Chugai Seiyaku Kabushiki Kaisha

Osaka University

Tokyo Ohka Kogyo Co., Ltd.

Hiroyuki Tsunoda

Tomoyuki Igawa

Tatsuhiko Tachibana

Taichi Kuramochi

Hirotake Shiraiwa

Masayuki Tsuchiya

Atsuhiko Maeda

Tetsuro Orita

Kiyotaka Nakano

Takeshi Baba

Keiko Esaki

Toshihiko Ohtomo

Tetsuo Kojima

Yuichi Hirata

Shinya Ishii

Yasuko Kinoshita

Takahiro Ishiguro

Naohiro Yabuta

Mika Sakurai

Yoshinobu Higuchi

Yasuo Sekimori

Atsushi Kumanogoh

Keiko Kasutani

Yuri Teranishi

Souhei Ohyama

Reiko Irie

Ryusuke Omiya

Sachiyo Suzuki

Yoshinori Watanabe

Masami Suzuki

Toshiyuki Ogata

Yasuo Suzuki

Atsushi Matsuo

Atsuhiko Kato

Jun-Ichi Nezu

Takashi Ohsaka

Shigeto Kawai

Hajime Miyamoto

Takuya Noguchi

Noriyuki Takahashi

Izumi Sugo

Shigero Tamba

Yasufumi Kikuchi

Kiyoshi Habu

Takeshi Yoshino

Hiroko Konishi

Megumi Tanaka

Etsuko Takeiri

Eri Hashimoto

Akiko Koga

The present inventors provide methods for modifying the isoelectric point of an antibody while retaining its antigen-binding activity, comprising modifying the charge of at least one exposable amino acid residue on the surface of the complementarity determining region (CDR). The present invention also provides methods for purifying multispecific antibodies, comprising modifying isoelectric point, and methods for improving the plasma pharmacokinetics of antibodies, comprising modifying isoelectric point. The present invention further provides antibodies with a modified isoelectric point, pharmaceutical compositions comprising the antibodies as an active ingredient, and methods for producing the antibodies and compositions.

Method of modifying isoelectric point of antibody via amino acid substitution in CDR

Provided is a novel anti-Plexin-A1 agonist antibody that promotes dendritic cell contraction. Also provided is a pharmaceutical composition comprising such an antibody and a pharmaceutically acceptable carrier.

AntiPlexin A1 agonist antibody

In an aspect, the present invention provides a structure, a structure-manufacturing method, a cell-sorting method, or the like. In an aspect, a structure () of the present invention is a structure including a first substrate () and a second substrate () disposed to face one side of the first substrate (), in which the first substrate () has a plurality of depressions () which are open to the other side of the first substrate and each of which has a size that enables each of the depressions to capture one unit of a cell, at least some of the depressions () have communication holes () which communicate with one side and the other side of the first substrate and each of which has a size that enables secretions secreted from the cell to move through the communication holes, the second substrate () can include accumulation portions () in which the secretions moving through the communication holes () are accumulated, and the accumulation portions () can correspond to the depressions ().

Substrate, structure, structure-manufacturing method, cell-sorting method, cell-manufacturing method, and secretion-producing method

In the course of the present invention, it was discovered that one could regulate association between polypeptides by modifying amino acid residues that form the interface during the association to amino acids carrying the same type of charge. In this context, the present invention enables efficient formation of heterologous molecules. For example, the present invention can be suitably applied to the preparation of bispecific antibodies.

Methods for producing polypeptides by regulating polypeptide association

The present inventors discovered that the half-life in blood of an IgG antibody which is a polypeptide comprising an FcRn-binding domain can be controlled by controlling the surface charge through modification of residues exposed on the surface among residues in the variable regions of the IgG antibody. Antibodies whose half-life in blood had been controlled by the methods of the present invention were confirmed to actually retain the original activity. The methods of the present invention are widely applicable to polypeptides comprising an FcRn-binding domain, such as IgG antibodies, which are recycled via the FcRn salvage pathway regardless of the type of target antigen.

Methods for controlling blood pharmacokinetics of antibodies

The present inventors devised methods for efficiently purifying bispecific antibodies using a chromatography column based on the difference in isoelectric points between the H chains of two types of antibodies, wherein the difference is introduced by modifying the amino acids present on the surface of the antibody variable regions of two types of antibodies that constitute a bispecific antibody. Furthermore, the inventors devised methods for efficiently purifying bispecific antibodies using a chromatography column by linking respective antigen binding sites (heavy chain variable regions) to the antibody constant regions having different isoelectric points, and then coexpressing these antibodies.

Methods of modifying antibodies for purification of bispecific antibodies

The present invention provides pharmaceutical compositions comprising second-generation molecules that are superior than TOCILIZUMAB, by altering the amino acid sequences of the variable and constant regions of TOCILIZUMAB, which is a humanized anti-IL-6 receptor IgG1 antibody, to enhance the antigen-neutralizing ability and increase the pharmacokinetics, so that the therapeutic effect is exerted with a less frequency of administration, and the immunogenicity, safety and physicochemical properties (stability and homogeneity) are improved. The present invention also provides methods for producing these pharmaceutical compositions. The present inventors have successfully generated second-generation molecules that are superior to TOCILIZUMAB by appropriately combining amino acid sequence alterations in the CDR domains, variable regions, and constant regions.

Methods of reducing IL-6 activity for disease treatment

Provided are methods for modifying the isoelectric point of an antibody while retaining its antigen-binding activity, comprising modifying the charge of at least one exposable amino acid residue on the surface of the complementarity determining region (CDR). Also provided are methods for purifying multispecific antibodies, comprising modifying isoelectric point, and methods for improving the plasma pharmacokinetics of antibodies with a modified isoelectric point. Antibodies with a modified isoelectric point, pharmaceutical compositions comprising the antibodies as an active ingredient, and methods for producing the antibodies and compositions are also disclosed.

The inventors successfully produced anti-Epiregulin antibodies showing cross-species reactivity between cynomolgus monkey (non-human animals) and human, anti-Epiregulin antibodies with suppressed chemical degradation, anti-Epiregulin antibodies with lowered isoelectric point, anti-Epiregulin antibodies with increased thermal denaturation midpoint temperature, and anti-Epiregulin antibodies with reduced amount of aggregate by performing appropriate amino acid residue substitutions in the variable-region sequences of the humanized EP27 antibody which inhibits growth of cancer cells by exhibiting cytotoxic activity and neutralizing activity against human Epiregulin-expressing cancer cells.

Humanized anti-epiregulin antibody, and cancer therapeutic agent comprising said antibody as active ingredient

Structural isomers in sc(Fv)2 compositions of anti-human Mpl antibody and humanized anti-human Mpl antibody were separated, and the obtained structural isomers were cleaved at their linkers to confirm that the structural isomers are of single chain diabody type and bivalent scFv type. In addition, the agonistic activities of these structural isomers were revealed to be significantly different. Furthermore, the present inventors discovered that the content ratio of the structural isomers in sc(Fv)2 compositions could be regulated by altering temperature, modifying lengths of the linkers of sc(Fv)2, or amino acids in their variable regions.

Structural isomers of sc(Fv)2

Growing community of inventors

Shizuoka, Japan

Hiroyuki Tsunoda