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WO2008150981A2 - Nouveaux peptides courts inhibant l'ouverture de jonctions serrées mammaliennes - Google Patents

Nouveaux peptides courts inhibant l'ouverture de jonctions serrées mammaliennes Download PDF

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Publication number
WO2008150981A2
WO2008150981A2 PCT/US2008/065268 US2008065268W WO2008150981A2 WO 2008150981 A2 WO2008150981 A2 WO 2008150981A2 US 2008065268 W US2008065268 W US 2008065268W WO 2008150981 A2 WO2008150981 A2 WO 2008150981A2
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Prior art keywords
peptide
seq
tight junctions
antagonist
acid sequence
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Ceased
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PCT/US2008/065268
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WO2008150981A3 (fr
Inventor
Amir Tamiz
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9 Meters Biopharma Inc
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Alba Therapeutics Corp
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Publication of WO2008150981A2 publication Critical patent/WO2008150981A2/fr
Publication of WO2008150981A3 publication Critical patent/WO2008150981A3/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/08—Tripeptides
    • C07K5/0802—Tripeptides with the first amino acid being neutral
    • C07K5/0804—Tripeptides with the first amino acid being neutral and aliphatic
    • C07K5/0806—Tripeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atoms, i.e. Gly, Ala
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
    • C07K14/28—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria from Vibrionaceae (F)
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/08—Tripeptides
    • C07K5/0819—Tripeptides with the first amino acid being acidic
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/10—Tetrapeptides
    • C07K5/1002—Tetrapeptides with the first amino acid being neutral
    • C07K5/1005—Tetrapeptides with the first amino acid being neutral and aliphatic
    • C07K5/1008—Tetrapeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atoms, i.e. Gly, Ala
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/10—Tetrapeptides
    • C07K5/1002—Tetrapeptides with the first amino acid being neutral
    • C07K5/1005—Tetrapeptides with the first amino acid being neutral and aliphatic
    • C07K5/101—Tetrapeptides with the first amino acid being neutral and aliphatic the side chain containing 2 to 4 carbon atoms, e.g. Val, Ile, Leu
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • Tight junctions act as a barrier between apical and basolateral compartments, selectively regulating the passive diffusion of ions and water-soluble solutes through the paracellular (between cells) pathway (Gumbiner, Am. J. Physiol., 253 (Cell Physiol. 22):C749-C758 (1987)). This barrier maintains any gradient generated by the activity of pathways associated with the transcellular route (Diamond, Physiologist, 20:10-18 (1977)).
  • Tight junction dysfunction occurs in a variety of clinical conditions, including food allergies, infections of the gastrointestinal tract, autoimmune diseases, celiac disease and inflammatory bowel diseases (Fasano A. 2001. Pathological and therapeutical implications of macromolecule passage through the tight junction. In Tight Junctions, CRC Press, Inc., Boca Raton, FL. 697-722.). Healthy, mature gut mucosa with its intact tight junction serves as the main barrier to the passage of macromolecules. During the healthy state, small quantities of immunologically active antigens cross the gut host barrier. These antigens are absorbed across the mucosa through at least two pathways.
  • Celiac disease is a chronic autoimmune disease that is HLA-DQ2/DQ8 haplotype restricted.
  • Glutens the major protein fraction of wheat, and related proteins in rye and barley are the triggering agents of the disease.
  • Ingested gluten or its' derivative fractions gliadin and subunits
  • tTG tissue transglutaminase
  • IBD Inflammatory bowel disease
  • Crohn's disease Crohn's disease
  • UC ulcerative colitis
  • ZOT Zonula occludens toxin
  • Fasano et al. Proc. Natl. Acad. Sci., USA, 8:5242-5246 (1991)
  • ZOT increases the intestinal permeability of rabbit ileal mucosa by modulating the structure of intercellular tight junctions.
  • Mammalian proteins that are immunologically and functionally related to ZOT have been identified. See US 5,945,510. These proteins, referred to as "zonulin,” function as the physiological effector of mammalian tight junctions. These proteins are useful for enhancing absorption of therapeutic agents across tight junction of intestinal and nasal mucosa, as well as across tight junction of the blood brain barrier.
  • Peptide antagonists of tight junction opening were described in U.S. Patent 6,458,925, which is incorporated by reference herein in its entirety, which corresponds to WO 00/07609.
