Preparation of AHTG-DNR conjugates and their antitumor effect in vitro

100%, 75%, 50%, 25% and 12.5% oxidized dextran T10 (Dex T10) were used as intermediate carriers for conjugating drug daunorubicin (DNR) and antibody anti-human thymocytic globulin (AHTG), to form different immunoconjugates, AHTG:Dex:DNR. It was demonstrated that the conjugate with 25% oxidized Dex T10 as intermediate carrier linked more DNR molecules than the others. The degree of its substitution was 10-11 moles of DNR per mole of AHTG. Moreover, because the amount to reducing agent sodium borohydride (NaBH4), required for the reduction reaction, was relatively small, its damaging effect on AHTG and DNR was lessened accordingly. The antitumor effect of AHTG:Dex:DNR in vitro was tested by using 24-h cytotoxicity assay, with CEM as target cell. Cytotoxic effect of the conjugate was proven and the LD50 was 10.68 micrograms/ml. However, it showed only slight cytotoxic effect on non-target cell K562. When 10 min cytotoxicity assay was performed to show the specific tumor-killing effect of the conjugate, it revealed an obvious cytotoxic activity toward CEM, with the LD50 being 14.79 micrograms/ml, but hardly toward K562. These results suggest that AHTG:Dex:DNR possesses specific cytotoxic effect.