Chemistry: molecular biology and microbiology – Vector – per se
Reexamination Certificate
1997-06-03
2001-03-13
Priebe, Scott D. (Department: 1632)
Chemistry: molecular biology and microbiology
Vector, per se
C435S455000, C435S456000, C424S093200
Reexamination Certificate
active
06200799
ABSTRACT:
BACKGROUND OF THE INVENTION
This invention relates to materials and methods for the construction of recombinant viral, particularly adenoviral, delivery systems that provide lens epithelial cell type specific regulation of expression of proteins which inhibit cellular proliferation. This invention also relates to the use of these recombinant viral delivery systems for inhibiting or preventing the formation of secondary cataracts following surgical intervention on the eye.
Cataract operations are the second most frequent operation in the western world. The cataract is an affliction characterized by an opacification of the crystalline lens of the eye which reduces the visual acuity of the patient. The surgical technique currently utilized for removing cataracts consists of removing the crystalline with a small aspirator. Access to the interior of the crystalline capsule is achieved by a circular opening at the anterior face of the lens capsule. This permits the surgeon to insert a new lens, which will restore the eye's ability to focus incoming light on the surface of the retina.
Secondary cataracts constitute the most frequent complication following this operation, occurring in up to 50% of adult eyes and even more frequently following surgery of children with congenital cataracts (Nischi et al., 1986). (Surgical interventions on the eyes of rabbits also result in secondary cataract formation (100%) in the weeks following the operation (Blomstedt. et al., 1987)). Human secondary cataracts are characterized by a secondary opacification of the posterior capsule appearing in the months following surgery. They are the consequence of the stimulated proliferation at the level of the germinative epithelium of the lens, which are the only cells with division potential in the mature eye. Laser treatment, which is the accepted means of treating secondary cataracts, unfortunately risks additional complications, the most notable of which is detachment of the retina (Salvenson et al., 1991).
SUMMARY OF THE INVENTION
In accordance with this invention is provided a replication-defective recombinant virus that can infect an eye of a patient and that contains:
an Open Reading Frame (ORF) which, when expressed in lens epithelial cells of the eye, suppresses, at the level of the germinative epithelium of the lens of the eye, cellular proliferation which is stimulated by eye surgery and which would otherwise result in secondary cataract formation in the eye;
the ORF, to be expressed, being under the control of a promoter sequence which is active exclusively in human lens epithelial cells.
The preferred replication-defective recombinant virus is a replication-defective recombinant adenovirus, especially an adenovirus lacking E1a, 1b and E4 ORF 6.
The preferred ORF, to be expressed, is one of the following:
a) an ORF encoding a protein that regulates the cell cycle, especially a non-phosphorylatable retinoblastoma (Rb) gene;
b) a dominant negative mutant of an ORF encoding a protein involved in signal transduction, especially a dominant negative mutant of a RAS gene, particularly the codon 116 mutant of the human RAS gene; or
c) an ORF encoding a protein which will severly disrupt DNA replication in human lens epithelial cells, especially a thymidine kinase gene of a Herpes virus, particularly the Herpes Simplex type 1 thymidine kinase (HSTK) gene (to be used in conjunction with a treatment of the patient with acyclovir or a nucleoside analogue thereof).
The preferred promoter sequence is one of the following:
a) a promoter of a human Major Intrinsic Protein (MIP) gene, preferably the portions of the MIP gene from −259 nt to +34 nt, or
b) a promoter of a human A3/A1-crystallin gene, preferably the portions of the -crystallin gene from −345 nt to +45 nt, or preferably
c) a composite promoter comprising either the MIP promoter or the -crystallin promoter or portions thereof, in combination with elements of an early growth response gene promoter such as the rat Early Growth Response-1 (EGR-1) gene promoter from −518 nt to −236 nt of the EGR-1 gene.
Also in accordance with this invention are provided:
a) purified and isolated DNA sequences of the composite EGR-1/MIP and composite EGR-1/-crystallin promoters and
b) vectors for the formation of replication-defective recombinant viruses containing the gene, to be expressed, under the control of the promoter sequence which is expressed exclusively in human lens epithelial cells.
Further in accordance with this invention is provided a method for the treatment of an eye, undergoing eye (e.g. cataract) surgery, in order to reduce the incidence of cellular proliferation in the eye following the eye surgery, and thereby prevent the formation of secondary cataracts, comprising the step of:
treating the eye with the replication-defective recombinant virus, particularly adenovirus, of this invention, preferably during the eye surgery.
REFERENCES:
Orkin et al., “Report and recommendations of the panel to assess the NIH investment in research on gene therapy”, issued by the U.S. National Institutes of Health, Dec. 1995.
Verma et al., “Gene therapy-promises, problems and prospects”, Nature 389: 239-242, Sep. 1997.
Murata et al., “Ocular gene therapy: Experimental studies and clinical possibilities”, Ophthalmic Res. 29: 242-251, Sep. 1997.
Borrás et al., “Ocular adenovirus gene transfer varies in efficiency and inflammatory response”, Invest. Ophthalmol. Vis. Sci. 37: 1282-1293, Jun. 1996.
Munier Francis
Sahli Roland
Shaw Phillip Herbert
Sickenberg Michel
Elrifi Ivor R.
Konzak Kristin E.
Mintz Levin Cohn Ferris Glovsky & Popeo
Priebe Scott D.
University of Lausanne
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