+

WO2002024887A2 - Echangeurs a specificite de ligands/recepteurs permettant le reacheminement d'anticorps vers des recepteurs sur un agent pathogene - Google Patents

Echangeurs a specificite de ligands/recepteurs permettant le reacheminement d'anticorps vers des recepteurs sur un agent pathogene Download PDF

Info

Publication number
WO2002024887A2
WO2002024887A2 PCT/IB2001/002327 IB0102327W WO0224887A2 WO 2002024887 A2 WO2002024887 A2 WO 2002024887A2 IB 0102327 W IB0102327 W IB 0102327W WO 0224887 A2 WO0224887 A2 WO 0224887A2
Authority
WO
WIPO (PCT)
Prior art keywords
seq
ligand
receptor
receptor specificity
specificity exchanger
Prior art date
Application number
PCT/IB2001/002327
Other languages
English (en)
Other versions
WO2002024887A3 (fr
Inventor
Matti Sallberg
Jan-Ingmar Flock
Original Assignee
Tripep Ab
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
Priority to EP01985271A priority Critical patent/EP1354033A2/fr
Priority to AU2002223973A priority patent/AU2002223973B2/en
Priority to JP2002529482A priority patent/JP2004509621A/ja
Priority to KR10-2003-7003990A priority patent/KR20030034194A/ko
Priority to AU2397302A priority patent/AU2397302A/xx
Priority to CA002421877A priority patent/CA2421877A1/fr
Application filed by Tripep Ab filed Critical Tripep Ab
Publication of WO2002024887A2 publication Critical patent/WO2002024887A2/fr
Priority to US10/372,735 priority patent/US6933366B2/en
Publication of WO2002024887A3 publication Critical patent/WO2002024887A3/fr
Priority to US11/121,282 priority patent/US20060094862A1/en
Priority to US11/412,181 priority patent/US20070026502A1/en
Priority to AU2007203138A priority patent/AU2007203138B2/en

