+

WO1997026009A1 - Composes capables d'inhiber l'infection due au vih-1 - Google Patents

Composes capables d'inhiber l'infection due au vih-1 Download PDF

Info

Publication number
WO1997026009A1
WO1997026009A1 PCT/US1997/000758 US9700758W WO9726009A1 WO 1997026009 A1 WO1997026009 A1 WO 1997026009A1 US 9700758 W US9700758 W US 9700758W WO 9726009 A1 WO9726009 A1 WO 9726009A1
Authority
WO
WIPO (PCT)
Prior art keywords
hiv
cell
infection
monoclonal antibody
molecule
Prior art date
Application number
PCT/US1997/000758
Other languages
English (en)
Inventor
Virginia M. Litwin
Graham P. Allaway
Paul J. Maddon
Original Assignee
Progenics Pharmaceuticals, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Progenics Pharmaceuticals, Inc. filed Critical Progenics Pharmaceuticals, Inc.
Priority to AU17504/97A priority Critical patent/AU1750497A/en
Publication of WO1997026009A1 publication Critical patent/WO1997026009A1/fr
Priority to US09/891,062 priority patent/US7118859B2/en
Priority to US11/544,346 priority patent/US7345153B2/en
Priority to US12/074,993 priority patent/US20080226651A1/en

