EP1427755A2 - Use of peptides comprising post-translational modifications in the treatment of autoimmune pathologies - Google Patents

Use of peptides comprising post-translational modifications in the treatment of autoimmune pathologies

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Publication number
EP1427755A2
EP1427755A2 EP02798771A EP02798771A EP1427755A2 EP 1427755 A2 EP1427755 A2 EP 1427755A2 EP 02798771 A EP02798771 A EP 02798771A EP 02798771 A EP02798771 A EP 02798771A EP 1427755 A2 EP1427755 A2 EP 1427755A2
Authority
EP
European Patent Office
Prior art keywords
lysine
sequence seq
serine
acetylated
phosphorylated
Prior art date
Legal status (The legal status 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 status listed.)
Ceased
Application number
EP02798771A
Other languages
German (de)
French (fr)
Inventor
Sylviane Muller
Fanny Sylvie Michèle MONNEAUX
Jean-Paul Briand
Gilles Guichard
Jean-Gérard Guillet
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Centre National de la Recherche Scientifique CNRS
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Centre National de la Recherche Scientifique CNRS
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Priority to EP10183969A priority Critical patent/EP2338908A1/en
Publication of EP1427755A2 publication Critical patent/EP1427755A2/en
Ceased legal-status Critical Current

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/04—Drugs for skeletal disorders for non-specific disorders of the connective tissue
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • 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
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A61P31/20—Antivirals for DNA viruses
    • A61P31/22—Antivirals for DNA viruses for herpes viruses
    • 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
    • A61P37/02—Immunomodulators
    • 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
    • A61P37/02—Immunomodulators
    • A61P37/06—Immunosuppressants, e.g. drugs for graft rejection
    • 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 subject of the present invention is new peptides transformed so as to include post-translational type modifications, such as phosphorylation or acetylation of one or more amino acids.
  • the invention also relates to their production methods, and to their uses in pharmaceutical compositions in the context of
  • the identification of the sequences of the 70K protein recognized by the self-reactive T cells began in 1998.
  • the inventors first synthesized 20 overlapping peptides covering the sequence of the 70K protein and studied the recognition of these peptides by the antibodies of MRL / lpr lupus mice as well as the self-reactive CD4 + T cells of these mice.
  • these 20 peptides they identified the peptide corresponding to the sequence 131-151 (RIHMVYSKRSGKPRGYAFIEY) of the protein 70K recognized very early by the antibodies of these mice (Monneaux et al, 2000).
  • This peptide is capable of stimulating in vitro the proliferation and the secretion of IL-2 of CD4 + T cells purified from the lymph nodes.
  • this sequence also represents a major epitope of the 70K protein in another lupus mouse model, the NZB / W mouse (FIG.
  • the peptide 131-151 is moreover capable of binding to various class II molecules of the major histocompatibility complex (both murine and human; FIG. 2).
  • This universal character constitutes an advantage for the use of this sequence in the development of therapeutic strategies in the context of human systemic lupus erythematosus.
  • the inventors then wished to determine the exact nature of the sequence of the 70K protein capable of activating the self-reactive T cells.
  • the inventors synthesized several peptides corresponding to the phosphorylated and acetylated forms on the serine and lysine residues of peptide 131-151, and studied the capacity of these peptides to be recognized by the T lymphocytes and the antibodies of lupus mice.
  • the present invention follows from the demonstration by the Inventors that these phosphorylated and acetylated peptides are recognized in an equal or even superior manner to the parent peptide which is not phosphorylated and not acetylated by CD4 + T cells and the antibodies of lupus mice, and that DNA administration of these phosphorylated and acetylated peptides decreases the production of high levels of antibodies directed against DNA, delays the onset of glomerulonephritis and prolongs the survival of animals, while the parent peptide does not, on the other hand, induce any statistically significant effect .
  • the object of the present invention is to provide new peptides which can be used for the preparation of medicaments in the context of the treatment of autoimmune diseases, and more particularly lupus, having the advantage of being significantly more effective than the peptides used up to now.
  • the modified peptides of the invention are specific for deleterious cells and target only these cells, unlike immunosuppressants, cytokines, or other molecules currently used which act globally on the immune system.
  • the subject of the invention is the use of peptides comprising epitopes of mammalian self proteins recognized by antibodies produced by the immune system of mammals suffering from autoimmune pathologies, and, where appropriate, by T cells. auxiliaries of said mammals, said epitopes being such that at least one of their amino acids comprises a modification of the post-translational type, for the preparation of a medicament intended for the prevention or treatment of said autoimmune pathologies.
  • post-translational type modification is meant in what precedes and what follows, any type of modification of the amino acids of a given protein capable of occurring in vivo in the cells of the organism, such as the phosphorylation processes. , acetylation, or other.
  • a more particular subject of the invention is the aforementioned use of peptides as defined above, comprising epitopes of which at least one of their amino acids is modified so that it is in a phosphorylated form, or acetylated.
  • the invention relates more particularly to the aforementioned use of peptides comprising epitopes derived from proteins of human or animal origin defined above, said proteins being chosen from nucleoproteins, proteins from the nucleosome, from the spliceosome, from the ribonucleoprotein particle Ro , or ribosome for example.
  • the invention also relates to the use of peptides as defined above, for the preparation of a medicament intended for prevention or treatment:
  • non-specific systemic organ diseases such as systemic lupus erythematosus (LED), rheumatoid arthritis, mixed connectivity, Sjôgren syndrome, or chronic juvenile arthritis .
  • autoimmune organ pathologies such as multiple sclerosis, insulin-dependent diabetes, Crohn's disease, or oily diseases.
  • a more particular subject of the invention is the use of peptides as defined above, for the preparation of a medicament intended for the prevention or treatment of
  • the invention relates more particularly to the aforementioned use of peptides comprising epitopes derived from the human or murine protein U1-70K of the spliceosome (described in particular in Klein Gunnewiek et al, 1997).
  • a more particular subject of the invention is also the above-mentioned use of peptides comprising the sequence delimited by amino acids 131 and 151 of the human or murine protein U1-70K, and corresponding to the following sequence SEQ ID NO: 1: RIHMVYSKRSGKPRGYAFIEY in which at least one of the amino acids comprises a modification of the post-translational type, in particular in which at least one of the amino acids is phosphorylated, or acetylated.
  • the invention relates more particularly to the aforementioned use of peptides comprising the sequence SEQ ID NO: 1 in which at least one of the serine residues in position 7 or 10 is phosphorylated, and / or at least one of the lysine residues in position 8 or 12 is acetylated. :
  • a more particular subject of the invention is the aforementioned use of peptides comprising the sequence SEQ ID NO: 1 in which:
  • the invention more particularly still relates to the aforementioned use of peptides chosen from the following:
  • a more particular subject of the invention is also the above-mentioned use of peptides chosen from the following:
  • sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated
  • the invention also relates to any pharmaceutical composition characterized in that it comprises at least one peptide chosen from those defined above, in combination with a pharmaceutically acceptable vehicle.
  • the invention more particularly relates to any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from those comprising the sequence SEQ ID NO: 1 in which at least one of the amino acids involves a modification of the post-translational type, in particular by phosphorylation, or acetylation.
  • the invention relates more particularly to any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from those comprising the sequence SEQ ID NO: 1 in which at least one of the serine residues in position 7 or 10 is phosphorylated, and / or at least one of the lysine residues in position 8 or
  • a more particular subject of the invention is any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from those comprising the sequence SEQ ID NO: 1 in which:
  • the serine in position 7 is phosphorylated, and / or the serine in position 10 is phosphorylated,
  • the invention more particularly still relates to any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from the following:
  • the invention more particularly relates to any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from the following: - the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
  • the abovementioned pharmaceutical compositions of the invention are characterized in that they are in a form which can be administered by the systemic route (namely by the intravenous, intramuscular, intraperitoneal, subcutaneous route), or non-invasive (for example intranasal route) , oral, or epicutaneous).
  • the abovementioned pharmaceutical compositions of the invention are characterized in that the daily doses of peptides in humans are from approximately 100 ng to approximately 5 mg.
  • the invention also relates to the peptides comprising the sequence delimited by amino acids 131 and 151 of the human U1-70K protein, or murine, and corresponding to the following sequence SEQ ID NO: 1: RIHMVYSKRSGKPRGYAFIEY in which one at least of the amino acid is phosphorylated, or acetylated.
  • a more particular subject of the invention is the above-mentioned peptides, comprising the sequence SEQ ID NO: 1 in which at least one of the serine residues in position 7 or 10 is phosphorylated, and / or one at least of the lysine residues in position 8 or 12 is acetylated.
  • the invention relates more particularly to the above-mentioned peptides, comprising the sequence SEQ ID NO: 1 in which:
  • the invention relates more particularly still to the following peptides: the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
  • sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated
  • the phosphorylated peptides PI 37 (corresponding to the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated), and P140 (corresponding to the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated), and the acetylated peptides Acl38 (corresponding to the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated), Acl42 (corresponding to the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated), and Acl38 + 142 (corresponding to the sequence SEQ ID NO: 1 in which the lysine at position 8 and that at position 12 are acetylated), as well as the scrambled peptide Se: YVSRYFGSAIRHEPKMKIYRG, and the scrambled peptide ScP corresponding to Se in which the serine in position 8 is phosphorylated, (used as negative controls in the tests which follow) corresponding respectively to the
  • the protein 70K being strongly phosphorylated in vivo (Woppmann et al, 1993), and although the number of phosphorylated sites and their identity are not known, the inventors have synthesized several peptides corresponding to the phosphorylated and acetylated forms respectively on the serine and lysine residues of peptide 131-151, and studied the capacity of these peptides to be recognized by T lymphocytes and antibodies from lupus mice. The inventors demonstrate in the context of the present invention that the phosphorylated peptide in position 140 is recognized in an equal or even superior manner to the non-phosphorylated peptide by CD4 + T cells and the antibodies of lupus mice (FIG. 13).