Links

Classifications

    • 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
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K19/00—Hybrid peptides, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes
    • 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
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • 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
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
    • 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
    • C07K14/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • 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 generally relates to compositions and methods for preventing and treating human diseases including, but not limited to, pathogens such as bacteria, yeast, parasites, fungus, viruses, and cancer. More specifically, embodiments described herein concern the manufacture and use of ligand/receptor specificity exchangers, which redirect existing antibodies in a subject to receptors present on pathogens. BACKGROUND OF THE INVENTION
  • receptors found on the surface of bacteria, yeast, parasites, fungus, viruses, and cancer cells, especially receptors that allow for attachment to a host cell or host protein (e.g., an extracellular matrix protein).
  • a host cell or host protein e.g., an extracellular matrix protein.
  • pathogenic bacteria e.g., Staphylococcus aureus
  • adhesion receptors e.g., ClfA, Efb, and FnBPA
  • a host's extracellular matrix proteins e.g., fibrinogen, fibronectin, and laminin.
  • investigators have shown that the adherence of some bacteria to host extracellular matrix proteins can be blocked by providing peptides that correspond to regions of the host extracellular matrix protein. (Pei et al., bifection and Immunity 67(9):4525-4530 (1999)).
  • viruses have receptors that interact with proteins present on the surface of host cells.
  • T4 glycoprotein a host cell protein
  • HAV human immunodeficiency virus
  • T4 peptide can be used to prevent or treat HIV infection.
  • many types of cancer cells express receptors that interact with host extracellular matrix proteins and investigators have shown that molecules that block integrin receptors can be used to inhibit tissue attachment, metastasis, angiogenesis, and tumor growth. (See e.g., U.S. Pat.
  • Embodiments include a ligand/receptor specificity exchanger having at least one specificity domain comprising a ligand for a receptor and at least one antigenic domain joined to said specificity domain, wherein said antigenic domain comprises an epitope of a pathogen or toxin.
  • the ligand/receptor specificity exchanger have a specificity domain that comprises at least three consecutive amino acids of a peptide selected from the group consisting of an extracellular matrix protein, a ligand for a receptor on a virus, and a ligand for a receptor on a cancer cell.
  • the peptide is an extracellular matrix protein selected from the group consisting of fibrinogen, collagen, vitronectin, laminin, plasminogen, thrombospondin, and fibronectin.
  • the extracellular matrix protein comprises at least 3 amino acids ofthe alpha-chain of fibrinogen and in the most preferred embodiments the ligand comprises the sequence Arginine-Glycine- Aspartate (RGD).
  • the peptide described above is a ligand for a receptor on a virus selected from the group consisting of T4 glycoprotein and hepatitis B viral envelope protein.
  • the peptide is a ligand for a receptor on a cancer cell selected from the group consisting of a ligand for HER-2/neu and a ligand for an integrin receptor.
  • Preferred embodiments have a specificity domain that comprises a sequence provided by one of SEQ. ID. Nos. 1-42.
  • the ligand/receptor specificity exchangers described herein interact with a receptor found on a pathogen.
  • the receptor is a bacterial adhesion receptor, for example, a bacterial adhesion receptor selected from the group consisting of extracellular fibrinogen binding protein (Efb), collagen binding protein, vitronectin binding protein, laminin binding protein, plasminogen binding protein, thrombospondin binding protein, clumping factor A (ClfA), clumping factor B (ClfB), fibronectin binding protein, coagulase, and extracellular adherence protein.
  • Efb extracellular fibrinogen binding protein
  • collagen binding protein vitronectin binding protein
  • laminin binding protein plasminogen binding protein
  • plasminogen binding protein thrombospondin binding protein
  • clumping factor A ClfA
  • clumping factor B ClfB
  • fibronectin binding protein coagulase, and extracellular adherence protein.
  • the antigenic domain comprises at least three amino acids of a peptide selected from the group consisting of a herpes simplex virus protein, a hepatitis B virus protein, a TT virus protein, and a poliovirus protein.
  • the ligand/receptor specificity exchanger has an antigenic domain that is a herpes simplex virus protein comprising a sequence selected from the group consisting of SEQ. ID. No. 53 and SEQ. ID. No. 54.
  • the antigenic domain is a hepatitis B virus protein comprising a sequence provided by one of SEQ. ID. No. 49, SEQ. ID. No. 50, SEQ. TO. No. 52, and SEQ.
  • Some ligand/receptor specificity exchangers also have an antigenic domain that is a TT virus protein comprising a sequence provided by one of SEQ. TD. Nos. 43-47 and SEQ. ID. Nos. 55-58.
  • the ligand/receptor specificity exchangers can also have an antigenic domain that is a polio virus protein comprising a sequence selected from the group consisting of SEQ. ID. No. 48 and SEQ. D). No. 51.
  • the ligand/receptor specificity exchanger has an antigenic domain that interacts with a high-titer antibody.
  • the antigenic domain specifically binds to an antibody present in animal serum that has been diluted to between approximately 1 : 100 to 1:1000 or greater.
  • the specificity exchangers of SEQ. ID. Nos. 60-105 are embodiments ofthe invention.
  • aspects of the invention also concern methods of treating or preventing a infection or proliferation of a pathogen.
  • One approach for example involves a method for treating and preventing bacterial infection. This method is practiced by providing a therapeutically effective amount of a ligand/receptor specificity exchanger to a subject, wherein said ligand/receptor specificity exchanger comprises a specificity domain that has a ligand that interacts with a receptor on a bacteria, and an antigenic domain that comprises an epitope for a pathogen or toxin.
  • a method of treating or preventing viral infection is also an embodiment.
  • a method of treating or preventing a viral infection is practiced by providing a therapeutically effective amount of a ligand/receptor specificity exchanger to a subject, wherein said ligand/receptor specificity exchanger comprises a specificity domain that has a ligand that interacts with a receptor on a virus, and an antigenic domain that comprises an epitope for a pathogen or toxin.
  • a method of treating or preventing cancer is an embodiment and this method can be practiced by providing a therapeutically effective amount of a ligand/receptor specificity exchanger to a subject, wherein said ligand/receptor specificity exchanger comprises a specificity domain that has a ligand that interacts with a receptor on a cancer cell, and an antigenic domain that comprises an epitope for a pathogen or toxin.
  • a ligand/receptor specificity exchanger comprises a specificity domain that has a ligand that interacts with a receptor on a cancer cell, and an antigenic domain that comprises an epitope for a pathogen or toxin.
  • antigen/antibody specificity exchanger is known in the art to refer to a molecule that comprises an amino acid sequence corresponding to an amino acid sequence of an antibody (e.g., a complementarity determining region) linked to an amino acid sequence to which a certain antibody binds (e.g., an epitope of a pathogen).
  • an antibody e.g., a complementarity determining region
  • Antigen/antibody specificity exchangers can redirect antibodies present in a subject to a pathogen and these exchanger agents have therapeutic and diagnostic use. (Id.).
  • ligand/receptor specificity exchangers Unlike antigen/antibody specificity exchangers, ligand/receptor specificity exchangers do not comprise a sequence found in an antibody. Instead, ligand/receptor specificity exchangers comprise a first domain that has a ligand for a receptor and a second domain that has an epitope of a pathogen or a toxin.
  • ligand/receptor specificity exchangers refers an exchanger agent that comprises a "specificity domain” that has at least one ligand for a receptor (a "ligand” is not an antibody or portion thereof) joined to an "antigenic domain” that has at least one epitope of a pathogen or toxin (e.g., pertussis toxin or cholera toxin).
  • the ligand/receptor specificity exchangers can comprise more than a specificity domain and an antigenic domain.
  • some ligand/receptor specificity exchangers comprise a plurality of specificity domains and/or antigenic domains.
  • Ligand/receptor specificity exchangers having multiple specificity domains and/or antigenic domains are said to be "multimerized” because more than one specificity domain and/or antigenic domain are fused in tandem.
  • Other embodiments concern ligand/receptor specificity exchangers that contain, in addition to a specificity domain and an antigenic domain, sequences that facilitate purification (e.g., a poly histidine tail), linkers (e.g., biotin and/or avidin or streptavidin or the flexible arms of 8 phage (8- linkers)), and sequences or modifications that either promote the stability of the ligand/receptor specificity exchanger (e.g., modifications that provide resistance to protease digestion) or promote the degradation of the ligand/receptor specificity exchanger (e.g., protease cleavage sites).
  • the specificity and antigenic domains are preferably peptides, some ligand/receptor specificity exchangers have specificity and antigenic domain
  • pathogen is used herein in a general sense to refer to an etiological agent of disease in animals including, but not limited to, bacteria, parasites, fungus, mold, viruses, and cancer cells.
  • receptor is used in a general sense to refer to a molecule (usually a peptide other than a sequence found in an antibody, but can be a carbohydrate, lipid, or nucleic acid) that interacts with a "ligand” (usually a peptide other than a sequence found in an antibody, or a carbohydrate, lipid, nucleic acid or combination thereof).
  • a "receptor”, as used herein, does not have to undergo signal transduction and can be involved in a number of molecular interactions including, but not limited to, adhesion (e.g., integrins) and molecular signaling (e.g., growth factor receptors).
  • desired specificity domains comprise a ligand that has a peptide sequence that is present in an extracellular matrix protein (e.g., fibrinogen, collagen, vitronectin, laminin, plasminogen, thrombospondin, and fibronectin) and some specificity domains comprise a ligand that interacts with a bacterial adhesion receptor (e.g., extracellular fibrinogen binding protein (Efb), collagen binding protein, vitronectin binding protein, laminin binding protein, plasminogen binding protein, thrombospondin binding protein, clumping factor A (ClfA), clumping factor B (ClfB), fibronectin binding protein, coagulase, and extracellular adherence protein).
  • Efb extracellular fibrinogen binding protein
  • CelfA clumping factor A
  • ClfB clumping factor B
  • fibronectin binding protein coagulase, and extracellular adherence protein
  • the specificity domain comprises a ligand that has a peptide sequence that interacts with a viral receptor (e.g., a fragment of T4 glycoprotein that binds gpl20 or a fragment of the preS domain, which binds gpl70 of the hepadnavirus family).
  • a viral receptor e.g., a fragment of T4 glycoprotein that binds gpl20 or a fragment of the preS domain, which binds gpl70 of the hepadnavirus family.
  • the specificity domain comprises a ligand that interacts with a receptor on a cancer cell (e.g., HER-2/neu (C-erbB2)) or an integrin receptor such as a vitronectin receptor, a laminin receptor, a fibronectin receptor, a collagen receptor, a fibrinogen receptor, an V431 receptor, an V63 ⁇ receptor, an V ⁇ ireceptor, an V ⁇ Bl receptor, and an V v 33receptor.
  • a cancer cell e.g., HER-2/neu (C-erbB2)
  • an integrin receptor such as a vitronectin receptor, a laminin receptor, a fibronectin receptor, a collagen receptor, a fibrinogen receptor, an V431 receptor, an V63 ⁇ receptor, an V ⁇ ireceptor, an V ⁇ Bl receptor, and an V v 33receptor.
  • Preferred embodiments have a specificity domain that comprises at least 8 amino acids of the alpha-chain of fibrinogen and/or the sequence Arginine-Glycine-Aspartic acid (RGD) and the most preferred embodiments have a specificity domain that comprises a sequence selected from the group consisting of SEQ. ID. Nos. 60-105.
  • RGD Arginine-Glycine-Aspartic acid
  • Desired antigenic domains have an epitope that is recognized by an antibody that already exists in a subject. For example, many people are immunized against childhood diseases including, but not limited to, small pox, measles, mumps, rubella, and polio. Thus, antibodies to epitopes on these pathogens can be produced by an immunized person. Desirable antigenic domains have an epitope that is found on one of these etiological agents.
  • Some embodiments have antigenic domains that interact with an antibody that has been administered to the subject.
  • an antibody that interacts with an antigenic domain on a ligand/receptor specificity exchanger can be co-administered with the ligand/receptor specificity exchanger.
  • an antibody that interacts with a ligand/receptor specificity exchanger may not normally exist in a subject but the subject has acquired the antibody by introduction of a biologic material (e.g., serum, blood, or tissue).
  • a biologic material e.g., serum, blood, or tissue.
  • subjects that undergo blood transfusion acquire numerous antibodies, some of which can interact with an antigenic domain of a ligand/receptor specificity exchanger.
  • the most desirable antigenic domains comprise an epitope that is recognized by a high titer antibody.
  • high titer antibody is meant an antibody that has high affinity for an antigen (e.g., an epitope on an antigenic domain).
  • an antigen e.g., an epitope on an antigenic domain.
  • a high titer antibody corresponds to an antibody present in a serum sample that remains positive in the assay after a dilution of the serum to approximately the range of 1:100-1:1000 in an appropriate dilution buffer, preferably, about 1:500.
  • the preferred antigenic domains have an epitope found on herpes simplex virus gG2 protein, hepatitis B virus s antigen (HBsAg), hepatitis B virus e antigen (HBeAg), hepatitis B virus c antigen (HBcAg), TT virus, and the poliovirus or combination thereof or comprise a sequence selected from the group consisting of SEQ. ID. Nos.43-59.
  • the ligand/receptor specificity exchangers described herein can be made by conventional techniques in recombinant engineering and/or peptide chemistry.
  • the specificity and antigenic domains are made separately and are subsequently joined together (e.g., through linkers or by association with a common carrier molecule). In other embodiments, the specificity domain and antigenic domain are made as part ofthe same molecule.
  • a ligand/receptor specificity exchanger having a specificity domain joined to an antigenic domain is made by a peptide synthesizer.
  • a nucleic acid encoding the specificity domain fused to an antigenic domain is cloned into an expression construct, transfected to cells, and the ligand/receptor specificity exchanger is purified or isolated from the cells or cell supernatent.
  • the ligand/receptor specificity exchanger can be screened to determine its ability to interact with the receptor on the pathogen and/or an antibody specific for the antigenic domain.
  • the term "characterization assay” is used to refer to an experiment or evaluation of the ability of a ligand/receptor specificity exchanger to interact with a receptor on a pathogen or cancer cell or fragment thereof and/or an antibody specific for the antigenic domain. Some characterization assays, for example, evaluate the ability of a ligand/receptor specificity exchanger to bind to a support having a receptor of a pathogen or fragment thereof disposed thereon or vice versa.
  • characterization assays assess the ability of a ligand/receptor specificity exchanger to bind to an antibody specific for the antigenic domain ofthe ligand/receptor specificity exchanger. Still other characterization assays evaluate the ability of the ligand/receptor specificity exchanger to effect infection by the pathogen or cancer cell proliferation in cultured cell lines or diseased animals.
  • the ligand/receptor specificity exchangers described herein can be used as the active ingredients in pharmaceuticals for the treatment and prevention of pathogenic infection, as well as cancer, in animals including humans.
  • the pharmaceutical embodiments can be formulated in many ways and may contain excipients, binders, emulsifiers, carriers, and other auxiliary agents in addition to the ligand/receptor specificity exchanger.
  • Pharmaceuticals comprising a ligand/receptor specificity exchanger can be administered by several routes including, but not limited to, topical, transdermal, parenteral, gastrointestinal, transbronchial, and transalveolar.
  • Ligand/receptor specificity exchangers can also be used as a coating for medical equipment and prosthetics to prevent infection or the spread of disease.
  • the amount of ligand/receptor specificity exchanger provided in a pharmaceutical, therapeutic protocol, or applied to a medical device varies depending on the intended use, the patient, and the frequency of administration.
  • Some ofthe methods disclosed concern the administration of a ligand/receptor specificity exchanger to a subject in need of treatment and/or prevention of bacterial infection, fungal infection, viral infection, and cancer.
