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- Unregulated Hazards ‘Naked’ and ‘Free’ Nucleic Acids

1. Traavik, T. (1999a) Too early may be too late: Ecological risks associated with the use of naked DNA as a biological tool for research, production and therapy. pp 29-31. Reported to the Directorate of Nature Management, Norway.

2. See Ho, M.W. (1998, 1999). Genetic Engineering Dream or Nightmare? The Brave New World of Bad Science and Big Business, Gateway Books, Bath 2nd ed., Gateway, Gill & Macmillan, Dublin.

3. As defined by Traavik, 1999a (note 1)

4. See Ho, 1998, 1999 (note 2); Ho, M.W., Traavik, T., Olsvik, R., Tappeser, B., Howard, V., von Weizsacker, C. and McGavin, G. (1998b). Gene Technology and Gene Ecology of Infectious Diseases. Microbial Ecology in Health and Disease 10, 33-59; Traavik, T. (1999b). An orphan in science: Environmental risks of genetically engineered vaccines. Reported to the Directorate of Nature Management, Norway.

5. See Ho et al, 1998 (note 4).

6. See Traavik, 1999b (note 4).

7. See Schindelhauer, D. (1999). Construction of mammalian artificial chromosomes: prospects for defining an optimal centromere. BioEssays 21, 76-83.

8. See Helin, V., Gottikh, M., Mishal, Z., Subra, F., Malvy, C. and Lavignon, M. (1999). Cell cycle-dependent distribution and specific inhibitory effect of vectorized antisense oligonucleotides in cell culture. Biochemical Pharmacology 58, 95-107; Campagno, D. and Toulme, J.-J. (1999). Antisense effects of ligonucleotides complementary to the hairpin of the Leishmania mini-exon RNA. Nucleosides & Nucleotides 18, 1701-1704.

9. See Hammann, C. and Tabler, M. (1999). Generation and application of asymmetric hammerhead ribozymes. Methods: a Companion to Methods in Enzymology 18, 273-380.

10. Han, Y., Zaks, T.Z., Wang, T.F., Irvine, D.R., Kammula, U.S., Marincola, F.M., Leitner, W.W. and Restifo, N.P. (1999). Cancer therapy using a self-replicating RNA vaccine. Nature Medicine 3, 823-827.

11. Beetham, P.R., Kipp, P.B., Sawycky, X.L., Arntzen, C.J. and May, G.D. (1999). A tool for functional plant genomics: Chimeric RNA/DNA oligonucleotides cause in vivo gene-specific mutations. PNAS 96, 8774-8778.

12. Reviewed by Lorenz, M.G. and Wackernagel, W. (1994). Bacterial gene transfer by natural genetic transformation in the environment. Microbiol. Rev. 58, 563-602; also, Ho, 1998,1999 (note 2); Ho, et al, 1998 (note 4); Traavik, 1999a (note 1).

13. This was said to M.W.H. by a spokesperson of the UK Health and Safety Executive when asked whether there is any recommended treatment for disposal of naked/free DNA.

14. See Lorenz and Wackernagel, 1994 (note 12); also Ho, 1998, 1999 (note 2); Ho, et al, 1998 (note 4).

15. Mercer, D.K., Scott, K.P., Bruce-Johnson, W.A. Glover, L.A. and Flint, H.J. (1999). Fate of free DNA and transformation of the oral bacterium Streptococcus gordonii DL1 by plasmid DNA in human saliva. Applied and Environmental Microbiology 65, 6-10.

16. Schubbert, R., Lettmann, C. and Doerfler, W. (1994). Ingested foreign (phage M13) DNA survives transiently in the gastrointestinal tract and enters the bloodstream of mice. Molecular and General Genetics 242, 495-504; Schubbert, R., Rentz, D., Schmitzx, B. and Doerfler, W. (1997). Foreign (M13 DNA ingested by mice reaches peripheral leukocytes, spleen and liver via the intestinal wall mucosa and can be covalently linked to mouse DNA. Proc. Nat. Acad. Sci. USA 94, 961-6.

17. Mercer et al, 1999 (note 15).

18. Brown, P. Naked DNA raises cancer fears for researchers. New Scientist, 6 October, 17 (1990).

19. Schubbert et al, 1997 (note 16).

20. Doerfler, W. and Schubbert, R. (1998). Uptake of foreign DNA from the environment: the gastroinestinal tract and the placenta as portals of entry, Wien Klin Wochenschr. 110, 40-44.

21. Doerfler and Schubbert, 1998, (note 20), p. 40.

22. Yei, S., Mittereder, N., Wert, S., Whitsett, J.A., Wilmott, R.W. and Trapnell, B.C. (1994). In vivo evaluation of the safety of adenovirus-mediated transfer of the human cystic fibrosis transmembrane conductance regulator cDNA to the lung. Hum. Gene Ther. 15, 731-744.