  • Peptide antagonists of tight junction opening may bind to the receptor utilized by the zonnula occludens toxin expressed by Vibrio cholerae, yet not function to physiologically modulate the opening of mammalian tight junctions.
  • the peptide antagonists may competitively inhibit the binding of ZOT and/or zonulin to the ZOT receptor, thereby inhibiting the ability of ZOT and/or zonulin to physiologically modulate the opening of mammalian tight junctions.
  • the present invention provides peptide antagonists of tight junctions.
  • Peptide antagonists of tight junctions according to the invention are preferably 7 amino acids or shorter in length.
  • peptide antagonists of the invention may comprise 3-7 amino acids.
  • a peptide antagonist of the invention may comprise 3 amino acids.
  • a peptide antagonist of the invention may consist of, or may consist essentially of, 3 amino acids.
  • a peptide antagonist of the invention may comprise the amino acid sequence Q-P-G (SEQ ID NO:6; corresponding to residues 6-8 of SEQ ID NO:1).
  • a peptide antagonist of the invention may consist of a peptide having the sequence Q-P-G.
  • a peptide antagonist of the invention may comprise the amino acid sequence G-G-V (SEQ ID NO: 11 ;corresponding to residues 1 -3 of SEQ ID NO: 1 ).
  • a peptide antagonist of the invention may consist of a peptide having the sequence G-G-V.
  • the invention provides methods of treating an excessive or undesirable permeability of a tissue containing tight junctions comprising administering to a subject in need thereof a composition comprising a tight junction antagonist.
  • a "subject" may be any mammal, for example, a human, dog, cat, horse, cow, etc.
  • a subject may be a human.
  • a subject may be a dog.
  • Compositions of the invention may comprise one or more peptide antagonists of tight junctions that may be 7 amino acids or shorter in length.
  • compositions of the invention may comprise one or more peptide antagonists that may comprise 3-7 amino acids.
  • compositions of the invention may comprise one or more peptide antagonists of the invention that may comprise 3 amino acids.
  • compositions of the invention may comprise a peptide antagonist of the invention that may consist of, or may consist essentially of, 3 amino acids.
  • compositions of the invention may comprise a peptide antagonist comprising the amino acid sequence Q-P-G (SEQ ID NO:6; corresponding to residues 6-8 of SEQ ID NO:1).
  • compositions of the invention may comprise a peptide antagonist that consists of a peptide having the sequence Q-P-G.
  • compositions of the invention may comprise a peptide antagonist comprising the amino acid sequence G-G-V (SEQ ID NO: 11 ; corresponding to residues 1 -3 of SEQ ID NO: 1 ).
  • compositions of the invention may comprise a peptide antagonist that consists of a peptide having the sequence G-G-V.
  • Compositions of the invention may comprise one or more tight junction antagonists and one or more therapeutic agents. Suitable therapeutic agents include, but are not limited to, aminosalicylates, corticosteroids, immunomodulators, antibiotics, cytokines, chemokines and biologic therapeutics.
  • the invention provides methods of treating celiac disease comprising administering to a subject in need thereof a composition comprising a tight junction antagonist.
  • a composition suitable for treating celiac disease may comprise a tight junction antagonist and may be a delayed release composition.
  • Compositions suitable for use in treating celiac disease may comprise one or more peptide antagonists of tight junctions that may be 7 amino acids or shorter in length.
  • compositions for use in treating celiac disease may comprise one or more peptide antagonists that may comprise 3-7 amino acids.
  • compositions for use in treating celiac disease may comprise one or more peptide antagonists of the invention that may comprise 3 amino acids.
  • compositions of the invention may comprise a peptide antagonist of the invention that may consist of, or may consist essentially of, 3 amino acids.
  • compositions for use in treating celiac disease may comprise a peptide antagonist comprising the amino acid sequence Q-P-G (SEQ ID NO:6;corresponding to residues 6-8 of SEQ ID NO:1).
  • compositions for use in treating celiac disease may comprise a peptide antagonist that consists of a peptide having the sequence Q-P-G.
  • compositions for use in treating celiac disease may comprise a peptide antagonist comprising the amino acid sequence G-G-V (SEQ ID NO: 11 ;corresponding to residues 1 -3 of SEQ ID NO: 1 ).
  • compositions for use in treating celiac disease may comprise a peptide antagonist that consists of a peptide having the sequence G-G-V.