  • the two peptides (phosphorylated at position 140 and not phosphorylated) were used for a study of restoration of self-tolerance. These two peptides were injected intravenously and intranasally into pre-auto mice, and the course of the disease in these mice was monitored.
  • the 3 acetylated peptides are at least as well or even better recognized than the parent peptide by the CD4 + T cells of normal mice immunized against the unmodified peptide: the proliferation rates are higher, the IL2 secretion rates are equivalent and the interferon ⁇ production rates are higher,
  • the 3 acetylated peptides are at least as well or even better recognized than the parent peptide by the CD4 + T cells of autoimmune mice: the proliferation rates are higher,
  • the 3 acetylated peptides are recognized by the antibodies of mice directed against the parent peptide.
  • the inventors have demonstrated that the peptides 131-151 and PI 40 are capable of binding various human MHC class II molecules (HLA-DR1, -DR4 and -DRU) (Figure 19).
  • CD4 + T lymphocytes (A) from MRL / lpr lupus mice; the graph on the left represents the proliferation of CD4 + T cells expressed in stimulation indices as defined above in the presence of the peptide 131-151, phosphorylated peptides PI 37 and PI 40 and two scrambled peptides phosphorylated or not (Se and ScP); the positivity limit corresponds to 2.0; the graph on the right represents the secretion of IL-2 (mU / ml) in the presence of peptide 131-151, peptides P137 and P140 and peptides Se and ScP.
  • A CD4 + T lymphocytes

Abstract

The invention concerns novel peptides transformed so as to comprise post-translational modifications, such as phosphorylation or acetylation of one or several amino acids. The invention also concerns methods for obtaining them, and their uses in pharmaceutical compositions for treating autoimmune pathologies.

Description

UTILISATION DE PEPTIDES COMPORTANT DES MODIFICATIONS DE TYPE POST-TRADUCTIONNEL DANS LE CADRE DU TRAITEMENT DE PATHOLOGLES AUTO-IMMUNES USE OF PEPTIDES COMPRISING POST-TRANSLATIONAL TYPE MODIFICATIONS IN THE TREATMENT OF AUTOIMMUNE PATHOLOGIES
La présente invention a pour objet de nouveaux peptides transformés de manière à comporter des modifications de type post-traductionnel, telles que phosphorylation ou acetylation d'un ou plusieurs acides aminés. L'invention concerne également leurs procédés d'obtention, et leurs utilisations dans des compositions pharmaceutiques dans le cadre duThe subject of the present invention is new peptides transformed so as to include post-translational type modifications, such as phosphorylation or acetylation of one or more amino acids. The invention also relates to their production methods, and to their uses in pharmaceutical compositions in the context of
10 traitement des pathologies auto-irnmunes.10 treatment of autoimmune pathologies.
Plusieurs études ont démontré l'intérêt de l'utilisation de peptides synthétiques pour la prévention dans des modèles murins du développement de pathologies auto-immunes telles que le lupus. Ces études ont été réalisées avec des peptides dérivés soit de séquences d'histones, soit d'anticorps anti-ADN. L'administration par voie intra-veineuse de cesSeveral studies have demonstrated the interest of using synthetic peptides for prevention in mouse models of the development of autoimmune pathologies such as lupus. These studies were carried out with peptides derived either from histone sequences or from anti-DNA antibodies. Intravenous administration of these
15 peptides a permis de diminuer la production d'anticorps anti-ADN typiques du lupus et de prolonger la survie des souris traitées. Aucune étude n'a été réalisée à partir de séquences de protéines du splicéosome, autre autoantigène du lupus.15 peptides made it possible to decrease the production of anti-DNA antibodies typical of lupus and to prolong the survival of the treated mice. No study has been carried out using protein sequences from the spliceosome, another autoantigen of lupus.
Les quelques études réalisées à ce jour ayant démontré une amélioration de la pathologie lupique chez des souris autoimmunes ont utilisé des peptides ne contenant pas deThe few studies carried out to date which have demonstrated an improvement in lupus pathology in autoimmune mice have used peptides containing no
?.o modifications post-traductionelles (Eilat et al. , 2000; Jouanne et al., 1999; Kaliyaperumal et al. , 1999; Marino et al, 2000).? .o post-translational modifications (Eilat et al., 2000; Jouanne et al., 1999; Kaliyaperumal et al., 1999; Marino et al, 2000).
Les modifications post-traductionnelles semblent jouer un rôle important dans l'émergence de la réponse auto-irnmune (Utz and Anderson, 1998). Afin de mettre en place des stratégies d'intervention efficaces, l'identification des cibles réellement responsables de laPost-translational modifications seem to play an important role in the emergence of the autoimmune response (Utz and Anderson, 1998). In order to set up effective intervention strategies, the identification of the targets really responsible for
25 rupture de tolérance au soi et reconnues ensuite par les cellules autoréactives constitue un intérêt majeur.25 breach of self-tolerance and then recognized by self-reactive cells is of major interest.
L'identification des séquences de la protéine 70K reconnues par les cellules T autoréactives a débuté en 1998. Les inventeurs ont dans un premier temps synthétisé 20 peptides chevauchants couvrant la séquence de la protéine 70K et étudié la reconnaissance de 0 ces peptides par les anticorps de souris lupiques MRL/lpr ainsi que par les cellules T CD4 + autoréactives de ces souris. Parmi ces 20 peptides, ils ont identifié le peptide correspondant à la séquence 131-151 (RIHMVYSKRSGKPRGYAFIEY) de la protéine 70K reconnu très précocement par les anticorps de ces souris (Monneaux et al, 2000). Ce peptide est capable de stimuler in vitro la prolifération et la sécrétion d'IL-2 des cellules T CD4+ purifiées à partir des ganglions.The identification of the sequences of the 70K protein recognized by the self-reactive T cells began in 1998. The inventors first synthesized 20 overlapping peptides covering the sequence of the 70K protein and studied the recognition of these peptides by the antibodies of MRL / lpr lupus mice as well as the self-reactive CD4 + T cells of these mice. Among these 20 peptides, they identified the peptide corresponding to the sequence 131-151 (RIHMVYSKRSGKPRGYAFIEY) of the protein 70K recognized very early by the antibodies of these mice (Monneaux et al, 2000). This peptide is capable of stimulating in vitro the proliferation and the secretion of IL-2 of CD4 + T cells purified from the lymph nodes.
Par la suite, les Inventeurs ont montré que cette séquence représentait aussi un épitope majeur de la protéine 70K dans un autre modèle de souris lupique, la souris NZB/W (figureSubsequently, the inventors have shown that this sequence also represents a major epitope of the 70K protein in another lupus mouse model, the NZB / W mouse (FIG.
1). Le peptide 131-151 est par ailleurs capable de se lier à diverses molécules de classe II du complexe majeur d'histocompatibilité (à la fois murines, et humaines; figure 2). Ce caractère universel constitue un avantage pour l'utilisation de cette séquence dans le développement de stratégies thérapeutiques dans le cadre du lupus érythémateux disséminé humain. Les Inventeurs ont ensuite souhaité déterminer la nature exacte de la séquence de la protéine 70K capable d'activer les cellules T autoréactives. Les Inventeurs ont synthétisé plusieurs peptides correspondant aux formes phosphorylées et acétylées sur les résidus serine et lysine du peptide 131-151, et étudié la capacité de ces peptides à être reconnus par les lymphocytes T et les anticorps de souris lupiques. La présente invention découle de la mise en évidence par les Inventeurs que ces peptides phosphorylés et acétylés sont reconnus de façon égale voire supérieure au peptide parent non phosphorylé et non acétylé par les cellules T CD4+ et les anticorps de souris lupiques, et que l'adn inistration de ces peptides phosphorylés et acétylés diminue la production de forts taux d'anticorps dirigés contre l'ADN, retarde l'apparition de la glomérulonéphrite et prolonge la survie des animaux, tandis que le peptide parent n'induit par contre aucun effet statistiquement significatif.1). The peptide 131-151 is moreover capable of binding to various class II molecules of the major histocompatibility complex (both murine and human; FIG. 2). This universal character constitutes an advantage for the use of this sequence in the development of therapeutic strategies in the context of human systemic lupus erythematosus. The inventors then wished to determine the exact nature of the sequence of the 70K protein capable of activating the self-reactive T cells. The inventors synthesized several peptides corresponding to the phosphorylated and acetylated forms on the serine and lysine residues of peptide 131-151, and studied the capacity of these peptides to be recognized by the T lymphocytes and the antibodies of lupus mice. The present invention follows from the demonstration by the Inventors that these phosphorylated and acetylated peptides are recognized in an equal or even superior manner to the parent peptide which is not phosphorylated and not acetylated by CD4 + T cells and the antibodies of lupus mice, and that DNA administration of these phosphorylated and acetylated peptides decreases the production of high levels of antibodies directed against DNA, delays the onset of glomerulonephritis and prolongs the survival of animals, while the parent peptide does not, on the other hand, induce any statistically significant effect .
