Development of a novel bi-specific monoclonal antibody approach for tumourtargeting

To overcome the disadvantages of bi-specific antibody methodologies in vivo , a novel antibody approach has been designed to improve tumour targeting a nd effector to target ratio. The technique involves biotinylated anti-CDS F ab fragments and streptavidinylated anti-tumour monoclonal antibodies (mAbs ) that can spontaneously form cross-links. We describe here a method for th e direct cross-linking of sulphydryl-conjugated HMFG1 (anti-MUC1 mucin mAb) to streptavidin by sulphosuccinimidyl-4-(N-maleimidomethyl) cyclohexanel-1 -carboxylate. Fab fragments generated by papain digestion of the 1452C11 an tibody (anti-CDS mAb without Pc to avoid peripheral activation of T-cells) were biotinylated with NHS-Iminobiotin. MUC1-transfected BALB/c breast canc er cell lines 413BCR and 425CCR and the parental cell line (410.4) were lab elled with streptavidinylated mouse anti-MUC1 mucin mAb. BALB/c effector T- cells were separately labelled with biotinylated anti-CD3 Fab fragments (14 52C11) and mixed with tumour cells in different effector to target ratios. Percentage of killing was assessed using the Cr-51 cytotoxicity assay. Seve nty per cent lysis was measured in the case of 413BCR (high MUC1 mucin expr essor) and 40% in the case of 425CCR (low expressor) cell line. No lysis wa s apparent in the MUC1 negative cell line. These results demonstrate that t he novel T-cell redirecting approach we have developed can produce effectiv e immune lysis of target cells in vitro. (C) 1999 Cancer Research Campaign.