Evaluation of multidrug resistance in in vitro treated cells of cancerous patients versus healthy donors by single cell gel electrophoresis assay (SCGE) assay.

P. Poli 1, A. Buschini 1, C. Alessandrini 1, A. Martino 1, L. Pasini 1, V. Rizzoli 2, S. Bonomini 2, C. Rossi 1.

1 Istituto di Genetica and 2 Cattedra di Ematologia, Università di Parma, 43100 Parma, Italy.

AIM. The emergence of drug-resistance during tumour treatment remains one of the major obstacles to the success of cancer chemotherapy. To confirm the hypothesis that chemoresistant cells show less DNA damage than sensitive cells and to evaluate the possible application of SCGE or comet assay in clinical practice, the response to different drugs in cells of both cancerous patients and healthy donors was compared. METHODS. Fresh leukocytes (healthy donors) or leukoblasts (patients) were treated with different antiblastic drugs for 1h. The DNA was allowed to unwind (20’, pH=13), and subjected to electrophoresis (20’, 0.78V/cm, 300mA). Ethidium bromide-stained cells were observed under a fluorescence microscope, using an image analysis system. RESULTS. Normal leukocytes show a dose-dependent response, with a linear increase of DNA migration after treatment with DNA directly acting drugs (correlation factor=0.90, dose=100, 10, and 0.05micrograms/ml for bleomycin, melphalan, and doxorubicin respectively); topotecan, inhibitor of DNA topoisomerase I, show a similar trend (dose=10micrograms/ml); mitoxantrone and etoposide, DNA topoisomerase II inhibitors, show a linear increase at the lowest doses (=1 and 0.05micrograms/ml respectively), with a “saturation” effect at higher doses; cytarabine, DNA polimerase beta inhibitor, does not appears to induce evident DNA damage at doses =10micrograms/ml. Results on patient’s leukoblasts show resistance against some drugs (in one subject, about 40%, 80%, and 60% inhibition of DNA damage induced in sensitive leukocytes by doxorubicin, melphalan and bleomycin respectively). CONCLUSIONS. The findings suggest the SCGE as one of methods useful in clinical practice to assess resistance against the drugs involved in DNA damaging during the tumour treatment

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Multidrug Resistance - MDR.