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
    • A01K2217/00—Genetically modified animals
    • A01K2217/05—Animals comprising random inserted nucleic acids (transgenic)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • This invention comprises a series of new therapeutics and targets for therapeutic intervention in HIV-1 infection.
  • Monoclonal antibodies have been identified that inhibit HIV-1 envelope-mediated membrane fusion and thereby inhibit virus infection.
  • the antibodies were discovered by screening panels of monoclonal antibodies generated by immunizing mice with human cells. The screening was initially performed using a resonance energy transfer (RET) assay of HIV-1 envelope-mediated membrane fusion. Antibodies which inhibited in this assay were further screened for inhibitory activity in a HIV-1 infection assay.
  • RET resonance energy transfer
  • inhibitory antibodies act by binding to molecules on the surface of cells, which are required for HIV-1 to fuse with and infect target cells.
  • the molecules are either previously unidentified or their role in HIV-1 entry was previously unrecognized.
  • the cell surface molecules are known as accessory molecules, since they are required for virus entry in addition to the HIV-1 receptor, CD4. While CD4 is required for HIV-1 attachment, the accessory molecules are required for the membrane fusion step of entry.
  • CD4 While CD4 is required for HIV-1 attachment, the accessory molecules are required for the membrane fusion step of entry.
  • These accessory molecules are generally expressed only on human cells, so HIV-1 does not infect non-human cells that have been engineered to express human CD4 (1,3,8,9) .
  • accessory molecules are required for HIV-1 fusion.
  • the precise nature of these co-receptors or accessory molecules has not yet been discerned. While some cell surface molecules have previously been implicated as fusion accessory molecules (7,10,11) , their role has not been confirmed (4) .
  • the fusion accessory molecules are found on a subset of human CD4 + cells and are required for infection by HIV-1 isolates with particular tropisms .
  • macrophage-tropic strains of HIV-1 such as HIV-l jR .p L may have different requirements for accessory molecules compared with T lymphotropic strains such as HIV-I LAJ . This phenomenon is, in part, responsible for differences in tropism between HIV-1 strains.
  • the current invention includes the monoclonal antibodies and the hybridomas which secret them; also their humanized equivalents, single chain antibodies or antigen binding fragments of the antibodies. These antibodies, single chain antibodies or antibody fragments have value as immunotherapeutics or immunoprophylactics for HIV-1 infection.
  • the invention also includes the genes encoding these antibodies, single chain antibodies and antibody fragments.
  • the invention includes the accessory molecules recognized by these monoclonal antibodies, or components of these accessory molecules and region(s) of HIV-1 gpl20/gp41 that interact with these accessory molecule (s) .
  • the invention includes the genes encoding the accessory molecules.
  • the accessory molecules or their fragments have value as therapeutic or prophylactic agents to inhibit HIV-1 infection.
  • This invention provides transgenic animals comprising DNA encoding these accessory molecules or fragments thereof. These transgenic animals are useful as animal models of HIV-1 infection.
  • the invention also includes the use of the antibodies and/or accessory molecules in drug screening assays to identify inhibitors of HIV-1 fusion. Finally, the invention includes the inhibitors identified using these drug screens.
  • This invention provides an antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein" cell with an appropriate CD4 + cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1.
  • This invention also provides a pharmaceutical composition comprising the complete or a portion of the above- described antibody and a pharmaceutically acceptable carrier.
  • This invention provides a method of inhibiting HIV-1 infection in a subject comprising administering an effective amount of the above pharmaceutical composition to the subject.
  • This invention also provides nucleic acid molecules encoding the complete or a portion of the light chain and the heavy chain protein of the above antibody.
  • This invention also provides vectors comprising these nucleic acid molecules operably linked to a promoter of RNA transcription.
  • This invention also provides host vector systems comprising one or more of these vectors in a suitable host cell.
  • This invention provides the molecule specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein * cell with an appropriate CD4 + cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and which capable of inhibiting infection by one or more strains of HIV-1.
  • this is a glycolipid molecule.
  • this is a polypeptide.
  • This invention also provides an isolated nucleic acid molecule encoding the complete or a portion of this polypeptide.
  • the molecule is a multichain polypeptide molecule.
  • This invention also provides a soluble protein which comprises a portion of the polypeptide or the multichain polypeptide molecule.
  • This invention also provides a pharmaceutical composition comprising an effective amount of the soluble protein to inhibit HIV-1 infection and a pharmaceutically acceptable carrier.
  • This invention also provides a method of inhibiting HIV-1 infection in a subject comprising administering an effective amount of the above pharmaceutical composition to the subject.
  • This invention provides an isolated nucleic acid molecule encoding the complete or a portion of a polypeptide of the above multichain polypeptide molecule.
  • This invention also provides a vector comprising the nucleic acid molecule encoding the complete or a portion of a polypeptide of the above multichain polypeptide molecule operably linked to a promoter of RNA transcription, and a host vector system comprising this vector in a suitable host cell.
  • This invention also provides a method for identifying inhibitors of HIV-1 infection comprising steps of: (a) contacting an effective amount of a compound with a system which contains HIV-1 gpl20, HIV-1 gp41 or a fragment thereof with the molecule specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein " cell with an appropriate CD4 + cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1 under conditions permitting formation of a complex between HIV-1 gpl20, HIV-1 gp41 or a fragment thereof and the molecule, so as to inhibit such formation; and (b) determining the amount of complex formed; and (c) comparing the amount determined in step (b) with the control which is without the addition of the compound, a decrease in the complex formation indicating that the compound is capable of inhibiting HIV-1 infection.
  • This invention also provides the identified compound and a pharmaceutical composition comprising the identified compound and a pharmaceutically acceptable carrier.
  • This invention provides a method of inhibiting HIV-1 infection in a subject comprising administering an effective amount of this pharmaceutical composition to the subject.
  • This invention provides a kit for identifying inhibitors of HIV-1 infection which comprises, in separate compartments: (a) purified HIV-1 gpl20, HIV-1 gp41 or a fragment thereof; and (b) the molecule specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein" cell with an appropriate CD4 + cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1.
  • this invention provides a transgenic nonhuman animal which comprises an isolated DNA molecule encoding the molecule specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein* cell with an appropriate CD4" cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and which inhibits infection by one or more strains of HIV-1.
  • This invention also provides the above-described transgenic nonhuman animal which further comprises a DNA molecule encoding human CD4. This invention further provides different uses of the transgenic animals for screening and development of HIV-1 drugs.
