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WO2014033266A1 - Anticorps anti-sr-bi pour inhiber une infection par le virus de l'hépatite c - Google Patents

Anticorps anti-sr-bi pour inhiber une infection par le virus de l'hépatite c Download PDF

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Publication number
WO2014033266A1
WO2014033266A1 PCT/EP2013/068008 EP2013068008W WO2014033266A1 WO 2014033266 A1 WO2014033266 A1 WO 2014033266A1 EP 2013068008 W EP2013068008 W EP 2013068008W WO 2014033266 A1 WO2014033266 A1 WO 2014033266A1
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Prior art keywords
hcv
infection
monoclonal antibody
antibody
antibodies
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PCT/EP2013/068008
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English (en)
Inventor
Thomas Baumert
Mirjam ZEISEL
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INSERM (Institut National de la Santé et de la Recherche Médicale)
Université De Strasbourg
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Publication of WO2014033266A1 publication Critical patent/WO2014033266A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/395—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum
    • A61K39/39533—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum against materials from animals
    • 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
    • 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
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/30—Immunoglobulins specific features characterized by aspects of specificity or valency
    • C07K2317/34—Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/70—Immunoglobulins specific features characterized by effect upon binding to a cell or to an antigen
    • C07K2317/76—Antagonist effect on antigen, e.g. neutralization or inhibition of binding
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/90—Immunoglobulins specific features characterized by (pharmaco)kinetic aspects or by stability of the immunoglobulin
    • C07K2317/92—Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value

Definitions

  • HCV Hepatitis C virus
  • Chronic HCV infection frequently results in serious liver disease, including fibrosis and steatosis (Chisari, Nature, 2005, 435: 930-932). About 20% of patients with chronic HCV infection develop liver cirrhosis, which progresses to hepatocellular carcinoma in 5% of the cases (Hoofnagle, Hepatology, 2002, 36: S21- S2).
  • Chronic HCV infection is the leading indication for liver transplantations (Seeff, Semin. Gastrointest., 1995, 6: 20-27).
  • liver transplantation is not a cure for hepatitis C; viral recurrence is an invariable problem and leading cause of graft loss (Brown, Nature, 2005, 436: 973-978).
  • Current therapies include administration of ribavirin and/or interferon-alpha (IFN-a), two non-specific anti-viral agents. Using a combination treatment of pegylated IFN-a and ribavirin, persistent clearance is achieved in about 50% of patients with chronic hepatitis C.
  • IFN-a interferon-alpha
  • HCVcc infectious particles in cell culture
  • HCVpp retroviral HCV pseudoparticles
  • HCV is a positive strand RNA virus classified in the Hepacivitus genus, within the Flaviviridae family. Translation of the major open reading frame of the HCV genome results in the production of an approximately 3000 amino acid long polyprotein, which is cleaved co- and post-translationally by the coordinated action of cellular and viral proteases into at least 10 mature proteins, including two envelope glycoproteins (El and E2). HCV initiates infection by attaching to molecules or receptors on the surface of hepatocytes. Current evidence suggests that at least three host cell molecules are important for HCV entry in vitro: the tetraspanin CD81 (Bartosch et al, J. Exp.
  • the present invention relates to targeted systems and strategies for the prevention and/or treatment of HCV infection and HCV-related diseases.
  • the present invention is directed to monoclonal antibodies that inhibit HCV infection and viral spread by interfering with HCV entry in one or more step(s) occurring following binding of the HCV envelope to the host cell membrane.
  • the monoclonal antibodies of the present invention recognize the extracellular domain of human SR-BI, and more specifically the N-terminal half of the extracellular domain of human SR-BI. These antibodies can be used in the prophylactic or therapeutic treatment of HCV infection (acute or chronic HCV infection) and HCV-related diseases or disorders ⁇ e.g., liver inflammation, cirrhosis, and hepatocellular carcinoma).
  • Monoclonal antibodies such as those provided herein that interfere with HCV entry into cells during post-binding steps are particularly attractive as antiviral therapeutics.
  • An inhibitor of HCV entry does not need to cross the plasma membrane or to be modified intracellularly.
  • antibody inhibitors of viral entry can be very potent and less susceptible to the development of viral resistance.
  • the monoclonal antibodies of the present invention are effective at inhibiting HCV variants that are resistant to direct-acting antivirals currently in clinical use.
  • the present invention provides hybridoma cell lines which secrete monoclonal antibodies that specifically bind to the N-terminal half of the extracellular domain of human SR-BI.
  • the present Applicants have deposited four of such hybridoma cell lines at the CNCM (Collection Nationale de Culture de Microorganismes, Institut Pasteur, 25 rue du Dondel Roux, 75724 Paris Cedex 15, France) on August 1, 2012. They were assigned Accession Numbers CNCM 1-4662, CNCM 1-4663, CNCM 1-4664 and CNCM 1-4665. The deposit was made pursuant to the provisions of the Budapest Treaty on the International recognition of the Deposit of Microorganism for the Purpose of Patent Procedure (Budapest Treaty).
  • the present invention provides a monoclonal antibody that is secreted by any one of the hybridoma cell lines deposited under Accession Numbers CNCM 1-4662, CNCM 1-4663, CNCM 1-4664 and CNCM 1-4665.
  • the monoclonal antibody may or may not be isolated and/or purified from hybridoma cultures.
  • the monoclonal antibody is an immunoglobulin of the rat IgG2 heavy (H) chain and kappa light (L) chain isotype.
  • the monoclonal antibody is an immunoglobulin of the mouse IgG2 heavy (H) chain and kappa light (L) chain isotype.
  • monoclonal antibodies secreted by the deposited hybridoma cell lines specifically bind to the extracellular domain of human SR-BI and more specifically to the N-terminal half of the extracellular domain of human SR-BI.
  • the Applicants have also shown that these monoclonal antibodies efficiently inhibit HCV infection in vitro by interfering with HCV entry during post- binding steps.
  • the present invention also encompasses any biologically active fragment of the inventive monoclonal antibodies, i.e., any fragment or portion that retains the ability of the monoclonal antibody to interfere with HCV-host cells interactions during post-binding steps, and/or to specifically bind to the extracellular domain of human SR-BI, in particular the N-terminal half of this extracellular domain, and/or to inhibit or block HCV entry into HCV- susceptible cells, and/or to inhibit or block HCV viral spread, and/or to reduce or prevent HCV infection of susceptible cells.
  • any biologically active fragment of the inventive monoclonal antibodies i.e., any fragment or portion that retains the ability of the monoclonal antibody to interfere with HCV-host cells interactions during post-binding steps, and/or to specifically bind to the extracellular domain of human SR-BI, in particular the N-terminal half of this extracellular domain, and/or to inhibit or block HCV entry into HCV- susceptible cells, and/or to inhibit or block HCV viral spread, and/or to
  • the present invention encompasses any molecule that comprises an inventive anti-SR-BI monoclonal antibody or a fragment thereof, including chimeric antibodies, humanized antibodies, de-immunized antibodies and antibody- derived molecules comprising at least one complementary determining region (CDR) from either a heavy chain or light chain variable region of an inventive anti-SR-BI monoclonal antibody as secreted by a hybridoma cell line, including molecules such as Fab fragments, F(ab') 2 fragments, Fd fragments, Fab fragments, Sc antibodies (single chain antibodies), diabodies, individual antibody light single chains, individual antibody heavy chains, chimeric fusions between antibody chains and other molecules, and antibody conjugates, such as antibodies conjugated to a diagnostic agent (detectable moiety) or therapeutic agent, so long as these antibody-related molecules retain at least one biologically relevant property of the inventive monoclonal antibody from which it is "derived".
  • CDR complementary determining region
  • the biologically relevant property may be the ability to interfere with HCV-host cells interactions during post-binding steps, to specifically bind to the extracellular domain of human SR-BI, in particular to the N-terminal half thereof, to inhibit or block HCV entry into HCV-susceptible cells, to inhibit or block HCV viral spread, and/or to reduce or prevent HCV infection of susceptible cells.
  • the monoclonal antibodies and antibody-related molecules of the present invention can find application in a variety of prophylactic and therapeutic treatments. Accordingly, in another aspect, the inventive monoclonal and antibody-related molecules are provided for use in preventing HCV infection of a cell (e.g., a susceptible cell or a population of susceptible cells); preventing or treating HCV infection or a HCV-related disease in a subject; and preventing HCV recurrence in a liver transplantation patient.
  • a cell e.g., a susceptible cell or a population of susceptible cells
  • the HCV infection may be due to HCV of a genotype selected from the group consisting of genotype 1, genotype 2, genotype 3, genotype 4, genotype 5 and genotype 6, or more specifically of a subtype selected from the group consisting of subtype la, subtype lb, subtype 2a, subtype 2b, subtype 2c, subtype 3a, subtype 4a-f, subtype 5a, and subtype 6a.
  • the HCV infection is due to a HCV that is resistant to at least one direct-acting antiviral.
  • the at least one direct-acting antiviral may be a protease inhibitor, such as boceprevir or telaprevir.
  • the present invention provides a method of reducing the likelihood of a susceptible cell of becoming infected with HCV as a result of contact with HCV, which comprises contacting the susceptible cell with an effective amount of an inventive antibody or antibody-related molecule. Also provided is a method of reducing the likelihood of a subject' s susceptible cells of becoming infected with HCV as a result of contact with HCV, which comprises administering to the subject an effective amount of an inventive antibody or antibody-related molecule.
  • the present invention also provides a method of treating or preventing HCV infection or a HCV-associated disease (e.g. , a liver disease or pathology) in a subject in need thereof which comprises administering to the subject an effective amount of an inventive antibody or antibody-related molecule.
  • a method of preventing HCV recurrence in a liver transplantation patient which comprises administering to the patient an effective amount of an inventive antibody or antibody-related molecule.
  • Administration of an inventive antibody or antibody-related molecule to a subject may be by any suitable route, including, for example, parenteral, aerosol, oral and topical routes.
  • the inventive antibody or antibody-related molecule may be administered alone or in combination with a therapeutic agent, such as an anti- viral agent.
  • inventive monoclonal antibodies and antibody-related molecules may be administered per se or as pharmaceutical compositions. Accordingly, in another aspect, the present invention provides for the use of an inventive monoclonal antibody or antibody-related molecule for the manufacture of medicaments, pharmaceutical compositions, or pharmaceutical kits for the treatment and/or prevention of HCV infection and HCV-associated diseases.
  • the present invention provides a pharmaceutical composition comprising an effective amount of an inventive antibody or antibody-related molecule and at least one pharmaceutically acceptable carrier or excipient.
  • the pharmaceutical composition is adapted for administration in combination with an additional therapeutic agent, such as an antiviral agent.
  • the pharmaceutical composition further comprises an additional therapeutic agent, such as an antiviral agent.
  • Antiviral agents suitable for use in methods and pharmaceutical compositions of the present invention include, but are not limited to, interferons (e.g.
  • interferon-alpha interferon-alpha
  • pegylated interferon-alpha ribavirin
  • anti-HCV monoclonal or polyclonal antibodies
  • RNA polymerase inhibitors protease inhibitors
  • IRES inhibitors helicase inhibitors
  • antisense compounds ribozymes, and any combination thereof.
  • the present invention provides a combination of at least one anti-SR-BI monoclonal antibody as described herein and:
  • anti-HCV envelope antibody selected from anti-El antibodies, anti- E2 antibodies, and anti-HCV IgGs from individuals chronically or previously infected with HCV, or
  • interferon selected from IFNcc-2a and IFNcc-2b, or
  • At least one host-targeting agent such as alisporivir or an anti-CLDNl monoclonal antibody
  • anti-SR-BI monoclonal antibody and anti-HCV envelope antibody or the anti-SR-BI monoclonal antibody and protein kinase inhibitor, or the anti-SR-BI monoclonal antibody and direct acting antiviral, or the anti-SR-BI monoclonal antibody and host-targeting agent act in synergy to inhibit HCV infection.
  • the combination is used for the treatment of HCV infection or a HCV-related disease in a subject; or for the control of chronic HCV infection in a subject; or for the prevention of HCV re-infection and recurrence in a liver transplantation patient.
  • HCV infection, chronic HCV infection and HCV reinfection may be due to HCV of any of the major genotypes or subtypes, as described above.
