WO2002022077A2 - Compositions and methods for inhibition of hiv-1 infection - Google Patents
Compositions and methods for inhibition of hiv-1 infection Download PDFInfo
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- WO2002022077A2 WO2002022077A2 PCT/US2001/028756 US0128756W WO0222077A2 WO 2002022077 A2 WO2002022077 A2 WO 2002022077A2 US 0128756 W US0128756 W US 0128756W WO 0222077 A2 WO0222077 A2 WO 0222077A2
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- hiv
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- antibody
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Classifications
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- 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
- C07K16/2866—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against receptors for cytokines, lymphokines, interferons
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/395—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum
- A61K39/39533—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum against materials from animals
- A61K39/39541—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum against materials from animals against normal tissues, cells
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
- A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
-
- 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
- A61P31/14—Antivirals for RNA viruses
- A61P31/18—Antivirals for RNA viruses for HIV
Definitions
- HIV-1 entry into target cells proceeds at the cell surface through a cascade of events that include (1) binding of the viral surface glycoprotein gpl20 to cell surface CD , which is the primary receptor for HIV-1, (2) env binding to fusion coreceptors such as CCR5 and CXCR4 , and (3) multiple conformational changes in gp41.
- gp41 adopts transient conformations that include a prehairpin fusion intermediate that ultimately folds into a conformation capable of mediating fusion. These events culminate in fusion of the viral and cellular membranes and the subsequent introduction of the viral genome into the target cell. A similar sequence of molecular events is required for infection to spread via fusion of infected and uninfected cells . Each stage of the viral entry process can be targeted for therapeutic intervention.
- HIV-1 attachment can be inhibited both by agents that bind the viral envelope glycoproteins and by agents that bind human CD4.
- HIV-1 attachment can be inhibited by compounds that incorporate the gpl20 -binding domains of human CD4 and molecular mimics thereof [1-7] . Because this interaction between gpl20 and CD4 is essential for virus infection, CD -based molecules have the potential to target most if not all strains of HIV-1. In addition, viruses have limited ability to develop resistance to such molecules.
- CD4 -based molecules A number of recombinant CD4 -based molecules have been developed and tested for clinical activity in man. The first of these contained the four extracellular domains (D1-D4) of CD4 but lacked the transmembrane and intracellular regions. This molecule, termed soluble CD4 (sCD4) , demonstrated excellent tolerability when administered to humans at doses ranging to 10 mg/kg [8,9] . Transient reductions in plasma levels of infectious HIV-1 were observed in certain patients treated with sCD4. The short half-life of sCD4 in humans (45 minutes following intravenous administration) was identified as one obstacle to using this agent for chronic therapy.
- sCD4 soluble CD4
- CD4-based proteins were developed with increased serum half-life. These CD4-immunoglobulin fusion proteins comprised the D1D2 domains of CD4 genetically fused to the hinge CH2 and CH3 regions of human IgG molecules. These divalent proteins derive HIV-1 neutralizing capacity from their CD4 domains and Fc effector functions from the IgG molecule.
- a CD4-IgGl fusion protein was shown to have excellent tolerability and improved pharmacokinetics in Phase I clinical testing [10] . The antiviral evaluations were inconclusive.
- CD4-IgG2 fusion protein comprises the D1D2 domains of CD4 genetically fused to the heavy and light chain constant regions of human IgG2.
- This agent binds the HIV-1 envelope glycoprotein gpl20 with nanomolar affinity [5] and may inhibit virus attachment both by receptor blockade and by detaching gpl20 from the virion surface, thereby irreversibly inactivating the virus.
- CD4-IgG2 neutralized primary HIV-1 isolates derived from all major subtypes and outlier groups.
- CD4-IgG2 was similarly effective in neutralizing HIV-1 obtained directly from the plasma of seropositive donors in an ex vivo assay, indicating that this agent is active against the diverse viral quasispecies that are encountered clinically [12] .
- CD4-IgG2 also provided protection against infection by primary isolates in the hu-PBL-SCID mouse model of HIV-1 infection [13] . Recent analyses have demonstrated that CD4- IgG2 ' s ability to neutralize primary viruses is independent of their coreceptor usage [14] .
- CD4- IgG2 Compared with mono- or divalent CD4-based proteins, CD4- IgG2 has consistently demonstrated as much as 100-fold greater potency at inhibiting primary HIV-1 isolates [5,12,14,15] .
- the heightened potency may derive from CD4- IgG2 ' s ability to bind virions with increased valency/avidity and its steric juxtaposition of two gpl20 binding sites on each Fab-like arm of the immunoglobulin molecule.
- the larger Fab-like arms of CD4-IgG2 are also more likely to span HIV-1 envelope spikes on the virion.
- CD4-IgG2's anti-HIV-1 activity has been shown to compare favorably with those of the rare human monoclonal antibodies that broadly and potently neutralize primary HIV-1 isolates [5,11,14,15] .
- CD4-IgG2 therapy is in principle less susceptible to the development of drug-resistant viruses than therapies employing anti-env monoclonal antibodies or portions of the highly mutable HIV-1 envelope glycoproteins.
- CD4-IgG2 may have clinical utility as an agent that neutralizes cell-free virus before it has the opportunity to establish new rounds of infection.
- CD4-IgG2 may have utility in preventing infection resulting from occupational, perinatal or other exposure to HIV-1.
- CD4-IgG2 demonstrated excellent pharmacology and tolerability. In addition, measurable antiviral activity was observed by each of two measures. First, a statistically significant acute reduction in plasma HIV RNA was observed following administration of a single 10 mg/kg dose. In addition, sustained reductions in plasma levels of infectious HIV were observed in each of two patients tested. Taken together, these observations indicate that CD4-IgG2 possesses antiviral activity in humans [16] . In addition to CD -based proteins and molecular mimics thereof, HIV-1 attachment can also be inhibited by antibodies and nonpeptidyl molecules.
- inhibitors include (1) anti-env antibodies such as IgGlbl2 and F105 [17,18], (2) anti-CD4 antibodies such as OKT4A, Leu 3a, and humanized versions thereof [19,20], and (3) nonpeptidyl agents that target either gpl20 or CD4 [21] , [22-24] .
- the latter group of compounds includes aurintricarboxylic acids, polyhydroxycarboxylates, sulfonic acid polymers, and dextran sulfates.
- agents that block HIV-1 infection by targeting gp41 fusion intermediates. These inhibitors may interact with the fusion intermediates and prevent them from folding into final fusogenic conformations.
- the first such agents to be identified comprised synthetic or recombinant peptides corresponding to portions of the gp41 ectodomain predicted to form hydrophobic alpha helices. One such region is present in both the amino and carboxy segments of the extracellular portion of gp41, and recent crystallographic evidence suggests that these regions interact in the presumed fusogenic conformation of gp41 [25,26]. HIV-1 infection can be inhibited by agents that bind to either N- or C-terminal gp41 epitopes that are exposed during fusion.
- T-20 (formerly known as DP178)
- T-1249 T-1249
- DP107 DP107
- N34, C28 various fusion proteins and analogues thereof
- Other studies have identified inhibitors that comprise non-natural D-peptides and nonpeptidyl moieties [34,35] .
- Clinical proof-of-concept for this class of inhibitors has been provided by T-20, which reduced plasma HIV RNA levels by as much as 2 logs in Phase I/II human clinical testing [36] .
- the broad antiviral activity demonstrated for this class of inhibitors reflects the high degree of gp41 sequence conservation amongst diverse strains of HIV-1.
- This invention provides a composition which comprises an admixture of two compounds, wherein: (a) one compound is an antibody or portion thereof which binds to a CCR5 receptor; and (b) one compound retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate; wherein the relative mass ratio of the compounds in the admixture ranges from about 100:1 to about 1:100, the composition being effective to inhibit HIV-1 infection of the CD4+ cell.
- This invention provides a composition which comprises an admixture of three compounds, wherein: (a) one compound is an antibody or portion thereof which binds to a CCR5 receptor; (b) one compound retards attachment of HIV-1 to a CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell; and (c) one compound retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate; wherein the relative mass ratio of any two of the compounds in the admixture ranges from about 100:1 to about 1:100, the composition being effective to inhibit HIV-1 infection of the CD4+ cell.
- This invention provides a method of inhibiting HIV-1 infection of a CD4+ cell which comprises contacting the CD4+ cell with an amount of the composition of the subject invention effective to inhibit HIV-1 infection of the CD4+ cell so as to thereby inhibit HIV-1 infection of the CD4+ cell .
- This invention provides a method of inhibiting HIV-1 infection of a CD4+ cell which comprises contacting the CD4+ cell with (1) an amount of an antibody which binds to a CCR5 receptor and (2) an amount of a compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope- on a gp41 fusion intermediate, so as to thereby inhibit HIV-1 infection of the CD4+ cell.
- This invention provides a method of inhibiting HIV-1 infection of a CD4+ cell which comprises contacting the CD4+ cell with (1) an amount of an antibody which binds to a CCR5 receptor, (2) an amount of a compound which retards attachment of HIV-1 to the CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell effective to inhibit HIV-1 infection of the CD4+ cell, and (3) an amount of a compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate, so as to thereby inhibit HIV-1 infection of the CD4+ cell.
- CD4-IgG2, T-20 and fixed mass ratios thereof were analyzed in the RET assay for the ability to inhibit env-mediated membrane fusion.
- the 25:1 (high) combination examined 10 three-fold serial dilutions of 250 ⁇ g/ml CD4-IgG2, 10 ⁇ g/ml T-20 and their combination.
- the 25:1 (low) combination examined 10 threefold serial dilutions of 50 ⁇ g/ml CD4-IgG2, 2 ⁇ g/ml T-20 and their combination.
- the 5:1 combination examined 10 three-fold serial dilutions of 50 ⁇ g/ml CD4-IgG2, 2 ⁇ g/ml T-20, and their combination.
- the 1:1 combination examined 10 three-fold serial dilutions of 10 ⁇ g/ml CD4-IgG2, 10 ⁇ g/ml T-20 and their combination.
- Inhibition data from three or more independent assays were averaged prior to analysis.
- Dose-response curves for the various inhibitors and combinations were fit to Equation (1) , which was then rearranged to calculate the inhibitor concentrations required to effect a given percent inhibition.
- the concentrations of the individual agents in an inhibitory mixture were calculated from their known mass ratios. These values were then used to calculate the Combination Index
- CI 1 indicates additive effects, and CI> 1 indicates antagonism.