  • FIG. 1 Flow cytometric analysis of human T lymphoblas oid cell lines with Ab PA-3, PA-5, PA-6 and PA-7. Cells were incubated for 15 min at 4°C with lOO ⁇ l of the indicated hybridoma supernatant (open histograms) , then washed 3 times in PBS containing 0.05% NaN 3 . Following a second incubation with FITC-conjugated goat anti-mouse antibody and additional washes the cells were analyzed by flow cytometry. Controls (shaded histograms) were stained with FITC-conjugated goat anti-mouse antibody only.
  • Fluorescence intensity is shown on the X-axis (four decade log scale) and the relative number of cells on the Y-axis.
  • Columns 1-4 are stained with mAb PA-3, PA-5, PA-6 and PA-7, respectively.
  • Rows 1-4 represent the cell lines PM-1, HUT 78, CEM and SUP-T1, respectively.
  • FIG.2A-2E Cell surface expression of antigens recognized by the mAb PA-3, PA-5, PA-6 and PA-7 on CD4- transfected cell lines and non-transfected counterparts.
  • a comparison of cell surface staining with PA-3, PA-5 and PA-6 and PA-7 was conducted on HeLa or HeLa-CD4 cells (Fig.2A-2E) and the Chinese hamster ovary cell line, DG44 or DG44 transfected with human CD4, DG44#3 (Fig.2F-2J) .
  • the CD4 expression on the transfectants and parental cells was demonstrated by staining with the CD4 specific mAb OKT4A.
  • PA-3, PA-5, VL125-6D1 recognize the same antigen.
  • PM-1 cells were surface labeled with biotin and immunoprecipitated with hybridoma supernatants PA-3, PA-5, VL125-6D1 and the CD4 specific antibody OKT4A (Ortho Diagnostics, Raritan, New Jersey) as described in the experimental methods.
  • Precipitated antigens were resolved on a 4-15% gradient polyacrylamide gel under reducing conditions. Gels were scanned using a Molecular Dynamics (Sunnyvale, CA) densitometer. Molecular weight markers are as indicated in the far left lane.
  • PA-3, PA-5 and VL125-6D1 precipitate two proteins of molecular weights approximately 158Kd and 87Kd.
  • Figure 4. Antigens recognized by PA-3 but not PA-7 co- migrate with LFA-1 (CDlla/CD18) .
  • PM-1 cells were surface labeled with biotin and i munoprecipitated with hybridoma supernatants PA-3 and PA-7 and the CD3 specific UCHTI
  • Antigens recognized by UCHTI did not resolve on this gel. Molecular weight markers are as indicated in the far left lane.
  • Monoclonal antibodies PA-6 and PA-7 recognize the same antigen. PM-1 cells were surface labeled with biotin and immunoprecipitated with hybridoma supernatants PA-3, PA-6, PA-7 and the CD4 specific antibody OKT4A Ortho Diagnostics) as described in the experimental methods. Precipitated antigens were resolved on a 4-15% gradient polyacrylamide gel under reducing conditions Gels were scanned using a Molecular Dynamics (Sunnyvale, CA) densitometer. molecular weight marker are as indicated in the far left lane. Monoclonal antibodies PA-6 and PA-7, precipitate two proteins of molecular weights approximately 24.8Kd and 19.1Kd. Figure 6. Antigens recognized by PA-7 co-migrate with HLA class II. PM-1 cells were surface labeled with biotin and immunoprecipitated with hybridoma supernatants PA-7 and the CD4 specific OKT4A (Ortho) and the HLA class II specific TU39
  • This invention provides an antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein * cell with an appropriate CD4 * cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1.
  • the antibody is a monoclonal antibody.
  • This invention also provides a hybridoma cell line producing the monoclonal antibody.
  • the antibody is a chimeric monoclonal antibody.
  • the antibody is a humanized monoclonal antibody.
  • the antibody is a human monoclonal antibody.
  • a "chimeric" monoclonal antibody is a murine monoclonal antibody comprising constant region fragments (F c ) from a different species.
  • the chimeric monoclonal antibody comprises human F c and murine F ab .
  • a "humanized" monoclonal antibody is a murine monoclonal antibody in which human protein sequences have been substituted for all the murine protein sequences except for the murine complementarity-determining regions (CDR) of both the light and heavy chains.
  • This invention also provides single chain antibodies or an antigen binding antibody fragment capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein * cell with an appropriate CD4 * cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1.
  • the fragments include but are not limited to Fab and Fab' .
  • the methods to generate the single chain antibodies and antigen binding antibody fragments with particular binding activities are well- known in the art (See for example, Crawley, P (1995) Antibody Patents, in Monoclonal Antibodies: Principles and Applications, Wiley-Liss, Inc. NY, pages 299-335) .
  • This invention also provides a monoclonal antibody capable of competitively inhibiting the binding of the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein * cell with an appropriate CD4 * cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1 to its target molecule.
  • the monoclonal antibody is labelled with a detectable marker, for example, a radioactive isotope, enzyme, dye or biotin.
  • a detectable marker for example, a radioactive isotope, enzyme, dye or biotin.
  • This invention provides a pharmaceutical composition comprising the complete or a portion of the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein* cell with an appropriate CD4 * cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1 and a pharmaceutically acceptable carrier.
  • pharmaceutically acceptable carriers means any of the standard pharmaceutical carriers.
  • suitable carriers are well known in the art and may include, but are not limited to, any of the standard pharmaceutical carriers such as phosphate buffered saline solutions, emulsions such as oil/water emulsion, and various type of wetting agents.
  • Other carriers may also include sterile solutions, tablets, coated tablets, and capsules.
  • Such carriers typically contain excipients such as starch, milk, sugar, certain types of clay, gelatin, stearic acid or salts thereof, magnesium or calcium stearate, talc, vegetable fats or oils, gums, glycols, or other known excipients.
  • excipients such as starch, milk, sugar, certain types of clay, gelatin, stearic acid or salts thereof, magnesium or calcium stearate, talc, vegetable fats or oils, gums, glycols, or other known excipients.
  • Such carriers may also include flavor and color additives or other ingredients.
  • Compositions comprising such carriers are formulated by well known conventional methods.
  • the monoclonal antibodies described and claimed herein are useful for isolating the compound to which the monoclonal antibodies bind.
  • This invention provides a method of inhibiting HIV-1 infection in a subject comprising administering an effective amount of the above-described pharmaceutical composition to the subject.
  • nucleic acid molecule encoding the complete or a portion of the light chain of the monoclonal antibody.
  • the nucleic acid molecule is a DNA molecule.
  • the DNA molecule is a cDNA molecule.
  • the DNA molecule is a genomic DNA molecule .
  • nucleic acid molecule encoding the complete or a portion of the heavy chain of the monoclonal antibody.
  • the nucleic acid molecule is a DNA molecule.
  • the DNA molecule is a cDNA molecule.
  • the DNA molecule is a genomic DNA molecule.
  • This invention provides an isolated nucleic acid molecule encoding the above-described single chain antibody.
  • the DNA molecules of the subject invention also include DNA molecules coding for polypeptide analogs, fragments or derivatives of polypeptides which differ from naturally-occurring forms in terms of the identity or location of one or more amino acid residues (deletion analogs containing less than all of the residues specified for the protein, substitution analogs wherein one or more residues specified are replaced by other residues and addition analogs wherein one or more amino acid residues is added to a terminal or medial portion of the polypeptides) and which share some or all of the properties of the naturally-occurring forms.
  • codons "preferred" for expression by selected non-mammalian hosts include: the incorporation of codons "preferred" for expression by selected non-mammalian hosts; the provision of sites for cleavage by restriction endonuclease enzymes; and the provision of additional initial, terminal or intermediate DNA sequences that facilitate construction of readily expressed vectors.
