WO1992016234A1 - Procede de modulation de la reaction de lymphocytes t de mammiferes - Google Patents

Procede de modulation de la reaction de lymphocytes t de mammiferes Download PDF

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Publication number
WO1992016234A1
WO1992016234A1 PCT/US1992/002419 US9202419W WO9216234A1 WO 1992016234 A1 WO1992016234 A1 WO 1992016234A1 US 9202419 W US9202419 W US 9202419W WO 9216234 A1 WO9216234 A1 WO 9216234A1
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peptide
mhc
cell
antigen
binding
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PCT/US1992/002419
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English (en)
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William V. Williams
Donald H. Rubin
David B. Weiner
Mark I. Greene
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The Trustees Of The University Of Pennsylvania
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Priority to JP4510619A priority Critical patent/JPH06507630A/ja
Priority to AU17950/92A priority patent/AU669329B2/en
Publication of WO1992016234A1 publication Critical patent/WO1992016234A1/fr

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    • 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
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • C07K14/70503—Immunoglobulin superfamily
    • C07K14/70539—MHC-molecules, e.g. HLA-molecules
    • 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
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • This invention relates to the field of mammalian therapeutics. More particularly, the invention relates to novel methods of modulating mammalian T-cell response restricted by an MHC and methods of treating MHC-linked diseases in a mammal with compounds capable of binding with a T-cell antigen receptor that recognize the MHC bound to an antigen.
  • MHC major histocompatibility complex
  • HLA in the human has been divided into five major gene loci, which according to World Health Organization nomenclature are designated HLA-A, HLA-B, HLA-C, HLA-D, and HLA-DR.
  • the A, B, and C loci are single gene loci.
  • the D and DR loci are multi-gene loci.
  • the A, B and C loci encode the classical transplantation antigens, whereas the D and DR loci encode products that control immune responsiveness. More recent definitions divide the gene products of the HLA loci into three classes (I, II, and III) based on structure and function.
  • Class I encompasses the products of the HLA-A, HLA- B, and HLA-C loci and the Qa/TL region.
  • the products of the HLA-D and HLA-DR related genes fall in Class II.
  • the Class II antigens are believed to be heterodimers composed of an alpha (approx. 34,000 daltons) glycopeptide and a beta (approx. 29,000 daltons) glycopeptide.
  • the number of loci and the gene order of Class II are tentative.
  • the third class. Class III includes components of complement.
  • MHC is intended to include the above described loci as well as loci that are closely linked thereto.
  • the class II antigen products are essential in the normal immune response for the triggering of the activation steps which lead to immunity. Even when the immune system is activated inappropriately, and attacks normal tissue, causing autoimmunity, these class II molecules play an essential role in the immune activation which leads to disease.
  • the target is assumed to reside in the synovial lining of the joints.
  • T-cells are derived from the thymus and accordingly they are called T-cells. They circulate freely through the blood and lymphatic vessels of the body, and so are able to detect and react against foreign invaders, i.e., viruses, allergens, tumors and autoantigens. Despite their uniform morphology under microscope, T-cells consist of a heterogeneous population of cells with several distinct functional subsets includinghelpers, suppressors and killers. Through a recognition system called the T-cell antigen receptor (TCR) , T-cells are able to detect the presence of invading pathogens and direct release of multiple.
  • TCR T-cell antigen receptor
  • T-cell factors which instruct B lymphocytes to initiate or suppress antibody production, and regulate the white blood system in producing more phagocytes and other white cells to neutralize the pathogens, and destroy tumor cells and virally infected cells.
  • T-cell factors T-cell antigen (Ag) receptors (TCRs) . These are believed to bind to antigenic peptides held in the groove of MHC molecules.
  • the Ag-MHC complex is formed on antigen presenting cells (APCs) following internalization and processing of the Ag into a form that can associate with MHC molecules. Both antigenic peptide and MHC molecule are required for T-cell activation. Together they form a trimolecular complex which is somewhat unique in receptor biology. Most ligand-receptor or receptor-receptor interactions are bimolecular. The trimolecular nature of the TCR-Ag-MHC complex has made the interactions involved particularly difficult to dissect.
