TNF-α converting enzyme inhibitory agents and method of using same
Inventors
Levine, Stewart J. • Buckley, Caitriona A. • Rouhani, Farshid N. • Kaler, Maryann
Assignees
SECRETARY OF DEPARTMENT OF HEALTH AND HUMAN SERVICES GOVERNMENT OF United States, AS REPRESENTED BY • US Department of Health and Human Services
Publication Number
US-7655752-B2
Publication Date
2010-02-02
Expiration Date
2024-09-24
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Abstract
The invention provides peptides, variants of peptides, peptide fragments, and peptidomimetics that can inhibit the protease activity of tumor necrosis factor alpha converting enzyme. The invention also provides coupled proteins containing a partner protein coupled to a peptide, peptide fragment, or peptidomimetic. The invention also provides polyproteins containing at least two peptides, peptide fragments, or coupled proteins that are connected through a linker. Isolated nucleic acid segments, expression cassettes, and nucleic acid constructs are also provided by the invention. The invention also provides antibodies and aptamers. Pharmaceutical compositions are provided by the invention. Methods to lower or increase levels of active tumor necrosis factor alpha in a mammal are also provided.
Core Innovation
The invention provides peptides, variants of peptides, peptide fragments, and peptidomimetics that inhibit the protease activity of tumor necrosis factor alpha converting enzyme (TACE). It also provides coupled proteins containing a partner protein coupled to such peptides, peptide fragments, or peptidomimetics, and polyproteins comprising at least two of these elements connected through a linker. Moreover, the invention includes isolated nucleic acid segments encoding these peptides, expression cassettes, nucleic acid constructs, antibodies, and aptamers. Pharmaceutical compositions containing these agents and methods to modulate levels of active tumor necrosis factor alpha in a mammal are also described.
The problem addressed by the invention relates to the regulation of TACE, a zinc metalloprotease that is important in inflammation, immune regulation, and cellular proliferation. TACE cleaves membrane-bound precursors of tumor necrosis factor alpha (TNF-α) and other cell surface proteins, playing a key role in multiple disease processes. Controlling TACE activity is necessary to prevent excessive or unanticipated cleavage and shedding of target membrane proteins, which are implicated in diseases such as arthritis, sepsis, inflammatory bowel disease, tumor metastasis, and more. Prior approaches lacked agents to specifically inhibit or activate TACE through an independent mechanism.
The invention is based on the discovery that the amino-terminus of the TACE prodomain possesses TACE inhibitory activity independent of the previously known cysteine-switch mechanism. A novel, endogenous mRNA sequence encoding a soluble protein named N-TACE, corresponding to the first 54 amino acids of TACE including the signal peptide but excluding the cysteine-switch motif, was identified. Peptides derived from N-TACE, particularly a leucine-rich 19-amino acid segment (positions 30-48), were found to specifically inhibit TACE protease activity in vitro and in cell-based assays, decreasing shedding of TNF-α receptor substrates.
Claims Coverage
The patent includes multiple claims with several independent claims focused on isolated peptides, including specific sequences and fragments, polyproteins, coupled proteins, and pharmaceutical compositions, as well as methods for lowering tumor necrosis factor alpha levels in mammals.
Isolated peptides that inhibit TACE protease activity
Isolated peptides consisting of sequences SEQ ID NOs: 3-8 or inhibitory fragments thereof specifically inhibit the protease activity of tumor necrosis factor alpha converting enzyme. Particular fragments comprising SEQ ID NOs: 4, 7, or 8 are included.
Peptides with terminal blockers
Peptides may include an amino-terminal blocker, carboxyl-terminal blocker, or both to enhance stability and resistance to degradation.
Peptides chemically or recombinantly coupled to partner proteins
Isolated peptides can be chemically coupled or fused to partner proteins such as carrier proteins (e.g., keyhole limpet hemacyanin, bovine serum albumin, ovalbumin, serum albumins) or other proteins that enhance immunogenicity, solubility, detection, separation, or cellular export/localization. Such coupled proteins may lack cysteine switch motifs.
Polyproteins comprising multiple peptides connected by cleavable linkers
Polyproteins include at least two peptides connected by linkers cleavable via chemicals or proteases and optionally linked to a partner protein through a separate cleavable linker.
Pharmaceutical compositions containing the isolated peptides
Pharmaceutical compositions comprising pharmaceutically acceptable carriers and isolated peptides as defined are claimed.
Method for lowering levels of active tumor necrosis factor alpha
A method to lower active tumor necrosis factor alpha levels in a mammal comprising administering a tumor necrosis factor alpha lowering amount of the isolated inhibitory peptides is claimed.
The claims cover isolated peptides and peptide fragments that inhibit TACE protease activity, their formulations in pharmaceutical compositions, coupling to partner proteins, assembly into polyproteins with cleavable linkers, and methods for lowering active TNF-α levels in mammals through administration of these peptides.
Stated Advantages
Provides peptides that inhibit TACE activity via a novel cysteine-switch independent mechanism.
Specific inhibition of TACE to modulate TNF-α levels, potentially reducing excessive inflammation and associated diseases.
Peptides can be formulated with terminal blockers and coupled to proteins to enhance stability, immunogenicity, and delivery.
Methods enabled to both lower and increase TACE activity, allowing therapeutic flexibility.
Documented Applications
Treatment of inflammatory diseases associated with elevated TNF-α such as arthritis, sepsis, ulcerative colitis, Crohn's disease, septic shock, graft rejection, and inflammatory bowel disease.
Modulation of growth hormone related pathologies by regulating soluble growth hormone binding protein levels via TACE inhibition or stimulation.
Reduction of tumor cell proliferation, migration, invasion, and metastasis by inhibiting TACE mediated release of epidermal growth factor receptor ligands.
Inducing apoptosis of cancer cells by increasing TNF-α levels through stimulation of TACE activity.
Alleviation of Alzheimer's disease by stimulating TACE activity to increase amyloid precursor protein cleavage, thus reducing amyloid beta formation.
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