In vitro cytotoxicity following specific activation of amygdalin by beta-glucosidase conjugated to a bladder cancer-associated monoclonal antibody.
We describe a novel version of antibody-directed enzyme prodrug therapy (ADEPT), with the use of amygdalin as prodrug. Amygdalin is a naturally occurring cyanogenic glycoside, which can be cleaved by sweet almond beta-glucosidase to yield free cyanide. If amygdalin could be activated specifically at the tumour site, then malignant cells would be killed without the systemic toxicity usually associated with chemotherapy. To this end, we have conjugated beta-glucosidase to a tumour-associated monoclonal antibody (MAb) (HMFG1) and the conjugate has been tested in vitro for specificity and cytotoxicity subsequent to activation of amygdalin. Amygdalin was cytotoxic to HT1376 bladder cancer cells only at high concentrations, whereas the combination of amygdalin with HMFG1-beta-glucosidase enhanced the cytotoxic effect of amygdalin by 36-fold. When 2 concentrations of HMFG1-beta-glucosidase were compared, the toxic effect was dose dependent. The cytotoxicity of amygdalin was also enhanced by the MAb-enzyme conjugate even when the unbound conjugate was removed from the medium prior to exposure to amygdalin and the cells were washed. In addition to the cytotoxic effect, we also demonstrated specificity, using a MAb-enzyme conjugate that does not recognise the HT1376 bladder cancer cells. Finally, we studied the cytotoxic effect of the conjugate in co-culture of HMFG1-positive and-negative cell lines (HT 1376 and U87MG cells). We demonstrated that the rate of surviving cells corresponds well to the percentage of U87MG (HMFG1-negative) cells in the flask. Our findings indicate that ADEPT is more effective than non-directed enzyme activation of a prodrug and can result in a non-toxic cancer therapy.
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Department of Clinical Oncology, Imperial College of Science, Technology and Medicine, Hammersmith Hospital Campus, London, UK.
Rowlinson-Busza
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D000678:Amygdalin / Q000008:administration & dosage*; D000911:Antibodies, Monoclonal / Q000008:administration & dosage*; D000918:Antibody Specificity; D000951:Antigens, Neoplasm; D002470:Cell Survival; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D011355:Prodrugs / Q000008:administration & dosage*; D014407:Tumor Cells, Cultured; D001749:Urinary Bladder Neoplasms / Q000188:drug therapy* / Q000276:immunology; D001617:beta-Glucosidase / Q000008:administration & dosage*
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D011355:Prodrugs; D000678:Amygdalin; D001617:beta-Glucosidase