- CD4-IgG2 and T-20 Dose reductions observed for synergistic combinations of CD4-IgG2 and T-20.
- CD4-IgG2, T-20 and a 25:1 fixed mass ratio thereof were tested in the RET assay for the ability to inhibit env-mediated membrane fusion.
- Inhibition data from six independent .assays were averaged.
- K and m were determined by curve-fitting the dose-response curves, and Equation (1) was rearranged to allow calculation of c for a given f for the single agents and their combination.
- Dose Reduction is the ratio of the inhibitor concentrations required to achieve a given degree of inhibition when the inhibitor is used alone v. in a synergistic combination.
- CD4-IgG2, PRO 140, PA12 and T-20 were tested individually and in combination for the ability to inhibit HIV env-mediated membrane fusion in the RET assay.
- PRO 140, T-20 and a 1:30 fixed molar ratio thereof where Dose Reduction is the ratio of the inhibitor concentrations required to achieve a given degree of inhibition when the inhibitor is used alone v.
- PRO 542, PRO 140 and T-20 Synergistically Blocks HIV-1 Entry.
- PRO 542, PRO 140 and T- 20 were used alone and in -3:1:30 molar combination to inhibit HIV-1 JR _ PL env-mediated cell-cell fusion. The methodology for this assay is described in Litwin et al . (67) .
- CD4-IgG2 (-- ⁇ --), T-20 (--•--), and a 25:1 CD4-IgG2 :T-20 combination (•••A---) were analyzed for inhibition of HIV-1 entry in an env-mediated membrane fusion (RET) assay.
- Inhibitors were added to a mix of HeLa-Eny JR - FL and PM1 cells previously labeled with F18 and R18 respectively. Fluorescence RET was measured after 4h of incubation, and percent inhibition was calculated as described [19] . Results are mean values from three independent experiments. The data were analyzed according to the median effect principle described in Equation (1) .
- Figure 7 Combination indices for inhibition of HIV-1 entry by combinations of CD4-IgG2 and T-20.
- CD4-IgG2, T-20 and fixed mass ratios thereof were analyzed in the RET assay for the ability to inhibit env-mediated membrane fusion.
- the 25:1 (high) combination examined 10 three-fold serial dilutions of 250 ⁇ g/ml CD4-IgG2, 10 ⁇ g/ml T-20 and their combination.
- the 25:1 (low) combination examined 10 threefold serial dilutions of 50 ⁇ g/ml CD4-IgG2, 2 ⁇ g/ml T-20 and their combination.
- the 5:1 combination examined 10 three-fold serial dilutions of 50 ⁇ g/ml CD4-IgG2, 2 ⁇ g/ml T-20, and their combination.
- the 1:1 combination examined 10 three-fold serial dilutions of 10 ⁇ g/ml CD4-IgG2, 10 ⁇ g/ml T-20 and their combination.
- Inhibition data from three or more independent assays were averaged prior to analysis.
- Dose-response curves for the various inhibitors and combinations were fit to Equation (1) , which was then rearranged to calculate the inhibitor concentrations required to effect a given percent inhibition.
- the concentrations of the individual agents in an inhibitory mixture were calculated from their known mass ratios . These values were then used to calculate the Combination Index (CI) according to Equation (2) .
- CI ⁇ 1 indicates synergy
- CI> 1 indicates antagonism.
- CD4-IgG2 and T-20 Dose reductions observed for synergistic combinations of CD4-IgG2 and T-20.
- CD4-IgG2, T-20 and a 25:1 fixed mass ratio thereof were tested in the RET assay for the ability to inhibit env-mediated membrane fusion.
- Inhibition data from six independent assays were averaged.
- K and m were determined by curve-fitting the dose-response curves, and Equation (1) was rearranged to allow calculation of c for a given f for the single agents and their combination.
- Dose Reduction is the ratio of the inhibitor concentrations required to achieve a given degree of inhibition when the inhibitor is used alone v. in a synergistic combination.
- FIG. 9 Inhibition of HIV-1 JR .
- the PRO 542: T-20 molar ratio was 1:2 (A and C) or 1:10 (B) .
- the concentrations of PRO 542 and T-20 in the mixtures are plotted for the purposes of illustrating dose reductions (A and B) or lack thereof (C) .
- the drugs were pre-incubated with HeLa-Env cells for 2 h at 25 °C prior to washing, warming and addition of PM1 cells.
- Figure 10 Combination index values and dose reductions for inhibition if HIV-1 entry with combinations of PR0542 and T-20.
- HIV-1 entry involves at least three steps that provide targets for therapy: gpl20-CD4 attachment; gpl20-coreceptor interactions; gp41-mediated fusion.
- Figure 12 PRO 542 (CD4-IgG2) attachment inhibitor.
- RET Resonance Energy Transfer
- PRO140, PR0542 and T-20 Triple Combination Synergistically Blocks HIV-1 Entry PRO140, PR0542 and T-20 used in a 1:3:30 molar ratio.
- PR0542 does not potentiate T-20 activity in the absence of coreceptor. PR0542 alone ( ⁇ • ⁇ A---), T-20 alone (--•--), PRO542:T-20 cocktail (-- ⁇ --),. ( ) PR0542 in cocktail .
- PR0542 increases the half- life of the T-20 targets.
- Synergistic inhibition of HIV-1 entry PRO 542, T-1249, and a 1.-10 molar combination (4:1 mass combination) of PRO 542:T-1249 were analyzed for inhibition of HIV-1 entry in an env-mediated membrane fusion (RET) assay. Inhibitors were added to a mix of HeLa-EnvJR-FL + and PM1 cells previously labeled with F18 and R18 respectively. Fluorescence RET was measured after 4 hours of incubation, and percent inhibition was calculated as described in Litwin, et al . (J. Virol. 70: 6437, 1996). The concentrations of PRO 542 and T-1249 present in the cocktail are derived from the curve fit of the data and are shown for the purposes of illustrating dose reductions.
- RET env-mediated membrane fusion
- the plasmids CD4-IgG2-HC-pRcCMV and CD4-kLC-pRcCMV were deposited pursuant to, and in satisfaction of, the requirements of the Budapest Treaty on the International Recognition of the Deposit of Microorganisms (the "Budapest Treaty") for the Purposes of Patent Procedure with the American Type Culture Collection (ATCC) , 10801 University Boulevard, Manassas, VA 20110-2209 under ATCC Accession Nos. 75193 and 75194, respectively.
- the plasmids were deposited with ATCC on Jan. 30, 1992.
- the plasmid designated pMA243 was similarly deposited in accordance with the Budapest Treaty with ATCC under Accession No. 75626 on December 16, 1993.
- the murine hybridomas PA8, PA9, PA10, PA11, PA12 and PA14 were deposited pursuant to, and in satisfaction of, the requirements of the Budapest Treaty on the International Recognition of the Deposit of Microorganisms (the "Budapest Treaty") for the Purposes of Patent Procedure with the American Type Culture Collection (ATCC) , 10801 University Boulevard, Manassas, VA 20110-2209 under the following ATCC Accession Nos. : PA8 (ATCC No. HB-12605) , PA9 (ATCC No . HB- 12606), PA10 (ATCC No. HB-12607) , PAH (ATCC No . HB-12608) , PA12 (ATCC No. HB-12609) and PA14 (ATCC No. HB-12610) .
- the hybridomas were deposited on December 2, 1998.
- This invention provides a composition which comprises an admixture of two compounds, wherein: (a) one compound is an antibody or portion thereof which binds to a CCR5 receptor; and (b) one compound retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate; wherein the relative mass ratio of the compounds in the admixture ranges from about 100:1 to about 1:100, the composition being effective to inhibit HIV-1 infection of the CD4+ cell.
- This invention provides a composition which comprises an admixture of three compounds, wherein: (a) one compound is an antibody or portion thereof which binds to a CCR5 receptor; (b) one compound retards attachment of HIV-1 to a CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell; and (c) one compound retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate; wherein the relative mass ratio of any two of the compounds in the admixture ranges from about 100:1 to about 1:100, the composition being effective to inhibit HIV-1 infection of the CD4+ cell.
- This invention provides a composition which comprises an admixture of two compounds, wherein one compound retards attachment of HIV-1 to a CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell and the other compound retards gp41 from adopting a conformation capable of mediating fusion of HIV- 1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate, wherein the relative mass ratio of the compounds in the admixture ranges from about 100:1 to about 1:100, the composition being effective to inhibit HIV-1 infection of the CD4+ cell.
- HIV-1 means the human immunodeficiency virus type-1. HIV-1 includes but is not limited to extracellular virus particles and the forms of HIV-1 associated with HIV-1 infected cells.
- HIV-I JR . FL is a strain that was originally isolated at autopsy from the brain tissue of an AIDS patient [47] . The virus was co-cultured with lectin-stimulated normal human peripheral blood mononuclear cells. The virus has been cloned and the DNA sequences of its envelope glycoproteins are known (Genbank Accession #U63632) . In terms of sensitivity to inhibitors of viral entry, HIV-1 JR . PL is known to be highly representative of primary HIV-1 isolates [11,14,15,48-50].
- gp41 fusion intermediates includes structures, conformations, and oligomeric states that are preferentially and transiently presented or exposed on the HIV-1 envelope glycoprotein gp41 during the process of HIV- 1 env-mediated membrane fusion. These intermediates may form upon interaction of HIV-1 with cellular receptors or may be present in partially or fully occluded states on HIV-1 prior to its interaction with cellular receptors. "gp41 fusion intermediates” do not include fusogenic gp41 conformations that cannot provide targets for therapeutic intervention .
- the gp41 fusion intermediates may contain multiple epitopes that are transiently exposed during fusion and can provide targets for therapeutic intervention.
- an "N-terminal gp41 epitope” may comprise all or portions of the sequences from amino acid A541 to Q590.
- a "C-terminal gp41 epitope” may comprise all or portions of the sequences from amino acid W628 to L663. These epitopes have the potential to form coiled-coils of interacting alpha helical segments by virtue of heptad (sequence of seven amino acids) repeats containing hydrophobic amino acids at positions 1 and 4 of the heptad.
- the amino acid numbering system is for the HxB2 isolate of HIV-1 (Genbank Protein Accession No. AAB50262) . Because of the sequence variability of HIV-1 envelope proteins, the composition, size and precise location of such sequences may be different for different viral isolates.
