Patent ID: 7494647
Filing Date: 2009-02-24
Classification: A61K,A61P,C07K

Abstract:
1. A process of preparing a natural humanized antibody that has complementarity determining regions (CDRs) derived from non-humans and a framework region (FR) derived from a natural human antibody and that has a reduced immunogenicity, said method comprising the steps of: (1) preparing a non-human monoclonal antibody responsive to an antigen of interest; (2) preparing a plurality of human antibodies having a high homology with the amino acid sequences of FRs of heavy chain (H chain) or light chain (L chain) in the monoclonal antibodies of the step (1); (3) preparing a first humanized antibody by steps comprising: a) replacing all FRs of H chain of the non-human antibody prepared in step (1) with corresponding FRs H chain of one human antibody prepared in step (2); b) replacing all FRs of L chain of the non-human antibody prepared in step (1) with corresponding FRs of L chain of one human antibody prepared in step (2); and c) changing constant region of the non-human antibody prepared in step (1) to constant region of a human antibody; (4) generating second antibodies by: a) in an H chain V region of the first humanized antibody prepared on said (3)a), replacing 1 to 3 FRs having at least one of FR2 and FR3, with corresponding FRs of a human antibody different from the antibody used in (3)a) selected from the human antibodies prepared in the step (2); b) in an L chain V region of the first humanized antibody prepared on said (3)b), replacing 1 to 3 FRs having at least one of FR2 and FR3, with corresponding FRs of a human antibody different from the antibody used in (3)b) selected from the human antibodies prepared in the step (2); and c) obtaining a second humanized antibody comprising an H chain obtained in a), an L chain obtained in b, and a constant region of a human antibody in (3)c); (5) determining ability of the first and the second humanized antibodies to bind to the antigen or ability of the first and the second humanized antibodies to neutralize a biological activity of the antigen; (6) comparing the ability of the first and the second antibodies determined in step (5), and selecting antibodies which have a higher antigen binding ability or higher ability to neutralize a biological activity of an antigen; and (7) repeating the steps of (4) to (6) while using the antibody selected in step (6) as a first humanized antibody in step (4) until a humanized antibody is selected having the same level or more of ability as the non-human monoclonal antibody in step (1), wherein said ability is to bind to the antigen or to neutralize a biological activity of the antigen.