Biosafety in Handling Gene Transfer Vectors

Scott Swindle1

1 Occupational Medicine and Research Safety, The University of Alabama at Birmingham, Birmingham
Publication Name:  Current Protocols in Human Genetics
Unit Number:  Unit 12.1
DOI:  10.1002/cphg.54
Online Posting Date:  January, 2018
GO TO THE FULL TEXT: PDF or HTML at Wiley Online Library


This unit is devoted to safety issues that must be considered when generating and working with the most common vectors under development for human gene therapy today. © 2018 by John Wiley & Sons, Inc.

Keywords: gene transfer; vectors; biosafety

PDF or HTML at Wiley Online Library

Table of Contents

  • Introduction
  • Background
  • Running A BL‐2 Lab
  • Specific Vectors
  • CRISPR/CAS9‐Based Genome Editing
  • Getting a Useful Vector into Clinical Trials
  • Obtaining Biosafety Committee Approval
  • Further Reading
  • Acknowledgements
  • Literature Cited
  • Figures
  • Tables
PDF or HTML at Wiley Online Library


PDF or HTML at Wiley Online Library



Literature Cited

  Abken, H., Butzler, C., & Willecke, K. (1987). Adenovirus type 5 persisting in human lymphocytes is unlikely to be involved in immortalization of lymphoid cells by fusion with cytoplasts or by transfection with DNA of mouse L cells. Anticancer Research, 7, 553–558.
  Bassin, R. H., Tuttle, N., & Fischinger, P. J. (1971). Rapid cell culture assay technique for murine leukaemia viruses. Nature, 229, 564–566. doi: 10.1038/229564b0.
  Beatty, M. S., & Curiel, D. T. (2012). Chapter two–Adenovirus strategies for tissue‐specific targeting. Advances in Cancer Research, 115, 39–67. doi: 10.1016/B978‐0‐12‐398342‐8.00002‐1.
  Bergelson, J. M., Cunningham, J. A., Droguett, G. et al. (1997). Isolation of a common receptor for Coxsackie B viruses and adenoviruses 2 and 5. Science, 275, 1320–1323. doi: 10.1126/science.275.5304.1320.
  Burns, P. A., Jack, A., Neilson, F., Haddow, S., & Balmain, A. (1991). Transformation of mouse skin endothelial cells in vivo by direct application of plasmid DNA encoding the human T24 h‐ras oncogene. Oncogene, 6, 1973–1978.
  Bushman, F., Lewinski, M., Ciuffi, A., Barr, S., Leipzig, J., Hannenhalli, S., & Hoffmann, C. (2005). Genome‐wide analysis of retroviral DNA integration. Nature Reviews. Microbiology, 3, 848–858. doi: 10.1038/nrmicro1263.
  Cattoglio, C., Facchini, G., Sartori, D., Antonelli, A., Miccio, A., Cassani, B., … Mavilio, F. (2007). Hot spots of retroviral integration in human CD34+ hematopoietic cells. Blood, 110(6), 1770–1778. doi: 10.1182/blood‐2007‐01‐068759.
  Chirmule, N., Propert, K., Magosin, S., Qian, Y., Qian, R., & Wilson, J. (1999). Immune responses to adenovirus and adeno‐associated virus in humans. Gene Therapy, 6(9), 1574–1583. doi: 10.1038/
  Cotten, M., Baker, A., Birnstiel, M. L., Zatloukal, K., & Wagner, E. (2001). Preparation of adenovirus‐polylysine‐DNA complexes. Current Protocols in Human Genetics, 11, 12.3.1–12.3.33. doi: 10.1002/0471142905.hg1203s11.
  Hardee, Cinnamon L., Arévalo‐Soliz, Lirio Milenka, Hornstein, Benjamin D., & Zechiedrich, Lynn (2017). Advances in non‐viral DNA vectors for gene therapy. Genes, 8, 65. doi: 10.3390/genes8020065.