  • HCV infection, chronic HCV infection and HCV reinfection are due to a HCV that is resistant to at least one direct-acting antiviral.
  • the at least one direct-acting antiviral may be a protease inhibitor, such as boceprevir or telaprevir.
  • the HCV infection, HCV-related disease or HCV reinfection is caused by a Hepatitis C virus that is resistant to a direct acting antiviral and/or that is transmitted by cell-cell transmission.
  • FIG. 1 Binding of monoclonal anti-SR-BI antibodies to human hepatocytes and inhibition of HCV infection.
  • Huh7.5.1 cells and PHH primary human hepatocytes (PHH) were incubated with anti-SR-BI mAbs and antibody binding was assessed using flow cytometry. Results are expressed as net mean fluorescence intensity (AMFI) of a representative experiment.
  • C Inhibition of HCVcc infection by anti-SR-BI mAbs.
  • Huh7.5.1 cells were preincubated for 1 hour at 37°C with anti-SR-BI or control mAbs (100 ⁇ g/mL) before infection with HCVcc (Luc-Jcl) for 4 hours at 37 °C.
  • HCV infection was assessed by lucif erase activity in lysates of infected Huh7.5.1 cells 72 hours post-infection. Results are expressed as means + SD % HCVcc infectivity in the absence of antibody of three independent experiments.
  • D Dose-dependent inhibition of HCVcc infection by anti-SR-BI mAbs. Huh7.5.1 cells were preincubated for 1 hour at 37°C with anti-SR-BI or control mAbs at the indicated concentrations before infection with HCVcc (Luc-Jcl) for 4 hours at 37°C. HCV infection was assessed by luciferase activity in lysates of infected Huh7.5.1 cells 72 hours post-infection.
  • Results are expressed as mean + SD % HCVcc infectivity in the absence of antibody of three independent experiments performed in triplicate. *, P ⁇ 0.01.
  • Figure 2. Monoclonal anti-SR-BI antibodies do not interfere with HCV binding to SR-BI but inhibit HCV entry at post-binding steps.
  • Huh7.5.1 cells were pre-incubated with heparin (100 ⁇ g/mL), anti-SR-BI or control (CTRL) serum (1:50) or anti-SR-BI or control (CTRL IgG) mAbs (20 ⁇ g/mL) for 1 hour prior to incubation with HCVcc (Jcl) at 4°C in the presence of compounds.
  • HCVcc (Luc- Jcl) binding to Huh7.5.1 cells was performed in the presence or absence of anti-CD81 (5 ⁇ g/mL), anti-SR-BI (20 ⁇ g/mL) or control mAbs (20 ⁇ g/mL) or proteinase K (50 ⁇ g/mL) for 1 hour at 4°C, before cells were washed and incubated for 4 hours at 37°C with compounds added at different time-points during infection. Compounds were then removed and cells were cultured for an additional 48 hours. Dashed lines indicate the time intervals where compounds were present. (C) HCV entry kinetics.
  • HCV infection was assessed by luciferase activity in lysates of infected Huh7.5.1 cells 48h post-infection. Results are expressed as mean % HCVcc infectivity in the absence of antibody of three independent experiments performed in triplicate. *, P ⁇ 0.01 Figure 3.
  • the SR-BI post-binding function is relevant for HCV cell-to-cell transmission and viral spread.
  • A Quantification of HCV-infected target cells (Ti) after co-cultivation with HCV (Jcl) producer cells (Pi) during incubation with control or anti-SR-BI mAbs (10 ⁇ g/mL) in the presence of E2-neutralizing antibody AP33 (25 ⁇ g/mL) by flow cytometry. Data are expressed as % infected target cells and represent means + SD of three independent experiments.
  • B Quantification of HCV cell-to-cell transmission in parental target cells compared to target cells overexpressing mouse (m) or human (h) SR-BI. Data are expressed as means + SD from three different experiments.
  • C-D Long-term analysis of HCVcc (Luc-Jcl) infection in the presence or absence of control (10 ⁇ g/mL) or anti-SR-BI mAb (C) QQ-4G9-A6 or (D) NK-8H5-E3 at the indicated concentrations.
  • Antibodies were added 48 hours after HCVcc infection and control medium or medium containing antibodies were replenished every 4 days. Luciferase activity was determined in cell lysates every 2 days. Data are expressed as LoglO RLU and represent means + SD of one representative out of three different experiments performed in duplicate.
  • E-F Cell spread in the presence or absence of anti-SR-BI mAbs.
  • Antibodies were added 48 hours after HCVcc (Jcl) infection and control medium or medium containing antibodies were replenished every 4 days. HCV-infected cells were visualized 7 days post-infection by immunofluorescence using (E) anti-NS5A or (F) anti-E2 (CBH23) antibodies. The percentage of infected cells was calculated as the number of infected cells relative to the total number of cells as assessed by DAPI staining of the nuclei. *, P ⁇ 0.01
  • FIG. 1 Monoclonal anti-SR-BI antibodies block HCV cell-to-cell transmission and spread.
  • A-B Quantification of HCV-infected target cells (Ti) after co-cultivation with HCV producer cells (Pi) during incubation with (A) control or anti-SR-BI mAb QQ-4G9-A6 (10 ⁇ g/mL) or (B) control or anti-SR-BI mAb QQ- 2A10-A5 (10 ⁇ g/mL) in the presence of E2-neutralizing antibody AP33 (25 ⁇ g/mL) by flow cytometry.
  • C Cell viability after long-term exposure to anti-SR-BI mAbs.
  • BRL3- hSR-BI cells were incubated in the presence or absence of anti-SR-BI mAbs (20 ⁇ g/mL) or polyclonal serum (1:50) or respective controls, prior to Cy5-HDL binding for 1 hour at 4°C. Bound Cy5-HDL was quantified using flow cytometry. Results represent mean + SD of three different experiments performed in duplicate.
  • B Lipid uptake by Huh7 cells. Huh7 cells were incubated with a mixture of anti-SR- BI mAbs (20 ⁇ g/mL) and H-CE-labeled HDL for 5 hours before incubation with unlabelled HDL for 30 minutes.
  • FIG. 9 Competition of monoclonal anti-SR-BI antibodies for cellular binding.
  • Huh7.5.1 cells were incubated with 0.1 ⁇ g/mL of biotinylated anti-SR-BI mAb (A) QQ-4A3-A1, (B) QQ-2A10-A5, (C) QQ-4G9-A6 or (D) NK-8H5-E3, together with increasing concentrations of unlabeled control or anti-SR-BI mAb (QQ- 4A3-A1, QQ-2A10-A5, QQ-4G9-A6, NK-8H5-E3) as competitors. Following washing of cells with PBS, binding of labelled mAbs was determined by flow cytometry and is shown % binding relative to biotinylated mAb incubated in the absence of antibody.
  • Results are expressed as means + SD net mean fluorescence intensity (AMFI).
  • D BRL3A cells engineered to express wild-type human SR-BI (SR-BI wt) or human SR-BI point mutants (Q420R, Q402R, E418R, and Q402R-E418R) were first incubated with monoclonal anti-SR-BI antibodies (20 ⁇ g/mL) for lh at RT before bound antibodies were detected using PE-labelled secondary antibodies.
  • Results are expressed as means + SD net mean fluorescence intensity (AMFI).
  • Figure 11 Western blot analysis of anti-SR-BI mAb binding to endogenous SR-BI expressed in Huh7.5.1 cells.
  • Huh7.5.1 cells were pre-incubated for 1 hour with serial concentrations of (A) IFN- a2a or (B) IFN-a2b or (C) protease inhibitors telaprevir, boceprevir, TMC-435 or danoprevir, NS5A inhibitor daclatasvir or polymerase inhibitors mericitabine or GS- 7977and 0.01 ⁇ g/ml anti-SRBI mAb before incubation with HCVcc Luc-Jcl in the presence of both compounds. HCVcc infection was analyzed by lucif erase reporter gene expression. The CI for an IC 50 was calculated and is indicated in Table 2.
  • Huh7.5.1 cells were pre-incubated with serial concentrations of alisporivir and 0.01 ⁇ g/ml of anti-CD81, anti-SRBI or anti-CLDNl mAbs or 0.1 ⁇ erlotinib or dasatinib before incubation with HCVcc Luc-Jcl in the presence of both compounds.
  • Huh7.5.1 cells were pre-incubated with serial concentrations of anti-CD81, anti- SR-BI or anti-CLDNl mAbs and 0.1 ⁇ erlotinib or dasatinib before incubation with
  • preventing, inhibiting or blocking HCV infection when used in reference to an inventive antibody or antibody-related molecule, means reducing the amount of HCV genetic information introduced into a susceptible cell or susceptible cell population as compared to the amount that would be introduced in the absence of the antibody or antibody-related molecule.
  • the monoclonal antibodies may be prepared by any other suitable method known in the art.
  • an inventive anti-SR-BI monoclonal antibody may be prepared by recombinant DNA methods. These methods generally involve isolation of the genes encoding the desired antibody, transfer of the genes into a suitable vector, and bulk expression in a cell culture system.
  • Antibody fragments of the present invention may be produced by any suitable method known in the art including, but not limited to, enzymatic cleavage (e.g. , proteolytic digestion of intact antibodies) or by synthetic or recombinant techniques.
  • F(ab') 2 , Fab, Fv and ScFv (single chain Fv) antibody fragments can, for example, be expressed in and secreted from mammalian host cells or from E. coli.
  • Antibodies can also be produced in a variety of truncated forms using antibody genes in which one or more stop codons have been introduced upstream of the natural stop site.
  • the various portions of antibodies can be joined together chemically by conventional techniques, or can be prepared as a contiguous protein using genetic engineering techniques.
  • molecular entities are attached at positions on the antibody molecule that do not interfere with the binding properties of the resulting conjugate, i.e., positions that do not participate in the specific binding of the antibody to the N-terminal half of the extracellular domain of human SR-BI.
  • the antibody molecule and molecular entity are covalently, directly linked to each other.
  • the direct covalent binding can be through a linkage such as an amide, ester, carbon-carbon, disulfide, carbamate, ether, thioether, urea, amine or carbonate linkage.
  • Covalent binding can be achieved by taking advantage of functional groups present on the antibody and the molecular entity.
  • An activating agent such as a carbodiimide, can be used to form a direct linkage.
  • the antibody molecule and the molecular entity are covalently linked to each other through a linker group. This can be accomplished by using any of a wide variety of stable bifunctional agents well known in the art, including homofunctional and heterofunctional linkers.
  • an antibody of the present invention (or a biologically active fragment thereof) is conjugated to a therapeutic moiety.
  • a therapeutic moiety Any of a wide variety of therapeutic moieties may be suitable for use in the practice of the present invention including, without limitation, cytotoxins (e.g., cytostatic or cytocidal agents), therapeutic agents, and radioactive metal ions (e.g., alpha-emitters and alpha-emitters attached to macrocyclic chelators such as DOTA).
  • Cytotoxins or cytotoxc agents include any agent that is detrimental to cells.
  • Therapeutic agents include, but are not limited to, antimetabolites (e.g., methotrexate, 6-mercaptopurine, 6-thioguanine, cytarabine, 5-fluorouracil decarbazine), alkylating agents (e.g., mechlorethamine, thioepa chlorambucil, melphalan, carmustine (BSNU) and lomustine (CCNU), cyclothosphamide, busulfan, dibromomannitol, streptozotocin, mitomycin C, and cisdichlorodiamine platinum (II) (DDP) cisplatin), anthracyclines (e.g., antimetabolites (e.g., methotrexate, 6-mercaptopurine, 6-thioguanine, cytarabine, 5-fluorouracil decarbazine), alkylating agents (e.g., mechlorethamine, thioepa chlorambuci
  • lymphokines interleukin-1 (IL- 1), interleukin-2 (IL-2), interleukin-6 (IL-6), granulocyte macrophage colony stimulating factor (GM-CSF), granulocyte colony stimulating factor (G-CSF), or other growth factors).
  • IL-1 interleukin-1
  • IL-2 interleukin-2
  • IL-6 interleukin-6
  • GM-CSF granulocyte macrophage colony stimulating factor
  • G-CSF granulocyte colony stimulating factor
  • each of the inventive anti-SR-BI monoclonal antibodies described in the Examples was produced from a hybridoma cell line provided herein and was selected for its ability to inhibit HCVcc infection of Huh7.5.1 cells.