- the gp4 1 fusion intermediates may also present other linear or conformational epitopes that are transiently expressed during HIV-1 entry.
- An inhibitor may target multiple epitopes present on gp41 fusion intermediates. Alternatively, separate inhibitors may be used in combination to target one or more epitopes present on gp41 fusion intermediates.
- HIV-1 fusion coreceptor means a cellular receptor that mediates fusion between the target cell expressing the receptor and HIV-1 or an HIV-1 envelope glycoprotein* cell. HIV-1 fusion co-receptors include but are not limited to CCR5, CXCR4 and other chemokine receptors.
- fusion means the joining or union of the lipid bilayer membranes found on mammalian cells or viruses such as HIV-1. This process is distinguished from the attachment of HIV-1 to a target cell. Attachment is mediated by the binding of the HIV-1 exterior glycoprotein to the human CD4 receptor, which is not a fusion co-receptor.
- “retards” means that the amount is reduced.
- attachment means the process that is mediated by the binding of the HIV-1 envelope glycoprotein to the human CD4 receptor, which is not a fusion coreceptor.
- CD4 means the mature, native, membrane-bound CD4 protein comprising a cytoplasmic domain, a hydrophobic transmembrane domain, and an extracellular domain which binds to the HIV-1 gpl20 envelope glycoprotein .
- epitope means a portion of a molecule or molecules that form a surface for binding antibodies or other compounds.
- the epitope may comprise contiguous or noncontiguous amino acids, carbohydrate or other nonpeptidyl moities or oligomer-specific surfaces.
- the compounds of the subject invention have shown to demonstrate a synergistic effect.
- synergistic means that the combined effect of the compounds when used in combination is greater than their additive effects when used individually.
- the compound which retards attachment of HIV-1 to the CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell is a CD4-based protein.
- CD4-based protein means any protein comprising at least one sequence of amino acid residues corresponding to that portion of CD4 which is required for
- CD4 to form a complex with the HIV-1 gpl20 envelope glycoprotein.
- the CD4-based protein is a CD4- immunoglobulin fusion protein.
- the CD4- immunoglobulin fusion protein is CD4-IgG2, wherein the CD4- IgG2 comprises two heavy chains and two lights chains, wherein the heavy chains are encoded by an expression vector designated CD4-IgG2HC-pRcCMV (ATCC Accession No. 75193) and the light chains are encoded by an expression vector designated CD4-kLC-pRcCMV (ATCC Accession No. 75194) .
- CD4-IgG2 is also referred to as PRO 542.
- the compound which retards attachment of HIV-1 to the CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD44- cell is a protein, the amino acid sequence of which comprises that of a protein found in HIV-1 as an envelope glycoprotein.
- the protein binds to an epitope of CD4 on the surface of the CD4+ cell.
- the envelope glycoprotein is selected from the group consisting of gpl20, gpl60, and gpl40.
- the compound which retards the attachment of HIV-1 to the CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell is an antibody or portion of an antibody.
- the antibody is a monoclonal antibody.
- the monoclonal antibody is a human, humanized or chimeric antibody.
- the portion of the antibody is a Fab fragment of the antibody.
- the portion of the antibody comprises the variable domain of the antibody.
- the portion of the antibody comprises a CDR portion of the antibody.
- the monoclonal antibody is an IgG, IgM, IgD, IgA, or IgE monoclonal antibody.
- antibody means an immunoglobulin molecule comprising two heavy chains and two light chains and which recognizes an antigen.
- the immunoglobulin molecule may derive from any of the commonly known classes, including but not limited to IgA, secretory IgA, IgG and IgM.
- IgG subclasses are also well known to those in the art and include but are not limited to human IgGl, IgG2 , IgG3 and IgG4. It includes, by way of example, both naturally occurring and non-naturally occurring antibodies.
- antibody includes polyclonal and monoclonal antibodies, and monovalent and divalent fragments thereof.
- antibody includes chimeric antibodies, wholly synthetic antibodies, single chain antibodies, and fragments thereof.
- the antibody may be a human or nonhuman antibody.
- a nonhuman antibody may be humanized by recombinant methods to reduce its immunogenicity in man. Methods for humanizing antibodies are known to those skilled in the art.
- humanized describes antibodies wherein some, most or all of the amino acids outside the CDR regions are replaced with corresponding amino acids derived from human immunoglobulin molecules. In one embodiment of the humanized forms of the antibodies, some, most or all of the amino acids outside the CDR regions have been replaced with amino acids from human immunoglobulin molecules but where some, most or all amino acids within one or more CDR regions are unchanged. Small additions, deletions, insertions, substitutions or modifications of amino acids are permissible as long as they would not abrogate the ability of the antibody to bind a given antigen.
- Suitable human immunoglobulin molecules would include IgGl, IgG2 , IgG3 , IgG4, IgA and IgM molecules.
- a "humanized" antibody would retain a similar antigenic specificity as the original antibody, i.e., in the present invention, the ability to bind CCR5.
- United States Patent No. 5,225,539 describes another approach for the production of a humanized antibody.
- This patent describes the use of recombinant DNA technology to produce a humanized antibody wherein the CDRs of a variable region of one immunoglobulin are replaced with the CDRs from an immunoglobulin with a different specificity such that the humanized antibody would recognize the desired target but would not be recognized in a significant way by the human subject's immune system.
- site directed mutagenesis is used to graft the CDRs onto the framework.
- WO 90/07861 which describe methods for producing humanized immunoglobulins . These have one or more CDRs and possible additional amino acids from a donor immunoglobulin and a framework region from an accepting human immunoglobulin. These patents describe a method to increase the affinity of an antibody for the desired antigen. Some amino acids in the framework are chosen to be the same as the amino acids at those positions in the donor rather than in the acceptor. Specifically, these patents describe the preparation of a humanized antibody that binds to a receptor by combining the CDRs of a mouse monoclonal antibody with human immunoglobulin framework and constant regions. Human framework regions can be chosen to maximize homology with the mouse sequence.
- a computer model can be used to identify amino acids in the framework region which are likely to interact with the CDRs or the specific antigen and then mouse amino acids can be used at these positions to create the humanized antibody.
- the above patents 5,585,089 and 5,693,761, and WO 90/07861 (62) also propose four possible criteria which may be used in designing the humanized antibodies. The first proposal was that for an acceptor, use a framework from a particular human immunoglobulin that is unusually homologous to the donor immunoglobulin to be humanized, or use a consensus framework from many human antibodies .
- the second proposal was that if an amino acid in the framework of the human immunoglobulin is unusual and the donor amino acid at that position is typical for human sequences, then the donor amino acid rather than the acceptor may be selected.
- the third proposal was that in the positions immediately adjacent to the 3 CDRs in the humanized immunoglobulin chain, the donor amino acid rather than the acceptor amino acid may be selected.
- the fourth proposal was to use the donor amino acid reside at the framework positions at which the amino acid is predicted to have a side chain atom within 3A of the CDRs in a three dimensional model of the antibody and is predicted to be capable of interacting with the CDRs.
- the above methods are merely illustrative of some of the methods that one skilled in the art could employ to make humanized antibodies.
- the monoclonal antibody binds to an HIV- 1 envelope glycoprotein.
- the HIV-1 envelope glycoprotein is selected from the group consisting of gpl20 and gpl60.
- the HIV-1 envelope glycoprotein is gpl20 and the monoclonal antibody which binds to gpl20 is IgGlbl2 or F105.
- IgGlbl2 is listed as item #2640 in the NIH AIDS Research and Reference Reagent Program Catalog.
- F105 is listed as item #857 in the NIH AIDS Research and Reference Reagent Program Catalog.
- the antibody binds to an epitope of CD4 on the surface of the CD4+ cell.
- the compound which retards attachment of HIV-1 to the CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell is a peptide.
- the compound which retards attachment of HIV-1 to the CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell is a nonpeptidyl agent.
- nonpeptidyl means that the agent does not consist in its entirety of a linear sequence of amino acids linked by peptide bonds. A nonpeptidyl agent may, however, contain one or more peptide bonds.
- the compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate is an antibody.
- the antibody is a monoclonal antibody.
- the antibody is a polyclonal antibody.
- the compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate is a peptide.
- the compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate is a fusion protein which comprises a peptide which inlcudes but is not limited to T-20 (SEQ ID NO: 1) , DP107 (SEQ ID NO: 2), N34 (SEQ ID NO: 3), C28 (SEQ ID NO: 4), and N34(L6)C28 (SEQ ID NO: 5).
- the peptide is selected from the group consisting of T-20 (SEQ ID NO: 1) , DP107 (SEQ ID NO: 2), N34 (SEQ ID NO: 3), C28 (SEQ ID NO: 4), and N34(L6)C28 (SEQ ID NO: 5).
- the peptide is selected from the group consisting of T-20 (SEQ ID NO: 1) , DP107 (SEQ ID NO: 2), N34 (SEQ ID NO: 3), C28
- the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20 (SEQ ID NO: 6). In one embodiment, the peptide is T-20
- T-20 and DP178 are used interchangeably to denote a peptide having the following amino acid sequence: YTSLIHSLIEESQNQQE NEQELLELDKWASLWNWF (SEQ ID NO: 1) and as described [29,32].
- DP107 has the following amino acid sequence: NNLLRAIEAQQHLLQLTVWGIKQLQARILAV ⁇ RYLKDQ
- N34 has the following amino acid sequence:
- C28 has the following amino acid sequence:
- N34(L6)C28 has the following amino acid sequence: SGIVQQQNNLLRAIEAQQHLLQLTVWGIKQLQARSGGRGGWMEWDREINNYTSLIHS LIEESQNQQEK (SEQ ID NO: 5) .
- T-1249 has the following amino acid sequence : WQEWEQKITALLEQAQIQQEKNEYELQ LDKWASLWEWF (SEQ ID NO: 6) .
- the peptide is a mutant peptide which (1) consists of amino acids having a sequence identical to that of a wildtype peptide selected from the group consisting of T-20 (SEQ ID NO: 1), DP-107 (SEQ ID NO: 2), N34 (SEQ ID NO: 3), C28 (SEQ ID NO: 4), N34(L6)C28 (SEQ ID NO: 5) and T-1249 (SEQ ID NO: 6), except for an addition of at least one glycine residue to a 5 ' end of the peptide, to a 3 ' end of the peptide, or to both ends of the peptide and (2) retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate.