  • a subject suffering from bacterial infection is provided a ligand/receptor specificity exchanger that comprises a specificity domain, which interacts with a bacterial receptor.
  • a subject suffering from a viral infection can be provided a ligand/receptor specificity exchanger that comprises a specificity domain that interacts with a viral receptor and a subject suffering from cancer is provided a ligand/receptor specificity exchanger that comprises a specificity domain that interacts with a receptor on the cancer cells.
  • Methods of treatment and prevention of disease are provided in which a subject suffering from disease or a subject at risk for contracting a disease is identified and then is provided a therapeutically effective amount of a ligand/receptor specificity exchanger that interacts with a receptor present on the etiological agent.
  • a subject suffering from disease or a subject at risk for contracting a disease is identified and then is provided a therapeutically effective amount of a ligand/receptor specificity exchanger that interacts with a receptor present on the etiological agent.
  • subjects suffering from a bacterial infection, fungal infection, viral infection, or cancer are identified by conventional clinical and diagnostic evaluation and are provided a therapeutically effective amount of a ligand/receptor specificity exchanger that interacts with the particular pathogen or cancer cell.
  • the ligand/receptor specificity exchangers described herein can be administered to all animals at risk of disease for prophylactic purposes, it may be desired to administer the ligand/receptor specificity exchangers only to those individuals that are in a high risk category (e.g., infants, the elderly, and those that come in close contact with pathogens). As stated above, high risk individuals are identified by currently available clinical and diagnostic techniques.
  • the section below provides more description of various types of ligand/receptor specificity exchangers that interact with receptors on bacteria, parasites, fungus, mold, viruses, and cancer cells.
  • Ligand/receptor specificity exchangers that interact with receptors on a pathogen
  • the ligand/receptor specificity exchangers that interact with receptors on a pathogen have a variety of chemical structures but, in a general sense, they are characterized as having at least one region that binds to the receptor (the specificity domain) and at least one region that interacts with an antibody that is specific for an epitope of a pathogen or toxin (the antigenic domain).
  • Preferred ligand/receptor specificity exchangers are peptides but some embodiments comprise derivatized or modified peptides or a peptidomimetic structure.
  • a typical peptide-based ligand/receptor specificity exchanger can be modified to have substituents not normally found on a peptide or to have substituents that are normally found on a peptide but are incorporated at regions that are not normal.
  • a peptide-based ligand/receptor specificity exchanger can be acetylated, acylated, or aminated and the substituents that can be included on the peptide so as to modify it include, but are not limited to, H, alkyl, aryl, alkenyl, alkynl, aromatic, ether, ester, unsubstituted or substituted amine, amide, halogen or unsubstituted or substituted sulfonyl or a 5 or 6 member aliphatic or aromatic ring.
  • the term "ligand/receptor specificity exchanger" is a broad one that encompasses modified or unmodified peptide structures, as well as peptidomimetics and chemical structures.
  • Synthetic peptides can be prepared employing conventional synthetic methods, utilizing L-amino acids, D-amino acids, or various combinations of amino acids ofthe two different configurations. Synthetic compounds that mimic the conformation and desirable features of a peptide but that avoid the undesirable features, e.g., flexibility (loss of conformation) and bond breakdown are known as a "peptidomimetics". (See, e.g., Spatola, A. F. Chemistry and Biochemistry of Amino Acids.
  • the design and synthesis of a peptidomimetic that resembles a ligand/receptor specificity exchanger involves starting with the sequence of the ligand/receptor specificity exchanger and conformation data (e.g., geometry data, such as bond lengths and angles) of a desired ligand/receptor specificity exchanger (e.g., the most probable simulated peptide), and using such data to determine the geometries that should be designed into the peptidomimetic.
  • conformation data e.g., geometry data, such as bond lengths and angles
  • a desired ligand/receptor specificity exchanger e.g., the most probable simulated peptide
  • Numerous methods and techniques are known in the art for performing this step, any of hich could be used. (See, e.g., Farmer, P. S., Drug Design, (Ariens, E. J. ed.), Vol. 10, pp.
  • Some embodiments comprise a plurality of specificity domains and/or a plurality of antigenic domains.
  • One type of ligand/receptor specificity exchanger that has a plurality of specificity domains and/or antigenic domains is referred to as a "multimerized ligand/receptor specificity exchanger" because it has multiple specificity domains and/or antigenic domains that appear in tandem on the same molecule.
  • a multimerized specificity domain may have two or more ligands that interact with one type of receptor, two or more ligands that interact with different types of receptors on the pathogen, and two or more ligands that interact with different types of receptors on different pathogens.
  • a multimerized antigenic domain can be constructed to have multimers of the same epitope of a pathogen or different epitopes of a pathogen, which can also be multimerized. That is, some multimerized antigenic domains are multivalent because the same epitope is repeated. In contrast, some multimerized antigenic domains have more than one epitope present on the same molecule in tandem but the epitopes are different. In this respect, these antigenic domains are multimerized but not multivalent. Further, some multimerized antigenic domains are constructed to have different epitopes but the different epitopes are themselves multivalent because each type of epitope is repeated.
  • Some ligand/receptor specificity exchangers comprise other elements in addition to the specificity domain and antigenic domain such as sequences that facilitate purification, linkers that provide greater flexibility and reduce steric hindrance, and sequences that either provide greater stability to the ligand/receptor specificity exchanger (e.g., resistance to protease degradation) or promote degradation (e.g., protease recognition sites).
  • the ligand/receptor specificity exchangers can comprise cleavable signal sequences that promote cytoplasmic export of the peptide and/or cleavable sequence tags that facilitate purification on antibody columns, glutathione columns, and metal columns.
  • Ligand/receptor specificity exchangers can comprise elements that promote flexibility of the molecule, reduce steric hindrance, or allow the ligand/receptor specificity exchanger to be attached to a support or other molecule. These elements are collectively referred to as "linkers".
  • avidin or streptavidin or their ligand — biotin
  • multiple ligand/receptor specificity exchangers can be joined together (e.g., through a support, such as a resin, or directly) or individual specificity domains and antigenic domains can be joined.
  • linker that can be included in a ligand/receptor specificity exchanger is referred to as a "8 linker" because it has a sequence that is found on 8 phage.
  • Preferred 8 sequences are those that correspond to the flexible arms of the phage.
  • These sequences can be included in a ligand/receptor specificity exchanger (e.g., between the specificity domain and the antigenic domain or between multimers of the specificity and/or antigenic domains) so as to provide greater flexibility and reduce steric hindrance.
  • ligand/receptor specificity exchangers can include sequences that either confer resistance to protease degradation or promote protease degradation. By incorporating multiple cysteines in a ligand/receptor specificity exchanger, for example, greater resistance to protease degradation can be obtained. These embodiments of the ligand/receptor specificity exchanger are expected to remain in the body for extended periods, which may be beneficial for some therapeutic applications. In contrast, ligand/receptor specificity exchangers can also include sequences that promote rapid degradation so as to promote rapid clearance from the body. Many sequences that serve as recognition sites for serine, cysteine, and aspartic proteases are known and can be included in a ligand/receptor specificity exchanger.
  • desired specificity domains comprise at least one ligand that has a peptide sequence that is present in an extracellular matrix protein. That is, a specificity domain can have a ligand that has a peptide sequence found in, for example, fibrinogen, collagen, vitronectin, laminin, plasminogen, thrombospondin, and fibronectin.
  • ligands that make up the specificity domain have an amino acid sequence that has been identified as being involved in adhesion to an extracellular matrix protein.
  • random fragments of known ligands for any receptor on a pathogen can be used to generate ligand/receptor specificity exchangers and these candidate ligand/receptor specificity exchangers can be screened in the characterization assays described infra to identify the molecules that interact with the receptors on the pathogen.
  • Some specificity domains have a ligand that interacts with a bacterial adhesion receptor including, but not limited to, extracellular fibrinogen binding protein (Efb), collagen binding protein, vitronectin binding protein, laminin binding protein, plasminogen binding protein, thrombospondin binding protein, clumping factor A (ClfA), clumping factor B (ClfB), fibronectin binding protein, coagulase, and extracellular adherence protein.
  • Efb extracellular fibrinogen binding protein
  • ClfA clumping factor A
  • ClfB clumping factor B
  • Ligands that have an amino acid sequence corresponding to the C-terminal portion of the gamma-chain of fibrinogen have been shown to competitively inhibit binding of fibrinogen to ClfA, a Staphylococcus aureus adhesion receptor.
  • preferred specificity domains comprise at least 3 amino acids of a sequence present in a molecule (e.g., fibrinogen) that can bind to a bacterial adhesion receptor.
  • Specificity domains can also comprise a ligand that interacts with a viral receptor.
  • a viral receptor Several viral receptors and corresponding ligands are known and these ligands or fragments thereof can be incorporated into a ligand/receptor specificity exchanger.
  • Tong et al. has identified an Hepadnavirus receptor, a 170kd cell surface glycoprotein that interacts with the pre-S domain of the duck hepatitis B virus envelope protein (U.S. Pat No. 5,929,220) and Maddon et al., has determined that the T cell surface protein CD4 (or the soluble form termedT4) interacts with gpl20 of HIV (U.S. Pat No. 6,093,539).
  • specificity domains that interact with a viral receptor can comprise regions of the pre-S domain of the duck hepatitis B virus envelope protein (e.g., amino acid residues 80-102 or 80-104) or regions of the T cell surface protein CD4 (or the soluble form termedT4) that interacts with gpl20 of HIV (e.g., the extracellular domain of CD4/T4 or fragments thereof).
  • regions of the pre-S domain of the duck hepatitis B virus envelope protein e.g., amino acid residues 80-102 or 80-104
  • regions of the T cell surface protein CD4 or the soluble form termedT4 that interacts with gpl20 of HIV (e.g., the extracellular domain of CD4/T4 or fragments thereof).
  • CD4 or the soluble form termedT4
  • Specificity domains can also comprise a ligand that interacts with a receptor present on a cancer cell.
  • the proto-oncogene HER-2/neu (C-erbB2) encodes a surface growth factor receptor of the tyrosine kinase family, pl85HER2.
  • Twenty to thirty percent of breast cancer patients over express the gene encoding HER-2/neu (C-erbB2), via gene amplification.
  • ligand/receptor specificity exchangers comprising a specificity domain that encodes a ligand for HER-2/neu (C- erbB2) are desirable embodiments.
  • Many types of cancer cells also over express or differentially express integrin receptors.
  • Many preferred embodiments comprise a specificity domain that interacts with an integrin receptor.
  • Ligands for integrin receptors include, but are not limited to, molecules that interact with a vitronectin receptor, a laminin receptor, a fibronectin receptor, a collagen receptor, a fibrinogen receptor, an V4 1 receptor, an V ⁇ 3 ⁇ receptor, an V33 preceptor, an V531 receptor, and an V v 33receptor.
  • TABLE I also lists several preferred specificity domains. The specificity domains described above are provided for illustrative purposes only and in no way should be construed to limit the scope of specificity domains that can be used with the embodiments described herein. The next section describes antigenic domains in greater detail. TABLE I
  • ILYFYALLFLSTCVAYVAT (SEQ . ID . No • 3)
  • RDNCCILDERFGSYCPTTCG (SEQ. ID. No. 5)
  • FGSYCPTTCGIADFLSTYQT (SEQ. ID. No. 6)
  • IADFLSTYQTKVDKDLQSLE SEQ. ID. No. 7
  • KVDKDLQSLEDILHQVENKT (SEQ. ID. No. 8)
  • DI HQVENKTSEVKQLIKAI (SEQ. ID. No. 9)
  • KPNMIDAATLKSRIMLEEIM SEQ. ID. No. 12
  • KSRIMLEEIMKYEASILTHD SEQ. ID. No. 13
  • TSHNGMQFSTWDNDNDKFEG SEQ. ID. No. 34
  • DNDNDKFEGNCAEQDGSG SEQ. ID. No. 35
  • NCAEQDGSGWWMNKCHAGHL SEQ. ID. No. 36
  • MNKCHAGHLNGVYYQGGTY SEQ. ID. No. 37
  • NGVYYQGGTYSKASTPNGYD (SEQ. ID. No. 38)
  • NGIIWATWKTRWYSMKKTTM SEQ. ID. No. 40
  • antigenic domains that can be used in the ligand/receptor specificity exchangers are also quite large because a pathogen or toxin can present many different epitopes. That is, the antigenic domains that can be incorporated into a ligand/receptor specificity exchanger include epitopes presented by bacteria, fungus, plants, mold, virus, cancer cells, and toxins. Desired antigenic domains comprise an epitope of a pathogen that already exists in a subject by virtue of naturally acquired immunity or vaccination. Epitopes of pathogens that cause childhood diseases, for example, can be used as antigenic domains.
  • Some embodiments have antigenic domains that interact with an antibody that has been administered to the subject.
  • an antibody that interacts with an antigenic domain on a specificity exchanger can be co-administered with the specificity exchanger.
  • an antibody that interacts with a ligand/receptor specificity exchanger may not no ⁇ nally exist in a subject but the subject has acquired the antibody by introduction of a biologic material (e.g., serum, blood, or tissue).
  • a biologic material e.g., serum, blood, or tissue.
  • subjects that undergo blood transfusion acquire numerous antibodies, some of which can interact with an antigenic domain of a ligand/receptor specificity exchanger.
  • Some preferred antigenic domains for use in a ligand/receptor specificity exchanger comprise viral epitopes including, but not limited to, the herpes simplex virus, hepatitis B virus, TT virus, and the poliovirus.
  • the antigenic domains comprise an epitope of a pathogen or toxin that is recognized by a "high-titer antibody”. Approaches to determine whether the epitope of a pathogen or toxin is recognizable by a high titer antibody are provided infra.
  • Epitopes of a pathogen that can be included in an antigenic domain of a ligand/receptor specificity exchanger include epitopes on peptide sequences disclosed in Swedish Pat No. 9901601-6; U.S. Pat. No. 5,869,232; Mol. Immunol. 28: 719-726 (1991); and J. Med. Virol. 33:248-252 (1991). TABLE II provides the amino acid sequence of several preferred antigenic domains.
  • LEDPASRDLV SEQ. ID. No. 52
  • VLICGENTVSRNYATHS (SEQ. ID. No. 55)
  • the specificity and antigenic domains are made separately and are subsequently joined together (e.g., through linkers or by association with a common carrier molecule) and in other embodiments, the specificity domain and antigenic domain are made as part of the same molecule.
  • any of the specificity domains listed in TABLE I can be joined to any of the antigenic domains of TABLE BE.
  • the specificity and antigenic domains could be made separately and joined together through a linker or carrier molecule (e.g., a complex comprising a biotinylated specificity domain, streptavidin, and a biotinylated antigenic domain), it is preferred that the ligand/receptor specificity exchanger is made as a fusion protein.
  • linker or carrier molecule e.g., a complex comprising a biotinylated specificity domain, streptavidin, and a biotinylated antigenic domain
  • the ligand/receptor specificity exchanger is made as a fusion protein.
  • preferred embodiments include fusion proteins comprising any of the specificity domains listed in TABLE I joined to any of the antigenic domains of TABLE II.
  • Ligand/receptor specificity exchangers can be generated in accordance with conventional methods of protein engineering, protein chemistry, organic chemistry, and molecular biology. Additionally, some commercial enterprises manufacture made-to-order peptides and a ligand/receptor specificity exchanger can be obtained by providing such a company with the sequence of a desired ligand/receptor specificity exchanger and employing their service to manufacture the agent according to particular specifications (e.g., Bachem AG, Switzerland). Preferably, the ligand/receptor specificity exchangers are prepared by chemical synthesis methods (such as solid phase peptide synthesis) using techniques known in the art, such as those set forth by Merrifield et al., J. Am. Chem. Soc.
  • solid phase peptide synthesis is performed using an Applied Biosystems 430A peptide synthesizer (Applied Biosystems, Foster City, CA). Each synthesis uses a p- methylbenzylhydrylamme solid phase support resin (Peptide International, Louisville, KY) yielding a carboxyl terminal amide when the peptides are cleaved off from the solid support by acid hydrolysis.
  • Applied Biosystems 430A peptide synthesizer Applied Biosystems, Foster City, CA.