  • the HCV infection inhibitory effect of other antibodies and antibody-related molecules of the invention may also be assessed using a HCVcc infection system.
  • the inhibitory effect of antibodies and antibody-related molecules on HCV infection may, alternatively or additionally, be assessed using retroviral HCV pseudotyped particles (HCVpp) as known in the art.
  • HCVpp retroviral HCV pseudotyped particles
  • an antibody or antibody-related molecule of the present invention will be shown to inhibit HCV infection of susceptible cells by HCVcc or HCVpp in a dose-dependent manner.
  • Binding specificity testing may be performed using the antibody or antibody-related molecule against a panel of cells, e.g., human cells, including, without limitation, liver cell lines (such as, for example, Huh7, Hep3b or HepG2), embryonic kidney cells (293T), fibroblasts (HeLa), B cells, T cells (e.g., Molt-4, Sup-TI, or Hut-78), monocytic cells (THP-I), astrocytic cells (U87), hepatoma cells (PLC/PRF:5) or other liver cell types, e.g., the liver adenocarcinoma SkHepI, human peripheral blood cells and various fractionated subtypes thereof including lymphocytes and monocytes or other cell lines including CaCo2 cells.
  • Flow cytometry analysis can reveal binding specificity of the antibody or antibody-related molecule for SR-BI on various cell types. Cells from non-human mammals may also be used in such assays.
  • IC 50 values may be determined for the antibodies and antibody-related molecules of the present invention. These values, which give an indication of the concentration of antibody or antibody-related molecule required for 50% inhibition of viral infectivity, provide meaningful and significant quantitative criteria and allow comparison of the infection inhibiting activity of different antibodies and antibody-related molecules.
  • Anti-SR-BI antibodies of the present invention may be used in therapeutic and prophylactic methods to treat and/or prevent HCV infection, or to treat and/or prevent a liver disease or a pathological condition affecting HCV- susceptible cells, such as liver cells, lymphoid cells, or monocytes/macrophages.
  • An inventive anti-SR-BI antibody interferes with HCV-host cells interactions during post-binding steps by binding to the extracellular domain of SR-BI on a cell surface, thereby reducing, inhibiting, blocking or preventing HCV entry into the cell and/or HCV infection of the cell.
  • Methods of treatment of the present invention may be accomplished using an inventive antibody or a pharmaceutical composition comprising an inventive antibody (see below). These methods generally comprise administration of an effective amount of at least one inventive anti-SR-BI antibody, or a pharmaceutical composition thereof, to a subject in need thereof. Administration may be performed using any of the methods known to one skilled in the art.
  • the antibody or composition may be administered by various routes including, but not limited to, aerosol, parenteral, oral or topical route.
  • an inventive antibody or composition will be administered in an effective amount, i.e. an amount that is sufficient to fulfill its intended purpose.
  • an effective amount is one that inhibits or prevents HCV from entering into a subject's susceptible cells and/or infecting a subject's cells, so as to thereby prevent HCV infection, treat or prevent liver disease or another HCV- associated pathology in the subject.
  • Antibodies and compositions of the present invention may be used in a variety of therapeutic or prophylactic methods.
  • the present invention also provides a method for treating or preventing a HCV- associated disease or condition (including a liver disease) in a subject, which comprises administering to the subject an effective amount of an inventive antibody (or composition thereof) which inhibits HCV from entering or infecting the subject's cells, so as to thereby treat or prevent the HCV-associated disease or condition in the subject.
  • the antibody or composition is administered to a subject diagnosed with acute hepatitis C.
  • the antibody or composition is administered to a subject diagnosed with chronic hepatitis C.
  • Administration of an inventive antibody or composition according to such methods may result in amelioration of at least one of the symptoms experienced by the individual including, but not limited to, symptoms of acute hepatitis C such as decreased appetite, fatigue, abdominal pain, jaundice, itching, and flu-like symptoms; symptoms of chronic hepatitis C such as fatigue, marked weight loss, flu-like symptoms, muscle pain, joint pain, intermittent low-grade fevers, itching, sleep disturbances, abdominal pain, appetite changes, nausea, diarrhea, dyspepsia, cognitive changes, depression, headaches, and mood swings; symptoms of cirrhosis such as ascites, bruising and bleeding tendency, bone pain, varices (especially in the stomach and esophagus), steatorrhea, jaundice and hepatic encephalopathy; and symptoms of extrahepatic manifestations associated with HCV such as thyroiditis, porphyria cutanea tarda, cryoglobulinemia, glomerulonephritis, sicca syndrome, thrombocytopenia,
  • administration of an inventive antibody or composition according to such methods may slow, reduce, stop or alleviate the progression of HCV infection or an HCV-associated disease, or reverse the progression to the point of eliminating the infection or disease.
  • Administration of an inventive antibody or composition according to such methods may also result in a reduction of the number of viral infections, reduction of the number of infectious viral particles, and/or reduction in the number of virally infected cells.
  • the effects of a treatment according to the invention may be monitored using any of the assays known in the art for the diagnosis of HCV infection and/or liver disease.
  • assays include, but are not limited to, serological blood tests, liver function tests to measure one or more of albumin, alanine transaminase (ALT), alkaline phosphatase (ALP), aspartate transaminase (AST), and gamma glutamyl transpeptidase (GGT), and molecular nucleic acid tests using different techniques such as polymerase chain reaction (PCR), transcription mediated amplification (TMA), or branched DNA (bDNA).
  • PCR polymerase chain reaction
  • TMA transcription mediated amplification
  • bDNA branched DNA
  • Antibodies and compositions of the present invention may also be used in immunization therapies.
  • the present invention provides a method of reducing the likelihood of susceptible cells of becoming infected with HCV as a result of contact with HCV.
  • the method comprises contacting the susceptible cells with an effective amount of an inventive antibody or composition which inhibits HCV from entering or infecting the susceptible cells, so as to reduce the likelihood of the cells to become infected with HCV as a result of contact with HCV.
  • the present invention also provides a method of reducing the likelihood of a subject's susceptible cells of becoming infected with HCV as a result of contact with HCV.
  • contacting the susceptible cells with the inventive antibody or composition may be performed by administrating the antibody or composition to the subject.
  • Reducing the likelihood of susceptible cells or of a subject of becoming infected with HCV means decreasing the probability of susceptible cells or a subject to become infected with HCV as a result of contact with HCV.
  • the decrease may be of any significant amount, e.g., at least a 2-fold decrease, more than a 2-fold decrease, at least a 10-fold decrease, more than a 10-fold decrease, at least a 100-fold decrease, or more than a 100-fold decrease.
  • the subject is infected with HCV prior to administration of the inventive antibody or composition.
  • the subject is not infected with HCV prior to administration of the inventive antibody or composition.
  • the subject is not infected with, but has been exposed to, HCV.
  • the subject may be infected with HIV or HBV.
  • the methods of the present invention may be used to reduce the likelihood of a subject's susceptible cells of becoming infected with HCV as a result of liver transplant.
  • serum viral levels plummet.
  • virus levels rebound and can surpass pre-transplant levels within a few days (Powers et ah, Liver TranspL, 2006, 12: 207-216).
  • Liver transplant patients may benefit from administration of an inventive antibody that binds to N- terminal half of the extracellular domain of human SR-BI on the surface of hepatocytes and thereby reduce, inhibit, block or prevent HCV entry into the cells. Administration may be performed prior to liver transplant, during liver transplant, and/or following liver transplant.
  • an inventive antibody or composition include, but are not limited to, babies born to HCV-infected mothers, in particular if the mother is also HIV-positive; health-care workers who have been in contact with HCV-contaminated blood or blood contaminated medical instruments; drug users who have been exposed to HCV by sharing equipments for injecting or otherwise administering drugs; and people who have been exposed to HCV through tattooing, ear/body piercing and acupuncture with poor infection control procedures.
  • an inventive antibody or composition include, but are not limited to, subjects that exhibit one or more factors that are known to increase the rate of HCV disease progression. Such factors include, in particular, age, gender (males generally exhibit more rapid disease progression than females), alcohol consumption, HIV co-infection (associated with a markedly increased rate of disease progression), and fatty liver.
  • Therapeutic agents that may be administered in combination with an inventive antibody or composition may be selected among a large variety of biologically active compounds that are known to have a beneficial effect in the treatment or prevention of HCV infection, or a HCV-associated disease or condition.
  • Such agents include, in particular, antiviral agents including, but not limited to, interferons (e.g., interferon- alpha, pegylated interferon- alpha), ribavirin, anti-HCV (monoclonal or polyclonal) antibodies, RNA polymerase inhibitors, protease inhibitors, IRES inhibitors, helicase inhibitors, antisense compounds, ribozymes, and any combination thereof.
  • interferons e.g., interferon- alpha, pegylated interferon- alpha
  • ribavirin e.g., anti-HCV (monoclonal or polyclonal) antibodies
  • RNA polymerase inhibitors e.g., protease inhibitors,
  • An inventive antibody (optionally after formulation with one or more appropriate pharmaceutically acceptable carriers or excipients), in a desired dosage can be administered to a subject in need thereof by any suitable route.
  • Various delivery systems are known and can be used to administer antibodies of the present invention, including tablets, capsules, injectable solutions, encapsulation in liposomes, microparticles, microcapsules, etc.
  • Methods of administration include, but are not limited to, dermal, intradermal, intramuscular, intraperitoneal, intralesional, intravenous, subcutaneous, intranasal, pulmonary, epidural, ocular, and oral routes.
  • the amount of monoclonal antibody administered will preferably be in the range of about 1 ng/kg to about 100 mg/kg body weight of the subject, for example, between about 100 ng/kg and about 50 mg/kg body weight of the subject; or between about 1 g/kg and about 10 mg/kg body weight of the subject, or between about 100 g/kg and about 1 mg/kg body weight of the subject.
  • anti-SR-BI antibodies (and related molecules) of the invention may be administered per se or as a pharmaceutical composition.
  • the present invention provides pharmaceutical compositions comprising an effective amount of an inventive antibody described herein and at least one pharmaceutically acceptable carrier or excipient.
  • the composition further comprises one or more additional biologically active agents.
  • compositions of the present invention may be formulated in dosage unit form for ease of administration and uniformity of dosage.
  • unit dosage form refers to a physically discrete unit of an inventive anti-SR-BI antibody for the patient to be treated. It will be understood, however, that the total daily dosage of the compositions will be decided by the attending physician within the scope of sound medical judgement.
  • sterile injectable preparations for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents, and suspending agents.
  • the sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a non-toxic parenterally acceptable diluent or solvent, for example, as a solution in 2,3-butanediol.
  • acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S. P. and isotonic sodium chloride solution.
  • sterile, fixed oils are conventionally employed as a solution or suspending medium.
  • any bland fixed oil can be employed including synthetic mono- or di-glycerides.
  • Fatty acids such as oleic acid may also be used in the preparation of injectable formulations.
  • Sterile liquid carriers are useful in sterile liquid form compositions for parenteral administration.
  • In order to prolong the effect of an active ingredient it is often desirable to slow the absorption of the ingredient from subcutaneous or intramuscular injection.
  • Delaying absorption of a parenterally administered active ingredient may be accomplished by dissolving or suspending the ingredient in an oil vehicle.
  • Injectable depot forms are made by forming microencapsulated matrices of the active ingredient in biodegradable polymers such as polylactide-polyglycolide. Depending upon the ratio of active ingredient to polymer and the nature of the particular polymer employed, the rate of ingredient release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations can also be prepared by entrapping the active ingredient in liposomes or microemulsions which are compatible with body tissues.
  • Liquid dosage forms for oral administration include, but are not limited to, pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups, elixirs, and pressurized compositions.
  • the liquid dosage form may contain inert diluents commonly used in the art such as, for example, water or other solvent, solubilising agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cotton seed, ground nut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols, and fatty acid esters of sorbitan and mixtures thereof.
  • inert diluents commonly used in the art such as, for example,
  • the oral compositions can also include adjuvants such as wetting agents, suspending agents, preservatives, sweetening, flavouring, and perfuming agents, thickening agents, colors, viscosity regulators, stabilizes or osmo-regulators.
