Methods For Treating Diseases and HCV Using Antibodies To Aminophospholipids

Abstract

Disclosed are surprising discoveries concerning the role of anionic phospholipids and aminophospholipids in tumor vasculature and in viral entry and spread, and compositions and methods for utilizing these findings in the treatment of cancer and viral infections. Also disclosed are advantageous antibody, immunoconjugate and duramycin-based compositions and combinations that bind and inhibit anionic phospholipids and aminophospholipids, for use in the safe and effective treatment of cancer, viral infections and related diseases.

1 - 31 . (canceled) 
     
     
         32 . A method of quantifying viral load in a biological sample, comprising contacting said biological sample with a first antibody, or an antigen-binding fragment thereof, in an amount effective to bind viruses in said biological sample and quantifying the bound viruses; wherein said first antibody binds to phosphatidylserine on the luminal surface of tumor vascular endothelial cells when administered to an animal with a solid tumor and wherein said first antibody exhibits significant binding to an ELISA plate coated with phosphatidylserine in an ELISA conducted in the presence of serum, but no detectable binding to an ELISA plate coated with phosphatidylcholine in an ELISA conducted in the presence of serum, wherein said ELISA conducted in the presence of serum comprises the steps of
 (a) coating a first ELISA plate with phosphatidylserine to prepare a PS-coated ELISA plate and coating a second ELISA plate with phosphatidylcholine to prepare a PC-coated ELISA plate;   (b) blocking said PS-coated ELISA plate and said PC-coated ELISA plate with a blocking buffer comprising 10% serum;   (c) adding said first antibody diluted in said blocking buffer to said PS-coated ELISA plate and said PC-coated ELISA plate under conditions effective to allow binding of said first antibody to said PS-coated ELISA plate and said PC-coated ELISA plate in the presence of serum; and   (d) detecting the binding of said first antibody to said PS-coated ELISA plate and said PC-coated ELISA plate in the presence of serum using a secondary antibody that binds to said first antibody; wherein said first antibody exhibits significant binding to said PS-coated ELISA plate, but no detectable binding to said PC-coated ELISA plate.   
     
     
         33 . The method of  claim 32 , wherein said first antibody effectively competes with the 3G4 antibody (monoclonal antibody 3G4 deposited as ATCC PTA 4545) in binding to an ELISA plate coated with phosphatidylserine in a competition ELISA conducted in the presence of serum, wherein said competition ELISA comprises:
 (a) coating an ELISA plate with phosphatidylserine to prepare a PS-coated ELISA plate;   (b) blocking said PS-coated ELISA plate with a blocking buffer comprising 10% serum;   (c) adding said 3G4 antibody diluted in said blocking buffer to said PS-coated ELISA plate under conditions effective to allow binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum;   (d) detecting the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum using a secondary antibody that binds to said 3G4 antibody; and   (e) identifying a first antibody that effectively competes with said 3G4 antibody in said competition ELISA by selecting a first antibody that substantially reduces the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum.   
     
     
         34 . The method of  claim 32 , wherein said antibody is bound to a solid support. 
     
     
         35 . The method of  claim 32 , wherein said biological sample is a blood sample. 
     
     
         36 . A method of purging a virus from a biological sample, comprising contacting said biological sample with a first antibody, or an antigen-binding fragment thereof, in an amount effective to bind and remove said virus from said biological sample; wherein said first antibody binds to phosphatidylserine on the luminal surface of tumor vascular endothelial cells when administered to an animal with a solid tumor and wherein said first antibody exhibits significant binding to an ELISA plate coated with phosphatidylserine in an ELISA conducted in the presence of serum, but no detectable binding to an ELISA plate coated with phosphatidylcholine in an ELISA conducted in the presence of serum, wherein said ELISA conducted in the presence of serum comprises the steps of:
 (a) coating a first ELISA plate with phosphatidylserine to prepare a PS-coated ELISA plate and coating a second ELISA plate with phosphatidylcholine to prepare a PC-coated ELISA plate;   (b) blocking said PS-coated ELISA plate and said PC-coated ELISA plate with a blocking buffer comprising 10% serum;   (c) adding said first antibody diluted in said blocking buffer to said PS-coated ELISA plate and said PC-coated ELISA plate under conditions effective to allow binding of said first antibody to said PS-coated ELISA plate and said PC-coated ELISA plate in the presence of serum; and   (d) detecting the binding of said first antibody to said PS-coated ELISA plate and said PC-coated ELISA plate in the presence of serum using a secondary antibody that binds to said first antibody; wherein said first antibody exhibits significant binding to said PS-coated ELISA plate, but no detectable binding to said PC-coated ELISA plate.   
     