  • nucleic acid sequences described and claimed herein are useful for generating new viral and circular plasmid vectors described below.
  • This invention provides the vector, for example a plasmid or a viral vector, comprising a nucleic acid molecule encoding the complete or a portion of light chain protein of the monoclonal antibody operably linked to a promoter of RNA transcription.
  • This invention also provides a vector, for example a plasmid or a viral vector, comprising a nucleic acid molecule encoding the complete or a portion of heavy chain protein of the monoclonal antibody operably linked to a promoter of RNA transcription.
  • This invention also provides a vector comprising a nucleic acid molecule encoding the above-described single chain antibody operably linked to a promoter of RNA transcription.
  • This invention also provide a vector comprising the nucleic acid molecules encoding the complete or a portion of light chain protein and the complete or a portion of heavy chain protein of the monoclonal antibody each operably linked to a promoter of RNA transcription.
  • This invention also provide a host vector system comprising one or more vectors which comprise either the complete or portion of the light chain or a complete or portion of the heavy chain or a combination thereof in a suitable host cell.
  • This invention further provides the above host vector system, wherein the suitable host cell is selected from a group consisting of a bacterial cell, an insect cell, a yeast cell or a mammalian cell.
  • This invention provides the molecule specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein* cell with an appropriate CD4 * cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1.
  • the molecule is a glycolipid.
  • the molecule is a polypeptide.
  • This invention also provides an isolated nucleic acid molecule encoding the complete or a portion of the polypeptide which is specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein* cell with an appropriate CD4 * cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1.
  • This invention provides a multichain polypeptide molecule comprising the polypeptide which is specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein* cell with an appropriate CD4 * cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1.
  • This invention also provides an isolated nucleic acid molecule encoding the complete or a portion of a polypeptide of the above-described multichain polypeptide molecule.
  • This invention also provides vectors comprising the nucleic acid molecule encoding the complete or a portion of a polypeptide of the above- described multichain polypeptide molecule operably linked to a promoter of RNA transcription.
  • This invention also provides vectors comprising the nucleic acid molecule encoding the complete or a portion of the above polypeptide molecule operably linked to a promoter of RNA transcriptio .
  • This invention also provides a host vector system comprising the above vectors in a suitable host cell.
  • the suitable host cell includes but is not limited to a bacterial cell, an insect cell, a yeast cell, a plant cell or a mammalian cell.
  • the above-described molecules specifically recognized by the antibody may be used to vaccinate healthy subjects for prevention or therapy of HIV infection. These molecules are also useful in identifying specific sites to which these molecules bind on the HIV-1 envelope glycoproteins gpl20 and gp41. The sites revealed may be used to generate more specific antibodies directed to these sites. Moreover, this information may be used to design drugs which inhibit the binding of the HIV-1 envelope glycoproteins gpl20 or gp41 to the molecule.
  • This invention also provides a soluble protein which comprises a portion of the polypeptide or the multichain polypeptide molecule.
  • a "soluble protein” is a protein free of cell membranes and other cellular components.
  • the soluble protein is labelled with a detectable marker, for example, a radioactive isotope, enzyme, dye or biotin.
  • the soluble protein is valuable as a product for making a new and useful pharmaceutical composition.
  • This invention also provides a pharmaceutical composition comprising an effective amount of the above soluble protein to inhibit HIV-1 infection and a pharmaceutically acceptable carrier.
  • an effective amount is well known to those skilled in the art. Simple titration experiments using different amounts of the soluble protein administered to different animal models of HIV-1 infection or to HIV-1 infected human subjects may be performed to determine such an effective amount. The amount administered to the animal model of HIV-1 infection or HIV-1 infected humans which results in a reduction in HIV-1 infection is an effective amount.
  • This invention also provides a method of inhibiting HIV-1 infection in a subject comprising administering an effective amount of the pharmaceutical composition of the above soluble protein to the subject.
  • administration means any of the standard methods of administering a pharmaceutical composition known to those skilled in the art. Examples include, but are not limited to, intravenous, intraperitoneal or intramuscular administration.
  • nucleic acid molecule encoding the soluble protein.
  • the nucleic acid molecule is a DNA molecule.
  • the DNA molecule is a cDNA molecule.
  • the nucleic acid sequences which encode the soluble protein are useful for generating new viral and circular plasmid vectors described below.
  • the nucleic acid molecules are valuable in a new and useful method of gene therapy, i.e., by stably transforming cells isolated from a patient with the nucleic acid molecules and then readministering the stably transformed cells to the patient.
  • Methods of isolating cells include any of the standard methods of withdrawing cells from an animal. Suitable isolated cells include, but are not limited to, bone marrow cells.
  • Methods of readministering cells include any of the standard methods of readministering cells to a patient.
  • United States Patent 5,366,346, entitled, "Gene Therapy" describes different gene therapy procedures, the contents of which are incorporated herein by reference.
  • This invention also provides a vector, for example, a plasmid vector or a viral vector, comprising the isolated nucleic acid molecule operably linked to a promoter of RNA transcription.
  • a vector for example, a plasmid vector or a viral vector, comprising the isolated nucleic acid molecule operably linked to a promoter of RNA transcription.
  • the vectors described and claimed herein are valuable as products useful for generating stably transformed eukaryotic host cells, and thereby in new and useful methods of growing such host cells under conditions suitable for the production of the soluble protein.
  • This invention further provides a host vector system comprising the vector having the sequence which encodes the soluble protein in a suitable host cell.
  • the suitable host cell is a stably transformed eukaryotic cell, for example, a stably transformed eukaryotic yeast or mammalian cell.
  • the stably transformed cell is a mammalian cell.
  • the host vector system is valuable as a product useful for the large scale synthesis of the soluble protein by growing the host vector system under conditions suitable for the production of protein and recovering the protein so produced.
  • a method of producing the soluble protein is also provided.
  • This invention further provides the soluble protein produced by this method.
  • This invention provides a method for identifying inhibitors of HIV-1 infection comprising steps of: (a) contacting an effective amount of a compound with a system which contains HIV-1 gpl20, HIV-1 gp41 or a fragment thereof with the molecule specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein* cell with an appropriate CD4 * cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1 under conditions permitting formation of a complex between the HIV-1 gpl20, HIV-1 gp41 or a fragment thereof with the molecule, so as to inhibit such formation; (b) determining the amount of complex formed; and (c) comparing the amount determined in step (b) with the control which is without the addition of the compound, a decrease in the complex formation indicating that the compound is capable of inhibiting HIV-1 infection.