  • suitable liquid carriers for oral administration include water (potentially containing additives as above, e.g. , cellulose derivatives, such as sodium carboxymethyl cellulose solution), alcohols (including monohydric alcohols and polyhydric alcohols such as glycols) and their derivatives, and oils (e.g. , fractionated coconut oil and arachis oil).
  • the liquid carrier can be halogenated hydrocarbon or other pharmaceutically acceptable propellant.
  • excipients suitable for solid formulations include surface modifying agents such as non-ionic and anionic surface modifying agents.
  • surface modifying agents include, but are not limited to, poloxamer 188, benzalkonium chloride, calcium stearate, cetostearyl alcohol, cetomacrogol emulsifying wax, sorbitan esters, colloidal silicon dioxide, phosphates, sodium dodecylsulfate, magnesium aluminum silicate, and triethanolamine.
  • the dosage form may also comprise buffering agents.
  • Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatine capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like.
  • the solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings, release controlling coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a composition such that they release the active ingredient(s) only, or preferably, in a certain part of the intestinal tract, optionally, in a delaying manner.
  • Examples of embedding compositions which can be used include polymeric substances and waxes.
  • an inventive composition may be desirable to administer an inventive composition locally to an area in need of treatment (e.g. , the liver). This may be achieved, for example, and not by way of limitation, by local infusion during surgery (e.g., liver transplant), topical application, by injection, by means of a catheter, by means of suppository, or by means of a skin patch or stent or other implant.
  • the composition is preferably formulated as a gel, an ointment, a lotion, or a cream which can include carriers such as water, glycerol, alcohol, propylene glycol, fatty alcohols, triglycerides, fatty acid esters, or mineral oil.
  • carriers such as water, glycerol, alcohol, propylene glycol, fatty alcohols, triglycerides, fatty acid esters, or mineral oil.
  • Other topical carriers include liquid petroleum, isopropyl palmitate, polyethylene glycol, ethanol (95%), polyoxyethylenemonolaurat (5%) in water, or sodium lauryl sulphate (5%) in water.
  • Other materials such as antioxidants, humectants, viscosity stabilizers, and similar agents may be added as necessary.
  • the inventive compositions may be disposed within transdermal devices placed upon, in, or under the skin.
  • transdermal devices include patches, implants, and injections which release the active ingredient by either passive or active release mechanisms.
  • Transdermal administrations include all administration across the surface of the body and the inner linings of bodily passage including epithelial and mucosal tissues. Such administrations may be carried out using the present compositions in lotions, creams, foams, patches, suspensions, solutions, and suppositories (rectal and vaginal).
  • Transdermal administration may be accomplished through the use of a transdermal patch containing an active ingredient (i.e. , an inventive anti-SR-BI monoclonal antibody) and a carrier that is non-toxic to the skin, and allows the delivery of the ingredient for systemic absorption into the bloodstream via the skin.
  • the carrier may take any number of forms such as creams and ointments, pastes, gels, and occlusive devices.
  • the creams and ointments may be viscous liquid or semisolid emulsions of either the oil-in- water or water-in-oil type.
  • Pastes comprised of absorptive powders dispersed in petroleum or hydrophilic petroleum containing the active ingredient may be suitable.
  • a variety of occlusive devices may be used to release the active ingredient into the bloodstream such as a semi-permeable membrane covering a reservoir containing the active ingredient with or without a carrier, or a matrix containing the active ingredient.
  • Suppository formulations may be made from traditional materials, including cocoa butter, with or without the addition of waxes to alter the suppository' s melting point, and glycerine.
  • Water soluble suppository bases such as polyethylene glycols of various molecular weights, may also be used.
  • the pharmaceutical composition may further comprise vaccine carriers known in the art such as, for example, thyroglobulin, albumin, tetanus toxoid, and polyamino acids such as polymers of D-lysine and D-glutamate.
  • vaccine carriers known in the art such as, for example, thyroglobulin, albumin, tetanus toxoid, and polyamino acids such as polymers of D-lysine and D-glutamate.
  • the vaccine may also include any of a variety of well known adjuvants such as, for example, incomplete Freund' s adjuvant, alum, aluminium phosphate, aluminium hydroxide, monophosphoryl lipid A (MPL, GlaxoSmithKline), a saponin, CpG oligonucleotides, montanide, vitamin A and various water-in-oil emulsions prepared from biodegradable oils such as squalene and/or tocopherol, Quil A, Ribi Detox, CRL- 1005, L- 121 and combinations thereof.
  • Materials and methods for producing various formulations are known in the art and may be adapted for practicing the subject invention. Suitable formulations for the delivery of antibodies can be found, for example, in "Remington's Pharmaceutical Sciences", E.W. Martin, 18 th Ed., 1990, Mack Publishing Co.: Easton, PA.
  • an inventive anti-SR-BI monoclonal antibody is the only active ingredient in a pharmaceutical composition of the present invention.
  • the pharmaceutical composition further comprises one or more biologically active agents.
  • suitable biologically active agents include, but are not limited to, vaccine adjuvants and therapeutic agents such as anti-viral agents (as described above), anti-inflammatory agents, immunomodulatory agents, analgesics, antimicrobial agents, antibacterial agents, antibiotics, antioxidants, antiseptic agents, and combinations thereof.
  • the anti-SR-BI monoclonal antibody and additional therapeutic agent(s) may be combined in one or more preparations for simultaneous, separate or sequential administration of the anti-SR-BI antibody and therapeutic agent(s). More specifically, an inventive composition may be formulated in such a way that the antibody and therapeutic agent(s) can be administered together or independently from each other.
  • an anti-SR-BI antibody and a therapeutic agent can be formulated together in a single composition. Alternatively, they may be maintained ⁇ e.g., in different compositions and/or containers) and administered separately.
  • the at least one anti-SR-BI monoclonal antibody and at one active ingredient act in synergy to inhibit HCV infection. Accordingly, the present invention also provides synergistic combinations.
  • the present invention provides a combination comprising at least one anti- SR-BI monoclonal antibody according to the invention and at least one additional agent for use in the treatment or prevention of HCV infection, wherein the at least one anti-SR-BI monoclonal antibody and at least one additional agent act in synergy to inhibit HCV infection.
  • the at least one anti-SR-BI monoclonal antibody according to the invention decreases the IC 50 for the inhibition of HCV infection by the additional agent by a factor of at least 10 fold or at least 15 fold, preferably at least 20 fold or at least 25 fold, more preferably at least 30 fold or 40 fold, and even more preferably 50 fold or more than 50 fold.
  • the concentration of the additional agent necessary to obtain a 50% inhibition of HCV entry is at least 10 times or at least 15 times, preferably at least 20 times or at least 25 times, more preferably at least 30 times or at least 40 times, and even more preferably 50 times lower or more than 50 times lower than the concentration of the additional agent that would be necessary to obtain the same HCV entry inhibition in the absence of anti-SR-BI monoclonal antibody.
  • the additional agent decreases the IC 50 for the inhibition of HCV infection by the at least one anti-SR-BI monoclonal antibody according to the invention by a factor of at least 5 times or 10 fold or at least 15 fold, preferably at least 20 fold or at least 25 fold, more preferably at least 30 fold or 40 fold, and even more preferably 50 fold or more than 50 fold.
  • the concentration of anti-SR-BI monoclonal antibody necessary to obtain a 50% inhibition of HCV entry is at least 5 times or at least 10 times or at least 15 times, preferably at least 20 times or at least 25 times, more preferably at least 30 times or at least 40 times, and even more preferably 50 times lower or more than 50 times lower than the concentration of anti-SR-BI monoclonal antibody that would be necessary to obtain the same HCV entry inhibition in the absence of additional agent.
  • the at least one additional agent is an anti- HCV envelope antibody.
  • the at least one anti-HCV envelope antibody is preferably selected from anti-El antibodies, anti-E2 antibodies, anti-HCV IgGs from individuals chronically or previously infected with HCV and mixtures thereof. The Applicants have shown that such combinations act in synergy to inhibit HCV entry into susceptible cells.
  • the one additional agent is a protein kinase inhibitor.
  • protein kinase inhibitor refers to any molecule that specifically blocks the action of one or more protein kinases. Protein kinase inhibitors are subdivided by the amino acids whose phosphorylation is inhibited. Most kinases act on both serine and threonine, the tyrosine kinases act on tyrosine, and a number (dual- specificity) kinases act on all three.
  • the term “serine/threonine kinase inhibitor” refers to a molecule that specifically blocks the action of one or more serine and/or threonine kinases.
  • the term “tyrosine kinase inhibitor” refers to a molecule that specifically blocks the action of one or more tyrosine kinases.
  • erlotinib and dasatinib act in synergy with an anti-SRBI antibody of the invention to inhibit HCV infection.
  • Both erlotinib and dasatinib are tyrosine kinase inhibitors. Therefore, in certain preferred embodiments, the at least one protein kinase inhibitor present in a combination according to the invention is a tyrosine kinase inhibitor.
  • HCV entry cofactors EGFR (epidermal growth factor receptor) and EphA2 (ephrin type-A receptor A)
  • EGFR epidermal growth factor receptor
  • EphA2 ephrin type-A receptor A
  • erlotinib and dasatinib broadly impaired infection by all major HCV genotypes and viral escape variants in vitro and in the human liver-chimeric Alb- uPA/SCID mouse model (Lupberger et al, Nature Medicine, 2011, 17: 589-595). They showed that erlotinib and dasatinib interfere with CD81-CLDN1 co-receptor interactions and with glycoprotein-dependant viral fusion.
  • the at least one protein kinase inhibitor present in a combination of the invention is a tyrosine kinase inhibitor that acts on the epidermal growth factor receptor (EGFR).
  • EGFR epidermal growth factor receptor
  • the target protein EGFR is also sometimes referred to as Herl or ErbB-1.
  • tyrosine kinase inhibitors that act on EGFR include, but are not limited to, erlotinib, gefitinib, vandetanib, and lapatinib.
  • the at least one tyrosine kinase inhibitor that acts on EGFR is erlotinib.
  • Erlotinib is marketed under the tradename TARCEVA ® by Genentech and OSI pharmaceuticals in the United States and by Roche elsewhere. Erlotinib binds in a reversible fashion to the adenosine triphosphate (ATP) binding site of the epidermal growth factor receptor. For the signal to be transmitted, two members of the EGFR family need to come together to form a homodimer.
  • the at least one tyrosine kinase inhibitor that acts on EGFR is gefitinib.
  • Gefitinib (tradename IRESSA ® ) is marketed by AstraZeneca and Teva. In Europe, gefitinib is indicated in advanced non-small cell lung cancer in all lines of treatment for patients harboring EGFR mutations.
  • the at least one tyrosine kinase inhibitor that acts on EGFR is vandetanib.
  • Vandetanib also known as ZD6474 is being developed by AstraZeneca. It is an antagonist of the epidermal growth factor receptor (EGFR) and the vascular endothelial growth factor receptor (VEGFR).
  • EGFR epidermal growth factor receptor
  • VEGFR vascular endothelial growth factor receptor
  • Vandetanib became the first drug to be approved by the FDA for the treatment of late-stage (metastatic) medullatory thyroid cancer in adult patients who are ineligible for surgery.
  • Lapatinib in the form of lapatinib ditosylate (tradenames TYKERB ® in the U.S. and TYVERB ® in Europe) is marketed by GlaxoSmithKline.
  • Lapatinib is a dual tyrosine kinase inhibitor, which inhibits the tyrosine kinase activity associated with EGFR and HER2/neu (Human EGFR type 2).
  • lapatinib received accelerated approval as front-line therapy in triple positive breast cancer.
  • tyrosine kinase inhibitors that act on EGFR and that are suitable for use in the present invention include molecules that are currently under development for human use, including, but not limited to, neratinib (also known as HKI-272, being developed by Pfizer), which is under investigation for the treatment of breast cancer and other solid tumors; and afatinib (also known as BIBW 2992, being developed by Boehringer Ingelheim), which is a candidate drug against non-small cell lung carcinoma, both of which are dual inhibitors of EGFR and Her2.
  • neratinib also known as HKI-272, being developed by Pfizer
  • afatinib also known as BIBW 2992, being developed by Boehringer Ingelheim
  • tyrosine kinase inhibitors that act on EGFR include anti- EGFR antibodies, such as Cetuximab, Panitumumab, Matuzumab, Zalutumumab, Nimotuzumab, Necitumumab, and the like.
