Adenoviral vectors encoding an antibody fused to a CD4 extracellular domain

Abstract

Genetically modified cell implant comprising an exogenous nucleotide sequence coding for all or part of an antibody, method for the preparation of such an implant and its therapeutic use for the treatment or prevention of an acquired disease. The invention also concerns an adenoviral vector for the expression of one or more proteins capable of forming a multimer, viral particles and cells containing the adenoviral vector, a pharmaceutical composition and its therapeutic use.

1 . Implant of genetically modified cells comprising an exogenous nucleotide sequence encoding all or part of an antibody, said exogenous nucleotide sequence being placed under the control of the elements necessary for its expression and for the secretion of said antibody.  
     
     
         2 . Implant according to  claim 1 , wherein said antibody is selected from the group consisting of: 
 a native antibody,    a chimeric antibody,    an antibody fragment, especially a fragment Fab, F(ab′) 2 , Fc or scFv, and    a bispecific antibody.    
     
     
         3 . Implant according to  claim 1 , wherein said antibody is modified by a toxic or immunopotentiating substance.  
     
     
         4 . Implant according to  claim 3 , wherein said antibody may be modified by a toxic substance selected from a ribonuclease, and especially the ribonuclease from  Bacillus amyloliquefaciens , ricin, diphtheria toxin, cholera toxin, herpes simplex virus thymidine kinase, cytosine deaminase from  Escherichia coli  or from a yeast of the genus  Saccharomyces , exotoxin from  Pseudomonas  and human angiogenin or an analog of said substances.  
     
     
         5 . Implant according to  claim 1 , wherein cells are genetically modified by transfection of a vector derived from a plasmid, from a retrovirus or from a herpes virus, from an adenovirus, from an adenovirus-associated virus comprising said exogenous nucleotide sequence placed under the control of the elements necessary for its expression and for the secretion of the said antibody.  
     
     
         6 . Implant according to  claim 5 , wherein said vector is dicistronic.  
     
     
         7 . Implant according to  claim 6 , wherein said vector is retroviral and comprises from 5′ to 3′: 
 (a) a 5′ LTR derived from a retrovirus,    (b) an encapsidation region,    (c) an exogenous nucleotide sequence comprising: 
 an internal promoter,  
 a first sequence encoding the heavy chain of an antibody,  
 a ribosome entry initiation site,  
 a second sequence encoding the light chain of an antibody, and  
   (d) a 3′ LTR derived from a retrovirus.    
     
     
         8 . Implant according to  claim 7 , wherein said exogenous nucleotide sequence comprises, in addition, a third sequence encoding a toxic or immunopotentiating substance fused downstream and operably to the second sequence.  
     
     
         9 . Implant according to  claim 1 , comprising genetically modified autologous cells.  
     
     
         10 . Implant according to  claim 9 , comprising genetically modified fibroblasts.  
     
     
         11 . Implant according to  claim 1 , wherein from 10 6  to 10 12 , preferably from 10 7  to 10 11  genetically modified cells.  
     
     
         12 . Method for the preparation of an implant according to  claim 1 , wherein the genetically modified cells and an extracellular matrix are placed in contact.  
     
     
         13 . A pharmaceutical composition intended for the treatment or for the prevention of an acquired disease comprising the implant according to  claim 1 .  
     
     
         14 . A pharmaceutical composition intended for the treatment or for the prevention of an infectious disease cancer comprising the implant according  claim 1 .  
     
     
         15 . Recombinant adenoviral vector comprising an exogenous nucleotide sequence encoding all or part of an antibody and placed under the control of the elements necessary for its expression, wherein said antibody is modified by a toxic or immunopotentiating substance.  
     
     
         16 . Recombinant adenoviral vector according to  claim 15 , the wherein said antibody is selected from the group consisting of a native antibody, a chimeric antibody, an antibody fragment and especially a fragment F(ab′) 2 , Fc or scFv and a bispecific antibody.  
     