  Crystal, R. G., McElvaney, N. G., Rosenfeld, M. A., Chu, C. S., Mastrangeli, A., Hay, J. G., … Danel, C. (1994). Administration of an adenovirus containing the human CFTR cDNA to the respiratory tract of individuals with cystic fibrosis. Nature Genetics, 8, 42–51. doi: 10.1038/ng0994‐42.
  Deyle, D. R., & Russel, D. W. (2009). Adeno‐ associated virus vector integration. Current Opinion in Molecular Therapy, 11(4), 442–447.
  Doerfler, W. (1991). Abortive infection and malignant transformation by adenoviruses: Integration of viral DNA and control of viral gene expression by specific patterns of DNA methylation. Advances in Virus Research, 39, 89–128. doi: 10.1016/S0065‐3527(08)60793‐9.
  Fallaux, F. J., Bout, A., van der Velde, I., van den Wollenberg, D. J. M., Hehir, K., Keegan, J., … Hoeben, R. C. (1998). New helper cells and matched early region 1‐deleted adenovirus vectors prevent generation of replication‐competent adenoviruses. Human Gene Therapy, 9, 1909–1917. doi: 10.1089/hum.1998.9.13‐1909.
  Flotte, T., Carter, B., Conrad, C., Guggino, W., Reynolds, T., Rosenstein, B., … Wetzel, R. (1996). A phase I study of an adeno‐associated virus‐CFTR gene vector in adult CF patients with mild lung disease. Human Gene Therapy, 7, 1145–1159. doi: 10.1089/hum.1996.7.9‐1145.
  Foreman, P. M., Friedman, G. K., Cassady, K. A., & Markert, J. M. (2017). Oncolytic Virotherapy for the Treatment of Malignant Glioma Neurotherapeutics, 14(2), 333–344. doi: 10.1007/s13311‐017‐0516‐0.
  Fraefel, C., Song, S., Lim, F., Lang, P., Yu, L., Wang, Y., … Geller, A. I. (1996). Helper virus‐free transfer of herpes simplex virus type 1 plasmid vectors into neural cells. Journal of Virology, 70, 7190–7197.
  Gaj, T1, Gersbach, C. A., & Barbas, C. F. III. (2013). ZFN, TALEN, and CRISPR/Cas‐based methods for genome engineering. Trends Biotechnol, 31(7), 397–405. doi: 10.1016/j.tibtech.2013.04.004.
  Gallimore, P. H. (1972). Tumor production in immunosupressed rats with cells transformed in vitro by Ad2. The Journal of General Virology, 62, 414–4126.
  Glover, D. J., Lipps, H. J., & Jans, D. A. (2005). Towards safe, non‐viral therapeutic gene expression in humans. Nature Reviews. Genetics, 6, 299–310. doi: 10.1038/nrg1577.
  Gómez, C. E., Nájera, J. L., Domingo‐Gil, E., Ochoa‐Callejero, L., González‐Aseguinolaza, G., & Esteban, M. (2007). Virus distribution of the attenuated MVA and NYVAC poxvirus strains in mice. The Journal of General Virology, 88(Pt 9), 2473–2478. doi: 10.1099/vir.0.83018‐0.
  Fox, J. P., Brandt, C. D., Wassermann, F. E., Hall, C. E., Spigland, I., Kogon, A., & Elveback, L. R. (1969). The virus watch program: A continuing surveillance of viral infections in metropolitan New York families. VI. Observations of adenovirus infections: Virus excretion patterns, antibody response, efficiency of surveillance, patterns of infections, and relation to illness. American Journal of Epidemiology, 89, 25–50. doi: 10.1093/oxfordjournals.aje.a120913.