- T-20 SEQ ID NO: 1
- DP-107 SEQ ID NO: 2
- N34 SEQ ID NO: 3
- C28 SEQ ID NO: 4
- the compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV- 1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate is a fusion protein which comprises a peptide selected from the group consisting of T-20 (SEQ ID NO: 1), DP107 (SEQ ID NO: 2), N34 (SEQ ID NO: 3), C28 (SEQ ID NO: 4), N34(L6)C28 (SEQ ID NO: 5), and T- 1249 (SEQ ID NO:6) .
- the peptide is selected from the group consisting of T-20 (SE Q ID NO: 1), DP107 (SEQ ID NO: 2), N34 (SEQ ID NO: 3), C28 (SEQ ID NO: 4), N34(L6)C28 (SEQ ID NO: 5), and T-1249 (SEQ ID NO : 6) .
- the peptide is T-20 (SEQ ID NO: 1) . In one embodiment of the compositions described herein, the peptide is T-1249 (SEQ ID NO: 6)
- the compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp4l fusion intermediate is a non-peptidyl agent.
- a non-peptidyl agent of the subject invention is a small molecule.
- the nonpeptidyl agent is a compound having a molecular weight less than 500 daltons.
- the antibody which binds to a CCR5 receptor includes but is not limited to PA8 (ATCC Accession No. HB-12605) , PA10 (ATCC Accession No.12607), PAH (ATCC Accession No. HB-12608) , PA12 (ATCC Accession No. HB-12609) , and PA14 (ATCC Accession No. HB-12610) .
- the antibody is a monoclonal antibody.
- the monoclonal antibody is a human, humanized or chimeric antibody.
- the portion of the antibody is a Fab fragment of the antibody.
- the portion of the antibody comprises the variable domain of the antibody.
- the portion of the antibody comprises a CDR portion of the antibody.
- the monoclonal antibody is an IgG, IgM, IgD, IgA, or IgE monoclonal antibody.
- the relative mass ratio of each such compound in the admixture ranges from about 25:1 to about 1:1. In one embodiment, the mass ratio is about 25:1. In one embodiment, the mass ratio is about 5:1. In one embodiment, the mass ratio is about 1:1.
- the composition is admixed with a carrier.
- the carriers of the subject invention include but are not limited to aerosol, intravenous, oral or topical carriers.
- Pharmaceutically acceptable carriers are well known to those skilled in the art .
- Such pharmaceutically acceptable carriers may include but are not limited to aqueous or non- queous solutions, suspensions, and emulsions.
- non-aqueous solvents are propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate.
- Aqueous carriers include water, alcoholic/aqueous solutions, emulsions or suspensions, saline and buffered media.
- Parenteral vehicles include sodium chloride solution, Ringer's dextrose, dextrose and sodium chloride, lactated Ringer's or fixed oils.
- Intravenous vehicles include fluid and nutrient replenishers, electrolyte replenishers such as those based on Ringer's dextrose, and the like. Preservatives and other additives may also be present, such as, for example, antimicrobials, antioxidants, chelating agents, inert gases and the like.
- This invention provides a method of inhibiting HIV-1 infection of a CD4+ cell which comprises contacting the CD4+ cell with an amount of the composition of the subject invention effective to inhibit HIV-1 infection of the CD4+ cell so as to thereby inhibit HIV-1 infection of the CD4+ cell .
- the CD4+ cell is present in a subject and the contacting is effected by administering the composition to the subject.
- subject includes any animal or artificially modified animal capable of becoming HIV- infected. Artificially modified animals include, but are not limited to, SCID mice with human immune systems. The animals include but are not limited to mice, rats, dogs, cats, guinea pigs, ferrets, rabbits, and primates. In the preferred embodiment, the subject is a human.
- administering may be effected or performed using any of the methods known to one skilled in the art, which includes intralesional , intraperitoneal, intramuscular, subcutaneous, intravenous, liposome mediated delivery, transmucosal , intestinal, topical, nasal, oral, anal, ocular or otic delivery.
- the compounds may be administered separately (e.g., by different routes of administration, sites of injection, or dosing schedules) so as to combine in synergistically effective amounts in the subject .
- the dose of the composition of the invention will vary depending on the subject and upon the particular route of administration used. Dosages can range from 0.1 to 100,000 ⁇ g/kg. Based upon the composition, the dose can be delivered continuously, such as by continuous pump, or at periodic intervals. For example, on one or more separate occasions. Desired time intervals of multiple doses of a particular composition can be determined without undue experimentation by one skilled in the art.
- effective dose means an amount in sufficient quantities to either treat the subject or prevent the subject from becoming infected with HIV-1.
- a person of ordinary skill in the art can perform simple titration experiments to determine what amount is required to treat the subject.
- the effective amount of the composition comprises from about 0.000001 mg/kg body weight to about 100 mg/kg body weight of the subject.
- This invention provides a method of inhibiting HIV-1 infection of a CD4+ cell which comprises contacting the CD4+ cell with (1) an amount of an antibody which binds to a CCR5 receptor and (2) an amount of a compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate, so as to thereby inhibit HIV-1 infection of the CD4+ cell.
- This invention provides a method of inhibiting HIV-1 infection of a CD4+ cell which comprises contacting the CD4+ cell with (1) an amount of an antibody which binds to a CCR5 receptor, (2) an amount of a compound which retards attachment of HIV-1 to the CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell effective to inhibit HIV-1 infection of the CD4+ cell, and (3) an amount of a compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate, so as to thereby inhibit HIV-1 infection of the CD4+ cell.
- This invention provides a method of inhibiting HIV-1 infection of a CD4+ cell which comprises contacting the CD4+ cell with (1) an amount of a compound which retards attachment of HIV-1 to the CD4+ cell by retarding binding of HIV-1 gpl20 envelope glycoprotein to CD4 on the surface of the CD4+ cell effective to inhibit HIV-1 infection of the CD4+ cell and (2) an amount of a compound which retards gp41 from adopting a conformation capable of mediating fusion of HIV-1 to a CD4+ cell by binding noncovalently to an epitope on a gp41 fusion intermediate so as to thereby inhibit HIV-1 infection of the CD4+ cell.
- the CD4+ cell is present in a subject' and the contacting is effected by administering the compounds to the subj ect .
- the compounds are administered to the subject simultaneously.
- the compounds are administered to the subject at different times.
- the compounds are administered to the subject by different routes of administration.
- the subject invention has various applications which includes HIV treatment such as treating a subject who has become afflicted with HIV.
- afflicted with HIV-1 means that the subject has at least one cell which has been infected by HIV-1.
- treating means either slowing, stopping or reversing the progression of an HIV-1 disorder. In the preferred embodiment, “treating” means reversing the progression to the point of eliminating the disorder.
- treating also means the reduction of the number of viral infections, reduction of the number of infectious viral particles, reduction of the number of virally infected cells, or the amelioration of symptoms associated with HIV-1. Another application of the subject invention is to prevent a subject from contracting HIV.
- HIV-1 infection means the introduction of HIV-1 genetic information into a target cell, such as by fusion of the target cell membrane with HIV-1 or an HIV-1 envelope glycoprotein 4 " cell.
- the target cell may be a bodily cell of a subject. In the preferred embodiment, the target cell is a bodily cell from a human subject.
- inhibit HIV-1 infection means reducing the amount of HIV-1 genetic information introduced into a target cell population as compared to the amount that would be introduced without said composition.
- CD4-IgG2 protein was produced by Progenies Pharmaceuticals, Inc. from plasmids CD4-IgG2-HC- pRcCMV and CD4-kLC-pRcCMV (ATCC Accession Nos. 75193 and 75194, respectively) as described [5] .
- HeLa-env cells were prepared by transfecting HeLa cells (ATCC Catalog # CCL-2) with HIV-1 gpl20/gp41 env-expressing plasmid pMA243 as described [51] .
- PM1 cells are available from the National Institutes of Health AIDS Reagent Program (Catalog #3038) .
- the T-20 peptide was synthesized using standard solid-phase Fmoc chemistry and purified and characterized as described [31,32] .
- F18 -labeled envelope-expressing cells and octadecyl rhodamine (R18) -labeled PM1 cells were plated in 96-well plates in 15% fetal calf serum in phosphate buffered saline and incubated for 4h at 37 (C in the presence of varying concentrations of CD4-IgG2, T-20 or combinations thereof.
- Fluorescence RET was measured with a Cytofluor plate-reader
- HIV-1 inhibition data were analyzed according to the Combination Index method of Chou and Talay [52,53]. The data are modeled according to the median-effect principle, which can be written
- K is equal to the IC 50 value.
- K and m are determined by curve-fitting the dose-response curves.
- Equation (1) is rearranged to allow for calculation of c for a given f .
- the resulting table of values e.g., Figure X
- CI Combination Index
- Equation (2) is used when the molecules are mutually nonexclusive, i.e., have different sites of action. Since this is the likely scenario for inhibitors of HIV-1 attachment and gp41 fusion intermediates, Equation (2) was used for all Combination Index calculations.
- Mutually nonexclusive calculations provide a more conservative estimate of the degree of synergy that mutually exclusive calculations, for which the c lm c 2m /c 1 c 2 term is dropped.
- CI ⁇ 1 indicates synergy
- CI > 1 indicates antagonism. In general, CI values are most relevant at the higher levels of inhibition that are required to achieve a measurable clinical benefit.
- Combinations of inhibitors of HIV-1 attachment and gp41 fusion intermediates were first tested for the ability to inhibit HIV-1 env-mediated membrane fusion in the RET assay.
- This assay has proven to be a highly successful model of the HIV-1 entry process.
- env- dependent coreceptor usage patterns and cellular tropisms of the parental viruses are accurately reproduced [19] .
- the assay was instrumental in demonstrating that CCR5 functions as a requisite fusion coreceptor and acts at the level of viral entry [54] .
- the fusion assay and infectious virus are similarly sensitive to inhibition by metal chelators and agents that target the full complement of viral and cellular receptors [19,46,55] .
- FIG. 1 illustrates representative dose-response curves obtained in the membrane fusion assay for CD4-IgG2, T-20, and combinations of the two. The curve for the combination is highly displaced towards lower inhibitor concentrations and provides qualitative evidence that CD4- IgG2 and T-20 act in a synergistic manner.
- gp41 fusion intermediates are inter-dependent steps.
- attachment inhibitors when used at suboptimal concentrations, may slow but not abrogate the binding of gpl20 to CD4.
