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WO2004091493A2 - Vaccins ameliores a base de proteines de choc thermique et immunotherapies associees - Google Patents

Vaccins ameliores a base de proteines de choc thermique et immunotherapies associees Download PDF

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Publication number
WO2004091493A2
WO2004091493A2 PCT/US2004/010983 US2004010983W WO2004091493A2 WO 2004091493 A2 WO2004091493 A2 WO 2004091493A2 US 2004010983 W US2004010983 W US 2004010983W WO 2004091493 A2 WO2004091493 A2 WO 2004091493A2
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WIPO (PCT)
Prior art keywords
seq
leu
arg
gly
phe
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PCT/US2004/010983
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WO2004091493A3 (fr
Inventor
Jessica Baker Flechtner
Kenya Prince-Cohane
Sunil Mehta
Paul Slusareqicz
Sofija Andjelic
Brian H. Barber
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Agenus Inc
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Antigenics LLC
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Priority claimed from US10/776,521 external-priority patent/US7420037B2/en
Priority claimed from US10/820,067 external-priority patent/US7309491B2/en
Priority to MXPA05010881A priority Critical patent/MXPA05010881A/es
Application filed by Antigenics LLC filed Critical Antigenics LLC
Priority to JP2006509858A priority patent/JP2007525448A/ja
Priority to BRPI0409321-6A priority patent/BRPI0409321A/pt
Priority to EP04759338A priority patent/EP1617804A4/fr
Priority to AU2004229458A priority patent/AU2004229458B2/en
Priority to CA002521809A priority patent/CA2521809A1/fr
Publication of WO2004091493A2 publication Critical patent/WO2004091493A2/fr
Anticipated expiration legal-status Critical
Publication of WO2004091493A3 publication Critical patent/WO2004091493A3/fr
Ceased legal-status Critical Current

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    • 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
    • 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
    • A61K39/39—Medicinal preparations containing antigens or antibodies characterised by the immunostimulating additives, e.g. chemical adjuvants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/04—Immunostimulants
    • 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
    • A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
    • A61K2039/55511—Organic adjuvants
    • A61K2039/55516—Proteins; Peptides
    • 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
    • A61K2039/60—Medicinal preparations containing antigens or antibodies characteristics by the carrier linked to the antigen
    • A61K2039/6031—Proteins
    • A61K2039/6043—Heat shock proteins
    • 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
    • A61K2039/62—Medicinal preparations containing antigens or antibodies characterised by the link between antigen and carrier
    • A61K2039/622—Medicinal preparations containing antigens or antibodies characterised by the link between antigen and carrier non-covalent binding
    • 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

  • the present invention relates to methods and compositions for inducing an immune response in a subject, wherein the subject is administered an effective amount of at least one or more defined hybrid antigens optionally in combination with one or more heat shock proteins. These methods and compositions may be used in the treatment of infectious diseases and cancers.
  • Heat shock proteins were originally observed to be expressed in increased amounts in mammalian cells which were exposed to sudden elevations of temperature, while the expression of most cellular proteins is significantly reduced. It has since been determined that such proteins are produced in response to various types of stress, including glucose deprivation.
  • heat shock protein will be used to encompass both proteins that are expressly labeled as such as well as other stress proteins, including homologues of such proteins that are expressed constitutively (i.e., in the absence of stressful conditions).
  • heat shock proteins include BiP (also referred to as grp78), hsp70, hsc70, gp96 (grp94), hsp60, hsp40 and hsp90.
  • Heat shock proteins have the ability to bind other proteins in their non-native states, and in particular to bind nascent peptides emerging from ribosomes or extruded into the endoplasmic reticulum. Hendrick and Hartl, Ann. Rev. Biochem. 62:349-384 (1993); Hartl, Nature 381:571-580 (1996). Further, heat shock proteins have been shown to play an important role in the proper folding and assembly of proteins in the cytosol, endoplasmic reticulum and mitochondria; in view of this function, they are referred to as "molecular chaperones.” Frydman et al., N ⁇ f ⁇ re 370:111-117 (1994); Hendrick and Hartl, Ann. Rev. Biochem. 62:349-384 (1993); Hartl, Nature 381:571-580 (1996).
  • the protein BiP a member of a class of heat shock proteins referred to as the hsp70 family
  • hsp70 a member of a class of heat shock proteins referred to as the hsp70 family
  • gp96 is a member of the hsp90 family of stress proteins which localizes in the endoplasmic reticulum. Li and Srivastava, EMBO J. 12:3143-3151 (1993); Mazzarella and Green, J. Biol Chem. 262:8875-8883 (1987). It has been proposed that gp96 may assist in the assembly of multi-subunit proteins in the endoplasmic reticulum. Wiech et al., Nature 358:169-170 (1992).
  • PCT US96/13363 describes hybrid antigens comprising an antigenic domain and a heat shock protein binding domain that, in a complex with a heat shock protein, induces immunological responses to antigens and are thus useful for treatment of cancer and infectious diseases.
  • PCT/US98/22335 describes additional heat shock protein binding domains for similar uses, as well as the ability for hybrid antigen administered alone to induce an immune response. It has now been discovered that improvements in the peptide linker present between the at least one antigenic domain and at least one heat shock protein binding domain in a hybrid antigen leads to an increase in biological activity. This increase is also found to provide an increase in inducing an immune response against the antigenic portion of the hybrid antigen. It is towards these improved peptide linkers, hybrid peptides containing them and their uses with and without heat shock protein, that the present application is directed.
  • the present invention relates to methods and compositions for inducing an immune response in a subject, wherein at least one defined hybrid antigen optionally in a complex with a heat shock protein is administered to the subject.
  • the hybrid antigen comprises at least one antigenic domain and at least one heat shock protein binding domain, and at least one peptide linker there between.
  • Induction of an immune response to an antigen associated with a disease such as an infectious disease or tumor is useful for treatment of the disease.
  • the antigenic or immunogenic domain of the hybrid antigen may be an entire protein or peptide antigen, or may be only a portion of the selected antigen, for example a selected epitope of the antigen.
  • the heat shock protein binding domain is a peptide that binds to a heat shock protein, preferably a peptide of 7-15 amino acids that binds to a heat shock protein, more preferably a hydrophobic peptide that binds to a heat shock protein, and most preferably a hydrophobic peptide of 7-15 amino acids that binds to a heat shock protein.
  • the linker has a sequence from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (OLE), Ala Lys Val Leu (AKVL; SEQ ID NO:1001); Lys Asn (KN); Arg Lys (RK); or AA ⁇ -AA 2 -AA -leucine, wherein AAi is A, S, V, E, G, L, or K, preferably V, more preferably S, and most preferably A ; AA2 is K, V, or E, preferably E, more preferably V and most preferably K; and AA3 is V, S, F, K, A, E, or T,
  • Gin Leu Lys (QLK), Arg Lys (RK) and Ala Lys Val Leu (AKVL; SEQ ID NO: 1001) are preferred, and Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000) is most preferred.
  • the present invention provides for methods of administering such hybrid antigens alone, as well as heat shock protein/hybrid antigen compositions, the latter comprising (i) combining one or more heat shock protein with one or more hybrid antigens in vitro, under conditions wherein binding of hybrid antigen to heat shock protein occurs to form a hybrid antigen/heat shock protein complex; and (ii) administering the hybrid antigen, bound to heat shock protein, in an effective amount to a subject in need of such treatment.
  • hybrid antigens optionally in combination with heat shock protein may be introduced into a subject by administering to the subject a nucleic acid encoding the hybrid antigen, optionally with nucleic acid encoding the heat shock protein.
  • the invention is directed to a hybrid antigen consisting essentially of an antigenic domain of an infectious agent or tumor antigen, a binding domain that non-covalently binds to a heat shock protein, and a peptide linker separating the antigenic domain and the binding domain, and wherein the peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO:1001); Lys Asn (KN); Arg Lys (RK); or AA ⁇ -AA 2 -AA 3 -leucine,
  • Phe Phe Arg Lys (FFRK; SEQ ID NO:1000) is most preferred.
  • the invention is directed to a hybrid antigen consisting essentially of a plurality of antigenic domains of one or more infectious agents or one or more tumor antigens, at least one binding domain that non-covalently binds to a heat shock protein, and at least one peptide linker separating the antigenic domains and the at least one binding domain, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1001); Lys Asn (KN); Arg Lys (RK);
  • Gin Leu Lys (QLK), Arg Lys (RK) and Ala Lys Val Leu (AKVL; SEQ ID NO: 1001) are preferred, and Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000) is most preferred.
