Gene Therapy pp 47-137 | Cite as

Methods for Gene Delivery

  • Mauro Giacca


The success of any gene transfer procedure, either through in vivo inoculation of the genetic material or after gene transfer into the patient’s cells ex vivo, strictly depends upon the efficiency of nucleic acid internalization by the target cells. As a matter of fact, making gene transfer more efficient continues to represent the most relevant challenge to the clinical success of gene therapy.


Gene Delivery Bovine Leukemia Virus Helper Virus Immediate Early Packaging Cell Line 
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3.1 Cellular Barriers to Gene Delivery

Further Reading

  1. Conner SD, Schmid SL (2003) Regulated portals of entry into the cell. Nature 422:37–44PubMedGoogle Scholar
  2. Doherty GJ, McMahon HT (2009) Mechanisms of endocytosis. Annu Rev Biochem 78:857–902PubMedGoogle Scholar
  3. Sandvig K, van Deurs B (2005) Delivery into cells: lessons learned from plant and bacterial toxins. Gene Ther 12:865–872PubMedGoogle Scholar

Selected Bibliography

  1. Kerr MC, Teasdale RD (2009) Defining macropinocytosis. Traffic 10:364–371PubMedGoogle Scholar
  2. Kirkham M, Parton RG (2005) Clathrin-independent endocytosis: new insights into caveolae and non-caveolar lipid raft carriers. Biochim Biophys Acta 1745:273–286PubMedGoogle Scholar
  3. Medina-Kauwe LK (2007) “Alternative” endocytic mechanisms exploited by pathogens: new avenues for therapeutic delivery? Adv Drug Deliv Rev 59:798–809PubMedGoogle Scholar
  4. Nichols B (2003) Caveosomes and endocytosis of lipid rafts. J Cell Sci 116:4707–4714PubMedGoogle Scholar
  5. Pelkmans L, Puntener D, Helenius A (2002) Local actin polymerization and dynamin recruitment in SV40-induced internalization of caveolae. Science 296:535–539PubMedGoogle Scholar
  6. Plemper RK, Wolf DH (1999) Retrograde protein translocation: ERADication of secretory proteins in health and disease. Trends Biochem Sci 24:266–270PubMedGoogle Scholar
  7. Roth MG (2006) Clathrin-mediated endocytosis before fluorescent proteins. Nat Rev Mol Cell Biol 7:63–68PubMedGoogle Scholar
  8. Torgersen ML, Skretting G, van Deurs B, Sandvig K (2001) Internalization of cholera toxin by different endocytic mechanisms. J Cell Sci 114:3737–3747PubMedGoogle Scholar

Direct Inoculation of DNAs and RNAs Selected Bibliography

  1. Braun S (2008) Muscular gene transfer using nonviral vectors. Curr Gene Ther 8:391–405PubMedGoogle Scholar
  2. Herweijer H, Wolff JA (2003) Progress and prospects: naked DNA gene transfer and therapy. Gene Ther 10:453–458PubMedGoogle Scholar

Physical Methods Further Reading

  1. Frenkel V (2008) Ultrasound mediated delivery of drugs and genes to solid tumors. Adv Drug Deliv Rev 60:1193–1208PubMedGoogle Scholar
  2. Hynynen K (2008) Ultrasound for drug and gene delivery to the brain. Adv Drug Deliv Rev 60:1209–1217PubMedGoogle Scholar
  3. Wells DJ (2004) Gene therapy progress and prospects: electroporation and other physical methods. Gene Ther 11:1363–1369PubMedGoogle Scholar

