Modified anti-cd52 antibody

Abstract

The present invention provides for modified forms of anti-CD52 antibodies with reduced numbers of potential T-cell epitopes that are expected to display decreased immunogenicity.

1 - 22 . (canceled) 
     
     
         23 . An expression vector comprising:
 a) a first nucleic acid sequence coding for a V-region heavy chain comprising a substituted variant of SEQ ID NO: 1 with one or more of the following substitutions wherein the numbering of amino acid residues relates to those of SEQ ID NO: 1:
 substitution of Lys at amino acid residue 3 with Gln; 
 Leu at amino acid residue 5 with Ala, Cys, Asn, Asp, Gln, Glu, Gly, His, Lys, Pro, Arg, Ser, or Thr; 
 Val at amino acid residue 12 with Asn, Asp, Glu, His, Lys, Pro, Gln, Arg, Ser, or Thr; 
 Gln at amino acid residue 13 with Ala, Phe, His, Lys, Asn, Pro, Gln, Arg, Ser, Thr; 
 Gly at amino acid residue 15 with Asp, His, Pro, Gln, Arg, Ser, Thr; 
 Ser at amino acid residue 17 with Gly, Met, Pro or Trp; 
 Met at amino acid residue 18 with Arg, Gly, Pro, Leu; 
 Arg at amino acid residue 19 with Ala, Cys, Phe, Gly, Ile, Leu, Met, Pro, Val, Trp, or Tyr; 
 Leu at amino acid residue 20 with Ala, Cys, Phe, Gly, His, Ile, Lys, Asn, Asp, Met, Gln, Glu, Pro, Arg, Ser, Thr, Val Trp, or Tyr; 
 Ser at amino acid residue 21 with Pro; 
 Ala at amino acid residue 23 with Asn, Asp, Glu, Gln, Gly, His, Lys, Pro, Arg, Ser, or Thr; 
 Ser at amino acid residue 25 with Phe, Gly, Leu, Pro, Trp or Tyr; 
 Gly at amino acid residue 26 with Asp, Asn, Glu, Gln, His, Lys, Pro, Arg, Ser, Thr, Trp or Tyr; 
 Asp at amino acid residue 31 with Ala, Phe, Gly, Ile, Met, Pro, Val, Trp, or Tyr; 
 Tyr at amino acid residue 33 with Ala, Gly, Met, or Pro; 
 Asn at amino acid residue 35 with Pro; 
 Trp at amino acid residue 36 with Ala, Asp, Glu, Gly, His, Lys, Asn, Pro, Gln, Arg, Ser, or The; 
 Ile at amino acid residue 37 with Val; 
 Arg at amino acid residue 38 with Phe, His, Pro, or Tyr; 
 Pro at amino acid residue 40 with Ala; 
 Ala at amino acid residue 41 with Asp, Asn, Glu, Gln, His, Lys, Pro, Arg, Ser, The or Trp; 
 Gly at amino acid residue 42 with Ile, Pro, Thr, Tyr; 
 Ala at amino acid residue 44 with Gly, His, Asn, Pro, Gln, Ser, Thr, Trp, Tyr, 
 Pro at amino acid residue 45 with Leu; 
 Leu at amino acid residue 48 with Val or Ile, 
 Thr at amino acid residue 71 with Phe, Leu, Pro, Trp, or Tyr; 
 Ile at amino acid residue 72 with Asp, Glu, His, Lys, Asn, Pro, Gln, Arg, Ser, or Thr; 
 Ser at amino acid residue 73 with Ala, Gly, or Pro; 
 Arg at amino acid residue 74 with Ala, Phe, Gly, Ile, Met, Pro, Trp, or Tyr; 
 Thr at amino acid residue 77 with Ala, His, Ile, Pro or Ser; 
 Gln at amino acid residue 78 with Lys; 
 Asn at amino acid residue 79 with Ala, Phe, Gly, Ile, Met, Pro, Val, Trp or Tyr; 
 Met at amino acid residue 80 with Ala, Asp, Glu, Gly, His, Lys, Asn, Pro, Gln, Arg, Thr, or Ser; 
 Tyr at amino acid residue 82 with Ala, Asp, Glu, Gly, His, Lys, Asn, Pro, Gln, Arg, Ser or Thr; 
 Gln at amino acid residue 84 with Ala, Phe, Gly, Ile, Leu, Met, Pro, Val, Trp or Tyr; 
 Met at amino acid residue 85 with Ala, Asp, Glu, Gly, His, Lys, Asn, Pro, Gln, Arg, Ser or Thr; 
 Thr at amino acid residue 87 with Ser; 
 Leu at amino acid residue 88 with Asp, Glu, Gly, His, Lys, Asn, Pro, Gln, Arg, Ser, or Thr; 
 Arg at amino acid residue 89 with Phe, Pro, Trp, Tyr; 
 Ala at amino acid residue 90 with Asn, Asp, Glu, Gln, His, Lys, Pro, Arg, Ser, Thr, Trp or Tyr; 
 Glu at amino acid residue 91 with Pro; 
 Asp at amino acid residue 92 with Ala, Phe, Gly, Ile, Leu, Met, Pro, Val, Trp, Tyr; 
