DIABODIES - SMALL BIVALENT AND BISPECIFIC ANTIBODY FRAGMENTS

Bivalent and bispecific antibodies and their fragments have immense po tential for practical application. Here we describe the design of smal l antibody fragments with two antigen-binding sites. The fragments com prise a heavy-chain variable domain (V(H)) connected to a light-chain variable domain (V(L)) on the same polypeptide chain (V(H)-V(L)). By u sing a linker that is too short to allow pairing between the two domai ns on the same chain, the domains are forced to pair with the compleme ntary domains of another chain and create two antigen-binding sites. A s indicated by a computer graphic model of the dimers, the two pairs o f domains can pack together with the antigen-binding sites pointing in opposite directions. The dimeric antibody fragments, or ''diabodies,' ' can be designed for bivalent or bispecific interactions. Starting fr om the monodonal antibodies NQ11.7.22 (NQ11) and D1.3 directed against the hapten phenyloxazolone and hen egg lysozyme, respectively, we bui lt bivalent fragments (V(H)NQ11-V(L)NQ11)2 and (V(H)D1.3-V(L)D1.3)2 an d bispecific fragments V(H)NQ11-V(L)D1.3 and V(H)D1.3-V(L)NQ11. The fr agments were expressed by secretion from bacteria and shown to bind sp ecifically to the hapten and/or antigen. Those with 5- and 15=residue linkers had similar binding affinities to the parent antibodies, but a fragment with the V(H) domain joined directly to the V(L) domain was found to have slower dissociation kinetics and an improved affinity fo r hapten. Diabodies offer a ready means of constructing small bivalent and bispecific antibody fragments in bacteria.