Synthesis of a novel duocarmycin derivative DU-257 and its application to immunoconjugate using poly(ethylene glycol)-dipeptidyl linker capable of tumor specific activation

Novel anti-tumor agent, duocarmycin derivative DU-257, was designed and syn thesized to prepare immunoconjugate in order to confirm the feasibility of enzymatically cleavable linker consisting of poly(ethylene glycol) (PEG) an d dipeptide. L-alanyl-L-valine. Oxyethylamine arm was introduced at 4-metho xy position of segment B of DU-86 to form DU-257 and evaluated its propel t y. DU-2.57 retained similar stability and potency with DU-86 while enhanced hydrophilicity suggested. DU-257 was condensed to the PEC-dipeptidyl linke r through carboxyl terminal of dipeptide, and enzymatic release of DU-257 u sing a model enzyme, thermolysin, similar enzyme of which was shown to be o verexpressed at various tumor sites, was evaluated by HPLC analysis. Cleava ge between the linker amino acids by the model protease and release of DU-2 57 as valine conjugated form was confirmed. The enzymatically released form of DU-257 expressed its cytotoxicity without loss of the potency for HeLaS 3 and SW1116 tumor cell lines, although the efficacy was different in indiv idual cells. DU-257 was then conjugated through the linker to KM231 monoclo nal antibody specifically reactive to GD3 antigen which was shown to be exp ressed on the surface of many malignant tumors such as SW1116. The conjugat e retained its binding specificity for SW1116 cell with a similar activity with KM231. Furthermore, the conjugate showed significant growth inhibition on SW1116 cell at a concentration of 75 mu g/mL while no effect on antigen negative cell, HeLaS3. These results suggest that the conjugate retained i ts anti-tumor effect only when it bound on and was activated at the target cell, simultaneously. DU-257 will be one of the candidate of anti-tumor age nt for application to immunoconjugate and its conjugate with KM231 via PEG- dipeptidyl linker will be a useful entity for cancer therapy related to sLe (a) expression. (C) 2000 Elsevier Science Ltd. All rights reserved.