Dimeric and trimeric antibodies: high avidity scFvs for cancer targeting

Recombinant antibody fragments can be engineered to assemble into stable mu ltimeric oligomers of high binding avidity and specificity to a wide range of target antigens and haptens. This review describes the design and expres sion of diabodies (dimers), triabodies (trimers) and tetrabodies (tetramers ). In particular we discuss the role of linker length between V-domains and the orientation of the V-domains to direct the formation of either diabodi es (60 kDa), triabodies (90 kDa) or tetrabodies (120 kDa), and how the size , flexibility and valency of each molecules is suited to different applicat ions for in vivo imaging and therapy. Single chain Fv antibody fragments jo ined by polypeptide linkers of at least 12 residues irrespective of V-domai ns orientation predominantly form monomers with varying amounts of dimer an d higher molecular mass oligomers in equilibrium. A scFv molecule with a li nker of 3-12 residues cannot fold into a functional Fv domain and instead a ssociates with a second, scFv molecule to form a bivalent dimer (diabody, s imilar to 60 kDa). Reducing the linker length below three residues can forc e scFv association into trimers (triabodies, similar to 90 kDa) or tetramer s (similar to 120 kDa) depending on linker length, composition and V-domain orientation. A particular advantage for tumour targeting is that molecules of 60-100 kDa have increased tumour penetration and fast clearance rates c ompared with the parent Ig (150 kDa). We highlight a number of cancer-targe ting scFv diabodies that have undergone successful pre-clinical trials for in vivo stability and efficacy. We also briefly review the design of multi- specific Fv modules suited to cross-link two or more different target antig ens. Bi-specific diabodies formed by association of different scFv molecule s have been designed as cross-linking reagents for T-cell recruitment into tumours (immunotherapy), viral retargeting (gene therapy) and as red blood cell agglutination reagents (immuno diagnostics). The more challenging tris pecific multimers (triabodies) remain to be described. (C) 2001 Elsevier Sc ience B.V. All rights reserved.