Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Having -c- – wherein x is chalcogen – bonded directly to...
Reexamination Certificate
1998-06-23
2002-11-19
O'Sullivan, Peter (Department: 1621)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Having -c-, wherein x is chalcogen, bonded directly to...
C514S355000, C514S400000, C514S419000, C514S448000, C514S471000, C514S478000, C514S562000, C514S563000, C514S566000, C546S316000, C548S338100, C548S495000, C549S069000, C549S493000, C560S013000, C560S027000, C560S041000, C562S430000, C562S432000
Reexamination Certificate
active
06482849
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to small molecules that are potent inhibitors of &agr;
4
&bgr;
1
mediated adhesion to either VCAM or CS-1 and which could be useful for the treatment of inflammatory diseases.
2. Description of Related Art
The extracellular matrix (ECM) is the major component of connective tissue which provides structural integrity, and promotes cell migration and differentiation. As part of these functions, extracellular matrix molecules such as fibronectin, collagen, laminin, von Willebrand factor, thrombospondin, fibrinogen, and tenascin have been shown to support adhesion of cells in vitro. This adhesive interaction is critical for a number of biological processes including hemostasis, thrombosis, wound healing, tumor metastasis, immunity and inflammation.
Fibronectin (FN) is the prototype ECM molecule. The major cell attachment site in the fibronectin molecule has been reproduced synthetically with the amino acid sequence arginine-glycine-aspartic acid, or RGD using single letter nomenclature. Peptides containing the RGD sequence which either inhibit or promote cell adhesion have been described (U.S. Pat. Nos. 4,589,881; 4,661,111; 4,517,686; 4,683,291; 4,578,079; 4,614,517; and 4,792,525). Changes in the peptide as small as the exchange of alanine for glycine or glutamic acid for aspartic acid, which constitute the addition of a single methyl or methylene group to the tripeptide, eliminates these activities (Pierschbacher et al.,
Proc. Natl. Acad. Sci. USA
81:5985 (1984)). Recently, a second FN cell binding domain has been identified within the alternatively spliced region of the A chain of the molecule, known as the connecting segment 1 (CS-1). The most active cell-binding site within this alternatively spliced region is composed of 25 amino acids where the carboxy terminus contains the sequence EILDVPST. The amino acid sequence EILDVPST forms a recognition motif on FN for cell surface receptors. (Wayner et al.,
J. Cell Biol.
109:1321 (1989); Guan et al.,
Cell
60:53 (1990)).
The receptors which recognize these sites on FN belong to a gene superfamily called integrins which consist of heterodimeric complexes of non-covalently associated alpha and beta subunits. A common &bgr; subunit combines with unique &agr; subunits to form an adhesion receptor of defined specificity. To date, 8 &bgr; subunits have been identified which can dimerize with 16 distinct &agr; subunits forming 22 distinct integrins. The &bgr;1 subfamily, also known as the VLA family (Very Late Activation Antigens), binds to ECM molecules such as FN, collagen and laminin. For reviews, see, Hynes,
Cell
48:549 (1987); Hemler,
Annu. Rev. Immunol.
8:365 (1990). Leukocyte interaction with FN at the two spatially separate binding domains is mediated by two distinct integrins. The RGD site is recognized by the integrin &agr;
5
&bgr;
1
, while, EILDV is recognized by &agr;
4
&bgr;
1
(Pytela et al.,
Cell
40:191 (1985); Wayner et al.,
J. Cell Biol.
109:1321 (1989); Guan et al,
Cell
60:53 (1990)).
Vascular endothelial cells form the interface between blood and tissues and control the passage of leukocytes as well as plasma fluid into tissues. A variety of signals generated at the site of inflammation can activate both endothelial cells as well as circulating leukocytes so that they become more adhesive to one another. Following this initial adhesion the leukocytes migrate into the tissues to perform host defense functions. Several adhesion molecules have been identified which are involved in leukocyte-endothelial interactions.
In the &bgr;
1
subfamily, in addition to binding to fibronectin, &agr;
4
&bgr;
1
interacts with a cytokine inducible protein on endothelial cells termed vascular cell adhesion molecule (VCAM). Further involved in the leukocyte-endothelial adhesion process is the &bgr;
2
integrin subfamily. &bgr;
2
integrins include CD11a/CD18, CD11b/CD18, and CD11c/CD18. In addition, the &bgr;
7
subunit associates with &agr;
4
to form a unique &agr;
4
&bgr;
7
heterodimer which binds to FN, to VCAM, and to Mucosal Addressin Cell Adhesion Molecule-1 (MAdCAM) (Ruegg et al,
J. Cell.Biol.
117:179 (1992); Andrew et al.,
J. Immunol.
153:3847 (1994); Briskin et al.,
Nature
363:461 (1993); Shyjan et al,
J. Immunol.
156:2851 (1996)). &agr;
4
integrins are widely expressed on different cell types including hematopoietic progenitors, lymphocytes, natural killer cells, monocytes, eosinophils, basophils, and mast cells (Helmer, M. E.,
Annu. Rev. Immunol.
