Immunoconjugates of toxins directed against malignant cells

Abstract

This invention provides for new recombinant ribonuclease proteins which are active when expressed by bacteria. This allows the recombinant ribonucleases of this invention to be fused in-frame with ligand binding moieties to form cytotoxic fusion proteins. Furthermore, these proteins are more active than ribonucleases currently available even though the proteins of this invention lack an N-terminal pyroglutamic acid, which has been found to be necessary for ribonucleolytic activity. Because these proteins are recombinant proteins, mutations which increase cytotoxicity can be engineered.

1 . A cytotoxic reagent comprising a ribonuclease encoded by a nucleic acid comprising SEQ ID NO: 14 and conservative variants thereof, 
 wherein the amino acid sequence is selected from the group consisting of SEQ ID NO: 15, SEQ ID NO:17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:24, and SEQ ID NO:26,    said ribonuclease being linked to an antibody directed against a cell surface antigen present on a cancerous B cell.    
     
     
         2 . The cytotoxic reagent of  claim 1 , wherein said antibody is directed against CD22.  
     
     
         3 . The cytotoxic reagent of  claim 2 , wherein said antibody is LL2.  
     
     
         4 . A method of killing malignant B cells comprising contacting cells to be killed with a cytotoxic reagent expressed by recombinant DNA, comprising a sequence selected from the group consisting of SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:8, SEQ ID NO: 1, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO:19, SEQ ID NO:21, SEQ ID NO:24 and SEQ ID NO:26 covalently linked to a ligand binding moiety, said ligand binding moiety being directed against a cell surface antigen on the malignant B cells.  
     
     
         5 . The method of  claim 4 , wherein said ligand binding moiety is an antibody.  
     
     
         6 . The method of  claim 5 , wherein said antibody is a single chain antibody.  
     
     
         7 . The method of  claim 5 , wherein said ligand binding moiety is an antibody directed against CD22.  
     
     
         8 . The method of  claim 7 , wherein said antibody is LL2.