  • the compound tested is not previously known. This invention also provide the compound identified by the above method.
  • This invention also provides a pharmaceutical composition
  • a pharmaceutical composition comprising the compound identified by the above method and a pharmaceutically acceptable carrier.
  • This invention provides a method of inhibiting HIV-1 infection in a subject comprising administering an effective amount of the above pharmaceutical composition comprising the compound identified by the above method to the subject.
  • This invention provides a kit for identifying inhibitors of HIV-1 infection which comprises, in separate compartments: (a) purified HIV-1 gpl20, HIV-1 gp41 or a fragment thereof; and (b) the molecule specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein* cell with an appropriate CD4* cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1.
  • This invention provides a transgenic nonhuman mammal which comprises an isolated DNA molecule encoding a molecule specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein * cell with an appropriate CD4* cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1.
  • One means available for producing a transgenic animal is as follows: Female mice are mated, and the resulting fertilized eggs are dissected out of their oviducts. The eggs are stored in an appropriate medium such as M2 medium (Hogan B. et al. Manipulating the Mouse Embryo, A Laboratory Manual, Cold Spring Harbor Laboratory (1986) ) . DNA or cDNA encoding the molecule specifically recognized by the monoclonal antibody capable of specifically inhibiting the fusion of an HIV-1 envelope glycoprotein * cell with an appropriate CD4" cell without cross reacting with the HIV-1 envelope glycoprotein or CD4 and capable of inhibiting infection by one or more strains of HIV-1 is purified from a vector by methods well known in the art.
  • M2 medium Hogan B. et al. Manipulating the Mouse Embryo, A Laboratory Manual, Cold Spring Harbor Laboratory (1986)
  • Inducible promoters may be fused with the coding region of the DNA to provide an experimental means to regulate expression of the trans ⁇ gene.
  • tissue specific regulatory elements may be fused with the coding region to permit tissue-specific expression of the transgene.
  • the injected egg is then transferred into the oviduct of a pseudopregnant mouse (a mouse stimulated by the appropriate hormones to maintain pregnancy but which is not actually pregnant) , where it proceeds to the uterus, implants, and develops to term.
  • pseudopregnant mouse a mouse stimulated by the appropriate hormones to maintain pregnancy but which is not actually pregnant
  • microinjection is not the only method for inserting DNA into the egg cell, and is used here only for exemplary purposes.
  • CD4 may be expressed in the above transgenic animal system such that the animal will be susceptible to HIV-1 infection. Accordingly, this invention also provides the above transgenic nonhuman animals further comprising an isolated DNA molecule encoding the full-length or portion of the CD4 molecule sufficient for binding the HIV-1 envelope glycoprotein. These animal model systems are useful for screening compounds which are capable of inhibiting HIV-1 infection. Moreover, they are useful in predicting or evaluating possible therapeutic applications of HIV drugs. Therefore, this invention also provides a method for screening compounds using these transgenic animals.
  • This invention describes mAb which recognize accessory molecules required for HIV-1 membrane fusion and tropism. mAb to accessory molecules are first selected by the ability to inhibit HIV-1 envelope glycoprotein-mediated membrane fusion in the RET assay and then analyzed for the ability to inhibit HIV-1 infection in vi tro .
  • the immunizing cell line is washed extensively in PBS.
  • Five female six week old Balb/c mice (Charles River Laboratories) are immunized intraperitoneally (IP) with 5 x 10 ⁇ cells. Animals receive a minimum of two IP inoculations followed by a three week rest interval. Three days prior to splenectomy one animal receives an intravenous (IV) injection of 0.5 x 10 6 cells. The additional animals continue to receive immunizations according to this schedule.
  • IP intraperitoneally
  • IV intravenous
  • Hybridomas are plated in 96-well flat bottom tissue culture plates, selected in 1 ⁇ g/ml azaserine (Sigma) , 5 M hypoxanthine
  • RET Assay RET Assay .
  • the RET assay has been adapted to measure HIV-1 envelope glycoprotein-mediated membrane fusion mediated by the laboratory-adapted T cell-tropic strain HIV-l- j ⁇ and the macrophage-tropic clinical isolate HIV-l ⁇ . ⁇ .
  • HeLa-env ⁇ cells are able to fuse with a variety of CD4-expressing cells; whereas, HeLa-env JR _ FL cells are only able to fuse with PM1 cells, of the panel of cell lines tested.
  • HeLa cells stably expressing HIV-1 gpl20/gp41 are labeled with octadecyl fluorescein (F18) and mixed with octadecyl rhodamine (R18) -labeled CD4* target cells.
  • Membrane fusion of the labeled cells is measured by exciting F18 and measuring emission from R18 (RET) which occurs only when the dyes are closely associated in the same membrane.
  • RET octadecyl fluorescein
  • RET octadecyl rhodamine
  • Results from the RET assay are quantitatively determined using a fluorescence plate reader, data are transferred directly into a spreadsheet and the %RET and % inhibition of RET are automatically calculated. Hybridoma cell supernatants which achieve 50% inhibition or greater in the RET assay are selected for further analysis.
  • the percent inhibition of RET is defined as follows:
  • Max RET is the %RET value obtained at four hours with HeLa-env cells and CD4-expressing cells in the absence of an inhibitory compound
  • Exp RET is the %RET value obtained for the same cell combination in the presence of an inhibitory compound
  • Min RET is the background %RET value obtained using HeLa cells in place of HeLa envelope glycoprotein-expressing cells.
  • the novel mAb generated in this study are characterized. First mAb are tested for reactivity to human CD4. Any mAb found to be specific for human CD4 are not further analyzed and are not considered part of this invention.
  • the hybridoma cell lines which secrete mAb that do not react with CD4 are cloned and used for ascites production, and the mAb are isotyped and purified and tested for ability to inhibit HIV-1 infection.
  • Immulon 1 plates (Dynatech Laboratories, Chantilly, VA) are coated overnight with sCD4 (Progenies) at 120 ng/well in carbonate buffer at pH 9.4 at 4°C. Plates are then washed three times in PTB (PBS containing 0.5% Tween 20, 1% FCS and 0.1% BSA) . Following blocking with 5% BSA and three additional washes, plates are incubated for one hour in the presence of 100 ⁇ l of hybridoma supernatant or a standard. The anti-CD4 mAb OKT4A is used as the standard and added at concentrations ranging from 60 ng/ml to 3.25 ng/ml . Plates are washed three times before and after the addition of 100 ng/well of horseradish peroxidase-conjugated goat anti-mouse (Kirkegaard and Perry Laboratories, Gaithersburg, MD) .
  • O-phenylenediamine at 1.2 mg/ml in solution (50 mM Na 2 HP0 4 , 25 M citric acid and 1.2% H 2 0 2 ) is then added and the plates developed in the dark for 10-20 min. The reaction is terminated by the addition of 100 ⁇ l of 1.3 M H 2 S0 4 .
  • the A 492 is measured on a SLT 400 ATC plate reader (Tecan US, Research Triangle Park, NO and the data analyzed with the winSeLect software program (Tecan) .
  • Hybridoma cell lines are cloned twice by the limiting dilution method. Cells are adjusted in hybridoma culture media to a concentration of five cells per ml by serial dilution. To a 96-well round bottom microliter plate, a volume of 200 ⁇ l is pipetted, achieving a density of one cell per well. Supernatants are screened in the RET assay when the cells have reached confluency. Three clones from each of the original cell lines are selected based both on growth rate and the ability to inhibit HIV-1 envelope glycoprotein-mediated membrane fusion and are cryopreserved.
  • the best of the three clones from each group is re-cloned by limiting dilution. Again, the three best clones are cryopreserved. One clone from each of the original hybridomas is selected for ascites production. Prior to ascites production, the isotype of the clones is determined.
  • the isotype of the monoclonal antibody secreted by each of the twice-cloned hybridoma cell lines is determined using the ImmunoType kit (Sigma) . This is an ELISA-based method performed on nitrocellulose strips coated with anti-murine isotype specific antibody.
  • mice For each cloned hybridoma cell line, fifteen female six week old Balb/c mice (Charles River Laboratories) are primed with 0.5 ml of pristane (Sigma) . Following a ten day rest period, mice are injected intraperitoneally with 1 x 10 6 hybridoma cells. As ascites fluid accumulates the peritoneal cavity is drained and the fluid collected. Antibody purification
  • Antibody is purified directly from ascites fluid using the protein A-based ImmunoPure Plus purification kit (Pierce, Rockford, IL) . Ascites fluid is clarified by high speed centrifugation, is diluted and applied to a protein A agarose column. After the column has been extensively washed, antibody is eluted in a high pH buffer. Following neutralization and concentration, the A 280 is measured and protein concentration determined.
  • protein A-based ImmunoPure Plus purification kit Pieris, Rockford, IL
  • the antigens recognized by the novel mAb are characterized. Flow cytometric analysis is conducted in order to evaluate the distribution of antigen expression in hematopoietic cell populations, in cell lines known to be HIV-1 membrane fusion competent or resistant, and in non-primate cell lines. A biochemical analysis of the antigens recognized by the novel mAb is performed.
  • Candidate target epitopes include amino-acid epitopes on proteins, or carbohydrate epitopes on glycoproteins or glycolipids. These possibilities are distinguished by the following analyses. First, antigens are immunoprecipitated from cells in which the surface proteins have been labeled, followed by SDS-PAGE.
  • mAbs are recognizing carbohydrate epitopes
  • cells are treated with neuraminidase prior to FACS analysis and immunoprecipitation.
  • inhibition of mAb binding to cells by lectins is examined. If these methods indicate the mAb do not recognize proteins but do recognize a carbohydrate epitope, thin layer chromatography is used to determine if the antigens are glycolipids. Determination of cellular distribution of the molecules by flow cytometry.
  • PBMC peripheral blood mononuclear cell
  • the mononuclear cells are separated from peripheral blood using a Ficoll-Hypaque (Pharmacia) density gradient .
  • PBMC are stained with the hybridoma cell supernatants and antigen expression in the lymphoid and myeloid populations determined by forward and side-scatter gating.
  • the expression of the molecules on various cell lines is compared to the ability of these cell lines to fuse with HIV-1 envelope glycoprotein-expressing cells.
  • the distribution of expression in non-primate cell lines is also evaluated.
  • Immunoprecipitation analysis is used to determine whether proteins are recognized by the novel mAb. A non-radioactive immunodetection with ECL Western blotting (Amersham Life Science, Arlington
  • Biotin-labeled cells are lysed in 0.5% NP-40 Lysis Buffer (0.05 M Tris-HCl pH 8.0 containing 0.5% NP-40, 0.15 M NaCl, 0.02% NaN 3 , 20 KlU/ml aprotinin, 1 mM PMSF) .
  • Immune complexes are precipitated with rabbit anti-mouse (Pel-Frez) coated pansorbin (Calbiochem, La Jolla, CA) .
  • Electrophoresis is performed under reducing and non-reducing conditions to determine if the molecules exist in a monomeric or complexed form. ECL protein molecular weight markers are used to estimate the molecular weights of the molecules.
  • the carbohydrate modifications of these proteins are evaluated.
  • the immunoprecipitated-antigens are treated with neuraminidase (Sigma) , N-glycanase (Boehringer Mannheim) and O-glycanase (Genzyme, Cambridge, MA) , respectively.
  • neuraminidase Sigma
  • N-glycanase Boehringer Mannheim
  • O-glycanase Gene, Cambridge, MA
  • HIV-1 inhibition studies with the mAb are performed with a panel of diverse viruses, including both laboratory-adapted strains and primary isolates. Purified mAb are used in the infection experiments.
  • CD4-IgG2 a fusion protein between CD4 and IgG2 which neutralizes all strains of HIV-1, is used as a positive control inhibitor. Initially, three concentrations (50, 25 and 12.5 ⁇ g/ml) of the candidate monoclonal antibodies are tested for the ability to inhibit infection by the primary isolate and laboratory-adapted strain of HIV-1. If inhibition is evident, a dose response relationship is determined with serial five-fold dilutions, starting at 50 ⁇ g/ml, of the candidate mAb, CD4-IgG2 or other positive or negative control antibodies.
  • a comparison of the p24 antigen concentration in the mAb-treated and untreated cultures is used to determine the percent neutralization, IC 50 and IC 90 values by linear regression analysis.
  • the infection experiments are extended to include a larger panel of HIV-1 isolates if promising results are achieved in the preliminary experiments.
  • Alternative target cell lines may be used in place of PBMCs, for example, PM1 cells may be used for HIV-l JR _ FL infection experiments.
  • Applicants' initial strategy was to immunize with the human carcinoma cell line, HeLa, and screen for monoclonal antibodies (mAb) which inhibited HIV-1 envelope glycoprotein-mediated membrane fusion as detected in the RET assay using HeLa-env ⁇ and HeLa-CD4 cells. Following the first fusion of splenocytes from the HeLa-immunized mice to the murine melanoma fusion partner, SP2/0, hybridoma supernatants were screened in the RET assay but no inhibitory mAb were identified. This scheme was repeated twice more but no inhibitory mAb were identified.
  • the third strategy involved immunization with proteinase K digested human erythrocytes and screening with the RET assay using HeLa-env ⁇ and HeLa-CD4 cells.
  • This approach was used following a published study which had found that when human erythrocytes were fused to mouse CD4 * cells using polyethylene glycol, the heterokaryons became competent targets for HIV-1 envelope-mediated membrane fusion, suggesting that erythrocyte membranes contain fusion accessory molecules (6) .
  • proteinase K treatment of the erythrocytes did not abrogate this complementation.
  • HeLa cells HeLa-env JR . F which stably express the envelope glycoprotein from the JR-FL strain of HIV-1 were generated.
  • the RET assay was then adapted to measure the fusion of HeLa-env JR . FL with PM1 cells. PM1 cells were then used for immunizations.
  • the resulting hybridomas were screened in the RET assay using HeLa-env JR _ FL and PM1 cells. Several hybridomas which inhibited fusion by 50% or greater were selected for further analysis. Hybridomas were analyzed by RET assay, flow cytometry and ELISA assay.
  • hybridomas selected for further analysis secreted mAb which continue to inhibit in the RET assay using HeLa-env JR _ FL and PM1 cells.
  • Four of these hybridomas designated PA-3, PA-5, PA-6 and PA-7 have been cloned twice by the limiting dilution method and ascites fluid has been produced. mAb are currently being purified from the ascites fluid.
  • PA-6 nor PA-7 recognized CEM or SUP-T1 or significantly inhibited fusion of HeLa-e u v i with CEM or Sup-Tl.
  • PA-6 nor PA-7 recognized CEM or SUP-T1 or significantly inhibited fusion of HeLa-e u v i with CEM or Sup-Tl.
  • none of the four novel mAb recognized cell surface antigens on HeLa cells or significantly inhibited HeLa-CD4 fusion with HeLa-env ⁇ ]-.
  • PA-6 and PA-7 both precipitate two proteins of molecular weights approximately 24.8 Kd and 19.1 Kd and thus appear to recognize the same antigen ( Figure 5) . However it is unclear whether they recognize the same epitope. When a pan specific HLA class II mAb is used for immunoprecipitation, bands of similar mobilities to those precipitated with PA-6 and PA-7 are resolved ( Figure 6) . Thus PA-6 and PA-7 may recognize HLA class II. Inhibition of HrV-l NM _ 3 and HIV-1 JR _ FL infection by PA-3, PA-5 and CD4-IgG2.
  • Table 2 Inhibition of HIV-1 ⁇ . 3 and HlV-l ⁇ ⁇ . infection by PA-3, PA-5 and CD4-IgG2.
  • CD4-IgG2 is a fusion protein between CD4 and hviman IgG2 which neutralizes all strains of HIV- 1.
  • the T4 gene encodes the AIDS virus receptor and is expressed in the immune system and the brain. Cell 47:333-348.