  • the at least one protein kinase inhibitor present in a combination of the invention is a tyrosine kinase inhibitor that acts on the ephrin type- A receptor 2 (EphA2).
  • EphA2 ephrin type- A receptor 2
  • Examples of such tyrosine kinase inhibitors include, for example, dasatinib.
  • Dasatinib (BMS-354825) is sold under the tradename SPRYCEL ® by Bristol- Myers Squibb. Dasatinib is a multi-targeted kinase inhibitor mainly developed for Bcr-Abl and Src family kinases, but which also inhibits multiple Eph kinases, including EphA2. Dasatinib is approved for use in patients with chronic myelogenous leukemia (CML) after imatinib treatment, and Philadelphia chromosome-positive acute lymphoblastic leukemia. It is being evaluated for use in numerous other cancers, including advanced prostate cancer.
  • CML chronic myelogenous leukemia
  • tyrosine kinase inhibitors that act on EphA2 include anti- EphA2 antibodies, such as those developed by Medlmmune Inc.
  • the at least one protein kinase inhibitor present in a combination of the present invention is an inhibitor that acts on a receptor tyrosine kinase (RTK) other than EGFR and EphA2, for example an anti-RTK monoclonal antibody.
  • RTK receptor tyrosine kinase
  • anti-RTK monoclonal antibodies include, but are not limited to, anti-VEGF antibodies such as Bevacizumab and Ranibizumab; anti-Erb2 antibodies such as Trastuzumab; anti-HER2/neu antibodies such as Trastuzumab, Ertimaxomab, and Pertuzumab; anti-VEGFR2 antibodies such as Ramucirumab and Alacizumab pegol; anti-VEGF-A antibodies such as Ranibizumab and Bevacizumab; anti-PDGF-R antibodies such as Olaratumab; and anti-IGF-1 receptor antibodies such as Figitumumab; Robatumumal and Cixutumumab.
  • anti-VEGF antibodies such as Bevacizumab and Ranibizumab
  • anti-Erb2 antibodies such as Trastuzumab
  • anti-HER2/neu antibodies such as Trastuzumab, Ertimaxomab, and Pertuzumab
  • the invention also provides a combination comprising at least one anti-SR-BI monoclonal antibody according to the invention and at least one direct acting antiviral (DAA) for use in the treatment or prevention of HCV infection, wherein the at least one anti-SR-BI monoclonal antibody and at least one DAA in synergy to inhibit HCV infection.
  • DAA direct acting antiviral
  • direct acting antiviral', direct acting antiviral agent, “DAA”, “specifically targeted antiviral therapy for hepatitis C” and "STAT-C” are used herein interchangeably. They refer to molecules that interfere with specific steps of the lifecycle of HCV and are thus useful in the prevention or treatment of HCV infection.
  • VX-950 also known as telaprevir
  • ITMN-191 also known as danoprevir
  • telaprevir telaprevir
  • boceprevir telaprevir
  • danoprevir telaprevir
  • TMC-435 HCV protease inhibitors
  • mericitabine also known as RG7128
  • GS-7977 HCV polymerase inhibitors
  • daclatasvir also known as BMS-790052
  • the at least one direct acting antiviral present in a combination according to the invention is a HCV protease inhibitor or a HCV polymerase inhibitor or an NS5A inhibitor.
  • Telaprevir also known as VX-950
  • INCIVEK ® a registered trademark of Lucent Technologies Inc.
  • Vertex Pharmaceuticals, Inc. and Johnson & Johnson the FDA approved telaprevir for the treatment of patients with genotype 1 chronic hepatitis C. ⁇ -191 (also known as R7227 or danoprevir) was being co- developed by Roche and InterMune Inc., but is now fully owned by Roche.
  • Boceprevir (initially developed by Schering-Plough, and then by Merck and marketed under the tradename VICTRELIS ® ) was approved by the FDA for the treatment of hepatitic C genotype 1 in May 2011.
  • BMS-650032 is being developed by Bristol- Myers-Squibb.
  • VX-985 is a NS3/4A protease inhibitor being developed by Vertex Pharmaceuticals, Inc. BI 201335 is being developed by Boehringer Ingelheim and is now in Phase III clinical trials in the United States. TMC435, a NS3/4A protease inhibitor being developed by Medivir/Tibotec/Johnson & Johnson, is also in Phase III clinical trials.
  • NS3/4A protease inhibitors that can be present in a combination according to the invention include, but are not limited to, NS3/4A protease inhibitors that are currently in phase II clinical trials such as GS 9256 and GS 9451 (being developed by Gilead), MK-7009 (also known as vaniprevir, being developed by Merck), ACH-1625 (being developed by Achillion), and ABT-450 (being developed by Abbott/Enanta); NS3/4A protease inhibitors that are currently in phase I clinical trials such as BMS-791325 (being developed by Bristol-Myers Squibb), VX-985 and VX-500 (being developed by Vertex pharmaceuticals), and PHX1766 (being developed by Phenomix); and NS3/4A protease inhibitors that are currently in preclinical trials such as VX-813 (being developed by Vertex), AVL-181 and AVL-192 (being developed by Avila Therapeutics), and ACH-2684 (being developed
  • Mericitabine (also known as RG7128 or RO5024048), is a prodrug of PSI-6130, an oral cytidine nucleoside analogue. It is being developed by Roche and Pharmasset. Mericitabine has shown in vitro activity against all of the most common HCV genotypes.
  • GS-7977 (also known as PSI-7977) is being developed by Gilead Sciences. It is currently in Phase III clinical trials. It is being studied as a treatment to be used in combination with ribavirin.
  • GS-78977 is a prodrug that is metabolized to the active antiviral agent 2'-deoxy-2'-a-fluoro-P-C-methyluridine-5'-monophosphate.
  • NS5B polymerase inhibitors that can be present in a combination according to the invention include, but are not limited to, nucleoside/nucleotide polymerase inhibitors that are currently in Phase II clinical trials such as IDX184 (being developed by Idenix) and PS 1-7977 (being developed by Pharmasset); non-nucleoside polymerase inhibitors that are currently in Phase II clinical trials such as VX-222 (initially developed by ViroChem, now owned by Vertex); PF-868554 (being developed by Pfizer); ABT-072 and ABT-333 (being developed by Abbott), GS 9190 (being developed by Gilead) and ANA598 (also known as setrobuvir, being developed by Anadys); nucleoside/nucleotide polymerase inhibitors that are currently in Phase I clinical trials such as BI 207127 (being developed by Boehringer Ingelheim), MK-0608 (being developed by Isis/Merck), TMC649128
  • NS5A inhibitors suitable for use in the context of the present invention include, in particular in particular daclatasvir (also known as BMS-790052), which was developed by Bristol-Myers-Squibb.
  • the invention also provides a combination comprising at least one anti-SR-BI monoclonal antibody according to the invention and at least one interferon for use in the treatment or prevention of HCV infection, wherein the at least one anti-SR-BI monoclonal antibody and at least one interferon in synergy to inhibit HCV infection.
  • interferon and ⁇ ⁇ FN , are used herein interchangeably. They refer to any interferon or interferon derivative ⁇ e.g., pegylated interferon) that can be used in the prevention or treatment of HCV infection and/or in the prevention or treatment of HCV-related diseases, in particular cirrhosis and liver cancer.
  • Interferons are a family of cytokines produced by eukaryotic cells in response to viral infection and other antigenic stimuli, which display broad-spectrum antiviral, antiproliferative and immunomodulatory effects.
  • Recombinant forms of interferons have been widely applied in the treatment of various conditions and diseases, such as viral infections ⁇ e.g., HCV, HBV and HIV), inflammatory disorders and diseases (e.g. , multiple sclerosis, arthritis, cystic fibrosis), and tumors (e.g. , liver cancer, lymphomas, myelomas, etc .).
  • the at least one interferon molecule present in a combination according to the invention is selected from the group consisting of IFN- a, IFN- ⁇ , IFN-CO, IFN- ⁇ , IFN- ⁇ , analogs thereof and derivatives thereof.
  • interferon and “IFN” more specifically refer to a peptide or protein having an amino acid substantially identical (e.g. , et least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or even 100% identical) to all or a portion of the sequence of an interferon (e.g., a human interferon), such as IFN- a, IFN- ⁇ , IFN-CO, IFN- ⁇ , and IFN- ⁇ that are known in the art.
  • an interferon e.g., a human interferon
  • Interferons suitable for use in the present invention include, but are not limited to, natural human interferons produced using human cells, recombinant human interferons produced from mammalian cells, E-co/i-produced recombinant human interferons, synthetic versions of human interferons and equivalents thereof.
  • Other suitable interferons include consensus interferons which are a type of synthetic interferons having an amino acid sequence that is a rough average of the sequence of all the known human IFN subtypes (for example, all the known IFN-a subtypes, or all the known IFN- ⁇ subtypes, or all the known IFN-CO subtypes, or all the known IFN- ⁇ subtypes, or all the known IFN- ⁇ subtypes.
  • interferons present in combinations according to the invention have been approved for human use. In other embodiments, interferons present in combinations according to the present are undergoing human clinical trials.
  • interferon and “IFN” also include interferon derivatives, i.e., molecules of interferon (as described above) that have been modified or transformed.
  • a suitable transformation may be any modification that imparts a desirable property to the interferon molecule. Examples of desirable properties include, but are not limited to, prolongation of in vivo half-life, improvement of therapeutic efficacy, decrease of dosing frequency, increase of solubility/water solubility, increase of resistance against proteolysis, facilitation of controlled release, and the like.
  • pegylated interferons have been produced (e.g. , pegylated IFN-a) and are currently used to treat hepatitis.
  • Pegylated interferons exhibit longer half-lifes, which allows for less frequent administration of the drug.
  • Pegylating an interferon molecule involves covalently binding the interferon to polyethylene glycol (PEG), an inert, nontoxic and biodegradable organic polymer. Therefore, in certain embodiments, the at least one interferon present in a combination according to the invention is a pegylated interferon.
  • Interferons have also been produced as fusion proteins with human albumin (e.g. , albumin-IFN-Cc).
  • the albumin-fusion platform takes advantage of the long half-life of human albumin to provide a treatment that allows the dosing frequency of IFN to be reduced in individuals with chronic hepatitis C.
  • the at least one interferon present in a combination according to the invention is an albumin-interferon fusion protein.
  • the terms "alpha interferon” , “interferon-alpha” , “interferon- of and “/FN- of are used herein interchangeably and refer to the family of highly homologous species- specific proteins (i.e. , glycoproteins) that are known in the art and inhibit viral replication and cellular proliferation, and modulate immune response.
  • IFN-a molecules suitable for use in the present invention include, but are not limited to, recombinant IFN-CC-2b (such as INTRON-A ® interferon available from Schering Corporation); recombinant IFN-CC-2a (such as ROFERON ® interferon available from Hoffman-La Roche); recombinant IFN-CC-2C (such as BEROFOR ® alpha 2 interferon available from Boehringer Ingelheim Pharmaceutical, Inc); IFN-a-nl, a purified blend of natural alpha interferons (such as SUMERIFERON ® available from Sumitomo, Japan or WELLFERON ® interferon alpha-nl (INS) available from Glaxo-Wellcome Ltd); IFN-a-n3, a mixture of natural alpha interferons (such as ALFERON ® made by Interferon Sciences); human leukocyte interferon-a obtained from the leukocyte fraction of human blood following induction with Sendai virus (such as MULTI
  • interferon alpha molecules include IFN-a derivatives, including, but not limited to, pegylated IFN-a-2a (such as PEGASYS ® available from Hoffman- La Roche); pegylated IFN-a-2b (such as PEGINTRON ® available from Schering Corporation); albumin IFN-a-2b also known as albinterferon (such as ALBUFERON ® available from Human Genome Sciences), and equivalents thereof.
  • pegylated IFN-a-2a such as PEGASYS ® available from Hoffman- La Roche
  • pegylated IFN-a-2b such as PEGINTRON ® available from Schering Corporation
  • albumin IFN-a-2b also known as albinterferon (such as ALBUFERON ® available from Human Genome Sciences), and equivalents thereof.
  • beta interferon refers to the family of highly homologous species-specific proteins (i.e., glycoproteins) that are known in the art and have the ability to induce resistance to viral antigens.
