Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Carbohydrate doai
Reexamination Certificate
1999-12-09
2002-07-02
Ketter, James (Department: 1632)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Carbohydrate doai
C435S320100, C435S325000, C435S352000, C435S366000, C424S093100, C424S093210, C536S023100, C536S023400
Reexamination Certificate
active
06413943
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
This invention relates to the inhibition of smooth muscle cell proliferation, particularly vascular smooth muscle cell proliferation following injury to a blood vessel from stenting of a blood vessel, angioplasty, or engraftment of a vein onto the arterial system.
2. Summary of the Related Art
Several disease phenotypes, including cancer, rheumatoid arthritis, macular degeneration, and restenosis (e.g., restenosis induced by angioplasty), involve the hyper-proliferation of cells. Cyclin-dependent kinase (CDK) complexes play a key role in the eukaryotic cell cycle (Draetta G. (1990)
Trends Biochem. Sci
. 15: 378-383; Sherr, C. J. (1993)
Cell
73: 1059-1065). The CDK complex activity is regulated by mechanisms such as stimulatory or inhibitory phosphorylations, as well as the synthesis and degradation of the kinase and cyclin subunit themselves. While p21
Waf1/Cip1
and p27
Kip1
inhibit all the CDK/cyclin complexes tested, p16
Ink4/MTS1
, p15, p18, and p19 block exclusively the activity of the CDK4/cyclin D and CDK6/cyclin D complexes in the early G
1
phase by either preventing the interaction of CDK4 and Cyclin D1, or indirectly preventing catalysis (Serrano et al. (1993)
Nature
366: 704-707).
Gene therapy offers the opportunity to prevent the disease pathology. In patients with occlusion of the coronary and/or peripheral vasculature, one of the leading treatments is disassociation of the atherosclerotic plaque through angioplasty. In response to angioplasty, the normal vessel architecture is disrupted and a complex cascade of events takes place, concluding with the hyper-proliferation of the vascular smooth muscle cells and eventual reocclusion of the vessel in approximately 30% of patients (Lange et al. (1993)
Am. J. Med. Sci
. 306:265-275).
Vein grafts are also used as a conduit for peripheral and aurotocoronary bypass grafting. However, more than 50% of coronary veinous bypass grafts fail within 10 years of implantation, often due to restenosis and intimal hyperplasia, which results from undesired smooth muscle cell proliferation (Channon et al. (1997)
Cardiovascular Research
35:505-513). Antiplatelet agents have had limited impact in reducing vein graft failure (Channon et al., supra).
Stent placement, a procedure for the prevention of immediate vessel closure or elastic rebound following angioplasty, where a stent is placed into a blood vessel, also causes injury to the blood vessel, which can promote restenosis.
Several groups have demonstrated that over-expression of the p21
Waf1/CiP1
, p27
Kip1
, p53, or Rb genes, or cdc-2 and cdk-2 anti-sense genes in the cells at the site of vessel injury will inhibit the hyper-proliferation of vascular smooth muscle cells induced by balloon injury (Chang et al. (1995)
J. Clin. Invest
. 96:2260-2268; Fukui et al. (1997)
Atherosclerosis
132:53-59; Chen et al. (1997)
J. Clin. Invest
. 99:2334-2341; Chang et al. (1995)
Science
267:518-522; Yang et al (1996)
Proc. Natl. Acad. Sci
. (
USA
) 93:7905-7910; Yonemitsu et al. (1998)
Circ. Res
. 82:147-156; Kaneda et al. (1997)
Ann. N.Y. Acad. Sci
. 811:308-310; Suzuki et al. (1997)
Nat. Med
. 8:900-903). However, the efficiency of intra-vascular gene therapy in patients may limit over-expression of these cell cycle regulators.
Therefore, there still exists a need for very potent anti-proliferative reagents to reduce undesired smooth muscle cell proliferation while overcoming the shortcomings of known cell proliferation-inhibiting reagents. Such novel reagents and methods for using them are useful for treating conditions associated with smooth muscle cell proliferation including, without limitation, restenosis due to angioplasty, stent placement, or vein engraftment.
