JP-2021513323-A5 -

Dates

Publication Date

20230516

Application Date

20190211

Description

The antibody molecule or its antigen-binding portion may contain one or more human variable domain framework skeletons , in which a CDR is inserted. For example, the VH region, the VL region, or both the VH and VL regions may contain amino acid sequences of one or more human framework regions (FRs). The antibody molecule or its antigen-binding portion may contain the IGHV1-46 human germline skeleton , in which the corresponding HCDR sequence is inserted. The antibody molecule or its antigen-binding portion may contain a VH region containing the amino acid sequence of the IGHV1-46 human germline skeleton , in which the corresponding sets of HCDR1, HCDR2, and HCDR3 amino acid sequences are inserted. The antibody molecule or its antigen-binding portion may contain the IGKV1-16 human germline skeleton , in which the corresponding LCDR sequence is inserted. The antibody molecule or its antigen-binding portion may contain a VL region containing the amino acid sequence of the IGKV1-16 human germline skeleton , in which the corresponding sets of LCDR1, LCDR2, and LCDR3 amino acid sequences are inserted. The antibody molecule or its antigen-binding portion may contain the IGHV1-46 human germline skeleton , with the corresponding HCDR sequence inserted therein, and may also contain the IGKV1-16 human germline skeleton , with the corresponding LCDR sequence inserted therein. The antibody molecule or its antigen-binding portion may contain a VH region containing the amino acid sequence of the IGHV1-46 human germline skeleton , with the corresponding sets of HCDR1, HCDR2, and HCDR3 amino acid sequences inserted therein, and may also contain a VL region containing the amino acid sequence of the IGKV1-16 human germline skeleton , with the corresponding sets of LCDR1, LCDR2, and LCDR3 amino acid sequences inserted therein. The amino acid sequences of HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 may be those of any one of the clones listed in Table 4 or Table 6 (all six CDR sequences are derived from the same clone). Results and Discussion <br/> CDR Transplantation to Preferred Human Germline V-Genes The CDR of antagonist mouse anti-C-KIT IgG 37M (37M; see WO2014018625A1 and Table 2) was initially introduced into the human germline immunoglobulin v-domain framework sequence scaffold using the CDR transplantation method. To direct genetic engineering efforts towards the final lead therapeutic IgG compound with optimal drug-like performance, we chose to transplant the parental antibody CDR into the "preferred" germline scaffolds IGHV1-46 and IGKV1-16. These scaffolds are known to have good solubility and high physical stability and are frequently used in the expressed human antibody repertoire. The definitions of these skeletons and transplanted CDRs are summarized in Table 2. The heavy and light chain sequences of the chimeric anti-C-KIT antibody m37M and the humanized h37M are also shown in Table 2. Although the process of CDR transplantation is well known, it remains difficult to predict whether a given set of human v-domain sequences will function as an appropriate acceptor framework for non-human CDR transplantation. The use of an inappropriate framework can lead to loss of target binding function, protein stability problems, and even decreased final IgG expression. In fact, the inclusion of multiple mouse residues in the h37M framework region suggests that target binding affinity could not be fully maintained in direct CDR transplantation into the germline framework. Therefore, IGHV1-46/IGKV1-16 transplantation was adopted as a template for CDR mutation induction, and improved clones were selected.