     
         37 . The method of  claim 36 , wherein said first antibody effectively competes with the 3G4 antibody (monoclonal antibody 3G4 deposited as ATCC PTA 4545) in binding to an ELISA plate coated with phosphatidylserine in a competition ELISA conducted in the presence of serum, wherein said competition ELISA comprises:
 (a) coating an ELISA plate with phosphatidylserine to prepare a PS-coated ELISA plate;   (b) blocking said PS-coated ELISA plate with a blocking buffer comprising 10% serum;   (c) adding said 3G4 antibody diluted in said blocking buffer to said PS-coated ELISA plate under conditions effective to allow binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum;   (d) detecting the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum using a secondary antibody that binds to said 3G4 antibody; and   (e) identifying a first antibody that effectively competes with said 3G4 antibody in said competition ELISA by selecting a first antibody that substantially reduces the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum.   
     
     
         38 . The method of  claim 36 , wherein said antibody is bound to a solid support. 
     
     
         39 . The method of  claim 38 , wherein said method is an affinity chromatography method. 
     
     
         40 . The method of  claim 36 , wherein said biological sample is a blood sample. 
     
     
         41 . A method for treating a mammal with septic shock, comprising administering to said mammal a pharmaceutical composition comprising a first antibody, or an antigen-binding fragment thereof, in an amount effective to treat said septic shock; wherein said first antibody effectively competes with the 3G4 antibody (monoclonal antibody 3G4 deposited as ATCC PTA 4545) in binding to an ELISA plate coated with phosphatidylserine in a competition ELISA conducted in the presence of serum, wherein said competition ELISA comprises:
 (a) coating an ELISA plate with phosphatidylserine to prepare a PS-coated ELISA plate;   (b) blocking said PS-coated ELISA plate with a blocking buffer comprising 10% serum;   (c) adding said 3G4 antibody diluted in said blocking buffer to said PS-coated ELISA plate under conditions effective to allow binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum;   (d) detecting the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum using a secondary antibody that binds to said 3G4 antibody; and   (e) identifying a first antibody that effectively competes with said 3G4 antibody in said competition ELISA by selecting a first antibody that substantially reduces the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum.   
     
     
         42 . The method of  claim 41 , wherein said first antibody is a human, humanized, part-human, chimeric, recombinant or engineered antibody. 
     
     
         43 . The method of  claim 41 , wherein said antibody is an Fab dimer of said 3G4 antibody. 
     
     
         44 . The method of  claim 41 , wherein said mammal is a human patient. 
     
     
         45 . A method for treating a mammal with sickle cell anaemia, comprising administering to said mammal a pharmaceutical composition comprising a first antibody, or an antigen-binding fragment thereof, in an amount effective to treat said sickle cell anaemia; wherein said first antibody effectively competes with the 3G4 antibody (monoclonal antibody 3G4 deposited as ATCC PTA 4545) in binding to an ELISA plate coated with phosphatidylserine in a competition ELISA conducted in the presence of serum, wherein said competition ELISA comprises:
 (a) coating an ELISA plate with phosphatidylserine to prepare a PS-coated ELISA plate;   (b) blocking said PS-coated ELISA plate with a blocking buffer comprising 10% serum;   (c) adding said 3G4 antibody diluted in said blocking buffer to said PS-coated ELISA plate under conditions effective to allow binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum;   (d) detecting the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum using a secondary antibody that binds to said 3G4 antibody; and   (e) identifying a first antibody that effectively competes with said 3G4 antibody in said competition ELISA by selecting a first antibody that substantially reduces the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum.   
     