  • Compositions for use in treating celiac disease may comprise one or more tight junction antagonists and one or more therapeutic agents. Suitable therapeutic agents include, but are not limited to, aminosalicylates, corticosteroids, immunomodulators, antibiotics, cytokines, chemokines, and biologic therapeutics.
  • the invention provides methods of treating inflammatory bowel disease comprising administering to a subject in need thereof a composition comprising a tight junction antagonist.
  • a composition suitable for treating IBD may comprise a tight junction antagonist and may be a delayed release composition.
  • Compositions suitable for use in treating IBD may comprise one or more peptide antagonists of tight junctions that may be 7 amino acids or shorter in length.
  • compositions for use in treating IBD may comprise one or more peptide antagonists that may comprise 3-7 amino acids.
  • compositions for use in treating IBD may comprise one or more peptide antagonists of the invention that may comprise 3 amino acids.
  • compositions of the invention may comprise a peptide antagonist of the invention that may consist of, or may consist essentially of, 3 amino acids
  • compositions for use in treating IBD may comprise a peptide antagonist comprising the amino acid sequence Q-P-G (SEQ ID NO:6; corresponding to residues 6-8 of SEQ ID NO: 1).
  • compositions for use in treating IBD may comprise a peptide antagonist that consists of a peptide having the sequence Q-P-G.
  • compositions for use in treating IBD may comprise a peptide antagonist comprising the amino acid sequence G-G-V (SEQ ID NO:11; corresponding to residues 1-3 of SEQ ID NO: 1).
  • compositions for use in treating IBD may comprise a peptide antagonist that consists of a peptide having the sequence G-G-V.
  • Compositions for use in treating IBD may comprise one or more tight junction antagonists and one or more therapeutic agents. Suitable therapeutic agents include, but are not limited to, aminosalicylates, corticosteroids, immunomodulators, antibiotics, cytokines, chemokines, and biologic therapeutics.
  • the present invention provides methods and materials for treating Crohn's disease.
  • the invention provides methods of treating Crohn's disease comprising administering to a subject in need thereof a composition comprising a tight junction antagonist.
  • a composition suitable for treating Crohn's may comprise a tight junction antagonist and may be a delayed release composition.
  • Compositions suitable for use in treating Crohn's may comprise one or more peptide antagonists of tight junctions that may be 7 amino acids or shorter in length.
  • compositions for use in treating Crohn's may comprise one or more peptide antagonists that may comprise 3-7 amino acids.
  • compositions for use in treating Crohn's may comprise one or more peptide antagonists of the invention that may comprise 3 amino acids.
  • compositions of the invention may comprise a peptide antagonist of the invention that may consist of, or may consist essentially of, 3 amino acids.
  • compositions for use in treating Crohn's may comprise a peptide antagonist comprising the amino acid sequence Q-P-G (SEQ ID NO:6; corresponding to residues 6-8 of SEQ ID NO:1).
  • compositions for use in treating Crohn's may comprise a peptide antagonist that consists of a peptide having the sequence Q-P-G.
  • compositions for use in treating Crohn's may comprise a peptide antagonist comprising the amino acid sequence G-G-V (SEQ ID NO:11; corresponding to residues 1-3 of SEQ ID NO:1).
  • compositions for use in treating Crohn's may comprise a peptide antagonist that consists of a peptide having the sequence G-G-V.
  • Compositions for use in treating Crohn's may comprise one or more tight junction antagonists and one or more therapeutic agents. Suitable therapeutic agents include, but are not limited to, aminosalicylates, corticosteroids, immunomodulators, antibiotics, and biologic therapeutics.
  • the present invention provides methods and materials for treating ulcerative colitis.
  • the invention provides methods of treating ulcerative colitis comprising administering to a subject in need thereof a composition comprising a tight junction antagonist.
  • a composition suitable for treating ulcerative colitis may comprise a tight junction antagonist and may be a delayed release composition.
  • Compositions suitable for use in treating ulcerative colitis may comprise one or more peptide antagonists of tight junctions that may be 7 amino acids or shorter in length.
  • compositions for use in treating ulcerative colitis may comprise one or more peptide antagonists that may comprise 3-7 amino acids.
  • compositions for use in treating ulcerative colitis may comprise one or more peptide antagonists of the invention that may comprise 3 amino acids.