  • T-cell clones specific for fluorescein + MHC have been established, and these have low affinity binding interactions with fluorescein alone.
  • R.F. Siliciano et al. "Direct evidence for the existence of nominal antigen binding sites on T-cell surface Ti alpha-beta heterodimers of MHC- restricted T-cell clones," Cell 47:161-171 (1986). This implies a direct interaction of the TCR with Ag in some instances. This was also implied in studies of T-cell mediated association of antigenic peptides with MHC molecules utilizing fluorescence energy transfer, T.H. Watts et al.
  • H-2K b IL-2 production in response to H-2I ⁇ a10 was diminished by soluble H-2K b as well as a peptide derived from amino acids 163-174 of H-2I ⁇ but not a similar peptide derived from the H- 2K ***010 sequence.
  • CTLs allospecific cytotoxic T lymphocytes
  • This peptide also ⁇ _ j blocked lysis of H-2k targets but not H-2L targets by a single bulk CTL culture alloreactive for both specificities.
  • studies of a similar peptide derived from amino acids 111-122 of the H-2IC molecule revealed another potential explanation for these findings, W.R. Heath et al., supra
  • the role of antigen within the binding groove in enhancing interaction with the T-cell receptor can be at least two-fold.
  • the TCR has a low affinity for the MHC molecule alone, and the antigen functions chiefly by directly binding the TCR, enhancing the affinity of the TCR for the Ag-MHC complex.
  • the TCR has a low affinity for the MHC molecule which is due to some strong attractive interactions and some similarly strong repulsive interactions.
  • antigen functions byreducing repulsive interactions, for example by conformationally altering the orientation of repulsive residues.
  • a novel method of treating an MHC-linked disease in a mammal suspected of needing such treatment comprising administering to said mammal an effective amount of a peptide, which peptide has an amino acid sequence substantially corresponding to at least a portion of the antigen recognition site of said MHC, or a peptide mimetic wherein said peptide or peptide mimetic is capable of binding with a T-cell antigen receptor which unbound T-cell antigen receptor is capable of recognizing said MHC bound to an antigen.
  • a novel method of modulating T-cell response restricted by an MHC in a mammal suspected of needing such modulation comprising contacting said T-cells with an effective amount of a peptide, which peptide has an amino acid sequence substantially corresponding to at least a portion of the antigen recognition site of said MHC, or a peptide mimetic wherein said peptide or peptide mimetic is capable of binding with a T-cell antigen receptor which unbound T-cell antigen receptor is capable of recognizing said MHC bound to an antigen. It is believed that peptides and peptide mimetics derivable from the MHC antigen recognition site that bind to T-cell receptors are useful as biologically active, immunomodulatory substances as more precisely detailed herein.
  • Antibodies were prepared from ascites or culture supernatant by ammonium sulfate precipitation, dialyzed, and diluted in FACS buffer 1% BSA in PBS with .1% sodium azide. Solid phase radioimmunoassay (RIA) was utilized to study binding as described.
  • RIA Solid phase radioimmunoassay
  • Antibodies were preincubated with 1 mg/ l (A) or varying amounts (B) of peptides prior to use in FACS analysis for binding to IA k molecules expressed on RT4.15.HP cells.
  • A the ⁇ mean channel number is shown for cells stained with 10.2.16 versus 15-1-5P.
  • B the % decrease in ⁇ mean channel number is shown for 10.2.16 binding in the presence of increasing amounts of IA 68 _ 83 peptide.
  • FIG. 3 Inhibition of D10.G4 proliferation by IA 68 - 83 peptide.
  • A counts per minute (CPM) incorporated is shown versus increasing amounts of IA k peptide for specific antigen (conalbumin) and anti-TCR e antibody (2C11) .