     
         46 . The method of  claim 45 , wherein said first antibody is a human, humanized, part-human, chimeric, recombinant or engineered antibody. 
     
     
         47 . The method of  claim 45 , wherein said antibody is an Fab dimer of said 3G4 antibody. 
     
     
         48 . The method of  claim 45 , wherein said mammal is a human patient. 
     
     
         49 . A method for treating a mammal with a protozoan infection, comprising administering to said mammal a pharmaceutical composition comprising a first antibody, or an antigen-binding fragment thereof, in an amount effective to treat said protozoan infection; wherein said first antibody effectively competes with the 3G4 antibody (monoclonal antibody 3G4 deposited as ATCC PTA 4545) in binding to an ELISA plate coated with phosphatidylserine in a competition ELISA conducted in the presence of serum, wherein said competition ELISA comprises:
 (a) coating an ELISA plate with phosphatidylserine to prepare a PS-coated ELISA plate;   (b) blocking said PS-coated ELISA plate with a blocking buffer comprising 10% serum;   (c) adding said 3G4 antibody diluted in said blocking buffer to said PS-coated ELISA plate under conditions effective to allow binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum;   (d) detecting the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum using a secondary antibody that binds to said 3G4 antibody; and   (e) identifying a first antibody that effectively competes with said 3G4 antibody in said competition ELISA by selecting a first antibody that substantially reduces the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum.   
     
     
         50 . The method of  claim 49 , wherein said first antibody is a human, humanized, part-human, chimeric, recombinant or engineered antibody. 
     
     
         51 . The method of  claim 49 , wherein said mammal is a human patient. 
     
     
         52 . A method for treating a mammal with antiphospholipid syndrome, comprising administering to said mammal a pharmaceutical composition comprising a first antibody, or an antigen-binding fragment thereof, in an amount effective to treat said antiphospholipid syndrome; wherein said first antibody effectively competes with the 3G4 antibody (monoclonal antibody 3G4 deposited as ATCC PTA 4545) in binding to an ELISA plate coated with phosphatidylserine in a competition ELISA conducted in the presence of serum, wherein said competition ELISA comprises:
 (a) coating an ELISA plate with phosphatidylserine to prepare a PS-coated ELISA plate;   (b) blocking said PS-coated ELISA plate with a blocking buffer comprising 10% serum;   (c) adding said 3G4 antibody diluted in said blocking buffer to said PS-coated ELISA plate under conditions effective to allow binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum;   (d) detecting the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum using a secondary antibody that binds to said 3G4 antibody; and   (e) identifying a first antibody that effectively competes with said 3G4 antibody in said competition ELISA by selecting a first antibody that substantially reduces the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum.   
     
     
         53 . The method of  claim 52 , wherein said first antibody is a human, humanized, part-human, chimeric, recombinant or engineered antibody. 
     
     
         54 . The method of  claim 52 , wherein said mammal is a human patient. 
     
     
         55 . A method for competing with a pathogenic antibody from a patient with antiphospholipid syndrome for binding to its phospholipid-protein target, comprising contacting said pathogenic antibody bound to said phospholipid-protein target with a first antibody, or an antigen-binding fragment thereof, which displaces said pathogenic antibody from said phospholipid-protein target; wherein said phospholipid-protein target is a phosphatidylserine (PS)-β 2 -glycoprotein I target and wherein said first antibody effectively competes with the 3G4 antibody (monoclonal antibody 3G4 deposited as ATCC PTA 4545) in binding to an ELISA plate coated with phosphatidylserine in a competition ELISA conducted in the presence of serum, wherein said competition ELISA comprises:
 (a) coating an ELISA plate with phosphatidylserine to prepare a PS-coated ELISA plate; 
 (b) blocking said PS-coated ELISA plate with a blocking buffer comprising 10% serum; 
 (c) adding said 3G4 antibody diluted in said blocking buffer to said PS-coated ELISA plate under conditions effective to allow binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum; 
 (d) detecting the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum using a secondary antibody that binds to said 3G4 antibody; and 
 (e) identifying a first antibody that effectively competes with said 3G4 antibody in said competition ELISA by selecting a first antibody that substantially reduces the binding of said 3G4 antibody to said PS-coated ELISA plate in the presence of serum. 
 