- gp41 fusion intermediates may be formed and persist on the virus (or infected cell) for longer periods of time at levels below that required for membrane fusion and thus provide better targets for inhibitory agents.
- HIV-1 entry proceeds via a cascade of at least three sequential events: (1) the attachment of the HIV-1 surface glycoprotein gpl20 to CD4, which is the primary cellular receptor for HIV-1, (2) the interaction of the gpl20-CD4 complex with fusogenic coreceptors such as CCR5 and CXCR4, and (3) membrane fusion mediated by the HIV-1 transmembrane glycoprotein gp41.
- PRO 542 CD4-IgG2
- PRO 140 (PA14) and PA12 are monoclonal antibodies to CCR5 that block its function as an HIV-1 coreceptor.
- T-20 is a 36-mer peptide derived from the highly conserved C- terminal ectodomain of gp41. T-20 blocks gp41-mediated membrane fusion events.
- PRO 542 is thus an attachment inhibitor that blocks the first step of HIV-1 entry;
- PRO 140 and PA12 are both CCR5 coreceptor inhibitors that block the second step; and
- T-20 is a fusion inhibitor that blocks the third step.
- Attachment, coreceptor and fusion inhibitors are all members of a broad category of antiviral agents collectively know as HIV-1 entry inhibitors.
- CCR5 coreceptor inhibitors and CXCR4 coreceptor inhibitors reconstitute two distinct subclasses of coreceptor inhibitors .
- PRO 542 and T-20 When used individually, each of these compounds inhibit HIV-1 infection in vitro.
- PRO 542 and T-20 have also both demonstrated significant antiviral activity when used individually in human clinical trials, providing clinical proof-of-concept for inhibitors of HIV-1 entry (1,2).
- Trkola A. Pomales AP., Yuan H. et al . Cross-clade neutralization of primary isolates of human immunodeficiency virus type 1 by human monoclonal antibodies and tetrameric CD4-IgG2. Journal of Virology 1995; 69:6609-6617.
- Trkola A Ketas T, KewalRamani VN et al . Neutralization sensitivity of human immunodeficiency virus type 1 primary isolates to antibodies and CD4- based reagents is independent of coreceptor usage. J Virol 1998; 72:1876-1885.
- Burkly L, Mulrey N, Blumenthal R, Dimitrov DS Synergistic inhibition of human immunodeficiency virus type 1 envelope glycoprotein-mediated cell fusion and infection by an antibody to CD4 domain 2 in combination with anti-gpl20 antibodies. Journal of Virology 1995; 69:4267-4273.
- HIV-1 tropism for ononuclear phagocytes can be determined by regions of gpl20 outside the CD4 -binding domain.
- Trkola A Matthews J, Gordon C, Ketas T, Moore JP .
- Chou TC The median effect principle and the combination index for quantitation of synergism and antagonism. Synergism and Antagonism in Chemotherapy 1991; 61-102. 53. Chou TC, Talalay P. Quantitative analysis of dose- effect relationships: the combined effects of multiple drugs or enzyme inhibitors . Advances in Enzyme Regulation 1984; 22:27-55.
- HIV-1 entry into target cells proceeds at the cell surface through a cascade of events that include (1) binding of the viral surface glycoprotein gpl20 to cell surface CD4 , which is the primary receptor for HIV-1, (2) env binding to fusion coreceptors such as CCR5 and CXCR4 , and (3) multiple conformational changes in gp41.
- gp41 adopts transient conformations that include a prehairpin fusion intermediate that ultimately folds into a conformation capable of mediating fusion. These events culminate in fusion of the viral and cellular membranes and the subsequent introduction of the viral genome into the target cell. A similar sequence of molecular events is required for infection to spread via fusion of infected and uninfected cells. Each stage of the viral entry process can be targeted for therapeutic intervention .
- HIV-1 attachment can be inhibited both by agents that bind the viral envelope glycoproteins and by agents that bind human CD4.
- HIV-1 attachment can be inhibited by compounds that incorporate the gpl20 -binding domains of human CD4 and molecular mimics thereof [1-7] . Because this interaction between gpl20 and CD4 is essential for virus infection, CD4 -based molecules have the potential to target most if not all strains of HIV-1. In addition, viruses have limited ability to develop resistance to such molecules .
- CD4-based molecules A number of recombinant CD4-based molecules have been developed and tested for clinical activity in man. The first of these contained the four extracellular domains (D1-D4) of CD4 but lacked the transmembrane and intracellular regions. This molecule, termed soluble CD4 (sCD4) , demonstrated excellent tolerability when administered to humans at doses ranging to 10 mg/kg [8,9] . Transient reductions in plasma levels of infectious HIV-1 were observed in certain patients treated with sCD4. The short half-life of sCD4 in humans (45 minutes following intravenous administration) was identified as one obstacle to using this agent for chronic therapy.
- sCD4 extracellular domains
- CD4-based proteins were developed with increased serum half-life. These CD4-immunoglobulin fusion proteins comprised the D1D2 domains of CD4 genetically fused to the hinge CH2 and CH3 regions of human IgG olecules . These divalent proteins derive HIV-1 neutralizing capacity from their CD4 domains and Fc effector functions from the IgG molecule.
- a CD4-IgGl fusion protein was shown to have excellent tolerability and improved pharmacokinetics in Phase I clinical testing [10] . The antiviral evaluations were inconclusive.
- CD4-IgG2 fusion protein comprises the D1D2 domains of CD4 genetically fused to the heavy and light chain constant regions of human IgG2. This agent binds the
- HIV-1 envelope glycoprotein gpl20 HIV-1 envelope glycoprotein gpl20 with nanomolar affinity
- CD4-IgG2 neutralized primary HIV-1 isolates derived from all major subtypes and outlier groups.
- CD4-IgG2 was similarly effective in neutralizing HIV-1 obtained directly from the plasma of seropositive donors in an ex vivo assay, indicating that this agent is active against the diverse viral quasispecies that are encountered clinically [12] .
- CD4-IgG2 also provided protection against infection by primary isolates in the hu-PBL-SCID mouse model of HIV-1 infection [13] .
- Recent analyses have demonstrated that CD4- IgG2 ' s ability to neutralize primary viruses is independent of their coreceptor usage [14] .
- CD4- IgG2 Compared with mono- or divalent CD4-based proteins, CD4- IgG2 has consistently demonstrated as much as 100 -fold greater potency at inhibiting primary HIV-1 isolates [5,12,14,15]. The heightened potency may derive from CD4- IgG2 ' s ability to bind virions with increased valency/avidity and its steric juxtaposition of two gpl20 binding sites on each Fab-like arm of the immunoglobulin molecule. The larger Fab-like arms of CD4-IgG2 are also more likely to span HIV-1 envelope spikes on the virion.
- CD4-IgG2's anti-HIV-1 activity has been shown to compare favorably with those of the rare human monoclonal antibodies that broadly and potently neutralize primary HIV-1 isolates [5,11,14,15] .
- CD4-IgG2 therapy is in principle less susceptible to the development of drug-resistant viruses than therapies employing anti-env monoclonal antibodies or portions of the highly mutable HIV-1 envelope glycoproteins.
- CD4-IgG2 may have clinical utility as an agent that neutralizes cell-free virus before it has the opportunity to establish new rounds of infection.
- CD4-IgG2 may have utility in preventing infection resulting from occupational, perinatal or other exposure to HIV-1.
- CD4-IgG2 demonstrated excellent pharmacology and tolerability. In addition, measurable antiviral activity was observed by each of two measures. First, a statistically significant acute reduction in plasma HIV RNA was observed following administration of a single 10 mg/kg dose. In addition, sustained reductions in plasma levels of infectious HIV were observed in each of two patients tested. Taken together, these observations indicate that CD4-IgG2 possesses antiviral activity in humans [16] .
- HIV-1 attachment can also be inhibited by antibodies and nonpeptidyl molecules.
- Known inhibitors include (1) anti-env antibodies such as IgGlbl2 and F105
- anti-CD4 antibodies such as OKT4A, Leu 3a, and humanized versions thereof [19,20]
- nonpeptidyl agents that target either gpl20 or CD4 [21] , [22-24] .
- the latter group of compounds includes aurintricarboxylic acids, polyhydroxycarboxylates, sulfonic acid polymers, and dextran sulfates.
- agents that block HIV-1 infection by targeting gp41 fusion intermediates. These inhibitors may interact with the fusion intermediates and prevent them from folding into final fusogenic conformations.
- the first such agents to be identified comprised synthetic or recombinant peptides corresponding to portions of the gp41 ectodomain predicted to form hydrophobic alpha helices. One such region is present in both the amino and carboxy segments of the extracellular portion of gp41, and recent crystallographic evidence suggests that these regions ⁇ interact in the presumed fusogenic conformation of gp41 [25,26] .
- HIV-1 infection can be inhibited by agents that bind to either N- or C-terminal gp41 epitopes that are exposed during fusion.
- T-20 (formerly known as DP178)
- T-1249 T-1249
- DP107 DP107
- N34, C28 various fusion proteins and analogues thereof
- Other studies have identified inhibitors that comprise non-natural D-peptides and nonpeptidyl moieties [34,35] .
- Clinical proof-of-concept for this class of inhibitors has been provided by T-20, which reduced plasma HIV RNA levels by as much as 2 logs in Phase I/II human clinical testing [36] .
- the broad antiviral activity demonstrated for this class of inhibitors reflects the high degree of gp41 sequence conservation amongst diverse strains of HIV-1.
- CD4-IgG2 protein Purified recombinant CD4-IgG2 protein was produced by Progenies Pharmaceuticals, Inc. from plasmids CD4-IgG2-HC- pRcCMV and CD4-kLC-pRcCMV (ATCC Accession Nos. 75193 and 75194, respectively) as described [5].
- HeLa-env cells were prepared by transfecting HeLa cells (ATCC Catalog # CCL-2) with HIV-1 gpl20/gp41 env-expressing plasmid pMA243 as described [51] .
- PM1 cells are available from the National Institutes of Health AIDS Reagent Program (Catalog #3038) .
- the T-20 peptide was synthesized using standard solid-phase Fmoc chemistry and purified and characterized as described [31,32] .
- HIV-1 inhibition data were analyzed according to the Combination Index method of Chou and Talay [52,53]. The data are modeled according to the median-effect principle, which can be written
- K is equal to the IC 50 value. K and are determined by curve-fitting the dose-response curves.