  Harvey, B. G., Maroni, J., O'Donoghue, K. A., Chu, K. W., Muscat, J. C., Pippo, A. L., … Crystal, R. G. (2002). Safety of local delivery of low‐ and intermediate‐dose adenovirus gene transfer vectors to individuals with a spectrum of morbid conditions. Hum. Gene Therapy, 13, 15–63. doi: 10.1089/10430340152712638.
  Hacein‐Bey‐Abina, S., Le Deist, F., Carlier, F., Bouneaud, C., Hue, C., De Villartay, J. P., … Cavazzana‐Calvo, M. (2002). Sustained correction of X‐linked severe combined immunodeficiency by ex vivo gene therapy. The New England Journal of Medicine, 346, 1185–1193. doi: 10.1056/NEJMoa012616.
  Hacein‐Bey‐Abina, S., Von Kalle, C., Schmidt, M., McCormack, M. P., Wulffraat, N., Leboulch, P., … Cavazzana‐Calvo, M. (2003). LMO2‐associated clonal T cell proliferation in two patients after gene therapy for SCID‐X1. Science, 302(5644), 415–419. doi: 10.1126/science.1088547.
  Hacein‐Bey‐Abina, S., Pai, S. Y., Gaspar, H. B., Armant, M., Berry, C. C., Blanche, S., … Thrasher, A. J. (2014). A modified γ‐retrovirus vector for X‐linked severe combined immunodeficiency. The New England Journal of Medicine, 371(15), 1407–1417. doi: 10.1056/NEJMoa1404588.
  Haeussler, M., Schönig, K., Eckert, H., Eschstruth, A., Mianné, J., Renaud, J. B., … Concordet, J. P. (2016). Evaluation of off target and on‐target scoring algorithms and integration into the guideRNA selection tool CRISPOR. Genome Biol, 17, 148. doi: 10.1186/s13059‐016‐1012‐2.
  Harui, A., Suzuki, S., Kochanek, S., & Mitani, K. (1999). Frequency and stability of chromosomal integration of adenovirus vectors. Journal of Virology, 73, 6141–6146.
  He, T.‐C. (2004). Adenoviral vectors. Current Protocols in Human Genetics, 40, 12.4.1–12.4.25. doi: 10.1002/0471142905.hg1204s40.
  Ibelgaufts, H., & Jones, K. W. (1983). Papovavirus‐related RNA sequences in human neurogenic tumours. Acta Neuropathol, 56, 118–122. doi: 10.1007/BF00690582.
  Johnston, K. M., Jacoby, D., Pechan, P. A., Fraefel, C., Borghesani, P., Schuback, D., … Breakefield, X. O. (1997). HSV/AAV hybrid amplicon vectors extend transgene expression in human glioma cells. Human Gene Therapy, 8, 359–370. doi: 10.1089/hum.1997.8.3‐359.
  Keeler, A. M. 1, ElMallah, M. K. 1, & Flotte, T. R. 1 (2017). Gene Therapy 2017: Progress and Future Directions. Clinical and Translational Science, 10(4), 242–248. doi: 10.1111/cts.12466.
  Kennedy, J. S., & Greenberg, R. N. (2009). IMVAMUNE: Modified vaccinia Ankara strain as an attenuated smallpox vaccine. Expert Review of Vaccines, 8(1), 13–24. doi: 10.1586/14760584.8.1.13.
  Kochanek, S., Clemens, P. R., Mitani, K., Chen, H. H., Chan, S., & Caskey, C. T. (1996). A new adenoviral vector: Replacement of all viral coding sequences with 28 kb of DNA independently expressing both full‐length dystrophin and β‐galactosidase. Proceedings of the National Academy of Sciences of the United States of America, 93, 5731–5736. doi: 10.1073/pnas.93.12.5731.
  Martuza, R. L., Malick, A., Markert, J. M., Ruffner, K. L., & Coen, D. M. (1991). Experimental therapy of human glioma by means of a genetically engineered virus mutant. Science, 252, 854–856. doi: 10.1126/science.1851332.
  McElvaney, N. G., & Crystal, R. G. (1995). IL‐6 release and airway administration of human CFR cDNA adenovirus vector. Nature Medicine, 1, 182–184. doi: 10.1038/nm0395‐182b.