  • B % inhibition of proliferation is shwown for CPM incorporated in the presence of increasing amounts of IA 68 _ 83 peptide.
  • FIG. 4 Antigen presenting cell (APC) dose dependence of IA k 68 _ 83 peptide inhibition of D10.G4 proliferation.
  • D10.G4 cells were stimulated with conalbumin and two doses of APCs as described in materials and methods, in the presence of varying amounts of IA peptide.
  • % maximal ⁇ CPM incorporated is shown for increasing doses of peptide. Maximal ⁇ CPM incorporated with 5 x 10 APCs was approximately 15,000, and with 5 x 10 APCs was approximately 5,000.
  • IA 68 _ 83 peptide D10.G4 were preincubated with IA 68 . 83 peptide
  • FIG. 7 Binding of IA k 68 . 83 peptide-BSA conjugates to D10.G4 cells.
  • the peptide-BSA conjugates were fluorsceinated as noted in Materials and Methods. Cells were incubated with a 1:10 dilution of fluoroscein isothiocyonate (FITC)-peptide-BSA in FACS buffer for 45 minutes at room temperature, washed twice and analyzed. D10.G4 or 22.D11 cells were incubated with either FITC-1S1 peptide-BSA (left) , or with FITC-IA 68 _ 83 peptide-BSA (right) . The mean channel number is shown for the different cell lines incubated with the conjugates.
  • FITC fluoroscein isothiocyonate
  • Figure 8 Inhibition of FITC-peptide-BSA binding to cells.
  • A Cells were preincubated with 100 ⁇ l unfluorsceinated peptide-BSA conjugates at 1 mg/ml for 45 minutes at room temperature. The FITC-IA 68 . 83 peptide-BSA conjugate was then added for an additional 45 minutes at room temperature, the cells washed twice and analyzed.
  • B Cells were preincubated with 100 ⁇ l of supraoptimal concentrations of each antibody (undiluted ammonium sulfate cuts) for 45 minutes at room temperature.
  • the FITC-IA k 68 - 83 peptide-BSA conjugate was then added for an additional 45 minutes at room temperature, the cells washed twice and analyzed.
  • the mean channel number is shown for the different cell lines incubated with the conjugates.
  • the % decrease in ean channel number compared with cells incubated with FITC-IA 68 _ 83 peptide- BSA alone is shown for each condition.
  • Methods of modulating mammalian T-cell response restricted by an MHC and methods of treating an MHC-linked disease in a mammal suspected of requiring such modulation or treatment are provided by the invention.
  • the methods comprise treating the mammal or contacting the T-cells respectively with an effective amount of a peptide, which peptide has an amino acid sequence substantially corresponding to at least a portion of the antigen recognition site of said
  • MHC or a peptide mimetic wherein said peptide or peptide mimetic is capable of binding with a T-cell antigen receptor which unbound T-cell antigen receptor is capable of recognizing said MHC bound to an antigen.
  • MHC-linked disease refers to those mammalian diseases where the relative risk for an individual expressing a particular MHC antigen to develop the disease is at least twice the risk of the population at large. Wherein the relative risk is computed from the following:
  • HLA antigens and diseases showing the most highly associated antigens in white populations.
  • peptide mimetic refers to any compound that functionally mimics the peptides described herein. That is, a peptide mimetic must be capable of binding with a T-cell antigen receptor which T-cell antigen receptor recognizes the MHC bound to an antigen, i.e. the T- cell antigen receptor is capable of binding with the MHC-Ag.
  • the T-cell antigen receptor is of the type that specifically binds the MHC-antigen fragment complex.
  • the "antigen recognition site" of the MHC refers to that portion of the MHC that is responsible for normal antigen presentation to the T-cell receptor. It is generally believed that the antigen binding site approximates a "groove” formed from two alpha helices lined on the bottom by ⁇ pleated sheets as described in Brown, J.H. et al., "A hypothetical model of the foreign antigen binding site of Class II histocompatibility molecules". Nature, 332:845-850 (28 April 1988) .
  • Peptides useful in this invention have an amino acid sequence which substantially corresponds to at least a portion of the antigen recognition site. It is only necessary that the peptide or peptide mimetic are capable of binding to the T-cell antigen receptor, which receptor in its unbound state, is capable of binding (recognizing) with an antigen-MHC complex.
  • the amino acid sequence of the peptide will preferably substantially correspond to at least a portion of the alpha helices of the antigen recognition site. Examples of methods to select peptides and peptide mimetics suitable for use in this invention are discussed below.
  • MHC antigens for the MHC-linked diseases shown in Table I have been characterized, i.e. the amino acid sequence of the MHC has been determined.
  • the known sequences are published and/or available from a variety of commercial data bases, such as GenBank.
  • a DNA molecule is synthesized which encodes a partial amino acid sequence of the MHC or which represents the complementary DNA strand to such a DNA molecule which encodes a partial amino acid sequence.
  • This synthetic DNA molecule may then be used to probe for DNA sequence homology in DNA sequences derived from the genomic DNA of the mammal or derived from cDNA copies of mRNA molecules isolated from cells or tissues of a mammal.
  • DNA molecules of fifteen (15) nucleotides or more are required for unique identification of an homologous DNA, said number requiring unique determination of at least five (5) amino acids in sequence.
  • the number of different DNA molecules which can encode the determined amino acid sequence may be very large since each amino acid may be encoded for by up to six (6) unique trinucleotide DNA sequences or codons. Therefore, it is impractical to test all possible synthetic DNA probes individually and pools of several such DNA molecules can be used concomitantly as probes.
  • the production of such pools which are referred to as "degenerate" probes is well known in the art. While only one DNA molecule in the probe mixture will have an exact sequence homology to the gene of interest, several of the synthetic DNA molecules in the pool may be capable of uniquely identifying the gene since only a high degree of homology is required. Therefore, successful isolation of the gene of interest may be accomplished with synthetic DNA probe pools which do not contain all possible DNA probe sequences. In fact, a single sequence DNA probe may be produced by including only the DNA codons most frequently utilized by the organism for each amino acid, although, it will be appreciated that this approach is not always successful.
  • PCR Polymerase Chain Reaction
  • a deoxythymidylate-tailed oligonucleotide is then used as a primer in order to reverse transcribe the RNA into cDNA.
  • a synthetic DNA molecule or mixture of synthetic DNA molecules as in the degenerate probe described above is then prepared which can encode the amino- terminal amino acid sequence of the MHC protein as previously determined. This DNA mixture is used together with the deoxythymidylate-tailed oligonucleotide to prime a PCR reaction. Because the synthetic DNA mixture used to prime the PCR reaction is specific to the desired mRNA sequence, only the desired cDNA will be effectively amplified. The resultant product represents an amplified cDNA which can be ligated to any of a number of known cloning vectors.
  • MHC peptides may exist in mammals which will have similar amino acid sequences and that in such cases, the use of mixed oligonucleotide primer sequences may result in the amplification of one or more of the related cDNAs encoding these related peptides.
  • the produced cDNA sequence can be cloned into an appropriate vector using conventional techniques, analyzed and the nucleotide base sequence determined. A direct amino acid translation of these PCR products will reveal that they corresponded to the complete coding sequence for the MHC protein.
  • sequence alignment As described in Brown, J.H. et al., "A hypothetical model of the foreign antigen binding site of Class II histo ⁇ ompatibility molecues". Nature , 332:845-850 (28 April 1988); or by determining the three- dimensional structure of the HLA molecule crystallographically as described in Bjorkman, P.J. , et al., "Structure of the human class I histocompatibility antigen, HLA-A2", Nature , 329:506-511, (8 October 1987) and Bjorkman, P.J.
  • the sequence alignment method is preferred. Once the MHC antigen sequence is known, the MHC sequence can then be aligned for maximal homology, as taught in Brown et al., with HLA-A2 (or other crystallographically known HLA antigen) sequence.
  • the sequences which correspond to the antigen recognition site are those which comprise the alpha helicies described in Brown et al., supra (1988). This are the helices lining the groove, and includes amino acid residues 60-86 and 140-174 of the HLA-A2 allele, and those sequences from other HLA types which align with these sequences as described in Brown et al., supra (1988).
  • the antigen recognition site of the MHC of interest is identified, at least a portion of the amino acid sequence of the site can be selected for its suitability for use in the method of the invention. It is expected that peptides substantially corresponding to the alpha helices will be particularly useful. For example, the entire sequence of one helix of the recognition site can be employed for testing (eg. residues 60-86 of HLA-A2) . Shorter, overlapping peptides for the entire recognition site can be synthesized for testing (eg. HLA-A260-70, 65-75, 70-80, etc.).
  • Regions of particular interest can be synthesized for testing, for example a region in the HLA DR4 ⁇ chain (Brown et al., supra (1988)) which is associated with rheumatoid arthritis, while the QK residues are invariably absent in non-rheumatoid arthritis associated alleles.
  • a region in the HLA DR4 ⁇ chain (Brown et al., supra (1988)) which is associated with rheumatoid arthritis, while the QK residues are invariably absent in non-rheumatoid arthritis associated alleles.
  • peptides or peptide mimetics as described herein bind to the TCR and inhibihit the cascade of host defense actions triggered by the formation of the tertiary TCR-Ag-MHC complex.
  • the peptides useful in the methods of this invention can be prepared synthetically or recombinantly by ways known to those in the art.
  • Peptides or peptide mimetics suitable for use in this invention can be screened for their ability to bind with a T-cell antigen receptor which T-cell antigen receptor recognizes the MHC bound to an antigen by any method known to those in the art.
  • Standard immunological assays for such binding include: binding by flow microfluorimetry to relevant cell lines; tritiated thymidine incorporation assays or similar assays to measure T-cell proliferation in the presence of the peptides; release of cytokines (such as interleukins) as determined by immunoassay or biological response assays (such as proliferation of cytokine dependent cell lines to the cytokines) in the presence of the peptides; Chromium-51 release or similar assays to measure cytotoxic T-cell activity; direct binding to T-cell receptors by standard ligand-binding assays or by competition; inhibition or stimulation of T-cell activation and/or growth; binding to MHC haplotype-specific antibodies.
  • Other screening methods also are believed useful such as influencing the course of an experimental model of an autoimmune disease in vivo or in vitro.
  • peptides that are suitable for use in this invention can be as short as two amino acids in length or the alpha helices which is generally expected to be about 60 amino acids in length.
  • the peptides and peptide mimetics can be formulated into a pharmological composition containing an effective amount of the peptide in a usual nontoxic carrier. See e.g. Gennaro, A., .Remington's Pharmaceutical Sciences, 17th edition, Mack Publishing Co., Easton, PA (1985).
  • the composition can be administered via a route suited to the form of the composition.
  • compositions are, for example, in the form of usual liquid preparations including solution, suspension, emulsion and the like, which are generally given orally, intravenously, subcutaneously, intramuscularly or topically.
  • the composition can also be provided as a dry preparation which can be reconstructed to a liquid for use by addition of a suitable liquid carrier.
  • the amount of the composition to be administered will vary with the age and sex of the patient, the type and severity of the MHC-linked disease, etc.
  • An effective amount of the peptide or peptide mimetic is that amount capable of treating an MHC-linked disease or that amount capable of modulating T-cell response to an MHC in an animal. It is expected that the composition will be administered at doses of about 0.01 to about 5000 mg/kg/day, calculated as protein, preferably in divided doses.
  • Peptides were purified by passage over sephadex G25 columns, or by HPLC on a TSK 3000 column (Waters) in 50% acetonitrile 50% water in an isocractic run. Peptides were lyophilized prior to use. For cell culture, all peptides were sterilized by irradiation with 10,000 rads (Cobalt source) prior to use. Peptides utilized are shown in Table 1.
  • Peptide-BSA conjugates were fluorsceinated as follows. Fluorscein isothiocyanate (FITC) (Sigma, St. Louis, MO), was dissolved at 1 mg/ml in 0.1 M Na 2 C0 3 . To this solution lyophilized peptide-BSA conjugate was added at a final concentration of 4 mg/ml. The solution was stirred at 4°C overnight and dialyzed against phosphate buffered saline (PBS) prior to use.
  • FITC Fluorscein isothiocyanate
  • PBS phosphate buffered saline
  • mice AKR female mice aged 6-8 weeks were obtained from the National Cancer Institute (Bethesda, MD) and were maintained in accordance with the National Institutes of Health and University of Pennsylvania guidelines.
  • D10.G4 cells were obtained from The American Type Culture Collection (ATCC) and grown in RPMI 1640 withaddedpenicillin/streptomycin, L-glutamine, non-essential amino acids, sodium pyruvate, 5xl ⁇ "5 M /3-mercaptoethanol, (all from GIBCO) and 10% fetal calf serum (FCS) (Hyclone) .
  • Conalbumin was purchased from Sigma (St. Louis, MO) . Cells were passaged at 5xl0 4 /n_l with antigen presenting cells (APCs) (2500 R irradiated AKR spleen cells) at 5X10 5 /_D,1 and conalbumin at 200 ⁇ g/ml.
  • APCs antigen presenting cells
  • Antibodies The following monoclonal antibodies were utilized: 15-1-5P anti-H-2K ⁇ D ⁇ (murine lgG2b) and 10.2.16 anti-IA (murine lgG2b) (both from the American Type Culture Collection, Rockville, MD (ATCC) ; 3D3 anti-D10.G4 clonotype (murine lgGl) , J. Kaye et al. , "Both a monoclonal antibody and antisera specific for determinants unique to individual to cloned helper T-cell lines can substitute for antigen and antigen-presenting cells in the activation of T-cells," J. Exp. Med. 158:836-856 (1983); J.M.
  • Hybridomas were grown in culture media and supernatants filter sterilized prior to use. Some antibodies were further subjected to ammonium sulfate precipitation and dialysis against phosphate buffered saline (PBS) prior to filter sterilization and use, W. Williams et al., supra (1988a) . Radioimmunoassay: This was as previously described, W.V. Williams et al., supra (1989).
  • Peptides were suspended in distilled water at varying concentrations and 50 ⁇ l/well evaporated onto 96 well V bottom plates (Dynatech Labs) .
  • the wells were washed in PBS, blocked with FACS buffer (1% BSA in PBS with 0.1% sodium azide) , and antibodies added at varying dilutions in FACS buffer, 50 ⁇ l/well.
  • Antibodies were incubated overnight at 4°C, the wells washed with PBS, and 1 5I-goat anti-mouse added, 50,000-100,000 counts per minute (CPM) per well, and incubated for >1 hour at 37"C or overnight at 4 ⁇ C.
  • the wells were washed lOx in tap water, cut out, and counted.
  • D10.G4 cells (10 4 /well) with 2500 rad irradiated AKR spleen cells (see figures for dosages) were cultured for 72 hours with various stimuli.
  • the wells were then pulsed with tritiated thymidine (l ⁇ Ci/well) for an additional 18 hours, the cells harvested onto glass fiber filters, and counted in a standard liquid scintillation system.
  • FACS Analysis This was as previously described, W. Williams et al., supra (1988a). Briefly, cells were resuspended at 10 7 / ⁇ l in FACS buffer and for D10.G4 cells, preincubated with peptides, conjugates or antibodies for 30-60 minutes at 23 ⁇ C. For IA expressing L cells, antibodies were preincubated with peptides at 23*C for 30-60 " minutes prior to addition of cells. Antibodies or FITC-peptide-BSA conjugates and cells were combined, and incubated for 20 minutes at 4'C. The cells were resuspended in 500 ul FACS buffer, spun down and washed prior to addition of secondary antibody (where indicated) .
  • IJI C goat anti-mouse lg (Fisher) was added for 20 minutes at 4"C, the cells washed twice, and analyzed as described, W. Williams et al., supra (1988).
  • Antibodies were utilized as follows: 15-1-5P, 10.2.16, 3D3, 500A2, and 145-2C11 were prepared as ammonium sulfate cuts of culture supernatant, and were utilized at a 1:50 dilution. C193.5 was utilized as culture supernatant undiluted.
  • the peptides utilized in this study are shown in Table I.
  • the IA 68 . 83 peptide corresponds to a region predicted to be an alpha helix lining the Ag binding groove of the IA molecule. This site contains polymorphic residues potentially involved in recognition by haplotype-specific antibodies directed to the IA K molecule, J.H. Brown et al., "A hypothetical model of the foreign antigen binding site of class II histocompatibility molecules," Nature 332:845-850 (1988) .
  • the control peptide (designated 1S1) was designed to have an identical net charge and hydrophobicity as the IA peptide. Amino terminal cysteine residues were added to each sequence to allow dimerization of the peptides, thereby increasing their avidity for various receptor structures.
  • IA ⁇ . ⁇ peptide The ability of IA ⁇ . ⁇ peptide to mimic a portion of the intact IA molecule suggests that this peptide might also interact with the TCR on IA restricted T lymphocytes.
  • T-cell clone D10.G4 was utilized, a murine TH 2 clone which responds to IA K + conalbumin, J. Kaye et al., supra (1983); J. Kaye et al., "Growth of a cloned helper T-cell line induced by a monoclonal antibody specific for the antigen receptor: interleukin 1 is required for the expression of receptors for interleukin 2.," J. Immunol . 133:1339-1345 (1984).
  • peptide-BSA conjugates were fluorsceinated, and the resultant complexes were utilized to stain D10.G4 cells as well as 22.D11 cells (a murine T-cell hybridoma specific for pigeon cytochrome c in the context of I-E ⁇ ) . These cell lines were incubated with the different fluorsceinated conjugates, washed and analyzed for fluorscence intensity ( Figure 7) .
  • the IA 68 - 83 peptide displayed biological effects in blocking D10.G4 activation in response to conalbumin + IA K ( Figure 3&4) .
  • the inhibition of activation seen in these experiments was of interest when compared with results utilizing the peptides without the addition of specific antigen. It is noteworthy that the peptides utilized bore an amino terminal cysteine residue, which should result in the formation of dimeric peptides.

Abstract

Procédés de modulation de la réaction de lymphocytes T de mammifères limitée par un complexe majeur d'histocompatibilité et procédés de traitement d'une maladie liée au complexe majeur d'histocompatibilité chez un mammifère dont on pense qu'il nécessite une telle modulation ou un tel traitement. Les procédés consistent à traiter le mammifère ou à mettre en contact les lymphocytes T respectivement avec une dose efficace d'un peptide, lequel peptide a une séquence d'acides aminés correspondant à au moins une partie du site de reconnaissance d'antigènes dudit complexe majeur d'histocompatibilité, ou un peptide mimétique, ledit peptide ou peptide mimétique est capable de se lier à un récepteur d'antigènes de lymphocytes T, lequel récepteur d'antigènes de lymphocytes T non liés est capable de reconnaître ledit complexe d'histocompatibilité lié à un antigène.
