A sensitive new method to assay
bleomycin (BLM) metabolism was developed using an ion-paired, reverse-phase high-pressure liquid chromatography technique in conjunction with fluorescence detection that allowed levels of BLM less than 20 ng/ml to be detected. The metabolism of
bleomycin B2 in homogenates from benign and malignant human
tumors was studied, and all 14
tumors were capable of metabolizing
bleomycin to desamidobleomycin B2. Metabolites other than desamidobleomycin B2 were not detected. BLM
hydrolase activities in individual
tumors varied more than 7-fold. The importance of BLM
hydrolase in limiting the therapeutic effectiveness of BLM was examined by measuring BLM
hydrolase activity and response of human
tumors to BLM in culture. Response to BLM in culture was measured by dissociating human
tumors to form single-cell
suspensions and exposing the cells to 0.05 or 1 microgram/ml of BLM for 1 hr. Colony formation after BLM treatment was determined in soft
agar and when compared to that of untreated cells, varied by more than 100-fold. No correlation was observed, however, between BLM
hydrolase activity and response to BLM in soft
agar. Thus, human
tumors can metabolize BLM, and while BLM
hydrolase activity may be important in
tumor resistance to the
drug, these data suggest that either (a) the
enzyme activity in the
tumor homogenate does not reflect that in the clonogenic cells or (b) other mechanisms of resistance may be operative.