La présente invention a pour but de fournir de nouveaux peptides utilisables pour la préparation de médicaments dans le cadre du traitement de maladies auto-immunes, et plus particulièrement du lupus, présentant l'avantage d'être nettement plus efficaces que les peptides utilisés jusqu'à présent, et de ne pas présenter d'effets secondaires importants tels que ceux rencontrés avec les techniques actuelles de traitement, dans la mesure où les peptides modifiés de l'invention sont spécifiques des cellules délétères et ne ciblent que ces cellules, à la différence des immunosuppresseurs, cytokines, ou autres molécules utilisées actuellement qui agissent de manière globale sur le système immunitaire. L'invention a pour objet l'utilisation de peptides comprenant des épitopes de protéines du soi des mammifères reconnus par des anticorps produits par le système immunitaire des mammifères atteints de pathologies auto-immunes, et, le cas échéant, par les cellules T auxiliaires desdits mammifères, lesdits épitopes étant tels que l'un au moins de leurs acides aminés comporte une modification de type post-traductionnel, pour la préparation d'un médicament destiné à la prévention ou au traitement desdites pathologies auto-immunes.The object of the present invention is to provide new peptides which can be used for the preparation of medicaments in the context of the treatment of autoimmune diseases, and more particularly lupus, having the advantage of being significantly more effective than the peptides used up to now. now, and not to have significant side effects such as those encountered with current treatment techniques, insofar as the modified peptides of the invention are specific for deleterious cells and target only these cells, unlike immunosuppressants, cytokines, or other molecules currently used which act globally on the immune system. The subject of the invention is the use of peptides comprising epitopes of mammalian self proteins recognized by antibodies produced by the immune system of mammals suffering from autoimmune pathologies, and, where appropriate, by T cells. auxiliaries of said mammals, said epitopes being such that at least one of their amino acids comprises a modification of the post-translational type, for the preparation of a medicament intended for the prevention or treatment of said autoimmune pathologies.
Par modification de type post-traductionnel, on entend dans ce qui précède et ce qui suit, tout type de modification des aminoacides d'une protéine donnée susceptible de se produire in vivo dans les cellules de l'organisme, tels que les processus de phosphorylation, d' acetylation, ou autre.By post-translational type modification is meant in what precedes and what follows, any type of modification of the amino acids of a given protein capable of occurring in vivo in the cells of the organism, such as the phosphorylation processes. , acetylation, or other.
L'invention a plus particulièrement pour objet l'utilisation susmentionnée de peptides tels que définis ci-dessus, comprenant des épitopes dont l'un au moins de leurs acides aminés est modifié de manière à ce qu'il soit sous une forme phosphorylée, ou acétylée.A more particular subject of the invention is the aforementioned use of peptides as defined above, comprising epitopes of which at least one of their amino acids is modified so that it is in a phosphorylated form, or acetylated.
L'invention concerne plus particulièrement l'utilisation susmentionnée de peptides comprenant des épitopes issus des protéines d'origine humaine ou animale définies ci- dessus, lesdites protéines étant choisies parmi les nucléoprotéines, les protéines du nucléosome, du splicéosome, de la particule ribonucléoprotéique Ro, ou du ribosome par exemple.The invention relates more particularly to the aforementioned use of peptides comprising epitopes derived from proteins of human or animal origin defined above, said proteins being chosen from nucleoproteins, proteins from the nucleosome, from the spliceosome, from the ribonucleoprotein particle Ro , or ribosome for example.
L'invention concerne également l'utilisation de peptides tels que définis ci-dessus, pour la préparation d'un médicament destiné à la prévention ou au traitement :The invention also relates to the use of peptides as defined above, for the preparation of a medicament intended for prevention or treatment:
- de pathologies auto-immunes de la famille des connectivités (maladies systémiques non spécifiques d'organes), telles que le lupus érythémateux disséminé (LED), la polyarthrite rhumatoïde, la connectivité mixte, le syndrome de Sjôgren, ou l'arthrite chronique juvénile,- autoimmune pathologies of the family of connectivities (non-specific systemic organ diseases), such as systemic lupus erythematosus (LED), rheumatoid arthritis, mixed connectivity, Sjôgren syndrome, or chronic juvenile arthritis ,
- ou de pathologies auto-immunes spécifiques d'organes, telles que la sclérose en plaques, le diabète insulino-dépendant, la maladie de Crohn, ou les maladies huileuses.- or specific autoimmune organ pathologies, such as multiple sclerosis, insulin-dependent diabetes, Crohn's disease, or oily diseases.
L'invention a plus particulièrement pour objet l'utilisation de peptides tels que définis ci-dessus, pour la préparation d'un médicament destiné à la prévention ou au traitement duA more particular subject of the invention is the use of peptides as defined above, for the preparation of a medicament intended for the prevention or treatment of
LED.LED.
A ce titre, l'invention concerne plus particulièrement l'utilisation susmentionnée de peptides comprenant des épitopes issus de la protéine humaine ou murine U1-70K du splicéosome (décrite notamment dans Klein Gunnewiek et al, 1997). L'invention a plus particulièrement pour objet encore l'utilisation susmentionnée de peptides comprenant la séquence délimitée par les acides aminés 131 et 151 de la protéine U1-70K humaine ou murine, et correspondant à la séquence SEQ ID NO : 1 suivante : RIHMVYSKRSGKPRGYAFIEY dans laquelle l'un au moins des acides aminés comporte une modification de type post- traductionnel, notamment dans laquelle l'un au moins des acides aminés est phosphorylé, ou acétylé. L'invention concerne plus particulièrement l'utilisation susmentionnée de peptides comprenant la séquence SEQ ID NO : 1 dans laquelle l'un au moins des résidus serine en position 7 ou 10 est phosphorylé, et/ou l'un au moins des résidus lysine en position 8 ou 12 est acétylé. : As such, the invention relates more particularly to the aforementioned use of peptides comprising epitopes derived from the human or murine protein U1-70K of the spliceosome (described in particular in Klein Gunnewiek et al, 1997). A more particular subject of the invention is also the above-mentioned use of peptides comprising the sequence delimited by amino acids 131 and 151 of the human or murine protein U1-70K, and corresponding to the following sequence SEQ ID NO: 1: RIHMVYSKRSGKPRGYAFIEY in which at least one of the amino acids comprises a modification of the post-translational type, in particular in which at least one of the amino acids is phosphorylated, or acetylated. The invention relates more particularly to the aforementioned use of peptides comprising the sequence SEQ ID NO: 1 in which at least one of the serine residues in position 7 or 10 is phosphorylated, and / or at least one of the lysine residues in position 8 or 12 is acetylated. :
L'invention a plus particulièrement pour objet l'utilisation susmentionnée de peptides comprenant la séquence SEQ ID NO : 1 dans laquelle :A more particular subject of the invention is the aforementioned use of peptides comprising the sequence SEQ ID NO: 1 in which:
- la serine en position 7 est phosphorylée,- the serine in position 7 is phosphorylated,
- et/ou la serine en position 10 est phosphorylée,- And / or the serine in position 10 is phosphorylated,
- et/ou la lysine en position 8 est acétylée,- And / or the lysine in position 8 is acetylated,
- et/ou la lysine en position 12 est acétylée. L'invention concerne plus particulièrement encore l'utilisation susmentionnée de peptides choisis parmi les suivants :- And / or the lysine in position 12 is acetylated. The invention more particularly still relates to the aforementioned use of peptides chosen from the following:
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, - the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 is acetylated
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 12 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 12 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 7 thus that the serine in position 10 are phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 12 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée, ,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées.- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated, - the sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
L'invention a plus particulièrement pour objet encore l'utilisation susmentionnée de peptides choisis parmi les suivants :A more particular subject of the invention is also the above-mentioned use of peptides chosen from the following:
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées. L'invention a également pour objet toute composition pharmaceutique caractérisée en ce qu'elle comprend au moins un peptide choisi parmi ceux définis ci-dessus, en association avec un véhicule pharmaceutiquement acceptable.- The sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated. The invention also relates to any pharmaceutical composition characterized in that it comprises at least one peptide chosen from those defined above, in combination with a pharmaceutically acceptable vehicle.
L'invention a plus particulièrement pour objet toute composition pharmaceutique telle que définie ci-dessus, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi ceux comprenant la séquence SEQ ID NO : 1 dans laquelle l'un au moins des acides aminés comporte une modification de type post-traductionnel, notamment par phosphorylation, ou acetylation.The invention more particularly relates to any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from those comprising the sequence SEQ ID NO: 1 in which at least one of the amino acids involves a modification of the post-translational type, in particular by phosphorylation, or acetylation.