  • at least one of the antigenic domains in the aforementioned hybrid antigen is a T helper epitope.
  • the invention is directed to a hybrid antigen comprising an antigenic domain of an infectious agent or tumor antigen and a binding domain that non- covalently binds to a heat shock protein, and a peptide linker there between, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO:1001); Lys Asn (KN); Arg Lys (RK); or AAi-AA2-AA 3 -leucine, wherein AAi is A, S, V, E
  • the aforementioned hybrid antigen has a peptide linker separating the antigenic domain and the binding domain.
  • the invention is directed to a hybrid antigen comprising a plurality of antigenic domains of one or more infectious agents or one or more tumor antigens and at least one binding domain that non-covalently binds to a heat shock protein, and at least one peptide linker there between, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO:1001); Lys Asn (KN); Arg Lys (RK); or AA ⁇ -AA 2 -AA 3 -leu
  • Gin Leu Lys (QLK), Arg Lys (RK) and Ala Lys Val Leu (AKVL; SEQ ID NO:1001) are preferred, and Phe Phe Arg Lys (FFRK; SEQ ID NO:1000) is most preferred.
  • at least one of the antigenic domains is a T helper epitope.
  • the invention is directed to a composition for inducing an immune response to an infectious agent or tumor antigen comprising at least one hybrid antigen, the hybrid antigen comprising an antigenic domain of the infectious agent or tumor antigen, a binding domain that non-covalently binds to a heat shock protein, and at least one peptide linker there between, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1001); Lys Asn (KN); Arg Lys (RK); or
  • the composition comprises a plurality of hybrid antigens, and one of the hybrid antigens can comprise a T helper epitope.
  • the invention is directed to a composition for inducing an immune response to an infectious agent or tumor antigen comprising at least one hybrid antigen, the hybrid antigen comprising a plurality of antigenic domains at least one of which is from the infectious agent or tumor antigen, at least one binding domain that non- covalently binds to a heat shock protein, and at least one peptide linker there between, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO:1001); Lys Asn (FFRK; SEQ ID NO:
  • Gin Leu Lys (QLK), Arg Lys (RK) and Ala Lys Val Leu (AKVL; SEQ ID NO:1001) are preferred, and Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000) is most preferred.
  • the invention is directed to a composition for inducing an immune response to an infectious agent or tumor antigen comprising at least one hybrid antigen, the hybrid antigen consisting essentially of an antigenic domain of the infectious agent or tumor antigen, a binding domain that non-covalently binds to a heat shock protein, and a peptide linker separating the antigenic domain and the binding domain, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1001); Lys Asn (KN
  • the aforementioned composition comprises a plurality of hybrid antigens.
  • at least one of the plurality of hybrid antigens comprises a T helper epitope.
  • the invention is directed to a composition for inducing an immune response to an infectious agent or tumor antigen comprising at least one hybrid antigen, the hybrid antigen consisting essentially of a plurality of antigenic domains at least one of which is from the infectious agent or tumor antigen, at least one binding domain that non-covalently binds to a heat shock protein, and at least one peptide linker separating the antigenic domain and the binding domain, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1000; Phe Arg
  • the invention is directed to a method for inducing an immune response to an infectious agent or tumor antigen comprising administering to a subject a complex of a heat shock protein and a hybrid antigen comprising at least one antigenic domain of the infectious agent or tumor antigen, at least one binding domain comprising a peptide that non-covalently binds to a heat shock protein, and a peptide linker there between; wherein the hybrid antigen and the heat shock protein are non-covalently bound, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE
  • the complex comprises a plurality of hybrid antigens.
  • at least one of the hybrid antigens is a T helper epitope.
  • the hybrid antigen comprises a plurality of antigenic domains, and at least one of the antigenic domains can be a T helper epitope.
  • the complex comprises a plurality of hybrid antigens, at least one of the hybrid antigens comprises a plurality of antigenic domains.
  • the heat shock protein is a hsp70.
  • the invention is directed to a method for inducing an immune response to an infectious agent or tumor antigen comprising administering to a subject a complex of a heat shock protein and a hybrid antigen, the hybrid antigen consisting essentially of at least one antigenic domain of an infectious agent or tumor antigen, a binding domain that non-covalently binds to a heat shock protein, and a peptide linker separating the antigenic domain and the binding domain, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (FFRK; SEQ ID NO: 1000;
  • the complex comprises a plurality of hybrid antigens.
  • at least one of the hybrid antigens is a T helper epitope.
  • the hybrid antigen comprises a plurality of antigenic domains.
  • at least one of the antigenic domains is a T helper epitope.
  • the complex comprises a plurality of hybrid antigens, at least one of the hybrid antigens comprising a plurality of antigenic domains.
  • the heat shock protein is a hsp70.
  • the invention is directed to a method for inducing an immune response to an infectious agent or tumor antigen comprising administering to a subject at least one hybrid antigen comprising at least one antigenic domain of the infectious agent or tumor antigen, at least one binding domain comprising a peptide that non-covalently binds to a heat shock protein, and at least one peptide linker there between, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1001); Lys Asn (K
  • the complex comprises a plurality of hybrid antigens.
  • at least one of the hybrid antigens is a T helper epitope.
  • the hybrid antigen comprises a plurality of antigenic domains.
  • at least one of the antigenic domains is a T helper epitope.
  • the complex comprises a plurality of hybrid antigens, at least one of the hybrid antigens comprising a plurality of antigenic domains.
  • a peptide linker separates the antigenic domain and the binding domain.
  • the invention is directed to a method for inducing an immune response to an infectious agent or tumor antigen comprising administering to a subject at least one hybrid antigen, the hybrid antigen consisting essentially of at least one antigenic domain of an infectious agent or tumor antigen, a binding domain that non- covalently binds to a heat shock protein, and a peptide linker separating the antigenic domain and the binding domain, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1000; Phe Arg Lys (FRK
  • the complex comprises a plurality of hybrid antigens.
  • at least one of the hybrid antigens is a T helper epitope.
  • the hybrid antigen comprises a plurality of antigenic domains.
  • at least one of the antigenic domains is a T helper epitope.
  • the complex comprises a plurality of hybrid antigens, at least one of the hybrid antigens comprising a plurality of antigenic domains.
  • the invention is directed to a method for treating an infectious disease or cancer comprising administering to a subject a complex of a heat shock protein and a hybrid antigen comprising at least one antigenic domain of an infectious agent or tumor antigen associated with the infectious disease or cancer, a binding domain comprising a peptide that non-covalently binds to a heat shock protein, and a peptide linker there between; and wherein the hybrid antigen and the heat shock protein are non-covalently bound, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE
  • the complex comprises a plurality of hybrid antigens.
  • at least one of the hybrid antigens is a T helper epitope.
  • the hybrid antigen comprises a plurality of antigenic domains.
  • at least one of the antigenic domains is a T helper epitope.
  • the complex comprises a plurality of hybrid antigens, at least one of the hybrid antigens comprising a plurality of antigenic domains.
  • a peptide linker separates the antigenic domain and the binding domain.
  • the heat shock protein is a hsp70.
  • the invention is directed to a method for treating an infectious disease or cancer comprising administering to a subject a complex of a heat shock protein and a hybrid antigen, the hybrid antigen consisting essentially of at least one antigenic domain of an infectious agent or tumor antigen associated with the infectious disease or cancer, at least one binding domain that non-covalently binds to a heat shock protein, and a peptide linker separating the antigenic domain and the binding domain, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID
  • AAi is A, S, V, E, G, L, or K, preferably V, more preferably S, and most preferably A ; AA 2 is K, V, or E, preferably E, more preferably V and most preferably K; and AA 3 is V, S, F, K, A, E, or T, preferably F, more preferably S and most preferably V.
  • the complex comprises a plurality of hybrid antigens.
  • at least one of the hybrid antigens is a T helper epitope.
  • the hybrid antigen comprises a plurality of antigenic domains.
  • at least one of the antigenic domains is a T helper epitope.
  • the complex comprises a plurality of hybrid antigens, at least one of the hybrid antigens comprising a plurality of antigenic domains.