  • compositions of the invention may comprise a peptide antagonist of the invention that may consist of, or may consist essentially of, 3 amino acids.
  • compositions for use in treating ulcerative colitis may comprise a peptide antagonist comprising the amino acid sequence Q-P-G (SEQ ID NO:6; corresponding to residues 6-8 of SEQ ID NO: 1).
  • compositions for use in treating ulcerative colitis may comprise a peptide antagonist that consists of a peptide having the sequence Q-P-G.
  • compositions for use in treating ulcerative colitis may comprise a peptide antagonist comprising the amino acid sequence G-G-V (SEQ ID NO:11; corresponding to residues 1-3 of SEQ ID NO:1).
  • compositions for use in treating ulcerative colitis may comprise a peptide antagonist that consists of a peptide having the sequence G-G-V.
  • compositions for use in treating ulcerative colitis may comprise one or more tight junction antagonists and one or more therapeutic agents. Suitable therapeutic agents include, but are not limited to, aminosalicylates, corticosteroids, immunomodulators, antibiotics, and biologic therapeutics.
  • the instant invention includes: (1) A peptide antagonist of tight junctions, wherein the peptide is 7 amino acids or less in length;
  • a peptide antagonist of tight junctions according to (1), wherein the peptide consists of 3 amino acids;
  • a method of treating an excessive or undesirable permeability of a tissue containing tight junctions comprising: administering to a subject in need thereof a composition comprising a peptide according to (1);
  • composition further comprises a therapeutic agent
  • a method of treating inflammatory bowel disease comprising: administering to a subject in need thereof a composition comprising a peptide according to (1);
  • composition further comprises a therapeutic agent
  • therapeutic agent is selected from the group consisting of aminosalicylates, corticosteroids, immunomodulators, antibiotics, cytokines, chemokines and biologic therapeutics;
  • a method of treating Crohn's disease comprising: administering to a subject in need thereof a composition comprising a peptide according to (1);
  • (31) A method according to (30), wherein the peptide is selected from the group consisting of Q-P-G and G-G-V;
  • composition further comprises a therapeutic agent
  • a method of treating ulcerative colitis comprising: administering to a subject in need thereof a composition comprising a peptide according to (1);
  • composition further comprises a therapeutic agent
  • therapeutic agent is selected from the group consisting of aminosalicylates, corticosteroids, immunomodulators, antibiotics, cytokines, chemokines and biologic therapeutics;
  • tight junction antagonists prevent, inhibit or reduce the opening of tight junctions, for example, the opening of tight junctions induced by a tight junction agonist.
  • a tight junction antagonist may bind to the receptor that mediates tight junction agonist induced opening of tight junctions.
  • a tight junction antagonist may bind to the ZOT receptor and prevent, inhibit, reduce or reverse the tight junction opening triggered by the tight junction agonist ZOT.
  • a subject is any animal, e.g., mammal, upon which methods of the invention may be practiced and/or to which materials of the present invention may be administered.
  • Subjects include, but are not limited to, humans.
  • Antagonists of the invention may comprise peptide antagonists.
  • An example of a peptide antagonist of tight junctions is a peptide that comprises the amino acid sequence Gly Gly VaI Leu VaI Gln Pro Gly (SEQ ID NO:1).
  • peptide antagonists of the invention include, but are not limited to, peptides that comprise amino acids 1 -3 of SEQ ID NO: 1 (SEQ ID NO:11), amino acids 1-4 of SEQ ID NO: 1 (SEQ ID NO:10), amino acids 1-5 of SEQ ID NO: 1 (SEQ ID NO:9), amino acids 1-6 of SEQ ID NO: 1 (SEQ ID NO:8), amino acids 1-7 of SEQ ID NO: 1 (SEQ ID NO:7), amino acids 2-8 of SEQ ID NO: 1 (SEQ ID NO:2), amino acids 3-8 of SEQ ID NO: 1 (SEQ ID NO:3), amino acids 4-8 of SEQ ID NO: 1 (SEQ ID NO:4), amino acids 5-8 of SEQ ID NO: 1 (SEQ ID NO:5), and amino acids 6-8 of SEQ ID NO: 1 (SEQ ID NO:6).
