Characterization of a CC49-based single-chain fragment-beta-lactamase fusion protein for antibody-directed enzyme prodrug therapy (ADEPT).
CC49 is a clinically validated antibody with specificity for TAG-72, a carbohydrate epitope that is overexpressed and exposed on the cell surface in a large fraction of solid malignancies. We constructed a single-chain fragment (scFv) based on CC49 and fused it to beta-lactamase (BLA). Following optimization of the scFv domain by combinatorial consensus mutagenesis (CCM) for increased expression and stability, we characterized the protein variant for binding, in vivo pharmacokinetics (PK), and antitumor efficacy. The fusion protein TAB2.5 possessed a similar binding specificity relative to the parent antibody CC49. TAB2.5 also showed prolonged retention (T(1/2) = 36.9 h) in tumor-bearing mice with tumor/plasma ratios of up to 1000. Preliminary evaluation of TAB2.5, in combination with a novel prodrug, GC-Mel, resulted in significant efficacy in a colorectal xenograft tumor model and supports the utility of the protein as an agent for tumor-selective prodrug activation.
Genencor International, a Danisco company, 925 Page Mill Road, Palo Alto, California 94304, USA.
Ralph F
RF
Alderson
Brian E
BE
Toki
Martin
M
Roberge
Wei
W
Geng
Joshua
J
Basler
Regina
R
Chin
Amy
A
Liu
Roanna
R
Ueda
Douglas
D
Hodges
Enrique
E
Escandon
Tianling
T
Chen
Tessi
T
Kanavarioti
Lilia
L
Babé
Peter D
PD
Senter
Judith A
JA
Fox
Volker
V
Schellenberger
D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000627:therapeutic use; D000912:Antibodies, Neoplasm / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000627:therapeutic use; D018906:Antineoplastic Agents, Alkylating / Q000737:chemistry / Q000378:metabolism / Q000627:therapeutic use; D000972:Antineoplastic Agents, Phytogenic / Q000627:therapeutic use; D002166:Camptothecin / Q000031:analogs & derivatives / Q000627:therapeutic use; D002511:Cephalosporins / Q000737:chemistry* / Q000378:metabolism* / Q000627:therapeutic use*; D015179:Colorectal Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D004317:Doxorubicin / Q000627:therapeutic use; D004337:Drug Carriers / Q000737:chemistry / Q000378:metabolism; D016503:Drug Delivery Systems; D005260:Female; D006801:Humans; D007128:Immunoglobulin Fragments / Q000235:genetics / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000378:metabolism; D000077146:Irinotecan; D008558:Melphalan / Q000737:chemistry / Q000378:metabolism / Q000627:therapeutic use; D051379:Mice; D008819:Mice, Nude; D015394:Molecular Structure; D009368:Neoplasm Transplantation; D009588:Nitrogen Mustard Compounds / Q000737:chemistry* / Q000378:metabolism* / Q000627:therapeutic use*; D011355:Prodrugs / Q000737:chemistry / Q000378:metabolism / Q000627:therapeutic use; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000627:therapeutic use*; D001618:beta-Lactamases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000627:therapeutic use
D000903:Antibiotics, Antineoplastic; D000912:Antibodies, Neoplasm; D018906:Antineoplastic Agents, Alkylating; D000972:Antineoplastic Agents, Phytogenic; C091202:B72.3 antibody; D002511:Cephalosporins; D004337:Drug Carriers; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D009588:Nitrogen Mustard Compounds; D011355:Prodrugs; D011993:Recombinant Fusion Proteins; C511565:TAB2.5 fusion protein; C511564:glutaryl C-Mel; D000077146:Irinotecan; D004317:Doxorubicin; D001618:beta-Lactamases; D008558:Melphalan; D002166:Camptothecin