  Meyer, H., Sutte, G., & Mayer, A. (1991). Mapping of deletions in the genome of the highly attenuated vaccinia virus MVA and their influence on virulence. The Journal of General Virology, 72, 1031–1038. doi: 10.1099/0022‐1317‐72‐5‐1031.
  Miller, A. D. (2014). Retroviral vector production. Current Protocols in Human Genetics, 80, 12.5.1–12.5.22. doi: 10.1002/0471142905.hg1205s80.
  NIH Advisory Committee Report. (2002). Assessment of adenoviral vector safety and toxicity: Report of the National Institutes of Health Recombinant DNA Advisory Committee. Human Gene Therapy, 13, 3–13. doi: 10.1089/10430340152712629.
  Parrino, J., & Graham, B. S. (2006). Smallpox vaccines: Past, present, and future. The Journal of Allergy and Clinical Immunology, 118(6), 1320–1326. doi: 10.1016/j.jaci.2006.09.037.
  Pesonen, S., Kangasniemi, L., & Hemminki, A. (2011). Oncolytic adenoviruses for the treatment of human cancer: Focus on translational and clinical data. Molecular Pharmacology, 8, 12–28. doi: 10.1021/mp100219n.
  Printz, M., Reynolds, J., Mento, S. J., Jolly, D., Kowal, K., & Sajjadi, N. (1995). Recombinant retroviral vector interferes with the detection of amphotropic replication competent retrovirus in standard culture assays. Gene Therapy, 2, 143–150.
  Ramos, C. A., Savoldo, B., Torrano, V., Ballard, B., Zhang, H., Dakhova, O., … Dotti, G. (2016). Clinical responses with T lymphocytes targeting malignancy‐associated κ light chains. The Journal of Clinical Investigation, 126, 2588–2596. doi: 10.1172/JCI86000.
  Ramesh, N., Ge, Y., Ennist, D. L., Zhu, M., Mina, M., Ganesh, S., … Yu, D. C. (2006). CG0070, a conditionally replicating granulocyte macrophage colony‐stimulating factor–armed oncolytic adenovirus for the treatment of bladder cancer. Clinical Cancer Research, 12, 305–313. doi: 10.1158/1078‐0432.CCR‐05‐1059.
  Raper, S. E., Yudkoff, M., Chirmule, N., Gao, G. P., Nunes, F., Haskal, Z. J., … Batshaw, M. L. (2002). A pilot study of in vivo liver‐directed gene transfer with an adenoviral vector in partial ornithine transcarbamylase deficiency. Human Gene Therapy, 13, 163–175. doi: 10.1089/10430340152712719.
  Rochlitz, C., Figlin, R., Squiban, P., Salzberg, M., Pless, M., Herrmann, R., … Acres, B. (2003). Phase I immunotherapy with a modified vaccinia virus (MVA) expressing human MUC1 as antigen‐specific immunotherapy in patients with MUC1‐positive advanced cancer. The Journal of Gene Medicine, 5(8), 690–699. doi: 10.1002/jgm.397.
  Russell, S. J., Peng, K. W., & Bell, J. C. (2012). Oncolytic virotherapy. Nature Biotechnology, 30, 658–670. doi: 10.1038/nbt.2287.
  Russell, S., Bennett, J., Wellman, J. A., Chung, D. C., Yu, Z. F., Tillman, A., … Maguire, A. M. (2017). Efficacy and safety of voretigene neparvovec (AAV2‐hRPE65v2) in patients with RPE65‐mediated inherited retinal dystrophy: A randomised, controlled, open‐label, phase 3 trial. Lancet, Jul 13. pii: S0140‐6736(17)31868‐8. doi: 10.1016/S0140‐6736(17)31868‐8
  Schaefer, K. A., Wu, W. H., Colgan, D. F., Tsang, S. H., Bassuk, A. G., & Mahajan, V. B. (2017). Unexpected mutations after CRISPR‐Cas9 editing in vivo. Nature Methods, 14(6), 547–548. doi: 10.1038/nmeth.4293.
