Gene-specific damage produced by topoisomerase inhibitors in human lung cancer cells and peripheral mononuclear cells as assayed by polymerase chain reaction-stop assay.

We have previously reported that the polymerase chain reaction (PCR)-stop assay is an useful technique for investigating gene-specific damage induced by cisplatin, and that the degree of gene-specific damage sustained by peripheral blood mononuclear cells (MNC) when exposed to cisplatin in vitro predicts the response to cisplatin-based chemotherapy. In the current study, we investigated whether PCR-stop assay was suitable for investigating gene- specific damage induced by the topoisomerase I inhibitor CPT-11 or the topoisomerase II inhibitor VP-16, in the human lung cancer cell lines PC-7 and H69, respectively. The cells were incubated with CPT-11 or VP-16 for 24 hours in vitro and the hypoxanthine-phosphoribosyl transferase gene was amplified by PCR. Amplification of a 2.7kb fragment of this gene was clearly inhibited by each drug in a dose dependent manner. The concentration which reduced amplification by 63% (D63, the dose that produces an average of one lesion per single strand of the 2.7kb fragment), was 318 micrograms/ml for PC-7 treated with CPT-11 and 35 micrograms/ml for H69 treated with VP-16. We also used PCR-stop assay to investigate gene-specific damage induced by CPT-11 or VP-16 in adenocarcinoma cells from pleural effusions and MNC from freshly isolated blood obtained from 4 patients with lung cancer. Between-patient variations in the extent of gene-specific damage were observed in both types of cells. These results suggest that PCR-stop assay is suitable for the analysis of interindividual variations in the extent of gene-specific damage induced by topoisomerase inhibitors in human cells.