Rationally designed, synthetic antibody libraries and uses therefor

Abstract

The present invention overcomes the inadequacies inherent in the known methods for generating libraries of antibody-encoding polynucleotides by specifically designing the libraries with directed sequence and length diversity. The libraries are designed to reflect the preimmune repertoire naturally created by the human immune system and are based on rational design informed by examination of publicly available databases of human antibody sequences.

1 - 72 . (canceled) 
     
     
         73 . A method of isolating one or more host cells expressing one or more antibodies, the method comprising:
 (i) expressing the one or more antibodies in one or more host cells, wherein the one or more antibodies comprise:
 (a) a CDRH3 amino acid sequence comprising:
 (A) an N1 amino acid sequence of 0 to about 3 amino acids, 
 
 wherein each amino acid of the N1 amino acid sequence is among the 12 most frequently occurring amino acids at the corresponding position in N1 amino acid sequences of CDRH3 amino acid sequences that are functionally expressed by B cells,
 (B) a human CDRH3 DH amino acid sequence, N- and C-terminal truncations thereof, or a sequence of at least about 80% identity to any of them, 
 (C) an N2 amino acid sequence of 0 to about 3 amino acids, 
 
 wherein each amino acid of the N2 amino acid sequence is among the 12 most frequently occurring amino acids at the corresponding position in N2 amino acid sequences of CDRH3 amino acid sequences that are functionally expressed by B cells; and
 (D) a human CDRH3 H3-JH amino acid sequence, N-terminal truncations thereof, or a sequence of at least about 80% identity to any of them; and 
 
 (b) a VKCDR3 amino acid sequence comprising about 1 to about 10 of the amino acids found at Kabat positions 89, 90, 91, 92, 93, 94, 95, 95A, 96, and 97, in selected VKCDR3 amino acid sequences derived from a particular IGKV or IGKJ germline sequence; or 
 (c) a VKCDR3 amino acid sequence of at least about 80% identity to a amino acid sequence represented by the following formula:
   [VK_Chassis]-[L3-VK]-[X]-[JK*], wherein: 
 (1) VK Chassis is an amino acid sequence selected from the group consisting of about Kabat amino acid 1 to about Kabat amino acid 88 encoded by IGKV1-05, IGKV1-06, IGKV1-08, IGKV1-09, IGKV1-12, IGKV1-13, IGKV1-16, IGKV1-17, IGKV1-27, IGKV1-33, IGKV1-37, IGKV1-39, IGKV1D-16, IGKV1D-17, IGKV1D-43, IGKV1D-8, IGKV2-24, IGKV2-28, IGKV2-29, IGKV2-30, IGKV2-40, IGKV2D-26, IGKV2D-29, IGKV2D-30, IGKV3-11, IGKV3-15, IGKV3-20, IGKV3D-07, IGKV3D-11, IGKV3D-20, IGKV4-1, IGKV5-2, IGKV6-21, and IGKV6D-41, or a sequence of at least about 80% identity to any of them; 
 (2) L3-VK is the portion of the VKCDR3 encoded by the IGKV gene segment; 
 (3) X is any amino acid residue; and 
 (4) JK* is an amino acid sequence selected from the group consisting of amino acid sequences encoded by IGJK1, IGJK2, IGJK3, IGJK4, and IGJK5, wherein the first residue of each IGJK amino acid sequence is not present; and 
 
   (ii) contacting the host cells with one or more antigens; and   (iii) isolating one or more host cells having antibodies that bind to the one or more antigens.   
     
     
         74 . The method of  claim 73 , further comprising isolating the one or more antibodies from the one or more host cells. 
     
     
         75 . The method of  claim 73 , further comprising the step of isolating one or more polynucleotide sequences encoding the one or more antibodies from the one or more host cells. 
     
     
         76 . The method of  claim 73 , wherein the one or more host cells are yeast cells. 
     
     
         77 . The method of  claim 76 , wherein the yeast cells are  S. cerevisiae  cells. 
     
     
         78 . An antibody isolated from the one or more host cells isolated according to the method of  claim 73 .