Selected Bibliography

  1. Andre F, Mir LM (2004) DNA electrotransfer: its principles and an updated review of its therapeutic applications. Gene Ther 11[Suppl 1]:S33–42Google Scholar
  2. Bigey P, Bureau MF, Scherman D (2002) In vivo plasmid DNA electrotransfer. Curr Opin Biotechnol 13:443–447PubMedGoogle Scholar
  3. Hagstrom JE (2003) Plasmid-based gene delivery to target tissues in vivo: the intravascular approach. Curr Opin Mol Ther 5:338–344PubMedGoogle Scholar
  4. Heller LC, Heller R (2006) In vivo electroporation for gene therapy. Hum Gene Ther 17:890–897PubMedGoogle Scholar
  5. Lewis DL, Wolff JA (2007) Systemic siRNA delivery via hydrodynamic intravascular injection. Adv Drug Deliv Rev 59:115–123PubMedGoogle Scholar
  6. Mennuni C, Calvaruso F, Zampaglione I et al (2002) Hyaluronidase increases electrogene transfer efficiency in skeletal muscle. Hum Gene Ther 13:355–365PubMedGoogle Scholar
  7. Mir LM (2008) Application of electroporation gene therapy: past, current, and future. Methods Mol Biol 423:3–17PubMedGoogle Scholar
  8. Newman CM, Bettinger T (2007) Gene therapy progress and prospects: ultrasound for gene transfer. Gene Ther 14:465–475PubMedGoogle Scholar
  9. Reed SD, Li S (2009) Electroporation advances in large animals. Curr Gene Ther (in press)Google Scholar
  10. Stein U, Walther W, Stege A et al (2008) Complete in vivo reversal of the multidrug resistance phenotype by jet-injection of anti-MDR1 short hairpin RNA-encoding plasmid DNA. Mol Ther 16:178–186PubMedGoogle Scholar
  11. Walther W, Siegel R, Kobelt D et al (2008) Novel jet-injection technology for nonviral intratumoral gene transfer in patients with melanoma and breast cancer. Clin Cancer Res 14:7545–7553PubMedGoogle Scholar
  12. Walther W, Stein U, Fichtner I et al (2001) Nonviral in vivo gene delivery into tumors using a novel low volume jet-injection technology. Gene Ther 8:173–180PubMedGoogle Scholar
  13. Walther W, Stein U, Fichtner I et al (2002) Intratumoral low-volume jet-injection for efficient nonviral gene transfer. Mol Biotechnol 21:105–115PubMedGoogle Scholar

Chemical Methods Further Reading

  1. Beerens AM, Al Hadithy AF, Rots MG, Haisma HJ (2003) Protein transduction domains and their utility in gene therapy. Curr Gene Ther 3:486–494PubMedGoogle Scholar
  2. Elouahabi A, Ruysschaert JM (2005) Formation and intracellular trafficking of lipoplexes and polyplexes. Mol Ther 11:336–347PubMedGoogle Scholar
  3. Fittipaldi A, Giacca M (2005) Transcellular protein transduction using the Tat protein of HIV-1. Adv Drug Deliv Rev 57:597–608PubMedGoogle Scholar
  4. Giacca M (2004) The HIV-1 Tat protein: a multifaceted target for novel therapeutic opportunities. Curr Drug Targets Immune Endocr Metabol Disord 4:277–285PubMedGoogle Scholar
  5. Park TG, Jeong JH, Kim SW (2006) Current status of polymeric gene delivery systems. Adv Drug Deliv Rev 58:467–486PubMedGoogle Scholar
  6. van Dillen IJ, Mulder NH, Vaalburg W et al (2002) Influence of the bystander effect on HSVtk/GCV gene therapy. A review. Curr Gene Ther 2:307–322PubMedGoogle Scholar
  7. Vile RG, Russell SJ, Lemoine NR (2000) Cancer gene therapy: hard lessons and new courses. Gene Ther 7:2–8PubMedGoogle Scholar
  8. Wasungu L, Hoekstra D (2006) Cationic lipids, lipoplexes and intracellular delivery of genes. J Control Release 116:255–264PubMedGoogle Scholar