  • the heat shock protein is a hsp70.
  • the invention is directed to a method for treating an infectious disease or cancer comprising administering to a subject at least one hybrid antigen comprising at least one antigenic domain of an infectious agent or tumor antigen associated with the infectious disease or cancer, a binding domain comprising a peptide that non- covalently binds to a heat shock protein, and a peptide linker there between, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1001); Lys Asn (KN);
  • the complex comprises a plurality of hybrid antigens.
  • at least one of the hybrid antigens is a T helper epitope.
  • the hybrid antigen comprises a plurality of antigenic domains.
  • at least one of the antigenic domains is a T helper epitope.
  • the complex comprises a plurality of hybrid antigens, at least one of the hybrid antigens comprising a plurality of antigenic domains.
  • a peptide linker separates the antigenic domain and the binding domain.
  • the invention is directed to a method for treating an infectious disease or cancer comprising administering to a subject at least one hybrid antigen, the hybrid antigen consisting essentially of at least one antigenic domain of an infectious agent or tumor antigen associated with an infectious disease or cancer, a binding domain that non-covalently binds to a heat shock protein, and a peptide linker separating the antigenic domain and the binding domain, and wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu
  • AAi is A, S, V, E, G, L, or K, preferably V, more preferably S, and most preferably A ;
  • AA2 is K, V, or E, preferably E, more preferably V and most preferably K;
  • AA 3 is V, S, F, K, A, E, or T, preferably F, more preferably S and most preferably V.
  • the complex comprises a plurality of hybrid antigens.
  • at least one of the hybrid antigens is a T helper epitope.
  • the hybrid antigen comprises a plurality of antigenic domains.
  • at least one of the antigenic domains is a T helper epitope.
  • the complex comprises a plurality of hybrid antigens, at least one of the hybrid antigens comprising a plurality of antigenic domains.
  • the invention is directed to a peptide from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1001); Lys Asn (KN); Arg Lys (RK); or AA ⁇ -AA 2 -AA 3 -leucine, wherein AAi is A, S, V, E, G, L, or K, preferably V, more preferably S, and most preferably A ; AA 2 is K, V, or E, preferably E, more preferably V and most preferably K; and AA 3 is V
  • the invention is directed to an immunogenic polypeptide comprising a plurality of antigenic domains, at least one heat shock protein binding domain and at leatst one peptide linker there between wherein at least one peptide linker is from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ LD NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1001); Lys Asn (KN); Arg Lys (RK); or AA ⁇ -AA 2 -AA 3 -leucine, wherein AAi is A, S, V, E, G, L, or K
  • Gin Leu Lys (QLK), Arg Lys (RK) and Ala Lys Val Leu (AKVL; SEQ ID NO: 1001) are preferred, and Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000) is most preferred.
  • the invention is directed to a polynucleotide encoding any of the hybrid antigens in the aforementioned first, second, third or fourth aspect.
  • the invention is directed to a method of inducing an immune response to an infectious disease or cancer comprising administering to a subject a polynucleotide encoding a hybrid antigen comprising an antigenic domain of an infectious agent or tumor antigen associated with the infectious disease or cancer, a heat shock protein binding domain, and a peptide linker there between from among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000); Phe Arg Lys (FRK); Phe Arg Lys Asn (FRKN, SEQ ID NO: 1002); Arg Lys Asn (RKN); Phe Phe Arg Lys Asn (FFRKN, SEQ ID NO: 1003); Phe Arg (FR), Gin Leu Lys (QLK), Gin Leu Glu (QLE), Ala Lys Val Leu (AKVL; SEQ ID NO: 1001); Lys Asn (KN); Arg Lys (RK); or AA ⁇ -AA 2 -AA
  • Gin Leu Lys (QLK), Arg Lys (RK) and Ala Lys Val Leu (AKVL; SEQ ID NO: 1001) are preferred, and Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000) is most preferred.
  • the invention is directed a method of inducing an immune response to an infectious disease or cancer comprising administering to a subject a polynucleotide encoding a hybrid antigen as mentioned above, and a polynucleotide encoding a heat shock protein.
  • the encoded heat shock protein is a hsp70.
  • the infectious disease antigen may be derived from an infectious agent such as a bacterium, virus, protozoan, mycoplasma, fungus, yeast, parasite, or prion, by way of non-limiting example.
  • a cancer or tumor antigen associated with cancer may be derived from a sarcoma, a lymphoma, a leukemia, or a carcinoma, a melanoma, carcinoma of the breast, carcinoma of the prostate, ovarian carcinoma, carcinoma of the cervix, colon carcinoma, carcinoma of the lung, glioblastoma, or astrocytoma, by way of non-limiting examples.
  • the antigenic domain of an infectious agent or cancer comprises an antigen derived from or associated with the infectious disease or tumor antigen, such that induction of an immune response to the antigen of the infectious agent or cancer antigen, respectively, is useful for treating the corresponding infectious disease or cancer.
  • Figure 1 shows the results of a tumor challenge study in which immunization using a hybrid antigen or complex of a hybrid antigen with a heat shock protein was performed, followed seven days later by challenge with a tumor expressing the antigen.
  • a hybrid antigen comprises at least one antigenic (immunogenic) domain, at least one heat shock protein-binding domain, and a peptide linker between at least two of these domains, wherein the peptide linker is among Phe Phe Arg Lys (FFRK; SEQ ID NO: 1000),
  • AAi-AA 2 -AA 3 -leucine wherein AAi is A, S, V, E, G, L, or K, preferably V, more preferably S, and most preferably A ; AA 2 is K, V, or E, preferably E, more preferably V and most preferably K; and AA 3 is V, S, F, K, A, E, or T, preferably F, more preferably S and most preferably V.
  • Gin Leu Lys (QLK), Arg Lys (RK) and Ala Lys Val Leu (AKVL; SEQ ID NO:1001) are preferred, and Phe Phe Arg Lys (FFRK; SEQ ID NO:1000) is most preferred.
  • the hybrid antigen serves at least two functions, namely (i) it contains an epitope capable of inducing the desired immune response; and (ii) it is capable of physically binding to a heat shock protein. As will be noted below, such binding may occur in vivo such that administration of the hybrid antigen alone will induce the desired immune response and provide the desired therapeutic effect.
  • antigen refers to a compound which may be composed of amino acids, carbohydrates, nucleic acids or lipids individually or in any combination.
  • hybrid antigen refers to a compound which binds to one or more heat shock proteins and which is representative of the immunogen toward which an immune response is desirably directed.
  • the immunogen is an influenza virus
  • the hybrid antigen may comprise a peptide fragment of the matrix protein of the influenza virus.
  • the term “immunogen” is applied to the neoplastic cell, infected cell, pathogen, or component thereof, towards which an immune response is to be elicited, whereas the hybrid antigen comprises a portion of that immunogen which can provoke the desired response and which binds to one or more heat shock proteins.
  • the antigenic domain of the hybrid antigen is selected to elicit an immune response to a particular disease or pathogen, including peptides obtained from MHC molecules, mutated DNA gene products, and direct DNA products such as those obtained from tumor cells.
  • immunogens of particular interest are those associated with, derived from, or predicted to be associated with a neoplastic disease, including but not limited to a sarcoma, a lymphoma, a leukemia, or a carcinoma, and in particular, with melanoma, carcinoma of the breast, carcinoma of the prostate, ovarian carcinoma, carcinoma of the cervix, colon carcinoma, carcinoma of the lung, glioblastoma, astrocytoma, etc. Selections of melanoma antigens useful in hybrid antigens of the present invention may be found, by way of non-limiting example, in
  • the immunogen may be associated with an infectious disease, and, as such, may be a bacterium, virus, protozoan, mycoplasma, fungus, yeast, parasite, or prion.
  • the immunogen may be a human papilloma virus (see below), a herpes virus such as herpes simplex or herpes zoster, a retrovirus such as human immunodeficiency virus 1 or 2, a hepatitis virus, an influenza virus, a rhinovirus, respiratory syncytial virus, cytomegalovirus, adenovirus, Mycoplasma pneutnoniae, a bacterium of the genus Salmonella, Staphylococcus, Streptococcus,
  • Immunogens may be obtained by isolation directly from a neoplasm, an infected cell, a specimen from an infected subject, a cell culture, or an organism culture, or may be synthesized by chemical or recombinant techniques.
