A Cre-expressing cell line and an E1/E2a double-deleted virus for preparation of helper-dependent adenovirus vector.

Adenoviral vectors are attractive for the delivery of transgenes into mammalian cells because of their efficient transduction, high titer, and stability. The major concerns with using E1-deleted adenoviral vectors in gene therapy are the pathogenic potential of the virus backbone and the leaky viral protein synthesis that leads to host immune responses and a short duration of transgene expression. Helper-dependent (HD) adenoviral vectors that are devoid of all viral protein-coding sequences have significantly increased the safety and reduced the immunogenicity of these vectors. Currently available HD vectors depend on an E1-deleted adenovirus as a helper to provide viral proteins in trans. As a consequence, contamination with helper virus cannot be avoided in the HD vector preparation though it can be decreased to 0.01% using a Cre/loxP mechanism. Since the presence of E1-deleted helper virus may have substantial unwanted effects, we have developed a new Cre-expressing cell line based on an E1- and E2a-complementing cell. This new cell line can efficiently cleave the packaging region in the helper virus genome. We have also developed an E1 and E2a double-deleted helper virus. By using the CreE cell with the helper virus deleted in both the E1 and the E2a genes it may be possible to further improve the safety of the vectors.

[1]  H. Zhou,et al.  Helper-dependent adenoviral vector-mediated gene transfer in aged rat brain. , 2001, Human gene therapy.

[2]  I. Verma.  Gene therapy: the need for basic science. , 2000, Molecular Therapy.

[3]  A. Beaudet,et al.  A new vector system with inducible E2a cell line for production of higher titer and safer adenoviral vectors. , 2000, Virology.

[4]  H. Zhou,et al.  Prolonged transgene expression mediated by a helper-dependent adenoviral vector (hdAd) in the central nervous system. , 2000, Molecular therapy : the journal of the American Society of Gene Therapy.

[5]  M. Metzker,et al.  Optimization of the helper-dependent adenovirus system for production and potency in vivo. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[6]  E Marshall,et al.  Gene Therapy Death Prompts Review of Adenovirus Vector , 1999, Science.

[7]  F. Graham,et al.  Effects of Stuffer DNA on Transgene Expression from Helper-Dependent Adenovirus Vectors , 1999, Journal of Virology.

[8]  A. Amalfitano,et al.  Next-generation adenoviral vectors: new and improved , 1999, Gene Therapy.

[9]  W. Poller,et al.  ‘Autoreplication’ of the vector genome in recombinant adenoviral vectors with different E1 region deletions and transgenes , 1999, Gene Therapy.

[10]  B. O’Malley,et al.  Adenovirus-mediated regulable target gene expression in vivo. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[11]  A. Beaudet,et al.  High doses of a helper-dependent adenoviral vector yield supraphysiological levels of alpha1-antitrypsin with negligible toxicity. , 1998, Human gene therapy.

[12]  P. Seth,et al.  Recombinant, Replication-Defective Adenovirus Gene Transfer Vectors Induce Cell Cycle Dysregulation and Inappropriate Expression of Cyclin Proteins , 1998, Journal of Virology.

[13]  C. Caskey,et al.  An adenoviral vector deleted for all viral coding sequences results in enhanced safety and extended expression of a leptin transgene. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[14]  M. Lusky,et al.  In Vitro and In Vivo Biology of Recombinant Adenovirus Vectors with E1, E1/E2A, or E1/E4 Deleted , 1998, Journal of Virology.

[15]  A. Beaudet,et al.  Genomic DNA transfer with a high-capacity adenovirus vector results in improved in vivo gene expression and decreased toxicity , 1998, Nature Genetics.

[16]  J. Chamberlain,et al.  Production and Characterization of Improved Adenovirus Vectors with the E1, E2b, and E3 Genes Deleted , 1998, Journal of Virology.

[17]  J. Mcghee,et al.  Adenoviral gene delivery elicits distinct pulmonary-associated T helper cell responses to the vector and to its transgene. , 1997, Journal of immunology.

[18]  F. Graham,et al.  Cloning and sequencing of the cellular-viral junctions from the human adenovirus type 5 transformed 293 cell line. , 1997, Virology.

[19]  A. Beaudet,et al.  Immune responses to reporter proteins and high viral dose limit duration of expression with adenoviral vectors: comparison of E2a wild type and E2a deleted vectors. , 1997, Human gene therapy.

[20]  M. Lusky,et al.  Gene therapy with recombinant adenovirus vectors: evaluation of the host immune response. , 1997, Immunology letters.

[21]  F. Graham,et al.  A helper-dependent system for adenovirus vector production helps define a lower limit for efficient DNA packaging , 1997, Journal of virology.

[22]  M. Mehtali,et al.  Conditional repression of the E2 transcription unit in E1-E3-deleted adenovirus vectors is correlated with a strong reduction in viral DNA replication and late gene expression in vitro , 1997, Journal of virology.

[23]  S. Kochanek,et al.  Persistence in muscle of an adenoviral vector that lacks all viral genes. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[24]  K. Hehir,et al.  Molecular characterization of replication-competent variants of adenovirus vectors and genome modifications to prevent their occurrence , 1996, Journal of virology.

