Binding characteristics and tumor targeting of a covalently linked divalent CC49 single-chain antibody.
Multivalency is a recognized means of increasing the functional affinity of single-chain Fvs (scFvs) for optimizing tumor uptake. A unique divalent single-chain Fv protein [sc(Fv)2], based on the variable regions of the monoclonal antibody (MAb) CC49, has been generated that differs from other dimeric single-chain constructs in that a linker sequence (L) is encoded between the repeated V(L) and V(H) domains (V(L)-L-V(H)-L-V(L)-L-V(H)). This construct was expressed in soluble form in Escherichia coli and purified by ion-exchange and gel-filtration chromatography. Purity and immunoreactivity were determined by SDS-PAGE, HPLC and competitive RIA. sc(Fv)2 exhibited a relative K(A) (3.34 x 10(7) M(-1)) similar to that of the native IgG (1.14 x 10(8) M(-1)) as determined by BIAcore analysis. Pharmacokinetic studies showed rapid blood clearance for sc(Fv)2, with a T(1/2) less than 40 min. Whole-body clearance analysis also revealed rapid clearance, suggesting no significant retention in the extravascular space or normal tissues. Biodistribution studies of radiolabeled sc(Fv)2 showed tumor uptake greater than 6% ID/g after 30 min, which remained at this level for 6 hr. High tumor uptake and retention of sc(Fv)2 coupled with rapid blood and whole-body clearance makes this dimeric scFv of MAb CC49 a strong candidate for imaging and therapeutic applications.
Beresford
G W
GW
Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha 68198-3135, USA.
Pavlinkova
G
G
Booth
B J
BJ
Batra
S K
SK
Colcher
D
D
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000302:isolation & purification / Q000493:pharmacokinetics*; D000912:Antibodies, Neoplasm / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D003110:Colonic Neoplasms / Q000378:metabolism*; D004926:Escherichia coli; D005260:Female; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D007457:Iodine Radioisotopes / Q000493:pharmacokinetics*; D008657:Metabolic Clearance Rate; D051379:Mice; D008819:Mice, Nude; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000493:pharmacokinetics; D014018:Tissue Distribution; D014183:Transplantation, Heterologous
D000911:Antibodies, Monoclonal; D000912:Antibodies, Neoplasm; C091202:B72.3 antibody; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D007457:Iodine Radioisotopes; D011994:Recombinant Proteins