- Equation (1) is rearranged to allow for calculation of c for a given f .
- the resulting table of values e.g., Figure X
- CI Combination Index
- Equation (2) is used when the molecules are mutually nonexclusive, i.e., have different sites of action. Since this is the likely scenario for inhibitors of HIV-1 attachment and gp41 fusion intermediates, Equation (2) was used for all Combination Index calculations.
- Mutually nonexclusive calculations provide a more conservative estimate of the degree of synergy that mutually exclusive calculations, for which the e lm c 2m /c 1 c 2 term is dropped.
- CI ⁇ 1 indicates synergy
- CI > 1 indicates antagonism. In general, CI values are most relevant at the higher levels of inhibition that are required to achieve a measurable clinical benefit .
- Combinations of inhibitors of HIV-1 attachment and gp41 fusion intermediates were first tested for the ability to inhibit HIV-1 env-mediated membrane fusion in the RET assay.
- This assay has proven to be a highly successful model of the HIV-1 entry process.
- env- dependent coreceptor usage patterns and cellular tropisms of the parental viruses are accurately reproduced [19] .
- the assay was instrumental in demonstrating that CCR5 functions as a requisite fusion coreceptor and acts at the level of viral entry [54] .
- the fusion assay and infectious virus are similarly sensitive to inhibition by metal chelators and agents that target the full complement of viral and cellular receptors [19,46,55].
- FIG. 6 illustrates representative dose-response curves obtained in the membrane fusion assay for CD4-IgG2, T-20, and combinations of the two. The curve for the combination is highly displaced towards lower inhibitor concentrations and provides qualitative evidence that CD4- IgG2 and T-20 act in a synergistic manner.
- Trkola A. Pomales AP., . Yuan H. et al . Cross-clade neutralization of primary isolates of human immunodeficiency virus type 1 by human monoclonal antibodies and tetrameric CD4-IgG2. Journal of
- Trkola A Ketas T, KewalRamani VN et al . Neutralization sensitivity of human immunodeficiency virus type 1 primary isolates to antibodies and CD4- based reagents is independent of coreceptor usage. J Virol 1998; 72:1876-1885.
- Burkly L, Mulrey N, Blumenthal R, Di itrov DS Synergistic inhibition of human immunodeficiency virus type 1 envelope glycoprotein-mediated cell fusion and infection by an antibody to CD4 domain 2 in combination with anti-gpl20 antibodies. Journal of Virology 1995; 69:4267-4273.
- HIV-1 tropism for mononuclear phagocytes can be determined by regions of gpl20 outside the CD4-binding domain. Nature 1990; 348:69-73.
- Trkola A Matthews J, Gordon C, Ketas T, Moore JP.
- HIV-1 entry proceeds via a cascade of events that afford promising targets for therapy.
- PRO 542 neutralizes HIV-1 by blocking its attachment to CD4, and T-20 blocks gp41- ediated fusion.
- Both drugs have shown promise in Phase I/II clinical trials.
- the drugs were tested individually and in combination in preclinical models of HIV-1 infection, and inhibition data were analyzed for cooperativity using the Combination Index method.
- Synergistic inhibition of virus-cell and cell-cell fusion was observed for phenotypically diverse viruses over a broad range of drug concentrations, often resulting in 10- fold or greater dose reductions in vitro. Additional mechanism-of-action studies probed the molecular basis of the synergies.
- the markedly enhanced activity observed for the PRO 542-.T-20 combination indicates that the multi-step nature of HIV-1 entry leaves the virus particularly vulnerable to combinations of entry inhibitors.
- HIV-1 entry comprises three steps that provide promising targets for therapy: (1) the attachment of the HIV-1 surface glycoprotein gpl20 to CD4 , (2) the interaction of the gpl20-CD4 complex with a coreceptor, and (3) membrane fusion mediated by the HIV-1 transmembrane glycoprotein gp41. Attachment, coreceptor, and fusion inhibitors are broadly referred to as entry inhibitors and are currently in clinical development [1-4] .
- the attachment inhibitor PRO 542 (CD4-IgG2) is a tetravalent CD4 -immunoglobulin fusion protein that potently neutralizes primary HIV-1 isolates [5] .
- PRO 542 has demonstrated excellent tolerability and pharmacology and has reduced viral loads as measured by plasma HIV RNA, plasma viremia, and cell-associated virus [1,2].
- PRO 542 is currently in Phase II clinical testing.
- the fusion inhibitor T-20 is a peptide derived from the C- ter inal ectodomain of gp41.
- T-20 possesses broad-spectrum antiviral activity mediated by its ability to act upon transient gp41 fusion intermediates [6,7], Although the precise structures of the fusion intermediates are not known, they are known as "pre-hairpin" intermediates based on the post-fusion conformation of gp41.
- T-20 mediated 1-2 log 10 reductions in HIV RNA when used as a single agent in Phase I/II trials and has been associated with prolonged viral suppression when used with FDA-approved antiretrovirals [3,4] .
- T-20 has entered Phase III clinical testing .
- PRO 542 and T-20 are attractive candidates for combination use because they are well -tolerated, "first-in-class" agents that target distinct stages of HIV-1 entry. Moreover, the multi-step, interdependent nature of HIV-1 entry suggests that these drugs might act in a non-additive manner. In this study, we evaluated these agents individually and in combination in vitro and analyzed the results for cooperative effects.
- the PRO 542.-T-20 combination potently and synergistically inhibited entry of phenotypically diverse HIV-1 isolates over a broad range of experimental conditions. Additional studies shed light on the molecular basis for the enhanced antiviral activity.
- Compounds PRO 542 was expressed in Chinese hamster ovary cells and purified by column chromatography by Progenies
- T-20 was synthesized by solid-phase fluroenylmethoxycarbonyl chemistry, purified by reversed phase chromatography and analyzed for purity and size by
- PRO 542 and T-20 were formulated in phosphate buffered saline and stored at -95°C
- HIV-1 virus-cell fusion assays NLluc + env " viruses were complemented in trans by the envelope glycoproteins (Env) from HIV-IJ R . FL (R5) or HIV-1 DH123 (R5X4) [8] .
- HeLa-CD4 + CCR5 + cells were infected with chimeric reporter viruses containing 50-100 ng/ l of p24 in the presence or absence of drugs. Cells were washed and fed fresh drug-containing media at 2 h and drug-free media at 12 h. After 72 h, cell lysates were assayed for luciferase activity [9] .
- HIV-1 cell-cell fusion assays Cell-cell fusion mediated by the envelope glycoproteins of HIV-1 JR - PL and HIV-l ⁇ was measured using a fluorescence resonance energy transfer (RET) assay [10] , which measures fusion between HeLa cells expressing HIV-1 envelope glycoproteins and CD4 + target cells .
- RET fluorescence resonance energy transfer
- the T cell lines PM1 and SupTl were used as target cells in this study.
- PM1 endogenously express CCR5 and CXCR4.
- Cells are membrane-labeled with fluorescent dyes that possess overlapping excitation and emission spectra, such that RET occurs when the dyes are placed in the same membrane following fusion.
- the RET signal is directly related to the extent of membrane fusion.
- HIV-1 virus-cell fusion PRO 542, T-20 and 1 : 2 molar combinations thereof were used to inhibit HIV-l ⁇ and HIV-1 DH123 entry.
- concentration required for 90% inhibition (IC 90 ) was 12 nM for PRO 542, 45 nM for T-20 and 4.1 nM for the combination (1.5 nM PRO 542 and 2.6 nM T-20).
- IC 90 concentration required for 90% inhibition
- the concentration required for 90% inhibition was 12 nM for PRO 542, 45 nM for T-20 and 4.1 nM for the combination (1.5 nM PRO 542 and 2.6 nM T-20).
- Favorable drug-drug interactions are evident from the fact that the IC 90 of the combination is substantially lower than that of either compound alone.
- PRO 542 and T-20 also synergistically inhibited HIV- 1 ⁇ (X4) Env-mediated fusion.
- CI values ranged from 0.45-0.28 at 50-95% inhibition.
- the dose reductions ranged from 12-fold to 30-fold for PRO 542 and from 2.5-fold to 5.4-fold for T-20. Synergy was also observed when the SupTl T cell line was used instead of PM1 (data not shown) .
- PRO 542.T-20 synergy T-20 does not interact with Env in its pre-fusion conformation but rather targets transient gp41 structures [7].
- PRO 542 neutralizes free virus and triggers Env conformational changes that are required for entry [12] .
- PRO 542 could conceivably trigger formation of gp41 fusion intermediates, enabling T-20 to act on free virus.
- PRO 542 and T-20 were pre- incubated alone and in combination for 1 h at 37 °C with HeLa-Env cells prior to washing and the addition of PM1 cells ( Figure 9c) . Whereas PRO 542 demonstrated 6-fold lower but nonetheless significant potency, T-20 was essentially inactive in this setting.
- HIV-1 entry is a promising target for a new generation of antiretroviral therapies, and PRO 542 and T-20 are "first- in-class" investigational agents that block distinct stages of this highly cooperative process. Recognizing that combinations of entry inhibitors may emerge as a new paradigm for HIV-1 therapy, this study explored PRO 542 :T- 20 combinations in two preclinical models of HIV-1 infection. Potent synergy was observed for both virus-cell and cell -cell fusion, for phenotypically diverse virus isolates, and over a broad range of drug concentrations. In many instances, the synergy translated into 10 -fold or greater dose reductions for PRO 542 and T-20 in vitro. Thus the multi-step nature of HIV-1 entry appears to leave the virus particularly vulnerable to cooperative inhibition. The findings provide a strong rationale for combination clinical trials designed to maximize the therapeutic potential of each compound.
- PRO 542 but not T-20 neutralizes cell-free virus
- T-20 is particularly effective in blocking cell-cell fusion [6-8] .
- PRO 542:T-20 combinations may prove to be particularly effective in vivo.
- PRO 542 and T-20 synergistically inhibited fusion mediated by R5 , X4 and R5X4 virus envelopes While the activity of PRO 542 against primary viruses is independent of their coreceptor usage [8], T-20's ability to block virus-cell fusion reportedly varies according coreceptor usage [14] . Additional studies with a broader panel of viruses are needed to determine if PRO 542:T-20 synergy is subtly affected by coreceptor usage.
