A Bioassay to Measure Cytotoxicity of Plasma from Patients Treated with Mitomycin C1

The unpredictable clinical toxicity observed in patients treated with mitomycin C and the observation that this agent must be reduced to an active form before alkylating target molecules have led to the development of a bioassay which is capable of detecting biologically active forms of mitomycin C in the plasma of drug-treated patients. The bioassay makes use of a repairdeficient mutant of Chinese hamster ovary cells, UV-20, which is 40 to 60 times more sensitive to mitomycin C than its wildtype parent. A standard curve relating in vitro cell colony-forming ability of UV-20 versus drug concentration in the plating medium has been determined. Mitomycin C levels in patient plasma as low as 1 ng/ml can be detected, compared to the 5-ng/ml limit of detection obtained with a high-pressure liquid chromatography assay for the parent compound. This assay has been utilized to detect active drug in plasma obtained from patients with colorectal cancer treated with mitomycin C as a single agent. At the completion of drug injection, serial blood samples were collected in heparinized tubes, and aliquots of plasma were extracted and assayed for mitomycin C levels by high-pressure liquid chroma tography, diluted and assayed directly for their toxicity for UV20 cells, or frozen at -20°C to be assayed at a later time. The activity detected by immediate bioassy was stable up to 2 mo in frozen samples. Plasma pharmacokinetics determined by the bioassay in seven patients were no different than those deter mined by high-pressure liquid chromatography. No stable, cytotoxic species other than the parent compound were detected by the bioassay in the plasma of patients treated with mitomycin C.