  • suitable antigenic peptides particularly for use in a hybrid antigen, for use against viruses, bacteria and the like can be designed by searching through their sequences for MHC class I restricted peptide epitopes containing HLA binding sequences such as but not limited to
  • HLA-A2 peptide binding sequences HLA-A2 peptide binding sequences:
  • Ser Tyr Val Pro Ser Ala Glu Gin lie (SEQ ID NO: 136); from cancers: Phe Glu Gin Asn Thr Ala Gin Pro(SEQ ID NO: 137);
  • epitopes are merely exemplary of selections available associated with various infectious diseases and cancer, and are provided without any intention whatsoever to be limiting.
  • a humoral immune response may be appropriate.
  • a cellular immune response is particularly desirable. Accordingly, particular epitopes associated with the activation of B cells, T helper cells, or cytotoxic T cells may be identified and selected for incorporation into the hybrid antigen.
  • hybrid antigen associated with an autoimmune disease or allergy may also be desirable to utilize hybrid antigen associated with an autoimmune disease or allergy.
  • a hybrid antigen may be administered, together with one or more heat shock proteins, in an amount sufficient to be tolerogenic or to inhibit a pre-existing immune response to the hybrid antigen in a subject.
  • the amount of heat shock protein required to inhibit the immune response is expected to be substantially greater than the amount required for stimulation.
  • the size of hybrid antigen may vary depending upon the heat shock protein used, in non-limiting embodiments of the invention, the hybrid antigen may be the size of a peptide having between 10 and 500 amino acid residues, and preferably be the size of a peptide having between 14 and 100, most preferably 18 and 50 amino acid residues. As such, it may be desirable to produce a fragment of an immunogen to serve as the antigenic domain of a hybrid antigen, or, alternatively, to synthesize a hybrid antigen by chemical or recombinant DNA methods.
  • a hybrid antigen may be prepared, and then tested for its ability to bind to heat shock protein, hi some instances, binding of hybrid antigen to a particular heat shock protein may be facilitated by the presence of at least one other protein, which may be a heat shock protein.
  • binding of hybrid antigen to a heat shock protein may be evaluated by labeling the hybrid antigen with a detectable label, such as a radioactive, fluorescent, enzymatic or pigmented label, combining the hybrid antigen with heat shock protein under conditions which would be expected to permit binding to occur, and then isolating the heat shock protein while removing any unbound hybrid antigen, and determining whether any labeled hybrid antigen had adhered to the heat shock protein.
  • a detectable label such as a radioactive, fluorescent, enzymatic or pigmented label
  • the ability of a hybrid antigen to bind to the heat shock protein BiP may be evaluated by combining 2 ⁇ g BiP with up to about 1150 pmole of radioactively labeled hybrid antigen in buffer containing 50 mM Tris HC1 (pH 7.5), 200 mM NaCl, and 1 M Na 2 EDTA, in a final volume of 50 ⁇ l, for 30 minutes at 37 degrees Centigrade. Unbound hybrid antigen may then be removed from bound BiP-hybrid antigen by centrifugation at lOOg by desalting through a 1 ml Sephadex-G column for 2 minutes. Penefsky, J. Biol. Chem. 252:2891 (1977).
  • columns may first be treated with 100 ⁇ l of bovine serum albumin in the same buffer and centrifuged as above. Bound hybrid antigen may then be quantitated by liquid scintillation counting. See Flynn et al., Science 245:385-390 (1989).
  • ATP hydrolysis drives the release of peptides from many known heat shock proteins
  • the amount of ATPase activity may often be used to quantitate the amount of hybrid antigen binding to heat shock protein.
  • An example of how such an assay may be performed is set forth in Flynn et al., Science 245:385-390 (1989).
  • the heat shock protein-binding domain is selected so that the hybrid antigen will bind in vitro or in vivo to a heat shock protein such as BiP, hsp70, gp96, or hsp90, or a member of the foregoing heat shock protein families, alone or in combination with accessory heat shock proteins such as hsp40, or hsp60.
  • a heat shock protein such as BiP, hsp70, gp96, or hsp90
  • a member of the foregoing heat shock protein families alone or in combination with accessory heat shock proteins such as hsp40, or hsp60.
  • Non-limiting examples of peptides which fulfill this criterion maybe identified by panning libraries of antigens known to bind well to one or more heat shock proteins as described in Blond-Elguindi et al., Cell 75:717-728 (1993):
  • Trp Asp Asp Leu Leu His Gly Arg (SEQ ID NO: 170);
  • Hy(T ⁇ /X)HyXHyXHy (SEQ ID NO: 29) where Hy represents a hydrophobic amino acid residue, particularly tryptophan, leucine or phenylalanine (SEQ ID NO:30), and X is any amino acid.
  • High affinity heat-shock protein- binding sequences inco ⁇ orating this motif include:
  • Heat shock binding motifs have also been identified as consisting of seven to fifteen residue long peptides which are enriched in hydrophobic amino acids.
  • heat shock protein binding peptides include: Gly Lys T ⁇ Val Tyr He (SEQ ID NO:295);
  • heat shock protein binding domains include Phe Tyr Gin Leu Ala Leu Thr(SEQ ID NO: ), Phe Tyr Gin Leu Ala Leu Thr T ⁇ (SEQ ID NO: ), Arg Lys Leu Phe Phe Asn Leu Arg (SEQ ID NO: ), Arg Lys Leu Phe Phe Asn Leu Arg T ⁇ (SEQ DO NO: ), Lys Phe Glu Arg Gin (SEQ DO NO: ), Asn He Val Arg Lys Lys Lys (SEQ ID NO: ), and Arg Gly Tyr Val Tyr Gin Gly Leu (SEQ ID NO: ).
  • heat shock protein binding domains include those described in W09922761.
  • Xaa represents any amino acid.
  • TETPYPTG (SEQ ID NO: 83);
  • LTTPFSSG (SEQ ID NO: 84);
  • GVPLTMDG (SEQ ID NO: 85);
  • KLPTVLRG (SEQ ID NO: 86); CRFHGNRG (SEQ ID NO: 87); YTRDFEAG (SEQ ID NO: 88);
  • SSAAGPRG SEQ TD NO: 89
  • DALMWP Xaa G (SEQ DO NO: 91); SS Xaa SLYIG (SEQ ID NO: 92);
  • VGSMESLG SEQ DO NO: 96
  • F Xaa PMI Xaa SG SEQ ID NO: 97
  • KPPLFQIG (SEQ ID NO: 100);
  • TTPPNFAG SEQ DO NO: 104
  • ISLDPRMG SEQ ID NO: 105
  • SLPLFGAG (SEQ ID NO: 106); NLLKTTLG (SEQ DO NO: 107);
  • DQNLPRRG (SEQ ID NO: 108);
  • TPQLHHGG (SEQ ID NO: 110);
  • APLDRITG SEQ ID NO: 111
  • FAPLIAHG SEQ ID NO: 112
  • SWIQTFMG (SEQ ID NO: 113);
  • NTWPHMYG (SEQ DO NO: 114);