  • IFN- ⁇ molecules suitable for use in the present invention include, but are not limited to, recombinant ⁇ - ⁇ -la (such as, REBIF ® available from Pfizer or AVONEX ® available from Biogen Idex), recombinant ⁇ - ⁇ -lb (such as BETAFERON ® /BETASERON ® available from Bayer HealthCare or EXTAVIA ® , the generic form of BETAFERON, available from Novartis, or ZIFERON ® , an interferon- ⁇ lb biosimilar, available from Zistdaru Danesh Ltd), IFN- ⁇ molecules described in U.S. Pat. Nos. 4,820,638 and 5,795,779) and, equivalents thereof.
  • recombinant ⁇ - ⁇ -la such as, REBIF ® available from Pfizer or AVONEX ® available from Biogen Idex
  • recombinant ⁇ - ⁇ -lb such as BETAFERON ® /BETA
  • interferon beta molecules include IFN- ⁇ derivatives, including, but not limited to, pegylated INF- ⁇ (such as TRK-560 being developed by Toray Industries, Inc.), pegylated ⁇ - ⁇ -la (such as BUBO 17 being developed by Biogen pie); pegylated ⁇ - ⁇ -lb (such as NU100 and NU400 being developed by Nuron Biotech); albumin- IFN- ⁇ fusion proteins such as those described in U.S. Pat. No. 7,572,437, and equivalents thereof.
  • pegylated INF- ⁇ such as TRK-560 being developed by Toray Industries, Inc.
  • pegylated ⁇ - ⁇ -la such as BUBO 17 being developed by Biogen pie
  • pegylated ⁇ - ⁇ -lb such as NU100 and NU400 being developed by Nuron Biotech
  • albumin- IFN- ⁇ fusion proteins such as those described in U.S. Pat. No. 7,572,437, and equivalents thereof.
  • IFN-CO molecules suitable for use in the present invention include, but are not limited to, IFN-CO described in European patent No. EP0 170 204, ITCA being developed by Intarcia Therapeutics, Inc., and equivalents thereof.
  • Other suitable interferon omega molecules include IFN-CO derivatives, including, but not limited to, pegylated INF-CO that can be obtained using a method described in U.S. Pat. Nos. 5,612,460; 5,711,944; 5,951,974 or 5,951,974; albumin-IFN-CO fusion proteins such as those described in U.S. Pat. No. 7,572,437, and equivalents thereof.
  • interferon omega molecules include IFN- ⁇ derivatives, including, but not limited to, pegylated INF- ⁇ that can be obtained using a method described in U.S. Pat. Nos. 5,612,460; 5,711,944; 5,951,974 or 5,951,974 albumin- IFN-CO fusion proteins such as those described in U.S. Pat. No. 7,572,437, and equivalents thereof.
  • IFN- ⁇ are used herein interchangeably and refer to the family of highly homologous species-specific proteins (i.e., glycoproteins) that are known in the art and have antiviral properties.
  • Typical IFN- ⁇ molecules suitable for use in the present invention include, but are not limited to, IFN- ⁇ , ⁇ - ⁇ 2 and ⁇ - ⁇ 3 molecules described in international patent applications number WO02/086087, WO2004/037995 and WO/2005/023862 and equivalents thereof.
  • interferon omega molecules include IFN- ⁇ derivatives including, but not limited to, pegylated ⁇ - ⁇ -la (such as BMS-914143 being developed by Bristol-Myers Squibb), albumin- IFN-CO fusion proteins such as those described in U.S. Pat. No. 7,572,437, and equivalents thereof.
  • pegylated ⁇ - ⁇ -la such as BMS-914143 being developed by Bristol-Myers Squibb
  • albumin- IFN-CO fusion proteins such as those described in U.S. Pat. No. 7,572,437, and equivalents thereof.
  • interferon and “/ N' also include interferon-like molecules, i.e., molecules that have functional and/or structural features exhibited by or similar to known interferons or interferon analogs, such as those described above.
  • the invention provides a combination comprising at least one anti-SR- BI monoclonal antibody according to the invention and the host-targeting agent, alisporivir.
  • Alisporivir also known as Debia 0.25, DEB025 or UNIL-025
  • Debia 0.25, DEB025 or UNIL-025 is a cyclophilin A inhibitor. It is under development by Debiopharm for Japan and by Novartis for the rest of the world since February 2010. It is being researched for potential use in the treatment of hepatitis C (Flisiak et ah, Hepatology, 2009, 49: 1460-1468), and also investigated for Duchenne muscular dystrophy (Reutenauer et al, Br. J. Pharmacol., 2008, 155: 574-584).
  • the HCV infection or HCV-related disease to be treated by a combination according the invention is caused by a Hepatitis C virus that is resistant to a direct acting antiviral and/or that is transmitted by cell-cell transmission.
  • the invention also provides pharmaceutical compositions comprising an effective amount of at least one combination of the invention and at least one pharmaceutically acceptable carrier or excipient, as described above.
  • the present invention provides a pharmaceutical pack or kit comprising one or more containers (e.g. , vials, ampoules, test tubes, flasks or bottles) containing one or more ingredients of an inventive pharmaceutical composition, allowing administration of an anti-SR-BI monoclonal antibody of the present invention.
  • containers e.g. , vials, ampoules, test tubes, flasks or bottles
  • a pharmaceutical pack or kit may be supplied in a solid (e.g. , lyophilized) or liquid form. Each ingredient will generally be suitable as aliquoted in its respective container or provided in a concentrated form. Pharmaceutical packs or kits may include media for the reconstitution of lyophilized ingredients. Individual containers of the kits will preferably be maintained in close confinement for commercial sale.
  • a pharmaceutical pack or kit includes one or more additional therapeutic agent(s) (e.g., one or more anti-viral agents, as described above).
  • Optionally associated with the container(s) can be a notice or package insert in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceutical or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration.
  • the notice of package insert may contain instructions for use of a pharmaceutical composition according to methods of treatment disclosed herein.
  • An identifier e.g. , a bar code, radio frequency, ID tags, etc.
  • the identifier can be used, for example, to uniquely identify the kit for purposes of quality control, inventory control, tracking movement between workstations, etc.
  • Antibodies of the present invention may be employed in a variety of non-therapeutic applications, such as purification and screening methods.
  • antibodies of the invention may be used as affinity purification agents.
  • an inventive antibody is immobilized on a solid phase such as Sephadex resin or filter paper, using methods well known in the art.
  • the immobilized antibody is contacted with a sample containing human SR-BI (or a fragment thereof) to be purified, and thereafter the support is washed with a suitable solvent that will remove substantially all the material in the sample except the SR-BI protein, which is bound to the immobilized antibody. Finally, the support is washed with another suitable solvent which will release the SR-BI protein from the antibody.
  • Anti-SR-BI monoclonal antibodies of the present invention may also be used in drug screening methods based on competitive binding assays. Such methods may involve the steps of allowing competitive binding between a test compound (e.g. , a test antibody) in a sample and a known amount of an inventive anti-SR-BI monoclonal antibody, for binding to cells to which the inventive antibody binds, and measuring the amount of the known monoclonal antibody bound.
  • a test compound e.g. , a test antibody
  • inventive monoclonal antibody is appropriately labeled, for example, with an enzymatic, chemiluminescent, or fluorescent label. Examples
  • HEK293T Chinese hamster ovary (CHO), Buffalo Rat Liver (BRL3A), Huh7, Huh7.5-GFP and Huh7.5.1 cells were cultured as described (Krieger et al., Hepatology, 2010, 51: 1144-1157; Pestka et al, Proc. Natl. Acad. Sci. USA, 2007, 104: 6025-6030; Dreux et al, PLoS Pathog., 2009, 5:el00031; Witteveldt et al, J. Gen. Virol., 2009, 90: 48-58).
  • Primary human hepatocytes were isolated as previously described (Krieger et al, Hepatology, 2010, 51: 1144-1157).
  • CHO and BRL3A cells expressing SR-BI were produced as described (Zeisel et al, Hepatology, 2007, 46: 1722-1731; Dreux et al, PLoS Pathog., 2009, 5:el00031).
  • Antibodies Polyclonal (Zeisel et al, Hepatology, 2007, 46: 1722-1731) and monoclonal antibodies (mAbs) directed against the extracellular loop of SR-BI were raised by genetic immunization of Wistar rats and Balb/c mice as described (Zeisel et al, Hepatology, 2007, 46: 1722-1731) according to proprietary technology (Aldevron GmbH, Freiburg, Germany). Anti-SR-BI mAbs were purified using protein G and selected by flow cytometry for their ability to bind to human SR-BI (Zeisel et al, Hepatology, 2007, 46: 1722-1731).
  • Huh7.5.1 cells were incubated with increasing concentrations of mAbs and binding was assessed using flow cytometry. Kd values were determined as half- saturating concentrations of the mAbs using SigmaPlot.
  • Anti-CD81 (JS-81), anti-SR-BI (CLA-1) and phycoerythrin (PE)-conjugated anti-mouse antibodies were from BD Biosciences.
  • Anti-His and FITC-conjugated anti-His antibodies were from Qiagen and rabbit anti-actin (AA20-30) antibodies from Sigma- Aldrich.
  • Anti-El Innogenetics
  • anti-E2 IGH461, Innogenetics
  • AP33 Genentech
  • CBH23 a kind gift from S. K. H. Foung
  • patient-derived anti-HCV IgG have been described (Haberstroh et al, Gastroenterology, 2008, 135: 1719-1728; Fofana et al, Gastroenterology, 2010, 39: 953-964; Fofana et al, Gastroenterology, 2012, 143: 223- 233).
  • HCVcc Cell Culture-derived HCV
  • HCVpp HCVpp
  • MLVpp MLVpp
  • VSV-Gpp VSV-Gpp
  • infection and kinetic experiments have been described (Zeisel et al, Hepatology, 2007, 46: 1722-1731; Harris et al, J. Virol., 2008, 82: 5007-5020; Pestka et al, Proc. Natl. Acad. Sci. USA, 2007, 104: 6025-6030; Fofana et al, Gastroenterology, 2010, 39: 953-964; Bartosch et al, J. Exp. Med., 2003, 197: 633-642).
  • HCVpp of genotypes 1-6 have been described (Lupberger et al, Nature Medicine, 2011, 17: 589- 595; Fofana et al, Gastroenterology, 2012, 143: 223-233).
  • Chimeric HCVcc of genotypes 1-4 have been described (Koutsoudakis et al, J. Virol., 2006, 80: 5308- 5320).
  • HCVcc experiments were performed using Luc-Jcl and infection was analyzed in cell lysates by quantification of luciferase activity (Koutsoudakis et al, J. Virol., 2006, 80: 5308-5320). For combination experiments, each antibody was tested individually or in combination with a second antibody.
  • Huh7.5.1 cells were pre-incubated with anti-SR-BI or control mAb for 1 hour and then incubated for 4 hours at 37 °C with HCVcc or HCVpp (P02VJ) (pre-incubated for 1 hour with or without anti-envelope antibodies).
  • Synergy was assessed using the combination index and the method of Prichard and Shipman (Zhu et al, J. Infect. Dis., 2012, 205: 656-662; Prichard et al, Antiviral Res., 1990, 14: 181-205).
  • Cell viability was assessed using a MTT test (Lupberger et al, Nature Medicine, 2011, 17: 589- 595). Cellular Binding of Envelope Glycoprotein E2.
  • sE2 His-tagged soluble E2
  • Huh7.5.1 cells were pre-incubated with control or anti-SR-BI serum (1:50), anti-SR-BI or control mAbs (20 ⁇ g/mL) for 1 hour at room temperature (RT) and then incubated with sE2 for 1 hour at RT. Binding of sE2 was revealed using flow cytometry as described (Krieger et al, Hepatology, 2010, 51: 1144-1157; Dreux et al, PLoS Pathog., 2009, 5:el00031).
  • Huh7.5.1 cells were pre-incubated with heparin (100 ⁇ g/mL), control or anti-SR-BI serum (1:50), anti-SR-BI or control mAbs (20 ⁇ g/mL) for 1 hour at 37°C prior to incubation with HCVcc as described (Krieger et al, Hepatology, 2010, 51: 1144-1157; Dreux et al, PLoS Pathog., 2009, 5:el00031).
