexation. Chem Eur J 5:2335–2347

    Article  CAS  Google Scholar 

  18. Dileep PV, Antony A, Bhattacharya S (2001) Incorporation of oxyethylene units between hydrocarbon chain and pseudoglyceryl backbone in cationic lipid potentiates gene transfection efficiency in the presence of serum. FEBS Lett 509:327–331

    Article  CAS  PubMed  Google Scholar 

  19. Bhattacharya S, Dileep PV (2004) Cationic oxyethylene lipids. Synthesis, aggregation, and transfection properties. Bioconjugate Chem 15:508–519

    Article  CAS  Google Scholar 

  20. Gao X, Huang L (1991) A novel cationic liposome reagent for efficient transfection of mammalian cells. Biochim Biophys Res Commun 179:280–285

    Article  CAS  Google Scholar 

  21. Ghosh YK, Visweswariah SS, Bhattacharya S (2000) Nature of linkage between the cationic headgroup and cholesteryl skeleton controls gene transfection efficiency. FEBS Lett 473:341–344

    Article  CAS  PubMed  Google Scholar 

  22. Hasegawa S, Hirashima N, Nakanishi M (2002) Comparative study of transfection efficiency of cationic cholesterols mediated by liposomes-based gene delivery. Bioorg Med Chem Lett 12:1299–1302

    Article  CAS  PubMed  Google Scholar 

  23. Nakanishi M (2003) Strategy in gene transfection by cationic transfection lipids with a cationic cholesterol. Curr Med Chem 10:1289–1296

    Article  CAS  PubMed  Google Scholar 

  24. Bajaj A, Mishra SK, Kondaiah P, Bhattacharya S (2008) Effect of the headgroup variation on the gene transfer properties of cholesterol based cationic lipids possessing ether linkage. Biochim Biophys Acta 1778:1222–1236

    Article  CAS  PubMed  Google Scholar 

  25. Bajaj A, Kondaiah P, Bhattacharya S (2008) Effect of the nature of the spacer on gene transfer efficacies of novel thiocholesterol derived gemini lipids in different cell lines: a structure–activity investigation. J Med Chem 51:2533–2540

    Article  CAS  PubMed  Google Scholar 

  26. Kumar K, Maiti B, Kondaiah P, Bhattacharya S (2015) Efficacious gene silencing in serum and significant apoptotic activity induction by survivin downregulation mediated by new cationic gemini tocopheryl lipids. Mol Pharm 12:351–361

    Article  CAS  PubMed  Google Scholar 

  27. Hirsch-Lerner D, Min Z, Eliyahu H, Ferrari ME, Wheeler CJ, Barenholz Y (2005) Effect of “helper lipid” on lipoplex electrostatics. Biochim Biophys Acta 1714:71–84

    Article  CAS  PubMed  Google Scholar 

  28. Kerner M, Meyuhas O, Hirsch-Lerner D, Rosen L, Min Z, Barenholz Y (2001) Interplay in lipoplexes between type of pDNA promoter and lipid composition determines transfection efficiency of human growth hormone in NIH3T3 cells in culture. Biochim Biophys Acta 1532:128–136

    Article  CAS  PubMed  Google Scholar 

  29. Simberg D, Danino D, Talmon Y, Minsky A, Ferrari M, Wheeler CJ, Barenholz Y (2001) Phase behavior, DNA ordering, and size instability of cationic lipoplexes. Relevance to optimal transfection activity. J Biol Chem 276:47453–47459

    Article  CAS  PubMed  Google Scholar 

  30. Candiani G, Pezzoli D, Ciani L, Chiesa L, Ristori S (2010) Bioreducible liposomes for gene delivery: from the formulation to the mechanism of action. PLoS One 5:e13430

    Article  PubMed  PubMed Central  Google Scholar 

  31. Sarah R, Paul P, Alain TR (2009) Physico-chemical characteristics of lipoplexes influence cell uptake mechanisms and transfection efficacy. PLoS One 4:e6058

    Article  Google Scholar 

  32. Hong K, Zheng W, Baker A, Papahadjopoulos D (1997) Stabilization of cationic liposome-plasmid DNA complexes by polyamines and poly(ethylene glycol)-phospholipid conjugates for efficient in vivo gene delivery. FEBS Lett 400:233–237

    Article  CAS  PubMed  Google Scholar 

  33. Bhattacharya S, Mandal SS (1998) Evidence of interlipidic ion-pairing in anion induced DNA release from cationic amphiphile-DNA complexes. Mechanistic implications in transfection. Biochemistry 37:7764–7777

    Article  CAS  PubMed  Google Scholar 

  34. Misra SK, Biswas J, Kondaiah P, Bhattacharya S (2013) Gene transfection in high serum levels: case studies with new cholesterol based cationic gemini lipids. PLoS One 8:e68305

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Authors and Affiliations

  1. Department of Organic Chemistry, Indian Institute of Science, Bangalore, 560 012, India

    Santosh K. Misra & Santanu Bhattacharya

  2. Department of Bioengineering, University of Illinois at Urbana-Champaign, Champaign, IL, 61801, USA

    Santosh K. Misra

  3. Indian Association for the Cultivation of Science, Jadavpur, Kolkata, 700032, India

    Santanu Bhattacharya

Authors
  1. Santosh K. Misra
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  2. Santanu Bhattacharya
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Correspondence to Santanu Bhattacharya .

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Editors and Affiliations

  1. Polytechnic University of Milan, Milan, Italy

    Gabriele Candiani

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Misra, S.K., Bhattacharya, S. (2016). Physical Chemical and Biomolecular Methods for the Optimization of Cationic Lipid-Based Lipoplexes In Vitro for the Gene Therapy Applications. 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_1

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  • DOI: https://doi.org/10.1007/978-1-4939-3718-9_1

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  • 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

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