     
         17 . Recombinant adenoviral vector according to  claim 15 , wherein said antibody may be modified by a toxic substance selected from a ribonuclease, and especially the ribonuclease from  Bacillus amyloliquefacienis , ricin, diphtheria toxin, cholera toxin, herpes simplex virus thymidine kinase, cytosine deaminase from  Escberichia coli  or from a yeast of the genus  Saccharomyces , exotoxin from  Pseudomonas  and human angiogenin or an analog of said substances.  
     
     
         18 . Recombinant adenoviral vector according to  claim 15 , wherein said antibody is modified by an immunopotentiating substance.  
     
     
         19 . Recombinant adenoviral vector comprising an exogenous nucleotide sequence encoding all or part of one or more protein(s) of interest capable of forming a multimer, such as a dimer or a tetramer, in a host cell; the said exogenous nucleotide sequence being placed under the control of the elements necessary for its expression, said vector being derived from an adenovirus of human, canine, avian, bovine, murine, ovine, porcine or simian origin or a hybrid comprising adenoviral genome fragments of different origins.  
     
     
         20 . Recombinant adenoviral vector according to  claim 15 , derived from an adenovirus of human, canine, avian, bovine, murine, ovine, porcine or simian origin or from a hybrid comprising adenoviral genome fragments of different origins.  
     
     
         21 . Recombinant adenoviral vector according to  claim 15 , wherein it is defective for replication.  
     
     
         22 . Recombinant adenoviral vector according to  claim 21 , wherein it at least lacks all or part of the E1 region and, optionally, all or part of the E3 region.  
     
     
         23 . Recombinant adenoviral vector according to  claim 21 , comprising an exogenous nucleotide sequence encoding the heavy chain of the 2F5 antibody, an IRES element and the light chain of the 2F5 antibody; said exogenous nucleotide sequence being placed under the control of the elements necessary for its expression.  
     
     
         24 . Recombinant adenoviral vector according to  claim 21 , comprising an exogenous nucleotide sequence encoding the signal sequence and the extracellular I and II domains of the CD4 protein operably fused to the constant γ3 region (hinge region-CH2 and CH3) of the heavy chain of the 2F5 antibody.  
     
     
         25 . Recombinant adenoviral vector according to  claim 21 , comprising an exogenous nucleotide sequence encoding the signal sequence and the extracellular I and II domains of the CD4 protein operably fused to the constant γ3 region (hinge region-CR2 and CR3) of the heavy chain of the 2F5 antibody and operably fused to the mature human angiogenin.  
     
     
         26 . Recombinant adenoviral vector according to  claim 15 , wherein the elements necessary for the expression comprise a promoter selected from the group consisting of the adenoviral early promoter E1A, the late promoter MLP (Major Late Promoter), the murine or human PGK (Phosphoglycerate kinase) promoter, the SV40 virus early promoter, the RSV (Rous Sarcoma virus) virus promoter, a promoter which is specifically active in tumor cells and finally a promoter which is specifically active in the infected cells.  
     
     
         27 . Infectious viral particle comprising a recombinant adenoviral vector according to  claim 15 .  
     
     
         28 . Eukaryotic host cell comprising a recombinant adenoviral vector according to  claim 15 .  
     
     
         29 . Pharmaceutical composition comprising a recombinant adenoviral vector according to  claim 15 , in association with a pharmaceutically acceptable carrier.  
     
     
         30 . Pharmaceutical composition according to  claim 29 , comprising 10 4  to 10 14  pfu.  
     
     
         31 . Pharmaceutical composition according to  claim 29  wherein it is in injectable form.  
     
     
         32 . A method for the preparation of a pharmaceutical composition intended for the treatment and/or prevention of the human or animal body by gene therapy comprising using the recombinant adenoviral vector according to  claim 15 .  
     
     
         33 . The method according to  claim 32 , wherein said pharmaceutical composition is intended for the treatment and/or prevention of acquired diseases and especially cancers and AIDS.  
     
     
         34 . The method according to  claim 33 , wherein said pharmaceutical composition administrable via the intravenous or intratumor route.