Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Carbohydrate doai
Reexamination Certificate
1999-12-17
2001-08-28
Fonda, Kathleen K. (Department: 1623)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Carbohydrate doai
C514S025000, C514S027000, C514S028000, C514S029000, C514S030000, C514S031000, C435S184000, C530S351000, C530S396000, C530S399000
Reexamination Certificate
active
06281197
ABSTRACT:
TECHNICAL FIELD
The present invention relates generally to inhibitors of leaderless protein export, and more specifically, to the use of cardiac glycosides and aglycone derivatives to inhibit export of leaderless proteins into extracellular spaces.
BACKGROUND OF THE INVENTION
Many proteins exert an effect on cell growth, differentiation, and inflammation through signal transduction, mediated by binding to a cell surface receptor. Yet other proteins such as factors that initiate or are necessary the blood clot formation act enzymatically in blood. While these actions are generally part of normal processes, under certain circumstances, it may be desirable to limit or inhibit the action of certain proteins and the effects of subsequent signaling. For example, tumor growth promoted by a growth factor, such as bFGF acting on melanoma cells, is deleterious and often leads to fatalities.
Approaches to inhibit specific proteins have concentrated primarily on interfering with protein-substrate or protein-receptor interactions. Typically, this involves using an antibody or other molecule that competitively binds the protein, by administration of competitors for receptor binding, or by protease digestion of the protein. An alternative approach, not generally pursued, is to reduce the level of the protein by inhibiting its expression at a transcriptional or translational level. Methods of reducing protein levels by inhibiting the trascription or translation of the protein have been difficult to achieve because of inherent problems of inhibiting the specific expression of one or a few proteins.
The discovery that certain proteins, such as growth factors, mediators of inflammation, and mediators of blood clotting, are exported through a nonclassical secretory pathway allows the development of specific inhibitors for these proteins. These proteins are identified by their lack of a hydrophobic leader sequence that mediates secretion by the classical Golgi/ER pathway. These proteins are believed to be exported from a cell by exocytosis.
This invention provides inhibitors of the export of these leaderless proteins, allowing control of undesired proliferation and inflammation, as well as other related advantages.
SUMMARY OF THE INVENTION
The present invention generally provides methods of inhibiting the export of a leaderless protein from a cell expressing the protein. In one aspect of the invention, export is inhibited by contacting a cell expressing the protein with a cardiac glycoside. In certain embodiments, the cardiac glycoside is selected from the group consisting of digoxin, strophantin K, digitoxin, lanatoside A and ouabain. Preferably the cardiac glycoside is ouabain or digoxin.
In another aspect of the invention, methods of inhibiting the export of a leaderless protein from a cell expressing the protein by contacting the cell with an aglycone derivative of a cardiac glycoside are provided. In certain embodiments, the aglycone derivative is selected from the group consisting of digoxigenin, digitoxigenin and uzarigenin. Preferably the aglycone derivative is digoxigenin.
In other aspects, methods are provided for inhibiting the export of FGF-2 from a cell expressing FGF-2, comprising contacting the cell with a cardiac glycoside or aglycone derivative of a cardiac glycoside. In yet other aspects, methods of treating an FGF-mediated pathophysiological condition in a patient are provided, comprising administering a therapeutically effective dosage of a cardiac glycoside or aglycone derivative of a cardiac glycoside, thereby reducing the amount of FGF-2 that is exported. In certain embodiments, the pathophysiological condition is melanoma, ovarian carcinoma, tetracarcinoma and neuroblastoma.
In yet other aspects, methods are provided for inhibiting proliferation of a cell bearing an FGF receptor, comprising contacting the cell with a cardiac glycoside or an aglycone derivative of a cardiac glucoside. In still other aspects, methods are provided for treating complications of diabetes, comprising contacting a cell with an inhibiting amount of a cardiac glycoside or aglycone derivative.
Methods are also provided for inhibiting export of leaderless proteins, comprising treating cells with a compound selected from the group consisting of formula 1, formula 2, formula 3, formula 4, or formula 5.
These and other aspects of the present invention will become evident upon reference to the following detailed description and attached drawings. Various references are set forth below which described in more detail certain procedures or compositions (e.g., plasmids, etc.). All of these references are incorporated by reference in their entirety.
REFERENCES:
patent: 5545623 (1996-08-01), Matsumori
patent: 5627195 (1997-05-01), Hu
patent: 5872103 (1999-02-01), Belletti
patent: WO 96/04291 (1996-02-01), None
Katahira et al.ChemotherapyJul. 1991, 39(7), 678-686.*
Callard et al. “The Cytokine FactsBook” Academic Press Limited, London, 1994, pp. 31-38.*
Brooks, D., “Protein Secretion by the Rat Epididymis Can Be Selectively Modified in Vitro by Local Anesthetics, Glucose Deprivation, Dinitrophenol, Ouabain and Ionophores,”J. Androl.5: 351-360, 1984.
Cooper and Barondes, “Evidence for Export of a Muscle Lectin from Cytosol to Extracellular Matrix and for a Novel Secretory Mechanism,”Journal of Cell Biology110:1681-1691, 1990.
Crutchely and Smariga, “Monovalent Cation Dependence of Tissue Plasminogen Activator Synthesis by HeLa Cells,”Journal of Biological Chemistry262(7): 3017-3021, 1987.
Florkiewicz et al., “Multiple Forms of bFGF: Differential Nuclear and Cell Surface Localizaton,”Growth Factors4:265-275, 1991.
Florkiewicz et al., “Quantitative Export of FGF-2 Occurs Through an Alternative, Energy-Dependent, Non-ER/Golgi Pathway,”Journal of Cellular Physiology162: 388-399, 1995.
Jackson et al., “Heath shock induces the release of fibroblast growth factor 1 from NIH 3T3 cells, ”Proc. Natl. Acad. Sci. USA 89: 10691-10695, 1992.
Jackson et al., “The Release of Fibroblast Growth Factor-1 from NIH 3T3 Cells In Response to Temperature Involves the Function of Cysteine Residues,”Journal of Biological Chemisty 270(1): 33-36, 1995.
Mignatti et al., “Basic Fibroblast Growth Factor, a Protein Devoid of Secretory Signal Sequence, Is Released by Cells via a Pathway Independent of the Endoplasmic Reticulum-Golgi Complex,”Journal of Cellular Physiology 151: 81-93, 1992.
Repke et al., “Potential suitablility of NA({circumflex over ( )}+) /K({circumflex over ( )}+) -transporting ATPase in pre-screens for anti-cancer agents,”Anti-Cancer Drug Design 10:177-187, 1995.
Rubartelli et al., “A novel secretory pathway for interleukin-1.beta., a protein lackin a signal sequence,”EMBO Journal 9(5): 1503-1510, 1990.
Rubartelli et al., “Post-Translational Regulation of Interleukin 1.beta. Secretion,”Cytokine5(2): 117-134, 1993.
Rubartelli et al., “Secretion of Thioredoxin by Normal and Neoplastic Cells through a Leaderless Secretory Pathway,”Journal of Biological Chemistry 267(34): 24161-14164, 1992.
Siders and Mizel, “Interleukin-1.beta. Secretion. A Possilbe Multistep Process that is Regulated in a Cell Type-Specific Manner,”Journal of Biological Chemistry 270(27):16258-16264, 1995.
Fonda Kathleen K.
Scripps Research Institute
Seed Intellectual Property Law Group PLLC
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