L'invention concerne plus particulièrement toute composition pharmaceutique telle que définie ci-dessus, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi ceux comprenant la séquence SEQ ID NO : 1 dans laquelle l'un au moins des résidus serine en position 7 ou 10 est phosphorylé, et/ou l'un au moins des résidus lysine en position 8 ouThe invention relates more particularly to any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from those comprising the sequence SEQ ID NO: 1 in which at least one of the serine residues in position 7 or 10 is phosphorylated, and / or at least one of the lysine residues in position 8 or
12 est acétylé. L'invention a plus particulièrement pour objet toute composition pharmaceutique telle que définie ci-dessus, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi ceux comprenant la séquence SEQ ID NO : 1 dans laquelle :12 is acetylated. A more particular subject of the invention is any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from those comprising the sequence SEQ ID NO: 1 in which:
- la serine en position 7 est phosphorylée, - et/ou la serine en position 10 est phosphorylée,the serine in position 7 is phosphorylated, and / or the serine in position 10 is phosphorylated,
- et/ou la lysine en position 8 est acétylée,- And / or the lysine in position 8 is acetylated,
- et/ou la lysine en position 12 est acétylée.- And / or the lysine in position 12 is acetylated.
L'invention concerne plus particulièrement encore toute composition pharmaceutique telle que définie ci-dessus, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi les suivants :The invention more particularly still relates to any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from the following:
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 12 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 7 thus that the serine at position 10 is phosphorylated, and the lysine at position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 12 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 ainsi que la lysine en position- the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 as well as the lysine in position
12 sont acétylées,12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,- the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 12 is acetylated, the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées.- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated, - the sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
L'invention a plus particulièrement pour objet encore toute composition pharmaceutique telle que définie ci-dessus, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi les suivants : - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée,The invention more particularly relates to any pharmaceutical composition as defined above, characterized in that it comprises at least one peptide chosen from the following: - the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées.- The sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
Avantageusement, les compositions pharmaceutiques susmentionnées de l'invention, sont caractérisées en ce qu'elles se présentent sous une forme administrable par voie systémique (à savoir par voie intraveineuse, intramusculaire, intrapéritonéale, sous- cutanée), ou non invasive (par exemple intranasale, orale, ou épicutanée). Avantageusement encore, les compositions pharmaceutiques susmentionnées de l'invention, sont caractérisées en ce que les doses journalières de peptides chez l'homme sont d'environ 100 ng à environ 5 mg.Advantageously, the abovementioned pharmaceutical compositions of the invention are characterized in that they are in a form which can be administered by the systemic route (namely by the intravenous, intramuscular, intraperitoneal, subcutaneous route), or non-invasive (for example intranasal route) , oral, or epicutaneous). Advantageously also, the abovementioned pharmaceutical compositions of the invention are characterized in that the daily doses of peptides in humans are from approximately 100 ng to approximately 5 mg.
L'invention concerne également les peptides comprenant la séquence délimitée par les acides aminés 131 et 151 de la protéine U1-70K humaine, ou murine, et correspondant à la séquence SEQ ID NO : 1 suivante : RIHMVYSKRSGKPRGYAFIEY dans laquelle l'un au moins des acides aminés est phosphorylé, ou acétylé.The invention also relates to the peptides comprising the sequence delimited by amino acids 131 and 151 of the human U1-70K protein, or murine, and corresponding to the following sequence SEQ ID NO: 1: RIHMVYSKRSGKPRGYAFIEY in which one at least of the amino acid is phosphorylated, or acetylated.
L'invention a plus particulièrement pour objet les peptides susmentionnés, comprenant la séquence SEQ ID NO : 1 dans laquelle l'un au moins des résidus serine en position 7 ou 10 est phosphorylé, et/ou l'un au moins des résidus lysine en position 8 ou 12 est acétylé. L'invention concerne plus particulièrement les peptides susmentionnés, comprenant la séquence SEQ ID NO : 1 dans laquelle :A more particular subject of the invention is the above-mentioned peptides, comprising the sequence SEQ ID NO: 1 in which at least one of the serine residues in position 7 or 10 is phosphorylated, and / or one at least of the lysine residues in position 8 or 12 is acetylated. The invention relates more particularly to the above-mentioned peptides, comprising the sequence SEQ ID NO: 1 in which:
- la serine en position 7 est phosphorylée, - et/ou la serine en position 10 est phosphorylée,- the serine in position 7 is phosphorylated, - And / or the serine in position 10 is phosphorylated,
- et/ou la lysine en position 8 est acétylée,- And / or the lysine in position 8 is acetylated,
- et/ou la lysine en position 12 est acétylée.- And / or the lysine in position 12 is acetylated.
L'invention concerne plus particulièrement encore les peptides suivants : - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée,The invention relates more particularly still to the following peptides: the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 12 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 12 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées. L'invention a plus particulièrement pour objet encore les peptides suivants :- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated, - the sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated. A more particular subject of the invention is also the following peptides:
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée,the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated, the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées.- The sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
L'invention sera davantage illustrée à l'aide de la description détaillée qui suit de la, synthèse des peptides modifiés de l'invention, ainsi que de l'étude de leurs propriétés biologiques.The invention is further illustrated by the following detailed description of, synthesis of modified peptides of the invention, as well as the study of their biological properties.
I) SynthèsesI) Summaries
Les peptides phosphorylés PI 37 (correspondant à la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée), et P140 (correspondant à la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée), et les peptides acétylés Acl38 (correspondant à la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée), Acl42 (correspondant à la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée), et Acl38 + 142 (correspondant à la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et celle en position 12 sont acétylées), ainsi que le peptide brouillé Se : YVSRYFGSAIRHEPKMKIYRG, et le peptide brouillé ScP correspondant à Se dans lequel la serine en position 8 est phosphorylée, (utilisés comme contrôles négatifs dans les tests qui suivent) correspondant respectivement à la séquence SEQ ID NO : 1 et à la séquence de P140 dans lesquelles les acides aminés sont dans un ordre différent et aléatoire, ont été synthétisés chimiquement en phase solide sur un synthétiseur automatique en utilisant la stratégie Fmoc (N-(9-fluorenyl)methoxycarbonyl). Afin d'introduire les résidus de serine phosphorylée à la place des résidus de serine ou les résidus de lysine acétylée à la place des résidus de lysine, un dérivé de la serine de type Fmoc- Ser(PO(Obz)OH)-OH, ou un dérivé de lysine de type Fmoc-Lys(Ac), ont été utilisés. Le temps de couplage est augmenté à 30 minutes et un second couplage est systématiquement effectué. Après le clivage en milieu acide, chaque peptide est précipité par de l'éther froid, solubilisé dans une solution d'eau et d'acétonitrile et enfin lyophilisé. Les peptides sont ensuite purifiés par RP-HPLC, leur intégrité et leur pureté ont été analysées par HPLC analytique et par spectrométrie de masse (Maldi-TOF).The phosphorylated peptides PI 37 (corresponding to the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated), and P140 (corresponding to the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated), and the acetylated peptides Acl38 (corresponding to the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated), Acl42 (corresponding to the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated), and Acl38 + 142 (corresponding to the sequence SEQ ID NO: 1 in which the lysine at position 8 and that at position 12 are acetylated), as well as the scrambled peptide Se: YVSRYFGSAIRHEPKMKIYRG, and the scrambled peptide ScP corresponding to Se in which the serine in position 8 is phosphorylated, (used as negative controls in the tests which follow) corresponding respectively to the sequence SEQ ID NO: 1 and to the sequence of P140 in which the amino acids are in different order ferent and random, were chemically synthesized in solid phase on an automatic synthesizer using the Fmoc (N- (9-fluorenyl) methoxycarbonyl) strategy. In order to introduce phosphorylated serine residues in place of serine residues or acetylated lysine residues in place of lysine residues, a serine derivative of the Fmoc-Ser type (PO (Obz) OH) -OH, or a lysine derivative of type Fmoc-Lys (Ac), were used. The coupling time is increased to 30 minutes and a second coupling is systematically carried out. After cleavage in an acid medium, each peptide is precipitated with cold ether, dissolved in a solution of water and acetonitrile and finally lyophilized. The peptides are then purified by RP-HPLC, their integrity and purity were analyzed by analytical HPLC and by mass spectrometry (Maldi-TOF).
Pureté Masse attendue Masse mesuréePurity Expected mass Measured mass
P137 71.1 % 2639 2637.04P137 71.1% 2639 2637.04
P140 90.2% 2639 2637.03P140 90.2% 2639 2637.03
Acl38 88.8% 2600 2602.3Acl38 88.8% 2600 2602.3
Acl42 83.4% 2600 2600.3Acl42 83.4% 2600 2600.3
Acl38 + 142 85.1 % 2643 2644.68Acl38 + 142 85.1% 2,643 2,644.68
Se 96.5% 2558 2559.56Se 96.5% 2558 2559.56
ScP 97.2% 2637 2637.06ScP 97.2% 2637 2637.06
Les profils HPLC des peptides P137, P140, Acl38, Acl42, Acl38 + 142, Se et ScP sont représentés respectivement sur les figures 3, 4, 5, 6, 7, 15 et 16 (matériel utilisé : colonne Nucifosil C IB 150x4.6 mm; débit : 1.2 ml/mn; détection UV : 210 nm; gradient utilisé : 5-65 en 20 min en eau + TFA 0,1% et acétonitrile + TFA 0,08%).The HPLC profiles of the peptides P137, P140, Acl38, Acl42, Acl38 + 142, Se and ScP are shown respectively in Figures 3, 4, 5, 6, 7, 15 and 16 (material used: column Nucifosil C IB 150x4.6 mm; flow rate: 1.2 ml / min; UV detection: 210 nm; gradient used: 5-65 in 20 min in water + 0.1% TFA and acetonitrile + 0.08% TFA).