  • Non-bound HCVcc were removed by washing of cells with PBS and cell bound HCV RNA was then quantified by RT-PCR (Krieger et al, Hepatology, 2010, 51: 1144-1157; Dreux et al, PLoS Pathog., 2009, 5:el00031).
  • HCV Cell-to-cell Transmission HCV cell-to-cell transmission was assessed as described (Lupberger et al., Nature Medicine, 2011, 17: 589-595; Witteveldt et al., J. Gen. Virol., 2009, 90: 48-58).
  • Producer Huh7.5.1 cells were electroporated with Jcl RNA (Pietschmann et al., Proc. Natl. Acad.
  • HCV E2-neutralizing antibody AP33, 25 ⁇ g/mL was added to block cell-free transmission (Witteveldt et al, J. Gen. Virol., 2009, 90: 48-58). After 24 hours of co-culture, cells were fixed with paraformaldehyde, stained with an NS5A-specific antibody and analyzed by flow cytometry (Lupberger et al., Nature Medicine, 2011, 17: 589-595; Witteveldt et al, J. Gen. Virol., 2009, 90: 48-58). Immunofluorescence. Cell spread was assessed by visualizing Jcl -infected
  • HDL Binding HDL was labeled using Amersham Cy5 Mono-Reactive Dye Pack (GE Healthcare). Unbound Cy5 was removed by applying labeled HDL on illustra MicroSpin G-25 Columns (GE Healthcare). Blocking of Cy5-HDL binding with indicated reagents was performed for 1 hour at RT prior to Cy5-HDL binding for 1 hour at 4°C on 10 6 target cells.
  • Lipid Transfer Assays Selective HDL-CE uptake and lipid efflux assays were performed as described (Dreux et ah, PLoS Pathog., 2009, 5:el00031; Le Goff et ah, J. Lipid Res., 2006, 47: 51-58).
  • HDL-CE uptake was assessed in the presence or absence of anti-SR-BI mAbs (20 ⁇ g/mL) and 3 H-CE-labelled HDL (60 ⁇ g protein) for 5 hours at 37°C. Selective uptake was calculated from the known specific radioactivity of radiolabeled HDL-CE and is denoted in ⁇ g HDL-CE ⁇ g cell protein.
  • Huh7 cells were labeled with H-cholesterol (1 ⁇ / ⁇ ⁇ ) and incubated at 37°C for 48 hours as described (Dreux et ah, PLoS Pathog., 2009, 5:el00031; de la Llera Moya et al, Arterioscler. Thromb.
  • the Applicants then aimed at characterizing the viral entry steps targeted by these anti-SR-BI mAbs. They first assessed the ability of these mAbs to interfere with viral binding. To reflect the complex interaction between HCV and hSR-BI during viral binding, they studied the effect of anti-SR-BI mAbs on HCVcc binding to Huh7.5.1 cells at 4°C. Incubation of Huh7.5.1 cells with anti-SR-BI mAbs prior to and during HCVcc binding did not inhibit virus particle binding (Figure 2A). These data suggest that, in contrast to previously described anti-SR-BI mAbs, these novel anti-SR-BI mAbs do not inhibit HCV binding but interfere with HCV entry during post-binding steps.
  • the Applicants assessed HCVcc entry kinetics into Huh7.5.1 cells in the presence of anti-SR-BI mAbs inhibiting HCV infection (QQ-4A3-A1, QQ-2A10-A5 and QQ-4G9-A6 and NK-8H5-E3) added at different time-points during or after viral binding (Figure 2(B)).
  • This assays was performed side-by-side with an anti-CD81 mAb inhibiting HCV post-binding (Zeisel et al, Hepatology, 2007, 46: 1722-1731; Krieger et al, Hepatology, 2010, 51: 1144-1157; Koutsoudakis et al, J. Virol., 2006, 80: 5308-5320) and proteinase K (Schwartz et al, J. Virol., 2009, 83: 12407-12414) to remove HCV from the cell surface.
  • HCVcc binding to Huh7.5 cells was performed for 1 hour at 4°C in the presence or absence of compounds.
  • these antibodies are the first molecules exclusively targeting the post-binding function of SR-BI and thus represent a unique tool to more thoroughly assess the relevance of this function for HCV infection.
  • a Post-binding Function of SR-BI is Essential for Cell-to-cell Transmission and Viral Spread. HCV disseminates via direct cell-to-cell transmission (Witteveldt et al, J. Gen. Virol., 2009, 90: 48-58; Brimacombe et al, J. Virol., 2011, 85: 596- 605).
  • the Applicants first investigated the ability of the anti-SR-BI mAbs to interfere with neutralizing antibody-resistant viral spread by studying direct HCV cell-to-cell transmission in the presence of anti-SR-BI mAbs QQ-2A10-A5 and QQ-4G9-A6.
  • Viral "producer” cells containing replicating HCV Jcl (Pi) are co-cultured with GFP- expressing "target” cells (T) in the presence of E2-neutralizing mAb (AP33, 25 ⁇ g/mL) to prevent cell-free HCV transmission (Witteveldt et ah, J. Gen. Virol., 2009, 90: 48-58), AP33 reduces cell-free transmission by >90 and infectivity of producer cell supernatants is minimal at the time of co-culture; viral transmission thus occurs predominantly by cell-to-cell transmission (Lupberger et ah, Nature Medicine, 2011, 17: 589-595).
  • HCV cell-to-cell transmission was assessed by quantifying HCV- infected, GFP-positive target cells (Ti) by flow cytometry (Lupberger et ah, Nature Medicine, 2011, 17: 589-595).
  • Both anti-SR-BI mAbs (10 ⁇ g/mL) efficiently blocked HCV cell-to-cell transmission ( Figure 3(A), Figure 4(A)-(B)) indicating that these antibodies may prevent viral spread in vitro.
  • these anti-SR-BI mAbs do not block HCV-SR-BI binding ( Figure 2(A)) but inhibit HCV entry during post-binding steps ( Figure 2C), these data suggest that a SR-BI post-binding function plays an important role during HCV cell-to-cell transmission.
  • the Applicants thus used Huh7.5 cells or Huh7.5 cells overexpressing either mSR-BI, which is unable to bind E2, or hSR-BI, which is able to bind E2, as target cells.
  • Cell-to-cell transmission was enhanced in Huh7.5 cells overexpressing either hSR-BI (2.04 + 0.03 fold overexpression) or mSR-BI (1.92 + 0.19 fold overexpression) as compared to parental cells ( Figure 3(B)).
  • SR-BI Determinants Relevant for HCV Post-Binding Steps May be Linked to the Lipid Transfer Function of the Entry Factor.
  • the SR-BI ectodomain has been demonstrated to be important for both HDL binding and CE uptake but the determinants involved in these processes have not been precisely defined yet.
  • anti-SR-BI mAbs inhibiting HCV post-binding steps affect HDL binding to SR-BI, the Applicants studied Cy5-labeled HDL binding to hSR-BI in the presence or absence of anti-SR-BI mAbs.
  • the Applicants also assessed the ability of the anti-SR-BI mAbs to bind to SR-BI mutants reported to modulate lipid transfer (Dreux et ah, PLoS Pathog., 2009, 5:el00031). Taken together, these data suggest that SR-BI determinants involved in HCV post-binding events do not mediate HDL binding but may contribute to lipid transfer, in line with the reported link between the SR-BI lipid transfer function and HCV infection (Bartosch et al, J. Virol, 2005, 79: 8217-8229; Dreux et al, PLoS Pathog., 2009, 5:el00031).
  • Combination of anti-SR-BI and anti- HCV envelope antibodies resulted in a synergistic effect on inhibition of HCVpp P02VJ entry and HCVcc infection as assessed using the combination index (combination index of 0.06-0.67) and the method of Prichard and Shipman (Zhu et al., J. Infect. Dis., 2012, 205: 656-662; Prichard et al., Antiviral Res., 1990, 14: 181-205).
  • These combinations reduced the IC 50 of anti-SR-BI mAb by up to 100-fold ( Figure 8(A)-(D)).
  • the marked synergy may be explained by the fact that the E2- and SR-BI- specific antibodies target highly complementary steps during HCV entry. Taken together, these data indicate that interfering with SR-BI post-binding function may hold promise for the design of novel antiviral strategies targeting HCV entry factors.
  • the Applicants investigated their ability to bind to human-mouse SR-BI chimeras, where part of the mouse SR-BI ectodomain was replaced by the corresponding human sequence. While the HHH and MMM SR-BI constructs refer to the wild- type human (H) and mouse (M) SR-BI molecules, respectively, the human/mouse SR-BI chimeras were denominated according to the origin of either SR-BI region, e.g., HMM bears region 1 from human SR-BI and regions 2 and 3 from murine SR-BI ( Figure 10(A)). The overall homology between human and mouse SR-BI is 80% (54 aa difference).
  • the three rat anti-SR-BI mAbs QQ- 4A3-A1, QQ-2A10-A5, QQ-4G9-A6 bind to HMM SR-BI, i.e. aa 38-215, with high affinity and also to MHM, i. e. 216-398, to a lesser extent while the mouse mAb NK- 8H5-E3 only recognizes HMM SR-BI with high affinity ( Figure 10(B)).
  • the Applicants have generated novel anti-SR-BI mAbs specifically inhibiting HCV entry during post-binding steps that enabled them for the first time, using endogenous SR-BI, to explore and validate the hypothesis that SR-BI has a multifunctional role during HCV entry and to elucidate the functional role of SR-BI post-binding activity for HCV infection.
  • the present data demonstrate that the HCV post-binding function of hSR-BI can indeed be dissociated from its E2-binding function.
  • the post-binding activity of SR- BI is of key relevance for cell-free HCV infection as well as cell-to-cell transmission.
  • SR-BI mediates uptake of HDL-CE in a two-step process including HDL binding and subsequent transfer of CE into the cell without internalization of HDL.
  • SR-BI also participates in HCV binding and entry into target cells.
  • SR-BI is able to directly bind E2 and virus-associated lipoproteins but additional function(s) of SR-BI have been reported to be at play during HCV infection (Bartosch et al, J. Virol., 2005, 79: 8217-8229; Zeisel et al, Hepatology, 2007, 46: 1722-1731; Dreux et al, PLoS Pathog., 2009, 5:el00031).
  • the results from this study highlight the importance of a SR-BI post-binding function for HCV entry and further extend the relevance of this function for HCV cell-to-cell transmission.
  • novel anti-SR-BI mAbs are the first anti-SR-BI mAbs that do not inhibit HDL binding to SR-BI. These data suggest that HCV entry and dissemination can be inhibited without blocking HDL-SR-BI binding. The further characterization of the SR-BI post-binding function will allow to determine whether the SR-BI-mediated post-binding steps of HCV entry and dissemination are directly linked to its lipid transfer function.
  • the present data suggest that the SR-BI post-binding function is a highly relevant target for antivirals.
  • Therapeutic options for a large proportion of HCV- infected patients are still limited by drug resistance and adverse effects (Pawlotsky, Hepatology, 2011, 53: 1742-1751).
  • a strategy for prevention of HCV liver graft infection is absent.
  • Antivirals targeting essential host factors required for the HCV life cycle are attractive since they may increase the genetic barrier for antiviral resistance (Lupberger et al., Nature Medicine, 2011, 17: 589-595; Zeisel et al., J. Hepatol., 2011, 54: 566-576).
  • a human anti-SR-BI mAb has been reported to inhibit HDL binding, to interfere with cholesterol efflux and to decrease HCVcc entry during attachment steps without having a relevant impact on SR-BI mediated post-binding steps (Catanese et al, J. Virol., 2010, 84: 34-43; Catanese et al, J. Virol., 2007, 81: 8063-8071).
  • a codon-optimized version of this mAb has been demonstrated to prevent HCV spread in vivo (Meuleman et al, Hepatology, 2012, 55: 364-372) underscoring the potential of SR-BI as an antiviral target.
  • the mAbs generated in the present study are highly novel in their function as they do not interfere with HCV-SR-BI binding but inhibit HCV entry during post-binding steps of cell-free infection and cell-to-cell transmission. Furthermore, in contrast to previously described anti-SR-BI mAbs (Catanese et al, J. Virol., 2007, 81: 8063-8071), these mAbs do not hinder HDL-SR- BI binding and only partially inhibit lipid transfer at concentrations significantly inhibiting HCV infection.