Selected Bibliography

  1. Dass CR (2004) Lipoplex-mediated delivery of nucleic acids: factors affecting in vivo transfection. J Mol Med 82:579–591PubMedGoogle Scholar
  2. Dincer S, Turk M, Piskin E (2005) Intelligent polymers as nonviral vectors. Gene Ther 12[Suppl1]:S139–145Google Scholar
  3. Dufes C, Uchegbu IF, Schatzlein AG (2005) Dendrimers in gene delivery. Adv Drug Deliv Rev 57:2177–2202PubMedGoogle Scholar
  4. Duncan R, Izzo L (2005) Dendrimer biocompatibility and toxicity. Adv Drug Deliv Rev 57:2215–2237PubMedGoogle Scholar
  5. Fittipaldi A, Giacca M (2005) Transcellular protein transduction using the Tat protein of HIV-1. Adv Drug Deliv Rev 57:597–608PubMedGoogle Scholar
  6. Hoekstra D, Rejman J, Wasungu L et al (2007) Gene delivery by cationic lipids: in and out of an endosome. Biochem Soc Trans 35:68–71PubMedGoogle Scholar
  7. Lange, A, Mills RE, Lange CJ et al (2007) Classical nuclear localization signals: definition, function, and interaction with importin alpha. J Biol Chem 282:5101–5105PubMedGoogle Scholar
  8. Pedroso de Lima MC, Simoes S, Pires P et al (2001) Cationic lipid-DNA complexes in gene delivery: from biophysics to biological applications. Adv Drug Deliv Rev 47:277–294PubMedGoogle Scholar
  9. Rainov NG (2000) A phase III clinical evaluation of herpes simplex virus type 1 thymidine kinase and ganciclovir gene therapy as an adjuvant to surgical resection and radiation in adults with previously untreated glioblastoma multiforme. Hum Gene Ther 11:2389–2401PubMedGoogle Scholar
  10. Ram Z, Culver KW, Oshiro EM et al (1997) Therapy of malignant brain tumors by intratumoral implantation of retroviral vector-producing cells. Nat Med 3:1354–1361PubMedGoogle Scholar
  11. Svenson S (2009) Dendrimers as versatile platform in drug delivery applications. Eur J Pharm Biopharm 71:445–462PubMedGoogle Scholar
  12. Trask TW, Trask RP, Aguilar-Cordova E et al (2000) Phase I study of adenoviral delivery of the HSV-tk gene and ganciclovir administration in patients with current malignant brain tumors. Mol Ther 1:195–203PubMedGoogle Scholar
  13. Wolff JA, Rozema DB (2008) Breaking the bonds: non-viral vectors become chemically dynamic. Mol Ther 16:8–15PubMedGoogle Scholar
  14. Zuhorn IS, Engberts JB, Hoekstra D (2007) Gene delivery by cationic lipid vectors: overcoming cellular barriers. Eur Biophys J 36:349–362PubMedGoogle Scholar

Viral Vectors Further Reading

  1. Bessis N, GarciaCozar FJ, Boissier MC (2004) Immune responses to gene therapy vectors: influence on vector function and effector mechanisms. Gene Ther 11[Suppl 1]:S10–17Google Scholar
  2. Bestor TH (2000) Gene silencing as a threat to the success of gene therapy. J Clin Invest 105:409–411PubMedGoogle Scholar
  3. Chang AH, Sadelain M (2007) The genetic engineering of hematopoietic stem cells: the rise of lentiviral vectors, the conundrum of the ltr, and the promise of lineage-restricted vectors. Mol Ther 15:445–456PubMedGoogle Scholar
  4. Coffin JM, Hughes H, Varmus HE (1997) Retroviruses. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, NY, USAGoogle Scholar
  5. Daniel R, Smith JA (2008) Integration site selection by retroviral vectors: molecular mechanism and clinical consequences. Hum Gene Ther 19:557–568PubMedGoogle Scholar
  6. Kay MA, Glorioso JC, Naldini L (2001) Viral vectors for gene therapy: the art of turning infectious agents into vehicles of therapeutics. Nat Med 7:33–40PubMedGoogle Scholar
  7. Knipe DM, Roizman B, Howley PM et al (2006) Fields’ virology, 5th edn. Lippincott Williams & Wilkins, Philadelphia, PA, USAGoogle Scholar
  8. Schambach A, Baum C (2008) Clinical application of lentiviral vectors: concepts and practice. Curr Gene Ther 8:474–482PubMedGoogle Scholar
  9. St George JA (2003) Gene therapy progress and prospects: adenoviral vectors. Gene Ther 10:1135–1141Google Scholar
  10. Thomas CE, Ehrhardt A, Kay MA (2003) Progress and problems with the use of viral vectors for gene therapy. Nat Rev Genet 4:346–358PubMedGoogle Scholar
  11. Wu Z, Asokan A, Samulski RJ (2006) Adeno-associated virus serotypes: vector toolkit for human gene therapy. Mol Ther 14:316–327PubMedGoogle Scholar
  12. Zentilin L, Giacca M (2008) Adeno-associated virus vectors: versatile tools for in vivo gene transfer. Contrib Nephrol 159:63–77PubMedGoogle Scholar