  • Each synthesis uses a p- methylbenzylhydrylamme solid phase support resin (Peptide International, Louisville, KY) yielding a carboxyl terminal amide when the peptides are cleaved off from the solid support by acid hydrolysis.
  • the carboxyl terminal amide can be removed and the ligand/receptor specificity exchangers can be purified by high performance liquid chromatography (e.g., reverse phase high performance liquid chromatography (RP-HPLC) using a PepS-15 C18 column (Pharmacia, Uppsala, Sweden)) and sequenced on an Applied Biosystems 473A peptide sequencer.
  • high performance liquid chromatography e.g., reverse phase high performance liquid chromatography (RP-HPLC) using a PepS-15 C18 column (Pharmacia, Uppsala, Sweden)
  • RP-HPLC reverse phase high performance liquid chromatography
  • An alternative synthetic approach uses an automated peptide synthesizer (Syro, Multisyntech, Tubingen, Germany) and 9-fluorenylmethoxycarbonyl (fmoc) protected amino acids (Milligen, Bedford, MA).
  • ligand/receptor specificity exchangers can be chemically synthesized, it can be more efficient to produce these polypeptides by recombinant DNA technology using techniques well known in the art. Such methods can be used to construct expression vectors containing nucleotide sequences encoding a ligand/receptor specificity exchanger and appropriate transcriptional and translational control signals. These methods include, for example, in vitro recombinant DNA techniques, synthetic techniques, and in vivo genetic recombination. Alternatively, RNA capable of encoding a ligand/receptor specificity exchanger can be chemically synthesized using, for example, synthesizers. See, for example, the techniques described in Oligonucleotide Synthesis, 1984, Gait, M. J. ed., IRL Press, Oxford.
  • a variety of host-expression vector systems can be utilized to express the ligand/receptor specificity exchangers.
  • the ligand/receptor specificity exchanger is a soluble molecule it can be recovered from the culture, i.e., from the host cell in cases where the peptide or polypeptide is not secreted, and from the culture media in cases where the peptide or polypeptide is secreted by the cells.
  • the expression systems also encompass engineered host cells that express membrane bound ligand/receptor specificity exchangers. Purification or enrichment of the ligand/receptor specificity exchangers from such expression systems can be accomplished using appropriate detergents and lipid micelles and methods well known to those skilled in the art.
  • the expression systems include, but are not limited to, microorganisms such as bacteria (e.g., E. coli or B. subtilis) transformed with recombinant bacteriophage DNA, plasmid DNA or cosmid DNA expression vectors containing nucleotide sequences encoding a ligand/receptor specificity exchanger; yeast (e.g., Saccharomyces, Pichid) transformed with recombinant yeast expression vectors containing nucleotide sequences encoding ligand/receptor specificity exchangers; insect cell systems infected with recombinant virus expression vectors (e.g., Baculovirus) containing nucleic acids encoding the ligand/receptor specificity exchangers; or mammalian cell systems (e.g., COS, CHO, BHK, 293, 3T3) harboring recombinant expression constructs containing nucleic acids encoding ligand/receptor specificity exchangers.
  • a number of expression vectors can be advantageously selected depending upon the use intended for the ligand/receptor specificity exchanger. For example, when a large quantity is desired (e.g., for the generation of pharmaceutical compositions of ligand/receptor specificity exchangers) vectors that direct the expression of high levels of fusion protein products that are readily purified can be desirable.
  • vectors include, but are not limited, to the E.
  • coli expression vector pUR278 (Ruther et al., EMBO J., 2: 1791 (1983), in which the ligand/receptor specificity exchanger coding sequence can be ligated individually into the vector in frame with the lacZ coding region so that a fusion protein is produced; pIN vectors (Inouye & Inouye, Nucleic Acids Res., 13:3101-3109 (1985); Van Heeke & Schuster, J. Biol. Chem., 264:5503-5509 (1989)); and the like.
  • pGEX vectors can also be used to express foreign polypeptides as fusion proteins with glutathione S-transferase (GST).
  • fusion proteins are soluble and can be purified from lysed cells by adsorption to glutathione-agarose beads followed by elution in the presence of free glutathione.
  • the PGEX vectors are designed to include thrombin or factor Xa protease cleavage sites so that the cloned target gene product can be released from the GST moiety.
  • AcNPV Autographa californica nuclear polyhedrosis virus
  • the virus grows in Spodoptera frugiperda cells.
  • the ligand/receptor specificity exchanger gene coding sequence can be cloned individually into non- essential regions (for example the polyhedrin gene) of the virus and placed under control of an AcNPV promoter (for example the polyhedrin promoter). Successful insertion of ligand/receptor specificity exchanger gene coding sequence will result in inactivation of the polyhedrin gene and production of non-occluded recombinant virus, (i.e., virus lacking the proteinaceous coat coded for by the polyhedrin gene). These recombinant viruses are then used to infect Spodoptera frugiperda cells in which the inserted gene is expressed. (E.g., see Smith et al., J. Virol.
  • adenovirus In mammalian host cells, a number of viral-based expression systems can be utilized.
  • a nucleic acid sequence encoding a ligand/receptor specificity exchanger can be ligated to an adenovirus transcription/translation control complex, e.g., the late promoter and tripartite leader sequence. This chimeric gene can then be inserted in the adenovirus genome by in vitro or in vivo recombination.
  • Insertion in a non- essential region ofthe viral genome will result in a recombinant virus that is viable and capable of expressing the ligand/receptor specificity exchanger gene product in infected hosts.
  • a recombinant virus that is viable and capable of expressing the ligand/receptor specificity exchanger gene product in infected hosts.
  • Specific initiation signals can also be required for efficient translation of inserted ligand/receptor specificity exchanger nucleotide sequences (e.g., the ATG initiation codon and adjacent sequences). In most cases, an exogenous translational control signal, including, perhaps, the ATG initiation codon, should be provided.
  • initiation codon should be in phase with the reading frame of the desired coding sequence to ensure translation of the entire insert.
  • exogenous translational control signals and initiation codons can be of a variety of origins, both natural and synthetic.
  • the efficiency of expression can also be enhanced by the inclusion of appropriate transcription enhancer elements, transcription terminators, etc. (See Bittner et al., Methods in Enzymol, 153:516-544 (1987)).
  • a host cell strain can be chosen that modulates the expression of the inserted sequences, or modifies and processes the gene product in the specific fashion desired. Such modifications (e.g., glycosylation) and processing (e.g., cleavage) of protein products can be important for some embodiments.
  • Different host cells have characteristic and specific mechanisms for the post-translational processing and modification of proteins and gene products. Appropriate cell lines or host systems can be chosen to ensure the correct modification and processing of the foreign protein expressed.
  • eukaryotic host cells that possess the cellular machinery for proper processing ofthe primary transcript, glycosylation, and phosphorylation ofthe gene product can be used.
  • mammalian host cells include, but are not limited to, CHO, VERO, BHK, HeLa, COS, MDCK, 293, 3T3, and WI38.
  • cell lines that stably express the ligand/receptor specificity exchangers described above can be engineered.
  • host cells can be transformed with DNA controlled by appropriate expression control elements (e.g., promoter, enhancer sequences, transcription terminators, polyadenylation sites, etc.), and a selectable marker.
  • appropriate expression control elements e.g., promoter, enhancer sequences, transcription terminators, polyadenylation sites, etc.
  • engineered cells are allowed to grow for 1-2 days in an enriched media, and then are switched to a selective media.
  • the selectable marker in the recombinant plasmid confers resistance to the selection and allows cells to stably integrate the plasmid into their chromosomes and grow to form foci which in turn are cloned and expanded into cell lines.
  • This method is advantageously used to engineer cell lines which express a ligand/receptor specificity exchanger.
  • a number of selection systems can be used, including but not limited to the herpes simplex virus thymidine kinase (Wigler, et al., Ce/7 11:223 (1977)), hypoxanthine-guanine phosphoribosyltransferase (Szybalska & Szybalski, Proc. Natl. Acad. Sci. USA 48:2026 (1962)), and adenine phosphoribosyltransferase (Lowy, et al., Cell 22:817 (1980)) genes can be employed in tk.sup.-, hgprt.sup.- or aprt.sup.- cells, respectively.
  • anti etabolite resistance can be used as the basis of selection for the following genes: dhfr, which confers resistance to methotrexate (Wigler, et al., Proc. Natl. Acad. Sci. USA 77:3567 (1980)); O ⁇ are, et al., Proc. Natl. Acad. Sci. USA 78:1527 (1981)); gpt, which confers resistance to mycophenolic acid (Mulligan & Berg, Proc. Natl. Acad. Sci. USA 78:2072 (1981)); neo, which confers resistance to the aminoglycoside G-418 (Colberre-Garapin, et al., J. Mol. Biol. 150:1 (1981)); and hygro, which confers resistance to hygromycin (Santerre, et al., Gene 30:147 (1984)).
  • ligand/receptor specificity exchangers are analyzed for their ability to interact with a receptor and/or the ability to interact with an antibody that may be present in a subject.
  • characterization assay refers to an assay, experiment, or analysis made on a ligand/receptor specificity exchanger, which evaluates the ability of a ligand/receptor specificity exchanger to interact with a receptor (e.g., a surface receptor present in bacteria, virus, mold, or fungi) or an antibody (e.g., an antibody that recognizes an epitope found on a pathogen), or effect the proliferation of a pathogen.
  • characterization assay are binding studies (e.g., enzyme immunoassays (EIA), enzyme-linked immunoassays (ELISA), competitive binding assays, computer generated binding assays, support bound binding studies, and one and two hybrid systems), and infectivity studies (e.g., reduction of viral infection, propagation, and attachment to a host cell).
  • EIA enzyme immunoassays
  • ELISA enzyme-linked immunoassayssays
  • competitive binding assays e.g., computer generated binding assays, support bound binding studies, and one and two hybrid systems
  • infectivity studies e.g., reduction of viral infection, propagation, and attachment to a host cell).
  • Preferred binding assays use multimeric agents.
  • One form of multimeric agent concerns a composition comprising a ligand/receptor specificity exchanger, or fragments thereof disposed on a support.
  • Another form of multimeric agent involves a composition comprising a receptor or an antibody specific for the antigenic domain of a ligand/receptor specificity exchanger disposed on a support.
  • a "support” can be a carrier, a protein, a resin, a cell membrane, or any macromolecular structure used to join or immobilize such molecules.
  • Solid supports include, but are not limited to, the walls of wells of a reaction tray, test tubes, polystyrene beads, magnetic beads, nitrocellulose strips, membranes, microparticles such as latex particles, animal cells, Duracyte®, artificial cells, and others.
  • a ligand/receptor specificity exchanger can also be joined to inorganic supports, such as silicon oxide material (e.g. silica gel, zeolite, diatomaceous earth or aminated glass) by, for example, a covalent linkage through a hydroxy, carboxy, or amino group and a reactive group on the support.
  • the macromolecular support has a hydrophobic surface that interacts with a portion of the ligand/receptor specificity exchanger, receptor or ligand by a hydrophobic non-covalent interaction.
  • the hydrophobic surface of the support is a polymer such as plastic or any other polymer in which hydrophobic groups have been linked such as polystyrene, polyethylene or polyvinyl.
  • a ligand/receptor specificity exchanger, receptor or an antibody specific for the antigenic domain of a ligand/receptor specificity exchanger can be covalently bound to supports including proteins and oligo/polysaccarides (e.g.
  • a reactive group on the molecule such as a hydroxy or an amino group, is used to join to a reactive group on the carrier so as to create the covalent bond.
  • Additional multimeric agents comprise a support that has other reactive groups that are chemically activated so as to attach the ligand/receptor specificity exchanger, receptor, or antibody specific for the antigenic domain of a ligand/receptor specificity exchanger.
  • cyanogen bromide activated matrices epoxy activated matrices, thio and thiopropyl gels, nitrophenyl chloroformate and N-hydroxy succinimide chlorformate linkages, or oxirane acrylic supports can be used. (Sigma).
  • a liposome or lipid bilayer (natural or synthetic) is contemplated as a support and a ligand/receptor specificity exchanger, receptor, or an antibody specific for the antigenic domain of a ligand/receptor specificity exchanger can be attached to the membrane surface or are incorporated into the membrane by techniques in liposome engineering.
  • liposome multimeric supports comprise a ligand/receptor specificity exchanger, receptor, or an antibody specific for the antigenic domain of a ligand/receptor specificity exchanger that is exposed on the surface.
  • linkers e.g., " ⁇ linkers” engineered to resemble the flexible regions of ⁇ phage
  • linkers e.g., " ⁇ linkers” engineered to resemble the flexible regions of ⁇ phage
  • linkers e.g., " ⁇ linkers” engineered to resemble the flexible regions of ⁇ phage
  • linkers e.g., " ⁇ linkers” engineered to resemble the flexible regions of ⁇ phage
  • linkers e.g., " ⁇ linkers” engineered to resemble the flexible regions of ⁇ phage
  • the determination of an appropriate length of linker that allows for optimal binding can be found by screening the attached molecule with varying linkers in the characterization assays detailed herein.
  • ligand/receptor specificity exchangers employ a multimeric described above.
  • support-bound ligand/receptor specificity exchanger can be contacted with "free" adhesion receptors and an association can be determined directly (e.g., by using labeled adhesion receptors) or indirectly (e.g., by using a labeled ligand directed to an adhesion receptor).
  • candidate ligand/receptor specificity exchangers are identified as bona fide ligand/receptor specificity exchangers by virtue of the association of the receptors with the support-bound candidate ligand/receptor specificity exchanger.
  • support-bound adhesion receptors can be contacted with "free" ligand/receptor specificity exchangers and the amount of associated ligand/receptor specificity exchanger can be determined directly (e.g., by using labeled ligand/receptor specificity exchanger) or indirectly (e.g., by using a labeled antibody directed to the antigenic domain ofthe ligand/receptor specificity exchanger).
  • an antibody specific for the antigenic domain of a ligand/receptor specificity exchanger disposed on a support and labeled ligand/receptor specificity exchanger (or a secondary detection reagent, e.g., a labeled receptor or antibody to the ligand/receptor specificity exchanger) the ability of the antibody to bind to the antigenic domain of the ligand/receptor specificity exchanger can be determined.
  • Some characterization assays evaluate the ability of the ligand/receptor specificity exchanger to interact with the target receptor and the redirecting antibody while other characterization assays are designed to determine whether a ligand/receptor specificity exchanger can bind to both the target receptor and the redirecting antibody.
  • the characterization assays can be classified as: (1) in vitro characterization assays, (2) cellular characterization assays, and (3) in vivo characterization assays. A discussion of each type of characterization assay is provided in the following sections. In vitro characterization assays
  • ligand/receptor specificity exchanger binds to a particular receptor and whether an antibody found in a subject can bind to the ligand/receptor specificity exchanger.
  • the receptor is bound to a support (e.g., a petri dish) and the association of the ligand/receptor specificity exchanger with the receptor is monitored directly or indirectly, as described above.
  • a primary antibody directed to the antigenic domain of a ligand/receptor specificity exchanger can be bound to a support and the association of a ligand/receptor specificity exchanger with the primary antibody can be determined directly (e.g., using labeled ligand/receptor specificity exchanger) or indirectly (e.g., using labeled receptor or a labeled secondary antibody that interacts with an epitope on the ligand/receptor specificity exchanger that does not compete with the epitope recognized by the primary antibody).
  • Another approach involves a sandwich-type assay, wherein the receptor is bound to a support, the ligand/receptor specificity exchanger is bound to the receptor, and the primary antibody is bound to the ligand/receptor specificity exchanger. If labeled primary antibody is used, the presence of a receptor/specificity exchanger/primary antibody complex can be directly determined. The presence of the receptor/specificity exchanger/primary antibody complex can also be determined indirectly by using, for example, a labeled secondary antibody that reacts with the primary antibody at an epitope that does not interfere with the binding ofthe primary antibody to the ligand/receptor specificity exchanger.