8:365 (1990)). Other molecules on endothelial cells which bind to the leukocytes include ICAM-1, ICAM-2, E-selectin and P-selectin (Carlos and Harlan,
Immunol. Rev.
114:1 (1990); Osborn, L.,
Cell
62:3 (1990); Springer T.,
Nature
346:425 (1990); Geng et al.,
Nature
347:757 (1990); Stoolman,
Cell
56:907 (1989)).
A number of in vitro and in vivo studies indicate that &agr;
4
&bgr;
1
plays a critical role in the pathogenesis of a variety of diseases. Monoclonal antibodies directed against &agr;
4
have been tested in a variety of disease models. Anti-&agr;
4
antibodies block adhesion of lymphocytes to synovial endothelial cells; this adhesion plays a potential role in rheumatoid arthritis (van Dinther-Janssen et al,
J. Immunol.
147:4207 (1991)). &agr;
4
has also been implicated with respect to rheumatoid arthritis in separate studies (Laffon et al,
J. Clin. Invest.
88:546 (1991); Morales-Ducret et al,
J. Immunol.
149:1424 (1992)). A significant number of studies have evaluated the role of &agr;
4
in allergy and asthma. For example, monoclonal antibodies to &agr;
4
block adhesion of basophils and eosinophils to cytokine activated endothelial cells (Walsh et al,
J. Immunol.
146:3419 (1991); Bochner et al,
J. Exp. Med.
173:1553 (1991)). Monoclonal antibodies to &agr;
4
were also effective in several lung antigen challenge models (Abraham et al,
J. Clin. Invest.
93:776 (1994); Weg et al,
J. Exp. Med.
177:561 (1993)). The cotton-top tamarin, which experiences spontaneous chronic colitis, showed a significant attenuation of their colitis when anti-&agr;
4
antibody was administered (Podolsky et al,
J. Clin. Invest.
92:372 (1993); Bell et al,
J. Immunol.
151:4790 (1993)). In a rat and mouse model, autoimmune encephalomyelitis was blocked by anti-&agr;
4
antibody (Yednock et al,
Nature
356:63 (1992); Baron et al,
J. Exp. Med.
177:57 (1993)). Anti-&agr;
4
monoclonal antibodies also inhibit insulitis and delay the onset of diabetes in the non-obese diabetic mouse (Baron et al,
J. Clin. Invest.
93:1700 (1994); Yang et al,
Proc. Natl. Acad. Sci. USA
90:10494 (1993); Burkly et al,
Diabetes
43:529 (1994)). &agr;4 is also implicated in atherosclerosis due to its endothelial expression during atherogenesis (Cybulsky et al,
Science
251:788 (1991)). The migration of leukocytes to an inflammatory site can also be blocked by anti-&agr;
4
antibodies. In addition to the blocking of migration, inhibitors of leukocyte endothelial adhesion may block the costimulatory signals mediated by integrins and thus inhibit overproduction of inflammatory cytokines. In a separate set of experiments not using anti-&agr;
4
antibodies, the peptides GRDGSP or EILDV were tested against contact hypersensitivity response. The contact hypersensitivity response was found to be blocked by GRDGSP or EILDV suggesting that both &agr;
4
&bgr;
1
and &agr;
5
&bgr;
1
are involved in this inflammatory response.
Other ailments which may involve &agr;
4
&bgr;
1
-mediated conditions include the inflammatory disorders rheumatoid arthritis, allergic disorders, asthma, spontaneous chronic colitis, insulitis, contact hypersensitivity response, atherosclerosis and autoimmune encephalomyelitis. These studies illustrate that small molecules that are potent inhibitors of &agr;
4
&bgr;
1
mediated adhesion to either VCAM-1 or CS-1 may be used as a form of treatment in numerous inflammatory diseases. However, these inflammatory conditions could be expanded to include adult respiratory distress
Chrusciel Robert A.
Fisher Jed F.
Lobl Thomas J.
Polinsky Alexander
Rishton Gilbert M.
Birch & Stewart Kolasch & Birch, LLP
O'Sullivan Peter
Tanabe Seiyaku Co. Ltd.
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