A bivalent single-chain Fv fragment against CD47 induces apoptosis for leukemic cells.
We constructed a single-chain antibody fragment (scFv) of murine monoclonal antibody, MABL, which specifically bound to human CD47 (hCD47) and induced apoptosis of the leukemic cells. The scFv of MABL antibody with a 15-residue linker (MABL scFv-15) formed both dimer (Mr 50 kDa) and monomer (Mr 25 kDa). Both MABL scFv-15 dimer and monomer had binding activity for hCD47. MABL scFv-15 dimer strongly induced apoptosis of hCD47-introduced mouse leukemic cells in vitro and exhibited anti-tumor effect in a myeloma transplanted mice model. However, MABL scFv-15 monomer scarcely exhibited these activities. These results strongly demonstrate that the ligation of CD47 antigen by two antigen-binding sites of MABL dimer is needed for inducing apoptosis. The parent MABL antibody caused hemagglutination due to the CD47 expressed on erythrocytes. Interestingly, MABL scFv-15 dimer did not cause hemagglutination. This apoptosis-inducing dimer appears to be a lead candidate for novel leukemic therapy.
Chugai Pharmaceutical Co., Ltd., Fuji-Gotemba Research Laboratories, 1-135 Komakado, Gotemba-shi, Shizuoka-ken 412-8513, Japan. kikuchiysf@chugai-pharm.co.jp
Yasufumi
Y
Kikuchi
Shinsuke
S
Uno
Yasushi
Y
Yoshimura
Koji
K
Otabe
Shin-ichiro
S
Iida
Masayoshi
M
Oheda
Naoshi
N
Fukushima
Masayuki
M
Tsuchiya
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D015703:Antigens, CD / Q000276:immunology* / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects*; D001665:Binding Sites; D051928:CD47 Antigen; D016466:CHO Cells; D002352:Carrier Proteins / Q000276:immunology* / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D019281:Dimerization; D005434:Flow Cytometry / Q000379:methods; D006386:Hemagglutination Tests; D006801:Humans; D007128:Immunoglobulin Fragments / Q000235:genetics / Q000276:immunology* / Q000378:metabolism / Q000494:pharmacology*; D007074:Immunoglobulin G / Q000097:blood; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology; D007939:Leukemia L1210 / Q000276:immunology / Q000473:pathology* / Q000628:therapy; D008297:Male; D051379:Mice; D016513:Mice, SCID; D009101:Multiple Myeloma / Q000276:immunology / Q000473:pathology / Q000628:therapy; D009368:Neoplasm Transplantation / Q000276:immunology; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D051928:CD47 Antigen; C497894:CD47 protein, human; D002352:Carrier Proteins; C497895:Cd47 protein, mouse; D007128:Immunoglobulin Fragments; D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins