WO1996022067A2 - Banques de peptides a encliquetage pour vaccins et therapeutiques induisant des lymphocytes t cytotoxiques - Google Patents

Banques de peptides a encliquetage pour vaccins et therapeutiques induisant des lymphocytes t cytotoxiques Download PDF

Info

Publication number
WO1996022067A2
WO1996022067A2 PCT/US1995/016290 US9516290W WO9622067A2 WO 1996022067 A2 WO1996022067 A2 WO 1996022067A2 US 9516290 W US9516290 W US 9516290W WO 9622067 A2 WO9622067 A2 WO 9622067A2
Authority
WO
WIPO (PCT)
Prior art keywords
library
ratchet
ctl
amino acids
peptide
Prior art date
Application number
PCT/US1995/016290
Other languages
English (en)
Other versions
WO1996022067A3 (fr
Inventor
Peter J. Kuebler
Douglas F. Nixon
Original Assignee
United Biomedical, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by United Biomedical, Inc. filed Critical United Biomedical, Inc.
Priority to AU58497/96A priority Critical patent/AU5849796A/en
Publication of WO1996022067A2 publication Critical patent/WO1996022067A2/fr
Publication of WO1996022067A3 publication Critical patent/WO1996022067A3/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/82—Translation products from oncogenes
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
    • C07K1/04—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length on carriers
    • C07K1/047—Simultaneous synthesis of different peptide species; Peptide libraries
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/005—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
    • 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
    • C07K14/44—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from protozoa
    • C07K14/445—Plasmodium
    • 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
    • C07K14/4701—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals not used
    • C07K14/4727—Mucins, e.g. human intestinal mucin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/16011—Human Immunodeficiency Virus, HIV
    • C12N2740/16111—Human Immunodeficiency Virus, HIV concerning HIV env
    • C12N2740/16122—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
    • C12N2760/00011—Details
    • C12N2760/16011—Orthomyxoviridae
    • C12N2760/16111—Influenzavirus A, i.e. influenza A virus
    • C12N2760/16122—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • Ratchet libraries composed of related peptides synthesized simultaneously in a single peptide synthesis.
  • Ratchet libraries are derived from a longer template peptide by sequentially "ratcheting" the template sequence into the shorter ratchet length and are used for cytotoxic T lymphocyte (CTL) induction or stimulation if the CTL epitope is known. If the CTL epitope is unknown, then the ratchet libary can be used for identification of CTL epitopes.
  • CTL cytotoxic T lymphocyte
  • the ratchet libraries can be prepared from any protein sequence to which an immune CTL response is desired and can be formulated for delivery as a vaccine or therapeutic for the treatment or prevention of disease or malignancy.
  • a ratchet library can be used in the prevention and treatment of infectious or malignant diseases including HIV, influenza, malaria, breast, ovarian, lung and colon cancers.
  • CTL cytotoxic T lymphocytes
  • CTL are a vital component of the natural immune response against infectious organisms and malignant cells.
  • CTL are CD8' thymus derived lymphocytes which appear early in an immune response and help in the elimination of, for example, virus-infected cells or tumor cells by lysis of the target cells and by secretion of chemical immunomodulators termed cytokines, such as interferons.
  • CTL have been detected following many viral infections, including HIV infection, and extensive evidence points to a major role for CTL in control of virus infections [McMichael et al. (1983) New Eng. J. Med. 301:13; Nixon et al. (1992) Immunology 76:515).
  • adoptive transfer of specific CTL to influenza [Taylor et al. (1986) Immunology 58:417] or para yxovirus simian virus 5-infected mice cleared the virus from the lungs [Young et al. (1990) J. Virol. 64:5403].
  • Tumor specific CTL have also been shown to clear tumors caused by mouse retroviruses [Cerundolo et al.
  • CTL-inducing vaccine An essential step in the design of a CTL-inducing vaccine is in the identification of the antigenic sites to which CTL react.
  • CTL recognize infected or malignant cells through the interaction of their specific T-cell receptor with a complex displayed on the surface of the target cell.
  • the complex consists of an antigenic peptide specific to the virus or tumor, for example, and a major histocompatibility complex (MHC) class I molecule encoded by the Class I MHC genes of the host [Townsend et al. (1986) Cell 44:959].
  • MHC major histocompatibility complex
  • Clusters of closely linked MHC alleles are characteristically inherited as a genetic unit termed the "haplotype".
  • haplotype Clusters of closely linked MHC alleles are characteristically inherited as a genetic unit termed the "haplotype".
  • individual MHC molecules associate with and present different antigenic protein fragments, so that one fragment of an antigenic protein is recognized by CTL of a specific MHC haplotype, while a different MHC haplotype requires another fragment of the antigen for recognition, i.e., recognition of individual antigenic fragments is MHC-restricted.
  • recognition of individual antigenic fragments is MHC-restricted.
  • the MHC alleles are highly polymorphic between diverse genetic groups, a large number of distinct peptides may be needed to insure CTL stimulation across diverse human populations.
  • CTL epitope The exact fragment(s) of a virus or tumor antigen or other potential antigenic site (i.e., CTL epitope) recognized by a specific CTL was thought to be between 7 and 25 amino acids, but recent characterization of viral peptides naturally processed in virus-infected cells and displayed by Class I MHC molecules have identified the CTL epitopes as peptides of between 7 to 11 amino acids in length [R ⁇ tzschke et al. (1991) Immunology Today 12:447] with the majority of these peptides being of 9 amino acids (nonomers) . Identification of CTL epitopes in a protein sequence has been achieved by using synthetic peptides to map i munogenic sites.
  • CTL epitopes have been defined from HIV through an in vitro testing process of the human immune response to HIV infection [Nixon et al. (1988) Nature 336:484-487; Nixon et al. U.K. Patents GB 2,255,093, 2,273,709, 2,273,710]. While many HIV CTL epitopes have been identified in animals, few have been identifed in humans. However, because CTL epitopes are simultaneously recognized by a T-cell receptor that is specific for both the virally-encoded peptide and the host-encoded MHC for clearance of an infected cell to occur, CTL-epitopes are species specific.
  • human CTL epitopes may not be reliably predicted from animal studies. While vaccine development has led to successful vaccine against many infectious diseases, (e.g. polio, measles) , there are several important pathogens for which vaccines are either ineffective or simply non-existant, for example HIV, hepatitis C virus (HCV) and herpes simplex virus (HSV) . Moreover, there are no vaccines for treatment of malignancies. The identification of CTL epitopes makes it feasible to design CTL-stimulating vaccines and other immunotherapeutics for prevention or treatment of disease by the clearance of virally-infected cells or malignant cancer cells.
  • infectious diseases e.g. polio, measles
  • HCV hepatitis C virus
  • HSV herpes simplex virus
  • the EBNA 1 protein of Epstein-Barr virus (EBV) .
  • the present invention provides a solution to this problem because the ratchet libraries can encompass extensive CTL antigenic regions and eliminate the need to precisely map CTL epitopes, or even to map the CTL epitopes at all.
  • the ratchet libraries can be used to map antigenic sites.
  • CTL responses can be primed by administration of lipid-derivatized peptides [Deres et al.
  • ratchet library peptides can be formulated into an appropriate vehicle to elicit CTL responses in vivo.
  • the ratchet library peptides provide a major advance since several epitopes can be incorporated into the ratchet libraries rather than relying upon mixtures individually synthesized immunogenic peptides.
  • Antigenic variation is a recurrent problem among certain pathogens contributing to unsuccessful or limited success of vaccines.
  • Extensive antigenic variation for example, is a hallmark of HIV (AIDS) , rhinovirus (the common cold) , influenza virus (flu) , plasmodium falciparum (malaria) .
  • HIV HIV
  • rhinovirus the common cold
  • influenza virus flu
  • plasmodium falciparum plasmodium falciparum
  • Ratchet libraries can be constructed to embody known antigenic variation and escape mutations to pre-empt these problems.
  • the ratchet library method of CTL induction provides a solution to obstacles in the development of vaccines and therapeutics for such pathogens or cancers.
  • This invention is directed to a ratchet library of peptides comprising at least one immunostimulatory cytotoxic T lymphocyte (CTL) epitope.
  • the peptides are of length 1.
  • the sequences of the peptides in the library are determined from a template peptide of length from 1+1 to n amino acids such that each position x in the library has all the amino acids present in the template peptide at positions x to n-(l-x), inclusive and the ratio of amino acids at each position x is determined by the relative prevalence of amino acids at that position x.
  • 1 is from about 7 to about 25 amino acids, preferably 8-10 and more preferably 9; n is from 1+1 to about 100, preferably from 1+1 to about 75 and more preferably from 1+1 to about 50; and x is from 1 to 1. Moreover, if a position x is identified as part of an MHC-binding motif of a CTL epitope, then that position x in the ratchet library is fixed as one or more amino acids of the MHC-binding motif in an equimolar ratio.
  • the CTL epitopes of this invention are from a virus, bacterium, parasite, tumor antigen, allergen or other protein antigen.
  • the ratchet library can be constructed from the template peptides of any one of SEQ ID NOS: 1-11.
  • the peptides can have a covalently attached N-terminal tripalmitoyl-5-glycerol-cysteine moiety or be linked to a branched core sequence, polymerized or conjugated to a carrier molecule.
  • Another aspect of this invention provides a pharmaceutical or vaccine composition comprising the subject ratchet libraries including emulsion or a microparticle formulation with or without the addition of free Pam 3 Cys or a derivative thereof.
  • compositions are useful in a method of treating or preventing a disease or a malignancy which comprises administering an amount of the ratchet library as a vaccine or pharmaceutical composition to a mammal effective to stimulate a CTL response against the disease or the malignancy associated with the CTL epitope present in the library.
  • Still another aspect of the invention is directed to a method of constructing a library of related peptides to provide a ratchet library which comprises identifying a template peptide; calculating a distribution of amino acids at each position x having those amino acids present in the template peptide at positions x to n-(l-x), inclusive, wherein 1 is from about 7 to about 25, n is from 1+1 to about 100, and x is from 1 to 1; and synthesizing said ratchet library.
  • Fig. 1 depicts malaria ratchet libraries 1 and 2 from Plasmodium berghei circumsporozoite (CS) protein and their construction.
  • Fig IA shows the template peptide with the known CTL epitope (CS 252-260) indicated by a box. Below the template peptide is the corresponding set of overlapping nonmer peptides.
  • Fig. IB provides an example of a sequence alignment for malaria ratchet library 1 (top panel) , its corresponding amino acid (AA) distribution by position (middle panel) and the percent of each amino acid at each position in the library (bottom panel) .
  • Fig. 2 is a graphic illustration of malarial-speci ic CTL activity against CS 252-260.
  • the graph shows the percent specific cell lysis as a function of the effector to target cell (E:T) ratio in mice immunized with 100 ⁇ g doses of malaria ratchet library 1 in microparticles.
  • Fig. 3 is a graphic illustration of malarial-specific CTL activity against CS 252-260.
  • the graph shows the percent specific cell lysis as a function of E:T ratio in mice immunized with 1 mg doses of malaria ratchet library 2 in microparticles.
  • Fig. 4 is a graphic illustration of malarial-specific CTL activity against CS 247-266.
  • the graph shows the percent specific cell lysis as a function of E:T ratio in mice immunized with 1 mg doses of malaria ratchet library 2 in microparticles.
  • Fig. 3 is a graphic illustration of malarial-specific CTL activity against CS 252-260.
  • the graph shows the percent specific cell lysis as a function of E:T ratio in mice immunized with 1 mg doses of
  • FIG. 5 is a graphic illustration of malarial-specific CTL activity against CS 252-260.
  • the graph shows the percent specific cell lysis as a function of E:T ratio in mice immunized with 10 ⁇ g doses of malaria ratchet library 1 as lipopeptides.
  • Fig. 6 is a graphic illustration of the lack of malarial-specific CTL activity against a self peptide.
  • the graph shows the percent specific cell lysis as a function of E:T ratio in mice immunized with 100 ⁇ g doses of malaria ratchet library 1 as lipopeptides.
  • Fig. 7 depicts an MHC-restricted malaria ratchet library constructed from malarial ratchet library 1 (top panel) .
  • the middle panel shows the known anchor residues for four MHC haplotypes, K d , D b , K b and L d . These anchor residues are at positions 2 for K d and L d , 5 for D b , K b and L d , 8 for K b and 9 for ⁇ D b and L d .
  • the bottom panel shows the resulting MHC-restricted malaria ratchet library.
  • Fig. 8 depicts an HIV ratchet library from a 35 amino acid sequence of the HIV-1 gpl20 V3 loop region (residues 305-339) .
  • Fig. 8A provides the sequence of 15 HIV-1 variants from this region (top panel) and the corresponding SSAL from those sequences.
  • Fig. 8B provides the amino acid (AA) distribution by position (middle panel) and the percent of each amino acid at each position in the library (bottom panel) in a ratchet library constructed from the SSAL. Amino acids divergent from the consensus B sequence are shown as upper case letters and conserved amino acids in the consensus sequences are shown as lower case letters.
  • Fig. 9 depicts an HIV-1 gag peptide linear ratchet library containing a mouse HIV CTL epitope prepared from a 100 amino acid template (top panel) .
  • Fig. 10 depicts an HIV-1 gag peptide linear ratchet library containing a mouse HIV CTL epitope prepared from a 40 amino acid template (top panel) .
  • the amino acid (AA) distribution by position (middle panel) and the percent of each amino acid at each position in the library (bottom panel) is shown.
  • the D h restricted epitope (at gag residues 390-398) is indicated by the box.
  • FIG. 11 depicts an HIV-1 gag peptide linear ratchet library containing a mouse HIV CTL epitope prepared from a 20 amino acid template (top panel) .
  • the amino acid (AA) distribution by position (middle panel) and the percent of each amino acid at each position in the library (bottom panel) is shown.
  • the D h restricted epitope (at gag residues 390-398) is indicated by the box.
  • Fig. 12 is a graphic illustration of HIV-specific CTL activity against HIV-1 gag residues 390-398. The graph shows the percent specific cell lysis as a function of E:T ratio in mice immunized with 100 ⁇ g doses of the HIV ratchet library from the 40-mer template in an emulsion.
  • Fig. 12 is a graphic illustration of HIV-specific CTL activity against HIV-1 gag residues 390-398. The graph shows the percent specific cell lysis as a function of E:T ratio in mice immunized with 100 ⁇ g doses of the HIV ratchet library
  • FIG. 13 depicts a mucin ratchet library from a 20-mer repeating sequence (top line) .
  • the next two lines illustrate two alternate template peptides for construction of this mucin ratchet library.
  • the boxed residues are the additional sequences added at the termini to allow representation of all possible nonomers of the 20 amino acid repeat sequence.
  • the amino acid (AA) distribution by position (middle panel) and the percent of each amino acid at each position in the library (bottom panel) is shown.
  • Fig. 14 depicts a mutant p53 ratchet library constructed from a template peptide of amino acids 124-151 (top panel) . There are additional amino acids, which represent known p53 mutants incorporated at positions 9- 12.
  • the boxed residues are 10 amino acid CTL epitope identified in Balb/C mice.
  • the amino acid (AA) distribution by position (middle panel) and the percent of each amino acid at each position in the library (bottom panel) is shown.
  • Fig. 15 depicts influenza ratchet library 1 constructed from a 25-mer template sequence of residues 139-163 of influenza A A/34/PR8 nucleoprotein (top panel) .
  • the known K d -restricted epitope of residues 147-155 is indicated by a box.
  • the amino acid (AA) distribution by position (middle panel) and the percent of each amino acid at each position in the library (bottom panel) is shown.
  • FIG. 16 depicts influenza ratchet library 2 from a template peptide which is a linkage of 3 CTL epitopes in order from N to C terminus of residues 50-58, 147-155 and 366-374 of the nucleoprotein.
  • Each CT1 epitope is indicated by a box.
  • the amino acid (AA) distribution by position (middle panel) and the percent of each amino acid at each position in the library (bottom panel) is shown.
  • the present invention is directed to a ratchet library of peptides comprising at least one immunostimulatory cytotoxic T lymphocyte (CTL) epitope.
  • CTL cytotoxic T lymphocyte
  • the peptides of the ratchet library are of length 1 and the sequences of the peptides in the library are determined from a template peptide having a length from 1+1 to n amino acids.
  • Each position x in the library peptides has those amino acids which are present in the template peptide at positions x to n-(l-x), inclusive. Accordingly, the ratio of the individual amino acids at each position x is determined from the relative numbers (or prevalence) of the amino acids at that position x.
  • 1 is from about 7 to about 25
  • n is from 1+1 to about 100
  • x is from 1 to 1.
  • position 1 contains all the amino acids of the template peptide at positions 1 to n-(x-l)
  • position 2 contains all the amino acids of the template peptide at positions 2 to n-(x-2)
  • position 3 contains all the amino acids of the template peptide at positions 3 to n-(x-3)
  • position x contains all the amino acids of the template peptide at positions x to n.
  • a malaria ratchet library showing the relative ratio of amino acids at each position in the ratchet library derived from a longer malaria template peptide as well as the percentage of amino acids required for synthesis at each position of the ratchet library. More specifically, the malaria template peptide is divided up into sequential 9-mers which are aligned at the amino terminus of the template peptide to create a set of overlapping peptides from which to calculate the ratchet library composition. After calculation of its composition, the ratchet library is prepared in a single synthesis based on the calculated amino acid distributions at each position. The size of the ratchet 1, or ratchet length, can be determined from the actual or approximate size of the target CTL epitope.
  • CTL epitopes have been identified which vary in length from 7 to 25 residues. However, the majority of CTL peptides are from 8-10 amino acids, and many are 9 amino acids. While the actual size of the CTL inducing peptide is preferred to determine the length 1 of the ratchet library, e.g. 9 amino acids, the ratchet length can be determined by other means, including an arbitrary selection of size within the range of 7 to 25 amino acids.
  • the size of the template peptide ranges from 1+1 to about 100 amino acids, and preferably from 1+1 to about 75 amino acids, and more preferably from 1+1 to about 50.
  • the template peptide length is an important factor in determining the overall complexity of the ratchet library, so that shorter template peptides tend to yield less complex ratchet libraries, i.e., have fewer peptides in the library. If the CTL epitope is known, then the template peptide should be of a length n such that the CTL epitope is flanked by sufficient adjoining sequences, preferably at least 1-1, to insure that the CTL epitope is represented in the ratchet library. If the CTL epitope is not known, then the template peptide can have a length which covers a significant region of the protein being tested.
  • a length can range from about 20 to about 100 residues, and preferably ranges up to 50 or 75 residues.
  • the template peptide can also be selected on the basis of clustering of epitopes, of hydrophobicity, of stretches containing basic amino acids or of another biological characteristic. Selection of a longer template peptide is useful in identifying unknown CTL epitopes.
  • MHC binding motifs have identified particular amino acids residues within CTL epitopes which are important in peptide binding to the MHC recepter. When the MHC binding motif is known for a particular CTL epitope, then the ratchet library can be simplified by replacing the calculated distribution of amino acids at a particular site with the ratio of known amino acids from the MHC binding motif at that site.
  • human HLA binding motifs for 9-mer or 10-mer peptides typically have the designated amino acids at the indicated positions: for HLA-A2, leucine at position 2, valine or leucine at the C terminus; for HLA-B35, proline at postion 2, tyrosine at the C terminus; for HLA-B53, proline at position 2, phenylalanine or tryptophan at the C terminus; for HLA-B8, lysine at position 3, lysine at position 5, isoleucine at the C terminus; for HLA-B27, arginine at position 2, lysine or arginine at the C terminus; for HLA-B7, alanine at position 1, proline at position 2, arginine at position 3, leucine or valine at the C terminus; for HLA-A68, thre
  • the template peptide length n can be equal to the length of the CTL epitope plus 1-1 residues where the 1-1 residues carboxy-terminal amino acids of the epitope are placed at its amino -_ strict, repeat PCT/US95/16290 22067
  • the 1-1 amino-terminal residues of the epitope are placed at its carboxyl terminus, for example as shown in Fig. 5 for the mucin ratchet library.
  • the ratchet library can be constructed from a template peptide which is itself a "structured synthetic antigen library" or SSAL.
  • SSALs are defined and exemplified in U.S. Serial No. 08/143,412, filed October 26, 1993, which is incorporated herein by reference. Briefly, the sequence of an SSAL is determined by aligning the primary amino acid sequences of a related family of CTL epitopes and identifying the invariant and variant loci within the alignment.
  • the invariant loci generally represent the structural framework of the SSAL.
  • the degeneracy within the SSAL is determined by the loci within the alignment that harbor different amino acid residue types relative to an arbitrary prototype sequence. After determining which amino acids are to be at each position, the degree of degeneracy for the multiresidue position in the SSAL library is determined from the number of variants each individual amino acid represents by one of three methods. Thus in a simple manner, the specific amino acids and their frequency of appearance at each position within the SSAL is defined by the primary sequences of the different CTL antigens or molecules in the alignment of multiple primary sequences.
  • the degeneracies for the variant amino acid positions used for an SSAL can be determined in one of three ways.
  • the identity and ratio of residues is determined by the relative prevalence of the amino acids in a compilation of known sequences for the epitope.
  • identity of the amino acids at the variant position is determined from the compilation of known sequences for the epitope but the ratio of amino acids is set to be equimolar.
  • identity and 1 ratio of amino acids at a variant position can be
  • the ratchet libraries can be prepared as a ensemble
  • 29 branched core sequences conjugated to a carrier or 30 polymerized.
  • branched cores e.g. poly-L-lysine
  • 34 can be composed of an amino acid or an amino acid analog
  • amino acids are lysine or a lysine analog such as ornithine.
  • the amino acid analog can be an ⁇ -amino acid, a ⁇ -amino acid, or any other either natural or non-natural amino acid with two amino groups and one carboxyl group available for forming peptide bonds.
  • Preferred branched peptides of the invention are dimers, tetramers and octamers, especially those having a branching core structure composed of lysine such as a heptalysine core.
  • the branched cores can contain other residues interspersed among the branching residues as depicted, for example, in Fig. 12 of U.S. Serial No. 143,412.
  • the library can have a C-terminal methionine as the residue that is attached to the branched core.
  • the methionine provides a cleavable site to facilitate analysis of the ratchet library.
  • the ratchet can have one or more lysine residues (added at the amino terminus) to increase peptide solubility, cysteine and haloacylated residues can be added to facilitate directed coupling to carrier molecules, and methionine can be added for cyanogen bromide cleavage if necessary.
  • Pani j Cys, or a similar lipid tail, can be added to create a lipopeptide.
  • the subject ratchet libraries can also be used to form conjugates, i.e., the ratchet library, either in branched or linear form, can be coupled directly or indirectly, by methods known in the art, to carriers such as bovine serum albumin (BSA) , human serum albumin (HSA) , or other proteins, red blood cells or latex particles.
  • BSA bovine serum albumin
  • HSA human serum albumin
  • a ratchet library can be polymerized to homo- or hetero-di ers or higher oligomers by cysteine oxidation, by induced disulfide cross-linking, or by use of homo- or hetero-functional multivalent cross-linking reagents.
  • a CTL epitope is a fragment of an antigen which binds to the peptide-binding cleft of an MHC molecule such that the fragment-MHC complex is recognized by a T cell antigen-specific receptor (TCR) and thereby stimulates a CTL response.
  • TCR T cell antigen-specific receptor
  • the protein sequence selected for a ratchet library can be from a protein with known immunogenic CTL epitopes, or from a protein whose CTL-stimulating ability has not been determined, in which case the ratchet library method can be used to identify CTL epitopes.
  • Ratchet libraries can be constructed from CTL epitopes (or putative CTL epitopes ) of viruses, bacteria, parasites, tumor antigens, allergens, amino acid sequences deduced from an intron or exon/intron mixtures, or from abberrant proteins often associated with malignancy and generated by frameshift mutations (i.e. frameshift sequences), or any other proteins known to stimulate a CTL response.
  • ratchet libraries can be prepared from the following proteins or proteins from the listed organisms or diseases (with the cited references indicating known CTL response to those proteins) : melanoma proteins [Bakker et al. (1994) J. Exp. Med. 179:1005] including MAGE-1, -2, and -3 [Gaugler et al. (1994) J.Exp. Med. 179:921]; proteins associated with renal cell carcinoma; proteins associated with colon carcinoma [Townsend et al. (1994) Nature: 371:662]; proteins associated with prostate cancer (malignant or benign) including PSA; tyrosinase [Brichard et al. (1993) J. Exp. med.
  • oncogenes such as the HER-2/neu proto-oncogene; ras [Gedde-Dahl et al. (1994) Eur. J. Immunol. 24:410]; MUC1 [Barnd et al. (1989) Proc. Natl. Acad. Sci. USA 86:7159]; p53 [Mijman et al. (1994) Immunol. Lett. 40:171]; pl6; TL [Morita et al. (1994) J. Exp. Med.
  • proteins from HIV-1 or -2 including envelope, gag, pol, nef, tat, rev, vpx, vpu [Nixon et al. (1992)]; HTLV-I or -II including envelope, ga i pol# pX and TAX [Jacobson et al. (1990) Nature 348:245; ly phocytic chorio eningitis virus of mice (LCMV) [Aebisher et al. (1991) Proc. Natl. Acad. Sci.
  • LCMV ly phocytic chorio eningitis virus of mice
  • influenza A, B or C including PB1, PB2, PA, NSl, Ml, NP, HA [McMichael et al. (1978) Eur. J. Immunol. 8:705]; Epstein-Barr virus (EBV) including TETA, EENL, EBNA3, EBNA1 and LMP [Brooks et al. (1993) J. Exp. Med. 178:879]; respiratory syncytia virus (RSV) [Bangham et al. (1985) J. Virol. 56:55]; hepatitis B virus (HBV) [Bertoletti et al. (1993) J. Virol.
  • HCV hepatitis C virus
  • CMV herpes simplex virus
  • CMV cyto egalovirus
  • parainfluenza virus 1 including hemagglutinin, neuraminidase, phosphoprotein and nucleoprotein [Dave et al. (1994) Virology 199:376]
  • intracisternal A particle gag de Bergeyck et al. (1994) Eur. J.
  • the preferred ratchet libraries of this invention are those libraries provided in the Examples and the Figures.
  • CTL responses can be measured by conventional techniques known to ordinarily skilled artisans, including, for example, the dye exclusion test and the Cr- release assay described in Encyclopedia of Immunology. supra at page 451. Another method to assay CTL is described by McDonald et al. (1980) Immunol. Rev. 51:93- 123.
  • the ratchet libraries are prepared by chemical synthesis using standard techniques well known in the art such as the solid-phase synthetic route pioneered by Merrifield.
  • the coupling pf multiple amino acids at a given position is accomplished by providing a mixture of the desired amino acids at the ratios determined by the ratchet process. If necessary the ratio of amino acids in the mixture can be varied to account for different coupling efficiency of those amino acids.
  • an HIV ratchet library can be used as an HIV CTL vaccine (either as a vaccine component or as a therapeutic in the treatment of AIDS) , an HCV ratchet library as an HCV CTL vaccine, an influenza ratchet library as a flu CTL vaccine, a mutant p53 ratchet library as a cancer CTL vaccine and the like.
  • HIV CTL vaccine either as a vaccine component or as a therapeutic in the treatment of AIDS
  • HCV ratchet library as an HCV CTL vaccine
  • influenza ratchet library as a flu CTL vaccine
  • a mutant p53 ratchet library as a cancer CTL vaccine and the like.
  • Vaccine compositions containing one or more distinct ratchet libraries can be introduced into normal subjects to stimulate production of CTL by immunization protocols known in the art.
  • subject ratchet libraries one or more libraries
  • Adjuvants for use in this invention include incomplete Freunds' adjuvant (IFA) , alum, lipidic amino acids and Pairi j Cys (see description in the Examples) . These latter two adjuvants can be either covalently attached to the ratchet to produce a lipopeptide ratchet library or formulated together with the ratchet library for co-administration.
  • Vaccine formulations are readily determined by one of ordinary skill in the art and include formulations for immediate release and for sustained release. Formulations contemplated by this invention include microparticles, microcapsules, emulsions, liposomes, DMSO-glycerol and the like.
  • the present vaccines can be administered by any convenient route including subcutaneous, oral, intramuscular, intravenous, intra-der al, intraocular, vaginal, trans-dermal or other parenteral or enteral route. Similarly the vaccines can be administered as a single dose or divided into multiple doses for administration.
  • the vaccine compositions of the instant invention contain an im unoeffective amount of a ratchet library to treat or prevent disease or malignancy associated with the source of the CTL epitope in that ratchet library.
  • Preferred vaccine compositions are effective for CTL induction with respect to malaria, HIV, HCV, mucin, p53 and influenza and ther associated pathogenic conditions.
  • compositions in dosage unit form can contain about 10 ng to about 2 mg of the peptide (or mixture of peptides) per kg body weight.
  • the dosage unit form is conveniently divided into the appropriate amounts per dosage. Accordingly, another aspect of this invention provides a method of treating or preventing a disease or a malignancy which comprises administering an amount of the library of Claim 1 to a mammal effective to stimulate a CTL response against the disease or malignancy associated with a CTL epitope present in said library.
  • ratchet library i.e., the virus, bacterium or other organism from which the template peptide was derived or a protein associated with malignancy
  • the amount needed for delivery to obtain the desired therapeutic result that is, the amount of library to induce a CTL response of therapeutic benefit for the disease or condition under treatment.
  • these dosages ranges are as indicated above for the vaccine formulation.
  • an efficacious formulation for delivery of the ratchet library can be determined.
  • a ratchet peptide can be used to identify CTL epitopes within a protein sequence.
  • this invention is directed to a method of constructing a library of related peptides to provide a ratchet library which comprises identifying a template peptide; calculating a distribution of amino acids at each position x having those amino acids present in the template peptide at positions x to n-(l-x), inclusive, wherein 1 is from about 7 to about 25, n is from 1+1 to about 100, and x is from 1 to 1; synthesizing said ratchet library; and assaying said ratchet library for the ability to stimulate CTL activity.
  • the ratchet peptide is constructed and used to immunize animals, typically though not necessarily mice.
  • the immunized animals are sacrificed and splenocytes removed and cultured in vitro with a pool of overlapping individual peptides that span the ratcheted template.
  • the activate splenocytes are then tested on target cells pulsed with 50 ⁇ M (for example) pooled peptides, and if any CTL activity is present, the splenoytes are tested on the individual peptides derived from the region. If a single peptide derived from the pool of overlapping peptides is recognized, a new CTL epitope has been identified.
  • the examples serve to illustrate the present invention and are not to be used to limit the scope of the invention.
  • Ratchet libraries were synthesized by standard F-moc chemistry using solid phase peptide synthesis with an F- oc RINK MBHA resin [4-(2 ' ,4 •-dimethoxyphenyl-Fmoc- aminomethyl) -phenoxyacetamido-norleucyl-MBH ⁇ resin; MBHA is methylbenzhydrylamine] according to manufacturer's instructions on an ABI Model 433 peptide synthesizer or similar model.
  • the ratchet libraries were synthesized as linear peptides or as branched peptides using a heptalysyl core.
  • Ratchet libraries were formulated at the indicated concentrations and then used for immunization as microparticles, emulsions or lipopeptides.
  • Microparticles were prepared according to the water- in-oil-water solvent evaporation method described in U.S. Serial No. 08/263,841, filed June 22, 1994, which is incorporated herein by reference, using polylactide-co- glycolide polymer Resomer RG 505 (Boehringher Ingelheim) .
  • Microparticles containing 100 ⁇ g of ratchet library were suspended in 0.5 ml phosphate-buffered saline (PBS) for intraperitoneal immunization of mice on days 0, 10 and 20 followed by sacrifice of the animals 7 to 10 days later.
  • PBS phosphate-buffered saline
  • Emulsions were prepared so that the final preparations contained 100 ⁇ g of ratchet library and 50 ⁇ g Pairi j Cys-seryl—lysyl-lysyl-lysyl-lysyl (Pam 3 Cys-SKKKK) (SEQ ID NO:12) in a volume of 0.5 mL unless indicated otherwise.
  • 4 mg of ratchet library was dissolved in 16 mL H 2 0 and 240 mg of egg lecithin was dispersed therein by homogenization (Model STD 1 fitted with a 0.25" tubular head, Silverson Machines, East Longmeadow, MA) at 10,000 rpm for 5 min.
  • PairtjCys-SKKK (2 mg) was mixed with 4 g soya oil and then added to the library mixture by homogenization at 10,000 -23- rpm for 5 min. Further mixing was conducted by ultrasonic pulsation of the emulsion with an ultra sonic probe (Vibra cell, Sonics and Materialds, Inc., Danbury, CT) . The emulsions (0.5 mL) were injected intraperitoneally into mice on days 0 and 10 followed by sacrifice of the animals 7 to 10 days later.
  • the peptides of the ratchet library were covalently coupled to Pam 3 Cys (tripalmitoyl-5-glycerol-cysteine) as generally described (Deres et al.) to produce the corresponding lipopeptide ratchet library.
  • the lipopeptide ratchet libraries (100 ⁇ g) were suspended in a 0.5 mL of 1% DMSO in glycerol and injected intraperitoneally into mice at day 0. The animals were sacrificed 7-9 days later. Control mice were injected with 0.5 mL phosphate- buffered saline (PBS) using the corresponding injection schedule as that of the formualted ratchet library.
  • PBS phosphate- buffered saline
  • spleens Upon sacrifice, the spleens were removed and splenocytes were pooled and cultured in vitro with the indicated peptide at a concentration of 1 ⁇ g/mL for one week to produce activated splenocytes. CTL assays were then conducted according to the method of McDonald et al. (1980) as briefly described below.
  • a ratchet library was designed from a 20 amino acid sequence of residues 247-266 from the circumsporozoite protein of Plasmodium berghei (CS 247-266) . This sequence contains the nonomer CTL epitope designated as CS 252-260 [Eberl et al. (1993) Int. Immunol. 5:1489-1492).
  • Malarial ratchet library 1 from the template peptide of SEQ ID N0:1 (Fig.IB, bottom panel) was prepared in linear form and formulated in microparticles at a final concentration of 200 ⁇ g/mL.
  • Three BALB/C mice were immunized with 0.5 L per injection as described above.
  • H-2 a target cells mouse mastocytoma cell line P815 or A20.1 were JI Cr labeled for one h, washed and then incubated for one h in the presence of CS 252-260 peptide at 50 ⁇ M, in the presence of an unrelated control peptide, influenza nucleoprotein peptide NP 147-155 at 50 ⁇ M or in media (i.e., in the absence of a peptide antigen).
  • CTL activity was determined by incubating these target cells with activated splenocytes (effector cells) for 4 hours in round-bottomed 96-well plates at a range of effector:target (E:T) ratios of 100:1, 50:1, 25:1 and 12.5:1 and measuring the release of Cr.
  • IB bottom panel
  • a heptalysyl core was synthesized as branched octameric peptides on a heptalysyl core and formulated in microparticles at a final concentration of 2 mg/mL and injected into mice as described above.
  • Splenocytes were incubated and CTL activity was assayed as described above using E:T ratios of 100:1, 50:1 25:1 and 12.5:1.
  • the results are shown in Fig. 3.
  • Malaria ratchet library 2 was formulated and injected into mice as described in the preceding paragraph to determine CTL activity against the 20 residue CS 247-266 peptide.
  • Splenocytes were incubated with CS 252-260 peptide and CTL activity was determined as above, except that the target cells were incubated in the presence of 50 ⁇ M of CS 252-260 peptide, 50 ⁇ M of CS 247-266 peptide or in media using E:T ratios of 100:1, 50:1 25:1 and 12.5:1.
  • the results are shown in Fig. 4.
  • Malaria ratchet library 1 was formulated and injected as a lipopeptide except that the lipopeptide ratchet library was formulated at a concentration of 20 ⁇ g/mL.
  • Splenocytes were incubated CS 252-160 and CTL activity was assayed as described above using E:T ratios of 100:1, 50:1 25:1 and 12.5:1. The results are shown in Fig. 5.
  • Malaria ratchet library 1 was formulated and injected as a lipopeptide ratchet library at a concentration of 200 ⁇ g/mL. Splenocytes were incubated with CS 252-260 and CTL activity was as described above to determine CTL activity against a self peptide.
  • the target cells were incubated in the presence of 50 ⁇ M of CS 252-260 peptide, 50 ⁇ M of a K d - restricted self peptide (SYFPEITHI; SEQ ID NO: 13) or in media using E:T ratios of 100:1, 50:1 25:1 and 12.5:1.
  • SYFPEITHI K d - restricted self peptide
  • E:T ratios 100:1, 50:1 25:1 and 12.5:1.
  • the results are shown in Fig. 6.
  • These malaria ratchet libraries elicit malaria- specific CTL at immunogen doses ranging from 10 ⁇ g to 1 mg (Figs. 2-5).
  • the CTL activity is MHC restricted and is elicited when presented with the processed epitope as a longer peptide (Fig. 4) .
  • Fig. 7 bottom panel, illustrates an MHC-restricted malaria ratchet library from the template peptide of SEQ ID NO:l.
  • This library is constructed from malaria ratchet library 1 of Fig. IB (also top panel of Fig. 7) by replacing those positions which contain anchor residues for the K h , D h , K J , and L J molecules (see below) with an equal proportion of the anchor residues at the position in question.
  • the anchor residues are those amino acids which have been identified as necessary for binding to the MHC class 1 molecule for the given haplotype.
  • the anchor residues for the indicated haplotypes are shown in the middle panel of Fig. 7 and the MHC-restricted malaria ratchet library is shown in the bottom panel of Fig. 7.
  • the ratchet incorporates only those anchor amino acids shown in the middle panel.
  • position 2 contains 50% tyrosine Y and 50 % proline
  • position 5 contains 33% asparagine, 33% tyrosine and 33% phenylalanine
  • position 8 contains 100% leucine
  • position 9 contains 25% isoleucine, 25% leucine, 25% phenylalanine and 25% methionine.
  • this ratchet has the anchor residues involved in binding MHC class I molecules and stimulating CTL.
  • HIV ratchet libraries Can Accomodate Antigenic Diversity
  • An HIV ratchet library was constructed from the SSAL (Fig. 8A, bottom panel) of template peptide SEQ ID NO:2 and the additional 14 HIV-1 sequences (Fig. 8A) from a 35 amino acid sequence of the HIV-1 gpl20 V3 loop region, the priniciple neutralizing domain known to have extensive sequence variability. This region contains a D* restricted CTL epitope at amino acid positions 318-326.
  • the antigenic diversity of this region is acco odated by taking 15 HIV-1 consensus sequences including the sequence HIV-MVP5188 and constructing an SSAL library where the identity and ratio of amino acids at each position is determined by the relative prevalence of amino acids in those 16 sequences.
  • the SSAL library which is "ratcheted" to yield the HIV-1 ratchet library shown in the bottom panel of Fig. 8.
  • EXAMPLE 4 Peptide Ratchet Libraries from Different Length Template Peptides To examine the effect of template peptide length on the efficacy of CTL induction via ratchet libraries, three HIV-1 gag peptide linear ratchet libraries (Figs. 9-11, bottom panel) containing a mouse HIV CTL epitope were synthesized using template peptides of lengths 100, 40 or 20 amino acids of the gag sequence as shown in Figs. 9-11, respectively, and designated by SEQ ID NOS:3-5, respectively. These libraries were formulated with 100 ⁇ g in 0.5 mL as microparticles, emulsions or lipopeptides and injected as described in Example 1 with the following modifications: The immunized mice were C57BL/6 mice.
  • Activated splenocytes were prepared by culturing with 1 ⁇ g/mL HIV gag peptide 390-398 [Elvin et al. (1993) J. Immunol. Methods 158:161-171].
  • the target cells were EL4 and were incubated with 50 ⁇ M HIV gag peptide 390-398.
  • the results are presented in Table 1.
  • Fig. 12 show the specific cell lysis results for the emulsion formulation of the ratchet library from the 40 amino acid template.
  • the template peptide of the indicated amino acid length was ratcheted to 9-mers as described in Example .
  • Fig 13. (bottom panel) provides a mucin ratchet library constructed from the template peptide of SEQ ID NO:7 or SEQ ID NO:8.
  • Mucin is a large, heavily glycosylated molecule expressed and secreted by ductal epithelial cells and tumors. Mucin consists of multiple copies of a 20 amino acid tandem repeat (SEQ ID NO:6) which appears to elicit non-MHC restricted CTL responses. Because the 20-mer repeat will not contain every possible nonomer when used as a template, due to end effects in the ratchet, an alternative approach was used to generate all possible nonomers of the repeating 20-mer peptide.
  • the last eight carboxy-terminal amino acids of the 20-mer repeat peptide were placed at its amino terminus to yield a template peptide of 28 amino acids before calculation of the ratchet (Fig. 13, middle panel) .
  • the first eight amino-terminal amino acids were placed at its carboxyl terminus to yield a template peptide of 28 amino acids before calculation of the ratchet (Fig. 13, middle panel) .
  • the calulated mucin ratchet library is the same.
  • the protein p53 is a tumor supressor which fails to
  • Class 9 due to one or more point mutations in the protein.
  • the four ratchets are designed around mutation hot 3 spots in the protein: (1) template peptide of amino acids 4 124-151 (SEQ ID NO:9) as a 10-mer ratchet library (Fig. 5 14, bottom panel); (2) template peptide of amino acids 6 166-187 as a 9-mer ratchet library; (3) template peptide 7 of amino acids 228-256 as a 9-mer ratchet library; and (4) 8 template peptide of amino acids 264-289 as a 9-mer ratchet 9 library.
  • the ratchet library from the 166-187 template is 1 especially useful for treating colon cancer since this 2 mutation is frquently encountered with this malignancy.
  • influenza ratchet library 1 from a 25-mer template peptide of residues 139- 163 (SEQ ID NO: 10) of influenza A A/34/PR8 nucleoprotein. This library encompasses the known K''-restricted epitope of residues 147-155.
  • Fig. 16 shows influenza ratchet library 2 constructed from the template peptide of SEQ ID NO: 11 which is a linkage of 3 CTL epitopes in order from N to C terminus of residues 50-58, 147-155 and 366-374 of the nucleoprotein.
  • Lys lie Leu Glu Phe Val Lys Gin 15 20

Landscapes

  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biophysics (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Medicinal Chemistry (AREA)
  • Biochemistry (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Toxicology (AREA)
  • Zoology (AREA)
  • Virology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Analytical Chemistry (AREA)
  • Oncology (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

La présente invention concerne des banques à encliquetage constituées de peptides voisins synthétisés simultanément lors d'une synthèse unique de peptides. Les banques à encliquetage sont dérivées d'un peptide à matrice plus longue en 'encliquetant' séquentiellement la séquence de matrice dans la plus faible longueur d'encliquetage, et elles sont utilisées pour induire ou stimuler des lymphocytes T cytotoxiques (LTC), si l'épitope des LTC est connu. Si l'épitope des LTC est inconnu, on peut utiliser la banque à encliquetage pour identifier des épitopes de LTC. Les banques à encliquetage peuvent être préparées à partir de toute séquence protéique pour laquelle on désire une réaction LTC immunitaire, et on peut leur donner une formule pour l'administration en tant que vaccin ou que thérapeutique, dans le traitement ou la prévention d'une maladie ou d'une affection maligne. Par exemple, une banque à encliquetage peut être utilisée dans la prévention et le traitement de maladies infectieuses ou malignes, y compris le VIH, la grippe, le paludisme et les cancers du sein, des ovaires, du poumon et du côlon.
PCT/US1995/016290 1994-12-27 1995-12-15 Banques de peptides a encliquetage pour vaccins et therapeutiques induisant des lymphocytes t cytotoxiques WO1996022067A2 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU58497/96A AU5849796A (en) 1994-12-27 1995-12-15 Peptide ratchet libraries for ctl-inducing vaccines and therapeutics

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US36633294A 1994-12-27 1994-12-27
US08/366,332 1994-12-27

Publications (2)

Publication Number Publication Date
WO1996022067A2 true WO1996022067A2 (fr) 1996-07-25
WO1996022067A3 WO1996022067A3 (fr) 1996-11-28

Family

ID=23442575

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US1995/016290 WO1996022067A2 (fr) 1994-12-27 1995-12-15 Banques de peptides a encliquetage pour vaccins et therapeutiques induisant des lymphocytes t cytotoxiques

Country Status (2)

Country Link
AU (1) AU5849796A (fr)
WO (1) WO1996022067A2 (fr)

Cited By (19)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1997011958A1 (fr) * 1995-09-29 1997-04-03 The Scripps Research Institute Analyse de signature de proteine
WO1997041440A1 (fr) * 1996-04-26 1997-11-06 Rijksuniversiteit Te Leiden Procedes de selection et de production d'epitopes peptidiques de lymphocytes t et vaccins contenant lesdits epitopes selectionnes
WO1999023114A1 (fr) * 1997-10-31 1999-05-14 Biomira Inc. Derives de muc-1 et leur utilisation pour traiter l'immuno-depresseur induite par mucine muc-1 et associee au cancer
WO2000052046A1 (fr) * 1999-03-01 2000-09-08 Imperial Cancer Research Technology Limited Glycopeptide immunomodulatrice
WO2001016163A2 (fr) * 1999-08-27 2001-03-08 Eurodiagnostica Ab Melange peptidique et vaccin contre une infection virale chronique
FR2809402A1 (fr) * 2000-05-26 2001-11-30 Dev Des Antigenes Combinatoire Bibliotheques peptidiques combinatoires convergentes et leur application a la vaccination contre le virus de l'hepatite c
EP1369428A1 (fr) * 1997-10-31 2003-12-10 Biomira Inc. Dérivés de Muc-1 et leur utilisation dans le traitement d'immunosupression induite par le MUC-1 Mucin associé au cancer
US7252829B1 (en) * 1998-06-17 2007-08-07 Idm Pharma, Inc. HLA binding peptides and their uses
EP1950223A2 (fr) * 1998-03-09 2008-07-30 Zealand Pharma A/S Conjugués peptides pharmacologiquement actifs dotés d'une tendance réduite à l'hydrolyse enzymatique
US7462360B2 (en) * 1999-12-27 2008-12-09 Entopharm Co., Ltd. Alloferons—immunomodulatory peptides
US7572882B2 (en) 1999-12-10 2009-08-11 Pharmexa Inc. Inducing cellular immune responses to human papillomavirus using peptide and nucleic acid compositions
US7611713B2 (en) 1993-03-05 2009-11-03 Pharmexa Inc. Inducing cellular immune responses to hepatitis B virus using peptide compositions
DE102009034779A1 (de) 2009-07-25 2011-02-03 Emc Microcollections Gmbh Synthetische Analoga bakterieller Lipopeptide und ihre Anwendung zur Therapie und Prophylaxe allergischer Erkrankungen
US20130266599A1 (en) * 2010-12-13 2013-10-10 Cel - Sci Corporation Method for inducing an immune response against avian, swine, spanish, h1n1, h5n9 influenza viruses and formulations thereof
EP2341072A3 (fr) * 1997-05-08 2013-10-30 Oncothyreon Inc. Procédé pour l'obtention de cellules lymphocytes activées et de cellules présentatrices d'antigènes à impulsion antigénique
US8741576B2 (en) 1999-11-18 2014-06-03 Epimunne Inc. Heteroclitic analogs and related methods
US9266930B1 (en) 1993-03-05 2016-02-23 Epimmune Inc. Inducing cellular immune responses to Plasmodium falciparum using peptide and nucleic acid compositions
US10179174B2 (en) 2011-05-25 2019-01-15 Cel-Sci Corp. Method for inducing an immune response and formulations thereof
WO2023023523A1 (fr) * 2021-08-16 2023-02-23 Duke University Vaccins de nouvelle génération comprenant des banques antigéniques et leurs procédés de préparation et leurs méthodes d'utilisation

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1992000091A1 (fr) * 1990-07-02 1992-01-09 Bioligand, Inc. Banque de bio-oligomeres aleatoires, son procede de synthese et son mode d'emploi

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1992000091A1 (fr) * 1990-07-02 1992-01-09 Bioligand, Inc. Banque de bio-oligomeres aleatoires, son procede de synthese et son mode d'emploi

Non-Patent Citations (7)

* Cited by examiner, † Cited by third party
Title
ANNALES DE BIOLOGIE CLINIQUE, vol. 49, no. 4, April 1991, pages 231-242, XP002014808 R.H.MELOEN E.A.: "The use of peptides to reconstruct conformational determinants" *
BEHRING INSTITUTE MITTEILUNGEN, no. 94, July 1994, pages 48-60, XP002014809 J.RUPPERT E.A : "Class I MHC-peptide interaction: structural and functional aspects" *
EUR.J.IMMUNOL., vol. 24, November 1994, pages 2789-2795, XP002014810 J.ESTAQUIER E.A.: "The mixotope: a combinatorial peptide library as a T cell and B cell immunogen" *
J.A.SMITH E.A.: "PEPTIDES,chemistry and Biology; Proc.12th Am. Pept. Symp." 1992 , ESCOM , LEIDEN XP002014814 H.Grass-Masse e.a.: Synthetic vaccines: the mixotope strategy see page 842 - page 844 *
JOURNAL OF IMMUNOLOGICAL METHODS, vol. 173, no. 2, 1 August 1994, NEW YORK US, pages 253-263, XP002014811 E.BORGES E.A.: "efficacy of synthetic vaccines in the induction of cytotoxic t lymphocytes" *
NATURE, vol. 342, 30 November 1989, pages 561-564, XP002014812 K DERES E.A.: "In vivo priming of virus specific CTL's with synthetic peptides" cited in the application *
VACCINE, vol. 12, no. 9, 1994, pages 791-797, XP002014813 M.FRIEDE E.A.: "Selective induction of protection against influenza virus infection in mice by a lipid-peptide conjugate delivered in liposomesŸ" cited in the application *

Cited By (27)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7611713B2 (en) 1993-03-05 2009-11-03 Pharmexa Inc. Inducing cellular immune responses to hepatitis B virus using peptide compositions
US9266930B1 (en) 1993-03-05 2016-02-23 Epimmune Inc. Inducing cellular immune responses to Plasmodium falciparum using peptide and nucleic acid compositions
WO1997011958A1 (fr) * 1995-09-29 1997-04-03 The Scripps Research Institute Analyse de signature de proteine
WO1997041440A1 (fr) * 1996-04-26 1997-11-06 Rijksuniversiteit Te Leiden Procedes de selection et de production d'epitopes peptidiques de lymphocytes t et vaccins contenant lesdits epitopes selectionnes
EP2341072A3 (fr) * 1997-05-08 2013-10-30 Oncothyreon Inc. Procédé pour l'obtention de cellules lymphocytes activées et de cellules présentatrices d'antigènes à impulsion antigénique
AU749152B2 (en) * 1997-10-31 2002-06-20 Biomira Inc. MUC-1 derivatives and their use in treating cancer-associated MUC-1 mucin-induced immunosuppression
WO1999023114A1 (fr) * 1997-10-31 1999-05-14 Biomira Inc. Derives de muc-1 et leur utilisation pour traiter l'immuno-depresseur induite par mucine muc-1 et associee au cancer
EP1369428A1 (fr) * 1997-10-31 2003-12-10 Biomira Inc. Dérivés de Muc-1 et leur utilisation dans le traitement d'immunosupression induite par le MUC-1 Mucin associé au cancer
US7935786B2 (en) 1998-03-09 2011-05-03 Zealand Pharma A/S Pharmacologically active peptide conjugates having a reduced tendency towards enzymatic hydrolysis
EP1950223A3 (fr) * 1998-03-09 2009-05-13 Zealand Pharma A/S Conjugués peptides pharmacologiquement actifs dotés d'une tendance réduite à l'hydrolyse enzymatique
EP1950223A2 (fr) * 1998-03-09 2008-07-30 Zealand Pharma A/S Conjugués peptides pharmacologiquement actifs dotés d'une tendance réduite à l'hydrolyse enzymatique
US7252829B1 (en) * 1998-06-17 2007-08-07 Idm Pharma, Inc. HLA binding peptides and their uses
WO2000052046A1 (fr) * 1999-03-01 2000-09-08 Imperial Cancer Research Technology Limited Glycopeptide immunomodulatrice
WO2001016163A2 (fr) * 1999-08-27 2001-03-08 Eurodiagnostica Ab Melange peptidique et vaccin contre une infection virale chronique
WO2001016163A3 (fr) * 1999-08-27 2001-09-07 Eurodiagnostica Ab Melange peptidique et vaccin contre une infection virale chronique
US8741576B2 (en) 1999-11-18 2014-06-03 Epimunne Inc. Heteroclitic analogs and related methods
US7572882B2 (en) 1999-12-10 2009-08-11 Pharmexa Inc. Inducing cellular immune responses to human papillomavirus using peptide and nucleic acid compositions
US7462360B2 (en) * 1999-12-27 2008-12-09 Entopharm Co., Ltd. Alloferons—immunomodulatory peptides
FR2809402A1 (fr) * 2000-05-26 2001-11-30 Dev Des Antigenes Combinatoire Bibliotheques peptidiques combinatoires convergentes et leur application a la vaccination contre le virus de l'hepatite c
WO2001092311A3 (fr) * 2000-05-26 2002-05-02 Dev Des Antigenes Soc D Et Bibliotheques peptidiques combinatoires convergentes et leur applicaton a la vaccination contre le virus de l'hepatite c
WO2001092311A2 (fr) * 2000-05-26 2001-12-06 Societe D'etude Et De Developpement Des Antigenes Combinatoires - Sedac Therapeutics Bibliotheques peptidiques combinatoires convergentes et leur applicaton a la vaccination contre le virus de l'hepatite c
WO2011012240A2 (fr) 2009-07-25 2011-02-03 Emc Microcollections Gmbh Lipopeptide pour la thérapie et la prophylaxie de maladies allergiques
DE102009034779A1 (de) 2009-07-25 2011-02-03 Emc Microcollections Gmbh Synthetische Analoga bakterieller Lipopeptide und ihre Anwendung zur Therapie und Prophylaxe allergischer Erkrankungen
US20130266599A1 (en) * 2010-12-13 2013-10-10 Cel - Sci Corporation Method for inducing an immune response against avian, swine, spanish, h1n1, h5n9 influenza viruses and formulations thereof
US10238747B2 (en) * 2010-12-13 2019-03-26 Cel-Sci, Corp Method for inducing an immune response against avian, swine, spanish, H1N1, H5N9 influenza viruses and formulations thereof
US10179174B2 (en) 2011-05-25 2019-01-15 Cel-Sci Corp. Method for inducing an immune response and formulations thereof
WO2023023523A1 (fr) * 2021-08-16 2023-02-23 Duke University Vaccins de nouvelle génération comprenant des banques antigéniques et leurs procédés de préparation et leurs méthodes d'utilisation

Also Published As

Publication number Publication date
WO1996022067A3 (fr) 1996-11-28
AU5849796A (en) 1996-08-07

Similar Documents

Publication Publication Date Title
WO1996022067A2 (fr) Banques de peptides a encliquetage pour vaccins et therapeutiques induisant des lymphocytes t cytotoxiques
Defoort et al. Macromolecular assemblage in the design of a synthetic AIDS vaccine.
JP3738395B2 (ja) Ｈｌａ−制限型ｂ型肝炎ウィルスのｃｔｌエピトープ
US6419931B1 (en) Compositions and methods for eliciting CTL immunity
Hamley Peptides for vaccine development
CA2562784C (fr) Vecteurs fluorocarbones pour l'administration d'antigenes et constructions connexes
JP3926839B2 (ja) 万能ｄｒ−結合性ペプチドを用いる免疫応答の改変
JP3650110B2 (ja) Ｂ型肝炎ウイルスに対する細胞毒性ｔリンパ球応答を誘発するためのペプチド
KR102625645B1 (ko) B형 간염 바이러스에 대한 백신
CZ20013195A3 (cs) HIV peptidy, antigeny, vakcíny, souprava pro imunotest a způsob detekce protilátek indukovaných HIV
US5993823A (en) Cytotoxic T lymphocyte-inducing lipopeptides and methods of use
JPH10503473A (ja) 細胞傷害性ｔリンパ球刺激およびｈｃｖ曝露診断用ｃ型肝炎ウイルスコアペプチド
JP5901084B2 (ja) ペプチドアジュバント
JP2010029217A (ja) Ｈｉｖ特異的ｃｔｌを誘導し得るペプチド及び該ペプチドを含む抗ａｉｄｓ予防・治療剤
AU748700B2 (en) HIV virus mimotopes
AU727738B2 (en) Compositions and methods for eliciting CTL immunity
KR20080004445A (ko) 절단형 ｌｈｒｈ 포뮬레이션
EP1617865A2 (fr) Vaccins vih-1 ameliores et leurs procedes d'utilisation
Sundaram Evaluation of T-cell and B-cell epitopes and design of multivalent vaccines against HTLV-1 diseases
JP2010001303A (ja) Ｈｌａ結合ペプチド及びその使用

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A2

Designated state(s): AU CA FI JP NO

AL Designated countries for regional patents

Kind code of ref document: A2

Designated state(s): AT BE CH DE DK ES FR GB GR IE IT LU MC NL PT SE

121 Ep: the epo has been informed by wipo that ep was designated in this application
122 Ep: pct application non-entry in european phase
NENP Non-entry into the national phase

Ref country code: CA