Chimeric toxins for targeted therapy

Abstract

The present invention relates particularly to neoplastic cells targeted chimeric toxins comprising of cell targeting moieties and cell killing moieties for recognizing and for destroying the neoplastic cells, wherein the cell targeting moieties consist of gonadotropin releasing hormone homologues and the cell killing moieties consist of Pseudomonas Exotoxin A. The present invention further relates to pharmaceutical compositions containing as an active ingredient these neoplastic cells targeted chimeric toxins and to a method for the production of these chimeric toxins. The said invention also relates to a method for cancer therapy, treating malignant carcinoma cells and benign hyperplasia including uterine leiomyoma cells, extrauterine endometrial island cells, benign hyperplasia of prostate and breast and pituitary tumor adenoma cells, by the use of the above-mentioned chimeric toxins.

1 . A fused chimeric protein comprising a linear genetically engineered molecule consisting essentially of peptide bonds, produced by fusing, at the level of cDNA: 
 A. DNA encoding at least one cell targeting moiety consisting essentially of Met-GnRH or a Met-GnRH analog that specifically binds to GnRH binding sites on Caco2 adenocarcinoma cells; and    B. DNA encoding at least one cell killing moiety.    
     
     
         2 . A fused chimeric protein according to  claim 1 , produced by fusing at the cDNA level an oligonucleotide encoding gonadotropin releasing hormone (GnRH) or a GnRH analog, preceded by a Met, to a mutated DNA fragment of the full length  Pseudomonas  exotoxin (PE), encoding the protein Met-GnRH-PE66.  
     
     
         3 . A fused chimeric protein according to  claim 1 , produced by fusing at the cDNA level an oligonucleotide encoding gonadotropin releasing hormone (GnRH) or a GnRH analog, preceded by a Met, to a DNA fragment comprising domains II and III of the  Pseudomonas  exotoxin (PE), encoding the protein Met-GnRH-PE40.  
     
     
         4 . A method for the production of a chimeric protein as defined in  claim 3 , comprising ligating an oligonucleotide encoding gonadotropin releasing hormone (GnRH) or a GnRH analog, preceded by a Met, upstream of a DNA fragment encoding a mutated form of PE, under conditions sufficient to produce a chimeric protein comprising Met-GnRH-PE66.  
     
     
         5 . A method for the production of a chimeric protein as defined in  claim 4  that targets adenocarcinoma cells, comprising ligating an oligonucleotide encoding gonadotropin releasing hormone (GnRH) or a GnRH analog, preceded by a Met, upstream to a DNA fragment encoding domains II and III of the PE, under conditions sufficient to produce a chimeric protein comprising Met-GnRH-PE40.  
     
     
         6 . A composition useful for treatment in cancer therapy comprising as active ingredient, a chimeric protein as defined in  claim 1 .  
     
     
         7 . A plasmid comprising a promoter operably linked to a DNA molecule encoding a fused chimeric protein as defined in  claim 1 .  
     
     
         8 . A chimeric protein comprising a genetically engineered molecule comprising a fusion of 
 at least one cell targeting moiety consisting essentially of gonadotropin releasing hormone (GnRH) preceded by a Met (Met-GnRH) or a Met-GnRH analog that specifically binds to GNRH binding sites on Caco2 adenocarcinoma; and    at least one cell killing moiety.    
     
     
         9 . A fusion protein as claimed in  claim 8 , wherein said cell killing moiety comprises  Pseudomonas  exotoxin A.  
     
     
         10 . A fusion protein as claimed in  claim 8 , that is a single protein.  
     
     
         11 . A fusion protein as claimed in  claim 8 , that has no linking moiety between said cell killing moiety and said cell targeting moiety.  
     
     
         12 . A fusion protein as claimed in  claim 8 , wherein said chimeric protein recognizes and/or binds to GnRH-binding sites on adenocarcinoma and hepatocarcinoma cells.  
     
     
         13 . A fusion protein as claimed in  claim 8 , wherein said cell targeting moiety is a Met-GnRH analog having the sequence of Met-GnRH but having a glycine residue as the sixth amino acid of GnRH.