     
     
         56 . The method of  claim 55 , wherein said first antibody is a human, humanized, part-human, chimeric, recombinant or engineered antibody. 
     
     
         57 . A method for treating a mammal with a hepatitis C virus infection, comprising administering to said mammal a pharmaceutical composition comprising at least a first anti-viral agent in an amount effective to treat said hepatitis C virus infection; wherein said at least a first anti-viral agent is a chimeric antibody comprising antigen-binding fragments of mouse monoclonal antibody 3G4, produced by hybridoma ATCC PTA 4545, operatively attached to a human antibody constant region. 
     
     
         58 . The method of  claim 57 , wherein at least a second, distinct anti-viral agent is administered to said mammal. 
     
     
         59 . The method of  claim 58 , wherein said second, distinct anti-viral agent is interferon alpha or ribavirin. 
     
     
         60 . The method of  claim 59 , wherein said second, distinct anti-viral agent is ribavirin. 
     
     
         61 . The method of  claim 57 , wherein said at least a first anti-viral agent is administered to said mammal intravenously. 
     
     
         62 . The method of  claim 57 , wherein said mammal is a human patient. 
     
     
         63 . The method of  claim 62 , wherein said at least a first anti-viral agent is administered to said human patient in a dose of about 20 mgs or about 25 mgs per patient. 
     
     
         64 . The method of  claim 62 , wherein said at least a first anti-viral agent is administered to said human patient in a dose of about 200 mgs or about 225 mgs per patient. 
     
     
         65 . A method for treating a mammal with a hepatitis C virus infection, comprising administering to said mammal at least a first and second anti-viral agent in an amount effective to treat said hepatitis C virus infection; wherein said first anti-viral agent is a chimeric antibody comprising antigen-binding fragments of mouse monoclonal antibody 3G4, produced by hybridoma ATCC PTA 4545, operatively attached to a human antibody constant region; and wherein said second anti-viral agent is ribavirin. 
     
     
         66 . The method of  claim 65 , wherein said first and second anti-viral agents are administered to said mammal intravenously. 
     
     
         67 . The method of  claim 65 , wherein said mammal is a human patient. 
     
     
         68 . The method of  claim 67 , wherein said at least a first anti-viral agent is administered to said human patient in a dose of about 20 mgs or about 25 mgs per patient. 
     
     
         69 . The method of  claim 67 , wherein said at least a first anti-viral agent is administered to said human patient in a dose of about 200 mgs or about 225 mgs per patient. 
     
     
         70 . A method for treating a human patient with a hepatitis C virus infection, comprising administering to said human patient at least a first and second anti-viral agent in an amount effective to treat said hepatitis C virus infection; wherein said first anti-viral agent is a chimeric antibody comprising antigen-binding fragments of mouse monoclonal antibody 3G4, produced by hybridoma ATCC PTA 4545, operatively attached to a human antibody constant region; and wherein said second anti-viral agent is ribavirin. 
     
     
         71 . The method of  claim 70 , wherein said first and second anti-viral agents are administered to said human patient intravenously. 
     
     
         72 . The method of  claim 70 , wherein said at least a first anti-viral agent is administered to said human patient in a dose of about 20 mgs or about 25 mgs per patient. 
     
     
         73 . The method of  claim 70 , wherein said at least a first anti-viral agent is administered to said human patient in a dose of about 200 mgs or about 225 mgs per patient.