WO1994002445A1 - Succedanes non peptidiques de la sequence ldv et leur utilisation dans le traitement d'inflammations, de maladies autoimmunes et du developpement de tumeurs - Google Patents

Succedanes non peptidiques de la sequence ldv et leur utilisation dans le traitement d'inflammations, de maladies autoimmunes et du developpement de tumeurs Download PDF

Info

Publication number
WO1994002445A1
WO1994002445A1 PCT/US1993/007012 US9307012W WO9402445A1 WO 1994002445 A1 WO1994002445 A1 WO 1994002445A1 US 9307012 W US9307012 W US 9307012W WO 9402445 A1 WO9402445 A1 WO 9402445A1
Authority
WO
WIPO (PCT)
Prior art keywords
ldv
cell
compound
surrogates
cells
Prior art date
Application number
PCT/US1993/007012
Other languages
English (en)
Inventor
Ofer Lider
Noam Greenspoon
Rami Hershkoviz
Ronen Alon
Original Assignee
Yeda Research And Development Co. Ltd. At The Weizmann Institute Of Science
Rycus, Avigail
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Yeda Research And Development Co. Ltd. At The Weizmann Institute Of Science, Rycus, Avigail filed Critical Yeda Research And Development Co. Ltd. At The Weizmann Institute Of Science
Priority to JP6504747A priority Critical patent/JPH07509469A/ja
Priority to AU47860/93A priority patent/AU4786093A/en
Priority to EP93918388A priority patent/EP0652863A1/fr
Publication of WO1994002445A1 publication Critical patent/WO1994002445A1/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/50—Cyclic peptides containing at least one abnormal peptide link
    • C07K7/54—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring
    • C07K7/56—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring the cyclisation not occurring through 2,4-diamino-butanoic acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/08—Antiallergic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C237/00—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by amino groups
    • C07C237/02—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by amino groups having the carbon atoms of the carboxamide groups bound to acyclic carbon atoms of the carbon skeleton
    • C07C237/22—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by amino groups having the carbon atoms of the carboxamide groups bound to acyclic carbon atoms of the carbon skeleton having nitrogen atoms of amino groups bound to the carbon skeleton of the acid part, further acylated
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C323/00—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups
    • C07C323/50—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton
    • C07C323/51—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton
    • C07C323/60—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton with the carbon atom of at least one of the carboxyl groups bound to nitrogen atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/02—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing at least one abnormal peptide link
    • C07K5/0202—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing at least one abnormal peptide link containing the structure -NH-X-X-C(=0)-, X being an optionally substituted carbon atom or a heteroatom, e.g. beta-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