- PRO 542 alone was not able to trigger formation of the gp41 pre-hairpin intermediate, which formed only when Env, CD4 and coreceptor interacted at a fusion-permissive temperature. Thus synergy may derive from subtler events, such as the clustering of multiple gpl20/gp41 trimers to form the fusion pore [13] .
- PRO 542 may block a subset of Envs from interacting with target cells, while perhaps allowing other Envs to proceed to the pre-hairpin intermediate.
- the fusion intermediates may remain exposed and susceptible to T-20 for significantly longer periods of time, as required to recruit a critical number of free Envs to the site of the fusion pore. In a recent study, T-20 mediated 100% inhibition when added within the first 15 minutes of cell mixing but was essentially inactive ( ⁇ 10% inhibition) at 75 minutes [15] . Ongoing studies are examining the effects of PRO 542 during this time period.
- PRO 542 and T-20 were potently synergistic when used to inhibit HIV-1 entry in vitro. Cooperative inhibition of virus-cell and cell-cell fusion was observed for phenotypically diverse viruses and translated into meaningful dose reductions. These findings provide a strong rationale for evaluating combinations of these agents in vivo.
- HIV-1 entry proceeds via a cascade of events that provide promising targets for a new generation of antiviral therapies, including the gpl20-CD4 attachment inhibitor PR0542, the gpl20-coreceptor inhibitor PR0140, and the gp41 fusion inhibitors T-20 and T-1249.
- the multi-step nature of HIV-1 entry leaves the virus highly susceptible to inhibition by combinations of drugs that act at distinct stages of this process. It has been shown that double and triple drug cocktails of attachment, coreceptor and the fusion inhibitors potently and synergistically block HIV-1 entry over a wide range of experimental conditions in vitro.
- HIV-1 membrane fusion was monitored in real time using a semi-automatic fluorometric assay, and the inhibitory activities of the individual drugs and drug cocktails were evaluated in ti e-of-addition, washout, and stepwise temperature-controlled studies.
- the data were consistent with a model wherein the drugs act cooperatively to delay the recruitment of a critical number of fusion-active HIV-1 envelope glycoproteins to the site of the fusion pore.
- HIV-1 entry involves at least three steps that provide targets for therapy: (1) gpl20-CD4 attachment; (2) gpl20-coreceptor interactions; and (3) gp41 -mediated fusion.
- the CD4-IgG2 attachment inhibitor (PRO 542) , as shown in Figure 12. Is a tetravalent CD4-based protein. It broadly and potently neutralizes primary HIV isolates in vitro, ex vivo and in vivo, and acts prior to fusion. PR0542 has completed two phase I/II clinical trials, showing (1) excellent safety and phar acokinetic profiles and that it is non-immunogenic; (2) statistically significant reductions in viral load with a single dose; and (3) sustained reductions in free and cell-associated infectious virus. PRO 542 is in Phase II testing.
- the PRO 140 coreceptor inhibitor is a monoclonal antibody to CCR5. It potently neutralizes CCR5 -using virus. It protects primary T cells and macrophages, and its inhibition is genetic subtype- independent. It selectively blocks HIV entry without affecting chemokine-induced signaling. There has been no emergence of HIV resistance following 31 weeks in vitro culture .
- the T-20 fusion inhibitor as shown in 14, is a peptide derived from HIV-1 gp41. It broadly and potently blocks HIV-1 cell-cell and cell-virus fusion in vitro. It has completed Phase I/II clinical trials, showing (1) good tolerability; (2) pK supports twice daily subcutaneous dosing; and (3) clinically significant dose-dependent reductions in viral load.
- Combination therapy may be required to control HIV in vivo. HIV entry inhibitors may represent the next major advance in therapy. It is critical to determine drug-drug interactions earlier rather than later in clinical development to avoid antagonistic combinations and identify synergistic combinations. The clinical benefits of synergistic combinations include more potent antiviral effects and more durable antiviral effects.
- HIV-1 virus-cell fusion assay is described in Figure 15.
- Env-Luc+ HIV reporter viruses were prepared as described [1] .
- the viruses are complemented with gpl20/gp41 from an HIV strain of interest and thus are capable of fusing with target cells that express CD4 and fusion coreceptors, resulting in the introduction of the viral genes into the cell.
- the viruses encode the luciferase reporter enzyme rather than HIVenv, infection leads to the production of luciferase rather than new viral particles.
- the amount of luciferase is measured 72 hours post-infection using standard methods.
- Figures 16 and 17 demonstrate the synergistic inhibition of virus-cell fusion with PR0542 and T-20.
- FIG. 18 An HIV-1 cell-cell fusion assay is described in Figure 18. Such assay is a Resonance Energy Transfer (RET) assay of HIV-1 membrane fusion.
- RET Resonance Energy Transfer
- Cells expressing gpl20/gp41 are labeled with the fluorescent dye octadecyl fluorescsin (F18)
- T cell that express CD4 and fusion coreceptors CD4 cells
- CD4 cells octadecyl rhodamine
- the emission spectrum of F18 overlaps with the excitation spectrum of R18, allowing fluorescence RET to occur when the dyes are placed in the same membrane following fusion.
- RET is measured by stimulating F18 at 450nm and measuring the R18 emission at 590nm.
- the RET signal is directly related to the amount of HIV-1 fusion [2] .
- Figures 19 and 20 demonstrate the synergistic inhibition of cell-cell fusion with PR0542 and T-20.
- Figures 21 and 22 demonstrates that the PRO140, PR0542 and T-20 triple combination synergistically blocks HIV-1 entry.
- Figure 23 demonstrates that PR0542 does not potentiate T-20 activity in the absence of coreceptor.
- PR0542, T-20 or a 1:2 mixture were pre-incubated with HeLa-ENVJR-FL cells for 2 hours at 37°C prior to washing, addition of PM1 target cells, and completion of the fusion assay. Formation of the prehairpin intermediates may need CD4 , coreceptor and 37°C, as demonstrated in Figures 24 and 25.
- Figures 26 and 27 describe a possible mechanism of synergy, wherein PR0542 increases the half-life of the T-20 targets.
- Figure 28 demonstrates synergistic inhibition of HIV-1 entry using PR0542, T-1249 and in a 1:10 molar combination in an env-medicated membrane fusion (RET) assay.
- Figure 29 demonstrates the combination indices and dose reductions for a 1:10 molar combination of PR0542 and T-1249.
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WO2006103100A2 (en) | 2005-04-01 | 2006-10-05 | F. Hoffmann-La Roche Ag | Antibodies against ccr5 and uses thereof |
US7122185B2 (en) | 2002-02-22 | 2006-10-17 | Progenics Pharmaceuticals, Inc. | Anti-CCR5 antibody |
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Families Citing this family (19)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CA2224003C (en) * | 1995-06-07 | 2010-04-13 | Progenics Pharmaceuticals, Inc. | Fluorescence resonance energy transfer screening assay for the identification of hiv-1 envelope glycoprotein-medicated cell |
US7118859B2 (en) | 1996-01-17 | 2006-10-10 | Progenics Pharmaceuticals, Inc. | Methods for inhibiting HIV-1 infection |
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Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6107019A (en) * | 1996-06-14 | 2000-08-22 | Progenics Pharmaceuticals, Inc. | Method for preventing HIV-1 infection of CD4+ cells |
US6344545B1 (en) * | 1996-06-14 | 2002-02-05 | Progenics Pharmaceuticals, Inc. | Method for preventing HIV-1 infection of CD4+ cells |
Family Cites Families (59)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US4886743A (en) | 1985-04-24 | 1989-12-12 | California Institute Of Technology | Diagnostic reagents based on unique sequences within the variable region of the T cell receptor and uses thereof |
US5225539A (en) * | 1986-03-27 | 1993-07-06 | Medical Research Council | Recombinant altered antibodies and methods of making altered antibodies |
US5126433A (en) | 1986-08-21 | 1992-06-30 | The Trustees Of Columbia University In The City Of New York | Soluble forms of the t cell surface protein cd4 |
US5091513A (en) | 1987-05-21 | 1992-02-25 | Creative Biomolecules, Inc. | Biosynthetic antibody binding sites |
US5854400A (en) | 1987-05-29 | 1998-12-29 | Tanox, Inc. | Monoclonal antibodies which neutralize HIV-1 infection |
US5215913A (en) | 1987-11-30 | 1993-06-01 | Roger Williams General Hospital | IgG-1 human monoclonal antibody reactive with an HIV-1 antigen and methods of use |
US5071964A (en) | 1988-05-04 | 1991-12-10 | Dana-Farber Cancer Institute, Inc. | Protein micelles |
US5021409A (en) | 1989-12-21 | 1991-06-04 | Johnson Matthey Plc | Antiviral cyclic polyamines |
US5668149A (en) * | 1990-01-26 | 1997-09-16 | The United States Of America As Represented By The Department Of Health And Human Services | Inhibition of human immunodeficiency virus-1 infectivity in human cells |
WO1992001451A1 (en) | 1990-06-12 | 1992-02-06 | Board Of Regents, The University Of Texas System | Method for killing hiv-infected cells |
US5440021A (en) | 1991-03-29 | 1995-08-08 | Chuntharapai; Anan | Antibodies to human IL-8 type B receptor |