  • a labeled tertiary antibody may be used to react with an unlabeled secondary antibody, thus, forming a receptor/specificity exchanger/primary antibody/secondary antibody/labeled tertiary antibody complex.
  • the example below describes a characterization assay that was performed to determine whether a specificity domain derived from the C-terminal domain of fibrinogen inhibits the binding of clumping factor (Clf) to fibrinogen.
  • EXAMPLE 1 In this example, several peptides corresponding to the C-terminal domain of fibrinogen (Fib) were analyzed for their ability to block the binding of clumping factor (Clf) to fibrinogen. (See TABLE ID). These peptides were manufactured using standard techniques in peptide synthesis using fmoc chemistry (Syro, MultiSynTech, Germany). Preferably, the peptides are purified by reverse-phase HPLC. A competition enzyme immunoassay was then performed to determine whether the peptides were able to block the interaction between Clf and fibrinogen.
  • Ligand/receptor specificity exchangers having specificity domains (approximately 20 amino acids long) corresponding to various regions of the fibrinogen gamma-chain sequence were produced using standard techniques in peptide synthesis using fmoc chemistry (Syro, MultiSynTech, Germany) and these ligand/receptor specificity exchangers were analyzed for their ability to bind the ClfA receptor and an antibody specific for their antigenic domains.
  • the sequences of these ligand/receptor specificity exchangers are listed in TABLE IV and are provided in the Sequence listing (SEQ. ID. Nos. 60-103).
  • the ligand/receptor specificity exchangers used in this analysis have an antigenic domain that presents an epitope of herpes simplex virus gG2 protein, which is recognized by a monoclonal antibody for herpes simplex virus gG2 proteins.
  • Serial dilutions of these ligand/receptor specificity exchangers were made in phosphate buffered saline (PBS) containing 2:g/ml goat serum. (Sigma Chemicals, St. Louis, MO) and 0.5% Tween 20 (PBS-GT).
  • PBS phosphate buffered saline
  • the receptor ClfA was passively adsorbed at 10 ⁇ g/ml to 96 well microtiter plates in 50mM sodium carbonate buffer, pH 9.6, overnight at +4° C.
  • the diluted ligand/receptor specificity exchangers were then incubated on the plates for 60 minutes.
  • the ability of the ligand/receptor specificity exchanger to interact with the receptor was determined by applying a primary antibody to the plate and incubating for 60 minutes (a 1:1000 dilution of mAb for herpes simplex virus gG2 proteins).
  • the bound primary mAb was then indicated by a rabbit anti-mouse IgG (Sigma) secondary antibody and a peroxidase labeled goat anti-rabbit IgG (Sigma) tertiary antibody.
  • the plates were developed by incubation with dinitro- phenylene-diamine (Sigma) and the absorbance at 405 nm was analyzed.
  • Every ligand/receptor specificity exchanger provided in TABLE IV (SEQ. ID Nos. 60- 103) appreciably bound the immobilized ClfA and also allowed for the binding of the mAb specific for HSV gG2 protein.
  • the method described above for determining the affinity of a ligand/receptor specificity exchanger for an adhesion receptor and a primary antibody can be performed for any candidate ligand/receptor specificity exchanger comprising any specificity domain and any antigenic domain provided that the appropriate sequences and adhesion receptors are used.
  • TABLE IV describes another characterization assay that was performed to determine whether a ligand/receptor specificity exchanger interacts with bacteria having the ClfA receptor.
  • RDNCCILDERFGSYCPTTCGHRGGPEE (SEQ. ID. No. 66)
  • FGSYCPTTCGIADFLSTYQTHRGGPEE (SEQ. ID. No. 67)
  • IADFLSTYQTKVDKDLQSLEHRGGPEE (SEQ. ID. No. 68)
  • KVDKDLQSLEDILHQVENKTHRGGPEE (SEQ. ID. No. 69)
  • DILHQVENKTSEVKQLIKAIHRGGPEE (SEQ. ID. No. 70)
  • IHDITGKDCQDIANKGAKQSHRGGPEE (SEQ. ID. No. 79)
  • NQQFLVYCEIDGSGNGWTVFHRGGPEE (SEQ. ID. No. 82)
  • DGSGNGWTVFQKRLDGSVDFHRGGPEE (SEQ. ID. No. 83)
  • ALRVELED NGRTSTADYAMHRGGPEE (SEQ. ID. No. 89)
  • FKVGPEADKYRLTYAYFAGGHRGGPEE (SEQ. ID. No. 91)
  • TSHNGMQFSTWDNDNDKFEGHRGGPEE (SEQ. ID. No. 95)
  • DNDNDKFEGNCAEQDGSGWHRGGPEE (SEQ. ID. No. 96)
  • NCAEQDGSG WMNKCHAGHLHRGGPEE (SEQ. ID. No. 97) MNKCHAGHLNGVYYQGGTYHRGGPEE (SEQ. ID. No. 98)
  • NGVYYQGGTYSKASTPNGYDHRGGPEE (SEQ. ID. No. 99)
  • NGII AT KTR YSMKKTTMHRGGPEE SEQ. ID. No. 101
  • Ligand/receptor specificity exchangers having specificity domains that bind to clumping factor (Clf) and antigenic domains that correspond to an epitope derived from the polio virus were produced using standard techniques in peptide synthesis using fmoc chemistry (Syro, MultiSynTech, Germany). See TABLE V. These ligand/receptor specificity exchangers were analyzed for their ability to inhibit the interaction between CLF and fibrinogen. In these experiments, the ligand/specificity exchangers described in TABLE V were manufactured and various concentrations of these molecules were added to an enzyme competition immunoassay containing Clf and fibrinogen.
  • the lowest inhibiting concentration which is the lowest peptide concentration needed to inhibit the Clf/Fib interaction, was ascertained. Accordingly, the lower the concentration needed to inhibit the Fib/Clf interaction, the more effective the inhibitor. Additionally, the lowest solid-phase bound peptide concentration, which is the lowest tested concentration of peptide recognized by anti-poliovirus antibodies in the immunoassay, was determined.
  • Some of the peptides used e.g., CPALTAVETGCTNPLAAHHLGGAKQAG (SEQ ID No. 135), HHLGGAKQAG-AA-CPALTAVETGCTNPL (SEQ ID No. 137), CPALTAVETGC-TNPLHHLGGAKQAG (SEQ ID No.
  • HHLGGAKQAG-AA-CPALTAVETGCTNPL SEQ ID No. 137
  • HHLGGAKQAG- CPALTAVETGCTNPL SEQ ID No. 142
  • HHLGGAKQAG-PALTAVETGATNPL >625 >200
  • the next section describes several cellular-based characterization assays that can be performed to determine whether a ligand/receptor specificity exchanger has an effect on the proliferation of a pathogen.
  • a pathogen-based approach is used to evaluate the ability of a ligand/receptor specificity exchanger to interact with a pathogen and an antibody directed to the antigenic domain of the ligand/receptor specificity exchanger.
  • This analysis also reveals the ability of the ligand/receptor specificity exchanger to effect proliferation of a pathogen because, in the body of a subject, the interaction ofthe ligand/receptor specificity exchanger with a pathogen and an antibody directed to the antigenic domain of the ligand/receptor specificity exchanger is followed by humoral and cellular responses that purge the pathogen from the subject (e.g., complement fixation and macrophage degradation).
  • the pathogen-based characterization assays involve providing ligand/receptor specificity exchangers to cultured pathogens and monitoring the association of the ligand/receptor specificity exchanger with the cells or virus.
  • pathogen-based characterization assays can be used and the example below describes some ofthe preferred characterization assays in greater detail.
  • EXAMPLE 4 One type of pathogen-based characterization assay involves binding of a ligand/receptor specificity exchanger to bacteria disposed on a support. Accordingly, bacteria that produce Clfa (e.g., Staphylococcus aureus, or Escherichia coli.) are grown in culture or on a agar plate in a suitable growth media (e.g., LB broth, blood broth, LB agar or blood agar). The cells are then transferred to a membrane (e.g., nitrocellulose or nylon) by either placing the culture on the membrane under vacuum (e.g., using a dot-blot manifold apparatus) or by placing the membrane on the colonies for a time sufficient to permit transfer.
  • a membrane e.g., nitrocellulose or nylon
  • the cells that are bound to the membrane are then provided a serial dilution of a ligand/receptor specificity exchanger (e.g., 500ng, l:g, 5:g, 10:g, 25:g, and 50:g of ligand/receptor specificity exchanger in a total volume of 200:1 of PBS).
  • a ligand/receptor specificity exchanger e.g., 500ng, l:g, 5:g, 10:g, 25:g, and 50:g of ligand/receptor specificity exchanger in a total volume of 200:1 of PBS.
  • a ligand/receptor specificity exchanger e.g., 500ng, l:g, 5:g, 10:g, 25:g, and 50:g of ligand/receptor specificity exchanger in a total volume of 200:1 of PBS.
  • the ligand/receptor specificity exchangers listed in TABLE IV or V are used.
  • the non-bound ligand/receptor specificity exchangers are removed and the membrane is washed with PBS (e.g., 3 washes with 2ml of PBS per wash).
  • PBS e.g., 3 washes with 2ml of PBS per wash.
  • a 1:100 - 1:1000 dilution of a primary antibody that interacts with the antigenic domain of the ligand/receptor specificity exchanger e.g., mAb for herpes simplex virus gG2 protein
  • the membrane is washed with PBS (e.g., 3 washes with 2ml of PBS per wash) to remove unbound primary antibody.
  • Appropriate controls include the membrane itself, bacteria on the membrane without a ligand/receptor specificity exchanger, and bacteria on the membrane with ligand/receptor specificity exchanger but no primary antibody.
  • a secondary antibody e.g., rabbit anti-mouse IgG (Sigma)
  • a tertiary antibody e.g., a peroxidase labeled goat anti-rabbit IgG (Sigma)
  • a labeled secondary antibody that interacts with the primary antibody can be used as well.
  • the secondary antibody is contacted with the membrane for 60 minutes and the non-bound secondary antibody is washed from the membrane with PBS (e.g., 3 washes with 2ml of PBS per wash).
  • the tertiary antibody is contacted with the membrane for 60 minutes and the non-bound tertiary antibody is washed from the membrane with PBS (e.g., 3 washes with 2ml of PBS per wash).
  • PBS e.g., 3 washes with 2ml of PBS per wash.
  • the bound tertiary antibody can be detected by incubating the membrane with dinitro-phenylene- diamine (Sigma).
  • Another approach involves the use of an immobilized ligand/receptor specificity exchanger.
  • primary antibody e.g., mAb for herpes simplex virus gG2 protein
  • various dilutions of a ligand/receptor specificity exchanger e.g., a ligand/receptor specificity exchanger provided in TABLE IV or V
  • the ligand/receptor specificity exchanger is allowed to associate with the primary antibody for 60 minutes and the non-bound ligand/receptor specificity exchanger is washed away (e.g., three washes with 2ml of PBS).
  • Appropriate controls include petri dishes without primary antibody or ligand/receptor specificity exchanger.
  • a turbid solution of bacteria e.g., E. coli
  • bacteria e.g., E. coli
  • PBS e.g., 3 washes with 2ml of PBS
  • growth media e.g., LB broth
  • LB agar is added to the petri dish and the culture is incubated overnight.
  • An interaction between the ligand/receptor specificity exchanger and the bacteria can be observed visually (e.g., turbid growth media, which can be quantified using spectrophotometric analysis or the appearance of colonies on the agar).
  • a ligand/receptor specificity exchanger By modifying the approaches described above, one of skill in the art can evaluate the ability of a ligand/receptor specificity exchanger to interact with a virus. For example, soluble fragments of T4 glycoprotein have been shown to interact with a human immunodeficiency virus (HIV) envelope glycoprotein. (See e.g., U.S. Pat No. 6,093,539).
  • HIV human immunodeficiency virus
  • Ligand/receptor specificity exchangers having a specificity domain comprising a fragment of T4 glycoprotein that interacts with HIV envelope glycoprotein e.g., amino acids 1-419 ofthe T4 glycoprotein sequence provided in U.S. Pat No.
  • 6,093,539 or a portion thereof can be made by synthesizing a fusion protein having the specificity domain joined to an antigenic domain (e.g., an antigenic domain listed in TABLE II).
  • an antigenic domain e.g., an antigenic domain listed in TABLE II.
  • peptide chemistry can be used to make the ligand/receptor specificity exchanger, it is preferred that an expression construct having the fragment of T4 glycoprotein joined to an antigenic domain is made and transfected into a suitable cell.
  • the expression and purification strategies described in U.S. Pat No. 6,093,539 and above can also be employed.
  • a filter binding assay is performed. Accordingly, serial ten-fold dilutions of HIV inoculum are applied to a membrane (e.g. nitrocellulose or nylon) in a dot blot apparatus under constant vacuum. Then serial ten fold dilutions of the ligand/receptor specificity exchanger are applied to the bound HIV particles. The ligand/receptor specificity exchanger is contacted with the particles for 60 minutes before applying vacuum and washing with PBS (e.g., 3 washes with 2ml of PBS per wash)).
  • PBS e.g., 3 washes with 2ml of PBS per wash
  • the non-bound ligand/receptor specificity exchanger is removed, ten fold serial dilutions of the primary antibody, which binds to the antigenic domain, are added to the samples and the binding reaction is allowed to occur for 60 minutes. Then a vacuum is applied and the non-bound primary antibody is washed with PBS (e.g., 3 washes with 2ml of PBS per wash)). The detection of the bound primary antibody can be accomplished, as described above.
  • a ligand/receptor specificity exchanger to interact with a virus can also be evaluated in a sandwich-type assay. Accordingly, a primary antibody that interacts with the antigenic domain of the ligand/receptor specificity exchanger is immobilized in micro titer wells and serial dilutions of ligand/receptor specificity exchanger are added to the primary antibody so as to create a primary antibody/specificity exchanger complex, as described above. Next, ten fold serial dilutions of HIV inoculum are added and the binding reaction is allowed to occur for 60 minutes. Non-bound HIV particles are removed by successive washes in PBS. Detection of the bound HIV particles can be accomplished using a radiolabeled anti-HIV antibody (e.g., antibody obtained from sera from a person suffering with HIV infection).
  • a radiolabeled anti-HIV antibody e.g., antibody obtained from sera from a person suffering with HIV infection.
  • cancer cells that expresses an RGD-responsive integrin (e.g., M21 human melanoma cells) are cultured to confluency.
  • M21 cells can be grown in DMEM media with 10% fetal bovine serum, 20 mM Hepes, and 1 mM pyruvate.
  • the cells are stained with hydroethidine (Polysciences, Inc., Warrington, PA) at 20 ⁇ g/ml final concentration (2 x 10 ⁇ cells/ml) for 30 min at 37°C and then washed twice to remove excess dye.
  • Hydroethidine intercalates into the DNA resulting in a red fluorescent labeling of the cells and does not impair the cell's adhesive functions.
  • the staining provides a way to quantify the binding of a ligand/receptor specificity exchanger to the cells. That is, the total number of hydroethidine stained cells can be compared to the number of cells bound to a fluorescently labeled primary antibody/specificity exchanger complex so as to determine the binding efficiency.
  • the stained cells are incubated with various dilutions of a ligand/receptor specificity exchanger comprising a RGD sequence (e.g., GRGDSPHRGGPEE (SEQ. ID No. 104) or WSRGDWHRGGPEE (SEQ. ID No. 105)).
  • a ligand/receptor specificity exchanger comprising a RGD sequence (e.g., GRGDSPHRGGPEE (SEQ. ID No. 104) or WSRGDWHRGGPEE (SEQ. ID No. 105)).
  • RGD sequence e.g., GRGDSPHRGGPEE (SEQ. ID No. 104) or WSRGDWHRGGPEE (SEQ. ID No. 105)
  • the non-bound ligand/receptor specificity exchanger is removed by several washes in DMEM media with 10% fetal bovine serum, 20 mM Hepes, and 1 mM pyruvate (e.g., 3 washes of 5ml of media).
  • a 1:100 - 1:1000 dilution of a primary antibody that interacts with the antigenic domain of the ligand/receptor specificity exchanger e.g., mAb for herpes simplex virus gG2 protein
  • the binding reaction is allowed to occur for 60 minutes.
  • several washes in media are performed to remove any non-bound primary antibody.
  • Appropriate controls include stained cells without ligand/receptor specificity exchanger or stained cells without primary antibody.
  • a goat anti-mouse FITC labeled antibody (1:100 dilution) (Sigma) is added and binding is allowed to occur for 60 minutes. Again, several media washes are made to remove any non-bound secondary antibody. Analysis is made by flow cytometry with filter settings at 543/590 nm for hydroethidine and 495/525 nm for fluorescin. One will observe an appreciable binding of primary antibody to the ligand/receptor specificity exchanger/cell complex, which will demonstrate that the ligand/receptor specificity exchanger will have an effect on the cell.
  • the example below describes a characterization assay that verified that RGD-containing ligand/receptor specificity exchangers effectively bind to mammalian cells and redirect antibodies to these cells.
  • the peptide RGDSAATPPAYR (SEQ ID NO. 145) was manufactured using standard techniques in peptide synthesis using fmoc chemistry (Syro, MultiSynTech, Germany). This peptide has a specificity domain that binds integrin receptors, a spacer (the AA), and an antigenic domain that has an epitope recognized by the monoclonal antibody 57/8, an epitope present on the hepatitis B virus e antigen (HBeAg).
  • Murine myeloma cells were washed in serum free media and were then incubated with the RGDSAATPPAYR (SEQ ID NO. 145) peptide or a control peptide derived from hepatitis C virus (HCV) NS3 domain at a concentration of 50 ⁇ g/ml. The cells were then washed and the amount surface bound peptide was detected by labeling the cells with the the 57/8 antibody. Surface bound antibody was indicated by an FITC labelled anti-mouse IgG conjugate diluted 1/500 and the level of surface staining was determined by fluorescent microscopy.
  • RGDSAATPPAYR SEQ ID NO. 1405
  • HCV hepatitis C virus
  • Characterization assays also include experiments that evaluate ligand/receptor specificity exchangers in vivo. There are many animal models that are suitable for evaluating the ability of a ligand/receptor specificity exchanger to inhibit pathogenic infection. Mice are preferred because they are easy to maintain and are susceptible to bacterial infection, viral infection, and cancer. Chimpanzees are also preferred because of their close genetic relationship to humans.