  • the present invention relates to novel Leu-Asp-VaL (LDV) surrogates, to their preparation and to pharmaceutical compositions comprising them for treatment of several disorders.
  • LDV Leu-Asp-VaL
  • ECM extracellular matrix
  • the integrins are heterodimeric molecules consisting of an alpha ( ⁇ ) and a beta ( ⁇ ) subunits which are non-covalently linked. Eleven ⁇ and six ⁇ subunits have been identified. The pairing of ⁇ and ⁇ subunits may be cell-type specific and determines ligand specificity.
  • the integrins play an important role in linking the ECM with the intercellular actin-containing cytoskeleton: the extracellular portion mediates the binding of adhesive proteins, and the intracellular portion interacts with elements of the cytoskeleton.
  • the exit of cells from blood vessels and their ensuing tissue localization depends on receptor-mediated recognition of endothelial cells, their basement membranes and the ECM.
  • the ability of various cell types to recognize and attach to components of the ECM may therefore be considered as an essential physiological feature of homeostasis .
  • Fibronectin a well characterized ECM-derived cell-adhesive glycoprotein, is present in a variety of matrices and is synthesized and secreted by a variety of cell types as a 230-250 kDa cross-linked dimer composed of three types of internal repeats: I, II, and III.
  • FN is involved in processes that include wound healing, embryonic cell migration and differentiation, cell activation, proliferation, and adhesion .
  • Cell binding to immobilized FN is mediated primarily by surface integrins of the ⁇ 1 (CD29; very late antigens (VLA))-subfamily of receptors, including VLA-3, -4 and -5 .
  • LDV Leu-Asp-Val sequence
  • VLA-4 ou ⁇ 1 integrin
  • This domain was found to be recognized by integrin receptors of various cell types, such as those on tumor cells and lymphocytes (Guan et al, 1990; Shimizu et al, 1990; Postigo et al, 1991; Roldan et al, 1992).
  • VLA-4 integrin was also implicated in the recognition of vascular cell adhesion molecule-1 (VCAM-1) on endothelial cells, although this binding site was distinct from that recognized on FN (Elices et al,1990).
  • Inhibitors of VLA-4 recognition of FN may thus interfere with cell migration from the circulation to inflammatory sites. Indeed specific interference with VLA-4 activity, was found to inhibit experimental autoimmune encephalomyelitis as well as the elicitation of immune-cell mediated contact sensitivity reaction in mice.
  • the RGD surrogate effectively inhibited the elicitation of a CD4 + T cell-mediated inflammatory reaction in mice, indicating that the proteolytically stable RGD-mimetics may serve as useful therapeutic agents in a variety of pathologic processes which depend on RGD recognition .
  • LDV surrogates are effective inhibitors of cellular or molecular interactions which depend on LDV-sequence recognition. These LDV analogues are herein referred to as "LDV surrogates”.
  • the present invention thus relates to compounds of the general formula I
  • X 1 to X 5 are the same or different C, N, O or S atoms,at least two of them being C, and the X 1 - X 5 chain may be optionally substituted by radicals selected from halogen, hydrocarbyl, oxo, thioxo, amino and carboxyl, or the X 1 -X 5 chain or part thereof may form part of a heterocyclic ring, and pharmaceutically acceptable salts thereof.
  • X 1 , X 2 and X 4 represent carbon atoms
  • X 5 is NH and X 3 is NH or S
  • the X 1 -X 5 chain is substituted by one or two oxo groups.
  • X 1 , X 3 and X 4 are carbon atoms
  • X 2 is S and X 5 is NH.
  • the invention further relates to methods for the preparation of the LDV surrogates of the invention.
  • the LDV surrogates of the invention have various applications related to their inhibition of biological interactions dependent on LDV-sequence recognition, such as integrin-mediated cell functions.
  • the invention also relates to pharmaceutical compositions comprising the LDV surrogates for the treatment of several disorders, such as inflammatory disorders, and disorders involving interference with cell-matrix or cell-cell dependent immune processes, such as autoimmune diseases, allergy, graft-versus-host and related reactions, and inhibition of metastasis and tumor progression.
  • Fig.1 shows specificity of inhibition of T cell adhesion to fibronectin (FN) and laminin (LN) by the following inhibitory compounds: peptides GRGDSP AND EILDVPST, the LDV surrogates AC-16 and EG-45, and the comparison compounds AC-22 and BL-34.
  • Fig.2 shows analysis of inhibition of T cell adhesion to FN by the inhibitory compounds as in Fig.1, at concentrations of 800, 400 and 200 ⁇ g/ml.
  • Fig.3 shows dose-dependent inhibition of T cell adhesion to FN by the EILDVPST peptide (white symbols) and the corresponding EG-45 surrogate (black symbols).
  • the peptide amino and carboxyl end groups are not involved in recognition by its respective integrin and therefore do not affect its inhibitory activity, as is the case for RGD. It was also further considered that the major contribution of the LDV-containing peptides for binding to their putative integrin-recognition sites on integrins depends on the interaction of the carboxylate moiety of Asp (aspartic acid) inside an hydrophobic pocket in which the side chains of Leu are Val are bound.