  • EPLPTTLG (SEQ ID NO: 115);
  • HGPHLFNG (SEQ ID NO: 116); YLNSTLAG (SEQ ID NO: 117);
  • HLHSPSGG (SEQ ID NO: 118);
  • TLPHRLNG (SEQ ID NO: 119); SSPREVHG (SEQ DO NO: 120);
  • NQVDTARG (SEQ TD NO: 121);
  • HPAAFPWG SEQ DO NO: 123
  • LLPHSSAG SEQ ID NO: 124
  • KYVPLPPG SEQ ID NO: 126
  • APLALHAG (SEQ TD NO: 127);
  • GLATVKSG (SEQ TD NO: 130);
  • GATSFGLG (SEQ ID NO: 131);
  • KPPGPVSG (SEQ ID NO: 132);
  • TLYVSGNG (SEQ ID NO: 133); HAPFKSQG (SEQ ID NO: 134);
  • VAFTRLPG SEQ ID NO: 1357
  • RMNTEPPG SEQ TD NO: 138
  • KMTPLTTG SEQ TD NO: 139
  • ANATPLLG (SEQ TD NO: 140);
  • APLALHAG (SEQ ID NO: 146);
  • TAHDLTVG (SEQ ID NO: 147);
  • NMTNMLTG SEQ DO NO: 148
  • GSGLSQDG SEQ TD NO: 149
  • TPD TIYG (SEQ ID NO: 150);
  • SHLYRSSG (SEQ ID NO: 151); YTLVQPL (SEQIDNO: 152);
  • TPDITPK (SEQIDNO: 153);
  • TYPDLRY (SEQI NO: 154);
  • MAPSPPH SEQ DO NO: 160
  • SSFPDLL SEQ TD NO: 161
  • HLTHSQR (SEQIDNO: 163)
  • FATHHIG (SEQIDNO: 165); SMPEPLI (SEQ ID NO: 166);
  • IPRYHLI (SEQ ID NO: 167);
  • SAPHMTS SEQIDNO: 168
  • KAPVWAS (SEQIDNO: 169);
  • LPHWLLI SEQ HO NO: 170
  • ASAGYQI SEQIDNO: 171
  • VTPKTGS (SEQIDNO: 172);
  • EHPMPVL (SEQ ID NO: 173)
  • VSSFVTS (SEQ ID NO: 174);
  • STHFTWP (SEQ TD NO: 175); GQWWSPD (SEQ TD NO: 176);
  • GPPHQDS (SEQIDNO: 177);
  • NTLPSTI SEQIDNO: 178
  • YGNPLQP (SEQIDNO: 180); FHWWWQP (SEQIDNO: 181);
  • FHWPWLF (SEQ ID NO: 183); TAQDSTG (SEQ DO NO: 184);
  • FHWWWQP (SEQ DO NO: 185);
  • EPFFRMQ (SEQ HD NO: 187); TWWLNYR (SEQ DO NO: 188);
  • FHWWWQP (SEQ ID NO: 189);
  • FHWWWQP (SEQ ID NO: 192); HPSNQAS (SEQ ID NO: 193);
  • SWPFFDL (SEQ TD NO: 196);
  • TSPLSLL (SEQ ID NO: 200);
  • RPLHDPM SEQ TD NO: 202
  • WPSTTLF SEQ ID NO: 203
  • ATLEPVR (SEQ ID NO: 204);
  • APDLYVP (SEQ ⁇ ) NO: 207); RMPPLLP (SEQ ID NO: 208);
  • TPPLRIN SEQ DO NO: 210
  • DLNAYTH (SEQ ID NO: 212); VTLPNFH (SEQ ID NO: 213);
  • NSRLPTL (SEQ DO NO: 214);
  • YPHPSRS (SEQ ID NO: 215); GTAHFMY (SEQ DO NO: 216);
  • TSTLLWK (SEQ ID NO: 219); TSDMKPH (SEQ ID NO: 220);
  • NLYGPHD (SEQ ID NO: 222);
  • AYKSLTQ SEQ ID NO: 224
  • STSVYSS SEQ DO NO: 225
  • EGPLRSP SEQ ID NO: 226
  • TTYHALG SEQ TD NO: 227
  • THSHRPS SEQ ID NO: 229
  • ITNPLTT SEQ ID NO: 230
  • SLWTRLP SEQ DO NO:2344
  • NVYHSSL SEQ ro NO:235
  • NSPHPPT SEQ DO NO:2336
  • HNLHPNR SEQ ID NO:2378
  • YTTHRWL (SEQ ID NO:239); AVTAAIV (SEQ ID NO:240);
  • SHVPSMA (SEQ ID NO:244); HTTVYGA (SEQ ID NO:245);
  • GVPLTMD (SEQ ro NO:248)
  • KLPTVLR (SEQ ID NO:249)
  • YTRDFEA (SEQ ID NO: 251); SSAAGPR (SEQ DO NO: 252);
  • FNTSTRT (SEQ ID NO: 256); TVQHVAF (SEQ ID NO: 257);
  • APPRVTM (SEQ ID NO: 261); IATKTPK (SEQ ID NO: 262);
  • SSFATFL SEQ ID NO: 266
  • TTPPNFA SEQ ID NO: 267
  • NLLKTTL (SEQ ID NO: 270);
  • DQNLPRR SEQ ID NO: 271
  • SHFEQLL SEQ ID NO: 272
  • FAPLIAH (SEQ ID NO: 275);
  • SWIQTFM (SEQ DO NO: 276); NTWPHMY (SEQ TD NO: 277);
  • EPLPTTL (SEQ ID NO: 278);
  • HGPHLFN (SEQ TD NO: 279); YLNSTLA (SEQ ID NO: 280);
  • HPAAFPW SEQ DO NO: 286)
  • LETYTAS SEQ ID NO: 288
  • KYVPLPP SEQ ID NO: 289
  • APLALHA (SEQ ID NO: 290);
  • GLATVKS SEQ ro NO: 293
  • GATSFGL SEQ ID NO: 294
  • TLYVSGN SEQ ID NO: 296
  • HAPFKSQ SEQ ID NO: 297
  • VAFTRLP SEQ ID NO: 298
  • LPTRTPA SEQ ID NO: 299
  • ASFDLLI SEQ ID NO: 300
  • KMTPLTT (SEQ ro NO: 302)
  • ANATPLL (SEQ ID NO: 303); TIWPPPV (SEQ ID NO: 304);
  • NHAVFAS SEQ ro NO: 306
  • TWQPYFH (SEQ ID NO: 308); APLALHA (SEQ ID NO: 309);
  • TAHDLTV (SEQ ID NO: 310);
  • NMTNMLT (SEQ ID NO: 311); GSGLSQD (SEQ DO NO: 312); TP ⁇ CTIV (SEQ DO NO: 313); SHLYRSS (SEQ DO NO: 314); HGQAWQF (SEQ DO NO: 315); and FHWWW (SEQ ID NO: 317).
  • the aforementioned heat shock protein binding domains are merely exemplary of various peptides, among peptide and non-peptide heat shock protein binding molecules, that may be used in the practice of the present invention.
  • the heat shock protein binding domain may be directed to bind to a different part of the mammalian heat shock protein that those aforementioned, and the heat shock protein-binding domains of the invention are not limited to binding to any particular portion of the heat shock protein molecule.
  • the peptide ⁇ FAGDCKKAERADLIAYLKQATAK (Greene et al., 1995, J. Biol. Chem.
  • a heat shock protein-binding fragment of this peptide is used in any of the conjugates of the invention to facilitate the binding of a pre-selected molecule to a heat shock protein.
  • the binding may be achieved through the use of an organic molecule or compound with heat shock protein binding activity.
  • suitable molecules include members of the benzoquinone ansamycin antibiotics, such as herbimycin A, geldanamycin, macmimycin I, mimosamycin, and kuwaitimycin (Omura et al., 1979, J.
  • heat shock protein binding domains those preferred in the present invention as part of a hybrid antigen comprising an antigenic domain and peptide linker of the invention there between includes the following heat shock protein binding domains:
  • heat shock protein binding domains with a terminal T ⁇ residue useful for the various aspects of the present invention include: [0064] Asn Leu Leu Arg Leu Thr Gly T ⁇ (SEQ TD NO:350);
  • heat shock protein binding domains useful in the practice of the present invention include Phe Tyr Gin Leu Ala Leu Thr T ⁇ (SEQ ID NO: 501), Phe Tyr Gin Leu Ala Leu Thr T ⁇ (SEQ ID NO:502 ), Arg Lys Leu Phe Phe Asn Leu Arg T ⁇ (SEQ ED NO:503 ), Arg Lys Leu Phe Phe Asn Leu Arg T ⁇ (SEQ HD NO:504 ), Lys Phe Glu Arg Gin T ⁇ (SEQ ID NO:505), Asn He Val Arg Lys Lys Lys T ⁇ (SEQ ID NO:506), and Arg Gly Tyr Val Tyr Gin Gly Leu T ⁇ (SEQ ID NO:507).
  • other heat shock protein binding domains include those described in W09922761, and may have a terminal T ⁇ residue added to achieve the pmposes of the present invention.
  • Xaa represents any amino acid.
  • the heat shock protein binding domain Asn Leu Leu Arg Leu Thr Gly T ⁇ (SEQ ID NO: 350) is most preferred in the hybrid antigens of the invention.
  • the aforementioned heat shock protein binding domains are merely exemplary of various moieties, among peptide and non- peptide heat shock protein binding molecules, that may be used in the practice of the present invention.
  • the hybrid antigen of the invention inco ⁇ orates at least one antigenic (immunogenic) domain and at least one one heat shock protein-binding domain, separated by at least one peptide linker as described herein.