  • EXAMPLE 6 To test the ability of a ligand/receptor specificity exchanger to treat a bacterial infection the following characterization assay can be performed. Several female CF-1 outbred mice (Charles Rivers Laboratories) of approximately 8 weeks of age and 25 gram body mass are vaccinated with the antigenic domains of the ligand/receptor specificity exchangers to be tested. Preferably, the antigenic domains are coupled to a carrier and are administered with an adjuvant.
  • the antigenic domains can be fused to keyhole limpet hemocyanin or bovine serum albumin, which act as both a carrier and adjuvant or an adjuvant such as Freund's adjuvant, aluminum hydroxide, or lysolecithin can be used.
  • keyhole limpet hemocyanin or bovine serum albumin act as both a carrier and adjuvant or an adjuvant such as Freund's adjuvant, aluminum hydroxide, or lysolecithin can be used.
  • an adjuvant such as Freund's adjuvant, aluminum hydroxide, or lysolecithin
  • the immunized mice are inoculated intraperitoneally with overnight cultures of Staphylococcus aureus NTCC 10649. The inoculums are adjusted to yield approximately 100 x LD50 or log 6.6 for S. aureus.
  • Serial dilutions of ligand/receptor specificity exchangers are formulated in sterile water for injection and are administered by the subcutaneous (SC) or oral (PO) route at one and five hours post infection.
  • the challenge LD50 is validated by inoculation of untreated mice with log dilutions of the bacterial inoculum.
  • a five log dilution range of the bacterial challenges is inoculated into five groups often mice each (ten mice per log dilution). A mortality rate of 100% will be produced in all groups of untreated mice at the 100 X LD50 challenge inoculum.
  • mice are monitored daily for mortality for seven days.
  • the mean effective dose to protect 50% ofthe mice (ED50) can be calculated from cumulative mortality by logarithmic-probit analysis of a plotted curve of survival versus dosage as described in Antimicrob. Agents Chemother. 31: 1768-1774 and Proc. Soc. Exp. Biol. Med. 1994, 57, 261-264.
  • similar approaches can be used to test the ability of ligand/receptor specificity exchangers to inhibit viral infection and cancer.
  • the ligand/receptor specificity exchangers described herein can be formulated in pharmaceuticals and administered to subjects in need of an agent that inhibits the proliferation of a pathogen.
  • the section below describes several pharmaceuticals comprising ligand/receptor specificity exchangers that interact with a receptor on a pathogen.
  • compositions comprising a ligand/receptor specificity exchanger that interacts with a receptor on a pathogen
  • the ligand/receptor specificity exchangers described herein are suitable for incorporation into pharmaceuticals for administration to subjects in need of a compound that treats or prevents infection by a pathogen.
  • These pharmacologically active compounds can be processed in accordance with conventional methods of galenic pharmacy to produce medicinal agents for administration to mammals including humans.
  • the active ingredients can be incorporated into a pharmaceutical product with and without modification. Further, the manufacture of pharmaceuticals or therapeutic agents that deliver the pharmacologically active compounds of this invention by several routes are aspects of the present invention.
  • DNA, RNA, and viral vectors having sequences encoding a ligand/receptor specificity exchanger that interacts with a receptor on a pathogen are used with embodiments ofthe invention.
  • Nucleic acids encoding a ligand/receptor specificity exchanger can be administered alone or in combination with other active ingredients.
  • the compounds can be employed in admixture with conventional excipients, i.e., pharmaceutically acceptable organic or inorganic carrier substances suitable for parenteral, enteral (e.g., oral) or topical application that do not deleteriously react with the pharmacologically active ingredients described herein.
  • conventional excipients i.e., pharmaceutically acceptable organic or inorganic carrier substances suitable for parenteral, enteral (e.g., oral) or topical application that do not deleteriously react with the pharmacologically active ingredients described herein.
  • Suitable pharmaceutically acceptable carriers include, but are not limited to, water, salt solutions, alcohols, gum arabic, vegetable oils, benzyl alcohols, polyetylene glycols, gelatine, carbohydrates such as lactose, amylose or starch, magnesium stearate, talc, silicic acid, viscous paraffin, perfume oil, fatty acid monoglycerides and diglycerides, pentaerythritol fatty acid esters, hydroxy methylcellulose, polyvinyl pyrrolidone, etc.
  • auxiliary agents e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring, flavoring and/or aromatic substances and the like so long as the auxiliary agents does not deleteriously react with the ligand/receptor specificity exchangers.
  • the effective dose and method of administration of a particular pharmaceutical having a ligand/receptor specificity exchanger can vary based on the individual needs ofthe patient and the treatment or preventative measure sought.
  • Therapeutic efficacy and toxicity of such compounds can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., ED50 (the dose therapeutically effective in 50% of the population).
  • ED50 the dose therapeutically effective in 50% of the population.
  • the effective dose of a ligand/receptor specificity exchanger can be evaluated using the characterization assays described above. The data obtained from these assays is then used in formulating a range of dosage for use with other organisms, including humans.
  • the dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with no toxicity. The dosage varies within this range depending upon type of ligand/receptor specificity exchanger, the dosage form employed, sensitivity ofthe organism, and the route of administration.
  • Normal dosage amounts of a ligand/receptor specificity exchanger can vary from approximately 1 to 100,000 micrograms, up to a total dose of about 10 grams, depending upon the route of administration. Desirable dosages include about 250:g-lmg, about 50mg-200mg, and about 250mg-500mg.
  • the dose of a ligand/receptor specificity exchanger preferably produces a tissue or blood concentration or both from approximately 0.1 :M to 500mM. Desirable doses produce a tissue or blood concentration or both of about 1 to 800 ⁇ M. Preferable doses produce a tissue or blood concentration of greater than about 10 ⁇ M to about 500:M. Although doses that produce a tissue concentration of greater than 800:M are not preferred, they can be used. A constant infusion of a ligand/receptor specificity exchanger can also be provided so as to maintain a stable concentration in the tissues as measured by blood levels. The exact dosage is chosen by the individual physician in view ofthe patient to be treated.
  • Dosage and administration are adjusted to provide sufficient levels of the active moiety or to maintain the desired effect. Additional factors that can be taken into account include the severity ofthe disease, age ofthe organism being treated, and weight or size ofthe organism; diet, time and frequency of administration, drug combination(s), reaction sensitivities, and tolerance/response to therapy. Short acting pharmaceutical compositions are administered daily or more frequently whereas long acting pharmaceutical compositions are administered every 2 or more days, once a week, or once every two weeks or even less frequently.
  • Routes of administration of the pharmaceuticals include, but are not limited to, topical, transdermal, parenteral, gastrointestinal, transbronchial, and transalveolar.
  • Transdermal administration is accomplished by application of a cream, rinse, gel, etc. capable of allowing the ligand/receptor specificity exchangers to penetrate the skin.
  • Parenteral routes of administration include, but are not limited to, electrical or direct injection such as direct injection into a central venous line, intravenous, intramuscular, intraperitoneal, intradermal, or subcutaneous injection.
  • Gastrointestinal routes of administration include, but are not limited to, ingestion and rectal.
  • Transbronchial and transalveolar routes of administration include, but are not limited to, inhalation, either via the mouth or intranasally.
  • compositions having the ligand/receptor specificity exchangers described herein that are suitable for transdermal or topical administration include, but are not limited to, pharmaceutically acceptable suspensions, oils, creams, and ointments applied directly to the skin or incorporated into a protective carrier such as a transdermal device ("transdermal patch").
  • transdermal patch a transdermal device
  • suitable creams, ointments, etc. can be found, for instance, in the Physician's Desk Reference.
  • suitable transdermal devices are described, for instance, in U.S. Patent No. 4,818,540 issued April 4, 1989 to Chinen, e al.
  • compositions having pharmacologically active compounds that are suitable for parenteral administration include, but are not limited to, pharmaceutically acceptable sterile isotonic solutions.
  • Such solutions include, but are not limited to, saline and phosphate buffered saline for injection into a central venous line, intravenous, intramuscular, intraperitoneal, intradermal, or subcutaneous injection.
  • compositions having pharmacologically active compounds that are suitable for transbronchial and transalveolar administration include, but are not limited to, various types of aerosols for inhalation. Devices suitable for transbronchial and transalveolar administration of these are also embodiments. Such devices include, but are not limited to, atomizers and vaporizers. Many forms of currently available atomizers and vaporizers can be readily adapted to deliver compositions having the ligand/receptor specificity exchangers described herein.
  • Compositions having pharmacologically active compounds that are suitable for gastrointestinal administration include, but not limited to, pharmaceutically acceptable powders, pills or liquids for ingestion and suppositories for rectal administration. Due to the ease of use, gastrointestinal administration, particularly oral, is a preferred embodiment. Once the pharmaceutical comprising the ligand/receptor specificity exchanger has been obtained, it can be administered to an organism in need to treat or prevent pathogenic infection.
  • aspects of the invention also include a coating for medical equipment such as prosthetics, implants, and instruments.
  • Coatings suitable for use on medical devices can be provided by a gel or powder containing the ligand/receptor specificity exchanger or by a polymeric coating into which a ligand/receptor specificity exchanger is suspended.
  • Suitable polymeric materials for coatings of devices are those that are physiologically acceptable and through which a therapeutically effective amount ofthe ligand/receptor specificity exchanger can diffuse.
  • Suitable polymers include, but are not limited to, polyurethane, polymethacrylate, polyamide, polyester, polyethylene, polypropylene, polystyrene, polytetrafluoroethylene, polyvinyl-chloride, cellulose acetate, silicone elastomers, collagen, silk, etc.
  • Such coatings are described, for instance, in U.S. PatentNo. 4,612,337.
  • compositions comprising a ligand/receptor specificity exchanger can be administered to a subject in need to treat and/or prevent infection by a pathogen that has a receptor.
  • Such subjects in need can include individuals at risk of contacting a pathogen or individuals who are already infected by a pathogen. These individuals can be identified by standard clinical or diagnostic techniques.
  • a subject suffering from a bacterial infection is identified as a subject in need of an agent that inhibits proliferation of a pathogen.
  • This subject is then provided a therapeutically effective amount of ligand/receptor specificity exchanger.
  • the ligand/receptor specificity exchanger used in this method comprises a specificity domain that interacts with a receptor present on the bacteria (e.g., extracellular fibrinogen binding protein (Efb), collagen binding protein, vitronectin binding protein, laminin binding protein, plasminogen binding protein, thrombospondin binding protein, clumping factor A (ClfA), clumping factor B (ClfB), fibronectin binding protein, coagulase, and extracellular adherence protein).
  • Efb extracellular fibrinogen binding protein
  • collagen binding protein e.g., collagen binding protein, vitronectin binding protein, laminin binding protein, plasminogen binding protein, thrombospondin binding protein, clumping factor A (ClfA
  • the ligand/receptor specificity exchanger also comprises an antigenic domain that has an epitope of a pathogen or toxin, preferably, an epitope recognized by high titer antibodies present in the subject in need. It may also be desired to screen the subject in need for the presence of high titer antibodies that recognize the antigenic domain prior to providing the subject the ligand/receptor specificity exchanger. This screening can be accomplished by EIA or ELISA using immobilized antigenic domain or ligand/receptor specificity exchanger, as described above.
  • a subject in need of an agent that inhibits viral infection can be administered a ligand/receptor specificity exchanger that recognizes a receptor present on the particular etiologic agent. Accordingly, a subject in need of an agent that inhibits viral infection is identified by standard clinical or diagnostic procedures. Next, the subject in need is provided a therapeutically effective amount of a ligand/receptor specificity exchanger that interacts with a receptor present on the type of virus infecting the individual. As above, it may be desired to determine whether the subject has a sufficient titer of antibody to interact with the antigenic domain ofthe ligand/receptor specificity exchanger prior to administering the ligand/receptor specificity exchanger.
  • a subject in need of an agent that inhibits the proliferation of cancer can be administered a ligand/receptor specificity exchanger that interacts with a receptor present on the cancer cell.
  • a subject in need of an agent that inhibits proliferation of cancer is identified by standard clinical or diagnostic procedures; then the subject in need is provided a therapeutically effective amount of a ligand/receptor specificity exchanger that interacts with a receptor present on the cancer cells infecting the subject.
  • Ligand/receptor specificity exchangers described herein can also be administered to subjects as a prophylactic to prevent the onset of disease. Virtually anyone can be administered a ligand/receptor specificity exchanger described herein for prophylactic purposes, (e.g., to prevent a bacterial infection, viral infection, or cancer). It is desired, however, that subjects at a high risk of contracting a particular disease are identified and provided a ligand/receptor specificity exchanger. Subjects at high risk of contracting a disease include individuals with a family history of disease , the elderly or the young, or individuals that come in frequent contact with a pathogen (e.g., health care practitioners).
  • a pathogen e.g., health care practitioners
  • ligand/receptor specificity exchanger subjects at risk of becoming infected by a pathogen that has a receptor are identified and then are provided a prophylactically effective amount of ligand/receptor specificity exchanger.
  • One prophylactic application for the a ligand/receptor specificity exchangers described herein concerns coating or cross-linking the ligand/receptor specificity exchanger to a medical device or implant.
  • Implantable medical devices tend to serve as foci for infection by a number of bacterial species. Such device-associated infections are promoted by the tendency of these organisms to adhere to and colonize the surface of the device. Consequently, there is a considerable need to develop surfaces that are less prone to promote the adverse biological reactions that typically accompany the implantation of a medical device.
  • the medical device is coated in a solution of containing a ligand/receptor specificity exchanger.
  • medical devices e.g., a prosthetic valve
  • ligand/receptor specificity exchangers for example.
  • Medical devices can also be coated in a powder or gel having a ligand/receptor specificity exchanger.
  • gloves, condoms, and intrauterine devices can be coated in a powder or gel that contains a specificity exchanger that interacts with a bacterial or viral receptor.
  • the ligand/receptor specificity exchanger is immobilized to the medical device.
  • the medical device is a support to which a ligand/receptor specificity exchanger can be attached. Immobilization may occur by hydrophobic interaction between the ligand/receptor specificity exchanger and the medical device but a preferable way to immobilize a ligand/receptor specificity exchanger to a medical device involves covalent attachment.
  • medical devices can be manufactured with a reactive group that interacts with a reactive group present on the specificity exchanger.
  • a periodate is combined with a ligand/receptor specificity exchanger comprising a 2-aminoalcohol moiety to form an aldehyde-functional exchanger in an aqueous solution having a pH between about 4 and about 9 and a temperature between about 0 and about 50 degrees Celsius.
  • the aldehyde-functional exchanger is combined with the biomaterial surface of a medical device that comprises a primary amine moiety to immobilize the ligand/receptor specificity exchanger on the support surface through an imine moiety.
  • the imine moiety is reacted with a reducing agent to form an immobilized ligand/receptor specificity exchanger on the biomaterial surface through a secondary amine linkage.
  • Other approaches for cross-linking molecules to medical devices can be modified to immobilize the ligand/receptor specificity exchanger described herein.