 Thr at amino acid residue 95 with Val; 
 Asp at amino acid residue 109 with Ala, Phe, Gly, Ile, Leu, Met, Pro, Val, Trp or Tyr; 
 Trp at amino acid residue 111 with Ala, Asp, Glu, Gly, His, Lys, Asn, Pro, Gln, Arg, Ser, or Thr; 
 Gly at amino acid residue 114 with His, Pro, Ser, or Thr; 
 Val at amino acid residue 115 with Thr; 
 Met at amino acid residue 116 with Thr, Phe, Ile, Leu, Pro, Val, Trp or Tyr; 
 Val at amino acid residue 117 with Ala, Phe, Gly, Ile, Met, Pro, Trp or Tyr; and 
   b) a second nucleic acid sequence coding for a V-region light chain comprising a substituted variant of SEQ ID NO:2 with one or more of the following substitutions wherein the numbering of amino acid residues relates to those of SEQ ID NO: 2:
 substitution of Lys at amino acid residue 3 with Gln; 
 Phe at amino acid residue 10 with Ala, Asp, Asn, Glu, Gln, Gly, His, Lys, Pro, Arg, Ser or Thr; 
 Val at amino acid residue 15 with Ala, Gly, His or Pro; 
 Asp at amino acid residue 17 with Pro; 
 Val at amino acid residue 19 with Pro or Trp; 
 Leu at amino acid residue 21 with Pro or Ile; 
 Asn at amino acid residue 22 with Thr; 
 Lys at amino acid residue 24 with Arg; 
 Leu at amino acid residue 33 with Ala, Asp, Asn, Gln, Glu, Gly, His, Lys, Pro, Arg, Ser or Thr; 
 Leu at amino acid residue 40 with Asp, Asn, Gln, Glu, Gly, His, Lys, Pro, Arg, Ser or Thr; 
 Glu at amino acid residue 42 with Lys; 
 Ser at amino acid residue 43 with Ala; 
 Leu at amino acid residue 46 with Ser; 
 Thr at amino acid residue 56 with Ala, Phe, Gly, Ile, Met, Pro, Ser, Trp or Tyr; 
 Ile at amino acid residue 58 with Ala; Gly, Met, Pro or Val; 
 Ser at amino acid residue 60 with Ala, Phe; Gly, Ile, Met, Pro; Trp or Tyr; 
 Arg at amino acid residue 61 with Pro; 
 Ser at amino acid residue 63 with Phe, Leu, Pro, Trp or Tyr; 
 Gly at amino acid residue 64 with Asp, Asn, Gln, Glu, His, Lys, Pro, Arg, Ser or Thr; 
 Leu at amino acid residue 78 with Asp, Asn, Gln, Glu, Gly, His, Lys, Pro, Arg, Ser, Thr; 
 Val at amino acid residue 83 with Ala, Asp, Asn, Glu, Gln, Gly, His, Ile, Lys, Pro, Arg, Ser, Thr; 
 Phe at amino acid residue 87 with Tyr, 
   
       wherein the first nucleic acid sequence and the second nucleic acid sequence are operably linked to one or more expression control sequences. 
     
     
         24 . An expression vector comprising a nucleic acid sequence coding for SEQ ID NO: 71, SEQ ID NO: 73, SEQ ID NO: 74 or SEQ ID NO: 75, or a degenerate variant thereof. 
     
     
         25 . (canceled) 
     
     
         26 . The expression vector of  claim 23 , further comprising a nucleic acid sequence coding for a human IgG1 constant region domain. 
     
     
         27 . A cultured cell comprising the vector of  claim 23 . 
     
     
         28 . (canceled) 
     
     
         29 . An expression vector comprising a nucleic acid sequence coding for SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, or SEQ ID NO: 79, or a degenerate variant thereof. 
     
     
         30 . (canceled) 
     
     
         31 . The expression vector of  claim 23 , further comprising a nucleic acid sequence coding for a human kappa constant region domain. 
     
     
         32 - 33 . (canceled) 
     
     
         34 . A method of preparing an immunoglobulin, comprising culturing the cell of  claim 27  under conditions permitting expression under the control of the expression control sequences, and purifying the immunoglobulin from the medium of said cell, wherein the immunoglobulin has specificity for CD52 antigen.