US5817767A (en) * | 1993-02-24 | 1998-10-06 | Progenics Pharmaceuticals, Inc. | Synergistic composition of CD4-based protein and anti-HIV-1 antibody, and methods of using same |
US5449608A (en) | 1993-03-22 | 1995-09-12 | The United States Of America As Represented By The Department Of Health And Human Services | Parvovirus B19 receptor and parvovirus B19 detection |
US5464933A (en) | 1993-06-07 | 1995-11-07 | Duke University | Synthetic peptide inhibitors of HIV transmission |
US5464963A (en) * | 1993-08-27 | 1995-11-07 | Motoman Inc. | Sealing arrangement for a laser enclosure |
US5603933A (en) * | 1993-08-31 | 1997-02-18 | Board Of Regents, The University Of Texas | CD4 peptides for binding to viral envelope proteins |
US5504003A (en) | 1994-03-08 | 1996-04-02 | Human Genome Sciences, Inc. | Macrophage inflammatory protein-3 and -4 |
FR2727221B1 (en) | 1994-11-23 | 1996-12-27 | Kodak Pathe | METHOD FOR COMMUNICATING DIGITAL INFORMATION AND SYSTEM FOR IMPLEMENTING THE METHOD |
EP0815137A4 (en) | 1995-06-06 | 1998-08-05 | Human Genome Sciences Inc | Human g-protein chemokine receptor hdgnr10 |
MX9709849A (en) | 1995-06-06 | 1998-03-29 | Human Genome Sciences Inc | Human g-protein chemokine receptor hdgnr10 |
EP1145721A3 (en) | 1995-06-06 | 2003-12-03 | Human Genome Sciences, Inc. | Human G-protein chemokine receptor HDGNR10 (CCR5 receptor). Pharmaceutical composition |
US6025154A (en) | 1995-06-06 | 2000-02-15 | Human Genome Sciences, Inc. | Polynucleotides encoding human G-protein chemokine receptor HDGNR10 |
US20040151719A1 (en) | 1995-06-06 | 2004-08-05 | Human Genome Sciences, Inc. | Human G-protein chemokine receptor (CCR5) HDGNR10 |
US6743594B1 (en) | 1995-06-06 | 2004-06-01 | Human Genome Sciences, Inc. | Methods of screening using human G-protein chemokine receptor HDGNR10 (CCR5) |
CA2224003C (en) | 1995-06-07 | 2010-04-13 | Progenics Pharmaceuticals, Inc. | Fluorescence resonance energy transfer screening assay for the identification of hiv-1 envelope glycoprotein-medicated cell |
US6265184B1 (en) | 1995-12-20 | 2001-07-24 | Icos Corporation | Polynucleotides encoding chemokine receptor 88C |
WO1997026009A1 (en) | 1996-01-17 | 1997-07-24 | Progenics Pharmaceuticals, Inc. | Compounds capable of inhibiting hiv-1 infection |
US7118859B2 (en) | 1996-01-17 | 2006-10-10 | Progenics Pharmaceuticals, Inc. | Methods for inhibiting HIV-1 infection |
ES2227672T3 (en) | 1996-03-01 | 2005-04-01 | Euroscreen S.A. | C-C CKR-5, CC CHEMIOKIN RECEIVER, DERIVED FROM THE SAME AND ITS USES. |
AU728512B2 (en) | 1996-04-02 | 2001-01-11 | Progenics Pharmaceuticals, Inc. | Method for preventing HIV-1 infection of CD4+ cells |
US5939320A (en) | 1996-05-20 | 1999-08-17 | New York University | G-coupled receptors associated with macrophage-trophic HIV, and diagnostic and therapeutic uses thereof |
US6258527B1 (en) | 1996-05-20 | 2001-07-10 | The Aaron Diamond Aids Research Center | Methods of identifying g-coupled receptors associated with macrophage-trophic HIV, and diagnostic and therapeutic uses thereof |
WO1997045543A2 (en) | 1996-05-28 | 1997-12-04 | The Government Of The United States Of America, As Represented By The Secretary Of Health And Human Services, National Institutes Of Health | Cc chemokine receptor 5, antibodies thereto, transgenic animals |
US20040086528A1 (en) | 1996-06-14 | 2004-05-06 | Progenics Pharmaceuticals, Inc. | Uses of a chemokine receptor for inhibiting HIV-1 infection |
WO1997047319A1 (en) | 1996-06-14 | 1997-12-18 | Progenics Pharmaceuticals, Inc. | Uses of a chemokine receptor for inhibiting hiv-1 infection |
WO1997049424A1 (en) | 1996-06-25 | 1997-12-31 | The Trustees Of The University Of Pennsylvania | Antibodies directed against cellular coreceptors for human immunodeficiency virus and methods of using the same |
US5994515A (en) * | 1996-06-25 | 1999-11-30 | Trustees Of The University Of Pennsylvania | Antibodies directed against cellular coreceptors for human immunodeficiency virus and methods of using the same |
US6528625B1 (en) * | 1996-10-28 | 2003-03-04 | Millennium Pharmaceuticals, Inc. | Anti-CCR5 antibodies and kits comprising same |
US6100087A (en) | 1998-03-11 | 2000-08-08 | City Of Hope | Ribozymes targeted to human CCR5 mRNA |
US6258782B1 (en) * | 1998-05-20 | 2001-07-10 | Trimeris, Inc. | Hybrid polypeptides with enhanced pharmacokinetic properties |
US20040228869A1 (en) | 1998-12-16 | 2004-11-18 | Progenics Pharmaceuticals, Inc. | Synergistic inhibition of HIV-1 fusion and attachment, compositions and antibodies thereto |
EP1144006B1 (en) | 1998-12-16 | 2007-10-17 | Progenics Pharmaceuticals, Inc. | CCR5 Antibody PA14 |
US6692745B2 (en) | 2000-01-28 | 2004-02-17 | Arogenics Pharmaceuticals, Inc. | Compositions and methods for inhibition of HIV-1 infection |
KR20030032916A (en) | 2000-02-09 | 2003-04-26 | 휴먼 게놈 사이언시즈, 인크. | Antibodies to ccr5 |
AU2001247254A1 (en) | 2000-02-29 | 2001-09-12 | Aaron Diamond Aids Research Centre | Sulfated CCR5 peptides for HIV -1 infection |
US7138119B2 (en) | 2000-09-15 | 2006-11-21 | Progenics Pharmaceuticals, Inc. | Compositions and methods for inhibition of HIV-1 infection |
US7175988B2 (en) | 2001-02-09 | 2007-02-13 | Human Genome Sciences, Inc. | Human G-protein Chemokine Receptor (CCR5) HDGNR10 |
WO2002068608A2 (en) | 2001-02-28 | 2002-09-06 | Progenics Pharmaceuticals, Inc. | Sulfated ccr5 peptides for hiv-1 infection |
US20030003440A1 (en) | 2001-03-14 | 2003-01-02 | Lucia Lopalco | Novel CCR5 epitope and antibodies against it |
US7060273B2 (en) | 2001-04-06 | 2006-06-13 | Progenics Pharmaceuticals, Inc. | Methods for inhibiting HIV-1 infection |
US20020146415A1 (en) * | 2001-04-06 | 2002-10-10 | Olson William C. | Methods for inhibiting HIV-1 infection |
US6825210B2 (en) | 2001-04-19 | 2004-11-30 | Bristol-Myers Squibb Pharma Company | Tricyclic compounds useful as HIV reverse transcriptase inhibitors |
US7393934B2 (en) | 2001-12-21 | 2008-07-01 | Human Genome Sciences, Inc. | Human G-protein chemokine receptor (CCR5) HDGNR10 |
US7122185B2 (en) * | 2002-02-22 | 2006-10-17 | Progenics Pharmaceuticals, Inc. | Anti-CCR5 antibody |
AU2003217674C1 (en) | 2002-02-22 | 2009-03-26 | Abbvie Biotherapeutics Inc. | Anti-CCR5 antibody |
CN1805740A (en) | 2003-05-16 | 2006-07-19 | 马里兰大学生物技术研究所 | Compositions for down-regulation of CCR5 expression and methods of use thereof |
US20050131042A1 (en) | 2003-12-11 | 2005-06-16 | Flentge Charles A. | HIV protease inhibiting compounds |
EP1723178A4 (en) | 2004-03-12 | 2007-12-12 | Human Genome Sciences Inc | Human g-protein chemokine receptor (ccr5) hdgnr10 |
MX2008000984A (en) | 2005-07-22 | 2008-04-04 | Progenics Pharm Inc | Methods for reducing viral load in hiv-1-infected patients. |
-
2001
- 2001-07-25 US US09/912,824 patent/US7138119B2/en not_active Expired - Lifetime
- 2001-09-14 WO PCT/US2001/028756 patent/WO2002022077A2/en active Application Filing
- 2001-09-14 AU AU9092501A patent/AU9092501A/en active Pending
- 2001-09-14 AU AU2001290925A patent/AU2001290925C1/en not_active Expired
- 2001-09-14 EP EP01970984A patent/EP1322332A4/en not_active Withdrawn
- 2001-09-14 CA CA002422275A patent/CA2422275A1/en not_active Abandoned
- 2001-09-14 JP JP2002526332A patent/JP2004518624A/en not_active Withdrawn
-
2006
- 2006-09-12 US US11/520,556 patent/US7736649B2/en not_active Expired - Fee Related
Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6107019A (en) * | 1996-06-14 | 2000-08-22 | Progenics Pharmaceuticals, Inc. | Method for preventing HIV-1 infection of CD4+ cells |
US6344545B1 (en) * | 1996-06-14 | 2002-02-05 | Progenics Pharmaceuticals, Inc. | Method for preventing HIV-1 infection of CD4+ cells |
Non-Patent Citations (1)
Title |
---|
See also references of EP1322332A2 * |
Cited By (15)
Publication number | Priority date | Publication date | Assignee | Title |
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US7935797B2 (en) | 1996-04-01 | 2011-05-03 | Progenics Pharmaceuticals Inc. | CCR5 chemokine receptor-specific monoclonal antibodies capable of inhibiting HIV-1 cell fusion |
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 |
US7122185B2 (en) | 2002-02-22 | 2006-10-17 | Progenics Pharmaceuticals, Inc. | Anti-CCR5 antibody |
US7615216B2 (en) | 2005-04-01 | 2009-11-10 | Roche Palo Alto Llc | Antibodies against CCR5 and uses thereof |
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US7951920B2 (en) | 2006-08-17 | 2011-05-31 | Roche Palo Alto Llc | Conjugate of an antibody against CCR5 and an antifusogenic peptide |
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EP3077051A4 (en) * | 2013-12-02 | 2018-01-31 | Aaron Diamond Aids Research Center | Improved hiv-1-neutralizing antibody potency and breadth via cell receptor anchoring using bispecific antibodies with native architecture |
CN106102837B (en) * | 2013-12-02 | 2020-10-13 | 艾伦戴蒙德艾滋病研究中心 | Bispecific HIV-1-neutralizing antibodies |
Also Published As
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WO2002022077A3 (en) | 2002-06-20 |
AU9092501A (en) | 2002-03-26 |
AU2001290925C1 (en) | 2009-01-22 |
US7736649B2 (en) | 2010-06-15 |
US7138119B2 (en) | 2006-11-21 |
CA2422275A1 (en) | 2002-03-21 |
US20070020280A1 (en) | 2007-01-25 |
US20020106374A1 (en) | 2002-08-08 |
EP1322332A2 (en) | 2003-07-02 |
AU2001290925B2 (en) | 2006-07-13 |
EP1322332A4 (en) | 2006-06-21 |
JP2004518624A (en) | 2004-06-24 |
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