  Suter, M., Meisinger‐Henschel, C., Tzatzaris, M., Hülsemann, V., Lukassen, S., Wulff, N. H., … Chaplin, P. (2009). Modified vaccinia Ankara strains with identical coding sequences actually represent complex mixtures of viruses that determine the biological properties of each strain. Vaccine, 27(52), 7442–7450. doi: 10.1016/j.vaccine.2009.05.095.
  Ulmer, J. B., & Rappuoli, R. (2003). DNA vaccination. Current Protocols in Human Genetics, 37, 13.2.1–13.2.9. doi: 10.1002/0471142905.hg1302s37.
  Verheust, C., Goossens, M., Pauwels, K., & Breyer, D. (2012). Biosafety aspects of modified vaccinia virus Ankara (MVA)‐based vectors used for gene therapy or vaccination. Vaccine, 30(16), 2623–2632 doi: 10.1016/j.vaccine.2012.02.016.
  Vogels, R., Zuijdgeest, D., van Rijnsoever, R., Hartkoorn, E., Damen, I., de Bethune, M. P., … Havenga, M. (2003). Replication‐deficient human adenovirus type 35 vectors for gene transfer and vaccination: Efficient human cell infection and bypass of preexisting adenovirus immunity. Journal of Virology, 77, 8263–8271. doi: 10.1128/JVI.77.15.8263‐8271.2003.
  Wang, S., & Vos, J. (1996). A hybrid herpesvirus infectious vector based on Epstein‐Barr virus and herpes simplex virus type 1 for gene transfer into human cells in vitro and in vivo. Journal of Virology, 70, 8422–8430.
  Whitemore, M., Li, S., & Huang, L. (2001). Liposome vectors for in vivo gene delivery. Current Protocols in Human Genetics, 20, 12.8.1–12.8.9. doi: 10.1002/0471142905.hg1208s20.
  Yang, N.‐S., Burkholder, J., McCabe, D., Neumann, V., & Fuller, D. (2001). Particle‐mediated gene delivery in vivo and in vitro. Current Protocols in Human Genetics, 12, 12.6.1–12.6.14. doi: 10.1002/0471142905.hg1206s12.
  Yohn, D. S., Funk, C.A., & Grace, J. T. Jr (1967). Sex‐related resistance in hamsters to adenovirus‐12 oncogenesis. II. Influence of virus dose. Journal of Virology, 1, 1186–1192.
Internet Resources
  Biosafety in Microbiological and Biomedical Laboratories (BMBL), 5th edition, from the Centers for Disease Control (CDC) Office of Health Safety (OHS) and the NIH.
  FDA Center for Biologics Evaluation and Research (CBER) home page.
  The NIH Office of Science Policy.‐content/uploads/NIH_Guidelines.html#_APPENDIX_B._CLASSIFICATION
  APPENDIX B of the April 2002 NIH Recombinant DNA guidelines, listing recombinant DNA agents by risk groups.‐content/uploads/NIH_Guidelines.html#_Toc446948489
  APPENDIX M of the April 2002 NIH Recombinant DNA Guidelines, consisting of a series of screening questions concerning potential vectors and their application to human gene therapy.
  Resource for gene therapy clinical trials worldwide provided by the Journal of gene Medicine.
  NIH Design Requirements Manual 2016 (revised 4/17/2‐17).‐1)small.pdf
  Biosafety Manual from the University of Pennsylvania.‐research/biological‐safety/biosafety‐manual
  Biosafety Manual from Stanford University.‐manual
  Biosafety Manual from Princeton University.‐research/biological‐safety/biosafety‐manual
  National Gene Vector Biorepository (NGVB).
PDF or HTML at Wiley Online Library