Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Peptide containing doai
Reexamination Certificate
1999-02-05
2003-04-29
Carlson, Karen Cochrane (Department: 1653)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Peptide containing doai
C514S002600, C514S014800, C514S017400, C514S021800, C530S300000, C530S326000, C530S327000, C530S329000
Reexamination Certificate
active
06555522
ABSTRACT:
1. FIELD OF THE INVENTION
The present invention relates generally to the field of peptides and other small molecules (i.e. peptide mimetics) as pharmaceutical and/or therapeutic agents, and to methods for identification and design of peptides and peptide mimetics having desired functional activities. Specifically, peptides and other small molecules derived from regions of interacting intracellular signaling proteins are provided. More specifically, peptides and other small molecules derived from regions of the G&bgr; subunit of heterotrimeric GTP binding proteins are provided. Such molecules include specific agonists and antagonists of G&bgr; downstream effectors, including adenylyl cyclase and phospholipase C. Such molecules are targeted to predicted regions of interaction between intracellular signaling proteins and tested for activity in functional assays using methods of the invention. One major advantage of the invention is the incorporation of three-dimensional structural information in models used for predicting interaction surfaces between intracellular proteins. Another major advantage is the ability to distinguish, within a predicted interaction surface, a signal transfer region from a general binding domain. Resolution of such signal transfer regions from general binding domains is useful for prediction and validation of pharmacologic and therapeutic agonists and antagonists.
2. BACKGROUND OF THE INVENTION
The ability to target a desired drug intervention to a specific site in a biological system underlies the rational design of safe and effective drugs. Past drug design efforts have often focused on development of molecules believed to interact with cell surface receptors. For example, high-throughput assays have been used to screen synthetic organic compounds to identify molecules interacting with an extracellular domain of a cell surface receptor (Tian et al., 1998, A small, nonpeptidyl mimic of granulocyte-colony-stimulating factor, Science 281, 257-259). Further, methods have been developed for determining whether a candidate compound is an agonist of a peptide hormone receptor (see Kopin et al., U.S. Pat. No. 5,750,353, issued May 12, 1998, Assay for non-peptide agonists to peptide hormone receptors). Peptides and mimetics have also been developed based on the transmembrane domains of G-protein-coupled receptors (Bouvier et al., Jan. 8, 1998, Peptides and peptidomimetic compounds affecting the activity of G-protein-coupled receptors by altering receptor oligomerization, International Publication No. WO 98/00538). Examples of other extracellular ligands for which peptide mimetics have been developed include erythropoietin and TNF&agr; (Wrighton et al., 1997, Increased potency of an erythropoietin peptide mimetic through covalent dimerization, Nature Biotechnology 15, 1261-1265; Takasaki et al., 1997, Structure-based design and characterization of exocyclic peptidomimetics that inhibit TNF&agr; binding to its receptor, Nature Biotechnology 15, 1266-1270). Finally, distinct regions of peptide hormones have even been considered for design of receptor antagonists (Portoghese et al., 1990, Design of peptidomimetic &dgr; opioid receptor antagonists using the message-address concept, J. Med. Chem. 33, 1714-1720).
Heterotrimeric GTP-binding proteins (G proteins) consisting of G&agr;&bgr;&ggr; subunits are ubiquitous signal transduction proteins that play essential roles in intracellular communication (see e.g. DeVivo and Iyengar, 1994, G protein pathways: signal processing by effectors, Molec. Cell. Endocrinol. 100, 65-70). For example, the enzymatic production of cyclic AMP (cAMP) via adenylyl cyclases is regulated by G proteins (Smit and Iyengar, 1998, Mammalian adenylyl cyclases, Adv. Sec. Mess. Phosphoprot. Res. 32, 1-21; Iyengar, 1993, Multiple families of Gs-regulated adenylyl cyclases, Adv. Sec. Mess. Phosphoprot. Res. 28, 27-36; Pieroni et al., 1993, Signal recognition and integration by Gs-stimulated adenylyl cyclases, Curr. Opin. Neurobiol. 3, 345-351; Weng et al., 1996, G beta subunit interacts with a peptide encoding region 956-982 of adenylyl cyclase 2, cross-linking of the peptide to free G beta gamma but not the heterotrimer, J. Biol. Chem. 271, 26445-264488; Harry et al., 1997, Differential regulation of adenylyl cyclases by G alphas, J. Biol. Chem. 272, 19017-19021). G proteins provide a versatile system for investigation of intracellular protein-protein interactions by virtue of their interactions with multiple downstream effectors. For example, G protein &bgr;&ggr; subunits regulate the activity of not on adenylyl cyclase but also phospholipase C-&bgr;2, calcium channels, potassium channels, and &bgr;-adrenergic receptor kinase (see e.g. Ford et al., 1998, Molecular basis for interactions of G protein &bgr;&ggr; subunits with effectors, Science 280, 1271-1274).
Drug intervention beyond the cell surface, i.e. at intracellular protein-protein interaction sites, would broaden the array of potential targets for achieving a desired therapeutic effect. Intracellular targets may also provide intervention points having enhanced specificity compared to drugs targeted strictly at cell surface receptors. The ability to use intracellular interacting proteins as therapeutic targets for drug design has been less clearly established, however. One reason may be that an intracellular protein-protein interaction, unlike a typical cell surface hormone-receptor interaction, will often involve a multiplicity of proteins. Thus, resolution of specific interactions among three or more proteins will often be necessary to carry out design of safe and effective drugs. Accordingly, a need exists for a generally-applicable approach for identification of peptides and mimetics thereof having selective activity at a chosen intracellular site of action.
3. SUMMARY OF THE INVENTION
This invention provides peptides and other small molecules derived from regions of intracellular interacting proteins and methods for identification of such molecules. More specifically, the present invention provides peptides and other small molecules derived from regions of G&bgr; proteins which function as agonists or antagonists of adenylyl cyclase or phospholipase C-&bgr;2. The invention is based, at least in part, on the discovery of the inventors that it is possible to resolve, within a given intracellular signal transduction protein, a signal transfer region from a general binding domain. Such resolution provides a rational basis for design of agonists and antagonists of virtually any desired intracellular protein-protein interaction. The drug design methods of the invention utilize three-dimensional structural information for prediction of protein-protein interactions followed by evaluation of predictions in functional assays.
The present invention relates generally to the field of peptides and peptide mimetics as pharmaceutical and/or therapeutic agents. More particularly, the present invention relates to peptides and other small molecules (e.g. peptide mimetics) derived from regions of G&bgr; proteins and their use as pharmaceutical and/or therapeutic agents. For example, peptides and derivatives thereof for modulating adenylyl cyclase and phospholipase C-&bgr;2 activities are provided. Still further, methods for identification of peptides and derivatives thereof useful for modulating a chosen effector-of-interest among various effectors are provided. One advantage of the methods of the invention is the use of structural modeling information to predict and validate pharmacologic and therapeutic agents.
Predictions about effector interactions of G&bgr; proteins have been made using a combination of molecular modeling and experimental validation in which the predictions of the model are tested. Through an iterative process involving cycles of structural modeling followed by experimental testing, precise definition of individual effector domains within a G&bgr; signaling protein has been achieved. This validated procedure has general applicability for drug design targeted at other intracellular pr
Buck Elizabeth
Chen Yibang
Iyengar Srinivas Ravi V.
Weinstein Harel
Weng Gezhi
Carlson Karen Cochrane
Chism Billy D
Mount Sinai School of Medicine of the City of New York
Pennie & Edmonds LLP
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