  • the hybrid antigen of the invention may be synthesized using chemical peptide synthesis methods or it can be synthesized by expression of a nucleic acid construct containing linked sequences encoding the antigenic and heat shock protein binding domains.
  • One suitable technique utilizes initial separate PCR amplification reactions to produce separate DNA segments encoding the two domains, each with a linker segment attached to one end, followed by fusion of the two amplified products in a further PCR step. This technique is refe ⁇ ed to as linker tailing.
  • Suitable restriction sites may also be engineered into regions of interest, after which restriction digestion and ligation is used to produce the desired hybrid antigen-encoding sequence.
  • nucleic acid encoding a hybrid antigen of the invention is also suitable for therapeutic use by administration to the subject, where expression in vivo yields the hybrid antigen with the ability of inducing an immune response.
  • heat shock protein refers to any protein which exhibits increased expression in a cell when the cell is subjected to a stress.
  • the heat shock protein is originally derived from a eukaryotic cell; in more preferred embodiments, the heat shock protein is originally derived from a mammalian cell.
  • heat shock proteins which may be used according to the invention include BiP (also referred to as g ⁇ 78), hsp70, hsc70, gp96 (g ⁇ 94), hsp ⁇ O, hsp40, and hsp90, and members of the families thereof.
  • heat shock proteins are BiP, gp96, and hsp70, as exemplified below. Most preferred is a member of the hsp70 family.
  • Naturally occurring or reeombinantly derived mutants of heat shock proteins may also be used according to the invention.
  • the present invention provides for the use of heat shock proteins mutated so as to facilitate their secretion from the cell (for example having mutation or deletion of an element which facilitates endoplasmic reticulum recapture, such as KDEL or its homologues; such mutants are described in PCT Application No. PCT/US96/13233 (WO 97/06685), which is inco ⁇ orated herein by reference).
  • the heat shock protein may be prepared, using standard techniques, from natural sources, for example as described in Flynn et al., Science 245:385-390 (1989), or using recombinant techniques such as expression of a heat shock encoding vector in a suitable host cell such as a bacterial, yeast or mammalian cell. If pre-loading of the heat shock protein with peptides from the host organism is a concern, the heat shock protein can be incubated with ATP and then repurif ⁇ ed.
  • suitable host cell such as a bacterial, yeast or mammalian cell.
  • a nucleic acid encoding a heat shock protein may be operatively linked to elements necessary or desirable for expression and then used to express the desired heat shock protein as either a means to produce heat shock protein for use in a protein vaccine or, alternatively, in a nucleic acid vaccine.
  • Elements necessary or desirable for expression include, but are not limited to, promoter/enhancer elements, transcriptional start and stop sequences, polyadenylation signals, translational start and stop sequences, ribosome binding sites, signal sequences and the like.
  • genes for various heat shock proteins have been cloned and sequenced, including, but not limited to, gp96 (human: Genebank Accession No.
  • hybrid antigens of the invention or complexes of hybrid antigens and heat shock proteins may be administered to a subject using either a peptide-based, protein-based or nucleic acid vaccine, so as to produce, in the subject, an amount of complex which is effective in inducing a therapeutic immune response in the subject.
  • the subject may be a human or nonhuman subject.
  • the term "therapeutic immune response,” as used herein, refers to an increase in humoral and/or cellular immunity, as measured by standard techniques, which is directed toward the hybrid antigen.
  • the induced level of humoral immunity directed toward hybrid antigen is at least four-fold, and preferably at least 16-fold greater than the levels of the humoral immunity directed toward the antigen prior to the administration of the compositions of this invention to the subject.
  • the immune response may also be measured qualitatively, by means of a suitable in vitro or in vivo assay, wherein an arrest in progression or a remission of neoplastic or infectious disease in the subject is considered to indicate the induction of a therapeutic immune response.
  • heat shock protein hybrid antigen administered may depend on numerous factors including the immunogenicity of the particular vaccine composition, the immunocompetence of the subject, the size of the subject and the route of administration. Determining a suitable amount of any given composition for administration is a matter of routine screening. [0077] Furthermore, significant immunological efficacy was identified in studies in which the hybrid antigen was administered alone, i.e., without heat shock protein. While Applicants have no duty to disclose the theory by which the invention operates, and are not bound thereto, the results of these studies suggest that the hybrid antigens, upon injection into the subject, bind to endogenous heat shock proteins, and thus do not require the concomitant administration of heat shock protein for effectiveness.
  • the present invention extends to such utilities of the hybrid antigens of the invention, and moreover, to concomitant therapies or treatments that increase endogenous heat shock protein levels systemically or at the intended site of administration of the hybrid antigens of the invention.
  • concomitant therapies or treatments include but are not limited to local application of heat or local or systemic pharmaceutical agents that increase the expression of heat shock protein in the local tissue. Such agents and methods are known in the art.
  • Hybrid antigens that are administered in the absence of co-administration of a heat shock protein i.e., administered not in a complex with a heat shock protein
  • that comprise at least one antigenic domain and at least one heat shock protein binding domain comprise one of the peptide linkers mentioned hereinabove.
  • a hybrid antigen of the invention may include more than one immunogenic domain or more than one epitope.
  • compositions comprising hybrid antigen/heat shock protein or hybrid antigen alone as set forth above are referred to herein as "vaccines.”
  • the term vaccine is used to indicate that the compositions of the invention may be used to induce a prophylactic or therapeutic immune response.
  • a vaccine of the invention may comprise a hybrid antigen with a single antigenic domain or epitope, or a hybrid antigen with a plurality of antigenic domains or epitopes. Further, a vaccine may comprise an admixture of hybrid antigens with single or pluralities of antigenic domains or epitopes, or any combination of the foregoing.
  • the hybrid antigens or admixtures thereof may be complexed with one or more heat shock proteins before administration, or may be administered without heat shock protein.
  • a vaccine composition comprising one or more hybrid antigens optionally complexed to one or more heat shock proteins in accordance with the invention may be administered cutaneously, subcutaneously, intradermally, intravenously, intramuscularly, parenterally, intrapulmonarily, intravaginally, intrarectally, nasally or topically.
  • the vaccine composition may be delivered by injection, particle bombardment, orally or by aerosol.
  • Incubation with heating of the heat shock protein with the hybrid antigen will in general lead to loading of the antigen onto the heat shock protein.
  • hsp40 can facilitate loading of peptides onto hsp70.
  • Denaturants such as guanidinium HC1 or urea can be employed to partially and reversibly destabilize the heat shock protein to make the peptide binding pocket more accessible to the antigen.
  • a vaccine of the invention comprising a heat shock protein preferably also includes adenosine diphosphate (ADP), to promote the association between the heat shock protein and the heat shock protein binding domain prior to the complex reaching its destination.
  • ADP adenosine diphosphate
  • Other compounds with similar capabilities may used, alone or in combination with ADP.
  • Vaccine compositions in accordance with the invention may fiirther include various additional materials, such as a pharmaceutically acceptable carrier.
  • suitable carriers include any of the standard pharmaceutically accepted carriers, such as phosphate buffered saline solution, water, emulsions such as an oil/water emulsion or a triglyceride emulsion, various types of wetting agents, tablets, coated tablets and capsules.
  • An example of an acceptable triglyceride emulsion useful in intravenous and intraperitoneal administration of the compounds is the triglyceride emulsion commercially known as Intralipid®.
  • Such carriers typically contain excipients such as starch, milk, sugar, certain types of clay, gelatin, stearic acid, talc, vegetable fats or oils, gums, glycols, or other known excipients.
  • excipients such as starch, milk, sugar, certain types of clay, gelatin, stearic acid, talc, vegetable fats or oils, gums, glycols, or other known excipients.
  • Such carriers may also include flavor and color additives or other ingredients.
  • the vaccine composition of the invention may also include suitable diluents, preservatives, solubilizers, emulsifiers, adjuvants and/or carriers.
  • suitable diluents may be in the form of liquid or lyophilized or otherwise dried formulations and may include diluents of various buffer content (e.g., Tris-HCl, acetate, phosphate), pH and ionic strength, additives such as albumin or gelatin to prevent abso ⁇ tion to surfaces, detergents (e.g., Tween 20, Tween 80, Pluronic F68, bile acid salts), solubilizing agents (e.g.
