Generation and characterization of dimeric small immunoproteins specific for neuroblastoma associated antigen GD2

GD2 is a disialoganglioside expressed at high density on the surface of malignant cells of neuroectodermal origin, especially in neuroblastoma (NB) and melanoma. Since its expression in normal tissues is very restricted, GD2 represents an excellent target for neuroectodermal tumor targeting. Mini-antibody technology allows the production of dimeric single-chain antibodies, also called small immunoproteins (SIPs), which are composed of a scFv fused to a dimerizing domain of immunoglobulin heavy chains. Dimerization results in an increase of the total apparent affinity and a slower clearance in vivo than scFvs. These properties make SIPs very attractive molecules for tumor targeting. We isolated the variable regions from an anti-GD2 monoclonal antibody and exploited the SIP technology to generate two novel anti-GD2 SIPs. The first anti-GD2 SIP is a fully murine molecule containing the CH3 domain of mouse IgG1, whereas the second construct is a hybrid mouse-human molecule containing the CH4 domain of human IgE. Both mini-antibodies were successfully produced and shown to retain binding specificity as well as an affinity similar to that of the original antibody.