23. Noisakran S, Campbell IL, Carr DJ, (1999) Ecotopic expression of DNA encoding IFN-alpha 1 in the cornea protects mice from herpes simplex virustype 1-induced encephalitis. J Immunol 162, 4184-90.

24. Yamasoba T, Yagl M, Roessler BJ, Miller JM, Rapheal Y (1999) Inner ear transgene expression after adenoviral vecotr inoculation in the endolymphatic sac. Hum Gene Ther 10, 744-69.

25. See Hoffman, R.M. (2000). The hair follicle as a gene therapy target. Nature Biotechnology 18, 20-1.

26. Budker, V., Zhang, G., Danko, I., Williams P. and Wolff, J. (1998). The efficient expression of intravascularly delivered DNA in rat muscle. Gene Therapy 5, 272-6; Han, et al, 1999 (note 12).

27. Khavari, P.A. Cutaneous gene therapy. Advances in Clinical Research 15, 27-35 (1997);

28. During, M.J., Xu, R., Young, D., Kaplitt, M.G., Sherwin, R.S., Leone, P. (1998). Peroral gene therapy of lactose intolerance using an adeno-associated virus vector. Nat. Med. 4, 1131-5.

29. Spadafora, C. (1998). Sperm cells and foreign DNA: a controversial relation. BioEssays 20, 955-64.

30. Zhang, G. Vargo, D., Budker, V., Armstrong, N., Knechtle, S. and Wolf, J., Expression of naked DNA injected into the afferent and efferent vessels of rodents and dog livers. Human Gene Ther. 8,1763-72 (1997).

31. Hengge, U., Chan, E., Foster, R.,Walker, P. and Vogel, J., Cytokine gene expression in epidermis with biological effects following injection of naked DNA,. Nat. Genet 10,161-6 (1995)

32. Hengge, U., Walker, P. and Vogel, J. Expression of naked DNA in human, pig and mouse skin. J Clin Invest 97,2911-6 (1996).

33. Martin, T., Parker, S.E., Hedstrom, R., Le, Thong, Hoffman, S.L., Norman, J., Hobart, P. and Lew, D. (1999). Plasmid DNA malaria vaccine: the potential for genomic integration after intramuscular injection. Hum. Gene Ther. 10, 759-68.

34. Zhaqo,T., Robinson, M., Bowers, F. and Kindt,T. Infectivity of chimeric human T-cell leukaemia virus type I molecular clones assessed by naked DNA inoculation. Proc. Natnl. Acad Sci. USA 93,6653-8 (1996).

35. Rekvig, O.P. Fredriksen, K., Brannsether, B., Moens, U., Sundsfjord, A. and Traavik, T., Antibodies to eucaryotic, including autologous, native DNA are produced during BK virus infection, but not after immunization with non-infectious BK DNA. Scand. J. Immunol. 36, 487-495 (1992).

36. Brower, V. (1998). Naked DNA vaccines come of age. Nature Biotechnology 16, 1304-5;

37. see also Traavik, 1999b (note 4). See Traavik, 1999b (note 4).

38. See Verdier, F. and Descotes, J. (1999). Preclinical safety evaluation of human gene therapy products. Toxicological Sciences 47, 9-15; Jane, S.M., Cunningham, J.M. and Vanin, E.F. (1998). Vector development: a major obstacle in human gene therapy. Annals of Medicine 30, 413-5.

39. Putnam, L. (1998). Debate grows on safety of gene-therapy vectors. The Lancet 351, 808.

40. See Ho, et al, 1998 (note 4).

41. Baba, T.W. Liska, V., Khimani, A.H., Ray, N.B., Dailey, P.J., Penninck, D., Bronson, R., Greene, M.F., McClure, H.M., Martin, L.N. and Ruth M. Ruprecht, R.M. Live attenuated, multiply deleted simian immunodeficiency virus causes AIDS in infant and adult macaques. Nature Med. 5, 194, 203 (1999).

42. See Verdier and Desotes, 1999 (note 38).

43. See Coghlan, A. (1996). Gene shuttle virus could damage the brain. New Scientist 11 May, 6.

44. Nelson D & Weiss R. Gene research moves towards secrecy. Washington Post Nov 3, 1999

45. Suzuki, K., Mori, A., Ishii, K.J., Singer, D.S., Klinman, D.M., Krause, P.R. and Kohn, L.D. (1999). Activation of target-tissue immune-recognition molecules by double-stranded polynucleotides. Proc. Natl. Acad. Sci. USA 96, 2285-90.

46. Fong KM at al (1997) FHIT and FRA3B 3p14.2 allele loss are common in lung cancer and preneoplasic bronchial lesions and are associated with cancer related FHIT cDNA splicing abberations. Cancer Res. (CNF), 57 (11) ; 2256-67.