Les résultats sont indiqués dans les tableaux A, B, C, D, E, F et G ci-après correspondant respectivement aux profils HPLC des peptides P137, P140, Ac 138, Ac 142, Ac 138 + 142, Se et ScP.The results are indicated in Tables A, B, C, D, E, F and G below corresponding respectively to the HPLC profiles of the peptides P137, P140, Ac 138, Ac 142, Ac 138 + 142, Se and ScP.
Tableau ATable A
nombre de pics temps de rétention zone de pic pourcentage de zonenumber of peaks retention time peak area area percentage
1 12.16 164.7 71.1 2 12.39 60.5 26.11 12.16 164.7 71.1 2 12.39 60.5 26.1
3 12.63 6.6 2.33 12.63 6.6 2.3
Total 231.8 100.0 Tableau BTotal 231.8 100.0 Table B
nombre de pics temps de rétention zone de pic pourcentage de zone 1 11.93 8.9 3.1 2 12.13 254.4 90.2 3 12.45 11.5 4.1 4 12.72 7.3 2.6 Total 292.1 100.0number of peaks retention times peak area area percentage 1 11.93 8.9 3.1 2 12.13 254.4 90.2 3 12.45 11.5 4.1 4 12.72 7.3 2.6 Total 292.1 100.0
Tableau CTable C
nombre de pics temps de rétention zone de pic pourcentage de zone 1 12.67 171.6 88.8 2 12.83 21.7 11.2 Total 193.3 100.0number of peaks retention time peak area area percentage 1 12.67 171.6 88.8 2 12.83 21.7 11.2 Total 193.3 100.0
Tableau DTable D
nombre de pics temps de rétention zone de pic pourcentage de zone 1 11.39 1.9 0.8 2 12.75 203.1 83.4 3 12.92 34.8 14.2 4 13.22 3.8 1.6 Total 243.6 100.0number of peaks retention time peak area area percentage 1 11.39 1.9 0.8 2 12.75 203.1 83.4 3 12.92 34.8 14.2 4 13.22 3.8 1.6 Total 243.6 100.0
Tableau ETable E
nombre de pics temps de rétention zone de pic pourcentage de zonenumber of peaks retention time peak area area percentage
1 13.03 213.3 85.11 13.03 213.3 85.1
2 13.22 15.2 6.12 13.22 15.2 6.1
3 13.37 22.1 8.83 13.37 22.1 8.8
Total 250.6 100.0 Tableau FTotal 250.6 100.0 Table F
nombre de pics temps de rétention zone de pic pourcentage de zonenumber of peaks retention time peak area area percentage
1 12.08 219.3 96.5 2 12.67 2.5 1.11 12.08 219.3 96.5 2 12.67 2.5 1.1
3 12.63 5.5 2.43 12.63 5.5 2.4
I Total 227.2 100.0I Total 227.2 100.0
Tableau GTable G
nombre de pics temps de rétention zone de pic pourcentage de zonenumber of peaks retention time peak area area percentage
1 12.39 273.4 97.21 12.39 273.4 97.2
2 12.93 2.4 0.92 12.93 2.4 0.9
3 13.63 5.4 1.93 13.63 5.4 1.9
Total 281.2 100.0Total 281.2 100.0
Les spectres de masse des peptides P137, P140, Ac 138, Ac 142, Ac 138 + 142, Se et ScP sont représentés respectivement sur les figures 8, 9, 10, 11, 12, 17 et 18.The mass spectra of peptides P137, P140, Ac 138, Ac 142, Ac 138 + 142, Se and ScP are shown in Figures 8, 9, 10, 11, 12, 17 and 18 respectively.
II) Propriétés biologiques La protéine 70K étant fortement phosphorylée in vivo (Woppmann et al, 1993), et bien que le nombre de sites phosphorylés et leur identité ne soient pas connus, les Inventeurs ont synthétisé plusieurs peptides correspondant aux formes phosphorylées et acétylées respectivement sur les résidus serine et lysine du peptide 131-151, et étudié la capacité de ces peptides à être reconnus par les lymphocytes T et les anticorps de souris lupiques. Les Inventeurs démontrent dans le cadre de la présente invention que le peptide phosphorylé en position 140 est reconnu de façon égale voire supérieure au peptide non phosphorylé par les cellules T CD4+ et les anticorps de souris lupiques (figure 13). Les deux peptides (phosphorylé en position 140 et non phosphorylé) ont été utilisés pour une étude de restauration de tolérance au soi. Ces deux peptides ont été injectés par voie intraveineuse et voie intra-nasale à des souris pré-autoi munes, et l'évolution de la maladie chez ces souris a été suivie.II) Biological Properties The protein 70K being strongly phosphorylated in vivo (Woppmann et al, 1993), and although the number of phosphorylated sites and their identity are not known, the inventors have synthesized several peptides corresponding to the phosphorylated and acetylated forms respectively on the serine and lysine residues of peptide 131-151, and studied the capacity of these peptides to be recognized by T lymphocytes and antibodies from lupus mice. The inventors demonstrate in the context of the present invention that the phosphorylated peptide in position 140 is recognized in an equal or even superior manner to the non-phosphorylated peptide by CD4 + T cells and the antibodies of lupus mice (FIG. 13). The two peptides (phosphorylated at position 140 and not phosphorylated) were used for a study of restoration of self-tolerance. These two peptides were injected intravenously and intranasally into pre-auto mice, and the course of the disease in these mice was monitored.
Les Inventeurs ont mis en évidence que l'administration par voie intraveineuse mais pas par voie intra-nasale du peptide phosphorylé P140 diminue la production de forts taux d'anticorps dirigés contre l'ADN, retarde l'apparition de la glomérulonéphrite et prolonge la survie des animaux (figure 14), tandis que le peptide parent n'induit par contre aucun effet statistiquement significatif.The inventors have demonstrated that the administration, intravenously but not intranasally, of the phosphorylated peptide P140 decreases the production of high levels of antibodies directed against DNA, delays the appearance of glomerulonephritis and prolongs survival. animals (Figure 14), while the parent peptide does not induce any statistically significant effect.
De plus, des études avec 3 peptides acétylés sur les lysines 138, 142 et 138+ 142 ont été effectuées. Comme dans le cas des peptides phosphorylés, rien ne permet d'affirmer que ces positions sont réellement acétylées in vivo. Les premiers résultats ont montré que :In addition, studies with 3 acetylated peptides on lysines 138, 142 and 138+ 142 were carried out. As in the case of phosphorylated peptides, there is nothing to confirm that these positions are actually acetylated in vivo. The first results showed that:
- les 3 peptides acétylés sont au moins aussi bien voire mieux reconnus que le peptide parent par les cellules T CD4+ de souris normales immunisées contre le peptide non modifié : les taux de prolifération sont supérieurs, les taux de sécrétion d'IL2 sont équivalents et les taux de production d'interféron γ sont supérieurs,- the 3 acetylated peptides are at least as well or even better recognized than the parent peptide by the CD4 + T cells of normal mice immunized against the unmodified peptide: the proliferation rates are higher, the IL2 secretion rates are equivalent and the interferon γ production rates are higher,
- les 3 peptides acétylés sont au moins aussi bien voire mieux reconnus que le peptide parent par les cellules T CD4+ de souris autoimmunes : les taux de prolifération sont supérieurs,the 3 acetylated peptides are at least as well or even better recognized than the parent peptide by the CD4 + T cells of autoimmune mice: the proliferation rates are higher,
- les 3 peptides acétylés sont reconnus par les anticorps de souris dirigés contre le peptide parent.- The 3 acetylated peptides are recognized by the antibodies of mice directed against the parent peptide.
Enfin les Inventeurs ont démontré que les peptides 131-151 et PI 40 étaient capables de lier diverses molécules du CMH de classe II humaines (HLA-DR1, -DR4 et -DRU) (Figure 19). Finally, the inventors have demonstrated that the peptides 131-151 and PI 40 are capable of binding various human MHC class II molecules (HLA-DR1, -DR4 and -DRU) (Figure 19).
Références bibliographiquesBibliographic references
Andersen M.H. , Bonfill J.E. , Neisig A. , Arsequell G. , Sondergaard L , Valencia G. , Neefjes J. , Zeuthen J. , Elliot T. and Haururn J. S. (1999) Phosphorylated peptides can be transported by TAP molécules, presented by class I MHC molécules, and recognized by phosphorylated-specific CTL. J. Immunol. 163:3812-3818.Andersen MH, Bonfill JE, Neisig A., Arsequell G., Sondergaard L, Valencia G., Neefjes J., Zeuthen J., Elliot T. and Haururn JS (1999) Phosphorylated peptides can be transported by TAP molecules, presented by class I MHC molecules, and recognized by phosphorylated-specific CTL. J. Immunol. 163: 3812-3818.
