Preferential heterodimerization of a bispecific diabody based on a humanized anti-EGFR antibody 528.
We report the utility of in vitro refolding in the preparation of monomorphous hEx3 bispecific diabodies with epidermal growth factor receptor and CD3 retargeting from insoluble aggregates in Escherichia coli. Appropriate interaction between cognate variable heavy and light chains led to the formation of functional hEx3 heterodimers in a diabody format rather than inactive homodimers. The refolded hEx3 was found to exhibit almost the equivalent activity to the hEx3 and single-chain hEx3 (hEx3-scDb) prepared in a mammalian secretion system. We suggest that the preparation of hEx3 from bacterial insoluble material by means of in vitro refolding would be useful for industrial-scale production of the diabody for its potential use in clinical studies.
Asano
Ryutaro
R
Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University, Sendai 980-8579, Japan.
Sone
Yukiko
Y
Ikoma
Keiko
K
Hayashi
Hiroki
H
Nakanishi
Takeshi
T
Umetsu
Mitsuo
M
Katayose
Yu
Y
Unno
Michiaki
M
Kudo
Toshio
T
Kumagai
Izumi
I
D000818:Animals; D018033:Antibodies, Bispecific / Q000737:chemistry* / Q000276:immunology* / Q000378:metabolism / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000737:chemistry / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology; D016466:CHO Cells; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002850:Chromatography, Gel; D006224:Cricetinae; D003412:Cricetulus; D019281:Dimerization; D066246:ErbB Receptors / Q000276:immunology*; D015870:Gene Expression; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000378:metabolism; D007147:Immunoglobulin Light Chains / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000378:metabolism; D020673:Protein Renaturation; D020836:Protein Structure, Quaternary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology; D013379:Substrate Specificity
D018033:Antibodies, Bispecific; D000970:Antineoplastic Agents; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins; D066246:ErbB Receptors