  • this tripeptide is most probably dominated by the S-trans configuration of the two peptide bonds, and therefore the compounds prepared according to the invention have preferably two hydrophobic side chains and a carboxylate moiety in the same relative orientations they have in the LDV peptide.
  • LDV surrogates refers to novel compounds of the general formula I herein, and pharmaceutically acceptable salts thereof.
  • hydrocarbyl as employed herein includes straight or branched C 1 -C 12 alkyl and C 2 -C 12 alkenyl or alkynyl radicals, such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, pentyl, vinyl, allyl and ethynyl; C 3 -C 7 cycloalkyl and cycloalkenyl radicals, such as cyclopropyl, cyclopentyl, cyclohexyl and cyclohexenyl; and C 6 -C 14 aryl radicals, such as phenyl and naphthyl.
  • heterocyclic rings comprising the X 1 -X 5 chain or part thereof include saturated or unsaturated rings, optionally containing additional O, S and/or N atoms, and may be optionally substituted by halogen, hydrocarbyl, oxo, thioxo, amino or carboxyl.
  • An example is the compound herein designated NG-93, which contains a 13-membered ring.
  • one or more of X 1 to X 5 are NH and one or more are C atoms substituted by oxo groups, such as the compound herein designated AC-16 in which X 3 and X 5 are -NH, X 1 is >CH 2 and ⁇ 2 and X 4 are carbonyl groups, of the formula depicted in Scheme 1 herein.
  • one or more of X 1 to X 5 are -NH, one is O or S and one or more are C atoms substituted by oxo groups, such as the compound herein designated LB-1, in which X 3 is S, X 5 is -NH, X 1 and X 2 are >CH 2 and X 4 is carbonyl, and the compound herein designated EG-45, in which X 2 is S, X 5 is -NH, X 1 and X 3 are >CH 2 and X 4 is carbonyl (Scheme 1).
  • the atoms of the X 1 -X 5 chain together with the C atom adjacent to X 5 form part of a heterocyclic ring, such as the compound herein designated NG-93, in which X 3 and X 5 are -NH, X 2 and X 4 are carbonyl and they form part of a 13-membered heterocyclic ring (Scheme 1).
  • the compounds of the invention may be prepared by several procedures.
  • LDV analogues wherein X 4 is carbonyl and X 3 is nitrogen e.g. compound AC-16
  • protected L-Asp e.g.,t-butyloxycarbonyl Asp (O-benzyl) wherein amide bonds (i.e., X 5 is -NH and X 2 is carbonyl) are prepared by coupling with carbodiimide as shown in Scheme 2.
  • Ester bonds (X 5 is oxygen) and thioesters are also prepared by carbodiimide coupling.
  • LEV analogues, such as the compound AC-22 are prepared according to the same synthetic scheme using as the starting material protected L-Glu, e.g. t-butyloxycarbonyl Glu-(O-benzyl).
  • Cyclic peptides e.g. NG-93 are prepared by stepwise synthesis on an oxime resin starting from Val attached to the resin and coupling of Asp followed by Leu. The peptide is then cleaved from the resin by an excess of t-Boc ⁇ -amino acid. Removal of the t-butyl protecting groups followed by cyclization (DCC,l-hydroxybenzotriazole) affords the O-benzyl protected cyclic peptide which is purified by HPLC. Hydrogenolysis of the O-benzyl group (Pd/C 10%) at atmospheric pressure of H 2 gives the cyclic compound.
  • DCC t-butyl protecting groups followed by cyclization
  • LDV analogues wherein X 3 is sulfur and X 5 is nitrogen, oxygen or sulfur e.g. compound LB-1
  • X 3 is sulfur and X 5 is nitrogen, oxygen or sulfur
  • compound LB-1 are prepared by a Michael addition of 4-methyl-1-pentanethiol to benzyl maleate. The addition takes place preferentially ⁇ - to the ester group. Coupling of the free carboxyl group with isobutyl amine, or alcohol or thiol (dicyclohexyl-carbodiimide, 1-hydroxy-benzotriazole) affords the benzyl-protected analogue. The benzyl group is removed by catalytic hydrogenation and the product, e.g. compound LB-1, is purified, e.g. by recrystallysation.
  • the compound EG-45 (Scheme 1) is a LDV peptide surrogate that, in addition to lacking the two terminal groups, also has a peptide bond replaced by a peptide bond surrogate.
  • Pseudo-S-CH 2 was chosen since its successful synthesis provides easy access to a new peptide bond replacement for peptides containing an Asp moiety next to the N-terminus. It is expected to be resistant to proteases and its lipophilic character can offer advantages.
  • the synthesis of EG-45 was based on the Michael addition of a thiol to a bis-ester of itaconic acid, by the synthetic path outlined in Scheme 3 herein.
  • compound BL-34 (Scheme 1) was prepared in a way similar to that used for compound EG-45, with sec-butylamine used for the amide formation in the last step of the synthesis, and was used as a control in the cell adhesion studies.
  • LDV analogues wherein X 2 and X 4 are carbonyl and X 1 and X 5 are oxygen are prepared by Michael addition of malonic acid to diisobutyl maleate followed by decarboxylation.
  • X 1 and X 5 are nitrogen, Michael addition of malonic acid to maleic acid diisobutyl amide produces the desired compounds.
  • Pharmaceutically acceptable salts of the LDV surrogates of the invention include but are not limited to inorganic salts, such as sodium, potassium, calcium and the like, and organic salts with amines or organic bases, such as piperidine, morpholine and the like.
  • the LDV surrogates of the invention can inhibit biological interactions which are dependent on LDV recognition.
  • Examples of versatile recognition processes mediated by the LDV pattern encompassed by the present invention include cellular and molecular interactions involving the LDV sequence by the integrin VLA-4. Thus, they can also prevent metastasis.
  • the surrogates inhibit lymphocyte interaction with certain antigen-presenting cells and thus inhibit T cell activation and migration, thereby preventing autoimmune diseases.
  • the compounds of the invention can be administered to patients by any suitable route including oral and parenteral routes, e.g., intravenous, subcutaneous or intramuscular injection.
  • An effective but essentially non-toxic quantity of the compound is employed in the treatment. Effective amounts may be within the range of 0.01 to 1 mg/kg, preferably 0.5 mg/kg on a regimen in single or more divided daily doses.
  • the invention further provides a pharmaceutical composition
  • a pharmaceutical composition comprising as active ingredient a surrogate according to the invention and a pharmaceutically acceptable carrier.
  • the compositions may be in the form of tablet, capsule, solution or suspension containing from about 0.7 to 70 mg per unit of dosage of an active compound of the invention or mixtures thereof.
  • the compounds may be compounded in conventional manner with a physiologically acceptable vehicle or carrier, excipient, binder, preservative, stabilizer, etc.
  • injections for intravenous administration may be prepared in saline, at a pH level of e.g. 7.4.
  • N,N'-Dicyclohexylcarbodiimide (226 mg, 1.1 mmol) was added to a solution containing t-butyloxycarbonyl Asp (O-benzyl) (309 mg, 1.0 mmol) and 1-hydroxybenzotriazole (148 mg, 1.1 mmol) in CH 2 Cl 2 / THF 1:1 (v/v) solution (7 ml). The reaction was left overnight at room temperature. The solution was filtered to remove dicyclohexylurea (DCU) generated in the reaction, and washed with dry CH 2 CI 2 . Isobutyl amine (87 mg, 1.2 mmol) was added and the solution was stirred for 5 h at room temperature.
  • DCU dicyclohexylurea
  • the solution was poured into aqueous 0.1 HCl solution (10 ml) and the product was extracted twice with chloroform (50 ml). The chloroform was dried over Na 2 SO 4 and was removed under reduced pressure.
  • the crude coupling product was dissolved in CH 2 CI 2 (4 ml) and was cooled to 0°C, trifluoroacetic acid (4 ml) was added and the reaction mixture was allowed to stand 30 min at 0°C and 1 h at room temperature. The solvent and the acid were removed under reduced pressure, thus producing N-tert-butyloxycarbonyl-4-benzyloxy-1-aspartic acid isobutylamide.
  • the crude product was pure enough for the next step.
  • N,N'-Dicyclohexylcarbodiimide (226, mg, 1.1.mmol) was added to a solution containing isocaproic acid (135 mg, 1.0 mmol) and 1-hydroxybenzotriazole (148 mg, 1.1 mmol) in CH 2 CI 2 / THF 1:1 (v/v) solution (7 ml). The reaction was left overnight at room temperature. Dicyclohexyurea (DCU) was filtered off and washed with dry CH 2 CI 2 . The crude product from the previous step was added to that solution along with diisopropyl ethyl amine (260 mg, 2 mmol). The reaction mixture was stirred for 5 h at room temperature.
  • DCC N,N'-Dicyclohexylcarbodiimide
  • the LEV surrogate AC-22 was prepared by the same procedure as compound AC-16, but using t-butyloxycarbonyl Glu (O-benzyl) instead of the Asp derivative, and was used for comparison with the LDV surrogate compound AC-16.
  • Example 4 Preparation of 8-methyl-3-( 2-methylpropyl- amino- carbonyl)-5-thianonanoic acid (Compound EG-45)
  • the synthesis of compound EG-45 was carried out as depicted in Scheme 3 herein.Starting from itaconic acid 1, a monoallyl ester at the ⁇ -position 2 is formed under acidic conditions. This was verified by 13 C-NMR which indicates that of the two carbonyl signals of itaconic acid at 166.89 and 171.47 ppm, corresponding to the unsaturated and saturated carbonyls, respectively, only the 171.47 ppm resonance was shifted upfield to 169.58 ppm upon esterification.
  • Compound BL-34 used for comparison with EG-45, was prepared by the same procedure as compound EG-45 in Example 4, but using sec-butylamine instead of the isobutylamine. The resulting mixture of diastereomers was used without further separation.
  • N- ⁇ -tBoc-Valine was coupled to an oxime resin (DeGrado, W.F. and Kaiser,E.T., 1980, J. Org. Chem. 45:1295) using DCC as the coupling reagent.
  • oxime resin (2.00g) was added to a solution containing N- ⁇ -tBoc.-valine (1 mmol) and DCC (1 mmol) in methylene chloride (10 ml). The mixture was shaken for 15 h, washed with methylene chloride, DMF, isopropanol and methylene chloride and dried in vacuo.
  • VLA-4 integrin plays a major role in cellular interactions with immobilized FN. Indeed, this integrin was shown to participate in various processes, such as lympho-hemopoiesis, bone marrow cell differentiation, VCAM-1 recognition, cell activation, and the binding of FN to several cell types, including tumor cells and lymphocytes (Elices et al,1990; Shimizu et al,1990; Roldan et al, 1992).
  • the minimal recognition site on FN for the VLA-4 integrin appears to be restricted to the alternatively spliced V region and is blocked specifically by the LDV-containing peptides (Humphries et al, 1986).
  • CD4 + T cells were purified from the peripheral blood of healthy human donors. Mononuclear leukocytes were isolated using Ficoll gradients, washed and incubated in RPMI supplemented with 10% FCS and antibiotics in petri dishes at 37°C humidified CO 2 incubator. After 2 h, the non-adherent cells were isolated and applied on nylon-wool columns (1.5 h). The CD4 + T cells were then negatively selected by exposure of these cells to a mixture of anti-CD8, CD19, and CD14 monoclonal antibodies (mAb) conjugated to magnetic-beads (Advanced Magnetics, MA). Cells that did not bind to the beads were exposed to a second round of negative selection. The resulting cell population, which consisted of over 90% CD3 + CD4 + cells, as determined by FACScan analysis (not shown), was used as the source of human CD4 + T cells.
  • mAb monoclonal antibodies
  • ECM components were first bound to polystirene wells.
  • FN or laminin (LN) (Sigma; 1 mg/50 ml medium/well) was added to 96-well flat-bottom microtiter plates (Costar) and incubated for 2 hours. Unbound protein was removed by washing and the remaining binding sites on the plates were blocked by 1% BSA (Sigma) in PBS.
  • the T cells were radioactively labeled with 51 [Cr] (New England Nuclear) and activated with phorbol myristate acetate (PMA; Sigma; 10ng/ml).
  • these T cells were pretreated (30 min at 37°C) with anti-CD29 (anti- ⁇ 1 chain of VLA integrins; diluted 1/200, Serotec, Oxford, UK), anti-VLA-5, anti-VLA-4, or anti-VLA-6 (diluted 1/400; Telios Pharmaceuticals Research Inc.) mAb.
  • anti-CD29 anti- ⁇ 1 chain of VLA integrins; diluted 1/200, Serotec, Oxford, UK
  • anti-VLA-5 anti-VLA-4
  • anti-VLA-6 diluted 1/400; Telios Pharmaceuticals Research Inc.
  • the RGD peptide GRGDSPK was purchased from Sigma.
  • the LDV peptide EILDVPST was synthesized by an Applied Biosystems synthesizer at the Peptide Synthesis Unit, The Weizmann Institute of Science, Rehovot, Israel.
  • the 51 [Cr]-labeled CD4-T cells (2 ⁇ 10 5 /well) were then added to the coated microtiter plates and incubated for 30 min at 37°C in a 10% CO 2 humidified incubator. The non-adherent cells were then gently removed and the adherent cells were lyzed and collected. The percent of cells that were adhered was calculated as follows: [CPM of residual cells in the well/ (total CPM of cells added to the well-spontaneous release of 51 Cr)x 100]. Results are expressed as the mean percent of T cell binding derived from quadriplicate wells for each experimental group.
  • anti-CD29 mAb specific to the common ⁇ 1 -chain of VLA integrins abolished T cell adhesion to both adhesive glycoproteins.
  • VLA-5 (as) 18 ⁇ 4 53 ⁇ 3
  • the inhibitory effect of the LDV surrogates on cell adhesion to FN was measured in a way similar to that described for peptide inhibition.
  • the freshly isolated and purified human CD4 + T cells were pretreated with the inhibitory compounds (peptides GRGDSP and EILDVPST, the LDV surrogates AC-16 and EG-45, and comparison compounds AC-22 and BL-34) at a fixed concentration of 600 ⁇ g/ml.
  • the cells were then seeded in the FN or LN precoated wells. Percent cell adhesion was calculated thereafter. Values ⁇ SD are shown in Fig. 1 (one experiment representative of 3).
  • Fig. 1 The results shown in Fig. 1 indicate that surrogates AC-16 and EG-45 inhibited T cell adhesion to FN.
  • the AC-16 mimetic is almost as active as the LDV-containing peptide, while AC-22 is inactive, indicating that the amino and carboxyl terminal groups of the tripeptide LDV are not required for its activity.
  • EG-45 was a better inhibitor of T cell adhesion than both AC-16 and the LDV-containing peptide (54, 34, and 46%, respectively).
  • Compounds AC-22 and BL-34 did not inhibit T cell adhesion to neither ECM-glycoproteins.
  • AC-22 derived from glutamic acid
  • AC-22 can be regarded as an LEV surrogate, and its inability to inhibit cell adhesion reflects the situation observed with the LEV-NH 2 peptide (Humphries et al, 1986).
  • LEV-NH 2 peptide Humphries et al, 1986.
  • compound BL-34 a terminal methyl group from the "amide side” was "shifted" to a position closer to the nitrogen, presumably introducing steric hindrance and interfering with hydrophobic group binding to its receptor pocket. This probably accounts for the poor inhibitory potential of this compound on CD4-T cell adhesion to FN.
  • mice with a synthetic EILDVPST peptide inhibited the induction of contact sensitivity, a T cell mediated immune reaction in the dermal micro-environment of mice (Ferguson et al, 1991).
  • LDV surrogates may indeed serve as inhibitors of inflammatory reactions.
  • the integrin binding site of the LDV sequence is composed of two hydrophobic pockets that surround a binding site for the Asp-carboxylate group.
  • the Asp residue has been implicated to be involved in many integrin-binding ligands.
  • Our results suggest that the carboxyl and amino groups at the C and N terminals of the peptide make only a minor contribution to the inhibitory effect of LDV containing peptides. Accordingly, a replacement of at least one peptide bond does not reduce the affinity for the receptor. This means that the peptide backbone as well makes no direct contribution to the affinity of the integrin for the LDV ligand.
  • the backbone may serve to position the hydrophobic and hydrophilic groups in the appropriate orientation for integrin-recognition and binding.
  • DTH delayed-type hypersensitivity

Landscapes

  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Medicinal Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Veterinary Medicine (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Pharmacology & Pharmacy (AREA)
  • General Chemical & Material Sciences (AREA)
  • Public Health (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Engineering & Computer Science (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Genetics & Genomics (AREA)
  • Molecular Biology (AREA)
  • Immunology (AREA)
  • Pulmonology (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Pain & Pain Management (AREA)
  • Rheumatology (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Peptides Or Proteins (AREA)

Abstract

Succédanés non peptidiques de Leu-Asp-Val (LDV) de la formule (I) dans laquelle X1 à X5 représentent des atomes de C, N, O ou S identiques ou différents, au moins deux d'entre eux représentant C, et la chaîne X1-X5 peut éventuellement être substituée par des radicaux choisis parmi halogène, hydrocarbyle, oxo, thioxo, amino et carboxyle, ou la chaîne X1-X5 ou une partie de celle-ci peut faire partie d'un cycle hétérocyclique. Ces succédanés, ainsi que leurs sels pharmaceutiquemt acceptables, inhibent les interactions cellulaires ou moléculaires dépendantes de la recconnaissance de la séquence LDV, et peuvent être utilisés dans le traitement d'inflammations et de troubles causés par l'interférence avec des processus immunitaires cellule-cellule ou cellule-matrice, tels que des malaides autoimmunes et des allergies, et dans l'inhibition du développement des tumeurs et des métastases.
PCT/US1993/007012 1992-07-26 1993-07-26 Succedanes non peptidiques de la sequence ldv et leur utilisation dans le traitement d'inflammations, de maladies autoimmunes et du developpement de tumeurs WO1994002445A1 (fr)

Priority Applications (3)

Application Number Priority Date Filing Date Title
JP6504747A JPH07509469A (ja) 1992-07-26 1993-07-26 Ｌｄｖ配列の非ペプチド性代替物
AU47860/93A AU4786093A (en) 1992-07-26 1993-07-26 Non-peptidic surrogates of the ldv sequence and their use in the treatment of inflammation, autoimmune diseases and tumour progression
EP93918388A EP0652863A1 (fr) 1992-07-26 1993-07-26 Succedanes non peptidiques de la sequence ldv et leur utilisation dans le traitement d'inflammations, de maladies autoimmunes et du developpement de tumeurs

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
IL10264692A IL102646A (en) 1992-07-26 1992-07-26 Non-peptidic surrogates of the ldv sequence and pharmaceutical compositions comprising them
IL102646 1992-07-26

Publications (1)

Publication Number Publication Date
WO1994002445A1 true WO1994002445A1 (fr) 1994-02-03

Family

ID=11063866

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US1993/007012 WO1994002445A1 (fr) 1992-07-26 1993-07-26 Succedanes non peptidiques de la sequence ldv et leur utilisation dans le traitement d'inflammations, de maladies autoimmunes et du developpement de tumeurs

Country Status (6)

Country Link
EP (1) EP0652863A1 (fr)
JP (1) JPH07509469A (fr)
AU (1) AU4786093A (fr)
CA (1) CA2140931A1 (fr)
IL (1) IL102646A (fr)
WO (1) WO1994002445A1 (fr)

Cited By (19)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996020216A1 (fr) * 1994-12-24 1996-07-04 Zeneca Limited Inhibiteurs d'adhesion de la fibronectine
ES2133042A1 (es) * 1996-03-14 1999-08-16 Univ Valencia Politecnica Sintesis de zeolita y zeotipos isomorfos a la zeolita beta.
US6034057A (en) * 1995-07-06 2000-03-07 Zeneca Limited Peptide inhibitors of fibronectine
US6232522B1 (en) 1990-01-31 2001-05-15 Oklahoma Medical Research Foundation Non-human animal model for systemic lupus erythematosis
US6235711B1 (en) 1996-06-21 2001-05-22 Zeneca Limited Cell adhesion ihibiting compounds
US6239108B1 (en) 1996-07-11 2001-05-29 Biogen, Inc. Cell adhesion inhibitors
US6248713B1 (en) 1995-07-11 2001-06-19 Biogen, Inc. Cell adhesion inhibitors
US6306840B1 (en) 1995-01-23 2001-10-23 Biogen, Inc. Cell adhesion inhibitors
US6495525B1 (en) 1998-05-28 2002-12-17 Biogen, Inc. VLA-4 inhibitor: oMePUPA-V
US6552216B1 (en) 1996-07-25 2003-04-22 Biogen, Inc. Molecular model for VLA-4 inhibitors
US6630503B1 (en) 1999-08-13 2003-10-07 Biogen, Inc. Cell adhesion inhibitors
US6686350B1 (en) 1996-07-25 2004-02-03 Biogen, Inc. Cell adhesion inhibitors
US6875743B1 (en) 2000-11-28 2005-04-05 Biogen, Inc. Cell adhesion inhibitors
US7001921B1 (en) 1995-01-23 2006-02-21 Biogen Idec Ma Inc. Cell adhesion inhibitors
US7192715B2 (en) 1993-11-30 2007-03-20 Oklahoma Medical Research Foundation Diagnostics and therapy of epstein-barr virus in autoimmune disorders
US7196112B2 (en) 2004-07-16 2007-03-27 Biogen Idec Ma Inc. Cell adhesion inhibitors
US7273613B1 (en) 1997-01-13 2007-09-25 The Board of Regents, The University of Oklahoma Diagnostics and therapy of Epstein-Barr virus in autoimmune disorders
US7276341B2 (en) 1990-01-31 2007-10-02 Oklahoma Medical Research Foundation Methods and reagents for diagnosis of autoantibodies
US8067467B2 (en) * 2005-05-18 2011-11-29 Biogen Idec International Gmbh Thiosuccinic acid derivatives and the use thereof

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1991003252A1 (fr) * 1989-09-01 1991-03-21 Fred Hutchinson Cancer Research Center Inhibition de l'adhesion de lymphocytes sur l'endothelium vasculaire au moyen d'une nouvelle interaction entre le recepteur matriciel extra-cellulaire et son ligand

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1991003252A1 (fr) * 1989-09-01 1991-03-21 Fred Hutchinson Cancer Research Center Inhibition de l'adhesion de lymphocytes sur l'endothelium vasculaire au moyen d'une nouvelle interaction entre le recepteur matriciel extra-cellulaire et son ligand

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
KOMORIYA A ET AL: "The minimal essential sequence for a major cell type-specific adhesion site (CS1) within the alternatively spliced type III connecting segment domain of fibronectin is leucine-aspartic acid-valine", JOURNAL OF BIOLOGICAL CHEMISTRY, vol. 266, no. 23, 15 August 1991 (1991-08-15), BALTIMORE, MD US, pages 15075 - 15079 *

Cited By (26)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6232522B1 (en) 1990-01-31 2001-05-15 Oklahoma Medical Research Foundation Non-human animal model for systemic lupus erythematosis
US7276341B2 (en) 1990-01-31 2007-10-02 Oklahoma Medical Research Foundation Methods and reagents for diagnosis of autoantibodies
US7192715B2 (en) 1993-11-30 2007-03-20 Oklahoma Medical Research Foundation Diagnostics and therapy of epstein-barr virus in autoimmune disorders
WO1996020216A1 (fr) * 1994-12-24 1996-07-04 Zeneca Limited Inhibiteurs d'adhesion de la fibronectine
US6034056A (en) * 1994-12-24 2000-03-07 Zeneca Limited Fibronectin adhesion inhibitors
US6630512B2 (en) 1995-01-23 2003-10-07 Biogen, Inc. Cell adhesion inhibitors
US6306840B1 (en) 1995-01-23 2001-10-23 Biogen, Inc. Cell adhesion inhibitors
US6376538B1 (en) 1995-01-23 2002-04-23 Biogen, Inc. Cell adhesion inhibitors
US6624152B2 (en) 1995-01-23 2003-09-23 Biogen, Inc. Cell adhesion inhibitors
US7001921B1 (en) 1995-01-23 2006-02-21 Biogen Idec Ma Inc. Cell adhesion inhibitors
US6034057A (en) * 1995-07-06 2000-03-07 Zeneca Limited Peptide inhibitors of fibronectine
US6248713B1 (en) 1995-07-11 2001-06-19 Biogen, Inc. Cell adhesion inhibitors
US6596687B1 (en) 1995-07-11 2003-07-22 Biogen, Inc. Cell adhesion inhibitors
ES2133042A1 (es) * 1996-03-14 1999-08-16 Univ Valencia Politecnica Sintesis de zeolita y zeotipos isomorfos a la zeolita beta.
US6235711B1 (en) 1996-06-21 2001-05-22 Zeneca Limited Cell adhesion ihibiting compounds
US6239108B1 (en) 1996-07-11 2001-05-29 Biogen, Inc. Cell adhesion inhibitors
US6686350B1 (en) 1996-07-25 2004-02-03 Biogen, Inc. Cell adhesion inhibitors
US6949534B2 (en) 1996-07-25 2005-09-27 Biogen Idec Ma Inc. Cell adhesion inhibitors
US6552216B1 (en) 1996-07-25 2003-04-22 Biogen, Inc. Molecular model for VLA-4 inhibitors
US7273613B1 (en) 1997-01-13 2007-09-25 The Board of Regents, The University of Oklahoma Diagnostics and therapy of Epstein-Barr virus in autoimmune disorders
US6495525B1 (en) 1998-05-28 2002-12-17 Biogen, Inc. VLA-4 inhibitor: oMePUPA-V
US6630503B1 (en) 1999-08-13 2003-10-07 Biogen, Inc. Cell adhesion inhibitors
US7034043B2 (en) 1999-08-13 2006-04-25 Biogen Idec Ma Inc. Cell adhesion inhibitors
US6875743B1 (en) 2000-11-28 2005-04-05 Biogen, Inc. Cell adhesion inhibitors
US7196112B2 (en) 2004-07-16 2007-03-27 Biogen Idec Ma Inc. Cell adhesion inhibitors
US8067467B2 (en) * 2005-05-18 2011-11-29 Biogen Idec International Gmbh Thiosuccinic acid derivatives and the use thereof

Also Published As

Publication number Publication date
EP0652863A1 (fr) 1995-05-17
CA2140931A1 (fr) 1994-01-27
JPH07509469A (ja) 1995-10-19
AU4786093A (en) 1994-02-14
IL102646A (en) 1996-05-14
IL102646A0 (en) 1993-01-14

Similar Documents

Publication Publication Date Title
WO1994002445A1 (fr) Succedanes non peptidiques de la sequence ldv et leur utilisation dans le traitement d'inflammations, de maladies autoimmunes et du developpement de tumeurs
JP3248584B2 (ja) フィブロネクチン接着抑制剤
EP0617705B1 (fr) Succedanes non peptidiques de la sequence arg-gly-asp, et compositions pharmaceutiques les contenant
CA2393682C (fr) Depsipeptide et ses congeneres destines a etre utilises en tant qu'immunodepresseurs
US5710129A (en) Inhibitors of SH2-mediated processes
US5190922A (en) Terminally modified tri-, tetra- and pentapeptide anaphylatoxin receptor ligands
BG65755B1 (bg) Инхибитори на клетъчна адхезия
CZ359997A3 (cs) Pětičlenné heterocyklické sloučeniny jako inhibitory adheze leukocytů a antagonisté VLA-4 a farmaceutický přípravek, který je obsahuje
KR20010083065A (ko) α4β1 매개 세포 접착성 억제제
JPH05504762A (ja) 抗血栓性ペプチド及び偽ペプチド
CZ23299A3 (cs) Inhibitory adheze buněk a farmaceutický prostředek, který je obsahuje
JPH07179414A (ja) サイトカイン生成を誘発するウレタン類およびウレア類
BG63876B1 (bg) Инхибитори на клетъчна адхезия
AU8888598A (en) Novel cyclic tetrapeptide derivatives and pharmaceutical use thereof
JP2010209088A (ja) Ｐ−セレクチンに対する結合性化合物
CA3018344A1 (fr) Agonistes du recepteur du neuropeptide s (npsr)
AU8614898A (en) Novel 5-membered ring heterocycles, their preparation, their use and pharmaceutical preparations comprising them
NZ242441A (en) Alpha-substituted peptides and their therapeutic use
KR20150065718A (ko) 인돌린
RU2134695C1 (ru) Антитромботические азациклоалкилалканоилпептиды
JPH0649718B2 (ja) 多環状窒素含有構造を有する新規なペプチド誘導体
US7767645B2 (en) SH2 domain binding inhibitors
GREENSPOON et al. Novel ψ‐S‐CH2 peptide‐bond replacement and its utilization in the synthesis of nonpeptidic surrogates of the Leu‐Asp‐Val sequence that exhibit specific inhibitory activities on CD4+ T cell binding to fibronectin
KR20050026543A (ko) 항-인테그린 활성을 갖는 플루오로알킬사이클로펩티드유도체
IL100130A (en) Guanidine derivatives and pharmaceutical compositions comprising them

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AT AU BB BG BR BY CA CH CZ DE DK ES FI GB HU JP KP KR KZ LK LU MG MN MW NL NO NZ PL PT RO RU SD SE SK UA US VN

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): AT BE CH DE DK ES FR GB GR IE IT LU MC NL PT SE BF BJ CF CG CI CM GA GN ML MR NE SN TD TG

ENP Entry into the national phase

Ref document number: 1994 373216

Country of ref document: US

Date of ref document: 19940424

Kind code of ref document: A

DFPE Request for preliminary examination filed prior to expiration of 19th month from priority date (pct application filed before 20040101)
121 Ep: the epo has been informed by wipo that ep was designated in this application
WWE Wipo information: entry into national phase

Ref document number: 1993918388

Country of ref document: EP

WWE Wipo information: entry into national phase

Ref document number: 2140931

Country of ref document: CA

WWP Wipo information: published in national office

Ref document number: 1993918388

Country of ref document: EP

REG Reference to national code

Ref country code: DE

Ref legal event code: 8642

WWW Wipo information: withdrawn in national office

Ref document number: 1993918388

Country of ref document: EP