  • glycerol polyethylene glycerol
  • anti-oxidants e.g., ascorbic acid, sodium metabisulfite
  • preservatives e.g., Thimerosal, benzyl alcohol, parabens
  • bulking substances or tonicity modifiers e.g., lactose, mannitol
  • covalent attachment of polymers such as polyethylene glycol to the protein, complexing with metal ions, or inco ⁇ oration of the material into or onto particulate preparations of polymeric compounds such as polylactic acid, polyglycolic acid, hydrogels, etc.
  • compositions will influence the physical state, solubility, stability, rate of in vivo release, and rate of in vivo clearance.
  • the choice of compositions will depend on the physical and chemical properties of the vaccine. For example, a product derived from a membrane-bound form of a protein may require a formulation containing detergent.
  • Controlled or sustained release compositions include formulation in lipophilic depots (e.g. fatty acids, waxes, oils). Also comprehended by the invention are particulate compositions coated with polymers (e.g.
  • compositions of the invention inco ⁇ orate particulate forms protective coatings, protease inhibitors or permeation enhancers for various routes of administration, including intramuscular, parenteral, pulmonary, nasal and oral.
  • one or more polynucleotide constructs may be administered which encode the hybrid antigen, optionally with heat shock protein, in expressible form.
  • the expressible polynucleotide constructs are introduced into cells in the subject using ex vivo or in vivo methods. Suitable methods include injection directly into tissue and tumors, transfecting using liposomes (Fraley et al., Nature 370:111-117 (1980)), receptor-mediated endocytosis (Zatloukal et al., Ann. NY Acad. Sci.
  • polynucleotide vaccine may also be introduced into suitable cells in vitro which are then introduced into the subject.
  • an expressible polynucleotide a region encoding the heat shock protein and/or hybrid antigen is prepared as discussed above and inserted into a mammalian expression vector operatively linked to a suitable promoter such as the SV40 promoter, the cytomegalovirus (CMV) promoter or the Rous sarcoma virus (RSV) promoter.
  • a suitable promoter such as the SV40 promoter, the cytomegalovirus (CMV) promoter or the Rous sarcoma virus (RSV) promoter.
  • CMV cytomegalovirus
  • RSV Rous sarcoma virus
  • the nucleic acid polymer(s) could also be cloned into a viral vector.
  • Suitable vectors include but are not limited to refroviral vectors, adenoviras vectors, vaccinia virus vectors, pox virus vectors and adenovirus-associated vectors.
  • Specific vectors which are suitable for use in the present invention are pCDNA3 (InVitrogen), plasmid AH5 (which contains the SV40 origin and the adenovirus major late promoter), pRC/CMV (InVitrogen), pCMU II (Paabo et al., EMBO J. 5:1921-1927 (1986)), pZip-Neo SV (Cepko et al., Cell 37:1053-1062 (1984)) and pSR ⁇ (DNAX, Palo Alto, CA).
  • EXAMPLE 1 A variety of hybrid antigens were prepared, each comprising a heat shock protein binding domain and a cancer antigen epitope or the model Class I H2-K epitope from ovalbumin, SIINFEKL. A peptide linker was included between the two domains.
  • the heat shock protein binding domains used in these experiments were among the following: HWDFAWPW, NLLRLTGW, FYQLALTW and RKLFFNLRW. Linkers were among those desceibed hereinabove.
  • the cancer and model epitopes were among the following:
  • hybrid antigens comprising a heat shock protein binding domain, a cancer epitope, and a linker there between, were synthesized, in various orientations.
  • binding affinities between recombinant human or murine heat shock protein 70 (hsp70) and the various heat shock protein binding domains and antigenic peptides mentioned above, as well as between the hybrid antigens comprising an antigenic peptide and a heat shock protein binding domain described above, were determined by a binding inhibition assays (Hill plots) relative to the binding affinity of a reference, labeled hybrid antigen (tritiated or fluoresceinated ALFDIESKVGSGHWDFAWPW) to hsp70 as determined by Scatchard analysis (Kds of 22.64 ⁇ M and 10.75 ⁇ M, respectively). Binding studies were performed in 39% PBS; 20 mM THAM, pH 8; 37 mM NaCl, 5 mM MgCl 2 ; and l mM ADP.
  • EXAMPLE 3 For immunological studies in mice, a murine MHC H2-K(b) epitope from ovalbumin, SIINFEKL (amino acids 257-264), and a H2-K(b) peptide from the nucleoprotein of vesicular stomatitis virus (VSV), RGYVYQGL (amino acids 52-59) were used for the preparation of hybrid antigens.
  • VSV vesicular stomatitis virus
  • RGYVYQGL amino acids 52-59
  • mice were immunized s.c. at the base of the tail with hsp70 alone, hsp70 complexed with SIINFEKL, and hybrid SIINFEKL peptide with or without HSP70. The doses were adjusted such that each immunization contained the same amount of SIINFEKL, except for hsp70 alone. Seven days later, spleens were harvested and enriched for CD8+ T cells, which were put into an ex vivo IFN- ⁇ ELISPOT assay.
  • SIINFEKL SIINFEKL Responses after pulsing with SIINFEKL (“SIINFEKL”) were recorded in the following table, which includes the doses, and the number of spots (mean ⁇ standard error) per 4 x 10 5 CD8 T cells, of > four experiments with at least three mice per group.
  • VSV epitope used as a control in many of the foregoing experiments was used as the epitope in preparing further hybrid antigens of the invention, and evaluated for induction of an immune response in similar experiments as described above.
  • E.G7 E7 tumor cells modified to express ovalbumin
  • mice immunized with hsp70 NLLRLTGWFFRKSIINFEKL developed tumors, nor did mice immunized with SIINFEKL emulsified in Titermax adjuvant.
  • Three of 10 mice vaccinated with NLLRLTGWFFRKSIINFEKL alone had tumors.
  • the in-vitro antigen presentation assay described above was utilized further in order to evaluate the formulations of the invention.
  • the assay was performed with the following formulations, with the results indicated.
  • the "EMD" peptide epitope IMDQVPFSV from the human melanoma antigen gplOO was evaluated in a hybrid antigen of the invention at low and high dose in the HHD II model.
  • test peptides being the IMD peptide and, as a control, a peptide from the melanoma antigen tyrosinase, YMDGTMSQV ("YMD").
  • YMD a peptide from the melanoma antigen tyrosinase
  • Hybrid antigens containing SIINFEKL from ovalbumin and RGYVYQGL from VSV were admixed and immunized with hsp70. The results are as follow.
  • formulations containing a plurality of hybrid antigens comprising different antigenic epitopes may be formulated with one or more heat shock proteins for immunization in humans in order to elicit an effective immune response to treat or prevent a disease.
  • a formulation comprising 8 different melanoma epitopes may be prepared as hybrid antigens, and formulated, for example, with hsp70.
  • the heat shock protein binding domain NLLRLTGW at the N-terminus is used for all epitopes, linked to the epitope at the C-terminus using the peptide linker FFRK.
  • Other binding domains and linkers are embraced herein.
  • This particular formulation is useful for treating patients with the HLA-A2 haplotype.
  • a formulation comprises the following hybrid antigens with hsp70:
  • approximately equal amounts of the foregoing 8 hybrid antigens may be complexed with hsp70, and administered in saline.
  • a formulation comprises the first five hybrid antigens listed.
  • the aforementioned formulations containing heat shock protein in saline optionally may contain ADP to stabilize the complexes, as well as other components, such as excipients, diluents and carriers, as mentioned above.
  • an admixture of the foregoing 8 hybrid antigens, or the first 5 listed is formulated in saline for administration without a heat shock protein.
  • hybrid antigens containing SIINFEKL and the VSV peptide RGYVYQGL were used. (VSV/OVA-72-02)
  • hybrid antigens comprising heat shock protein binding domain NLLRLTGW, antigenic domain SIINFEKL (from ovalbumin) or RGYVYQGL (from VSV protein) and various linkers set forth in Example 32 were carried out as described in Example 17.
  • the antigenic domains alone had a Kd for hsp70 binding of 235 ⁇ M and 82 ⁇ M, respectively. The results are shown below.
