Bivalent and Bispecific Diabodies and Single-chain Diabodies

Abstract

Diabodies are small dimeric bivalent or bispecific antibody fragments formed by cross-over pairing of two single-chain VH-VL fragments (Holliger et al., 1993, Whitlow et al., 1994). Dimer formation is favoured by reducing the linker length between the VH-VL domains from 15–20 amino acids, normally used to generate scFv fragments, to approximately 5 amino acids (Fig. lA). Further reduction of the linker can result in the formation of trimeric or even tetrameric molecules (triabodies, tetrabodies) (Kortt et al., 1997; Le Gall et al., 1999). As shown by crystallographic studies, the two binding sites of a diabody molecule are facing away from each other (Perisic et al., 1994) (Fig. 1B). Bivalent diabodies are generated by dimeric assembly of two identical VH-VL chains (homodimers). Due to the presence of two antigen binding sites, bivalent diabodies exhibit an increased functional affinity (Fitzgerald et al., 1997). The expression of two fragments of the format VHA-VLB and VHB-VLA in the same cell results in formation of heterodimers recognising two different antigens, but may also lead to the formation of non-functional homodimers (Fig. 1 C).