Chemistry: molecular biology and microbiology – Treatment of micro-organisms or enzymes with electrical or... – Modification of viruses
Patent
1996-09-12
2000-03-07
Stucker, Jeffrey
Chemistry: molecular biology and microbiology
Treatment of micro-organisms or enzymes with electrical or...
Modification of viruses
424 932, 4353201, 4352351, 435 703, 435 704, 435 41, 435 691, 435 693, C12N 1500, C12N 700, A01N 6300, C12P 100
Patent
active
060338857
DESCRIPTION:
BRIEF SUMMARY
BACKGROUND OF THE INVENTION
The present invention relates to recombinant vectors of viral origin and their therapeutic use. More particularly, it relates to recombinant adenoviruses containing a cassette capable of becoming integrated into the genome of the infected cells. The invention also relates to the preparation of these vectors, the pharmaceutical compositions containing them and their use for the transfer of genes in vitro, ex vivo and in vivo, especially within the framework of gene and cell therapies.
Gene and cell therapy consists in correcting a deficiency or an abnormality (mutation, aberrant expression and the like) or in ensuring the expression of a protein of therapeutic interest by the introduction of a genetic information into the cell or the affected organ. This genetic information can be introduced either in vitro into a cell extracted from the organ, the modified cell then being reintroduced into the body, or directly in vivo into the appropriate tissue. Various techniques have been described for the transfer of this genetic information, amongst which are various transfection techniques involving complexes of DNA and DEAE-dextran (Pagano et al., J. Virol. 1 (1967) 891), of DNA and nuclear proteins (Kaneda et al., Science 243 (1989) 375), of DNA and lipids (Felgner et al., PNAS 84 (1987) 7413), of DNA and polylysine, the use of liposomes (Fraley et al., J. Biol. Chem. 255 (1980) 10431) and the like.
More recently, the use of viruses as vectors for the transfer of genes appeared as a promising alternative to these physicochemical transfection techniques. In this respect, various viruses have been tested for their capacity to infect certain cell populations, in particular retroviruses (RSV, HMS, MMS and the like), HSV virus, adeno-associated viruses, and adenoviruses. However, the viral vectors developed up until now do not make it possible to solve satisfactorily all the difficulties linked to the transfer of genes into the cells and/or the body. Thus, adenovirus, which possesses attractive properties for the transfer of genes (possibility of producing high titres, low pathogenicity) is an extrachromosomal virus. Because of this, in dividing cells, the recombinant virus is diluted over generations and eventually disappears completely from the daughter cells. On the other hand, whereas retroviral vectors or vectors derived from adeno-associated viruses (AAV) are capable of becoming integrated into the genome of the cells which they infect, they cannot be produced in large quantities nor for example incorporate large transgenes.
BRIEF SUMMARY OF THE INVENTION
The present invention provides an advantageous solution to these problems. The present invention resides indeed in the development of recombinant vectors which can be used in gene therapy, possessing the infecting properties of a recombinant adenovirus vector and permitting the integration of a heterologous sequence into the genome of the infected cell or organ.
A first subject of the invention relates more particularly to a defective recombinant adenovirus comprising a cassette capable of becoming integrated into the genome of the infected cells.
Generally, the cassette comprises a desired DNA sequence, which is most often heterologous in relation to the adenovirus, and elements permitting its integration into the genome of the infected cells. Advantageously, the elements permitting the integration are of viral origin. Thus, the vectors of the invention combine properties of two types of viruses: the adenoviruses and the integrative viruses.
The vectors of the invention are particularly advantageous since they can be produced at high titres, are not pathogenic, possess a broad host spectrum, are capable of incorporating large heterologous DNA sequences and of integrating the said sequences into the genome of the infected cells.
Furthermore, the vectors of the invention make it possible to limit the risks of dissemination of the DNA sequence which it is desired to transfer to the cell or the body. Once the said cell has been integ
REFERENCES:
Rosenfeld, et al., In Vivo Transfer of the Human Cystic Fibrosis Transmembrane Conductance Regulator Gene to the Airway Epithelium, Cell, vol. 68, 143-155, see p. 143, col. 1, "Summary" and col. 2, second paragraph, "Introduction", and p. 144, Res, Jan. 1992.
Walsh C. et al. Regulated high level expression of a human gamma-globin gene introduced into erythroid cells by an adeno-associated virus vector, Proc. Nat'l Acad. Sci. 89:7257-7261, Aug. 1992.
Flotte T. et al. Expression of the cystic fibrosis transmembrane conductance regulator from a novel adeno-associated virus promoter, Jol. Biol. Chem. 268(5):3781-3790, Feb. 1993.
Nahreini et al., Cloning and integration of DNA fragments in human cells via the inverted terminal repeats of the adeno-associated virus 2 genome, Gene 119, 265-272 (1992).
Denefle Patrice
Latta Martine
Perricaudet Michel
Vigne Emmanuelle
Park Honkyel
Rhone-Poulenc Rorer S.A.
Stucker Jeffrey
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