  • peptide antagonists of the invention include, but are not limited to, peptides that consist essentially of amino acids 1-3 of SEQ ID NO: 1 (SEQ ID NO: 11), amino acids 1-4 of SEQ ID NO: 1 (SEQ ID NO: 10), amino acids 1-5 of SEQ ID NO: 1 (SEQ ID NO:9), amino acids 1-6 of SEQ ID NO: 1 (SEQ ID NO:8), amino acids 1-7 of SEQ ID NO: 1 (SEQ ID NO:7), amino acids 2-8 of SEQ ID NO: 1 (SEQ ID NO:2), amino acids 3-8 of SEQ ID NO: 1 (SEQ ID NO:3), amino acids 4-8 of SEQ ID NO: 1 (SEQ ID NO:4), amino acids 5-8 of SEQ ID NO: 1 (SEQ ID NO:5), and amino acids 6-8 of SEQ ID NO: 1 (SEQ ID NO:6).
  • peptide antagonists of the invention include, but are not limited to, peptides that consist of amino acids 1-3 of SEQ ID NO: 1 (SEQ ID NO:11), amino acids 1-4 of SEQ ID NO: 1 (SEQ ID NO: 10), amino acids 1-5 of SEQ ID NO: 1 (SEQ ID NO:9), amino acids 1-6 of SEQ ID NO: 1 (SEQ ID NO:8), amino acids 1-7 of SEQ ID NO: 1 (SEQ ID NO:7), amino acids 2-8 of SEQ ID NO: 1 (SEQ ID NO:2), amino acids 3-8 of SEQ ID NO: 1 (SEQ ID NO:3), amino acids 4-8 of SEQ ID NO: 1 (SEQ ID NO:4), amino acids 5-8 of SEQ ID NO: 1 (SEQ ID NO:5), and amino acids 6-8 of SEQ ID NO: 1 (SEQ ID NO:6).
  • Peptide antagonists of tight junctions of the invention are preferably, 3, 4, 5, 6, or 7 amino acids in length.
  • the peptide antagonists can be chemically synthesized and purified using well- known techniques, such as described in High Performance Liquid Chromatography of Peptides and Proteins: Separation Analysis and Conformation, Eds. Mant et al., C.R.C. Press (1991), and a peptide synthesizer, such as Symphony (Protein Technologies, Inc); or by using recombinant DNA techniques, i.e., where the nucleotide sequence encoding the peptide is inserted in an appropriate expression vector, e.g. , an E. coli or yeast expression vector, expressed in the respective host cell, and purified therefrom using well-known techniques.
  • an appropriate expression vector e.g. , an E. coli or yeast expression vector, expressed in the respective host cell, and purified therefrom using well-known techniques.
  • compositions such as pharmaceutical compositions, comprising a tight junction antagonist (e.g., peptide antagonist) comprise a pharmaceutically effective amount of the antagonist.
  • the pharmaceutically effective amount of antagonist (e.g., peptide antagonist) employed in any given composition may vary according to factors such as the disease state, age, sex, and weight of the individual. Dosage regimens may be adjusted to provide the optimum therapeutic response. For example, a single bolus may be administered, several divided doses may be administered over time or the dose may be proportionally reduced or increased as indicated by the exigencies of the therapeutic situation.
  • the amount of antagonist used for preventing, ameliorating and/or treating a disease in a subject will be in the range of about 1.0 ⁇ g to 1 g, preferably about 1 mg to about 1000 mg, or from about 10 mg to about 100 mg, or from about 10 mg to about 50 mg, or from about 10 mg to about 25 mg of antagonist.
  • Compositions of the invention may comprise one or more tight junction antagonists at a level of from about 0.1 wt% to about 20 wt%, from about 0.1 wt% to about 18 wt%, from about 0.1 wt% to about 16 wt%, from about 0.1 wt% to about 14 wt%, from about 0.1 wt% to about 12 wt%, from about 0.1 wt% to about 10 wt%, from about 0.1 wt% to about 8 wt%, from about 0.1 wt% to about 6 wt%, from about 0.1 wt% to about 4 wt%, from about 0.1 wt% to about 2 wt%, from about 0.1 wt% to about 1 wt%, from about 0.1 wt% to about 0.9 wt%, from about 0.1 wt% to about 0.8 wt%, from about 0.1 wt% to about 0.7 wt%, from about 0.1
  • Compositions of the invention may comprise one or more tight junction antagonists at a level of about 0.1 wt%, about 0.2 wt%, about 0.3 wt%, about 0.4 wt%, about 0.5 wt%, about 0.6 wt%, about 0.7 wt%, about 0.8 wt%, or about 0.9 wt% based on the total weight of the composition.