BRIEF SUMMARY OF THE INVENTION
The invention provides methods and reagents for inhibiting hyperproliferation of smooth muscle cells. The invention provides treatments for vascular pathologies associated with smooth muscle cell hyperproliferation, including restenosis. Preferably, the invention provides methods and reagents for inhibiting vascular smooth muscle cell hyperproliferation in restenosis following angioplasty, stent placement, or vein engraftment.
It has been discovered that cyclin dependent kinase inhibitors (CDKi's) can inhibit smooth muscle cell proliferation. This discovery has been exploited to develop the present invention which, in a first aspect, provides a method for inhibiting smooth muscle cell hyperproliferation comprising transducing smooth muscle cells with an effective amount of a recombinant adenovirus that lacks a functional E4 region and a functional E1 region and that comprises a transgene encoding a cyclin dependent kinase inhibitor (CDKi), wherein hyperproliferation of the transduced smooth muscle cells is inhibited. In certain embodiments of the first aspect of the invention, the cyclin dependent kinase inhibitor is selected from the group consisting of a protein from the INK4 family or an active fragment thereof; a protein from the CIP/KIP family or an active fragment thereof; and a fusion protein comprising at least an active fragment of the protein from the INK4 family and at least an active fragment of the protein from the CIP/KIP family.
In certain embodiments of the first aspect of the invention, the cyclin dependent kinase inhibitor is a protein from the CIP/KIP family of CDKi's. In certain embodiments, the CDKi is a human p27 protein (SEQ ID NO:26) or an active fragment thereof, such as an active fragment selected from the group consisting of amino acids 25-93 of p27 protein and amino acids 12-178 of p27 protein.
In certain embodiments of the first aspect of the invention, the cyclin dependent kinase inhibitor is a protein from the INK4 family of CDKi's. In certain embodiments, the cyclin dependent kinase inhibitor is a human p16 protein (SEQ ID NO:28) or an active fragment thereof.
In certain preferred embodiments of the first aspect of the invention, the cyclin dependent kinase inhibitor is derived from a mammal (e.g., a human).
In certain preferred embodiments of the first aspect of the invention, where the cyclin dependent kinase inhibitor is a fusion protein comprising at least an active fragment of a protein from the INK4 family of CDKi's and at least an active fragment of a protein from the CIP/KIP family of CDKi's, the protein from the INK4 family of CDKi's is human p16 and the protein from the CIP/KIP family of CDKi's is human p27. In other embodiments, the cyclin dependent kinase inhibitor a fusion protein comprising an active fragment of human p27 and an active fragment of human p16, and is W3 (SEQ ID NO:4), W4 (SEQ ID NO:6), W5 (SEQ ID NO:8), W6 (SEQ ID NO:10), W7 (SEQ ID NO:14), W8 (SEQ ID NO16), W9 (SEQ ID NO:20), or W10 (SEQ ID NO:22). In preferred embodiments, the dependent kinase inhibitor is W7 or W9. In a particularly preferred embodiment, the cyclin dependent kinase inhibitor is W9. In a certain embodiment, the fusion protein comprises a linker positioned between the active fragment of the first cyclin dependent kinase inhibitor and the active fragment of the second cyclin dependent kinase inhibitor.
In various embodiments of the first aspect of the invention, the recombinant adenovirus additionally lacks a functional E2 region, or a functional E3 region. In preferred embodiments, the recombinant adenovirus is replication-deficient.
In some embodiments of the first aspect of the invention, the smooth muscle cells are cultured smooth muscle cells. In certain embodiments the smooth muscle cells are in a mammal. In certain embodiments, the smooth muscle cells were induced to hyperproliferate by vascular injury. In certain embodiments, the injury was induced by angioplasty, stent placement, or vein engraftment.
In a second aspect, the invention provides a recombinant lentivirus that comprises a transgene encoding a cyclin dependent kinase inhibitor (CDKi). In certain embodiments, the cyclin dependent kinase inhibitor is selected from the group consisting of a protein fr
Finer Mitchell H.
Gyuris Jeno
McArthur James
Cell Genesys Inc.
Ketter James
Li Q Janice
Roylance Abrams Berdo & Goodman L.L.P.
LandOfFree
Methods and reagents for inhibiting proliferation of smooth... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Methods and reagents for inhibiting proliferation of smooth..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Methods and reagents for inhibiting proliferation of smooth... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2844940