  • QQ-4A3-A1, QQ-2A10-A5, QQ-4G9-A6 and NK-8H5- E3 define a novel class of anti-SR-BI mAbs for prevention and treatment of HCV infection.
  • Example 2 Synergistic Effects of Combinations Comprising an anti-SR-BI mAb
  • Anti-SR-BI (NK-8H5-E3, QQ-4G9- A6 and QQ-A43-A1) mAbs were used in this series of experiments. Erlotinib and dasatinib were obtained from IC laboratories.
  • the Cyclophilin A inhibitor (alisporivir), protease inhibitors (telaprevir, boceprevir, danoprevir and TMC-435), NS5A inhibitor (daclatasvir) and polymerase inhibitors (mericitabine and GS-7977 (formally known as PSI-7977)) were synthesized by Acme Bioscience, Inc.
  • Anti- CLDN1 mAbs (OM-7D3-B3, OM-8A9-A4 and OM-6E1-B5) were used. Analysis of Antiviral Activity of Compounds and Combinations on HCV Infection. The in vitro antiviral activity of each compound was tested individually and in combination with a second compound using the HCVcc Huh7.5.1 cell culture described (Lupberger et al, Nature Medicine, 2011, 17: 589-595; Zhong et al., Proc. Natl. Acad. Sci. USA, 2005, 102: 9294-9299; Koutsoudakis et al., J. Virol., 2006, 80: 5308-5320).
  • HCVcc (Luc-Jcl; TCID 50 approximately 10 4 /ml) has been described (Koutsoudakis et al., J. Virol., 2006, 80: 5308-5320).
  • entry inhibitor anti-SRBI mAb
  • IFN-a DAAs
  • DAAs telaprevir, boceprevir, danoprevir, TMC-435, daclatasvir, mericitabine and GS-7977
  • Huh7.5.1 cells (culture in 96-well-plates) were pre-incubated with IFN-a, DAAs or alisporivir and the anti-SR-BI mAb for 1 hour at 37 °C before incubation for 4 hours at 37 °C with HCVcc in the presence of both compounds.
  • Huh7.5.1 cells were pre-incubated with both entry inhibitiors or control reagent for 1 hour at 37°C. The mix was removed and Huh7.5.1 cells were incubated for 4 hours at 37°C with HCVcc in the presence of both compounds.
  • HCVcc infection was analyzed two days later by lucif erase reporter gene expression as previously described (Krieger et al., Hepatology, 2010, 54: 1144-1157; Fofana et al., Gastroenterology, 2010, 139: 953-964; Koutsoudakis et al., J. Virol., 2006, 80: 5308-5320).
  • Synergy was assessed by two independent methods: the combination index (Fofana et al., Gastroenterology, 210, 139: 953-964; Koutsoudakis et al, J. Virol., 2006, 80: 5308-5320) and the method of Prichard and Shipman (Zhao et al, Clin. Cancer Res., 2004, 10: 7994-8004; Prichard et al, Antiviral Res., 1990, 14: 181-205).
  • a CI of less than 0.9 indicates synergy; a CI equal to 0.9-1.1 indicates additivity; and a CI of more than 1.1 indicates antagonism (Zhao et al, Clin.
  • Huh7.5.1 cells and primary human hepatocytes isolated and cultured as described were incubated with the compounds for 48 hours (Krieger et ah, Hepatology, 2010, 51: 1144-1157). Cytotoxic effects were analysed by the ability to metabolize 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) as described (Lupberger et al, Nat. Med., 2011, 17: 589-595). An anti-Fas antibody (10 g/m) was used as a positive control.
  • Anti-SR-BI mAb potentiates the antiviral activity of interferon-alpha in a synergistic manner. Since IFN-a is the key component of standard-of-care, the Applicants investigated whether the anti-SR-BI mAb could potentiate the antiviral activity of IFN-a by investigating the effect of combining the anti-SR-BI mAb with IFN-a2a or IFN-a2b on HCVcc infection. The antiviral effect of each molecule was tested alone or in combination to determine the combination index (CI) ( Figure 12(A- B)).
  • TMC-435 0.013 ⁇ 0.001 anti-SRBI 0.006 ⁇ 0.0007 0.4910.
  • danoprevir 0.006 ⁇ 0.003 anti-SRBI 0.0007 ⁇ 0.001 0.1510.02 daclatasvir 0.012 ⁇ 0.003 anti-SRBI 0.000210.0004 0.0310.004 mericitabine 0.12 ⁇ 0.03 anti-SRBI 0.01910.007 0.1710.
  • Second-generation protease inhibitors have been demonstrated to have a higher genetic barrier for resistance. However, single amino acid substitutions are able to confer drug resistance in vivo. Importanly, it has been demonstrated that several telaprevir- and boceprevir-resistance mutations confer cross-resistance to these second-generation protease inhibitors (Sarrazin et ah, J. Hepatol., 2012, 56(1): S88- 100).
  • GS-7977 is currenlty in clinical development and has been suggested as having the potential to become the conerstone of an efficacious, all-oral combination regimen for many patients with chronic HCV infection (Zeisel et ah, Front Biosci., 2009, 14: 3274-3285; Zeisel et ah, J. Hepatol., 2011, 54: 566-576).
  • the inventors investigated whether the anti-SR-BI mAb potentiates the antiviral activity of GS-7977.
  • HTAs host-targeting agents
  • Example 3 Effects of Monoclonal Anti-SR-BI Antibodies on Viruses Resistant to Direct-Acting Antivirals (DAAs)
  • Antiviral resistance remains a major challenge for treatment of chronic HCV infections.
  • the functional role of viral dissemination for emergence and maintenance of antiviral resistance is largely unknown.
  • HCV is transmitted via cell-free diffusion but also uses direct cell-cell transfer to infect neighboring cells (Meredith et ah, J. Hepatol, 2013, 53: 1074-1080; Timpe et al, Hepatology, 2008, 47: 17-24). While cell-free entry is most relevant for initiation of HCV infection, HCV neutralizing antibody-resistance cell-cell transmission is thought to play an important role in viral persistence (Zeisel et al, J. Hepatol., 2013, 58: 375-384).
  • HTEIs host-targeting entry inhibitors
  • Anti-SR-BI NK-8H5-E3 mAb was used in this study. Erlotinib was obtained from IC laboratories. Anti-E2 mAb (AP33, Genetech) and human anti-HCV IgG have been described (Witteveldt et al, J. Gen. Virol., 2009, 90: 48-58; Fofana et al, Gastroenterology, 2012, 143: 223-233). Mouse IgG was purchased from BD. NS 5 -A- specific mAb was obtained from Virostat. Inhibitors of HCV protease (telaprevir, boceprevir and simeprevir) and HCV NS5A (daclatasvir) were synthesized by Acme Bioscience, Inc..
  • HCVcc J4 (genotype 2a/lb) and HCVcc J4 NS5A-Y93H (Y2065H) have been described (Scheel et al., Gastroenterology, 2011, 140: 1032- 1042).
  • HCVcc (TCIDso approximately 10 3 /ml to 10 4 /ml) were produced as previously described (Lupberger et al, Nature Med., 2011, 17: 589-595).
  • Huh7.5.1 cells were pre-incubated with serial concentrations of protease inhibitors (telaprevir, boceprevir), NS5A inhibitor (daclatasvir), anti-SR-BI monoclonal antibody or control reagents for 1 hour at 37°C before incubation for 4 hours at 37°C with wild-type or chimeric HCVcc.
  • protease inhibitors telaprevir, boceprevir
  • NS5A inhibitor daclatasvir
  • anti-SR-BI monoclonal antibody or control reagents for 1 hour at 37°C before incubation for 4 hours at 37°C with wild-type or chimeric HCVcc.
  • Viral infection was analysed by assessing the intracellular luciferase activity (Lupberger et al, Nature Med., 2011, 17: 589-595; Fofana et al, Gastroenterology, 2010, 139: 953-964) or intracellular HCV RNA levels as previous described (Lupberger et al, Nature Med., 2011, 17: 589-595; Fofana et al, Gastroenterology, 2010, 139: 953-964; Zeisel et al, Hepatology, 2007, 46: 1722-1731) Absent HCV RNA quantification by RT-PCR was confirmed using Abbott RealTime HCV assay (LOD 48 IU/ml). Statistical Analysis. Unless otherwise stated, results are expressed as means + standard deviation (SD) from at least 3 independent experiments performed in triplicate. Statistical analyses were performed using Student t test.
  • SD standard deviation
  • DAA-resistant Viruses and their Sensitivity to Monoclonal Anti-SR-BI antibody.
  • the Applicants generated DAA-resistant viruses and assessed their infectivity in state-of-the-art infection models. They first used classical Huh7.5.1 cell infection assays and characterized the ability of DAA-resistant viruses to infect hepatoma cells and their sensitivity to DAAs and HTEIs.
  • HCVcc cell culture- derived HCV
  • Luc-Jcl gene 2a/2a
  • two mutations at positions 155 and 156 of the HCV NS3 protein Sarrazin et al, J. Hepatol., 2012, 56(Suppl. 1): S88-100; Hiraga et al, Hepatology, 2011, 54: 781-788, known to induce cross-resistance in vitro and in vivo to protease inhibitors (teleprevir and boceprevir).
  • Mutation Y2065H confers resistance to daclatasvir, an HCV HS5A inhibitor, among recombinant HCV variants with NS5A from genotypes 1-7 (30).
  • mutation NS5A- Y93H in HCVcc J4 increased the IC 50 of daclatasvir up to 10-fold.
  • the IC 50 of CLDN1 -specific mAb to J4 NS5A-Y93H remained unchanged (Fig. 2B).
  • HCV dissemination within the liver and establishment of chronic HCV infection may mainly occur by direct cell-cell transmission between adjacent hepatocytes (Timpe et al, Hepatology, 2008, 47: 17- 24).
  • Functional results obtained in cell culture and animal models demonstrated strong evidence that cell-cell transmission plays a relevant role in dissemination of several viruses including HBV, HIV, herpes simplex virus (HSV), measles virus or human T-lymphotropic virus type 1 (HTLV-1) (Sattentau et al, Nat. Rev.
  • mice persistently infected with HCVcc Jcl (genotype 2a/2a).
  • HCV Jcl -infected mice persistently infected for 24 to 50 days, received 4 weekly antibody doses of SR-BI-specific or control mAb.
  • the SR-BI- specific mAb-treated mouse showed a rapid decline in viral load with undetectable HCV RNA after 2 injections (Fig. 18(A)) that were sustained for at least 6 weeks.
  • Human albumin levels remained stable during and post antibody administration and were higher in the SR-BI mAb-treated mouse than in the control mAb-treated mouse following antibody-treatment (Fig. 18(B)), indicating the presence of viable and functional hepatocytes following mAb treatment and exclude that the antiviral activity was due to a toxic effect on hepatocytes.

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Abstract

La présente invention concerne des anticorps monoclonaux qui se lient spécifiquement à la moitié N-terminal du domaine extracellulaire de SR-BI humain sur la surface cellulaire, inhibant ainsi l'entrée de VHC dans des cellules prédisposées au cours des étapes post-liaison et prévenant une infection par le VHC de ces cellules ; et des lignées cellulaires d'hybridome qui produisent de tels anticorps monoclonaux. L'invention concerne également des réactifs qui comprennent de tels anticorps, des combinaisons qui comprennent de tels anticorps et des compositions pharmaceutiques comprenant de tels anticorps. L'invention concerne également des méthodes de traitement ou de prévention d'une infection par le VHC par l'administration d'un anticorps monoclonal de l'invention ou d'une composition pharmaceutique associée.
PCT/EP2013/068008 2012-08-31 2013-08-30 Anticorps anti-sr-bi pour inhiber une infection par le virus de l'hépatite c WO2014033266A1 (fr)

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EP3224276A4 (fr) * 2014-11-25 2018-05-02 Rappaport Family Institute for Research in the Medical Sciences Nouvel épitope en tant que cible pour la thérapie de maladies inflammatoires auto-immunes et du rejet de greffe
US10975122B2 (en) 2014-11-25 2021-04-13 Technion Research & Development Foundation Limited Epitope as a target for therapy of inflammatory autoimmune diseases and graft rejection

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