Eilat E. , Zinger H. , Nyska A. and Mozes E. (2000) Prévention of systemic lupus erythematosus-like disease in (NZBxNZW)Fl mice by treating with CDR1- and CDR3-based ι peptides of a pathogenic autoantibody. J. Clin. Immunol. 20 :268-278. Jouanne C , Avrameas S. and Payelle-Brogard B. (1999) A peptide derived from a polyreactive monoclonal anti-DNA natural antibody modulate lupus development in (NZBxNZW)Fl mice. Immunology 96:333-339.Eilat E., Zinger H., Nyska A. and Mozes E. (2000) Prévention of systemic lupus erythematosus-like disease in (NZBxNZW) Fl mice by treating with CDR1- and CDR3-based ι peptides of a pathogenic autoantibody. J. Clin. Immunol. 20: 268-278. Jouanne C, Avrameas S. and Payelle-Brogard B. (1999) A peptide derived from a polyreactive monoclonal anti-DNA natural antibody modulate lupus development in (NZBxNZW) Fl mice. Immunology 96: 333-339.
Kaliyaperumal A. , Michaels M.A. ans Datta S.K. (1999) Antigen-specific therapy of murine lupus nephritis using nucleosomal peptides: tolérance spreading impairs pathogenic function of autoimmune T and B cells. J. Immunol. 162:5775-5783.Kaliyaperumal A., Michaels M.A. ans Datta S.K. (1999) Antigen-specific therapy of murine lupus nephritis using nucleosomal peptides: tolerance spreading odd pathogenic function of autoimmune T and B cells. J. Immunol. 162: 5775-5783.
Klein Gunnewiek, J. M. T. Van De Putte, L.B.A. and van Venrooij, W. J., The Ul snRNP complex : an autoantigen in connective tissue disease. Clin. Exp. Rheumatol. 1997, 15 : 549-560.Klein Gunnewiek, J. M. T. Van De Putte, L.B.A. and van Venrooij, W. J., The Ul snRNP complex: an autoantigen in connective tissue disease. Clin. Exp. Rheumatol. 1997, 15: 549-560.
Marino M. , Ruvo M. , de Fales S. and Facsina G. (2000) Prévention of systemic lupus erythematosus in MRL/lpr mice by adrnmistration of an immunoglobulin-binding peptide. Nature Biotechn. 18 : 735-739.Marino M., Ruvo M., de Fales S. and Facsina G. (2000) Prévention of systemic lupus erythematosus in MRL / lpr mice by adrnmistration of an immunoglobulin-binding peptide. Nature Biotechn. 18: 735-739.
Monneaux F., Briand J.-P. and Muller S. (2000) B and T cell immune response to snRNP in lupus mice. Autoreactive CD4+ T cells recognize a T cell epitope located within the conserved RNP consensus séquence of the 70K protein. Eur. J. Immunol. 20 :2191- 2200.Monneaux F., Briand J.-P. and Muller S. (2000) B and T cell immune response to snRNP in lupus mice. Autoreactive CD4 + T cells recognize a T cell epitope located within the conserved RNP consensus sequence of the 70K protein. Eur. J. Immunol. 20: 2191-2200.
Monneaux F. and Muller S. (2000) Laboratory protocols for the identification of Th cell épitopes on self antigens in mice with systemic autoimmune diseases. /. Immunol. Meth. 244 : 195-204.Monneaux F. and Muller S. (2000) Laboratory protocols for the identification of Th cell epitopes on self antigens in mice with systemic autoimmune diseases. /. Immunol. Meth. 244: 195-204.
Singh R.R. , Ebling F.M., Sercarz E.E. and Hahn B.H. (1995) Immune tolérance to autoantibody-derived peptides delays development of autoimmunity in murine lupus. J. Clin.Singh R.R., Ebling F.M., Sercarz E.E. and Hahn B.H. (1995) Immune tolerance to autoantibody-derived peptides delays development of autoimmunity in murine lupus. J. Clin.
Invest. 96:2990-2996. Utz, P. J. , and P. Anderson. (1998). Posttranslational protein modifications, apoptosis, ans the bypass of tolérance to autoantigens, Arthritis Rheum 41 : 1152-1160.Invest. 96: 2990-2996. Utz, PJ, and P. Anderson. (1998). Posttranslational protein modifications, apoptosis, ans the bypass of tolerance to autoantigens, Arthritis Rheum 41: 1152-1160.
Woppmann, A. C. L. Will. U. Kornstadt, P. Zuo, J. L. Manley, and R. Lurhmann, (1993). Identification of an snRNP-associated kinase activity that phosphorylatres arginine/serine rich domains typical of slicing factors. Nucleic Acids Res 21 : 2815-2822.Woppmann, A. C. L. Will. U. Kornstadt, P. Zuo, J. L. Manley, and R. Lurhmann, (1993). Identification of an snRNP-associated kinase activity that phosphorylatres arginine / serine rich domains typical of slicing factors. Nucleic Acids Res 21: 2815-2822.
Zarling A. L., Ficarro S.B., Shabanowitz J., Hunt D. F. and Engelhard V.H. (2000) Phosphorylated peptides are naturally processed and presented by major histocompatibility complex class I molécules in vivo. /. Exp. Med. 192:1755-1762.Zarling A. L., Ficarro S.B., Shabanowitz J., Hunt D. F. and Engelhard V.H. (2000) Phosphorylated peptides are naturally processed and presented by major histocompatibility complex class I molecules in vivo. /. Exp. Med. 192: 1755-1762.
Légende des figuresLegend of figures
- Figure 1 : reconnaissance du peptide 131-151 de la protéine 70K par les cellules T CD4+ de souris lupiques MRL/lpr (colonne de droite) ou NZB/W (colonne de gauche); les concentrations en peptide sont indiquées en abscisse, la prolifération des cellules T CD4+ de souris MRL/lpr et BW, représentée en ordonnées des graphes de la première ligne, est mesurée ex vivo en présence des différentes concentrations de peptide 131-151 de la protéine 70K; la prolifération est exprimée en indices de stimulation correspondant à la radioactivité incorporée dans l'ADN des cellules (en coups par minutes) en présence de peptide sur l'incorporation de radioactivité en absence de peptide; la sécrétion d'IL-2, représentée en ordonnées des graphes de la deuxième ligne, est mesurée dans les surnageants après 24h de culture par un test biologique; la concentration en LL-2 est déteiminée grâce à une gamme étalon d'IL-2 recombinante et est exprimée en mU/ml.- Figure 1: recognition of the 131-151 peptide of the 70K protein by CD4 + T cells of lupus mice MRL / lpr (right column) or NZB / W (left column); the peptide concentrations are indicated on the abscissa, the proliferation of CD4 + T cells of MRL / lpr and BW mice, represented on the ordinates of the graphs in the first line, is measured ex vivo in the presence of the different concentrations of peptide 131-151 of the protein 70K; the proliferation is expressed in stimulation indices corresponding to the radioactivity incorporated into the DNA of the cells (in counts per minute) in the presence of peptide on the incorporation of radioactivity in the absence of peptide; the secretion of IL-2, represented on the ordinates of the graphs of the second line, is measured in the supernatants after 24 h of culture by a biological test; the LL-2 concentration is determined by means of a standard range of recombinant IL-2 and is expressed in mU / ml.
- Figure 2 : liaison du peptide 131-151 aux molécules du complexe majeur d'histocompatibilité de clase II murines (I-Ek, I-A\ I-Ed, I-Ad); des fibroblastes transfectés par les molécules I-E\ ou I-Ak, ou I-Ed, ou I-Ad sont pré-incubés avec différentes concentrations du peptide 131-151; après 30 min à 37 °C, les peptides analogues 12-26CI ou 16-33 du récepteur β2-adrénergique, ainsi que les hybridomes T respectifs qui reconnaissent ces peptides dans le contexte adapté (81 pour I-Ek, E7E9 pour I-Ak, 26.1 pour I-Ed, et 26.2 pour I-Ad) sont ajoutés; les surnageants sont récupérés après 24 h de culture et la sécrétion d'IL-2 est évaluée comme précédemment; les résultats sont exprimés en % d'inhibition représentant la capacité du peptide 131-151 à inhiber la liaison des peptides analogues 12-26 CI et 16-33β2 aux molécules des classes IL - Figure 3 : profil HPLC du peptide P137.- Figure 2: binding of peptide 131-151 to the molecules of the major murine clase II histocompatibility complex (IE k , IA \ IE d , IA d ); fibroblasts transfected with the molecules IE \ or IA k , or IE d , or IA d are pre-incubated with different concentrations of the peptide 131-151; after 30 min at 37 ° C., the peptides analogs 12-26CI or 16-33 of the β2-adrenergic receptor, as well as the respective T hybridomas which recognize these peptides in the appropriate context (81 for IE k , E7E9 for IA k , 26.1 for IE d , and 26.2 for IA d ) are added; the supernatants are recovered after 24 h of culture and the secretion of IL-2 is evaluated as above; the results are expressed in% of inhibition representing the capacity of the peptide 131-151 to inhibit the binding of analog peptides 12-26 CI and 16-33β2 to molecules of classes IL - Figure 3: HPLC profile of the P137 peptide.
- Figure 4 : profil HPLC du peptide P140.- Figure 4: HPLC profile of the P140 peptide.
- Figure 5 : profil HPLC du peptide Ac 138.- Figure 5: HPLC profile of the Ac 138 peptide.
- Figure 6 : profil HPLC du peptide Ac 142. - Figure 7 : profil HPLC du peptide Acl38 + 142.- Figure 6: HPLC profile of the Ac 142 peptide. - Figure 7: HPLC profile of the Acl38 + 142 peptide.
- Figure 8 : spectre de masse du peptide PI 37.- Figure 8: mass spectrum of the peptide PI 37.