  • Hybrid antigens were prepared comprising two antigens, separated by a linker as described above, such that the hybrid antigen has the following general structure:
  • the heat shock protein binding domain is at the N-terminal portion of the hybrid antigen, this is not necessarily the case and hybrid antigens with the heat shock protein binding domain at the C-terminus, or in-between the two antigenic domains, is embraced by the present invention.
  • the same linker peptide is used between the antigenic domains and between the antigenic domain proximal to the heat shock protein binding domain, this is not necessarily the case and different linker peptides may be used.
  • the presence of the linker in one or both positions is optional.
  • three or more antigenic peptides may be used.
  • such hybrid antigens with two or more antigenic domains is termed a tandem hybrid antigen.
  • tandem hybrid antigen compositions complexes of one or more tandem hybrid antigens and a heat shock protein, and methods of eliciting an immune response or preventing or treating a disease by administering one or more tandem hybrid antigens or complexes of at least one heat shock protein and at least one tandem hybrid antigen are fully embraced herein.
  • the following experiments compare the immunogenicity of the admixture of two hybrid antigens and a tandem hybrid antigen comprising the same antigens, and a dose response study. In one experiment, a peptide comprising two linkers and epitopes but no heat shock protein binding domain was included.
  • the epitope proximal to the heat shock protein binding domain exhibited the strongest immune response, and thus the positioning of the selected epitopes selected for the vaccine formulations of the invention may be positioned to contribute maximally to the overall immunogenicity of the formulation, whether administered in the absence of heat shock proteins or administered as complexes with heat shock proteins.
  • tandem hybrid antigens were evaluated for immunogenicity.
  • H2-K b Class I peptides from ovalbumin (SIINFEKL) and from VSV (RGYVYQGL)
  • H2-K b ⁇ -casein peptide IAYFYPEL and the Sendai virus peptide FAPGNYPAL were also used.
  • two tandem hybrid antigens with the same antigenic peptides in alternate configurations were admixed. Strong immune responses to four epitopes were elicited. All of the formulations herein included 1 mM ADP. In one experiment described below, ADP was omitted.
  • antigenic peptides are delivered and induce immunogenicity without co-administered HSP70, when administered as an admixture of two tandem hybrid antigens and a single hybrid antigen to B6 mice.
  • the tandem hybrid antigens included VSV and ovalbumin peptides in one, and ⁇ -casein and Sendai virus peptides in the other.
  • the single hybrid antigen contained NS2-114 influenza peptide (RTFSFQLI).
  • the immunogenicity of the foregoing single hybrid antigens administered without heat shock protein were evaluated in combination with helper T cell epitopes present in a hybrid antigen.
  • a H2-K b Class II epitope from ovalbumin, amino acids 323- 339, TEWTSSNVMEERKIKV was used (i.e., the hybrid antigen had a sequence of NLLRLTGWFFRKTEWTSSNVMEERKLKV).
  • Inclusion of the Class II peptide- containing hybrid antigen increased the response to the Class I epitope on the average of about seven fold.
  • the Class I peptides were either SSWDFITV or DAPIYTNV;
  • Class II peptides 10 included the ovalbumin peptide mentioned above, a Class II peptide from tetanus toxoid NNFTVSFWLRVPKVSASHL (i.e., the hybrid antigen has a sequence of NLLRLTGWFFRKNNFTVSFWLRVPKVSASHL), or a HBVc (amino acids 128-140) peptide, TPPAYRPPNAPIL. 250-72-13
  • helper T cell epitope may be included in a hybrid antigen as the only epitope, and administered as an admixture with other hybrid antigens containing Class I epitope(s), or the helper T cell epitope can be included in a tandem hybrid antigen as one of the epitopes.
  • hybrid antigens comprising a helper T cell epitope co- administered with at least one tandem hybrid antigen, in the absence of co-administration of a heat shock protein, were also carried out.
  • HLA-A2 human Class I epitopes
  • ELISPOT results in cells per 300,000 were: Medium, 1.33 ⁇ 0.58; splenocytes 1 ⁇ 0; splenocytes plus YMDGTMSQV 123 ⁇ 13; and splenocytes plus IMDQVPFSV 4 ⁇ 1.
  • HHDII mice an immunogenic HLA-A2 epitope from T ⁇ -2 was used (SVYDFFVWL). Because this epitope is also a H2-Kb epitope, and the HHDII mice are on a B6 mouse (H2-Kb) background, an immune response induced against the T ⁇ - 2 peptide represents a breaking of tolerance to a self-epitope in the mouse model. The results of this experiment demonstrated that tolerance to this self-epitope was broken, and the present invention is further directed to methods of breaking tolerance by administering the hybrid antigens and complexes of the invention.
  • HHDII mice were used to evaluate the immunogenicity of complexes of hsp70 and three hybrid antigens comprising certain of the HEV viral component epitopes set forth in Example 27.
  • Admixtures of hybrid antigens containing H2-Kb epitopes complexed with hsp70 were evaluated for immunogenicity in B6 mice as described above.

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Abstract

La présente invention concerne des antigènes hybrides comportant au moins un domaine antigénique, au moins un domaine de liaison à des protéines de choc thermique, et au moins un lieur peptidique amélioré entre les deux domaines, lesdits antigènes s'avérant utiles pour l'induction d'une réponse immunitaire vis-à-vis du domaine antigénique lorsqu'ils sont administrés seuls ou dans un complexe avec au moins une protéine de choc thermique. Ces antigènes hybrides et les complexes associés peuvent être utilisés pour traiter des maladies infectieuses et des cancers qui expriment un antigène du domaine antigénique.
PCT/US2004/010983 2003-04-11 2004-04-09 Vaccins ameliores a base de proteines de choc thermique et immunotherapies associees Ceased WO2004091493A2 (fr)

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CA002521809A CA2521809A1 (fr) 2003-04-11 2004-04-09 Vaccins ameliores a base de proteines de choc thermique et immunotherapies associees
AU2004229458A AU2004229458B2 (en) 2003-04-11 2004-04-09 Improved heat shock protein-based vaccines and immunotherapies
MXPA05010881A MXPA05010881A (es) 2003-04-11 2004-04-09 Vacunas e inmunoterapias a base de proteina de choque termico mejorada.
JP2006509858A JP2007525448A (ja) 2003-04-11 2004-04-09 改良された熱ショックタンパク質に基づくワクチンおよび免疫治療
BRPI0409321-6A BRPI0409321A (pt) 2003-04-11 2004-04-09 antìgeno hìbrido, competição e método para induzir uma resposta imune para um agente infeccioso ou antìgeno de tumor, método para tratar uma doença infecciosa ou cáncer, e, peptìdeo
EP04759338A EP1617804A4 (fr) 2003-04-11 2004-04-09 Vaccins ameliores a base de proteines de choc thermique et immunotherapies associees

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US50341703P 2003-09-16 2003-09-16
US60/503,417 2003-09-16
US10/776,521 US7420037B2 (en) 2003-02-13 2004-02-12 Heat shock protein-based vaccines and immunotherapies
US10/776,521 2004-02-12
USPCT/US04/04340 2004-02-13
PCT/US2004/004340 WO2004071457A2 (fr) 2003-02-13 2004-02-13 Vaccins et immunotherapies ameliores a base de proteine de choc thermique
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EP2407178A2 (fr) 2005-04-19 2012-01-18 Eli Lilly and Company Antigènes polysaccharides synthétiques monovalent et polyvalent pour une intervention immunologique dans une maladie
US10568948B2 (en) 2015-05-13 2020-02-25 Agenus Inc. Vaccines for treatment and prevention of cancer
US11065317B2 (en) 2018-04-26 2021-07-20 Agenus Inc. Heat shock protein-binding peptide compositions and methods of use thereof

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Cited By (4)

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Publication number Priority date Publication date Assignee Title
EP1670507A4 (fr) * 2003-09-12 2007-10-17 Antigenics Inc Vaccin pour le traitement et la prevention de l'infection provoquee par le virus de l'herpes simplex
EP2407178A2 (fr) 2005-04-19 2012-01-18 Eli Lilly and Company Antigènes polysaccharides synthétiques monovalent et polyvalent pour une intervention immunologique dans une maladie
US10568948B2 (en) 2015-05-13 2020-02-25 Agenus Inc. Vaccines for treatment and prevention of cancer
US11065317B2 (en) 2018-04-26 2021-07-20 Agenus Inc. Heat shock protein-binding peptide compositions and methods of use thereof

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