  • Compositions of the invention may comprise one or more tight junction antagonists at a level of from about 1 wt% to about 20 wt%, from about 1 wt% to about 18 wt%, from about 1 wt% to about 16 wt%, from about 1 wt% to about 14 wt%, from about 1 wt% to about 12 wt%, from about 1 wt% to about 10 wt%, from about 1 wt% to about 9 wt%, from about 1 wt% to about 8 wt%, from about 1 wt% to about 7 wt%, from about 1 wt% to about 6 wt%, from about 1 wt% to about 5 wt%, from about 1 wt% to about 4 wt%, from about 1 wt% to about 3 wt%, or from about 1 wt% to about 2 wt% of the total weight of the composition.
  • Compositions of the invention may comprise one or more tight junction effectors at a level of about 1 wt%, about 2 wt%, about 3 wt%, about 4 wt%, about 5 wt%, about 6 wt%, about 7 wt%, about 8 wt%, or about 9 wt% based on the total weight of the composition.
  • compositions of the invention may be formulated for pulmonary delivery (e.g., may be pulmonary dosage forms).
  • Such compositions may be provided as pharmaceutical aerosols, e.g., solution aerosols or powder aerosols.
  • pharmaceutical aerosols e.g., solution aerosols or powder aerosols.
  • Sciarra and Sciarra, Aerosols in Remington: The Science and Practice of Pharmacy, 20th Ed., Chapter 50, Gennaro et al. Eds., Lippincott, Williams and Wilkins Publishing Co., (2000).
  • compositions of the invention may be formulated for enteric delivery, for example, may comprise one or more coatings, for example, delayed release coating containing one or more enteric agents.
  • a delayed release coating is typically substantially stable in gastric fluid and substantially unstable (e.g., dissolves rapidly or is physically unstable) in intestinal fluid, thus providing for substantial release of the tight junction antagonist from the composition in the duodenum or the jejunum.
  • stable in gastric fluid or “stable in acidic environments” refers to a composition that releases 30% or less by weight of the total tight junction antagonist in the composition in gastric fluid with a pH of 5 or less, or simulated gastric fluid with a pH of 5 or less, in approximately sixty minutes.
  • compositions of the of the invention may release from about 0% to about 30%, from about 0% to about 25%, from about 0% to about 20%, from about 0% to about 15%, from about 0% to about 10%, 5% to about 30%, from about 5% to about 25%, from about 5% to about 20%, from about 5% to about 15%, from about 5% to about 10% by weight of the total tight junction antagonist in the composition in gastric fluid with a pH of 5, or less or simulated gastric fluid with a pH of 5 or less, in approximately sixty minutes.
  • "about” used to modify a numerical value means within 10% of the value.
  • compositions of the invention may release about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10% by weight of the total tight junction antagonist in the composition in gastric fluid with a pH of 5 or less, or simulated gastric fluid with a pH of 5 or less, in approximately sixty minutes.
  • the term “unstable in intestinal fluid” refers to a composition that releases 70% or more by weight of the total tight junction antagonist in the composition in intestinal fluid or simulated intestinal fluid in approximately sixty minutes.
  • the term “unstable in near neutral to alkaline environments” refers to a composition that releases 70% or more by weight of the total amount of tight junction antagonist in the composition in intestinal fluid with a pH of 5 or greater, or simulated intestinal fluid with a pH of 5 or greater, in approximately ninety minutes.
  • a composition that is unstable in near neutral or alkaline environments may release 70% or more by weight of a tight junction antagonist peptide in a fluid having a pH greater than about 5 (e.g., a fluid having a pH of from about 5 to about 14, from about 6 to about 14, from about 7 to about 14, from about 8 to about 14, from about 9 to about 14, from about 10 to about 14, or from about 11 to about 14) in from about 5 minutes to about 90 minutes, or from about 10 minutes to about 90 minutes, or from about 15 minutes to about 90 minutes, or from about 20 minutes to about 90 minutes, or from about 25 minutes to about 90 minutes, or from about 30 minutes to about 90 minutes, or from about 5 minutes to about 60 minutes, or from about 10 minutes to about 60 minutes, or from about 15 minutes to about 60 minutes, or from about 20 minutes to about 60 minutes, or from about 25 minutes to about 90 minutes, or from about 30 minutes to about 60 minutes.