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Organic Chemistry (AREA)
  • Immunology (AREA)
  • Medicinal Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Biophysics (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biochemistry (AREA)
  • Cell Biology (AREA)
  • Toxicology (AREA)
  • Zoology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Peptides Or Proteins (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)

Abstract

Cette invention se rapporte à un anticorps capable d'inhiber de manière spécifique la fusion d'une cellule de glycoprotéine+ d'enveloppe du VIH-1 avec une cellule CD4+ appropriée sans qu'il y ait réaction croisée avec la cellule glycoprotéine ou CD4 d'enveloppe du VIH-1, et d'inhiber l'infection par une ou plusieurs souches du VIH-1. Cet anticorps est également utilisé pour identifier une molécule qui a son importance dans l'infection due au VIH. Différentes utilisations de l'anticorps de la molécule sont également décrites.
PCT/US1997/000758 1996-01-17 1997-01-17 Composes capables d'inhiber l'infection due au vih-1 WO1997026009A1 (fr)

Priority Applications (4)

Application Number Priority Date Filing Date Title
AU17504/97A AU1750497A (en) 1996-01-17 1997-01-17 Compounds capable of inhibiting hiv-1 infection
US09/891,062 US7118859B2 (en) 1996-01-17 2001-06-25 Methods for inhibiting HIV-1 infection
US11/544,346 US7345153B2 (en) 1996-01-17 2006-10-05 Compounds capable of inhibiting HIV-1 infection
US12/074,993 US20080226651A1 (en) 1996-01-17 2008-03-07 Compounds capable of inhibiting hiv-1 infection

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US58745896A 1996-01-17 1996-01-17
US08/587,458 1996-01-17

Related Parent Applications (1)

Application Number Title Priority Date Filing Date
US58745896A Continuation-In-Part 1996-01-17 1996-01-17

Related Child Applications (1)

Application Number Title Priority Date Filing Date
US11841598A Continuation 1996-01-17 1998-07-17

Publications (1)

Publication Number Publication Date
WO1997026009A1 true WO1997026009A1 (fr) 1997-07-24

Family

ID=24349889

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US1997/000758 WO1997026009A1 (fr) 1996-01-17 1997-01-17 Composes capables d'inhiber l'infection due au vih-1

Country Status (2)

Country Link
AU (1) AU1750497A (fr)
WO (1) WO1997026009A1 (fr)

Cited By (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2000047216A1 (fr) * 1999-02-15 2000-08-17 Institut De Recherche Pour Le Developpement (I.R.D.) Compositions immunogenes utilisables comme vaccins
US6265184B1 (en) 1995-12-20 2001-07-24 Icos Corporation Polynucleotides encoding chemokine receptor 88C
US6528625B1 (en) 1996-10-28 2003-03-04 Millennium Pharmaceuticals, Inc. Anti-CCR5 antibodies and kits comprising same
US7060273B2 (en) 2001-04-06 2006-06-13 Progenics Pharmaceuticals, Inc. Methods for inhibiting HIV-1 infection
US7122185B2 (en) 2002-02-22 2006-10-17 Progenics Pharmaceuticals, Inc. Anti-CCR5 antibody
US7138119B2 (en) 2000-09-15 2006-11-21 Progenics Pharmaceuticals, Inc. Compositions and methods for inhibition of HIV-1 infection
US7211659B2 (en) 2001-07-05 2007-05-01 Chiron Corporation Polynucleotides encoding antigenic HIV type C polypeptides, polypeptides and uses thereof
US7282364B2 (en) 2001-08-31 2007-10-16 Novartis Vaccines And Diagnostics, Inc. Polynucleotides encoding antigenic HIV type B polypeptides, polypeptides and uses thereof
US7345153B2 (en) 1996-01-17 2008-03-18 Progenics Pharmaceuticals, Inc. Compounds capable of inhibiting HIV-1 infection
US7348177B2 (en) 1998-12-31 2008-03-25 Novartis Vaccines And Diagnostics, Inc. Expression of HIV polypeptides and production of virus-like particles
US7862994B2 (en) 1995-06-07 2011-01-04 Progenics Pharmaceuticals Inc. Methods for inhibiting HIV-1 envelope glycoprotein-medicated membrane fusion
US7943375B2 (en) 1998-12-31 2011-05-17 Novartis Vaccines & Diagnostics, Inc Polynucleotides encoding antigenic HIV type C polypeptides, polypeptides and uses thereof
US8821877B2 (en) 2005-07-22 2014-09-02 Cytodyn Inc. Methods for inhibiting HIV-1 replication involving the administration of an anti-CCR5 receptor monoclonal antibody and small molecule CCR5 receptor antagonist

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5071964A (en) * 1988-05-04 1991-12-10 Dana-Farber Cancer Institute, Inc. Protein micelles
US5091513A (en) * 1987-05-21 1992-02-25 Creative Biomolecules, Inc. Biosynthetic antibody binding sites

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5091513A (en) * 1987-05-21 1992-02-25 Creative Biomolecules, Inc. Biosynthetic antibody binding sites
US5071964A (en) * 1988-05-04 1991-12-10 Dana-Farber Cancer Institute, Inc. Protein micelles

Non-Patent Citations (9)

* Cited by examiner, † Cited by third party
Title
AIDS, December 1991, Vol. 5, BUSSO et al., "HIV-Induced Syncytium Formation Requires the Formation of Conjugates Between Virus-Infected and Uninfected T-Cells in Vitro", pages 1425-1432. *
EUR. J. IMMUNOL., 1994, Vol. 24, WANG et al., "Deletion of T Lymphocytes in Human CD4 Transgenic Mice Induced by HIV-gp120 and gp120-Specific Antibodies from AIDS Patients", pages 1553-1557. *
FEBS LETTERS, May 1989, Vol. 248, HATTORI et al., "Involvement of Tryptase-Related Cellular Protease(s) in Human Immunodeficiency Virus Type 1 Infection", pages 48-52. *
INTERNATIONAL CONFERENCE ON AIDS, 1992, Vol. 81, PA5 PoA 2017, SATO et al., "Anti-CD7 Reagents Inhibit HIV-1 Induced Syncytium Formation". *
J. OF IMMUNOLOGY, 01 September 1995, Vol. 155, SCHANBERG et al., "Characterization of Human CD7 Transgenic Mice", pages 2407-2418. *
J. OF IMMUNOLOGY, 15 January 1991, Vol. 146, ATTANASIO et al., "Anti-Idiotypic Antibody Response to Monoclonal Anti-CD4 Preparations in Nonhuman Primate Species", pages 507-514. *
J. OF IMMUNOLOGY, February 1990, Vol. 144, VALENTIN et al., "The Leukocyte Adhesion Glycoprotein CD18 Paricipates in HIV Induced Syncyia Formation in Monocytoid and T Cells", pages 934-937. *
PROC. NATL. ACAD. SCI. U.S.A., December 1989, Vol. 86, QUEEN et al., "A Humanized Antibody that Binds to the Interleukin 2 Receptor", pages 10029-10033. *
SCIENCE, 02 June 1989, Vol. 244, HILDRETH et al., "Involvement of a Leukocyte Adhesion Receptor (LFA-1) in HIV-Induced Syncytium Formation", pages 1075-1078. *