PCT/US1992/002419 1991-03-22 1992-03-20 Procede de modulation de la reaction de lymphocytes t de mammiferes WO1992016234A1 (fr)

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WO1996036881A2 (fr) * 1995-05-16 1996-11-21 Cancer Research Campaign Technology Limited Evaluation preliminaire d'inhibiteurs potentiels des interactions entre les recepteurs des lymphocytes t (rlt) et le complexe majeur d'histocompatibilite (cmh)
EP0759771A1 (fr) * 1995-03-07 1997-03-05 President And Fellows Of Harvard College Identification des auto-antigenes et des non auto-antigenes intervenant dans les affections auto-immunes
EP0828503A1 (fr) * 1995-05-12 1998-03-18 Sangstat Medical Corporation Traitement destine a inhiber la progression de maladies auto-immunes
EP0837691A1 (fr) * 1995-06-07 1998-04-29 Anergen Incorporated Vaccination par peptides de molecules du cmh de classe ii destinee au traitement de maladie auto-immune
US6509165B1 (en) 1994-07-08 2003-01-21 Trustees Of Dartmouth College Detection methods for type I diabetes
US7084247B2 (en) 2004-03-11 2006-08-01 Peptimmune, Inc. Identification of self and non-self antigens implicated in autoimmune diseases

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EP0674526A1 (fr) * 1992-12-17 1995-10-04 SRIRAM, Subramaniam Vaccination a l'aide d'un peptide de molecules de classe ii du complexe majeur d'histocompatibilite (mhc) pour le au traitement de maladies autoimmunes
US6045796A (en) * 1992-12-17 2000-04-04 Anergen, Inc. Vaccination with peptide of MHC class II molecules for treatment of autoimmune disease
EP0674526A4 (fr) * 1992-12-17 1997-01-15 Subramaniam Sriram Vaccination a l'aide d'un peptide de molecules de classe ii du complexe majeur d'histocompatibilite (mhc) pour le au traitement de maladies autoimmunes.
US6509165B1 (en) 1994-07-08 2003-01-21 Trustees Of Dartmouth College Detection methods for type I diabetes
EP0759771A4 (fr) * 1995-03-07 1999-12-29 Harvard College Identification des auto-antigenes et des non auto-antigenes intervenant dans les affections auto-immunes
EP0759771A1 (fr) * 1995-03-07 1997-03-05 President And Fellows Of Harvard College Identification des auto-antigenes et des non auto-antigenes intervenant dans les affections auto-immunes
US7255861B1 (en) 1995-03-07 2007-08-14 President And Fellows Of Harvard College Preparations for inducing immunotolerance and uses therefor
EP0828503A1 (fr) * 1995-05-12 1998-03-18 Sangstat Medical Corporation Traitement destine a inhiber la progression de maladies auto-immunes
EP0828503A4 (fr) * 1995-05-12 1999-10-13 Sangstat Medical Corp Traitement destine a inhiber la progression de maladies auto-immunes
WO1996036881A3 (fr) * 1995-05-16 1997-01-09 Cancer Res Campaign Tech Evaluation preliminaire d'inhibiteurs potentiels des interactions entre les recepteurs des lymphocytes t (rlt) et le complexe majeur d'histocompatibilite (cmh)
WO1996036881A2 (fr) * 1995-05-16 1996-11-21 Cancer Research Campaign Technology Limited Evaluation preliminaire d'inhibiteurs potentiels des interactions entre les recepteurs des lymphocytes t (rlt) et le complexe majeur d'histocompatibilite (cmh)
EP0837691A1 (fr) * 1995-06-07 1998-04-29 Anergen Incorporated Vaccination par peptides de molecules du cmh de classe ii destinee au traitement de maladie auto-immune
EP0837691A4 (fr) * 1995-06-07 1999-12-15 Anergen Inc Vaccination par peptides de molecules du cmh de classe ii destinee au traitement de maladie auto-immune
US7084247B2 (en) 2004-03-11 2006-08-01 Peptimmune, Inc. Identification of self and non-self antigens implicated in autoimmune diseases

Also Published As

Publication number Publication date
EP0579747A1 (fr) 1994-01-26
CA2106567A1 (fr) 1992-09-23
AU669329B2 (en) 1996-06-06
EP0579747A4 (en) 1994-07-06
AU1795092A (en) 1992-10-21
JPH06507630A (ja) 1994-09-01

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