47. Asch HL (1996) Comparative expression of the LINE-1 p40 protein in human breast carcinomas and normal breast tissues. Oncol. Res (BBN) 8 (6): 239-47.

48. Miki Y. (1992). Disruption of the ARC gene by retrotransposal insertion of L1 sequence in a colon cancer. Cancer Res (CNF), 52 (3):643-5

49. Buendia, M.A. (1992). Mammalian hepatitis B viruses and primary liver cancer. Semin. Cancer Biol. 3, 309-20.

50. See Verdier and Descotes, 1999 (note 38); Spadafora, 1998 (note 29).

51. Mao, J.R., Inouye, M. and Inouye, S. (1996). An unusual bacterial reverse transcriptase having LVDD in the YXDD box from Escherichia coli. Biochem. Biophys. Res. Commun. 227, 489-93.

52. Hoffman, T., Golz, C. & Schieder, O. (1994). Foreign DNA sequences are received by a wild-type strain of Aspergillus niger after co-culture with transgenic higher plants. Current Genetics 27: 70-6.

53. De Vries, J. and Wackernagel, W. (1998). Detection of nptII (kanamycin resistance) genes in genomes of transgenic plants by marker-rescue transformation. Mol. Gen. Genet. 257, 606-13.

54. Schluter, K., Futterer, J. & Potrykus, I. (1995). Horizontal gene-transfer from a transgenic potato line to a bacterial pathogen (Erwinia-chrysanthem) occurs, if at all, at an extremely low-frequency. Bio/Techology 13: 1094-8.

55. Timms-Wilson, T.M., Lilley, A.K. and Bailey, M.J. (1999). A Review of Gene Transfer from Genetically Modified Micro-organisms. Report to UK Health and Safety Executive.

56. Gebhard, F. and Smalla, K. (1998). Transformation of Acinetobacter sp. strain BD413 by transgenic sugar beet DNA. Appl. Environ. Microbiol. 64, 1550-4.

57. See Traavik, 1999a (note 1); Timms-Wilson, et al, 1999 (note 24).

58. Pace, N. (1997). A molecular view of microbial diversity and the biosphere. Science 276, 734-9.

59. See Ho, 1998, 1999 (note 1); Ho et al, 1998b (note 1); Traavik, 1999a (note 1).

60. See Kohli, A., Griffiths, S., Palacios, N., Twyman, R.M., Vain, P., Laurie, D.A. and Christou, P. (1999). Molecular characterization of transforming plasmid rearrangements in transgenic rice reveals a recombination hotspot in the CaMV 35S promoter and confirms the predominance of microhomology mediated recombination. The Plant Journal 17, 591-601; also Ho, M.W., Ryan, A. and Cummins, J. (1999). The CaMV promoter -- A recipe for disaster?, Microbial Ecology in Health and Disease (in press).

61. See Ho et al, 1998b (note 4) and references therein.

62. Letter from N. Tomlinson, Joint Food Safety and Standards Group, MAFF, to US FDA, 4 December, 1998.

63. See Ho, M.W. , 1998, 1999 (note 2)

64. This was the question asked by Ho et al, 1998 (note 4) who called for an urgent public enquiry; See also Ho, M.W., Traavik, T., Olsvik, O., Midtvedt, T., Tappeser, B., Howard, C.V., von Weizsacker, C. and McGavin, G. (1998). Gene Technology in he Etiology of Drug-resistant Diseases. TWN Biotechnology & Biosafety Series 2, Third World Network, Penang.

65. See Ho, 1998, 1999 (note 2)

66. See Jakowitsch, J., Mette, M.G., van der Winden, J., Matzke, M.A. and Matzke, A.J.M. (1999). Integrated pararetrovial sequences define a unique class of dispersed repetitive DNA in plants. Proc. Nat. Acad. Sci. USA 96, 13241-6.

67. Greene, A.E. and Allison, R.F. (1994). Recombination between viral RNA and transgenic plant transcripts. Science 263, 1423-5; Wintermantel, W.M. and Schoelz, J.E. (1996). Isolation of recombinant viruses between cauliflower mosaic virus and a viral gene in transgenic plants under conditions of moderate selection pressure. Virology 223, 156-64.

68. See Ho, 1998, 1999 (note 2) especially Chapter 13/12.

69. See Ho et al, 1999 (note 30).

70. See Robinson, W.P. and Lalande, M. (1995). Sex-specific meiotic recombination in the Prader-Willi/Angelman syndrome imprinted region. Hum. Mol. Genet. 4, 801-6; Wu,T.C. and Lichten, M. (1994). Meiosis-induced double-stranded break sites determined by yeast chromatin structure. Science 263, 515-8.

71. See Kohli , et al, 1999 (note 56).

72. Ewen, S.W.B. and Pusztai, A. (1999). Effect of diets containing genetically modified potatoes expressing Galanthus nivalis lectin on rat small intestine. The Lancet 354.



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