Cited By (20)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7862994B2 (en) 1995-06-07 2011-01-04 Progenics Pharmaceuticals Inc. Methods for inhibiting HIV-1 envelope glycoprotein-medicated membrane fusion
US6797811B1 (en) 1995-12-20 2004-09-28 Icos Corporation Antibodies to chemokine receptor 88C
US6268477B1 (en) 1995-12-20 2001-07-31 Icos Corporation Chemokine receptor 88-C
US7662548B2 (en) 1995-12-20 2010-02-16 Euroscreen S.A. ICOS Corporation Method of screening for modulators of HIV infection
US6265184B1 (en) 1995-12-20 2001-07-24 Icos Corporation Polynucleotides encoding chemokine receptor 88C
US7345153B2 (en) 1996-01-17 2008-03-18 Progenics Pharmaceuticals, Inc. Compounds capable of inhibiting HIV-1 infection
US6528625B1 (en) 1996-10-28 2003-03-04 Millennium Pharmaceuticals, Inc. Anti-CCR5 antibodies and kits comprising same
US7348177B2 (en) 1998-12-31 2008-03-25 Novartis Vaccines And Diagnostics, Inc. Expression of HIV polypeptides and production of virus-like particles
US7943375B2 (en) 1998-12-31 2011-05-17 Novartis Vaccines & Diagnostics, Inc Polynucleotides encoding antigenic HIV type C polypeptides, polypeptides and uses thereof
WO2000047216A1 (fr) * 1999-02-15 2000-08-17 Institut De Recherche Pour Le Developpement (I.R.D.) Compositions immunogenes utilisables comme vaccins
FR2789590A1 (fr) * 1999-02-15 2000-08-18 Inst Rechpour Le Dev I R D Compositions immunogenes utilisables comme vaccins
US7138119B2 (en) 2000-09-15 2006-11-21 Progenics Pharmaceuticals, Inc. Compositions and methods for inhibition of HIV-1 infection
US7060273B2 (en) 2001-04-06 2006-06-13 Progenics Pharmaceuticals, Inc. Methods for inhibiting HIV-1 infection
US7211659B2 (en) 2001-07-05 2007-05-01 Chiron Corporation Polynucleotides encoding antigenic HIV type C polypeptides, polypeptides and uses thereof
US8133494B2 (en) 2001-07-05 2012-03-13 Novartis Vaccine & Diagnostics Inc Expression cassettes endcoding HIV-1 south african subtype C modified ENV proteins with deletions in V1 and V2
US9598469B2 (en) 2001-07-05 2017-03-21 Novartis Vaccines And Diagnostics, Inc. HIV-1 south african subtype C env proteins
US7282364B2 (en) 2001-08-31 2007-10-16 Novartis Vaccines And Diagnostics, Inc. Polynucleotides encoding antigenic HIV type B polypeptides, polypeptides and uses thereof
US7122185B2 (en) 2002-02-22 2006-10-17 Progenics Pharmaceuticals, Inc. Anti-CCR5 antibody
US7666419B2 (en) 2002-02-22 2010-02-23 Progenics Pharmaceuticals Inc. Anti-CCR5 antibody
US8821877B2 (en) 2005-07-22 2014-09-02 Cytodyn Inc. Methods for inhibiting HIV-1 replication involving the administration of an anti-CCR5 receptor monoclonal antibody and small molecule CCR5 receptor antagonist

Also Published As

Publication number Publication date
AU1750497A (en) 1997-08-11

Similar Documents

Publication Publication Date Title
US7345153B2 (en) Compounds capable of inhibiting HIV-1 infection
JP7610302B2 (ja) Ｃｃｒ８抗体及びその用途
JP4642035B2 (ja) Ａｉｄｓ、ａｒｃおよびｈｉｖ感染の予防および治療に有用な抗ｃｄ４抗体ホモログ
Peterson et al. Genetic analysis of monoclonal antibody and HIV binding sites on the human lymphocyte antigen CD4
Mizukami et al. Binding region for human immunodeficiency virus (HIV) and epitopes for HIV-blocking monoclonal antibodies of the CD4 molecule defined by site-directed mutagenesis.
US5912176A (en) Antibodies against a host cell antigen complex for pre and post exposure protection from infection by HIV
CN106068275B (zh) 抗pd-1的人抗体
US5994515A (en) Antibodies directed against cellular coreceptors for human immunodeficiency virus and methods of using the same
US7060273B2 (en) Methods for inhibiting HIV-1 infection
WO1997026009A1 (fr) Composes capables d'inhiber l'infection due au vih-1
CZ261097A3 (cs) Chemokinové receptory 88-2B (CKR-3) a 88C a jejich protilátky
JPH03201995A (ja) 活性内皮細胞に対するモノクローナル抗体、該モノクローナル抗体およびそれらの抗原を用いた医薬品および治療法ならびに該モノクローナル抗体を用いた診断法
UA128600C2 (uk) Варіанти поліпептидів і їх застосування
US20020146415A1 (en) Methods for inhibiting HIV-1 infection
RU2322454C2 (ru) Антитело против ccr5
EP0910659B1 (fr) Anticorps diriges contre un complexe de cd4 et d'un domaine de recepteurs des chemokines, et leur utilisation pour lutter contre les infections a vih
WO2007094983A2 (fr) Procedes et compositions permettant l'inhibition d'une infection par le vih des lymphocytes t
PT1144006E (pt) Pa14: anticorpo contra o ccr5
WO1997049424A9 (fr) Anticorps anti-corecepteur cellulaire pour le virus de l'immunodeficience humaine et procedes d'utilisation
US7667006B2 (en) Anti-HIV antibody
MATSUSHITA et al. Characterization of a mouse/human chimeric monoclonal antibody (Cβ1) to a principal neutralizing domain of the human immunodeficiency virus type 1 envelope protein
Simon et al. Role of CD4 epitopes outside the gp120-binding site during entry of human immunodeficiency virus type 1
TW201927336A (zh) 細胞毒性ｔ細胞耗竭用組成物
JP2011024593A (ja) Ａｉｄｓ、ａｒｃおよびｈｉｖ感染の予防および治療に有用な抗ｃｄ４抗体ホモログ
EP1900376A2 (fr) Inhibition synergique de la fusion et la fixation de l'HIV-1 sur des compositions et anticorps

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AU CA JP US

AL Designated countries for regional patents

Kind code of ref document: A1

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

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

Ref country code: JP

Ref document number: 97526206

Format of ref document f/p: F

122 Ep: pct application non-entry in european phase
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