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Organic Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Genetics & Genomics (AREA)
  • General Chemical & Material Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Animal Behavior & Ethology (AREA)
  • Biochemistry (AREA)
  • Public Health (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Biophysics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Communicable Diseases (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Zoology (AREA)
  • Oncology (AREA)
  • Biomedical Technology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Wood Science & Technology (AREA)
  • Toxicology (AREA)
  • Physics & Mathematics (AREA)
  • Virology (AREA)
  • Plant Pathology (AREA)
  • Microbiology (AREA)
  • Peptides Or Proteins (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)

Abstract

La présente invention concerne des compositions et des procédés de prévention et de traitement de maladies humaines comportant, mais de manière non exclusive, des agents pathogènes tels que les bactéries, les levures, les parasites, les champignons, les virus et le cancer. Plus particulièrement, les modes de réalisation de l'invention ont trait à la fabrication et à l'utilisation d'échangeurs à spécificité de ligands/récepteurs, permettant le réacheminement d'anticorps existant chez un sujet vers des récepteurs présents sur des agents pathogènes.
PCT/IB2001/002327 1994-04-28 2001-09-19 Echangeurs a specificite de ligands/recepteurs permettant le reacheminement d'anticorps vers des recepteurs sur un agent pathogene WO2002024887A2 (fr)