Selected Bibliography

  1. Aghi M, Martuza RL (2005) Oncolytic viral therapies: the clinical experience. Oncogene 24:7802–7816PubMedGoogle Scholar
  2. Aiken C (1997) Pseudotyping human immunodeficiency virus type 1 (HIV-1) by the glycoprotein of vesicular stomatitis virus targets HIV-1 entry to an endocytic pathway and suppresses both the requirement for Nef and the sensitivity to cyclosporin A. J Virol 71:5871–5877PubMedGoogle Scholar
  3. Alba R, Bosch A, Chillon M (2005) Gutless adenovirus: last-generation adenovirus for gene therapy. Gene Ther 12[Suppl 1]:S18–27Google Scholar
  4. Argnani R, Lufino M, Manservigi M, Manservigi R (2005) Replication-competent herpes simplex vectors: design and applications. Gene Ther 12[Suppl 1]:S170–177Google Scholar
  5. Barnard RJ, Elleder D, Young JA (2006) Avian sarcoma and leukosis virus-receptor interactions: from classical genetics to novel insights into virus-cell membrane fusion. Virology 344:25–29PubMedGoogle Scholar
  6. Barquinero J, Eixarch H, Perez-Melgosa M (2004) Retroviral vectors: new applications for an old tool. Gene Ther 11[Suppl 1]:S3–9Google Scholar
  7. Berges BK, Wolfe JH, Fraser NW (2007) Transduction of brain by herpes simplex virus vectors. Mol Ther 15:20–29PubMedGoogle Scholar
  8. Berns KI, Linden RM (1995) The cryptic life style of adeno-associated virus. BioEssays 17:237–245PubMedGoogle Scholar
  9. Berto E, Bozac A, Marconi P (2005) Development and application of replication-incompetent HSV-1-based vectors. Gene Ther 12[Suppl 1]:S98–102Google Scholar
  10. Brunetti-Pierri N, Ng P (2008) Progress and prospects: gene therapy for genetic diseases with helper-dependent adenoviral vectors. Gene Ther 15:553–560PubMedGoogle Scholar
  11. Buning H, Ried MU, Perabo L et al (2003) Receptor targeting of adeno-associated virus vectors. Gene Ther 10:1142–1151PubMedGoogle Scholar
  12. Burton EA, Bai Q, Goins WF, Glorioso JC (2002) Replication-defective genomic herpes simplex vectors: design and production. Curr Opin Biotechnol 13:424–428PubMedGoogle Scholar
  13. Cereseto A, Giacca M (2004) Integration site selection by retroviruses. AIDS Rev 6:13–20PubMedGoogle Scholar
  14. Cervelli T, Palacios JA, Zentilin L et al (2008) Processing of recombinant AAV genomes occurs in specific nuclear structures that overlap with foci of DNA-damage-response proteins. J Cell Sci 121:349–357PubMedGoogle Scholar
  15. Chirmule N, Propert K, Magosin S et al (1999) Immune responses to adenovirus and adeno-associated virus in humans. Gene Ther 6:1574–1583PubMedGoogle Scholar
  16. Danthinne X, Imperiale MJ (2000) Production of first generation adenovirus vectors: a review. Gene Ther 7:1707–1714PubMedGoogle Scholar
  17. Dull T, Zufferey R, Kelly M et al (1998) A third-generation lentivirus vector with a conditional packaging system. J Virol 72:8463–8471PubMedGoogle Scholar
  18. Dutheil N, Shi F, Dupressoir T, Linden RM (2000) Adeno-associated virus site-specifically integrates into a muscle-specific DNA region. Proc Natl Acad Sci U S A 97:4862–4866PubMedGoogle Scholar
  19. Epstein AL (2005) HSV-1-based amplicon vectors: design and applications. Gene Ther 12[Suppl 1]:S154–158Google Scholar
  20. Flotte TR (2004) Gene therapy progress and prospects: recombinant adeno-associated virus (rAAV) vectors. Gene Ther 11:805–810PubMedGoogle Scholar
  21. Flotte TR, Berns KI (2005) Adeno-associated virus: a ubiquitous commensal of mammals. Hum Gene Ther 16:401–407PubMedGoogle Scholar
  22. Gao G, Vandenberghe LH, Wilson JM (2005) New recombinant serotypes of AAV vectors. Curr Gene Ther 5:285–297PubMedGoogle Scholar
  23. Gregorevic P, Blankinship MJ, Allen JM et al (2004) Systemic delivery of genes to striated muscles using adeno-associated viral vectors. Nat Med 10:828–834PubMedGoogle Scholar
  24. Grimm D, Kern A, Rittner K, Kleinschmidt JA (1998) Novel tools for production and purification of recombinant adenoassociated virus vectors Hum. Gene Ther 9:2745–2760Google Scholar
  25. Hendrie PC, Russell DW (2005) Gene targeting with viral vectors. Mol Ther 12:9–17PubMedGoogle Scholar
  26. Jooss K, Chirmule N (2003) Immunity to adenovirus and adeno-associated viral vectors: implications for gene therapy. Gene Ther 10:955–963PubMedGoogle Scholar
  27. Linden RM, Ward P, Giraud C et al (1996) Site-specific integration by adeno-associated virus. Proc Natl Acad Sci U S A 93:11288–11294PubMedGoogle Scholar
  28. Liu Q, Muruve DA (2003) Molecular basis of the inflammatory response to adenovirus vectors. Gene Ther 10:935–940PubMedGoogle Scholar
  29. Manganaro L, Lusic M, Gutierrez MI et al (2009) Concerted action of cellular JNK and Pin1 restricts HIV-1 genome integration to activated CD4+ T lymphocytes. Nat MedGoogle Scholar
  30. Marconi P, Argnani R, Berto E et al (2008) HSV as a vector in vaccine development and gene therapy. Hum Vaccin 4:91–105PubMedGoogle Scholar
  31. McCarty DM, Young SM Jr, Samulski RJ (2004) Integration of adeno-associated virus (AAV) and recombinant AAV vectors. Annu Rev Genet 38:819–845PubMedGoogle Scholar
  32. Merten OW, Geny-Fiamma C, Douar AM (2005) Current issues in adeno-associated viral vector production. Gene Ther 12[Suppl 1]:S51–61Google Scholar
  33. Miller AD (1990) Retrovirus packaging cells. Hum Gene Ther 1:5–14PubMedGoogle Scholar
  34. Miller AD (1996) Cell-surface receptors for retroviruses and implications for gene transfer. Proc Natl Acad Sci U S A 93:11407–11413PubMedGoogle Scholar
  35. Miller DG, Wang PR, Petek LM et al (2006) Gene targeting in vivo by adeno-associated virus vectors. Nature Biotech 24:1022–1026Google Scholar
  36. Mueller C, Flotte TR (2008) Clinical gene therapy using recombinant adeno-associated virus vectors. Gene Ther 15:858–863PubMedGoogle Scholar
  37. Naldini L, Blomer U, Gallay P et al (1996) In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector. Science 272:263–267PubMedGoogle Scholar
  38. Nienhuis AW, Dunbar CE, Sorrentino BP (2006) Genotoxicity of retroviral integration in hematopoietic cells. Mol Ther 13:1031–1049PubMedGoogle Scholar
  39. Russell DW, Hirata RK (1998) Human gene targeting by viral vectors. Nat Genet 18:325–330PubMedGoogle Scholar
  40. Russell WC (2000) Update on adenovirus and its vectors. J Gen Virol 81:2573–2604PubMedGoogle Scholar
  41. Russell WC (2009) Adenoviruses: update on structure and function. J Gen Virol 90:1–20PubMedGoogle Scholar
  42. Samulski RJ, Berns KI, Tan M, Muzyczka N (1982) Cloning of adeno-associated virus into pBR322: rescue of intact virus from the recombinant plasmid in human cells. Proc Natl Acad Sci USA 79:2077–2081PubMedGoogle Scholar
  43. Schroder AR, Shinn P, Chen H et al (2002) HIV-1 integration in the human genome favors active genes and local hotspots. Cell 110:521–529PubMedGoogle Scholar
  44. Sinn PL, Sauter SL, McCray PB Jr (2005) Gene therapy progress and prospects: development of improved lentiviral and retroviral vectors: design, biosafety, and production. Gene Ther 12:1089–1098PubMedGoogle Scholar
  45. Vasileva A, Jessberger R (2005) Precise hit: adeno-associated virus in gene targeting. Nat Rev Microbiol 3:837–847PubMedGoogle Scholar
  46. Vihinen-Ranta M, Suikkanen S, Parrish CR (2004) Pathways of cell infection by parvoviruses and adeno-associated viruses. J Virol 78:6709–6714PubMedGoogle Scholar
  47. Wang Z, Zhu T, Qiao C et al (2005) Adeno-associated virus serotype 8 efficiently delivers genes to muscle and heart. Nat Biotechnol 23:321–328PubMedGoogle Scholar
  48. Yamamoto T, Tsunetsugu-Yokota Y (2008) Prospects for the therapeutic application of lentivirusbased gene therapy to HIV-1 infection. Curr Gene Ther 8:1–8PubMedGoogle Scholar
  49. Zentilin L, Qin G, Tafuro S et al (2000) Variegation of retroviral vector gene expression in myeloid cells. Gene Ther 7:153–166PubMedGoogle Scholar
  50. Zufferey R, Dull T, Mandel RJ et al (1998) Self-inactivating lentivirus vector for safe and efficient in vivo gene delivery. J Virol 72:9873–9880PubMedGoogle Scholar

Copyright information

© Springer-Verlag Italia 2010

Authors and Affiliations

  • Mauro Giacca
    • 1
  1. 1.International Centre for Genetic Engineering and Biotechnology (ICGEB)TriesteItaly

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