  • compositions of the invention may further comprise one or more therapeutic agents.
  • therapeutic agents include, but are not limited to, steroids and other anti-inflammatory compounds.
  • Suitable therapeutic agents may include one or more of aminosalicylates, corticosteroids, immunomodulators, antibiotics, cytokines, chemokines and biologic therapies.
  • suitable therapeutic agents that maybe included in the compositions of the invention to treat IBD (e.g., Crohn's disease and/or ulcerative colitis) include, but are not limited to:
  • 5-ASA agents e.g., Sulfasalazine
  • Azulfidine® Asacol
  • Dipentum Asacol
  • Pentasa ® and others
  • Antibiotics for example, metronidazole (Flagyl®) and ciprofloxacin (Cipro®), although there are many others that may be effective in certain individuals;
  • Steroids e.g., corticosteroids.
  • Suitable steroids include, but are not limited to, prednisone, hydrocortisone, Medrol®, and budesonide multiple-release capsule MRC (EntocortREC®).
  • 6-mercaptopurine (6-MP, Purinethol®) and azathioprine (Imuran®); and antibodies against inflammatory cytokines, e.g., Infliximab (RemicadeTM).
  • compositions of the invention may also comprise one or more pharmaceutically acceptable excipients.
  • suitable excipients include, but are not limited to, buffers, buffer salts, bulking agents, salts, surface active agents, acids, bases, and binders.
  • compositions of the invention can be used for preventing, slowing the onset of, ameliorating and/or treating any disease associated with an excessive or undesirable permeability of tissues containing tight junctions.
  • diseases of this type include, but are not limited to, celiac disease and IBD (e.g., Crohn's disease and/or ulcerative colitis).
  • the present invention provides a method of treating celiac disease comprising administering to a subject in need thereof a composition comprising a tight junction antagonist.
  • the present invention provides a method of treating Crohn's disease comprising administering to a subject in need thereof a composition comprising a tight junction antagonist
  • the present invention provides a method of treating ulcerative colitis comprising administering to a subject in need thereof a composition comprising a tight junction antagonist.
  • compositions of the invention may be given repeatedly over a protracted period, i.e., may be chronically administered.
  • compositions may be administered one or more times each day in an amount suitable to prevent, reduce the likelihood of an attack of, or reduce the severity of an attack of the underlying disease condition (e.g., celiac disease, IBD etc.).
  • Such compositions may be administered chronically, for example, one or more times daily over a plurality of days.
  • compositions of the invention may be use to treat acute attacks of the underlying disease (e.g., celiac disease, IBD (e.g., Crohn's disease and/or ulcerative colitis)).
  • underlying disease e.g., celiac disease, IBD (e.g., Crohn's disease and/or ulcerative colitis)
  • embodiments of this type will require administration of the compositions of the invention to a subject undergoing an attack in an amount suitable to reduce the severity of the attack.
  • One or more administration may be used.
  • CaCo2 cells form monolayers that exhibit tight junctions between adjacent cells.
  • Treatment of CaCo2 monolayers with peptide FCIGRL enhanced 51 -fold Lucifer Yellow permeability through CaCo2 monolayers compared to vehicle alone.
  • Peptide FCIGRL decreased TEER 16-fold in CaCo2 monolayers compared to vehicle alone.
  • Antagonists of tight junctions can be identified by their ability to prevent or decrease the enhancement of the flux of compounds (e.g. lucifer yellow) through the monolayer induced by agonists of tight junctions (e.g., peptide FCIGRL also known as AT1002).
  • Antagonist of tight junctions can also be identified by their ability to prevent the decrease in TEER induced by agonists of tight junctions (e.g., peptide FCIGRL also known as AT1002).
  • Antagonist Treatment Solution Prepare stock solutions of Antagonist Treatment Solution by dissolving appropriate amounts of antagonist and agonist (for example, a peptide agonist such as a peptide having the sequence FCIGRL) in 7.5mM Lucifer Yellow solution in HBSS. Vortex or sonicate the solution until it is clear then adjust pH to 7.4 ⁇ 0.1 using 1N NaOH.