Priority Applications (10)

Application Number Priority Date Filing Date Title
AU2002223973A AU2002223973B2 (en) 2000-09-19 2001-09-19 Ligand/receptor specificity exchangers that redirect antibodies to receptors on a pathogen
JP2002529482A JP2004509621A (ja) 2000-09-19 2001-09-19 病原体上の受容体に対して抗体を再指向させるリガンド／受容体特異性変換体
KR10-2003-7003990A KR20030034194A (ko) 2000-09-19 2001-09-19 병원체 내에서 수용체에 대한 항체를 재유도하는리간드/수용체 특이성 교환기
AU2397302A AU2397302A (en) 2000-09-19 2001-09-19 Ligand/receptor specificity exchangers that redirect antibodies to receptors on a pathogen
CA002421877A CA2421877A1 (fr) 2000-09-19 2001-09-19 Echangeurs a specificite de ligands/recepteurs permettant le reacheminement d'anticorps vers des recepteurs sur un agent pathogene
EP01985271A EP1354033A2 (fr) 2000-09-19 2001-09-19 Echangeurs a specificite de ligands/recepteurs permettant le reacheminement d'anticorps vers des recepteurs sur un agent pathogene
US10/372,735 US6933366B2 (en) 1996-12-27 2003-02-21 Specificity exchangers that redirect antibodies to bacterial adhesion receptors
US11/121,282 US20060094862A1 (en) 1994-04-28 2005-05-03 Specificity exchangers that redirect antibodies to a pathogen
US11/412,181 US20070026502A1 (en) 1994-04-28 2006-04-26 Specificity exchangers that redirect antibodies to a pathogen
AU2007203138A AU2007203138B2 (en) 2000-09-19 2007-07-05 Ligand/receptor specificity exchangers that redirect antibodies to receptors on a pathogen

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US66402500A 2000-09-19 2000-09-19
US09/664,025 2000-09-19

Related Child Applications (1)

Application Number Title Priority Date Filing Date
US10/372,735 Continuation-In-Part US6933366B2 (en) 1994-04-28 2003-02-21 Specificity exchangers that redirect antibodies to bacterial adhesion receptors

Publications (2)

Publication Number Publication Date
WO2002024887A2 true WO2002024887A2 (fr) 2002-03-28
WO2002024887A3 WO2002024887A3 (fr) 2003-08-14

Family

ID=24664202

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/IB2001/002327 WO2002024887A2 (fr) 1994-04-28 2001-09-19 Echangeurs a specificite de ligands/recepteurs permettant le reacheminement d'anticorps vers des recepteurs sur un agent pathogene

Country Status (9)

Country Link
EP (1) EP1354033A2 (fr)
JP (1) JP2004509621A (fr)
KR (1) KR20030034194A (fr)
CN (1) CN100371442C (fr)
AU (2) AU2002223973B2 (fr)
CA (1) CA2421877A1 (fr)
PL (1) PL364788A1 (fr)
RU (1) RU2300391C2 (fr)
WO (1) WO2002024887A2 (fr)

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2003046001A3 (fr) * 2001-11-23 2003-09-25 Syn X Pharma Inc Marqueurs biopolymeres de proteines predictifs de l'insulinoresistance
US6660842B1 (en) 1994-04-28 2003-12-09 Tripep Ab Ligand/receptor specificity exchangers that redirect antibodies to receptors on a pathogen
WO2004069873A2 (fr) 2003-02-06 2004-08-19 Tripep Ab Echangeurs de specificite glycosyles
US6933366B2 (en) 1996-12-27 2005-08-23 Tripep Ab Specificity exchangers that redirect antibodies to bacterial adhesion receptors
US7335359B2 (en) 2003-02-06 2008-02-26 Tripep Ab Glycosylated specificity exchangers
US9284548B2 (en) 2009-10-05 2016-03-15 Tripep Ab High affinity adaptor molecules for redirecting antibody specifity

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101987866B (zh) * 2010-10-25 2013-01-23 中国人民解放军军事医学科学院基础医学研究所 金黄色葡萄球菌Efb蛋白C端抗原表位及其制备方法和用途

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH04310861A (ja) * 1991-04-09 1992-11-02 Kazuo Yanagi 抗ｅｂｎａ抗体の測定方法および抗ｅｂｎａ抗体測定キット
ES2296289T3 (es) * 1994-02-18 2008-04-16 The American National Red Cross Fibrinogeno transgenico.
SE9401460D0 (sv) * 1994-04-28 1994-04-28 Ferring Ab Antigen/antibody specificity exhanger
WO1999061041A1 (fr) * 1998-05-27 1999-12-02 Hadasit Medical Research Services & Development Ltd. Nouveaux peptides
WO2000026385A1 (fr) * 1998-11-05 2000-05-11 Powderject Vaccines, Inc. Produits de recombinaison d'acide nucleique pour immunisation genetique

Cited By (17)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7019111B2 (en) 1994-04-28 2006-03-28 Tripep Ab Glycosylated ligand/receptor specificity exchangers specific for bacterial adhesion receptors
US6660842B1 (en) 1994-04-28 2003-12-09 Tripep Ab Ligand/receptor specificity exchangers that redirect antibodies to receptors on a pathogen
US6933366B2 (en) 1996-12-27 2005-08-23 Tripep Ab Specificity exchangers that redirect antibodies to bacterial adhesion receptors
WO2003046001A3 (fr) * 2001-11-23 2003-09-25 Syn X Pharma Inc Marqueurs biopolymeres de proteines predictifs de l'insulinoresistance
US7052849B2 (en) 2001-11-23 2006-05-30 Syn X Pharma, Inc. Protein biopolymer markers predictive of insulin resistance
US7318926B2 (en) 2003-02-06 2008-01-15 Tripep Ab Glycosylated specificity exchangers
WO2004069873A3 (fr) * 2003-02-06 2004-10-07 Tripep Ab Echangeurs de specificite glycosyles
JP2007516157A (ja) * 2003-02-06 2007-06-21 トリペップ アクチ ボラゲット グリコシル化された抗原／抗体又はリガンド／受容体特異性交換体
WO2004069873A2 (fr) 2003-02-06 2004-08-19 Tripep Ab Echangeurs de specificite glycosyles
US7332166B2 (en) 2003-02-06 2008-02-19 Tripep Ab Glycosylated specificity exchangers
US7335359B2 (en) 2003-02-06 2008-02-26 Tripep Ab Glycosylated specificity exchangers
US7534435B2 (en) 2003-02-06 2009-05-19 Tripep Ab Glycosylated specificity exchangers
EP2230249A3 (fr) * 2003-02-06 2010-12-22 ChronTech Pharma AB Antigène/anticorps ou ligand/récepteur glycosyles avec un échangeur de spécificité
US8303956B2 (en) 2003-02-06 2012-11-06 Chrontech Pharma Ab Glycosylated specificity exchangers
US8658179B2 (en) 2003-02-06 2014-02-25 Chrontech Pharma Ab Glycosylated specificity exchangers
US9079962B2 (en) 2003-02-06 2015-07-14 Tripep Ab Glycosylated specificity exchangers
US9284548B2 (en) 2009-10-05 2016-03-15 Tripep Ab High affinity adaptor molecules for redirecting antibody specifity

Also Published As

Publication number Publication date
CA2421877A1 (fr) 2002-03-28
AU2397302A (en) 2002-04-02
WO2002024887A3 (fr) 2003-08-14
JP2004509621A (ja) 2004-04-02
KR20030034194A (ko) 2003-05-01
CN100371442C (zh) 2008-02-27
PL364788A1 (en) 2004-12-13
CN1476476A (zh) 2004-02-18
RU2300391C2 (ru) 2007-06-10
RU2003106428A (ru) 2005-01-20
AU2002223973B2 (en) 2007-04-05
EP1354033A2 (fr) 2003-10-22

Similar Documents

Publication Publication Date Title
US6933366B2 (en) Specificity exchangers that redirect antibodies to bacterial adhesion receptors
JP4216473B2 (ja) Ｈｉｖ感染および免疫疾患の予防および治療のペプチド組成物
US7019111B2 (en) Glycosylated ligand/receptor specificity exchangers specific for bacterial adhesion receptors
AU2012200059B2 (en) "Antigen/antibody or ligand/receptor glycosylated"
CS275838B6 (en) Method for production of hybridoms used for monoclonal antigen against hiv virus producing
AU2002223973B2 (en) Ligand/receptor specificity exchangers that redirect antibodies to receptors on a pathogen
AU2002223973A1 (en) Ligand/receptor specificity exchangers that redirect antibodies to receptors on a pathogen
EP1294891A2 (fr) Peptides de transport dont le peptide e?rns c-terminal et des analogues de celui-ci
US6827939B2 (en) Peptide having an affinity for gp120
AU2007203138B2 (en) Ligand/receptor specificity exchangers that redirect antibodies to receptors on a pathogen
US7534435B2 (en) Glycosylated specificity exchangers
JP3725899B2 (ja) Ｈｉｖに対して使用するための多分岐ペプチド構築物
JP3725899B6 (ja) Ｈｉｖに対して使用するための多分岐ペプチド構築物
CA2058876A1 (fr) Peptides cycliques, principaux determinants de neutralisation du vih
JPH05271276A (ja) 環式ｈｉｖ主要中和抗原決定基及びリポペプチドを含有する合成ペプチド
MXPA00011508A (en) Antigenic complex comprising immunostimulatory peptide, cd4, and chemokine receptor domain for hiv treatment and immune disorders

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A2

Designated state(s): AE AG AL AM AT AT AU AZ BA BB BG BR BY BZ CA CH CN CO CR CU CZ CZ DE DE DK DK DM DZ EC EE EE ES FI FI GB GD GE GH GM HR HU ID IL IN IS JP KE KG KP KR KZ LC LK LR LS LT LU LV MA MD MG MK MN MW MX MZ NO NZ PH PL PT RO RU SD SE SG SI SK SK SL TJ TM TR TT TZ UA UG US UZ VN YU ZA ZW

AL Designated countries for regional patents

Kind code of ref document: A2

Designated state(s): GH GM KE LS MW MZ SD SL SZ TZ UG ZW AM AZ BY KG KZ MD RU TJ TM AT BE CH CY DE DK ES FI FR GB GR IE IT LU MC NL PT SE TR BF BJ CF CG CI CM GA GN GQ GW ML MR NE SN TD TG

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

Ref document number: 2421877

Country of ref document: CA

WWE Wipo information: entry into national phase

Ref document number: 2002223973

Country of ref document: AU

WWE Wipo information: entry into national phase

Ref document number: 1020037003990

Country of ref document: KR

Ref document number: 2002529482

Country of ref document: JP

WWE Wipo information: entry into national phase

Ref document number: 018167802

Country of ref document: CN

ENP Entry into the national phase

Ref document number: 2003106428

Country of ref document: RU

Kind code of ref document: A

Format of ref document f/p: F

Country of ref document: RU

Kind code of ref document: A

Format of ref document f/p: F

WWE Wipo information: entry into national phase

Ref document number: 2001985271

Country of ref document: EP

WWP Wipo information: published in national office

Ref document number: 1020037003990

Country of ref document: KR

REG Reference to national code

Ref country code: DE

Ref legal event code: 8642

WWP Wipo information: published in national office

Ref document number: 2001985271

Country of ref document: EP

WWW Wipo information: withdrawn in national office

Ref document number: 2001985271

Country of ref document: EP

WWG Wipo information: grant in national office

Ref document number: 2002223973

Country of ref document: AU

点击 这是indexloc提供的php浏览器服务，不要输入任何密码和下载