- Figure 9 : spectre de masse du peptide PI 40.- Figure 9: mass spectrum of the PI 40 peptide.
- Figure 10 : spectre de masse du peptide Ac 138. i- Figure 10: mass spectrum of the Ac 138 peptide. I
- Figure 11 : spectre de masse du peptide Ac 142. - Figure 12 : spectre de masse du peptide Ac 138 + 142.- Figure 11: mass spectrum of the Ac 142 peptide. - Figure 12: mass spectrum of the Ac 138 + 142 peptide.
- Figure 13 : reconnaissance du peptide P140- Figure 13: recognition of the P140 peptide
. par les lymphocytes T CD4+ (A) de souris lupiques MRL/lpr ; le graphe de gauche représente la prolifération des cellules T CD4+ exprimée en indices de stimulation tels que définis ci-dessus en présence du peptide 131-151, des peptides phosphorylés PI 37 et PI 40 et de deux peptides .brouillés phosphorylés ou non (Se et ScP) ; la limite de positivité correspond à 2.0 ; le graphe de droite représente la sécrétion d'IL-2 (mU/ml) en présence du peptide 131-151, des peptides P137 et P140 et des peptides Se et ScP.. by CD4 + T lymphocytes (A) from MRL / lpr lupus mice; the graph on the left represents the proliferation of CD4 + T cells expressed in stimulation indices as defined above in the presence of the peptide 131-151, phosphorylated peptides PI 37 and PI 40 and two scrambled peptides phosphorylated or not (Se and ScP); the positivity limit corresponds to 2.0; the graph on the right represents the secretion of IL-2 (mU / ml) in the presence of peptide 131-151, peptides P137 and P140 and peptides Se and ScP.
. et par les anticorps (B) de souris lupiques MRL/lpr ; les résultats sont exprimés en titre d'anticorps correspondant à la dilution permettant d'obtenir en test ELISA une valeur de DO à 450nm égale à 0.2.. and by antibodies (B) from MRL / lpr lupus mice; the results are expressed as antibodies corresponding to the dilution making it possible to obtain, in the ELISA test, an OD value at 450 nm equal to 0.2.
- Figure 14 : effet de l'administration à des souris pré-autoimmunes MRL/lpr de la forme phosphorylée du peptide 131-151 de la protéine 70K . peptide PI 40) ; le graphe de gauche représente les groupes de souris qui ont reçu le peptide ou la solution saline par voie intraveineuse et le graphe de droite représente les souris qui ont reçu le peptide ou la solution saline par voie intranasale ; les résultats sont exprimés en pourcentage de survie en fonction de l'âge en semaines des souris lupiques utilisées (A), en pourcentage de protéinurie positive en fonction de l'âge en semaines des souris lupiques utilisées (B) et en pourcentage de taux élevés d'anticorps dirigés contre l'ADN en fonction de l'âge des souris lupiques utilisées (C) ; les dates d'administration sont représentées par des flèches, les symboles et barres vides correspondent aux souris ayant reçu le peptide P140 ; le groupe contrôle représenté par les symboles et barres pleins n'a reçu que la solution saline : dans le groupe injecté avec le peptide 131-151 nominal non phosphorylé, 25% des souris ont survécu à 35 semaines (p=0.2 par rapport aux souris contrôles), et la protéinurie était à peine diminuée.- Figure 14: effect of administration to pre-autoimmune MRL / lpr mice of the phosphorylated form of peptide 131-151 of the protein 70K. peptide PI 40); the graph on the left represents the groups of mice which have received the peptide or the saline solution by the intravenous route and the graph on the right represents the mice which have received the peptide or the saline solution by the intranasal route; the results are expressed as a percentage of survival as a function of the age in weeks of the lupus mice used (A), as a percentage of positive proteinuria as a function of the age in weeks of the lupus mice used (B) and as a percentage of high rates antibodies directed against DNA as a function of the age of the lupus mice used (C); the dates of administration are represented by arrows, the symbols and empty bars correspond to the mice having received the P140 peptide; the control group represented by the solid symbols and bars received only the saline solution: in the group injected with the non-phosphorylated nominal peptide 131-151, 25% of the mice survived 35 weeks (p = 0.2 compared to the control mice), and the proteinuria was hardly reduced.
- Figure 15 : profil HPLC du peptide Se- Figure 15: HPLC profile of the Se peptide
- Figure 16 : profil HPLC du peptide ScP- Figure 16: HPLC profile of the ScP peptide
- Figure 17 : spectre de masse du peptide Se- Figure 17: mass spectrum of the Se peptide
- Figure 18 : spectre de masse du peptide ScP- Figure 18: mass spectrum of the ScP peptide
- Figure 19 : liaison des peptides 131-151 et P140 aux molécules du complexe majeur d'histocompatibilité de classe II humaines ; les résultats sont exprimés en pourcentage de liaison des peptides aux molécules HLA-DRl, -DR4 et DRU ; le pourcentage d'inhibition est calculé en fonction des valeurs de DO mesurées en présence de différentes concentrations de peptides 131-151 et P140 (0.01-lOOμM). - Figure 19: binding of peptides 131-151 and P140 to molecules of the major human class II histocompatibility complex; the results are expressed as a percentage of binding of the peptides to the HLA-DR1, -DR4 and DRU molecules; the percentage of inhibition is calculated as a function of the OD values measured in the presence of different concentrations of peptides 131-151 and P140 (0.01-100 μM).

Claims

REVENDICATIONS
1. Utilisation de peptides comprenant des épitopes de protéines du soi des mammifères reconnus par des anticorps produits par le système immumtaire des mammifères atteints de pathologies auto-immunes, et, le cas échéant, par les cellules T auxiliaires desdits mammifères, lesdits épitopes étant tels que l'un au moins de leurs acides i aminés comporte une modification de type post-traductionnel, pour la préparation d'un médicament destiné à la prévention ou au traitement desdites pathologies auto-immunes.1. Use of peptides comprising epitopes of mammalian self proteins recognized by antibodies produced by the immune system of mammals suffering from autoimmune pathologies, and, where appropriate, by the helper T cells of said mammals, said epitopes being such that at least one of their amino acids comprises a modification of the post-translational type, for the preparation of a medicament intended for the prevention or the treatment of said autoimmune pathologies.
2. Utilisation de peptides selon la revendication 1, comprenant des épitopes dont l'un au moins de leurs acides aminés est modifié de manière à ce qu'il soit sous une forme phosphorylée, ou acétylée.2. Use of peptides according to claim 1, comprising epitopes of which at least one of their amino acids is modified so that it is in a phosphorylated or acetylated form.
3. Utilisation selon la revendication 1 ou 2, de peptides comprenant des épitopes issus des protéines d'origine humaine ou animale défîmes dans la revendication 1, choisies parmi les nucléoprotéines, les protéines du nucléosome, du splicéosome, de la particule ribonucléoprotéique Ro, ou du ribososme.3. Use according to claim 1 or 2, of peptides comprising epitopes derived from proteins of human or animal origin defined in claim 1, chosen from nucleoproteins, proteins of the nucleosome, of the spliceosome, of the ribonucleoprotein particle Ro, or ribososm.
4. Utilisation de peptides selon l'une des revendications 1 à 3, pour la préparation d'un médicament destiné à la prévention ou au traitement :4. Use of peptides according to one of claims 1 to 3, for the preparation of a medicament intended for the prevention or the treatment:
- de pathologies auto-immunes de la famille des connectivités (maladies systémiques non spécifiques d'organes), telles que le lupus érythémateux disséminé (LED), la polyarthrite rhumatoïde, la connectivité mixte, le syndrome de Sjôgren, ou l'arthrite chronique juvénile,- autoimmune pathologies of the family of connectivities (non-specific systemic organ diseases), such as systemic lupus erythematosus (LED), rheumatoid arthritis, mixed connectivity, Sjôgren syndrome, or chronic juvenile arthritis ,
- ou de pathologies auto-immunes spécifiques d'organes, telles que la sclérose en plaques, le diabète insulino-dépendant, la maladie de Crohn, ou les maladies huileuses.- or specific autoimmune organ pathologies, such as multiple sclerosis, insulin-dependent diabetes, Crohn's disease, or oily diseases.
5. Utilisation de peptides selon l'une des revendications 1 à 4, pour la préparation d'un médicament destiné à la prévention ou au traitement du LED. 5. Use of peptides according to one of claims 1 to 4, for the preparation of a medicament intended for the prevention or treatment of LED.
6. Utilisation selon la revendication 5, de peptides comprenant des épitopes issus de la protéine humaine ou murine U1-70K du splicéosome.6. Use according to claim 5, of peptides comprising epitopes derived from the human or murine protein U1-70K of the spliceosome.
7. Utilisation selon la revendication 5 ou 6, de peptides comprenant la séquence délimitée par les acides aminés 131 et 151 de la protéine U1-70K humaine ou murine, et correspondant à la séquence SEQ ID NO : 1 suivante :7. Use according to claim 5 or 6, of peptides comprising the sequence delimited by amino acids 131 and 151 of the human or murine U1-70K protein, and corresponding to the following sequence SEQ ID NO: 1:
RIHMVYSKRSGKPRGYAFIEYRIHMVYSKRSGKPRGYAFIEY
I dans laquelle l'un au moins des acides aminés comporte une modification de type post- traductionnelle, notamment par phosphorylation, ou acetylation.I in which at least one of the amino acids comprises a modification of the post-translational type, in particular by phosphorylation, or acetylation.