[25]  M. Rudnicki,et al.  A helper-dependent adenovirus vector system: removal of helper virus by Cre-mediated excision of the viral packaging signal. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[26]  F. Graham,et al.  Production and characterization of human 293 cell lines expressing the site-specific recombinase Cre , 1996, Somatic cell and molecular genetics.

[27]  M. Ng.  Is gene therapy in cystic fibrosis a realistic expectation , 1996 .

[28]  A. Beaudet,et al.  Development of a complementing cell line and a system for construction of adenovirus vectors with E1 and E2a deleted , 1996, Journal of virology.

[29]  J. Chamberlain,et al.  Encapsidated adenovirus minichromosomes allow delivery and expression of a 14 kb dystrophin cDNA to muscle cells. , 1996, Human molecular genetics.

[30]  R. Luthra,et al.  Elimination of both E1 and E2 from adenovirus vectors further improves prospects for in vivo human gene therapy , 1996, Journal of virology.

[31]  D. Bellinger,et al.  Lack of persistence of E1- recombinant adenoviral vectors containing a temperature-sensitive E2A mutation in immunocompetent mice and hemophilia B dogs. , 1996, Gene therapy.

[32]  A. Bradley,et al.  Chromosome engineering in mice , 1995, Nature.

[33]  C. Caskey,et al.  Rescue, propagation, and partial purification of a helper virus-dependent adenovirus vector. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[34]  Hanns Lochmüller,et al.  Emergence of Early Region 1-Containing Replication-Competent Adenovirus in Stocks of Replication-Defective Adenovirus Recombinants (ΔE1 + ΔE3) During Multiple Passages in 293 Cells , 1994 .

[35]  F. Graham,et al.  An efficient and flexible system for construction of adenovirus vectors with insertions or deletions in early regions 1 and 3. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[36]  Y Wang,et al.  A regulatory system for use in gene transfer. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[37]  J. Whitsett,et al.  Evaluation of the efficacy and safety of in vitro, adenovirus-mediated transfer of the human cystic fibrosis transmembrane conductance regulator cDNA. , 1994, Human gene therapy.

[38]  M. Gossen,et al.  Acceleration of the G1/S phase transition by expression of cyclins D1 and E with an inducible system. , 1994, Molecular and cellular biology.

[39]  S. Thorgeirsson,et al.  Adenovirus–mediated in vivo gene transfer and expression in normal rat liver , 1992, Nature Genetics.

[40]  J. Mandel,et al.  Adenovirus as an expression vector in muscle cells in vivo. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[41]  S. Chen‐Kiang,et al.  Interleukin 6 enhances a cellular activity that functionally substitutes for E1A protein in transactivation. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[42]  M. Perricaudet,et al.  Adenovirus-mediated transfer of a recombinant alpha 1-antitrypsin gene to the lung epithelium in vivo. , 1991, Science.

[43]  M. Hitt,et al.  Adenovirus E1A under the control of heterologous promoters: wide variation in E1A expression levels has little effect on virus replication. , 1990, Virology.

[44]  S. Nagata,et al.  Isolation and characterization of the human chromosomal gene for polypeptide chain elongation factor-1 alpha. , 1989, The Journal of biological chemistry.

[45]  G. Winberg,et al.  Dissection of overlapping functions within the adenovirus type 5 E1A gene. , 1984, The EMBO journal.

[46]  J. Nevins,et al.  Common control of the heat shock gene and early adenovirus genes: evidence for a cellular E1A-like activity , 1984, Molecular and cellular biology.

[47]  T. Shenk,et al.  Functional analysis of the nucleotide sequence surrounding the cap site for adenovirus type 5 region E1A messenger RNAs. , 1983, Journal of molecular biology.

[48]  F. Graham,et al.  Characteristics of a human cell line transformed by DNA from human adenovirus type 5. , 1977, The Journal of general virology.

[49]  E. Southern.  Detection of specific sequences among DNA fragments separated by gel electrophoresis. , 1975, Journal of molecular biology.

[50]  J. Maizel,et al.  The polypeptides of adenovirus. I. Evidence for multiple protein components in the virion and a comparison of types 2, 7A, and 12. , 1974, Virology.

[51]  D. Curiel,et al.  Interleukin-6 modulated conditionally replicative adenovirus as an antitumor/cytotoxic agent for cancer therapy. , 1999, Clinical cancer research : an official journal of the American Association for Cancer Research.

[52]  E. Schaefer,et al.  Characterization of replication-competent adenovirus isolates from large-scale production of a recombinant adenoviral vector. , 1999, Human gene therapy.

[53]  M. Hitt,et al.  Human adenovirus vectors for gene transfer into mammalian cells. , 1997, Advances in pharmacology.

[54]  M. Perricaudet,et al.  Direct gene transfer of human CFTR into human bronchial epithelia of xenografts with E1–deleted adenoviruses , 1993, Nature Genetics.

[55]  F. Graham,et al.  Manipulation of adenovirus vectors. , 1991, Methods in molecular biology.

[56]  M. Perricaudet,et al.  Evaluation of the transfer and expression in mice of an enzyme-encoding gene using a human adenovirus vector. , 1990, Human gene therapy.

[57]  L. G. Davis,et al.  Basic methods in molecular biology , 1986 .

[58]  T. Shenk,et al.  Genetic analysis of adenoviruses. , 1984, Current topics in microbiology and immunology.