Immunoconjugates and humanized antibodies specific for b-cell lymphoma and leukemia cells

Abstract

A chimeric LL2 monoclonal antibody is described in which the complementarity determining regions (CDRs) of the light and heavy chains of the murine LL2 anti-B-lymphoma, anti-leukemia cell monoclonal antibody has been recombinantly joined to the human kappa and IgG 1 constant region domains, respectively, which retains the immunospecificity and B-cell lymphoma and leukemia cell internalization capacity of the parental murine LL2 monoclonal antibody, and which has the potential of exhibiting reduced human anti-mouse antibody production activity. A humanized LL2 monoclonal antibody is described in which the CDRs of the light and heavy chains have been recombinantly joined to a framework sequence of human light and heavy chains variable regions, respectively, and subsequently linked to human kappa and IgG 1 constant region domains, respectively, which retains the immunospecificity and B-lymphoma and leukemia cell internalization capacities of the parental murine and chimeric LL2 monoclonal antibodies, and which has the potential for exhibiting reduced human anti-mouse antibody production activity. Vectors for producing recombinant chimeric and humanized chimeric monoclonal antibodies are provided. Isolated DNAs encoding the amino acid sequences of the LL2 variable light and heavy chain and CDR framework regions are described. Conjugates of chimeric and humanized chimeric LL2 antibodies with cytotoxic agents or labels find use in therapy and diagnosis of B-cell lymphomas and leukemias.

1 . A humanized immunoglobulin chain comprising four FRs (FR1 to FR4) from a human acceptor immunoglobulin chain and CDRs from a non-human donor immunoglobulin chain wherein not all of the FRs are from the same human acceptor immunoglobulin chains.  
     
     
         2 . The humanized immunoglobulin chain of  claim 1  wherein the humanized immunoglobulin chain is a heavy chain.  
     
     
         3 . The humanized immunoglobulin chain of  claim 1  wherein the humanized immunoglobulin chain is a light chain.  
     
     
         4 . The humanized immunoglobulin chain of  claim 1  wherein the humanized immunoglobulin chain comprises the variable domain.  
     
     
         5 . The humanized immunoglobulin chain of  claim 4  wherein the humanized immunoglobulin chain further comprises the constant domain.  
     
     
         6 . The humanized immunoglobulin chain of  claim 1  wherein FR4 is selected from a different immunoglobulin chain than FR1, FR2 and FR3.  
     
     
         7 . The humanized immunoglobulin chain of  claim 1  wherein at least one residue in the FR region of the FR-CDR interface is a residue from the non-human donor immunoglobulin chain.  
     
     
         8 . The humanized immunoglobulin chain of  claim 1  wherein potential FR-CDR interactions are identified by modelling and wherein non-human donor immunoglobulin FR residues which interact with a CDR are maintained in the humanized immunoglobulin chain.  
     
     
         8 . The humanized immunoglobulin chain of  claim 1  wherein FR residues within 4.5 Å of at least one CDR from the non-human donor immunoglobulin are non-human donor immunoglobulin residues.  
     
     
         9 . The humanized immunoglobulin chain of  claim 1  wherein at least 75 percent of the FR-CDR interface residues in the humanized immunoglobulin chain that are not conserved between the human acceptor immunoglobulin chain and the non-human donor immunoglobulin chain are residues from the non-human donor immunoglobulin chain.  
     
     
         10 . The humanized immunoglobulin chain of  claim 1  wherein all of the FR-CDR interface residues in the humanized immunoglobulin chain that are not conserved between the human acceptor immunoglobulin chain and the non-human donor immunoglobulin chain are residues from the non-human donor immunoglobulin chain.  
     
     
         11 . A humanized immunoglobulin chain comprising four human FRs (FR1 to FR4) from a human acceptor immunoglobulin chain and CDRs from a non-human donor immunoglobulin chain, wherein the non-human donor immunoglobulin chain further comprises non-human FRs and wherein the four human FRs are selected based on sequence homology to the non-human FRs.  
     
     
         12 . A humanized immunoglobulin chain according to  claim 11  wherein not all of the human FRs are selected from the same human acceptor immunoglobulin chain.  
     
     
         13 . A humanized immunoglobulin chain comprising four human FRs (FR1 to FR4) from a human immunoglobulin chain and CDRs from a murine immunoglobulin chain, wherein the murine immunoglobulin chain further comprises murine FRs and wherein the human FRs are selected based on sequence homology to the murine FRs.  
     
     
         14 . A humanized immunoglobulin chain according to  claim 13  wherein not all of the human FRs are selected from the same human immunoglobulin chain.  
     
     
         15 . A humanized antibody comprising light and heavy humanized immunoglobulin chains, the humanized immunoglobulin chains comprising four FRs (FR1 to FR4) from a human acceptor immunoglobulin chain and CDRs from a non-human donor immunoglobulin chain wherein, for at least one of the humanized immunoglobulin chains, not all of the FRs are from the same human acceptor immunoglobulin chains.  
     
     
         16 . A humanized antibody comprising light and heavy humanized immunoglobulin chains, the humanized immunoglobulin chains comprising four FRs (FR1 to FR4) from a human immunoglobulin chain and CDRs from a murine immunoglobulin chain wherein, for at least one of the humanized immunoglobulin chains, not all of the FRs are from the same human immunoglobulin chains.  
     
     
         17 . A method of producing a humanized antibody comprising: 
 comparing the variable (V) region framework (FR) sequences of a non-human antibody to the variable (V) region framework (FR) sequences of human antibodies to determine the degree of sequence homology between the non-human antibody FRs and the human antibody FRs; and    replacing FRs in the non-human antibody with the human antibody FRs which exhibit sequence homology to the non-human antibody FRs.    
     
     
         18 . A method according to  claim 17 , wherein the FRs in the non-human antibody are replaced with the FRs having the highest degree of sequence homology to the non-human FRs of the sequences that were compared.