8. Utilisation selon l'une des revendications 5 à 7, de peptides comprenant la séquence SEQ ID NO : 1 dans laquelle l'un au moins des résidus serine en position 7 ou 10 est phosphorylé, et/ou l'un au moins des résidus lysine en position 8 ou 12 est acétylé.8. Use according to one of claims 5 to 7, of peptides comprising the sequence SEQ ID NO: 1 in which at least one of the serine residues in position 7 or 10 is phosphorylated, and / or at least one of lysine residue in position 8 or 12 is acetylated.
9. Utilisation selon l'une des revendications 5 à 8, de peptides comprenant la séquence SEQ ID NO : 1 dans laquelle :9. Use according to one of claims 5 to 8, of peptides comprising the sequence SEQ ID NO: 1 in which:
- la serine en position 7 est phosphorylée,- the serine in position 7 is phosphorylated,
- et/ou la serine en position 10 est phosphorylée, - et/ou la lysine en position 8 est acétylée,- and / or the serine in position 10 is phosphorylated, - and / or the lysine in position 8 is acetylated,
- et/ou la lysine en position 12 est acétylée.- And / or the lysine in position 12 is acetylated.
10. Utilisation selon l'une des revendications 5 à 9, de peptides choisis parmi les suivants : - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée,10. Use according to one of claims 5 to 9, of peptides chosen from the following: - the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,- the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 12 is acetylated, the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 7 thus that the serine at position 10 is phosphorylated, and the lysine at position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,- the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 12 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées.- The sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
11. Utilisation selon l'une des revendications 5 à 10, de peptides choisis parmi les suivants :11. Use according to one of claims 5 to 10, of peptides chosen from the following:
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées. - The sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
12. Composition pharmaceutique caractérisée en ce qu'elle comprend au moins un peptide choisi parmi ceux définis dans l'une des revendications 1 à 11, en association avec un véhicule pharmaceutiquement acceptable.12. Pharmaceutical composition, characterized in that it comprises at least one peptide chosen from those defined in one of claims 1 to 11, in combination with a pharmaceutically acceptable vehicle.
13. Composition pharmaceutique selon la revendication 12, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi ceux comprenant la séquence SEQ ID NO : 1 dans laquelle l'un au moins des acides aminés comporte une modification de type post- traductionnel, notamment par phosphorylation, ou acetylation..13. Pharmaceutical composition according to claim 12, characterized in that it comprises at least one peptide chosen from those comprising the sequence SEQ ID NO: 1 in which at least one of the amino acids comprises a modification of post-translational type, in particular by phosphorylation, or acetylation.
14. Composition pharmaceutique selon la revendication 12 ou 13, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi ceux comprenant la séquence SEQ ID NO : 1 dans laquelle l'un au moins des résidus serine en position 7 où 10 est phosphorylé, et/ou l'un au moins des résidus lysine en position 8 ou 12 est acétylé.14. Pharmaceutical composition according to claim 12 or 13, characterized in that it comprises at least one peptide chosen from those comprising the sequence SEQ ID NO: 1 in which at least one of the serine residues in position 7 where 10 is phosphorylated , and / or at least one of the lysine residues in position 8 or 12 is acetylated.
15. Composition pharmaceutique selon l'une des revendications 12 à 14, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi ceux comprenant la séquence SEQ ID NO : 1 dans laquelle :15. Pharmaceutical composition according to one of claims 12 to 14, characterized in that it comprises at least one peptide chosen from those comprising the sequence SEQ ID NO: 1 in which:
- la serine en position 7 est phosphorylée,- the serine in position 7 is phosphorylated,
- et/ou la serine en position 10 est phosphorylée, - et/ou la lysine en position 8 est acétylée,- and / or the serine in position 10 is phosphorylated, - and / or the lysine in position 8 is acetylated,
- et/ou la lysine en position 12 est acétylée.- And / or the lysine in position 12 is acetylated.
16. Composition pharmaceutique selon l'une des revendications 13 à 15, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi les suivants : - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée,16. Pharmaceutical composition according to one of claims 13 to 15, characterized in that it comprises at least one peptide chosen from the following: - the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,- the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 12 is acetylated, the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 7 thus that the serine at position 10 is phosphorylated, and the lysine at position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,- the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 12 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées.- The sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
17. Composition pharmaceutique selon l'une des revendications 13 à 16, caractérisée en ce qu'elle comprend au moins un peptide choisi parmi les sμivants :17. Pharmaceutical composition according to one of claims 13 to 16, characterized in that it comprises at least one peptide chosen from the following:
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées. - The sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
18. Composition pharmaceutique selon l'une des revendications 13 à 17, caractérisée en ce qu'elle se présente sous une forme administrable par voie systémique, ou non invasive.18. Pharmaceutical composition according to one of claims 13 to 17, characterized in that it is in a form which can be administered systemically or non-invasively.
19. Composition pharmaceutique selon l'une des revendications 13 à 18, caractérisée en ce que les doses journalières de peptides chez l'homme sont d'environ 100 ng à environ 5 mg. i19. Pharmaceutical composition according to one of claims 13 to 18, characterized in that the daily doses of peptides in humans are from about 100 ng to about 5 mg. i
20. Peptides comprenant la séquence délimitée par les acides aminés 131 et 151 de la protéine U1-70K humaine, ou murine, et correspondant à la séquence SEQ ID NO : 1 suivante : RIHMVYSKRSGKPRGYAFIEY dans laquelle l'un au moins des acides aminés est phosphorylé, ou acétylé.20. Peptides comprising the sequence delimited by amino acids 131 and 151 of the human U1-70K protein, or murine, and corresponding to the following sequence SEQ ID NO: 1: RIHMVYSKRSGKPRGYAFIEY in which at least one of the amino acids is phosphorylated , or acetylated.
21. Peptides selon la revendication 20, comprenant la séquence SEQ ID NO : 1 dans laquelle l'un au moins des résidus serine en position 7 ou 10 est phosphorylé, et/ou l'un au moins des résidus lysine en position 8 ou 12 est acétylé.21. Peptides according to claim 20, comprising the sequence SEQ ID NO: 1 in which at least one of the serine residues in position 7 or 10 is phosphorylated, and / or at least one of the lysine residues in position 8 or 12 is acetylated.
22. Peptides selon la revendication 20 ou 21, comprenant la séquence SEQ ID NO : 1 dans laquelle : - la serine en position 7 est phosphorylée,22. Peptides according to claim 20 or 21, comprising the sequence SEQ ID NO: 1 in which: the serine in position 7 is phosphorylated,
- et/ou la serine en position 10 est phosphorylée,- And / or the serine in position 10 is phosphorylated,
- et/ou la lysine en position 8 est acétylée,- And / or the lysine in position 8 is acetylated,
- et/ou la lysine en position 12 est acétylée.- And / or the lysine in position 12 is acetylated.
23. Peptides selon l'une des revendications 20 à 22, choisis parmi les suivants :23. Peptides according to one of claims 20 to 22, chosen from the following:
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées,the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 12 is acetylated, the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 7 thus that the serine at position 10 is phosphorylated, and the lysine at position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 ainsi que la serine en position 10 sont phosphorylées, et la lysine en position 8 ainsi que la lysine en position! 12 sont acétylées, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 as well as the serine in position 10 are phosphorylated, and the lysine in position 8 as well as the lysine in position ! 12 are acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 12 est acétylée, - la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, et la lysine en position 8 ainsi que la lysine en position 12 sont acétylées,- the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 12 is acetylated, - the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, and the lysine in position 8 as well as the lysine in position 12 are acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées.- The sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
24. Peptides selon l'une des revendications 20 à 23, choisis parmi les suivants :24. Peptides according to one of claims 20 to 23, chosen from the following:
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 7 est phosphorylée,- the sequence SEQ ID NO: 1 in which the serine in position 7 is phosphorylated,
- la séquence SEQ ID NO : 1 dans laquelle la serine en position 10 est phosphorylée, - la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 est acétylée,the sequence SEQ ID NO: 1 in which the serine in position 10 is phosphorylated, the sequence SEQ ID NO: 1 in which the lysine in position 8 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 12 est acétylée,- the sequence SEQ ID NO: 1 in which the lysine in position 12 is acetylated,
- la séquence SEQ ID NO : 1 dans laquelle la lysine en position 8 et la lysine en position 12 sont acétylées. - The sequence SEQ ID NO: 1 in which the lysine in position 8 and the lysine in position 12 are acetylated.
EP02798771A 2001-09-18 2002-09-18 Use of peptides comprising post-translational modifications in the treatment of autoimmune pathologies Ceased EP1427755A2 (en)

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FR2931360B1 (en) 2008-05-20 2012-05-18 Centre Nat Rech Scient NANOPARTICLES CONTAINING A PEPTIDE, VECTORS CONTAINING SAME AND PHARMACEUTICAL USES OF SAID NANOPARTICLES AND VECTORS
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US10213482B2 (en) 2014-12-12 2019-02-26 Immupharma France Sa Methods of treating chronic inflammatory diseases
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US7872098B1 (en) 2011-01-18
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FR2829768B1 (en) 2003-12-12

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