  • antagonist and agonist for example, a peptide agonist such as a peptide having the sequence FCIGRL
  • Culture Medium DMEM supplemented with 10% fetal bovine serum, 1% NEAA, l% Penn/Strep
  • Buffers Hank's Balanced Salt Solution (HBSS) without calcium and magnesium
  • Flasks 100 X 20 mm Tissue culture dish Falcon.
  • Plates 12 well polycarbonate Transwell filters; 0.3uM pore size
  • the results of these assays are provided in table 1.
  • the first column of the table provides the sequences of the peptides tested, the second column indicates the activity of the peptide in attenuating the Lucifer yellow flux induced by AT 1002, and the third column indicates the activity of the peptide in attenuating the reduction of TEER induced by AT1002.
  • AT 1002 is a 6-mer peptide tight junction agonist having the sequence FCIGRL (SEQ ID NO: 12). See US patent publication US 2005/0059593 Al .
  • Table 1 Tight junction inhibition by SEQ ID NOs: 1-11
  • Gliadin treated with the peptidases pepsin and trypsin induces a cytoskeletal arrangement in CaCo2 cells grown in monolayers.
  • the rearrangement can be visualized using a Nikon-TE2000 epifluorescence microscope and a 40x objective and Alexa Fluor 555 conjugated phalloidin (Invitrogen, Carlsbad, CA), which binds specifically to F-actin.. Exposure times were identical for control and agonist/antagonist treated samples.
  • the figures were generated using Adobe Photoshop CS2 v 9.0.2. The concentration of antagonist was 10 mg/mL.
  • Tight junction antagonists can be identified by their ability to inhibit or decrease the cytoskeletal rearrangement induced by the tight junction agonist AT 1002.
  • Active indicates the peptide protected central actin network (measured by high intensity of fluorescence) compared to control cells treated with agonist alone
  • Inactive indicates the peptide provided no protection of central actin network compared to control treated with agonist alone (low total fluorescence intensity)
  • the columns entitled Blood, Plasma, Gastric Fluid, and Intestinal Fluid indicate if the shorter peptide having the indicated sequence is formed when incubated in the specified fluid.
  • Gastric fluid and intestinal fluid are simulated fluids, fore example, simulated gastric fluid and simulated intestinal fluid include, but are not limited to, those disclosed in the 2005 Pharmacopeia 23NF/28USP in Test Solutions at page 2858 and/or other simulated gastric fluids and simulated intestinal fluids known to those of skill in the art, for example, simulated gastric fluid and/or intestinal fluid prepared without enzymes.
  • the blood and plasma used were of human origin.

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Abstract

La présente invention concerne de nouveaux antagonistes de peptide de jonctions serrées qui inhibent et/ou réduisent l'ouverture de jonctions serrées mammaliennes. Les peptides comprennent de préférence 7 acides aminés ou moins en longueur. La présente invention propose également des procédés pour le traitement d'une perméabilité excessive ou non souhaitable d'un tissu par l'administration à un sujet souffrant d'une telle affection d'une composition comprenant un antagoniste de jonction serrée de peptide de l'invention.
PCT/US2008/065268 2007-05-30 2008-05-30 Nouveaux peptides courts inhibant l'ouverture de jonctions serrées mammaliennes Ceased WO2008150981A2 (fr)

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20200231626A1 (en) * 2008-05-06 2020-07-23 Alba Therapeutics Corporation Inhibition of gliadin peptides
US11608359B2 (en) 2018-02-23 2023-03-21 9 Meters Biopharma, Inc. Compounds and methods for treating tight junction permeabtility

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6458925B1 (en) * 1998-08-03 2002-10-01 University Of Maryland, Baltimore Peptide antagonists of zonulin and methods for use of the same
US20060127320A1 (en) * 2004-05-10 2006-06-15 Nastech Pharmaceutical Company Inc. Method of delivering parathyroid hormone to a human

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20200231626A1 (en) * 2008-05-06 2020-07-23 Alba Therapeutics Corporation Inhibition of gliadin peptides
US11149063B2 (en) 2008-05-06 2021-10-19 Alba Therapeutics Corporation Peptide inhibitors of tight junction permeability
US11608359B2 (en) 2018-02-23 2023-03-21 9 Meters Biopharma, Inc. Compounds and methods for treating tight junction permeabtility

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